Vector-borne diseases - Eurosurveillance

E u r o p e ’ s j o u r n a l o n i n f e c t i o u s d i s e a s e e p i d e m i o l o g y, p r e v e n t i o n a n d c o n t r o l 

Special edition: 

Vector-borne 

diseases 

Jan − Dec 2010 

• Almost a third of the recorded events related to emerging 

infectious diseases in Europe were due to vector-borne diseases 

during the decade 1990-2000 

• In this special edition Eurosurveillance presents a series of 

review articles with a particular focus on arthropod-borne 

diseases transmitted by mosquitoes and phlebotomine 

sandflies and reports on recent outbreaks caused by vectorborne 

diseases in Europe 

www.eurosurveillance.org 

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Editor-in-Chief 

Karl Ekdahl 

Managing Editor 

Ines Steffens 

Scientific Editors 

Kathrin Hagmaier 

Williamina Wilson 

Assistant Editors 

Alina Buzdugan 

Ingela Söderlund 

Associate Editors 

Andrea Ammon, ECDC, Stockholm, Sweden 

Tommi Asikainen, ECDC, Stockholm, Sweden 

Mike Catchpole, Health Protection Agency, 

London, United Kingdom 

Denis Coulombier, ECDC, Stockholm, Sweden 

Christian Drosten, Universitätsklinikum Bonn, Bonn, Germany 

Johan Giesecke, ECDC, Stockholm, Sweden 

Herman Goossens, Universiteit Antwerpen, Antwerp, Belgium 

David Heymann, World Health Organisation, Geneva, 

Switzerland 

Irena Klavs, National Institute of Public Health, Ljubljana, 

Slovenia 

Karl Kristinsson, Landspitali University Hospital, Reykjavik, 

Iceland 

Daniel Lévy-Bruhl, Institut de Veille Sanitaire, Paris, France 

Richard Pebody, Health Protection Agency, London, 

United Kingdom 

Panayotis T. Tassios, University of Athens, Athens, Greece 

Hélène Therre, Institut de Veille Sanitaire, Paris, France 

Henriette de Valk, Institut de Veille Sanitaire, Paris, France 

Sylvie van der Werf, Institut Pasteur, Paris, France 

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© Eurosurveillance, 2011 

Austria: Reinhild Strauss, Vienna 

Belgium: Koen De Schrijver, Antwerp 

Belgium: Sophie Quoilin, Brussels 

Bulgaria: Mira Kojouharova, Sofia 

Croatia: Borislav Aleraj, Zagreb 

Cyprus: Olga Poyiadji-Kalakouta, Nicosia 

Czech Republic: Bohumir Križ, Prague 

Denmark: Peter Henrik Andersen, Copenhagen 

England and Wales: Neil Hough, London 

Estonia: Kuulo Kutsar, Tallinn 

Finland: Hanna Nohynek, Helsinki 

France: Judith Benrekassa, Paris 

Germany: Jamela Seedat, Berlin 

Greece: Rengina Vorou, Athens 

Hungary: Ágnes Csohán, Budapest 

Iceland: Haraldur Briem, Reykjavik 

Ireland: Lelia Thornton, Dublin 

Italy: Paola De Castro, Rome 

Latvia: Jurijs Perevoščikovs, Riga 

Lithuania: Milda Zygutiene, Vilnius 

Luxembourg: Robert Hemmer, Luxembourg 

FYR of Macedonia: Elisaveta Stikova, Skopje 

Malta: Tanya Melillo Fenech, Valletta 

Netherlands: Paul Bijkerk, Bilthoven 

Norway: Hilde Klovstad, Oslo 

Poland: Malgorzata Sadkowska-Todys, Warsaw 

Portugal: Judite Catarino, Lisbon 

Romania: Daniela Pitigoi, Bucharest 

Scotland: Norman Macdonald, Glasgow 

Slovakia: Lukáš Murajda, Martin 

Slovenia: Alenka Kraigher, Ljubljana 

Spain: Elena Rodríguez Valín, Madrid 

Sweden: Aase Sten, Stockholm 

Turkey: Aysegul Gozalan, Istanbul 

European Commission: Paolo Guglielmetti, Luxembourg 

World Health Organization Regional Office for Europe: Nedret 

Emiroglu, Copenhagen

Contents 

Special focus: Vector-borne diseases 

Editorials 

A perspective on emerging mosquito and 

phlebotomine-borne diseases in Europe 3 

by G Hendrickx, R Lancelot 

West Nile virus: the need to strengthen 

preparedness in Europe 5 

by H Zeller, A Lenglet, W Van Bortel 

Surveillance and outbreak reports 

West Nile virus circulation in Emilia-Romagna, 

Italy: the integrated surveillance system 2009 7 

by P Angelini, M Tamba, A C Finarelli, R Bellini, A Albieri, 

P Bonilauri, F Cavrini, M Dottori, P Gaibani, E Martini, 

A Mattivi, A M Pierro, G Rugna, V Sambri, G Squintani, P Macini 

Phylogenetic analysis in a recent controlled 

outbreak of Crimean-Congo haemorrhagic fever in 

the south of Iran, December 2008 12 

by S Chinikar, S Mojtaba Ghiasi, M Moradi, M M Goya, 

M Reza Shirzadi, M Zeinali, E Mostafavi, M Pourahmad, A Haeri 

Reviews 

West Nile virus in Europe: understanding the 

present to gauge the future 16 

by P Reiter 

Yellow fever and Dengue: a threat to Europe? 23 

by P Reiter 

Rift valley fever - a threat for Europe? 30 

by V Chevalier, M Pépin, L Plée, R Lancelot 

Leishmaniasis emergence in Europe 41 

by P D Ready 

Arthropod-borne viruses transmitted by 

phlebotomine sandflies in Europe: a review 52 

by J Depaquit, M Grandadam, F Fouque, PE Andry, C Peyrefitte 

Perspectives 

Crimean-Congo hemorrhagic fever in Europe: 

current situation calls for preparedness 60 

by H C Maltezou, L Andonova, R Andraghetti, M Bouloy, 

O Ergonul, F Jongejan, N Kalvatchev, S Nichol, M Niedrig, 

A Platonov, G Thomson, K Leitmeyer, H Zeller 

Rapid communications 

Chikungunya infection in a French traveller 

returning from the Maldives, October, 2009 64 

by M Receveur, K Ezzedine, T Pistone, D Malvy 

Chikungunya fever in two German tourists 

returning from the Maldives, September, 2009 66 

by M Pfeffer, I Hanus, T Löscher, T Homeier, G Dobler 


A case of dengue type 3 virus infection imported 

from Africa to Italy, October 2009 70 

by C Nisii, F Carletti, C Castilletti, L Bordi, S Meschi, 

M Selleri, R Chiappini, D Travaglini, M Antonini, S Castorina, 

F N Lauria, P Narciso, M Gentile, L Martini, G Di Perri, 

S Audagnotto, R Biselli, M Lastilla, A Di Caro, 

M Capobianchi, G Ippolito 

Recent expansion of dengue virus serotype 3 in 

West Africa 73 

by L Franco, A Di Caro, F Carletti, O Vapalahti, C Renaudat, 

H Zeller, A Tenorio 

Dengue type 3 virus infections in European 

travellers returning from the Comoros and 

Zanzibar, February-April 2010 77 

by P Gautret, F. Simon, H Hervius Askling, O Bouchaud, 

I Leparc-Goffart, L Ninove, P Parola, for EuroTravNet 

Fatal and mild primary dengue virus infections 

imported to Norway from Africa and south-east 

Asia, 2008-2010 80 

by K Vainio, S Noraas, M Holmberg, H Fremstad, 

M Wahlstrøm, G Ånestad, S Dudman 

First two autochthonous dengue virus infections in 

metropolitan France, September 2010 84 

by G La Ruche, Y Souarès, A Armengaud, F Peloux-Petiot, 

P Delaunay, P Desprès, A Lenglet, F Jourdain, I Leparc- 

Goffart, F Charlet, L Ollier, K Mantey, T Mollet, J P Fournier, 

R Torrents, K Leitmeyer, P Hilairet, H Zeller, W Van Bortel, 

D Dejour-Salamanca, M Grandadam, M Gastellu-Etchegorry 

Dengue virus infection in a traveller returning from 

Croatia to Germany 89 

by J Schmidt-Chanasit, M Haditsch, I Schöneberg, 

S Günther, K Stark, C Frank 

Relapsing vivax malaria cluster in Eritrean refugees, 

Israel, June 2010 91 

by E Kopel, E Schwartz, Z Amitai, I Volovik 

First autochthonous malaria case due to 

plasmodium vivax since eradication, Spain, 

October 2010 94 

by P Santa-Olalla Peralta, M C Vazquez-Torres, E Latorre- 

Fandós, P Mairal-Claver, P Cortina-Solano, A Puy-Azón, 

B Adiego Sancho, K Leitmeyer, J Lucientes-Curdi, M J Sierra-Moros 

Infection with Mayaro virus in a French traveller 

returning from the Amazon region, Brazil, 

January, 2010 97 

by M C Receveur, M Grandadam, T Pistone, D Malvy 

Epidemiological analysis of mosquito-borne 

pogosta disease in Finland, 2009 100 

by J Sane, S Guedes, S Kurkela, O Lyytikäinen, O Vapalahti 

Ongoing outbreak of West Nile virus infections in 

humans in Greece, July – August 2010 103 

by A Papa, K Danis, A Baka, A Bakas, G Dougas, T Lytras,G 

Theocharopoulos, D Chrysagis, E Vassiliadou, F Kamaria, A 

Liona, K Mellou, G Saroglou, T Panagiotopoulos 

A head on view of a Culex pipiens mosquito, this species is an important vector in West Nile Virus. 

© Istockphoto 

1

Retrospective screening of solid organ donors in 

Italy, 2009, reveals unpredicted circulation of West 

Nile virus 108 

by M R Capobianchi, V Sambri, C Castilletti, A M Pierro, 

G Rossini, P Gaibani, F Cavrini, M Selleri, S Meschi, D Lapa, 

A Di Caro, P Grossi, C De Cillia, S Venettoni, M P Landini, 

G Ippolito, A Nanni Costa, on behalf of the Italian Transplant 

Network 

Case report: West-Nile virus infection in two Dutch 

travellers returning from Israel 113 

by N Aboutaleb, M FC Beersma, H F Wunderink, 

A CTM Vossen, L G Visser 

Introduction and control of three invasive 

mosquito species in the Netherlands, 

July-October 2010 114 

by E J Scholte, W Den Hartog, M Dik, B Schoelitsz, M Brooks, 

F Schaffner, R Foussadier, M Braks, J Beeuwkes 

2 www.eurosurveillance.org

Editorials 

A perspective on emerging mosquito and phlebotomineborne 

diseases in Europe 

G Hendrickx (info@avia-gis.be) 1 , R Lancelot 2 

1. Avia-GIS, Agro-Veterinary Information and Analysis, Zoersel, Belgium 

2. CIRAD, French Agricultural Research Centre for International Development, UMR CMAEE (Control of emerging and exotic animal 

diseases), Montpellier, France 

Citation style for this article: 

Citation style for this article: Hendrickx G, Lancelot R. A perspective on emerging mosquito and phlebotomine-borne diseases in Europe. Euro Surveill. 

2010;15(10):pii=19503. Available online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19503 

Emerging infectious diseases are of increasing concern 

worldwide and in particular in Europe. In a review, 

Jones et al. have shown that between 1940 and 2004, 

the majority of emerging infectious diseases occurred 

in areas with both a high mobility and high density of 

population, notably in Western Europe. Furthermore, 

nearly a third (29%) of the recorded events related to 

emerging infectious diseases were due to vector-borne 

diseases during the decade 1990-2000[1]. 

This issue of Eurosurveillance presents a series of 

review articles with a particular focus on arthropodborne 

diseases transmitted by mosquitoes and phlebotomine 

sandflies. Each of the papers addresses a 

series of issues of common interest such as the relevance 

of the disease in the given context, its transmission 

and epidemiology, including the current 

geographical distribution, and clinical symptoms, 

diagnostic methods, treatment strategies and prevention 

methods. Furthermore, the papers describe factors 

triggering changes in distribution of the vectors 

and disease and risk prediction models. 

A review on West Nile virus by Reiter includes a fresh 

and innovative viewpoint on the epidemiology and 

transmission of the disease [2], and the same author 

contributed further with a twin-review on two diseases 

which have much in common: yellow fever and 

dengue [3]. Most importantly, both have a history of 

occurrence in Europe and vectors and pathogens are 

spreading through increased movement of persons and 

transport of goods. Chevalier et al. present a review on 

Rift Valley fever a mosquito-borne disease at Europe’s 

fringes. The epidemiology of Rift Valley fever is fascinating 

because of its complex cycle where the virus 

may remain dormant for many years and outbreaks 

involve both vector related and direct transmission. 

The disease may become a risk in the future in countries 

bordering the Mediterranean Sea, mainly through 

increased livestock trade. 

Two authors have contributed reviews on viruses transmitted 

by phlebotomine sandflies. Ready has written 


This article has been published on 11 March 2010 

about Leishmania, a parasite of particular relevance 

to Europe because it is currently established around 

the Mediterranean Sea, but known to be spreading 

north [4]. Depaquit et al. have contributed a review on 

a number of less well known Phlebo-, Vesiculo- and 

Orbiviruses such as Sandfly fever Sicilian and Naples 

virus, Toscana virus Chandipura virus and others that 

are transmitted by sandflies in Europe and more specifically 

around the Mediterranean Sea [5]. The paper 

summarizes the current knowledge on these viruses 

which have a potential to spread throughout the distribution 

zone of their phlebotomine vectors in Europe. 

Both reviews provide a series of maps displaying 

country based information on the distribution of the 

disease. 

In addition to these papers, the issue features a perspective 

paper by Maltezou et al. presenting the 

present situation of Crimean-Congo hemorrhagic fever, 

a tick-borne disease, in Europe and emphasizing relevant 

aspects for preparedness concerning the potential 

spread of this disease in Europe in the future [6]. 

As will become clear from reading these reviews, often 

crucial knowledge is still missing which is needed to 

anticipate, prevent or prepare for the establishment 

and spread of vector-borne diseases. One of these is 

reliable information on the continent wide distribution 

of potential disease vectors. National presenceabsence 

maps, as shown by Ready [4] and Depaquit 

[5] are a first step in this direction and need further 

refinement. 

In 2007/08, the European Centre for Disease Prevention 

and Control funded the V-borne project with the aim to 

identify and document vector-borne diseases relevant 

for public health in Europe, provide an overview of 

the existing relevant resources, carry out a qualitative 

multi-disciplinary risk assessment within the limits of 

the available information and data, and identify priorities 

for future prevention and control of vector borne 

diseases in Europe. Building on the expertise from 

this network, ECDC created a European network for 

3

arthropod vector surveillance for human public health, 

the VBORNET in 2009 [7]. The network will establish 

pan-European state of the art maps of validated vector 

distributions that can be used as basis for risk 

assessment studies thus contributing to preparedness 

for the emerge or re-emerge of vector-borne disease in 

Europe. 

References 

1. Jones KE, Pate NG, Levy MA, Storeygard A, Balk D, Gittleman 

JL et al. Global trends in emerging infectious diseases. Nature. 

2008;451:990-3. Available from: http://dx.doi.org/10.1038/ 

nature06536 

2. Reiter P. West Nile virus in Europe: understanding the present 

to gauge the future. Euro Surveill. 2010;15(10):pii=19508. 

Available from: http://www.eurosurveillance.org/ViewArticle. 

aspx?ArticleId=19508 

3. Paul Reiter Dengue and Yellow fever Eurosurveillance. Euro 

Surveill. 2010;15(10):pii=19509. Available online: http://www. 

eurosurveillance.org/ViewArticle.aspx?ArticleId=19509 

4. Chevalier V, Pépin M, Plée L, Lancelot R. Rift Valley fever - a 

threat for Europe?. Euro Surveill. 2010;15(10):pii=19506. 



5. Depaquit J, Grandadam M, Fouque F, Andry P, 

Peyrefitte C. Arthropod-borne viruses transmitted by 

Phlebotomine sandflies in Europe: a review . Euro Surveill. 

2010;15(10):pii=19507. Available from: http://www. 


6. Maltezou HC, Andonova L, Andraghetti R, Bouloy M, Ergonul 

O, Jongejan F, Kalvatchev N, Nichol S, Niedrig M, Platonov A, 

Thomson G, Leitmeyer K, Zeller H. Crimean-Congo hemorrhagic 

fever in Europe: current situation calls for preparedness. Euro 

Surveill. 2010;15(10):pii=19504. Available online: http://www. 


7. European Centre for Disease prevention and Control (ECDC). 

Network of medical entomologists and public health experts 

(VBORNET). Available from: http://www.ecdc.europa.eu/en/ 

activities/diseaseprogrammes/Pages/VBORNET.aspx 


Editorials 

West Nile virus: the need to strengthen preparedness in 

Europe 

H Zeller (evd@ecdc.europa.eu) 1 , A Lenglet 1 , W Van Bortel 1 

1. European Centre for Disease Prevention and Control, Stockholm, Sweden 


Zeller H, Lenglet A, Van Bortel W. West Nile virus: the need to strengthen preparedness in Europe. Euro Surveill. 2010;15(34):pii=19647. Available online: http:// 

www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19647 

The ongoing outbreak of West Nile virus (WNV) infections 

in humans in Greece described in this issue of 

Eurosurveillance is a timely reminder that WNV is a reemerging 

pathogen in Europe [1]. So far, WNV has been 

documented in animals and humans in several countries 

across Europe, mainly in central Europe and in 

the Mediterranean region. Over the last 15 years, outbreaks 

in horses and/or humans were reported from 

Romania, Hungary and Portugal, Spain, France, Italy 

and Greece [2]. 

In 2010, a single probable human case was reported 

in July in Portugal. Outside the European Union (EU), 

WNV circulation has been documented in horses in 

Morocco and human cases have occurred in Russia 

(Volgograd Oblast) and in Israel. All these regions are 

located along the main routes of migratory birds. The 

current outbreak in humans in northern Greece, is the 

first recognised WN fever outbreak in humans in this 

country. However, studies suggest that WNV has probably 

been circulating in humans in the region of central 

Macedonia in northern Greece for many years [3,4]. 

West Nile fever is a viral disease transmitted by mosquitoes 

and is distributed worldwide. The primary cycle 

of WNV involves ornithophilic mosquitoes and birds; 

some mosquito species mostly from the Culex genus 

can bite infectious birds and subsequently transmit the 

virus to humans and/or horses during another blood 

meal. Humans and horses are considered as deadend 

hosts. The vast majority of human cases remain 

asymptomatic after infection and severe neuroinvasive 

illness is reported in less than 1% of the patients. The 

main risk factor for severe clinical presentation is to 

be an elderly person. In this age group, reported case 

fatality rates may reach 10% [5]. In addition the high 

number of non-symptomatic cases may increase the 

risk of WNV transmission through blood donation or 

organ transplants [6]. 

Each WNV outbreak is unique in that there is a complex 

interaction of different factors in space and time that 

contribute to the transmission of the virus to humans. 

These factors range from the introduction of infected 

migratory birds into native local bird populations, to 


Article published on 26 August 2010 

climatic factors that increase the abundance of competent 

mosquito vectors and bridge vectors, to changes 

in human behaviour that favour exposure to infected 

mosquitoes. It is this complexity that makes each WNV 

outbreak particular and that make development and 

implementation of preparedness plans for the prevention 

of cases in humans so difficult. 

The recently reported probable and confirmed cases 

of WNV infection in Portugal and Greece, respectively 

reconfirm that this virus is actively circulating in several 

countries in the EU and that transmission to humans 

can be expected on a regular basis during the mosquito 

season. Also, reports of sporadic cases from several 

regions in Hungary during previous years indicate 

that WNV activity is widely distributed throughout this 

country and not limited to a single focus [7]. A recent 

study in Italy linked to infected organ donors [8] draws 

the same conclusion, that the virus is being transmitted 

in areas previously thought to not be at risk or 

affected. Furthermore, the case report in this issue of a 

Dutch traveller returning from Israel with WN infection 

highlights the need for awareness among physicians 

and laboratory staff to consider WNV infections as a 

differential diagnosis in cases where patients return 

from areas where they may have been exposed to the 

virus [9]. 

The events described above strengthen the need for 

integrated multidisciplinary surveillance systems and 

response plans. This includes raising the awareness of 

clinicians and veterinarians of the clinical presentation 

of WNV disease in humans and horses (particularly 

during the mosquito season from June to October), primarily 

in areas considered as at major risk surrounding 

irrigated areas and river deltas. Furthermore, strengthening 

the understanding of suitable habitats for birds 

that would increase the bird-mosquito-human interface 

would be of value. In terms of entomology, a thorough 

understanding of competent vector species, their 

breeding ecology, their abundance and geographic 

range is of significant importance in establishing limits 

around WNV affected areas and in the identification of 

potential new at-risk areas. 

5

In addition, there is a need to have a better and more 

precise picture of WNV risk areas in Europe and neighbouring 

countries in order to implement appropriate 

control measures, especially guidelines for blood donation 

and organ transplants. Also, studies in Europe are 

required to better understand the cycle of transmission 

and the maintenance of WNV in the environment 

over the years to provide appropriate indicators for risk 

assessment. 

References 

1. Papa A, Danis K, Baka A, Bakas A, Dougas G, Lytras T, 

et al. Ongoing outbreak of West Nile virus infections in 

humans in Greece, July – August 2010. Euro Surveill. 



2. Calistri P, Giovannini A, Hubalek Z, Ionescu A, Monaco F, 

Savini G, et al. Epidemiology of West Nile in Europe and in the 

Mediterranean basin. Open Virol J. 2010 Apr 22;4:29-37. 

3. Antoniadis A, Alexiou-Daniel S, Malissiovas N, Doutsos J, 

Polyzoni T, LeDuc JW, et al. Seroepidemiological survey for 

antibodies to arboviruses in Greece. Arch Virol. 1990 [suppl 1]: 

277-285. 

4. Papa A, Perperidou P, Tzouli A, Castilletti C. West Nile virus 

neutralizing antibodies in humans in Greece. Vector Borne and 

Zoonotic Dis. Epub 2010 Aug 25 

5. O’Leary DR, Marfin AA, Montgomery SP, Kipp AM, Lehman 

JA, Biggerstaff BJ et al. The epidemic of West Nile virus 

in the United States, 2002. Vector Borne Zoonotic Dis. 

2004;4(1):61-70. 

6. Centers for Disease Control and Prevention (CDC). Update: 

Investigations of West Nile virus infections in recipients of 

organ transplantation and blood transfusion. MMWR Morb 

Mortal Wkly Rep. 2002 Sep 20;51(37):833-6. 

7. Krisztalovics K, Ferenczi E, Molnár Z, Csohán Á, Bán E, Zöldi 

V, Kaszás K. West Nile virus infections in Hungary, August– 

September 2008. Euro Surveill. 2008;13(45):pii=19030. 



8. Capobianchi M, Sambri V, Castilletti C, Pierro AM, Rossini G, 

Gaibani P et al, on behalf of the Italian Transplant Network. 

Retrospective screening of solid organ donors in Italy, 2009, 

reveals unpredicted circulation of West Nile virus. Euro 

Surveill. 2010;15(34):pii=19648. Available from: http://www. 


9. Aboutaleb N, Beersma MF, Wunderink HF, Vossen AC, Visser LG. 

Case report: West-Nile virus infection in two Dutch travellers 

returning from Israel. Euro Surveill. 2010;15(34):pii=19649. 





West Nile virus circulation in Emilia-Romagna, Italy: 

the integrated surveillance system 2009 

P Angelini (pangelini@regione.emilia-romagna.it) 1 , M Tamba 2 , A C Finarelli 1 , R Bellini 3 , A Albieri 3 , P Bonilauri 2 , F Cavrini 4 , 

M Dottori 2 , P Gaibani 4 , E Martini 5 , A Mattivi 1 , A M Pierro 4 , G Rugna 2 , V Sambri 4 , G Squintani 5 , P Macini 1 

1. Public Health Service, Emilia-Romagna Region, Bologna, Italy 

2. Istituto Zooprofilattico Sperimentale della Lombardia e dell’Emilia-Romagna, Brescia, Italy 

3. Centro Agricoltura Ambiente “G Nicoli”, Crevalcore, Italy 

4. Regional Reference Centre for Microbiological Emergencies (CRREM), Microbiology Unit, Azienda Ospedaliero-Universitaria di 

Bologna, Policlinico S. Orsola–Malpighi, Bologna, Italy 

5. Veterinary and Food Hygiene Service, Emilia-Romagna Region, Bologna, Italy 


Citation style for this article: Angelini P, Tamba M, Finarelli AC, Bellini R, Albieri A, Bonilauri P, Cavrini F, Dottori M, Gaibani P, Martini E, Mattivi A, Pierro 

AM, Rugna G, Sambri V, Squintani G, Macini P. West Nile virus circulation in Emilia-Romagna, Italy: the integrated surveillance system 2009. Euro Surveill. 


Following a large West Nile virus (WNV) epidemic in 

north-eastern Italy in 2008, human and animal surveillance 

activities were implemented in Emilia Romagna. 

Human surveillance was performed by serology or 

genome detection on blood and cerebrospinal fluid for 

all suspected cases suffering from acute meningoencephalitis 

in the regional territory. Animal surveillance 

consisted of passive and active surveillance of horses 

and active surveillance of wild birds and mosquitoes. 

Between 15 June and 31 October 2009, nine of 78 possible 

cases of West Nile neuroinvasive disease were 

confirmed (three fatal). From May to October, 26 cases 

of neurological West Nile disease were confirmed 

among 46 horses. The overall incidence of seroconversion 

among horses in 2009 was 13%. In 2009, 44 of 

1,218 wild birds yielded positive PCR results for WNV 

infection. The planned veterinary and entomological 

surveillance actions detected WNV activity from the 

end of July 2009, about 2-3 weeks before the onset of 

the first human neurological case. Passive surveillance 

of horses seems to be an early and suitable tool for 

the detection of WNV activity, but it will be less sensitive 

in the future, because an intensive programme of 

horse vaccination started in June 2009. 

Regional integrated surveillance system 

In Italy a national veterinary plan for the surveillance 

of West Nile virus (WNV) circulation was set up in 2001 

under the coordination of the National Reference Centre 

for Exotic Diseases of animals (CEntro Studi Malattie 

Esotiche; CESME). 

During the late summer of 2008 a large epidemic of 

WNV infection occurred in north-eastern Italy over an 

area exceeding 7,000 km 2 , in three regions, including 

Emilia-Romagna. Twenty-three horse cases and three 

human cases of the neuroinvasive form of West Nile 

disease (WND) were confirmed by laboratory tests 

[1,2]. After the first evidence of WNV circulation in 

horses was found, additional surveillance on horses, 

birds and mosquitoes was activated. 


This article has been published on 22 April 2010 

In Emilia-Romagna the WNV surveillance plan 2009 

adopted locally the surveillance measures indicated by 

the national plan. In particular, among the surveillance 

activities, the choice was to monitor wild non-migratory 

birds, such as corvids (the crow family), considered 

the most sensitive indicators among birds, which 

can be captured easily. As regards equine surveillance, 

the regional plan emphasised the education of veterinary 

practitioners, focusing on the inclusion of WNV 

in differential diagnosis and the achievement of rapid 

reporting. A major feature of this plan was to establish 

an extremely sensitive system of passive surveillance. 

In addition to passive surveillance, active monitoring 

of horses was implemented in the area involved in the 

2008 outbreak, including Ferrara and the neighbouring 

provinces [3]. 

Evidence of WNV circulation in 2008 was found in 

animals [1,4], humans [5], and mosquitoes. This 

highlighted the need to implement an integrated surveillance 

system which would describe the phenomenon 

comprehensively. Such a system facilitates the 

collection of data to evaluate spatial distribution and 

time trends of viral circulation and shares information. 

For this reason the 2009 Regional Surveillance Plan 

implemented activities beyond those of the National 

Plan, revised the surveillance system of human cases, 

activated intensive entomological monitoring, and 

enlarged the surveillance area to involve all the provinces 

along the Po River. 

Human surveillance 

The aim of the human surveillance system was the 

early detection of infection in humans and the estimation 

of its diffusion through the systematic analysis of 

newly emerging clinical cases, in order to manage specific 

interventions. 

7

The surveillance was performed throughout the 

regional territory, from 15 June to 31 October 2009, 

corresponding to the period of vector activity in Emilia- 

Romagna and adjusted locally according to weather 

conditions and vector activity reports. In 2009, the 

2008 case definition [6] was extended to include cases 

of all ages and not only those over 15 years of age. 

The human surveillance activity was performed 

by serology or genome detection on blood and 

Figure 1 

Map of municipalities with confirmed West Nile 

virus circulation and localisation of human West Nile 

neuroinvasive disease cases by probable infection site, 

Emilia-Romagna, Italy, 2009 

WNV: West Nile virus 

cerebrospinal fluid for all suspected cases suffering 

from acute meningoencephalitis in the regional territory. 

Active surveillance of people who live or work 

in areas of documented viral circulation was also performed. 

In addition blood and cerebrospinal fluid samples 

from subjects living or staying for at least one 

night in the regional area were sent to the Regional 

Reference Centre for Microbiological Emergencies 

(Centro di Riferimento Regionale per le Emergenze 

Microbiologiche; CRREM). In selected cases, positive 

specimens were confirmed by the National Health 

Institute (Istituto Superiore di Sanità; ISS) and by 

the National Institute for Infectious Diseases (Istituto 

Nazionale Malattie Infettive; INMI). 

From October 2008 to April 2009 a serosurvey performed 

on 9,177 healthy blood donors living in the 

province of Ferrara detected a total of 62 IgG positive 

subjects, corresponding to a seroprevalence of 0.68%. 

Animal surveillance 

The regional veterinary WNV surveillance system was 

activated from May to October, performing passive and 

active surveillance on horses and on non-migratory 

wild birds. 

Horse passive surveillance 

In Italy all suspected signs of WND in horses must be 

notified to the official veterinary services. Suspected 

cases were confirmed if resulted positive by reverse 

transcription – polymerase chain reaction (RT-PCR) performed 

on central nervous system [7] or to a WNV virus 

Figure 2 

Distribution of West Nile virus confirmed cases (mosquito pools, birds, horses, and human West Nile neuroinvasive 

disease) by date, Emilia-Romagna, Italy, July-October 2009 

Number of cases 

18 

16 

14 

12 

10 

8 

6 

4 

2 

0 

15/7-31/7 1/8-15/8 16/8-31/8 1/9-15/9 16/9-30/9 1/10-15/10 16/10-31/10 

Date 2009 

WNND: West Nile neuroinvasive disease 

Note: The figure does not include a magpie (found in early May) or a jay (found in early November). 

Mosquito pools 

8 www.eurosurveillance.org 

Birds 

Horses 

Human WNND

neutralisation (VN) test (cut-off titre 1:10) in microtitre 

plates and to IgM enzyme linked immunosorbent assay 

(ELISA) [8,9]. 

Horse active surveillance 

In the provinces of Ferrara, Bologna, Modena, Ravenna, 

and Reggio Emilia, every 1,600 km 2 , 28 seronegative 

unvaccinated equine sentinels, sufficient to detect an 

incidence above 10% (CI 95%), were selected in the 

spring of 2009. They were serologically tested twice 

after the selection, at the beginning of August and 

the beginning of September. Samples collected were 

screened by a home-made competitive ELISA [10]. 

Positive samples were confirmed by VN and IgM ELISA 

at the CESME in Teramo. A seroconversion was confirmed 

if VN titre was at least 1:10 and there was evidence 

of IgM antibodies. 

Wild bird surveillance 

Monitoring was carried out in all the provinces along 

the Po River, in the plain area of Emilia-Romagna. 

Every 1,600 km 2 , a monthly sample of about 40 wild 

birds caught or shot within specific wildlife population 

control programmes was collected. Samples of organs 

(brain, heart, and kidney) of each bird were pooled and 

examined by RT-PCR [7]. 

Entomological surveillance 

The surveillance system was based on the weekly to 

monthly (frequency depends on local resources) collection 

of mosquitoes in fixed stations and in the sites 

where birds, humans, or horses signalled WNV activity. 

Mosquito collections for WNV screening were conducted 

in six provinces: Ferrara, Ravenna, Bologna, 

Modena, Reggio Emilia, and Parma, using 92 CO 2 

baited traps positioned in fixed stations. Moreover, 

mosquito collections were performed promptly using 

CO 2 and gravid traps in sites where positive horses and 

human cases had been detected. 

The surveillance system was activated in the period 15 

April to 10 October. Collected mosquitoes were pooled 


(maximum 200) by species, date, and site of collection 

and examined by RT-PCR [7]. In addition, overwintered 

mosquito females were collected during the period 3 

March to 8 April by manual aspirator in rural buildings 

in the area where WNV was active in 2008. 

Virological analysis 

Human samples 

The detection of WNV genome in human plasma, cerebrospinal 

fluid, and serum samples obtained from 

patient suffering from clinical symptoms of meningoecephalitis 

was performed by an RT-PCR assay based on 

specific TaqMan probes [7]. 

Animal samples 

In horses RNA was extracted starting from 200 µl of 

serum or whole blood with EDTA as anticoagulant. In 

birds RNA was extracted from 200 µl of phosphate 

buffer saline homogenate (about 20% tissue g / buffer 

ml) of pooled brain, heart and kidney of each analysed 

bird. In mosquitoes RNA was extracted from 200 µl of 

a maximum of 200 pooled mosquitoes manual grinded 

by using copper stained beads, in 500-800 µl of PBS. 

cDNA was submitted to RT-PCR according to the method 

of Tang and colleagues [7]. Positive samples were confirmed 

by sequencing of partial nucleocapsin and premembrane 

protein M amplified according to [11]. Finally 

from each RT-PCR positive sample WNV was isolated 

on Vero and RK13 cell lines. 

Results 

Results of the integrated surveillance system are 

mapped in figure 1, with the sequence of events summarised 

in figure 2 (July-October), and discussed 

below. 

Human cases 

As of 31 October 2009, nine out of 78 possible cases 

of West Nile neuroinvasive disease (WNND) notified 

in Emilia-Romagna have been confirmed (8/9 males; 

median age 72 years, range: 62-78). Three patients 

Table 1 

Species distribution of wild birds tested for West Nile virus (n=1,218), Emilia-Romagna, Italy, May-October 2009 

Species Birds tested WNV RT-PCR positive % WNV positive 

European magpie (Pica pica) 607 27 4.4 

Carrion crow (Corvus corone cornix) 350 5 1.4 

European starling (Sturnus vulgaris) 98 5 5 

Eurasian jay (Garrulus glandarius) 96 2 2 

Common blackbird (Turdus merula) 30 0 - 

Strigiformes 11 2 18 

Charadriiformes 8 3 38 

Other Passeriformes 7 0 - 

Other bird orders 5 0 - 

Piciformes 4 0 - 

Columbiformes 2 0 - 

Total 1,218 44 3.6 

RT-PCR: reverse transcription-polymerase chain reaction; WNV: West Nile virus 

9

Table 2 

Species of mosquitoes tested for West Nile virus (n=190,516), Emilia-Romagna, Italy, May-October 2009 

Province Bologna Forlì Ferrara Modena Piacenza Parma Ravenna Reggio Emilia Total 

Mos- 

Species 

Pool 

quito 

Pool Mos- 

Pool 

+ quito 

Pool Mos- 

Pool 

+ quito 

Pool Mos- 

Pool 

+ quito 

Pool Mos- 

Pool 

+ quito 

Pool Mos- 

Pool 

+ quito 

Pool Mos- 

Pool 

+ quito 

Pool Mos- Pool Mos- 

Pool Pool 

+ quito + quito 

Pool 

+ 

Ae. albop- 

169 13 11 4 392 31 227 23 58 11 228 6 142 20 1,227 108 0 

ictus 

Ae. caspius 1,713 51 14 5 9,953 114 16,915 121 2 1 12 2 606 11 68 9 29,283 314 0 

Ae. detritus 1 1 4 1 5 2 0 

Ae. dorsalis 13 1 13 1 0 

Ae. geniculatus 6 2 2 1 8 3 0 

Ae.vexans 2 1 11 2 84 3 4,090 41 1 1 363 3 46 9 4,597 60 0 

An. maculip- 

4 1 59 3 14 5 1 1 4 4 82 14 0 

ennis 

An. plumbeus 2 2 2 2 0 

Cx. modes- 

7 3 114 10 117 12 8 1 246 26 0 

tus 

Cx. pipiens 84,225 645 9 158 17 52,973 396 5 6,664 78 7 331 13 6,926 50 1 911 10 2,865 50 5 155,053 1,259 27 

Total 86,120 714 9 194 28 0 63,575 557 5 28,048 285 7 393 27 0 7,530 62 1 1,517 21 0 3,139 95 5 190,516 1,789 27 

died, two living in Ferrara Province and one in Modena. 

In addition, not reported in figures, the local health 

units of Parma and Modena notified two other confirmed 

cases, both 72 year-old women resident in 

Mantua province (Lombardy region), treated in hospital 

in Emilia-Romagna. Another case of infection was that 

of a 78 year-old female liver donor. Before her death, 

she had spent two weeks visiting relatives in the WNV 

affected area (Reggio Emilia). 

Horse cases 

Passive surveillance 

From May to October, 26 cases of neurological WND 

were confirmed among 46 horses in which it was suspected. 

Four of the eight provinces involved in the 

regional surveillance system had WND cases in horses. 

The first symptoms in horses were detected in the second 

half of July in the provinces of Ferrara and Reggio 

Emilia, but the most cases were notified between mid- 

August and mid-September (figure 2). 

Active surveillance 

Seroconversions in sentinel horses were detected in 

three provinces (Ferrara, Modena, and Reggio Emilia). 

Early seroconversions were registered among the controls 

examined at the beginning of August. Serological 

controls around the stables with WND cases also confirmed 

recent WNV infections in the province of Parma. 

The overall incidence of seroconversion among horses 

in 2009 was 13% (95% CI: 10% to 16%), but in Ferrara it 

was 28% (95% CI: 19% to 39%). 

Wild birds 

Six of the eight provinces that took part in the regional 

surveillance system reported positive birds. In 2009, 

44 wild birds out of 1,218 (tested by PCR) yielded 

positive results for WNV infection. With the exception 

of a magpie caught in May, positive wild birds were 

detected from the end of July (figure 2). Most of the 

infected wild birds were corvids (Pica pica, Corvus corone 

cornix, Garrulus glandarius), collected within population 

control programmes in August and September, 

but WNV was detected also in other species, mainly 

found dead in wildlife recovery centres (table 1). 

Table 1. Species distribution of wild birds tested for 

West Nile V (n=1,218), Emilia-Romagna, Italy, May- 

October 2009 

Mosquitoes 

About 190,000 mosquitoes were collected, pooled and 

tested using PCR (1,789 pools of ≤200 individuals/ 

pool). Culex pipiens were the most abundant species 

(81.4%) followed by Aedes caspius (15.4%) and Aedes 

vexans (2.4%). Other collected species are shown in 

table 2. 

Twenty-seven pools, all consisting of Culex pipiens, 

yielded positive results for WNV. Early positive pools 

were collected in the province of Reggio Emilia at 

the end of July. Minimum infection rates (MIR: (no. of 


positive pools/no. of mosquitoes tested) x 1,000) [12] 

were calculated, with higher MIR values recorded in 

August in the provinces of Reggio Emilia (3.08) and 

Modena (1.44). 

Referring to the collection of overwintering mosquitoes, 

three mosquito species were collected: Culex pipiens 

(516 females, 52%), Anopheles maculipennis s.l. 

(475 females, 48%), Culiseta annulata (four females, 


Phylogenetic analysis in a recent controlled outbreak 

of Crimean-Congo haemorrhagic fever in the south of 

Iran, December 2008 

S Chinikar (chinikar@pasteur.ac.ir) 1 , S Mojtaba Ghiasi 1 , M Moradi 1 , M M Goya 2 , M Reza Shirzadi 2 , M Zeinali 2 , E Mostafavi 1 , 

M Pourahmad 3 , A Haeri 3,4 

1. National Reference Laboratory for Arboviruses and Viral Haemorrhagic Fevers, Pasteur Institute of Iran, Tehran, Iran 

2. Centre for Disease Control (CDC), Ministry of Health of Iran, Tehran, Iran 

3. Jahrom Faculty of Medicine 

4. Shahid Beheshti University of Medical Sciences, Tehran, Iran 


Chinikar S, Mojtaba Ghiasi S, Moradi M, Goya MM, Reza Shirzadi M, Zeinali M, Mostafavi E, Pourahmad M, Haeri A. Phylogenetic analysis in a recent controlled 

outbreak of Crimean-Congo haemorrhagic fever in the south of Iran, December 2008. Euro Surveill. 2010;15(47):pii=19720. Available online: http://www. 


Crimean-Congo haemorrhagic fever (CCHF) is a viral 

zoonotic disease with a high mortality rate in humans. 

The CCHF virus is transmitted to humans through the 

bite of Ixodid ticks or contact with blood or tissues 

of CCHF patients or infected livestock. In December 

2008, a re-emerging outbreak of CCHF occurred in the 

southern part of Iran. Five people were hospitalised 

with sudden fever and haemorrhaging, and CCHF was 

confirmed by RT-PCR and serological assays. One of 

the cases had a fulminant course and died. Livestock 

was identified as the source of infection; all animals in 

the incriminated herd were serologically analysed and 

more than half of them were positive for CCHFV. We 

demonstrated that two routes of transmission played 

a role in this outbreak: contact with tissue and blood 

of infected livestock, and nosocomial transmission. 

Phylogenetic analyses helped to identify the origin of 

this transmission. This outbreak should be considered 

as a warning for the national CCHF surveillance system 

to avoid further outbreaks through robust prevention 

and control programmes. 

Introduction 

Crimean-Congo haemorrhagic fever (CCHF) is a viral 

zoonotic haemorrhagic fever with up to 13-50% mortality 

rate in humans. Infected animals are unsymptomatic. 

The disease is caused by Crimean-Congo 

haemorrhagic fever virus (CCHFV) that belongs to the 

family Bunyaviridae, genus Nairovirus. The of negative 

single-stranded RNA genome consists of three segments, 

large (L), medium (M) and small (S), coding for 

the viral polymerase (L), the envelope glycoproteins (M) 

and the viral nucleoprotein (S) [1-5]. The typical course 

of CCHF progresses through four distinct phases: incubation, 

pre-haemorrhagic phase, haemorrhagic phase 

and convalescence [6-8]. After a incubation period of 

one to three days, the patient has sudden onset of 

fever, myalgia, nausea and severe headache. Within 

three to six days of the onset of illness, a petechial 

rash and haemorrhagic symptoms such as epistaxis, 

Article published on 25 November 2010 

haematemesis, and melaena may occur. The most 

severely ill patients develop multiorgan failure characterised 

by shock, haemorrhaging and coma [9-11]. 

The virus is transmitted to humans through the bite of 

Ixodid ticks or by contact with blood or tissues from 

infected livestock [12-14]. In addition to zoonotic transmission, 

CCHFV can be spread from person to person 

and is one of the rare haemorrhagic fever viruses able 

to cause nosocomial outbreaks in hospitals [15-20]. 

In the period from 1 January 2000 to 12 September 

2010, 738 confirmed cases of CCHF and 108 associated 

fatalities were notified in Iran [15,21]. The province 

reporting most infections was Sistan-va-Baluchistan, 

Isfahan and Fars (Figure 1). 

Figure 1 

Geographical distribution of Crimean-Congo 

haemorrhagic fever in Iran, 1 January 2000-12 September 

2010 (n=738) 


Table 

History and laboratory results of probable cases of Crimean-Congo haemorrhagic fever, Fars province, Iran, November–December 2008 (n=5) 

Clinical and paraclinical signs Ig M Ig G 

Date of sampling 

RT-PCR 


Second sample 

First sample 

Second sample 

First sample 

Proteinuria 

Thrombocytopenia 

Leukocytopenia 

Haemorrhage 

Petechia 

Fever 

Death 

Second sample 

First sample 

Date of fever 

Contact with suspected patient 

Contact with livestock 

Age (years)/sex 

Profession 

Patient 

Aa Housewife 46/F Yes No 18 Dec 2008 21 Dec 2008 Not taken Yes Yes No Yes Yes Yes No report Negative Not taken Negative Not taken Positive 

B Butcher 21/M Yes No 18 Dec 2008 20 Dec 2008 25 Dec 2008 No Yes No Yes Yes Yes Yes Positive Positive Negative Negative Negative 

C Butcher 24/M Yes Yes 19 Dec 2008 20 Dec 2008 25 Dec 2008 No Yes No Yes No Yes Yes Negative Positive Negative Positive Positive 

D Self-employed 28/M Yes No 21 Dec 2008 22 Dec 2008 Not taken No Yes No No Yes Yes Yes Positive Not taken Negative Not taken Negative 

E Nurse 26/F No Yes 27 Dec 2008 30 Dec 2008 Not taken No Yes Yes Yes No Yes No Positive Not taken Negative Not taken Positive 

F: female; M: male. 

a Index case. 

Outbreak description 

Here, we report a CCHF outbreak in Fars province, 

Iran, caused by contact of humans with blood or tissues 

of infected livestock, with additional nosocomial 

transmission. In total, five patients (A-E) were admitted 

to the regional hospital with similar presentations 

of a haemorrhagic condition in the period from 18 

December to 21 December 2008. This period coincides 

with the Muslim ceremony Eid-al-Adha (the ceremony 

of sacrificing livestock) which is celebrated in Islamic 

countries on 9 December. 

Patients A and D (who are part of the same family) 

bought a calf from a butchery run by two brothers, 

patients B and C, and hired them to sacrifice the animal. 

On the morning of 18 December 2008, Patient A, 

the index case of this outbreak, was admitted to hospital 

and died after a fulminant course of CCHF. In the 

evening of the same day, Patients B and C were admitted 

to the same hospital with fever and chill, severe 

headache, dizziness, photophobia. Patient D developed 

similar clinical signs on 21 December and was 

hospitalised. Nine days after the index case, the nurse 

caring of these four patients was also hospitalised 

with haemorrhagic symptoms (Patient E). 

Materials and methods 

Case definition 

The case definition for probable cases included patients 

admitted between 18 December and 27 December 2008 

in the regional hospital and presenting with a clinical 

picture compatible with CCHF, or contact with tissues 

or blood from a possibly infected animal, or a healthcare 

worker with a history of contact with a CCHF case. 

Probable cases with positive IgM serology and/or positive 

RT-PCR were considered as CCHF confirmed cases. 

Laboratory analysis 

Human and animal sera were analysed by ELISA for 

anti-CCHFV IgM and IgG as described [15,22]. Viral RNA 

was extracted from patient’s serum using QIAamp RNA 

Mini kit (QIAgen GmbH, Hilden, Germany) and analysed 

by gel-based and real-time RT-PCR with a one-step 

RT-PCR kit (QIAgen GmbH, Hilden, Germany). A 536 bp 

fragment of the S segment of the CCHFV genome was 

amplified [4,8,12] and sequenced. 

Phylogenetic analysis was performed with the neighbour-joining 

method based on Kimura two-parameter 

distances by using Mega 4 software. Bootstrap confidence 

limits were based on 500 replicates. Evolutionary 

divergence, distance matrix and subsequently sets of 

phylogenetic trees were calculated by the software 

[23]. 

Results and discussion 

As summarised in the Table, five probable CCHF cases 

in this outbreak were confirmed by serological and 

molecular methods. It is worth mentioning that no 

immunological response was detected in the fatal 

case that had a fulminant course, Patient A, and CCHF 

13

in this case was only confirmed by a strongly positive 

RT-PCR. There is evidence of other fatal cases lacking 

an immune response to CCHFV [6,24]. Patients C and E 

were positive both for viral RNA and antibodies against 

CCHFV, while Patients B and D were negative in the PCR 

and only confirmed by serological assay. Notably, ribavirin 

was administered to the patients in hospital. 

At the same time, serum samples were collected from 

50 animals in the herd from which the calf had been 

bought. In 30 of these samples antibodies to CCHFV 

were detected. Although CCHF is an asymptomatic disease 

in livestock that does not kill the animals, seroepidemiological 

surveys of animal populations in endemic 

areas and high risk regions could be useful in that they 

may complement the national surveillance system and 

serve as an early warning of CCHF in the area. 

In this outbreak, it was demonstrated that the main 

transmission route of CCHF was through handling 

blood or tissues of infected livestock (for patients 

A, B, C and D), while patient E had had no contact to 

livestock and was infected nosocomially. It is unclear 

why Patient A had such a fulminant course of disease 

and died. Patients B, C and D were infected through the 

same route, by direct contact with tissue and blood of 

the same animal, but had a milder course of disease 

and recovered. It is important in infectious disease outbreaks 

to investigate what factors determine the severity 

of the disease in different individuals [6]. There are 

published reports on the influence of cytokine levels 

on the immune response to CCHFV in different patients 

[24,25]. It has been shown that patients infected with 

a higher dose of virus develop more severe disease 

symptoms and outcomes [26-28]. Although we did not 

use quantitative RT-PCR, the band density of the PCR 

product obtained from patient A was much higher than 

that of the other patients. On the other hand, it seems 

likely that patients B and C presented a mild form of the 

disease because they may already have had antibodies 

against CCHFV due to their professional exposure. 

Moreover, no anti-CCHFV IgG antibody response was 

detected for the patient B, whereas a normal serological 

and molecular pattern was seen in patient C, which 

Figure 2 

Phylogenetic comparison of Crimean-Congo haemorrhagic fever virus isolates from Iran with isolates form patients in the 

recent outbreak in Fars province, Iran, December 2008 

A 

B 

Divergence 

1 2 3 4 

Percent identity 

5 6 7 8 9 

1 99.9 99.9 99.9 99.9 99.8 97.6 97.6 97.7 1 AY366373(partial) 

2 0.2 100.0 99.9 99.9 99.9 97.6 97.6 97.7 2 AY366379(partial) 

3 0.2 0.0 99.9 99.9 99.9 97.6 97.6 97.7 3 AY366378(partial) 

4 0.2 0.4 0.4 99.8 99.8 97.5 97.6 97.6 4 AY366375(partial) 

5 0.4 0.2 0.2 0.6 99.9 97.6 97.6 97.7 5 AY366374(partial) 

6 0.6 0.4 0.4 0.8 0.2 97.5 97.5 97.6 6 AY366376(partial) 

7 1.0 0.8 0.8 1.2 1.0 1.2 99.9 99.8 7 Patient A 

8 1.0 0.8 0.8 1.3 1.0 1.3 0.2 99.9 8 Patient C 

9 0.8 0.6 0.6 1.0 0.8 1.0 0.4 0.2 9 Patient E 

1 2 3 4 5 6 7 8 9 

A. The phylogeny tree of nucleotide sequences spanning described regions of the S-segment of CCHF virus genome which is detected in the 

outbreak. B. Nucleotide identity and divergence of CCHF virus genomes isolated from patients of the outbreak. 


might be interpreted to indicate that patient B was 

infected with a very low viral dose. Recent investigations 

have concluded a relationship linking the severity 

and outcome of CCHFV infections with the strength 

of the host immune response and the initial viral load 

[24,25,27]. 

Phylogenetic analysis of alignments of three partial 

genomic sequences (536 bp) of the CCHFV S segment 

indicates that the viruses isolated from patients A, C 

and E can be differentiated into two distinct branches, 

with a slightly lower identity between patients A and 

E. As illustrated in Figure 2, the sequences obtained 

in this outbreak are not clustered with other CCHFV 

sequences isolated in Iran (about 97.5% identity). It 

is possible that a new strain occurred in the outbreak 

region, and further phylogenetic analyses are required 

to identify the precise origin of this genetic variant. 

However, comparison of the isolates from our patients 

with isolates from other areas may give some indications 

as to the origin of this outbreak [12]. 

One of the factors that contributed to the control of 

this outbreak was the well-coordinated and efficient 

surveillance system for CCHF and other viral haemorrhagic 

fevers that is in place in Iran. The system is not 

only responsible for continuous monitoring of these 

diseases but also deals with outbreaks. Rapid and precise 

laboratory diagnosis of CCHF allowed controlling 

this outbreak. Nevertheless, a higher level of training 

and precautionary measures for healthcare workers 

(such as use of isolation chambers in hospital wards, 

mask and other medical shields during contact to CCHF 

patients) and other high risk professions could help 

to decrease the outbreak rate in the endemic areas. 

In conclusion, with Iran being an endemic country for 

CCHF in the Middle East and neighbouring Turkey an 

endemic country in Europe, efficient surveillance and 

control programmes on CCHF in Iran could prove beneficial 

also for the European region. 

Acknowledgements 

We would like to thank the other members of the National 

Reference Laboratory for Arboviruses and Viral Haemorrhagic 

Fevers at Pasteur Institute of Iran for their technical support. 

This study was performed funded by the budget of 

the National Reference Laboratory for Arboviruses and Viral 

Haemorrhagic Fevers at Pasteur Institute of Iran. 

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Virol. 2006;36(4):272-6. 

25. Weber F, Mirazimi A. Interferon and cytokine responses 

to Crimean-Congo hemorrhagic fever virus; an emerging 

and neglected viral zoonosis. Cytokine Growth Factor Rev. 

2008;19(5-6):395-404. 

26. Duh D, Saksida A, Petrovec M, Ahmeti S, Dedushaj I, Panning 

M, et al. Viral load as predictor of Crimean-Congo hemorrhagic 

fever outcome. Emerg Infect Dis. 2007;13(11):1769-72. 

27. Papa A, Drosten C, Bino S, Papadimitriou E, Panning M, Velo E, 

et al. Viral load and Crimean-Congo hemorrhagic fever. Emerg 

Infect Dis. 2007;13(5): 805-6. 

28. Wölfel R, Paweska JT, Petersen N, Grobbelaar AA, Leman PA, 

Hewson R, et al. Virus detection and monitoring of viral load in 

Crimean-Congo hemorrhagic fever virus patients. Emerg Infect 

Dis. 2007;13(7):1097-100. 

15

Review articles 

West Nile virus in Europe: understanding the present to 

gauge the future 

P Reiter (paul.reiter@pasteur.fr) 1 

1. Insects and Infectious Disease Unit, Institut Pasteur, Paris, France 


Citation style for this article: Reiter P. West Nile virus in Europe: understanding the present to gauge the future. Euro Surveill. 2010;15(10):pii=19508. Available 

online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19508 

The appearance of West Nile virus in New York in 1999 

and the unprecedented panzootic that followed, have 

stimulated a major research effort in the western hemisphere 

and a new interest in the presence of this virus 

in the Old World. This review considers current understanding 

of the natural history of this pathogen, with 

particular regard to transmission in Europe. 

Background 

West Nile virus (WNV) is by far the most widely distributed 

arbovirus. It belongs to the Japanese encephalitis 

antigenic complex of the family Flaviviridae, transmitted 

in an avian cycle by ornithophilic mosquitoes, 

chiefly of the genus Culex [1]. Mammals can also be 

infected, but are considered dead end hosts because 

viraemia is generally too low to infect mosquitoes [2]. 

Mosquitoes acquire infection by feeding on a viraemic 

host. Virus passes through the gut wall into the 

haemolymph, the ‘blood’ of the insect, after which replication 

occurs in most of the internal tissues. When 

the salivary glands are infected, the virus can pass to a 

new host via saliva injected into the skin by the insect 

when it takes a blood meal. The period from the infective 

blood meal to infectivity, the extrinsic incubation 

period, lasts 10-14 days depending on temperature. 

Ornithophilic vectors that also bite and infect mammals, 

including humans, are termed bridge vectors. 

Human infections attributable to WNV have been 

reported in many countries in the Old World for more 

than 50 years [3-5]. In recent years these have included 

Algeria 1994 (eight deaths) [6], Romania 1996-2000 

(21 deaths) [7], Tunisia 1997 (eight deaths) [8], Russia 

1999 (40 deaths) [9], Israel 2000 (42 deaths) [10], and 

Sudan 2004 (four deaths) [11]. By far the largest outbreaks 

occurred in Bucharest in 1996 (393 hospitalised 

cases, 17 deaths) and Volgograd in 1999 (826 hospitalised 

cases, 40 deaths). Both occurred in urban areas 

and were associated with cellars flooded with sewagepolluted 

water in poorly maintained apartment blocks, 

a highly productive breeding site for an effective vector, 

Culex pipiens [7,9,12]. Outbreaks on this scale have 

also occurred in Israel [13]. All three sites are on major 

migratory routes of birds that overwinter in Africa. 


In its original range, WNV is enzootic throughout Africa, 

parts of Europe, Asia and Australia, but it received little 

attention until 1999, when a topotype circulating 

in Tunisia and Israel appeared in the Bronx, New York 

[14,15], probably imported in a live bird. The epizootic 

that followed was spectacular and unprecedented: 

within five years, the virus appeared ubiquitous, sometimes 

common, in nearly all counties of every state east 

of the Rocky Mountains, as well as parts of western 

Nevada and southern California. Sizeable outbreaks 

were also observed in six Canadian provinces. It is now 

widely established from Canada to Venezuela. To date 

(1999-2009), 29,606 clinical cases and 1,423 deaths 

have been reported in humans, and more than 27,000 

cases in horses, with a case fatality rate of about 33% 

[16]. Two thirds of the horse population in the United 

States are now vaccinated, but no vaccine is available 

for humans. 

The virus 

Two lineages of WNV are widely recognised that are 

about 30% divergent [14]. Lineage I includes WNV 

strains from Africa, the Middle East, Europe, India, 

Australia (formerly Kunjin virus) and the Americas. 

The close relationship between isolates from Kenya, 

Romania and Senegal are evidence of the geographic 

mobility of the virus in migratory birds [17]. The virus 

isolated in the Bronx, New York in 1999 was closely 

related to Lineage I strains circulating in Israel and 

Tunisia a year earlier [18] and most probably imported 

in a wild bird. Until recently, all isolates of Lineage II 

were from Sub-Saharan Africa and Madagascar, but in 

2004, it was isolated from a goshawk in Hungary, and 

from several birds of prey in 2005 [19]. 

At least five new lineages have been proposed for 

strains isolated in central Europe, Russia and India [20- 

23]. This is not surprising, given that the original range 

of the virus spans Europe, Africa, Asia and Australia, 

the increasing accessibility of sequencing technology, 

and the enormous interest in the virus since its 

appearance in North America. Lastly, a new genotype 

was identified in the US in 2003 and may now be the 

dominant strain in North America [24,25]. 


Pathology 

Only a small portion of human infections are symptomatic, 

with the headache, tiredness, body aches and 

swollen lymph glands typical of many febrile diseases. 

Occasionally there is an abdominal rash. About one 

in 150 patients develop one or multiple indicators of 

neuro-invasive disease; neck stiffness, stupor, disorientation, 

coma, tremors, convulsions, muscle weakness, 

and paralysis. This can occur in people of any age, 

but those over 50 years are at highest risk [26]. In the 

past five years, 4.8% of laboratory-confirmed clinical 

infections reported in the US were fatal. Symptomatic 

infections in horses are also rare and generally mild, 

but can cause neurologic disease including fatal 

encephalomyelitis [27]. 

In the Old World, mortality in birds associated with 

WNV infection is rare [28], although significant numbers 

of storks and domestic geese died during epizootics 

in Israel [29]. In striking contrast, the virus is highly 

pathogenic for New World birds; the appearance of 

large numbers of dead or dying birds is often an indicator 

of local transmission [30]. In the early days following 

appearance of the virus in the US, it appeared that 

members of the crow family (Corvidae) were particularly 

susceptible, but virus has been detected in dead 

and dying birds of more than 250 species – with viraemia 

as high as 109 pfu/ml – as well as various species 

of mammals and even in alligators [1]. In rural Europe, 

in the absence of large-scale bird mortality, neurologic 

symptoms in horses are often the sole indication of 

local presence of the virus. 

Vectors 

Mosquitoes of the genus Culex are generally considered 

the principal vectors of WNV, both in the Old World and 

in the Americas. Studies with bird-baited traps in the 

wetlands of Mediterranean Europe indicate four such 

species are dominant. For example, in a region of the 

Danube delta that has enormous populations of resident 

and migrant birds, 82% of mosquito captures in 

2008 (>10,000 mosquitoes, 17 species) were of three 

species: Cx. pipiens (44%), Cx. torrentium (27%) and 

Cx. modestus (11%). Coquillettidia richardii (14%) and 

Anopheles maculipennis (3%) made up all but 1% of the 

remaining species (F-L Prioteasa, personal communication). 

In contrast, Cx. modestus and C. richardii were 

the dominant species captured on humans in the same 

area (35% and 34%, respectively), while Cx. pipiens 

was one of five species that contributed less than 2% 

of the catch. On the other hand, 93% of all mosquitoes 

captured by bird-baited traps in a village were Cx. pipiens, 

5% were Cx. torrentium, and neither Cx. modestus 

nor C. richardii were present. Similarly, in many urban 

areas, Cx. pipiens is the dominant species, and bloodmeal 

analysis confirms that it is highly ornithophilic 

[31]. These data illustrate the complex relationship 

between abundance, species composition, host preference 

and vector competence. It has been suggested 

that a decline in bird populations in the autumn migration 

season augments the incidence of mammal-biting, 


but this is not borne out by field studies in Chicago, 

Illinois, an area of intense transmission [32]. 

In a study in the Danube delta study WNV was indicated 

by RAMP kit (Response BiomedicaL Corporation, 

Canada; a commercial kit based on WNV-specific antibodies 

with high specificity and sensitivity, [33,34]) 

in 14 pools of mosquitoes: 11 of Cx. pipiens, two of 

Cx. torrentium and one of An. maculipennis (F-L 

Prioteasa, personal communication). In a laboratory 

study of mosquitoes from the Rhone delta, France, 

infection and transmission rates were 89.2% and 

54.5%, respectively, for Cx. modestus, and 38.5% and 

15.8%, respectively, for Cx. pipiens [35,36]. Coupled 

with this high potential as a vector, Cx. modestus is 

abundant in reed-beds that are very probably an important 

ecotope for WNV transmission. 

In New York following the appearance of the virus in 

the US, WNV RNA was detected in three pools of overwintering 

Cx. pipiens, and virus was isolated from 

one of these [37]. In the Czech Republic, virus was 

detected in overwintering Cx. pipiens by PCR, but not 

confirmed by isolation (Z Hubalek and Iwo Rolf, personal 

communication), and in the Danube delta region, 

four pools of Cx. pipiens and one of An. maculipennis 

tested positive by RAMP (F-L Prioteasa, personal communication). 

These results, although not confirmed by 

virus isolation, are particularly interesting because a 

field study of Culex species in Massachusetts, US, confirmed 

that females do not feed on blood before overwintering 

(P. Reiter, unpublished data). This implies 

that these insects acquire their infection by vertical 

transmission between generations via the egg stage. 

Moreover, in the spring of the year of the study, a few 

days after mosquitoes had exited their overwintering 

sites, a number of WNV-positive crows were collected 

in a neighbouring states, circumstantial evidence that 

infected overwintering females had transmitted virus 

to these birds in their first (post-winter) blood meal. 

Lastly, WNV has been isolated from male Cx. pipiens in 

Connecticut, US, further evidence of vertical transmission 

[38], and from larvae of Cx. univittatus s.l. in the 

Rift Valley, Kenya [39]. 

Transmission between vertebrates 

In a landmark study, 25 bird species representing 

17 families and 10 orders were exposed to WNV by 

infectious mosquito bite. Only four of 87 individuals 

did not develop a detectable viraemia [40]. The most 

competent species, judged by magnitude and duration 

of viraemia, were passerines (perching birds, 11 

species, including members of the crow family) and 

charadriiformes (a wader and a gull). In surviving 

birds, the infection persisted in certain organs in 16 

of 41 infected birds until euthanised on day 14 after 

infection. In addition, five of 15 species (representing 

11 families) became infected when virus was placed in 

the back of the oral cavity (either in suspension or as 

a single infected mosquito) and crows were infected 

17

when fed a dead infected sparrow. Furthermore, virus 

was observed in the faeces of 17 of 24 species and in 

the oral cavity of 12 of 14 species for up to 10 days after 

infection. Moreover, contact transmission between 

cage mates was observed in four species. In summary, 

birds can be infected by a variety of routes other than 

mosquito bites, and different species may have different 

potential for maintaining the transmission cycle. 

In the light of this complexity, the spectacular conquest 

of the New World by WNV demands attention. 

Mosquito-borne transmission involves both the extrinsic 

and intrinsic incubation periods; even at high ambient 

temperatures this takes a minimum of 10-14 days, 

so it is hard to imagine that the virus could have traversed 

an entire continent in a period of four or five 

summers by this mechanism alone. Importation by 

infected migrant birds returning from their overwintering 

grounds could explain the distribution. Indeed, a 

new region of transmission, separate from the northern 

states, did appear in Florida and adjoining states two 

years after the initial New York infestation, presumably 

introduced by infected migrant birds, but thereafter the 

virus progressed rapidly westward along a broad front 

stretching from Canada to the Gulf of Mexico [41]. By 

2003, by far the majority of counties east of the Rocky 

Mountains had reported confirmed WNV-positive dying 

birds. 

An alternative explanation for dispersal rests on oral 

infection: crows are scavengers and feed on carrion, 

including dead crows. They are social birds, roost in 

large crowded colonies, have a wide daily dispersal 

range of up to 20 km in all directions, and their feeding 

grounds overlap with crows from other roosts. They 

also exhibit “kin-based cooperative breeding” in which 

grown offspring remain with their parents to rear new 

young [42]. It is conceivable that: (i) crows that die away 

from their roost relay virus by oral infection to birds 

from neighbouring roosts; (ii) faecal-oral transmission 

is significant in crowded roosts, (iii) crows feeding on 

carcasses of other infected species/animals introduce 

the infection to others in their roosts, and (iv) viraemic 

adult and juvenile birds infect nestlings per os. In 

this way, bird-to-bird transmission, particularly among 

social birds, could be a major, even the principal driver 

of amplification and dispersal, with mosquito-borne 

transmission active at the local level. Modelling studies 

give some support for this hypothesis [43]. 

There is also good evidence that oral and faecal-oral 

infection may be important in transmission dynamics 

in other species. In the New World, mortality in many 

species of raptors is out of all proportion to their abundance 

in nature [44-46]. Fatal infections in Imperial 

Eagles in Spain and goshawks in Hungary [47], high 

seroprevalence in kestrels in Egypt [48], and high mortality 

in flocks of domestic geese in Israel [49] and 

Hungary [47] point to the same mechanism. 

Oral infection is not limited to consumption of dead 

or dying birds. For example, adult hamsters are readily 

infected by ingestion of infected material as well as 

by mosquito bite [50]. In these animals, virus is rapidly 

cleared from the blood, but can survive in the central 

nervous system for at least 86 days [51]. Moreover, as 

a chronic renal infection, virus is excreted in the urine 

for at least eight months [52]. Thus, even if viraemia 

in mammals is insufficient to infect mosquitoes, it may 

still contribute to infection of scavengers and raptors. 

Circumstantial evidence for this may be the high mortality 

of owls, which largely feed on nocturnal rodents 

and other small mammals. For example, an epizootic 

of 64 dead or dying Great Horned Owls received by a 

wildlife rehabitation centre in Ohio in the space of six 

weeks was attributed to WNV [53], and there are similar 

reports from other sites in the US. Lastly, large dieoff 

in an alligator farm in Georgia has been attributed 

to the alligators’ diet of horse meat [54]. 

In the 1950s, up to 100% of hooded crows (Corvus corone 

sardonius) and more than 80% of the human population 

sampled in a group of villages at the southern 

end of the Nile delta, 50 km north of Cairo, Egypt, were 

seropositive for WNV, and more than 80% of the human 

population were also seropositive [55]. Laboratory 

studies confirmed that the birds were highly susceptible 

to WNV infection with consistently high titres of 

viraemia. The African species is not markedly social in 

habits, but it may be that, as in the New World, carrion 

feeding contributes to the high infection rate, and 

it is tempting to speculate that the virus is particularly 

adapted to corvids and raptors. Moreover, these birds 

feed by tearing shreds of meat from carrion or prey and 

packing them into a large storage bolus in the crop, 

after which fragments of the bolus are moved, piece 

by piece, to the stomach. Virus will be destroyed by 

the low pH of the stomach, but presumably until then, 

infection can occur by contact with the walls of the 

crop. 

The contrast in pathogenicity between the Old and the 

New World is indicative of a long association between 

the virus and its avian hosts in its original range. 

Indeed, bird species with low mortality in the Americas 

are those that, like the virus, are exotics imported from 

the Old World. In this context, there is a clear parallel 

with another Old World flavivirus, yellow fever virus, 

which was transported to the Americas from Africa in 

the slave trade. In its original range, infections in wild 

primates, the enzootic hosts, are asymptomatic, but 

in the Americas, the virus is lethal to monkeys; local 

inhabitants recognise an epizootic when the rain forest 

goes ‘silent’ because of mass mortality among 

Howler monkeys. In both cases, the introduction of 

an exotic zoonotic virus that is not pathogenic in its 

original range (presumably because it has a long history 

of contact with its hosts) has had a catastrophic 

impact on the local fauna in its new habitat. This is an 

important point: it is probably inappropriate to suggest 

that WNV will emerge as a serious pathogen in the Old 


World on the basis of what has happened in the past 

decade in the Americas. 

Bird migration 

In a serosurvey along the entire Nile valley, from southern 

Sudan to the Nile delta, seropositive humans were 

present at 39 of 40 locations [48]. The river is one of 

the world’s major routes for migrating birds, and the 

continuation of this flyway into Europe, via the Levant, 

the Bosporus and into eastern Europe, is a pathway 

with a consistent history of equine and human cases of 

WNV. Indeed, more than 130 records of suspected and 

confirmed WNV infection, dating back to the 1950s, 

have recently been collected from archives of health 

reports in Romania (G. Nicolescu, personal communication). 

It is interesting, however, that although the 

seasonal pattern of West Nile fever cases in Egypt and 

Romania is roughly synchronous, the seroprevalence 

data suggest a much higher and more consistent rate 

of transmission south of the Mediterranean. Moreover, 

the Nile valley study, there was little indication of significant 

mortality in humans; WNV appeared to be a 

childhood infection and the majority of people in older 

cohorts, who are more vulnerable to central nervous 

system complications, were already immune. 

In a study of 25 species of birds captured in the 

Guadialquivir delta, southern Spain, trans-Saharan 

migrant species had higher seroprevalence and 

higher antibody titres than resident and short-distance 

migrants, evidence that the migrants are primarily 

exposed to WNV in areas with higher circulation 

of virus, rather than in Europe. Indeed, a study in 

Senegal, where several of these species overwinter, 

revealed seroprevalence in horses as high as 90% [56], 

recalling the high seroprevalence observed along the 

river Nile [48]. An interesting point regards infections 

in horses: morbidity and mortality has not been documented 

in Egypt or Senegal, perhaps an indication of 

innate immunocompetence in areas of high circulation. 

The same may to apply to Romania, which has a population 

of about a million horses but little evidence of 

symptomatic infections. 

Transport of virus 

As already stated, commonality between viral 

sequences in different geographic areas is clear evidence 

of transportation in birds. This raises the question: 

how is it possible that a bird, in which viraemia 

lasts at most seven our eight days, can carry virus over 

distances of thousands of kilometres in a flight that 

lasts many weeks? The simplest explanation is that 

migrants en route have refuelling stops where they rest 

and feed before continuing their journey; at these sites, 

virus could be transmitted between migrants, and to 

local resident species, so that stopovers become foci 

of infection. This is plausible at certain sites, for example 

at desert oases, but transmission in, for example, 

the Nile valley occurs in mid-summer, after the passage 

of spring migrants [48]. An alternative explanation 

is that ectoparasites, such as hippoboscids and 


ticks, may constitute the real reservoir, carrying the 

virus on their avian hosts, and somehow transferring 

it to new birds at the migration destination. Lastly, it 

has been suggested that migration is stressful, and 

that this stress may cause a recrudescence of virus in 

birds with chronic infection. There is no physiological 

evidence for such stress, and indeed corticosterone 

levels rise after migration is complete [57]. Moreover, it 

is unlikely that viraemia in immunocompromised birds 

would attain levels sufficient to infect mosquitoes. A 

more likely possibility is that latent virus enters the 

transmission cycles when migrants are consumed by 

scavengers or raptors, or when feeding their young. 

Vector control 

In the US, ultra-low-volume fogging with adulticidal 

aerosols of insecticides delivered from road vehicles is 

widely used to combat WNV vectors and nuisance mosquitoes 

in residential areas. Unfortunately, the efficacy 

of this technique is affected by spacing between buildings, 

distance between roads, amount and type of vegetation, 

wind, convection and many other factors, and 

realistic field evaluations have given markedly variable 

results [58,59]. Moreover, aerosols do not affect the 

aquatic stages of the insects, and mortality of adults is 

restricted to those that are in flight and exposed in the 

short time, a matter of minutes, that the aerosol is airborne 

in lethal concentrations. For this and many other 

reasons, the epidemiological impact of fogging is hard 

to assess [60], and may be minor at best. 

In Europe, most transmission is associated with wetland 

areas of high biodiversity where, apart from difficulty 

of access, the use of insecticides is undesirable. 

In urban areas, a logical approach is the elimination 

of larval habitat. Graded drainage systems and other 

measures of basic sanitation are key to eliminating 

the problem at source, but this is not always straightforward. 

For example, water in catch-basins (settling 

tanks below street-drains) can be a major source of 

Cx. pipiens during dry weather, but they are difficult to 

treat effectively because they are flushed by rainfall. 

The problem in poorly constructed apartment buildings, 

such as occurred in Bucharest, will require major 

reconstruction. 

Weather and WNV recrudescence 

A great deal of attention has been paid to the potential 

impact of climate change on the prevalence and incidence 

of mosquito-borne disease [61]. Given that WNV 

is rarely evident in the Old World, however, it is hard 

to assess the role of climatic factors in its transmission. 

In this context it is therefore pertinent to review 

knowledge about Saint Louis encephalitis virus (SLEV), 

a closely related counterpart in the New World that has 

been the subject of research in the Americas since the 

1930s, for the similarities to WNV are striking: 

• Both are flaviviruses in the Japanese encephalitis 

complex. 

19

• Both are transmitted between birds by ornithophilic 

mosquitoes, mainly of the genus Culex. 

• Transmission of SLEV is rarely evident because 

infections in birds are asymptomatic, as is the 

case with WNV in the Old World. 

• As in Europe, urban epidemics of Saint Louis 

encephalitis have occurred in areas of poor sanitation, 

where sewage-polluted ditches and other 

collections of organically rich water lead to large 

numbers of Culex mosquitoes. 

• Infection of humans and horses can cause encephalitis, 

sometimes fatal. 

• Mammals appear to be dead-end hosts; viraemia is 

insufficient to infect mosquitoes. 

In common with WNV and indeed many other arboviruses, 

SLEV can remain undetected over long periods 

of time. It is only at erratic intervals, sometimes separated 

by several decades, that a sudden recrudescence 

is observed, occasionally developing into a significant 

epizootic. For example, the last major outbreak of 

Saint Louis encephalitis in North America was in 1976, 

yet despite a massive increase in surveillance of mosquitoes 

and birds for WNV (with simultaneous testing 

for SLEV), only a few small outbreaks have been documented 

in the past 25 years. 

Attempts have been made to associate Saint Louis 

encephalitis outbreaks with specific weather conditions. 

In regions of the US where Cx. pipiens and a second 

species, Cx. restuans, are the principal vectors, a 

pattern of mild winters, cold wet springs and hot dry 

summers has been associated with epizootics and 

human cases [62]. In the period since the first major 

epidemics to be recognised (more than 1,000 cases in 

St. Louis, Missouri, in neighbourhoods with primitive 

sanitation and extensive sewage-polluted ditches in 

1932 and 1933) the pattern holds true for some, but by 

no means all outbreaks, nor for years with such conditions 

but no outbreaks. Hot dry summers are liable 

to result in large accumulations of organically polluted 

stagnant water, favoured breeding habitat for 

Cx. pipiens, but although a number of summers have 

fitted this description since the appearance of WNV, 

and despite an enormous increase in vigilance for 

WNV, evidence of SLEV transmission has been unusually 

low. Similarly in Europe, the summers of 1996 and 

1999 were unusually hot and dry and coincided with 

outbreaks in Bucharest and Volgograd, but even hotter 

and drier years have occurred since then without 

any accompanying transmission. In short, the causes 

for recrudescence of both viruses remain enigmatic, 

and it may well be impossible to associate periods of 

transmission with specific patterns of weather. Indeed, 

given that the cradle of transmission is almost certainly 

south of the Sahara, we may need to look to the African 

continent for clues; transmission in Europe may represent 

the tip of the iceberg which has its main mass in 

the tropics. 

Future of WNV in Europe 

As already stated, the spectacular panzootic of WNV 

in the Americas has drawn attention to this virus, 

and it has been suggested that it is also an emerging 

pathogen in the Old World. It is important to put 

this into perspective: even if we include the urban outbreaks 

in Romania and Russia, less than 200 deaths 

in humans have been recorded over the past decade, 

and the number of equine cases is in the same order of 

magnitude. While it is true that an increasing number 

of small outbreaks, mainly among horses, have been 

reported, at least part of this increase was probably 

due to increased awareness of the virus, and major 

improvements in surveillance and diagnostic facilities. 

One point is clear: the importation and establishment 

of vector-borne pathogens that have a relatively low 

profile in their current habitat is a serious danger to 

Europe and throughout the world. It is a direct result of 

the revolution of transport technologies and increasing 

global trade that has taken place in the past three decades. 

Modern examples include the global circulation 

of dengue virus serotypes [63], the intercontinental 

dissemination of Aedes albopictus and other mosquitoes 

in used tires [64,65], the epidemic of chikungunya 

virus in Italy [66], and the importation of bluetongue 

virus and trypanosomiasis into Europe [67,68]. Thus, if 

for example SLEV were to be introduced into the Old 

World, there is every reason to believe that it would 

spark a panzootic analogous to that of WNV in the 

western hemisphere. In short, globalisation is potentially 

a far greater challenge to public health in Europe 

than any future changes in climate [69]. 


I am grateful to Guy Hendrickx and Sarah Randolph for 

helpful suggestions. This review is based on a literature 

study conducted as part of the European Centre for Disease 

Prevention and Control funded V-borne project “Assessment 

of the magnitude and impact of vector-borne diseases in 

Europe”, tender n° OJ/2007/04/13-PROC/2007/003. It was 

compiled, edited and reviewed according to the required 

Euro-Surveillance scientific review format using EDEN funding, 

EU grant GOCE-2003-010284 EDEN. The paper is catalogued 

by the EDEN Steering Committee as EDEN0196 (http:// 

www.eden-fp6project.net/). The contents of this publication 

are the responsibility of the authors and do not necessarily 

reflect the views nor of the European Centre for Disease 

Prevention and Control, nor of the European Commission. 

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Yellow fever and dengue: a threat to Europe? 

P Reiter (paul.reiter@pasteur.fr) 1 

1. Insects and Infectious Disease Unit, Institut Pasteur, Paris, France 


Citation style for this article: Reiter P. Yellow fever and dengue: a threat to Europe?. Euro Surveill. 2010;15(10):pii=19509. Available online: http://www. 


The introduction and rapidly expanding range of Aedes 

albopictus in Europe is an iconic example of the growing 

risk of the globalisation of vectors and vectorborne 

diseases. The history of yellow fever and dengue 

in temperate regions confirms that transmission of 

both diseases could recur, particularly if Ae. aegypti, 

a more effective vector, were to be re-introduced. The 

article is a broad overview of the natural history and 

epidemiology of both diseases in the context of these 

risks. 

Background 

There is logic in dealing with yellow fever and dengue 

together, for they have much in common: 

• Both are caused by viruses of the family 

Flaviviridae, genus Flavivirus. 

• Both viruses are strictly primatophilic – they only 

infect primates, including man. 

• In their original habitat, both are zoonotic infections 

transmitted by forest-dwelling mosquitoes. 

• Both can cause haemorrhagic illness in humans, 

often with fatal consequences. 

• Both owe their importance as human pathogens to 

two forest mosquitoes that have become closely 

associated with the peridomestic environment. 

• The viruses and their urban vectors owe their 

worldwide distribution to transportation of goods 

and people. 

• Both diseases have a history of transmission in 

temperate regions, including Europe. 

According to the World Health Organization, there are 

currently 200,000 worldwide cases and 30,000 deaths 

from yellow fever per year, 90% of them in Africa [1], 

and as many as 50 million cases of dengue [2]. 

Epidemics of yellow fever, sometimes catastrophic, 

were once common in North America as far north as 

New York and Boston (Table), and in European ports 

as far north as Cardiff and Dublin [3]. Large epidemics 

of dengue occurred in the same regions from the 18th 

century onwards. A massive epidemic, estimated at 

one million cases, with at least 1,000 deaths, occurred 

in Greece in 1927-28 [4,5] 



Aedes aegypti, the primary urban vector for both 

viruses, was once established as far north in Europe 

as Brest and Odessa (Figure 1). It disappeared from 

the entire Mediterranean region in the mid-20th century, 

for reasons that are not clear. Ae. albopictus, generally 

regarded as a less important vector of dengue 

[7], is also capable of transmitting yellow fever. It was 

introduced to Europe in the 1970s, is well established 

in at least twelve countries (Figure 2) [8], and is likely 

to spread northwards, perhaps as far as Scandinavia. 

The number of persons who visit countries endemic for 

dengue and yellow fever is continually rising [11,12]. It 

is therefore cogent to consider whether introduction 

of these viruses is likely to lead to autochthonous and 

even endemic transmission in Europe. 

Transmission 

Five factors are key to the epidemiology of vector-borne 

diseases: the ecology and behaviour of the host, the 

ecology and behaviour of the vectors, and the degree 

of immunity in the population. A holistic view of this 

complexity is key to assessing the likelihood of transmission 

in Europe [13]. 

Origin of the viruses 

There is little doubt that the yellow fever virus (YFV) 

originated in Africa, and that viruses circulating in the 

New World are of African origin. Curiously, yellow fever 

has never been recorded in Asia, although Ae. aegypti 

is widespread there. 

There are four antigenically distinct DENV serotypes 

that cause very similar disease in humans. It is widely 

accepted that all four are of Asian origin [14], although 

DENV-2 is enzootic in Africa [15]. 

Zoonotic vectors and hosts 

In the Old World, the sylvatic vectors of yellow fever 

and dengue are canopy-dwelling mosquitoes of the 

genus Aedes and three subgenera, Stegomyia, Finlaya, 

and Diceromyia, that feed exclusively on monkeys. In 

the Americas, the principal zoonotic vectors of yellow 

fever are Sabethes and Haemogogus species; both are 

also strictly primatophilic [3]. 

23

Sylvatic transmission to humans 

Sylvatic infections are acquired when humans enter 

woodland where there is zoonotic transmission. In 

recent years, a number of unvaccinated tourists have 

died of yellow fever after visiting enzootic areas [16,17]. 

Vector-host specificity 

Host specificity is a characteristic of many vectors; it 

is conceivable that it improves the chances of locating 

hosts. This may be particularly useful in the sylvatic 

environment, where bands of monkeys roam between 

established sleeping sites. 

The specificity of DENV and YFV to primatophilic vectors 

may have evolved to exploit this relationship, 

and/or to surmount barriers to infection in the insect. 

Table 

Major epidemics of yellow fever in North America, north of Mexico 

Whatever the reason, given the absence of wild primates, 

it is unlikely that any vector species native to 

Europe is able to transmit these viruses. 

Peridomestic transmission 

Neither YFV nor DENV would have major importance as 

human pathogens in the absence of two mosquito species, 

Ae. (Stegomyia) aegypti and Ae. (S.) albopictus, 

both of which have become closely associated with the 

peridomestic environment. Infected humans returning 

from an enzootic area may initiate transmission to 

humans in human settlements if either of these species 

is present (although to date, no yellow fever infections 

have been attributed to Ae. albopictus). 

Year Year 

1668 New York, Philadelphia and other settlements 1803 Boston, Philadelphia 

1690 Charleston 1804 Philadelphia 

1691 Boston 1805 Philadelphia 

1693 Charleston, Philadelphia, Boston 1807 Charleston 

1694 Philadelphia, New York, Boston 1811 New Orleans, Florida, New Jersey 

1699 Charleston, Philadelphia 1817 New Orleans, Charleston, Baltimore 

1702 New York 1819 New Orleans, Charleston, Baltimore, Philadelphia, New York 

1703 Charleston 1820 New Orleans, Philadelphia 

1728 Charleston 1821 

New Orleans, Mississippi Valley, Alabama, Charleston, 

Baltimore, Philadelphia, New York, Boston 

1732 Charleston 1822 New Orleans, New York 

1734 Charleston, Philadelphia, New York, Albany, Boston 1823 Key West 

1737 Virginia 1824 New Orleans, Charleston 

1739 Charleston 1825 Mobile, Natchez, Washington 

1741 Virginia, Philadelphia, New York 1827 New Orleans, Mobile 

1743 Virginia, New York 1828 New Orleans, Memphis 

1745 Charleston, New York 1829 Key West, Mobile, Natchez 

1747 New Haven 1837 New Orleans, Mobile, Natchez 

1748 Charleston 1839 Galveston, Mobile, Charleston 

1751 Philadelphia, New York 1841 Key West, New Orleans 

1762 Philadelphia 1843 Galveston, Mobile, Mississippi Valley, Charleston 

1778 Philadelphia 1847 New Orleans, Mobile, Natchez 

1780 Philadelphia 1852 Charleston 

1783 Baltimore 1853 New Orleans 

1791 Philadelphia, New York 1854 New Orleans, Mobile, Alabama, Charleston 

1792 Charleston 1855 Mississippi Valley, Norfolk 

1793 Philadelphia 1856 New Orleans, Charleston 


1795 Philadelphia 1867 Key West, Galveston, New Orleans, Mobile, Philadelphia 

1796 Philadelphia 1870 New York 

1797 Philadelphia 1873 New Orleans, Mississippi Valley, Alabama, Memphis 


1799 Philadelphia 1877 Port Royal SC 

1800 Philadelphia 1878 

New Orleans, Memphis, Mississippi Valley to St Louis, Chattanooga, 

many other cities 

1801 Norfolk, New York, Massachussetts 1879 Memphis 

1802 Philadelphia 1905 New Orleans 

Reproduced from [6] with permission from Environmental Health Perspectives. 


Figure 1 

Historical distribution of Aedes aegypti 

Dark grey areas: maximum range distribution of Ae. aegypti, black 

lines: January 10°C isotherm in the northern hemisphere; mid grey 

lines: the July 10°C isotherm in the southern hemisphere. 

The distribution limit broadly fits the 10°C isotherm in the southern 

hemisphere, but far less so in the northern hemisphere. 

Source: adapted from a map published by Christophers [9]. 

Figure 2 

Current (2009) distribution of Aedes albopictus in Europe by administrative unit 

Overwintering expanding populations 

No recent data on mosquito fauna 

Source: [10]. 


Dengue is endemic in many urban and rural populations 

throughout the tropics. ‘Virgin soil’ epidemics in 

large cities are often explosive. In 1988, for example, 

there were an estimated 420,000 cases in four months 

in the coastal city of Guayaquil, Ecuador [18] 

The large urban outbreaks of yellow fever that were 

common until the early 20th century remain a real 

and constant danger in enzootic countries that do not 

enforce routine vaccination. Moreover, it is reasonable 

to assume that areas that are prone to dengue transmission 

are equally prone to yellow fever, so areas 

without history of the latter, including those in southeast 

Asia, may well be at risk. 


The yellow fever mosquito, Aedes aegypti 

Ae. aegypti is the quintessential urban vector of yellow 

fever and dengue. It is a remarkable species because 

Populations only observed indoors (in glass houses) 

No information on any mosquito studies 

Not detected in past 5 years 

Not included in this study 

25

the ‘domesticated’ form is rarely found more than 100 

m from human habitation and feeds almost exclusively 

on human blood. Nevertheless, like its forest ancestor, 

it remains day-active with a preference for heavy 

shade. It freely enters homes and other buildings and 

spends much of its time hidden in dark places, often 

among clothing, a stable microclimate with few predators. 

Its human host is abundant and lives under the 

same roof, an arrangement that minimises the hazards 

of questing for a blood meal. It lays eggs in man-made 

objects that contain water, from discarded tires and 

buckets to the saucers under flowerpots and waterstorage 

barrels. In short, humans are the perfect host: 

they provide safe shelter, plentiful food and abundant 

sites for procreation. Indeed, in most cities of the tropics, 

homes are so close together and breeding sites so 

abundant that they can be regarded as a single factory 

for mosquitoes in an urban jungle. In the past three 

decades, attempts to reduce populations of the species 

have rarely been successful and never sustained 

[19,20]. 

The Asian Tiger mosquito, Aedes albopictus 

Ae. albopictus is often abundant in the peridomestic 

environment, particularly in areas with plentiful vegetation. 

However, in addition to humans, it feeds freely 

on animals and birds, and so can exist far from human 

habitation. Since non-primates are not susceptible to 

the viruses, such blood meals do not contribute to the 

transmission cycle, and for this reason, Ae. albopictus 

has generally been regarded as a secondary vector [7]. 

Nevertheless, dengue epidemics have been recorded in 

places where Ae. albopictus is the only vector [21], and 

in recent years, the species has proved highly effective 

in urban transmission of another African sylvatic virus, 

chikungunya virus [22,23]. 

Globalisation of vectors and viruses 

Aedes aegypti 

Ae. aegypti and yellow fever arrived in the New World 

together, as passengers in the slave trade. Slave 

ships generally made the passage from Africa to the 

Americas in four to six weeks. The virus was enzootic 

in regions where the slave caravans captured local 

inhabitants, and urban transmission was rife in the 

ports of dispatch. The casks used for shipboard storage 

of water must have been prolific breeding sites for 

the mosquito, and the slaves were an abundant source 

of blood. With the slaves and the mosquito came the 

virus, and it was not uncommon for ships to arrive 

in port with large numbers of dying persons aboard, 

hence the yellow flag of quarantine. 

In the United States, the species has been recorded 

from 21 states (Alabama, Arkansas, Florida, District of 

Colombia, Georgia, Illinois, Indiana, Kansas, Kentucky, 

Louisiana, Maryland, Missouri, Mississippi, New York, 

North Carolina, Ohio, Oklahoma, South Carolina, 

Tennessee, Texas, and Virginia) [24]. In many of these, 

winter temperatures below -20°C are not unusual. 

Presumably the mosquitoes survive in sheltered sites, 

for they are not resistant to freezing. Thus there is no 

obvious climatic reason why the species, were it to 

be re-introduced, could not survive in most areas in 

Europe. 

Aedes albopictus 

In its original range, Ae. albopictus was present from 

Beijing and northern Japan to tropical Asia [25]. In 

1983, however, the mosquito was found in Memphis, 

Tennessee [26], and, two years later, a survey revealed 

that it was widely distributed, often common, in the 

southern United States. Investigation revealed a global 

trade in used tyres that were frequently infested 

with eggs and larvae of the species [27]. Japan was the 

principal exporter, and a study of winter diapause at 

various latitudes in Asia confirmed that the day-length 

that triggered diapause was identical in the southern 

United States and in southern Japan [28]. The mosquito 

is now widespread in the United States, and is a 

major nuisance species as far north as Nebraska and 

Illinois, where winter snowfall can be well above 200 

cm, average January night-time temperatures are -10 0 C, 

and temperatures as low as -33 0 C have been recorded. 

It is also established in Mexico and all the countries of 

Central and South America except Chile. In Africa it is 

well established in Nigeria, Gabon, Equatorial Guinea 

and Cameroon [29,30], and in Europe it has been 

reported from 16 countries [8]. Recent infestations in 

the Netherlands have been traced to imports of ‘lucky 

bamboo’ from sub-tropical China [31], but these mosquitoes 

do not appear to have survived the winter, perhaps 

because they have no winter diapause. 

Clinical features 

Yellow fever 

As with most viral diseases, yellow fever can present 

with a wide spectrum of symptoms, from mild to fatal. 

In clinical cases, there is generally a sudden onset of 

fever with severe headache, arthralgias, and myalgia. 

The striking yellowing of the eyes and skin, caused by 

hepatic dysfunction, may appear on the third day and 

indicates a poor prognosis. The fever often follows a 

‘saddleback’ curve, with a brief drop in temperature 

and symptoms after the third day, followed by a return 

with increased severity that can lead to spontaneous 

haemorrhage (‘coffee ground’ vomit), delirium, renal 

failure, coma and death. Fatality rates of clinical cases 

can be as high as 80% [3], on a par with Ebola, Marburg 

and other haemorrhagic viral infections. 

Dengue 

As many as 80% of all dengue infections are asymptomatic. 

Among clinical cases, early stages are similar 

to those of yellow fever, although with excruciating 

arthralgia and myalgia, hence the term ‘break-bone 

fever’. Fever and other symptoms rarely last more 

than seven days, but convalescence can be prolonged 

and debilitating. The later stages of the illness often 

include a widespread rash [32]. 


A portion of dengue cases, usually less than 5%, can be 

severe and a fraction of these may be fatal [33]. Severe 

dengue, commonly referred to as dengue haemorrhagic 

fever/dengue shock syndrome (DHF/DSS) to distinguish 

it from ‘classic’ dengue, is associated with spontaneous 

haemorrhage and an increase of vascular permeability 

that can lead to life-threatening hypovolemic 

shock. The causes of this condition have been debated 

for decades, but remain unresolved [34-36]. A widely 

held but hotly contested hypothesis is that after infection 

with one serotype, secondary infections by one or 

more of the others can precipitate the syndrome by a 

process referred to as antibody-dependent enhancement, 

but the occurrence of severe dengue in epidemics 

of primary infection, such as the Greek epidemic 

and a recent epidemic in Cape Verde [37], contradicts 

this hypothesis. An associated controversy is the validity 

of graded sets of criteria to categorise severity that 

are recommended by the World Health Organization, 

and these have been revised several times in recent 

years [38]. Both issues are of prime importance for the 

management and treatment of patients. 

It is a common misconception that DHF/DSS first 

appeared in the 1950s in south-east Asia. It is certainly 

true that the syndrome became a serious public health 

problem in that period, but it was not a new phenomenon: 

significant mortality associated with haemorrhagic 

symptoms had been described in the earliest epidemic 

of dengue-like disease on record, in Philadelphia in 

1789, as well as in later epidemics in East Africa and in 

Australia [14,39]. Moreover, as already mentioned, at 

least 1,000 people died in the Greek epidemic in 1927- 

28. In the years after the Second World War, however, 

rapid expansion of densely populated urban areas, 

coupled with enormous infestations of Ae. aegypti, led 

to a massive increase in the prevalence and incidence 

of the disease in south-east Asia, so a plausible explanation 

for the emergence of this ‘new’ syndrome is that 

escalating numbers of classic infections simply led to 

an increased awareness of the relatively rare manifestations 

– the ‘iceberg effect’. 

Treatment 

There is no specific treatment for yellow fever or dengue 

virus infections; supportive therapy is the only 

option, although there is active research into antiviral 

drugs against these diseases [40]. For dengue fevers, 

intravenous fluids are used to counter haemoconcentration, 

and platelet transfusions in the event of severe 

thrombocytopaenia [41]. Strict avoidance of anticoagulants, 

including aspirin, is important. 

Prevention 

Vaccination 

Yellow fever 

A safe, effective yellow fever vaccine, based on a live 

attenuated strain, has been available for more than half 

a century, and mass vaccination is a highly effective 

approach to prevent urban transmission, but the incidence 

of the disease, particularly in Africa, confirms 


that coverage is inadequate, and there is a real and 

present danger of a major urban epidemic. Moreover, 

there is good reason to believe that the 2.5 billion people 

who live in regions at risk of dengue infection are 

also at risk of yellow fever; if so, then, given the lax 

attitude towards vaccination of travellers in most countries, 

the danger of a catastrophic epidemic beyond 

regions generally associated with transmission is also 

real, and this could include parts of Europe infested 

with Ae. albopictus. If such an event were to occur, current 

stocks of vaccine would probably be inadequate to 

respond to worldwide demand. 

Dengue 

No vaccine against dengue is available, but attenuated 

virus vaccines and second-generation recombinant 

vaccines are in active development [42]. A large-scale 

trial (phase IIb) of a chimeric tetravalent vaccine [43] 

has been under way since February 2009 [44]. If successful, 

then a vaccine might be licenced within five 

years. 

Vector control 

At the beginning of the 20th century, urban yellow fever 

was eliminated from many countries by energetic campaigns 

to eliminate Ae. aegypti breeding sites. After 

the Second World War, focal application of the synthetic 

pesticide dichlorodiphenyltrichloroethane (DDT) 

to infested containers and their surroundings was an 

outstanding success; according to the Pan American 

Health Organization, the species was eradicated from 

22 countries in the Americas [45]. The reason for the 

efficacy of this method has only recently become 

apparent: ‘skip-oviposition’ (the deposition of small 

numbers of eggs in many different sites) made it highly 

probable that they would encounter treated sites [19]. 

No substitute for DDT is currently available, so many 

authorities resort to spraying insecticidal aerosols 

(ultra-low-volume) of organophosphates or pyrethroids 

from hand-held machines, road vehicles or aircraft. 

Unfortunately, the method is expensive and generally 

ineffective, at least against Ae. aegypti, because 

the species spends much of its time indoors at sites 

that are inaccessible to the aerosol [20,46]. Moreover, 

even if a large number of mosquitoes were to be eliminated 

by this treatment, the impact on adult mosquito 

populations would probably be too short for an effective 

impact on transmission [47]. Although the World 

Health Organization recommends that health authorities 

evaluate the technique under local circumstances 

[6], their principal strategy is community-based source 

reduction, the elimination of breeding sites by the community. 

Unfortunately, there is no evidence that this 

approach has been successful in any part of the world. 

Control of Ae. albopictus is probably even more difficult 

than for Ae. aegypti, given its ability to breed away 

from human habitation, but insecticidal aerosols may 

be more effective for Ae. albopictus because the mosquito 

tends to rest outdoors. 

27

The future in Europe 

Dengue is essentially an urban disease because of 

the urban ecology of its vectors and the behaviour of 

its hosts. Rapid urbanisation has made it an increasingly 

serious public health problem in the tropics [48]. 

Millions of people travel from the tropics to Europe 

and North America each year (for example, 1.2 million 

people who live in the UK visit the Indian subcontinent, 

with average stays of 29 days) and, after malaria, dengue 

infection is the second most frequent reason for 

hospitalisation after their return [11,12]. 

The history of dengue and yellow fever in Europe 

is evidence that conditions are already suitable 

for transmission. The establishment of 

Ae. albopictus has made this possible, and the possibility 

will increase as the species expands northwards, or 

if Ae. aegypti is re-established. The epidemic of chikungunya 

in northern Italy in 2007 [8,49] confirms that 

Ae. albopictus is capable of supporting epidemic transmission, 

although laboratory studies indicate that the 

strain of virus involved was particularly adapted to this 

species [50,51]. Nevertheless, it is not unreasonable 

to assume that climatic conditions that permit malaria 

transmission will also support transmission of yellow 

fever and dengue, in which case transmission could 

extend into northern Europe [52]. 

Lastly, it is widely stated that the incidence of vectorborne 

diseases will increase if global temperatures 

increase. While there is no doubt that temperature 

and rainfall play a role in their transmission, it is clear 

that many other factors are involved [6]. A more urgent 

emerging problem is the quantum leap in the mobility 

of vectors and pathogens that has taken place in 

the past four decades, a direct result of the revolution 

of transport technologies and global travel [53]. The 

potential impact of this globalisation of vector-borne 

diseases is a challenge for the future. 


This review is based on a literature study conducted as part 

of the European Centre for Disease Prevention and Control 

funded V-borne project “Assessment of the magnitude 

and impact of vector-borne diseases in Europe”, tender n° 

OJ/2007/04/13-PROC/2007/003. It was compiled, edited and 

reviewed according to the required Euro-Surveillance scientific 

review format using EDEN funding, EU grant GOCE-2003- 

010284 EDEN. The paper is catalogued by the EDEN Steering 

Committee as EDEN0198 (http://www.eden-fp6project.net/). 

The contents of this publication are the responsibility of the 

authors and don’t necessarily reflect the views nor of the 

European Centre for Disease Prevention and Control, nor of 

the European Commission. 

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29


Rift Valley fever - a threat for Europe? 

V Chevalier (veronique.chevalier@cirad.fr) 1 , M Pépin 2 , L Plée 3 , R Lancelot 4 

1. Centre International de Recherche Agronomique pour le Développement (CIRAD, International Centre of Agricultural Research 

for Development), Unit for animal and integrated risk management (UR AGIRs), Montpellier, France 

2. Agence française pour la sécurité sanitaire des aliments (AFSSA, French Agency for Food Safety), Lyon, France 

3. Agence française pour la sécurité sanitaire des aliments (AFSSA, French Agency for Food Safety), Unit for the evaluation of 

risks associated with food and animal health, Maisons-Alfort, France 

4. Centre International de Recherche Agronomique pour le Développement (CIRAD, International Centre of Agricultural Research 

for Development), Unit for the control of exotic and emerging animal diseases (UMR CMAEE), Montpellier, France 


Citation style for this article: Chevalier V, Pépin M, Plée L, Lancelot R. Rift Valley fever - a threat for Europe?. Euro Surveill. 2010;15(10):pii=19506. Available online: 

http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19506 

Rift Valley fever (RVF) is a severe mosquito-borne 

disease affecting humans and domestic ruminants, 

caused by a Phlebovirus (Bunyaviridae). It is widespread 

in Africa and has recently spread to Yemen 

and Saudi Arabia. RVF epidemics are more and more 

frequent in Africa and the Middle East, probably in 

relation with climatic changes (episodes of heavy rainfall 

in eastern and southern Africa), as well as intensified 

livestock trade. The probability of introduction 

and large-scale spread of RVF in Europe is very low, 

but localised RVF outbreaks may occur in humid areas 

with a large population of ruminants. Should this happen, 

human cases would probably occur in exposed 

individuals: farmers, veterinarians, slaughterhouse 

employees etc. Surveillance and diagnostic methods 

are available, but control tools are limited: vector control 

is difficult to implement, and vaccines are only 

available for ruminants, with either a limited efficacy 

(inactivated vaccines) or a residual pathogenic effect. 

The best strategy to protect Europe and the rest of the 

world against RVF is to develop more efficient surveillance 

and control tools and to implement coordinated 

regional monitoring and control programmes. 

Relevance of Rift Valley fever to public 

health in the European Union 

Rift Valley fever (RVF) is a zoonotic disease of domestic 

ruminants and humans caused by an arbovirus belonging 

to the Phlebovirus genus (family Bunyaviridae). It 

causes high mortality rates in newborn ruminants, 

especially sheep and goats, and abortion in pregnant 

animals. Human infection by the RVF virus (RVFV) may 

result from mosquito bites, exposure to body fluids of 

livestock or to carcasses and organs during necropsy, 

slaughtering, and butchering [1]. 

The public health impact of RVF can be severe. In Egypt 

in 1976, 200,000 people were infected and 600 fatal 

cases officially reported, among others in the River 

Nile delta [2]. Over 200 human deaths were reported in 

Mauritania in 1987 [3]. In 2007-2008, 738 human cases 

were officially reported in Sudan, including 230 deaths 


[4]. It is likely that the number of cases was underreported 

because RVF mostly affects rural populations 

living far from public health facilities. The occurrence 

of RVF in northern Egypt is evidence that RVF may occur 

in Mediterranean countries, thus directly threatening 

Europe. In the Indian Ocean, RVF has been introduced 

in the French island of Mayotte, with several clinical 

cases reported in humans [5] 

Transmission, epidemiology 

and clinical symptoms 

The RVFV transmission cycle involves ruminants and 

mosquitoes. Host sensitivity depends on age and animal 

species [6] (Table 1). Humans are dead-end hosts. 

The epidemiological cycle is made more complex by 

direct transmission from infected ruminants to healthy 

ruminants or humans, by transovarian transmission 

in some mosquito species, and by a large number of 

potential vectors with different bio-ecology [6]. The 

existence of wild reservoir hosts has not been clearly 

demonstrated to date (Figure 1). 

Transmission mechanisms 

The bite of infected mosquitoes is the main transmission 

mechanism of RVF in ruminants during inter-epizootic 

periods. More than 30 mosquito species were found to 

be infected by RVFV [6,7] (Table 2), belonging to seven 

genera of which Aedes and Culex are considered as the 

most important from the point of view of vector competence 

(other genera are Anopheles, Coquillettidia, 

Eretmapodite, Mansonia and Ochlerotatus). 

In mosquitoes, transovarian RVFV transmission has 

been observed in Aedes mcintoshi. It appears to be a 

likely phenomenon in several other species, including 

the widespread Ae. vexans species complex. In some 

of these Aedes species, infected, diapaused eggs may 

survive in dried mud during inter-epizootic and/or dry/ 

cold periods [8] and hatch infected imagos. 

Ruminant-to-human transmission is the main infection 

route for humans, although they can also be infected 


y mosquito bites [9]. Body fluids such as the blood 

(during slaughtering and butchering), foetal membranes 

and amniotic fluid of viraemic ruminants are 

highly infective for humans. Fresh and raw meat may 

be a source of infection for humans, but the virus is 

destroyed rapidly during meat maturation. Empirical 

Table 1 

Species susceptibility and sensibility to the Rift Valley fever virus 


field observations indicate that ruminants can also 

become infected by contact with material containing 

virus (e.g. fetus and fetal membranes after abortion), 

however, this route of transmission has not yet been 

confirmed [10]. 

Mortality >70% Mortality 10-70% 

Severe disease with low 

fatality rate (

Table 2 

Arthropods naturally infected by Rift Valley fever virus 

Genus Species Country (year) 

cumminsii 

Kenya (1981-1984) 

Burkina Faso (1983) 

dalzieli Senegal (1974, 1983) 

dentatus Zimbabwe (1969) 

Aedes (Aedimorphus) 

durbanensis Kenya (1937) 

ochraceus Senegal (1993) 

tarsalis Uganda (1944) 

vexans arabiensis 

Senegal (1993) 

Saudi Arabia (2000) 

circumluteolus 

Uganda (1955) 

South Africa (1955, 1981) 

Aedes (Neomelaniconion) 

mcintoshi 

Zimbabwe (1969) 

South Africa (1974-1975) 

Kenya (1981-1984) 

palpalis Central African Republic (1969) 

caballus South Africa (1953) 

Ochlerotatus (Ochlerotatus) 

caspius Suspected, Egypt (1993) 

juppi South Africa (1974-1975) 

Aedes (Stegomya) 

africanus 

demeilloni 

Uganda (1956) 

Uganda (1944) 

Aedes (Diceromya) furcifer group Burkina Faso (1983) 

Anopheles (Anopheles) 

coustani 


Madagascar (1979) 

fuscicolor Madagascar (1979) 

chrityi Kenya (1981-1984) 

Anopheles (Cellia) 

cinereus 

pauliani 

South Africa (1974-1975) 


pharoensis Kenya (1981-1984) 

spp. Madagascar (1979) 

antennatus 

Nigeria(1967-1970) 

Kenya (1981-1984) 

neavi South Africa (1981) 

pipiens Egypt (1977) 

Culex (Culex) 

poicilipes 

theileri 

Senegal (1998, 2003) 

South Africa (1970) 


tritaeniorhynchus Saudi Arabia (2000) 

vansomereni Kenya (1981-1984) 

zombaensis 


Kenya (1981-1984, 1989) 

Culex (Eumelanomya) rubinotus Kenya (1981-1984) 

chrysogaster Uganda (1944) 

Eretmapodites 

quinquevittatus 


Kenya (1981-1984) 

Coquillettidia 

fuscopennata 

grandidieri 

Uganda (1959) 


Uganda (1959, 1968) 

africana 

Central African Republic (1969) 

Mansonia (Mansoniodes) 

Kenya (1989) 

uniformis 

Uganda (1959) 


Other diptera Culicoides spp. Nigeria (1967) 

Adapted from [1]. 


Direct human-to-human transmission has not been 

reported, and RVF is not considered to be a nosocomial 

disease. Transplacental RVFV transmission may occur 

in vertebrates, including humans. It results in abortion 

and high newborn mortality rates [11]. 

Rodents may be infected during epizootic periods [12- 

15] but their epidemiological role in virus transmission 

and maintenance is not clear. Bat species also have 

been suspected [16]. Finally, wild ruminants may play 

a role in the epidemiology of RVF in areas where their 

population density is high [17]. 

Clinical features 

Animals 

Clinical manifestations vary depending on age and 

animal species. In sheep, a fever of up to 41-42°C is 

observed after a short incubation period. Newborn 

lambs (and sometimes kids) usually die within 36 to 

40 hours after the onset of symptoms, with mortality 

rates sometimes reaching 95%. Older animals (from 

two weeks to three months-old) either die or develop 

only a mild infection. In pregnant ewes, abortions are 

frequent, ranging from 5% to 100%. Twenty per cent of 

the aborting ewes die. Vomiting may be the only clinical 

sign presented by adult sheep and lambs older than 

three months. However, these animals may experience 

fever with depression, haemorrhagic diarrhoea, bloodstained 

muco-purulent nasal discharge, and icterus. 

Case-fatality rates vary between 20% and 30%. Adult 

goats develop a mild form of the disease, but abortions 

are frequent (80%). Mortality rates are generally 

low [10]. Calves often develop acute illness, with fever, 

fetid diarrhoea, and dyspnoea. Mortality rates may 

vary from 10% to 70%. Abortion is often the only clinical 

sign and mortality rates are low (10-15%). 

Humans 

In most cases, human infections remain unapparent, or 

with mild, influenza-like symptoms. However, infected 

people may experience an undifferentiated, severe, 

influenza-like syndrome and hepatitis with vomiting 

and diarrhoea. Complications may occur. Severe 

forms are manifested in three different clinical syndromes. 

The most frequent one is a maculo-retinitis, 

with blurred vision and a loss of visual acuity due to 

retinal haemorrhage and macular oedema. Encephalitis 

may also occur, accompanied by confusion and coma. 

This form is rarely fatal but permanent sequelae are 

encountered. The third and most severe form is a 

haemorrhagic fever, with hepatitis, thrombocytopenia, 

icterus, and multiple haemorrhages. This form is often 

fatal [10,18,19]. Human case-fatality rates have been 

lower than 1% in the past, however, an increase has 

been reported since 1970 [19]. In the RVF epidemic in 

Saudi Arabia in the year 2000, the fatality rate reached 

14% [20]. 


Diagnostic methods 

RVFV presents a high biohazard for livestock farmers, 

veterinarians, butchers, slaughterhouse employees, 

and laboratory staff handling infected biological samples. 

International public health agencies have set a 

bio-safety level (BSL) of BSL3 for facilities in Europe 

handling the virus and of BSL4 for facilities in the 

United States (US). 

Appropriate diagnostic samples are peripheral blood 

collected on EDTA, plasma or serum of infected animals 

or patients, and the liver, brain, spleen or lymph nodes 

of dead animals. When samples can be conveyed rapidly 

to a diagnostic laboratory (

100% (n = 502), 95% confidence interval: 99.3 to 100% 

[29]. 

Treatments 

There is no specific treatment for either humans or 

animals. 

Prevention 

Vaccines 

A human vaccine (inactivated with beta-propiolactone) 

has been produced in the US and was used to protect 

laboratory staff and military troops. However, its production 

has been stopped [30]. 

Given that domestic ruminants are involved in the epidemiological 

cycle and that humans mostly become 

infected after contact with viraemic animals, the vaccination 

of ruminants is the method of choice to prevent 

human disease. Both live and inactivated vaccines are 

available for livestock. 

The Smithburn vaccine is a live attenuated vaccine. It 

is inexpensive to prepare and immunogenic for sheep, 

goats, and cattle. It protects these species against 

abortion caused by a wild RVFV, and post-vaccinal 

immunity is life long. However, it has a residual pathogenic 

effect and may induce foetal abnormalities and/or 

abortion in ruminants. It is also pathogenic for humans 

(febrile syndrome). Despite these drawbacks, it is recommended 

by the Food and Agriculture Organization 

of the United Nations (FAO) [31] and remains the most 

widely used vaccine against RVF in Africa. 

The inactivated RVF vaccine provides a lower level 

of protection and its production is more expensive. 

Moreover, it requires at least two inoculations and 

frequent booster shots to induce the desired level of 

protection, rendering it inappropriate in countries 

where large portions of ruminant herds are nomadic. 

However, it was used by the Israeli veterinary services 

to prevent RVF introduction to Israel after the 1977- 

1978 epidemic in Egypt [32], as well as by the Egyptian 

veterinary services to prevent re-introduction of RVF 

from Sudan after an epidemic hit that country in 2007. 

Other candidate vaccines are being evaluated such as 

the so-called “clone 13” which is an attenuated strain of 

RVFV that was isolated from a moderately ill patient in 

the Central African Republic [33]. This vaccine induces 

neutralising antibodies against RVFV. New-generation 

vaccines are also under study: recombinant vaccines 

using a poxvirus or an Alphavirus-based vector [34,35] 

and DNA vaccines [34*,36*]. However, these vaccines 

are still in the preliminary stages of development. 

Smithburn and inactivated vaccines are produced and 

commercially available in Egypt, South Africa, and 

Kenya. There is no Community pharmaceutical legislation 

prohibiting companies from producing RVF 

vaccines on EU territory and there is no obligation to 

notify such production to the European Commission. 

Moreover, quoting Council Directive 2001/82/EC (EC 

2OO1b), “in the event of serious epizootic diseases, 

Member States may provisionally allow the use of 

immunological veterinary medicinal products without a 

marketing authorisation, in the absence of a suitable 

medicinal product and after informing the Commission 

of the detailed conditions of use (article 8)” [37*]. 

Insecticide treatments 

Larvicide treatments may provide a control alternative 

where mosquito breeding sites are well identified and 

cover limited surface areas. Both Methoprene, a hormonal 

larval growth inhibitor, and Bacillus thuringiensis 

israeliensis (BTI) preparations, a microbial larvicide, 

are commercially available and can be used successfully 

to treat temporary ponds and watering places 

where mosquitoes proliferate. Adulticide treatments 

(e.g. using pyrethroids) are expensive and difficult to 

implement. Moreover, because this usually involves 

treating large areas, the environmental and ecological 

consequences may be important. 

Other measures 

Preventive measures should also include restrictions 

on animal movements, the avoidance or control of 

the slaughter and butchering of ruminants, the use of 

insect repellents and bed nets during outbreaks, information 

campaigns, and increased and targeted surveillance 

of animals, humans and vectors. 

Current geographical distribution 

RVF is either enzootic, or is reported in most sub-Saharan 

African countries, Egypt and Madagascar (Figure 

2). 

During the first large epidemic, reported in Egypt in 

1977-1978, over 600 people died of RVF [39]. The epidemic 

reached the Mediterranean shore (Nile delta) but 

did not spread to neighbouring countries. In September 

2000, RVF was detected for the first time outside of 

the African continent in Saudi Arabia and Yemen, and 

led to human deaths and major livestock losses [40]. 

By the end of 2006, the disease had re-emerged in 

Kenya [41], followed by Tanzania and Somalia [42]. 

Another large epidemic hit the Sudan in 2007 in the 

Nile Valley around Khartoum [4]. In May 2007, RVF 

was diagnosed on the French island of Mayotte in a 

young boy who had been evacuated from Anjouan, one 

of the other islands of the Comoros archipelago. The 

RVFV was probably introduced there by the trade of 

live ruminants imported from Kenya or Tanzania during 

the 2006-2007 epidemics. Studies conducted after this 

first human case was reported have shown that 10% of 

cattle had antibodies against RVFV (ELISA, IgG and/or 

IgM) - without any clinical suspicions reported by the 

public and private veterinary services. A retrospective 

study was then conducted in 2008, using blood samples 

collected from clinically suspected human cases 

of dengue or chikungunya illness who had tested negative 

for these two diseases, between 1 September 

2007 and 31 May 2008. Ten human RVF cases were 


found (including IgM- and/or RT-PCR-positive samples), 

seven of them (70%) occurring from January to 

April, during the hot, rainy season [5]. This study has 

Figure 2 

Geographical distribution of Rift Valley fever 

Latitude (degrees) 

40 

20 

0 

-20 

Source: United States Centers of Disease Control and Prevention 

[38*]. 


0 20 40 60 

Longitude (degrees) 

RVF free (or unreported) 

Outbreaks and epidemics 

Sporadic cases and serological evidences 

Table 3 

Main outbreaks of Rift Valley fever and factors causing them 

demonstrated that RVF had been circulating in Mayotte 

at least since early 2007, probably introduced there by 

the illegal importation of live infected ruminants from 

other Comoros islands. 

In 2008, a RVF epidemic occurred in Madagascar with 

over 500 human cases [43]. Several outbreaks were 

reported in South Africa in late 2007 and 2008 without 

any reported human cases [44]. 

Factors of change 

Factors that could cause a change in the epidemiology 

of RVF are summarised in Table 3. Irrigated areas, 

including rice fields, constitute favourable breeding 

sites for many mosquito species. Dambos are temporary 

surface water bodies found in semi-arid eastern 

Africa. With heavy rainfall and consecutive flooding, 

considerable mosquito proliferation may occur (mostly 

Aedes and Culex spp.). Wadi are temporary rivers 

encountered in arid areas (e.g. Yemen or Saudi Arabia): 

when they stop flowing, surface water remains available 

in ponds and mosquitoes may proliferate. 

Livestock trade and the Mediterranean region 

Livestock trade and transport may affect the geographical 

distribution of RVF and contribute to a large scale 

– sometimes continental - spread of the disease and to 

the introduction of the virus into disease-free areas via 

livestock movements. RVF cases were reported in irrigated 

areas of the Sudan during the 1970s. Antibodies 

were detected in camels that crossed the border from 

Sudan to Egypt, suggesting that infected camels may 

have introduced RVFV into Egypt [39]. 

Year Country Ecosystem Vector Hosts Triggering factor 

1975 South Africa ? ? ? ? 

1976 Sudan Irrigated area ? Small ruminants Irrigation (?) 

1977 Egypt Irrigated area Culex pipiens 

Small ruminants, camels, 

humans 

Irrigation, cattle trade 

1987 Mauritania, Senegal Irrigated area Culex pipiens 

Small ruminants, cattle, 

camels, humans 

? 

1993 Egypt Irrigated area ? Small ruminants, humans Irrigation 

1997 Egypt Irrigated area ? Small ruminants, humans Irrigation 

1997-1998 Kenya Dambos 

Aedes spp. 

Culex zombaensis 

Small ruminants Rainfall 

2000 Yemen, Saudi Arabia Wadi 

Aedes vexans 

Culex tritaeniorhynchus 


camels, humans 

Rainfall and virus introduction 

2006-2007 

Kenya, Tanzania, 

Somalia 

Dambos ? 


humans 

Rainfall 

2007 Sudan Irrigated area ? 


humans 

? 

2007-2008 Mayotte Island ? 


humans 

Virus introduction 

2008 Madagascar 

Rice field in 

highlands 

Culex? 

Anopheles? 


humans 

? 

35

During the outbreak in Saudi Arabia in 2000, six viral 

strains of RVFV were isolated from Aedes mosquitoes. 

These strains were genetically close to the strain isolated 

in Kenya (1997-1998), suggesting that the virus 

was probably introduced into Saudi Arabia from the 

Horn of Africa by ruminants [45]. It remains unknown 

whether the virus has survived in Saudi Arabia since 

2000. In any event, the risk of re-introduction from the 

Horn of Africa is high. During the period of religious 

festivals in Mecca, 10 to 15 million small ruminants are 

imported from there to Saudi Arabia. 

A similar pattern in sheep trade is observed between 

sub-Saharan Africa and northern Africa. In the coming 

years, the Muslim feasts of Eid-ul-Fitr and Eid al-Adha 

will occur between September and November, i.e. when 

the activity of mosquito populations is high (end of the 

rainy season in Sahelian Africa) [46]. 

Therefore, the introduction of RVF-infected animals on 

the eastern and southern shores of the Mediterranean 

Sea is a likely event. Once introduced there, RVFV may 

find ruminant hosts, as well as competent mosquito 

species [47]. However, because livestock trade from 

northern Africa and the Middle East to Europe is forbidden, 

the introduction of RVF-infected animals to Europe 

looks unlikely [48]. 

Climate 

Climate warming is likely to have an impact on the 

geographical distribution of RVF. Higher temperatures 

increase mosquito feeding frequency and egg production 

and decrease the duration of their development 

cycle, as well as the extrinsic incubation period of RVFV 

in mosquitoes. Therefore, higher temperatures associated 

with increased rainfall may result in higher vector 

densities and vector competence and, subsequently, a 

higher RVFV transmission rate. In addition, transovarian 

transmission processes could be altered. 

If the virus were introduced to northern Africa or southern 

Europe, mosquitoes such as Ae. vexans could play 

a role as vectors in many Mediterranean countries. 

Several Ochlerotatus species, which breed in wetlands, 

might also be able to transmit the virus. Culex pipiens, 

a ubiquitous species, is locally abundant (in wetlands, 

rice fields, irrigated crops, sewers etc) and may act 

as an amplifier in the biological cycle. Increased temperatures 

could also have an impact on the vector 

competence and capacity of other endemic European 

mosquito species [49], although this is difficult to 

quantify (it has already been proved in controlled conditions 

with other arboviruses). Indeed, if introduced, 

several potential vector species that have so far not 

been investigated may become involved in the transmission 

of the RVFV. 

In East Africa, RVFV causes major epidemics at irregular 

intervals of 5-15 years. Climate models for this 

region predict an increase in the mean annual rainfall 

as well as an increase in the frequency and intensity 

of extreme rainfall events [50]. These changes may 

induce more severe and more frequent outbreaks in 

East Africa, which would thus represent a high risk 

area for neighbouring regions with livestock trade relationships 

such as the Indian Ocean islands. 


The flight capacities of Aedes and Culex mosquitoes are 

somewhat limited, ranging from a few hundred meters 

to more than 10 km [51,52]. However, these distances 

are long enough to allow a local spread of RVF. 

Wind transportation of infected mosquitoes has been 

reported for other arboviruses [53,54]. Presently, no 

information is available for RVFV vectors. Passive 

transportation of infected mosquitoes in boats or 

planes travelling from Africa has been reported for 

Anopheles mosquitoes infected by Plasmodium parasites 

[55]. However, for RVFV to be introduced this way, 

such infected mosquitoes would need to find susceptible 

hosts to initiate a local cycle. This event looks 

unlikely. 

Predictive models 

Risk mapping 

East Africa (Kenya) 

In Kenya, a correlation has been demonstrated between 

heavy rainfall events and the occurrence of RVF outbreaks. 

Maps of remotely sensed rainfall as well as 

vegetation index maps have been used together with 

ground data to monitor and predict vector population 

dynamics and RVFV activity and have established a 

correlation between these two parameters. The main 

advantage of remote sensing for the prediction of RVF 

occurrence in East Africa is the relatively low cost. It 

is readily available on a country and regional basis 

and its use may allow preventive measures to be taken 

such as the vaccination of susceptible livestock and 

the control of mosquito larvae [56,57]. 

Predictive models have been improved over the past 

decade through the addition of Pacific and Indian 

Ocean surface temperature anomalies and rainfall and 

normalised difference vegetation index (NDVI) data. An 

accuracy of 95-100% was estimated for the prediction of 

Kenyan epizootics of RVF, with a lead time of two to five 

months [57]. The FAO has used the technology to warn 

countries facing an increased risk of RVF. However, the 

geographic scope of these models is limited because 

ecological and epidemiological processes are different 

in other areas of Africa [58]. The outlook for the use 

of these models is even worse for the Mediterranean 

basin and Europe where climate determinants differ 

significantly from those of East Africa and the potential 

ecological and epidemiological processes are unknown 

as the disease has never been reported in these areas. 

West Africa (Senegal) 

RVF is endemic in the Ferlo area (northern Senegal) 

[59]. This area is characterised by a temporary pond 

ecosystem. These ponds are filled at the beginning 


of the rainy season (July) and dry up from October to 

January, according to their size and the intensity of 

rainfall, and are favourable environment to the development 

of Aedes mosquito populations. 

However, the East African model can not be applied 

in West Africa: abundant rainfall is not often associated 

with RVF outbreaks. The epidemiological process 

leading to RVF epidemics looks much more complex, 

involving the joint dynamics of hosts movements (tran- 


shumance), host immunity, and a vector population 

with brief activity during the rainy season. 

In this region, the risk of transmission was shown to 

be heterogeneous and linked to pond type [59]. A very 

high spatial resolution remote sensing image was used 

to characterise the temporary ponds and their environment 

and derive indices linked to mosquito biology 

[60]. However, this work is not advanced enough to be 

used in surveillance programmes. 

Table 4 

Competent mosquito vectors of Rift Valley fever virus with known distribution in the European Union and candidate 

countries 

Country Aedes vexans vexans Ochlerotatus caspius Culex theileri Culex pipiens Culex perexiguus 

Austria X X ? X ? 

Belgium X X ? X ? 

Bulgaria X X X X X 

Croatia 1 X X ? X ? 

Cyprus ? X ? X ? 

Czech Republic X X ? X ? 

Denmark X X ? X ? 

Estonia X X ? X ? 

Finland X X ? X ? 

France (mainland) X X X X ? 

France (Corsica) X X X X ? 

Germany X X ? X ? 

Greece X X X X X 

Hungary X X X X ? 

Ireland ? X ? X ? 

Italy (mainland) X X X X X 

Italy (Sardinia) X X X X ? 

Italy (Sicily) X X X X X 

Latvia X X ? X ? 

Lithuania X X ? X ? 

Luxembourg ? ? ? ? ? 

Former Yugoslav Republic of Macedonia 1 X X X X X 

Malta ? X ? X ? 

The Netherlands X ? ? X ? 

Poland X X ? X ? 

Portugal X X X X X 

Romania X X X X ? 

Slovakia X X X X ? 

Slovenia X X ? X ? 

Spain (mainland) X X X X X 

Spain (Balearic Islands) X ? ? X ? 

Sweden X X ? X ? 

Turkey 1 X X X X X 

United Kingdom X X ? X ? 

X: vector present; ?: unknown to the authors, or not found yet. 

1EU candidate country. 

Adapted from [1]. 

37

Risk analysis for Europe 

A detailed, qualitative risk analysis was performed in 

2005 by The European Food Safety Authority (EFSA) 

[1]. The main conclusions of this study are summarised 

below. 

Ruminant importations 

The importation of infected ruminants is the greatest 

hazard for RVF introduction to the European Union (EU). 

Clinical signs may not be observed rapidly in livestock 

living in remote, humid areas such as the Camargue 

region in France or the Danube delta in Romania. Such 

a scenario would allow RVFV to amplify and endemic 

foci to develop, if suitable ecological and entomological 

conditions were met [1]. 

Official RVF-free status is required for a country to 

export livestock and livestock meat to the EU. Such 

a status depends on a country’s ability – relying on 

observable evidences - to implement an efficient disease 

surveillance system and willingness to report possible 

RVF outbreaks. These constraints are the same as 

for foot-and-mouth disease and other epizootic diseases. 

They were instituted in 1972 (directive 72/462/ 

CEE [61], later modified to be more stringent). The practical 

consequence is that any introduction of live ruminants 

and their products from Africa and the Middle 

East to the European Union is forbidden. However, 

illegal and unknown ruminant importations probably 

occur between the Middle East and central Europe, and 

between northern Africa and southern Europe. This is 

also a major component of the risk of introduction of 

many other important animal and zoonotic diseases, 

like peste des petits ruminants, foot–and-mouth disease, 

bluetongue disease, Crimean-Congo haemorrhagic 

fever, etc. For instance, a risk analysis has 

recently been conducted to assess the risk of introduction 

of peste des petits ruminants virus (a Morbillivirus) 

from Maghreb to France. The conclusion was that the 

risk was extremely low, ranging from 0 to 2 on a scale 

from 0 (impossible event) to 9 (certain event) [48]. 


Several potential RVFV vectors are present in the EU 

(Tables 4 and 5). Differences in climate, seasonal variations 

of vector and host density, and genetic drift may 

result in differences in vector competence (the biological 

suitability of the vector to transmit the pathogen) 

and vectorial capacity (external factors such as 

number and lifespan of the vector, feeding preferences 

of the host) compared with the situation in Africa. 

Nevertheless, there is almost no doubt that several 

of the mosquito species in the EU, e.g. Cx. pipiens, 

would be competent vectors for RVF [62]. Moreover, 

the introduction and spread of new vector species 

represents a further risk. For example, Ae. albopictus 

can transmit RVFV [62-64], and many epidemiological 

concerns arise from this species’ current distribution 

in Europe: Albania, Bosnia and Herzegovina, Croatia, 

Italy (including Sicilia and Sardinia), south eastern continental 

France and Corsica, limited areas of Germany 

(north of the Alps), Greece, Monaco, Montenegro, the 

Netherlands (green houses), San Marino, Slovenia, 

eastern Spain, southern Switzerland, and the Vatican 

city [65]. 

Virus survival 

Blood, organs, fresh meat, fetal fluids and tissues as 

well as hides all represent a serious hazard to at-risk 

occupational groups (farmers, veterinarians, slaughterhouse 

employees, butchers, etc). The virus persists 

in the liver, spleen and kidneys, but rapidly disappears 

from meat as the pH decreases with meat maturation. 

The importance of blood, bone and offal meal products 

as a vehicle for RVFV has not been evaluated [4]. Milk 

is not considered to constitute a risk. However, due to a 

lack of data, transmission by ingestion of milk can not 

be definitively ruled out. 

Accidental RVF infections have been recorded in laboratory 

staff handling blood and tissues from infected 

animals. 

Conclusion 

Several national and Commission-supported analyses 

have been conducted to assess the risk of the introduction 

and spread of RVF within the EU. The conclusions 

have been that the overall risk was low. However, 

the recent reappearance of RVF in East Africa, including 

Sudan, the Nile Valley, and the Indian Ocean, has 

shown that the RVFV is very active and sensitive to 

climate and other environmental as well as socio-economic 

changes. These changes, together with growing 

human populations and an associated increased 

demand for meat, will promote greater controlled and 

uncontrolled movements of livestock. Consequently, 

the Mediterranean basin, central Europe, and the 

Middle East will probably be increasingly exposed to 

the risk of introduction of RVF. It is important to promote 

risk analyses that rely on accurate estimations 

of livestock movements between endemic and RVFfree 

areas. Moreover, high-risk ecosystems should be 

catalogued and the data updated on a regular basis to 

account for environmental changes. This latter activity 

has been initiated under the EU-funded Emerging 

Diseases in a changing European eNvironment (EDEN) 

project and should be continued once the project ends 

in 2010. Research programmes are needed to better 

characterise the bionomics of RVFV vectors in Europe 

and to develop RVFV introduction, installation, and 

spread models to improve disease surveillance and 

provide more efficient decision-making tools. 

Furthermore, more efficient vector and disease control 

methods are needed to enable the implementation of 

efficient contingency plans: 

- For vector control, a systematic assessment of existing 

methods and tools should be undertaken (laboratory 

and field experiments) and research programmes 

developing new technologies should be supported, 

including options for the development of genetically 

modified mosquitoes designed either to reduce 


population sizes or to replace existing populations 

with vectors unable to transmit the disease. 

- For disease control in European ruminants, the existing 

vaccines should be tested, preferably in collaboration 

with pharmaceutical companies. Because the 

cheapest and most efficacious existing vaccine (the 

Smithburn RVFV strain) has residual pathogenic effects 

in ruminants and humans, research on new-generation 

vaccines (e.g. recombinant, or reverse-genetic vaccines) 

should also be supported, both for human and 

animal populations. 

- Because a large-scale RVF epidemic appears unlikely 

in Europe (where a low proportion of people have direct 

contact with ruminants and their body fluids), human 

vaccination should target the population subgroups at 

high risk of exposure (farmers, veterinarians, slaughterhouse 

employees, butchers etc), once human vaccines 

have been developed. 

- Finally, the most relevant long term strategy is to 

control RVF where it is endemic. A substantial effort is 

needed to better understand the bio-ecology of RVFV 

vectors and viruses and epidemiological processes in 

Africa, to develop predictive and quantitative risk models 

and maps, and to implement risk-based surveillance 

and control methods. 


This review was conducted in the framework of the European 

Centre for Disease Prevention and Control funded V-borne 

project “Assessment of the magnitude and impact of vector-borne 

diseases in Europe”, tender no. OJ/2007/04/13- 

PROC/2007/003. It was compiled, edited and reviewed using 

EDEN and Arbozoonet funding, (EU grant GOCE-2003-010284 

EDEN; EU grant agreement 211757). The paper is catalogued 

by the EDEN steering committee as EDEN0197 (http://www. 

eden-fp6project.net/). The contents of this publication are 

the responsibility of the authors and do not necessarily reflect 

the views of either the European Centre for Disease 

Prevention and Control or the European Commission. 

* Erratum: This reference was corrected on 18 March. 

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Leishmaniasis emergence in Europe 

P D Ready (P.Ready@nhm.ac.uk) 1 

1. Department of Entomology, Natural History Museum, London, United Kingdom 


Citation style for this article: Ready PD. Leishmaniasis emergence in Europe. Euro Surveill. 2010;15(10):pii=19505. Available online: http://www.eurosurveillance. 

org/ViewArticle.aspx?ArticleId=19505 

Leishmaniasis emergence in Europe is reviewed, 

based on a search of literature up to and including 

2009. Topics covered are the disease, its relevance, 

transmission and epidemiology, diagnostic methods, 

treatment, prevention, current geographical distribution, 

potential factors triggering changes in distribution, 

and risk prediction. Potential factors triggering 

distribution changes include vectorial competence, 

importation or dispersal of vectors and reservoir 

hosts, travel, and climatic/environmental change. The 

risk of introducing leishmaniasis into the European 

Union (EU) and its spread among Member States was 

assessed for the short (2-3 years) and long term (15-20 

years). There is only a low risk of introducing exotic 

Leishmania species because of the absence of proven 

vectors and/or reservoir hosts. The main threat comes 

from the spread of the two parasites endemic in the EU, 

namely Leishmania infantum, which causes zoonotic 

visceral and cutaneous leishmaniasis in humans and 

the domestic dog (the reservoir host), and L. tropica, 

which causes anthroponotic cutaneous leishmaniasis. 

The natural vector of L. tropica occurs in southern 

Europe, but periodic disease outbreaks in Greece (and 

potentially elsewhere) should be easily contained by 

surveillance and prompt treatment, unless dogs or 

other synanthropic mammals prove to be reservoir 

hosts. The northward spread of L. infantum from the 

Mediterranean region will depend on whether climate 

and land cover permit the vectors to establish 

seasonal biting rates that match those of southern 

Europe. Increasing dog travel poses a significant risk 

of introducing L. infantum into northern Europe, and 

the threat posed by non-vectorial dog-to-dog transmission 

should be investigated. 

Leishmaniasis 

Leishmaniasis (or ‘leishmaniosis’) is a complex of 

mammalian diseases caused by parasitic protozoans 

classified as Leishmania species (Kinetoplastida, 

Trypanosomatidae) [1,2]. Natural transmission may 

be zoonotic or anthroponotic, and it is usually by the 

bite of a phlebotomine sandfly species (order Diptera, 

family Psychodidae; subfamily Phlebotominae) of 

the genera Phlebotomus (Old World) and Lutzomyia 

(New World) [2,3]. Primary skin infections (cutaneous 

leishmaniasis) sometimes resolve without treatment, 

with the host developing acquired immunity 



through cellular and humoral responses [4], but the 

infection can spread to produce secondary lesions in 

the skin (including diffuse cutaneous leishmaniasis), 

the mucosa (muco-cutaneous leishmaniasis) and the 

spleen, liver and bone marrow (visceral leishmaniasis, 

which is usually fatal if untreated) [1]. Worldwide, at 

least 20 Leishmania species cause cutaneous and/or 

visceral human leishmaniasis (HumL) [1,5]. Most foci 

occur in the tropics or subtropics, and only zoonotic 

L. infantum is transmitted in both the eastern and 

western hemispheres [5] (Table 1). 

Worldwide and European 

relevance of leishmaniasis 

The World Health Organization (WHO) reports that 

the public health impact of leishmaniasis worldwide 

has been grossly underestimated for many years [1]. 

In 2001 and 2004, Desjeux reported that in the previous 

decade endemic regions had spread, prevalence 

had increased and the number of unrecorded cases 

must have been substantial, because notification was 

compulsory in only 32 of the 88 countries where 350 

million people were at risk [5,6]. About two million 

new cases of HumL (half a million visceral) are considered 

to occur every year in the endemic zones of Latin 

America, Africa, the Indian subcontinent, the Middle 

East and the Mediterranean region [1]. 

Risks of emergence or re-emergence of leishmaniasis 

in Europe are associated with three main scenarios: 

1) the introduction of exotic Leishmania species or 

strains into Europe via the increasing worldwide travelling 

of humans [6] and domestic dogs [7], 

2) the natural spread of visceral and cutaneous leishmaniasis 

caused by L. infantum and L. tropica from 

the Mediterranean region of Europe, where these 

species are endemic [1,8,9], to neighbouring temperate 

areas where there are vectors without disease [2], 

3) the re-emergence of disease in the Mediterranean 

region of Europe caused by an increase in the number 

of immunosuppressed people. 

The high prevalence of asymptomatic human carriers 

of L. infantum in southern Europe [10-13] suggests that 

this parasite is a latent public health threat. This was 

demonstrated by the increase of co-infections with 

41

human immunodeficiency virus (HIV) and Leishmania 

that has been observed since the 1980s [14], with 

leishmaniasis becoming the third most frequent opportunistic 

parasitic disease after toxoplasmosis and 

cryptosporidiosis [15]. 

Disease transmission and epidemiology 

Visceral leishmaniasis (VL) is usually fatal if untreated, 

and so it is distinguished from cutaneous leishmaniasis 

(CL) in all sections of the current review. If untreated, 

uncomplicated CL is often disfiguring, but not fatal. 

In contrast, muco-cutaneous and diffuse cutaneous 

disease can lead to fatal secondary infections even 

if treated. Patient immunodeficiency is one factor for 

this, but in Latin America these diseases are associated 

with regional strains of the L. braziliensis and 

L. mexicana species complexes [1,5]. 

A female sandfly ingests Leishmania while blood-feeding, 

and then transmits the infective stages (usually 

accepted to be the metacyclic promastigotes) during a 

subsequent blood meal [16]. The infective promastigotes 

inoculated by the sandfly are phagocytosed in the 

mammalian host by macrophages and related cells, in 

which they transform to amastigotes and often provoke 

a cutaneous ulcer and lesion at the site of the bite. 

There are only two transmission cycles with proven 

long-term endemism in Europe [2,17]: zoonotic visceral 

and cutaneous HumL caused by L. infantum throughout 

the Mediterranean region; and, anthroponotic cutaneous 

HumL caused by L. tropica now occurring sporadically 

in Greece (Table 1, Figure 1). 

Worldwide, most transmission cycles are zoonoses, 

involving reservoir hosts such as rodents, marsupials, 

edentates, monkeys, domestic dogs and wild canids 

[2,5,6,18] (Table 1). However, leishmaniasis can be 

anthroponotic, with sandflies transmitting parasites 

between human hosts without the involvement of a 

reservoir host. Anthroponotic transmission is characteristic 

of species of the L. tropica complex and, 

except for L. infantum, of the L. donovani complex. 

One species (L. donovani sensu stricto) or two species 

(L. donovani and L. archibaldi) cause periodic epidemics 

of anthroponotic visceral leishmaniasis (‘Kala-azar’) 

in India and northeast Africa, respectively [19]. Sandfly 

vectors of both complexes (L. donovani and L. tropica) 

are abundant in southern Europe. 

The domestic dog is the only reservoir host of major 

veterinary importance, and in Europe there is a 

large market for prophylactic drugs and treatment of 

canine leishmaniasis (CanL) caused by L. infantum [2]. 

Domestic cats might be secondary reservoir hosts of 

L. infantum in southern Europe [13], because they are 

experimentally infectious to sandflies [20] and natural 

infections can be associated with feline retroviruses 

[21]. 

Fewer than 50 of the approximately 1,000 species of 

sandflies are vectors of leishmaniasis worldwide [3]. 

This is due to the inability of some sandfly species to 

support the development of infective stages in their 

gut [16] and/or a lack of ecological contact with reservoir 

hosts [22]. Our understanding of the fundamentals 

of leishmaniasis epidemiology has been challenged 

in the last 20 years. Firstly, HIV/Leishmania co-infections 

were recorded in 35 countries worldwide, and 

widespread needle transmission of L. infantum was 

inferred in southwest Europe [15], where Cruz et al. 

demonstrated Leishmania in discarded syringes [23]. 

Secondly, leishmaniasis has become more apparent 

in northern latitudes where sandfly vectors are either 

absent or present in very low densities, such as in the 

eastern United States (US) and Canada [24] as well as 

in Germany [25-27]. Most infections involve CanL, not 

HumL, and this might be explained by dog importation 

from, or travel to, endemic regions, followed by vertical 

transmission from bitch to pup or horizontal transmission 

by biting hounds [24]. Vertical transmission of 

HumL from mother to child has rarely been reported 

[28]. 

Diagnostic methods 

Most diagnoses are only genus-specific, being based 

on symptoms, the microscopic identification of parasites 

in Giemsa-stained smears of tissue or fluid, and 

serology [18,29]. Consequently, the identity of some 

causative agents has only been known relatively 

recently, following typing performed during limited ecoepidemiological 

surveys. For example, it was thought 

that all cutaneous leishmaniasis cases in Europe were 

caused by L. tropica, until Rioux and Lanotte reported 

L. infantum to be the causative agent in the western 

Mediterranean region [30]. 

Rioux and Lanotte used multi-locus enzyme electrophoresis 

(MLEE) to identify Leishmania species and 

strains [30], which remains the gold standard [1,18]. 

However, MLEE requires axenic culture [31] in which 

one strain can overgrow others in mixed infections. It 

is therefore more practical to identify the isoenzyme 

strains (or zymodemes) by directly characterising the 

enzyme genes [32]. Other molecular tests have been 

used to identify Leishmania infections in humans, 

reservoir hosts and sandfly vectors [33], including in 

the Mediterranean region [34], but there has been no 

international standardisation [29]. However, PCR of the 

internal transcribed spacer of the multi-copy nuclear 

ribosomal genes is often used [34,35]. A set of carefully 

chosen criteria must accompany PCR-based diagnosis, 

especially for immunocompromised patients 

[14,15]. Monoclonal antibodies have long been available 

for the identification of neotropical species [36] 

and the serotyping of Old World species [37] but they 

are not widely used. 

Most sensitive molecular techniques indicate only the 

presence of a few recently living Leishmania, not that 

the parasites were infectious. Therefore, serology is 


often more informative [29]. However, antigens prepared 

in different laboratories can cause test variation 

for the frequently used methods [29]: the indirect 

fluorescent antibody test (IFAT), the enzyme-linked 

immunosorbent assay (ELISA), the indirect haemagglutination 

assay (IHA) and the direct agglutination test 

(DAT). Some antigens are stable and produced commercially, 

such as the recombinant (r) K39 for a dipstick 

or strip test. Multi-centre studies of ‘Kala-azar’ diagnostics 

[38] showed that both the freeze-dried DAT and 

the rK39 strip test could exceed the 95% sensitivity and 

90% specificity target, but only for the strains found in 

some regions. Antibody detection tests should complement 

other diagnostic tests, because they do not usually 

distinguish between acute disease, asymptomatic 

infections, relapses and cured cases [38]. 

Delayed hypersensitivity is an important feature of 

all forms of human leishmaniasis [4] and is often 

measured by the leishmanin skin test (or Montenegro 

reaction) [29]. False-positivity is approximately 1% in 

otherwise healthy people. Other problems with this 

test include the absence of commercially available 

leishmanins, that there is complete cross-reactivity 

among most species and strains of Leishmania, and 

that for VL its applicability is limited to the detection 

of past infections, because a complete anergy is found 

during active disease. 

Treatment 

Pentavalent antimonials were the first-choice drugs 

for leishmaniasis worldwide [39,40]. Miltefosine, 

Paromomycin and liposomal Amphotericin B are 

gradually replacing antimonials and conventional 

Amphotericin B in some regions [40,41], especially 

where there is drug resistance or the need to develop 

combination therapy to prevent the emergence of 

resistance to new drugs [41]. 

Highly active anti-retroviral therapy (HAART) treatment 

has reduced the incidence of co-infections with 

Leishmania and HIV by preventing an asymptomatic 

infection with L. infantum from becoming symptomatic, 

but unfortunately it is not good at preventing visceral 

leishmaniasis relapses. The benefits of treatment are 

not as clear-cut as they are for other opportunistic diseases 

[42]. 

Prevention 

Most research on vaccines is strategic, not applied, for 

example targeting secretory-gel glycans of Leishmania 

[43] and some sandfly salivary peptides [44], both of 

which are injected into the mammalian host by the 

female sandfly during blood feeding. Therapeutic vaccine 

trials continue to use killed cultured parasites 

(often with BCG as adjuvant) in combination with antileishmanial 

drugs but with only 0-75% efficacy [45]. 

One second generation recombinant vaccine contains a 

trifusion recombinant protein (Leish-111f), and some of 

its epitopes are shared by L. donovani and L. infantum 

[46]. 


Research and development of vaccines against CanL 

has been stimulated by the economic importance of 

dogs and their role as reservoirs of HumL caused by 

L. infantum in the Americas and the Mediterranean 

region. Leishmune is the first licensed vaccine against 

CanL. It contains the fucose-mannose ligand (FML) 

antigen of L. donovani and has a reported efficacy of 

76-80% [47]. The industrialised formulation of FMLsaponin 

underwent safety trials in Brazil [48]. The 

vaccine LiESAp-MDP (excreted/secreted antigens with 

adjuvant) was reported to have an efficacy of 92% 

when tested on naturally exposed dogs in the south 

of France [49,50]. More recently, a modified vaccinia 

virus Ankara (MVA) vaccine expressing recombinant 

Leishmania DNA encoding tryparedoxin peroxidase 

(TRYP) was found to be safe and immunogenic in outbred 

dogs [51]. 

One means of controlling transmission is to reduce 

the biting rate of peri-domestic sandfly vectors of visceral 

HumL and CanL. This has been effective locally, 

by using repellents [52], insecticide-impregnated nets 

and bednets [52], topical applications of insecticides 

[53] and deltamethrin-impregnated dog collars [54,55]. 

The latter are favoured by many dog owners in southern 

Europe. 

Current geographical distribution 

Outside the European Union 

Table 1 (updated from Ready [2]) relates the distributions 

of each form of HumL to causative species and 

known reservoir hosts [1,5,6,17,18]. Most VL foci occur 

in India and neighbouring Bangladesh and Nepal, 

and in Africa (Sudan and neighbouring Ethiopia and 

Kenya), where anthroponotic ‘Kala-azar’ is caused by 

L. donovani and in north-eastern Brazil and parts of 

Central America, where zoonotic infantile visceral 

leishmaniasis is caused by L. infantum. Most CL foci 

occur in Latin America, North Africa, and the Middle 

East, and muco-cutaneous and diffuse cutaneous disease 

are frequent in South America [56]. 

Inside the European Union: main biomes 

Only two transmission cycles have been endemic in the 

European Union (EU) for a long time, and both are widespread 

in the adjoining Middle East and in North Africa: 

zoonotic cutaneous and visceral leishmaniasis caused 

by L. infantum throughout the Mediterranean region 

and anthroponotic cutaneous leishmaniasis caused 

by L. tropica, which occurs sporadically in Greece and 

probably neighbouring countries and poses a high 

risk of introduction by migrants and travellers into the 

rest of the EU [2,6,17] (Table 1, Figure 1). The former is 

endemic and sandfly-borne only in the Mediterranean 

region of the EU (‘Mediterranean forests’ biome), 

where its epidemiological significance is clear from 

published serological surveys [7,8]. However, the vectors 

of L. infantum [57] (Figure 2, Table 2, updated from 

Ready [2]) are also abundant in the adjoining parts of 

the temperate region (Temperate broadleaf forests’ 

biome), in northern Spain [58] and central France [59], 

43

Table 1 

European distribution of parasites causing most human leishmaniasis worldwide up to 2009 

Diffuse cutaneous 

leishmaniasis 

Muco-cutaneous 

leishmaniasis 

Cutaneous 

leishmaniasis 

Visceral 

leishmaniasis 

Visceral 

leishmaniasis 

Cutaneous 

leishmaniasis 

Cutaneous leishmaniasis 

(Diffuse and muco-) 

cutaneous leishmaniasis 

Human disease 

L. mexicana species 

complex 

L. braziliensis 

L. braziliensis species 

complex.; L. mexicana 

Other members of 

L. donovani 

species complex 

L. infantum (= L. chagasi 

in Neotropics) 

L. infantum (= L. chagasi 

in Neotropics) 

L. major 

L. tropica species 

complex 

Leishmania species 

species complex 

Rodents, marsupials 

Rodents, marsupials 

Edentates, primates, 

rodents, marsupials 

Anthroponotic 

Domestic dogs, wild 

canids 

Domestic dogs, wild 

canids 

Often anthroponotic Rodents 

Reservoir hosts (zoonosis) or anthroponosis 

Neotropical, Nearctic Neotropical Neotropical 

Afrotropical, Indo- 

Malayan 

Palaearctic, Afrotropical, 

Neotropical 


Neotropical 


Indo-Malayan 

Palaearctic, 

Afrotropical, Indo- 

Malayan 

World ecozone 

Absent Absent Absent Absent 

Mediterranean 

forests, temperate 

Mediterranean 

forests, temperate 

Absent 

Mediterranean 

forests 

EU biome 

broadleaf forest 


EU: Cyprus Absent Absent Present Present Present Absent Absent Absent 

EU: France Absent Absent Present Present Absent Absent Absent Absent 

EU: Germany Absent Absent Absent Absent Absent Absent Absent Absent 

EU: Greece Sporadic Absent Present Present Absent Absent Absent Absent 


EU: Hungary Absent Absent Absent Absent Absent Absent Absent Absent 

EU: Italy Absent Absent Present Present Absent Absent Absent Absent 

EU: Malta Absent Absent Present Present Absent Absent Absent Absent 

EU: Portugal Absent Absent Present Present Absent Absent Absent Absent 

Formerly sporadic? 


EU: Romania 

Absent 

Formerly present Absent Absent Absent Absent 

(untyped parasites) 


EU: Spain Absent Absent Present Present Absent Absent Absent Absent 

EU Overseas Territories: French 

Absent Absent Absent Absent Absent Present Present Present 

Guyana 

EU candidate: Former Yugoslav Formerly sporadic? 


Absent 

Present Absent Absent Absent Absent 

Republic of Macedonia 


(confirmed in Croatia) 

EU candidate: European Turkey 

Absent? (present) Absent (absent) Absent? (present) Absent? (present) Absent Absent Absent Absent 

(Asiatic Turkey) 

Other Europe: Albania Absent Absent Present Present Absent Absent Absent Absent 

Other Europe: Switzerland Absent Absent Absent Absent Absent Absent Absent Absent 

Adapted from a contribution [2] published in the OIE (World Organisation for Animal Health) Scientific and Technical Review: In Climate 

change: the impact on the epidemiology and control of animal diseases (S. de la Roque, ed.). Rev Sci Tech Off Int Epiz. 2008;27(2):399-412.

and small numbers occur as far north as Paris [60] and 

the upper Rhine valley in Germany [26]. The occurrence 

of ‘vectors without disease’ poses a significant risk for 

the emergence of leishmaniasis in temperate regions 

of Europe [2]. 

Potential factors triggering changes in 

distribution 

Climate change 

Most transmission of Leishmania is by the bite of permissive 

sandflies, and so climate change might affect 

leishmaniasis distribution directly, by the effect of 

temperature on parasite development in female sandflies 

[16], or indirectly by the effect of environmental 

variation on the range and seasonal abundances of the 

vector species. Female sandflies seek sheltered resting 

sites for blood meal digestion, and in southern Europe 

the temperatures of these micro-habitats are buffered 

but vary significantly with the external air temperature 

[2]. 

Based on molecular markers, European vectors of 

leishmaniasis have extended their ranges northward 

since the last ice age (approximately 12,000 years ago) 

[61,62], and the mapping of statistical measures of climate 

has permitted transmission cycles to be loosely 

associated with some Mediterranean bioclimates [63]. 

However, bioclimate zones and their vegetation indicators 

vary regionally, and ongoing climate change may 

alter the patterns of land cover and land use. The geographic 

information system (GIS)-based spatial modelling 

of the Emerging Diseases in a changing European 

Environment (EDEN) project is permitting an analysis of 


changes in climate and land cover [64] and their effects 

on sandflies. 

The project ‘climate Change and Adaptation Strategies 

for Human health in Europe’ (cCASHh) concluded: 

“There is no compelling evidence, due to lack of historical 

data, that sand fly and VL distributions in Europe 

have altered in response to recent climate change” [9]. 

There is now a published analysis of the northward 

spread of CanL and its vectors in Italy [65], but an association 

with climate change was only surmised. 

Capacity and competence of vectors in Europe 

Vectorial capacity has only been calculated indirectly. 

The average number of gonotrophic cycles (i.e. egg 

development following a bloodmeal) completed by 

P. ariasi in the south of France was only a little greater 

than one [66]. Therefore, relatively small changes 

in temperature could have a large effect on vectorial 

capacity, because transmission occurs only during the 

second or subsequent bloodmeals and temperature 

affects the level of activity of the sandfly and therefore 

the frequency of the bloodmeals. 

Alone, PCR detection of a natural infection of 

Leishmania in a sandfly does not identify a vector. It 

only indicates that the sandfly has fed on an infected 

mammalian host [35] because many parasites do not 

survive in a non-permissive sandfly after bloodmeal 

defecation [16]. 

Vectorial competence has been tested [67] or inferred 

based on finding naturally infected females of the more 

Figure 1 

Distribution by country of Leishmania species transmitted by phlebotomine sandflies in Europe up to 2009 

L. infantum L. tropica 

Absent Present Sporadic or untyped infections Untyped infections 

Presence in North Africa and Middle East not depicted. 

Source: V-borne project; reproduced with permission from the European Centre of Disease Prevention and Control. 

45

Figure 2 

Distribution of vectors of leishmaniasis in European countries up to 2009 

Phlebotomus ariasi P. perniciosus 

P. sergenti P. perfiliew 

P. neglectus P. tobbi 

Present 

Presence in North Africa and Middle East is not depicted. 

Absent Old record 

Source: V-borne project; reproduced with permission from the European Centre of Disease Prevention and Control. 


abundant human-biting species [3,57,68], from which it 

is concluded that the principal vectors of L. infantum in 

the Mediterranean region are members of the subgenus 

Larroussius (Table 2, Figure 2). The vectorial competence 

of Phlebotomus (Transphlebotomus) mascittii 

should be tested because this species is now known 

to be widespread in northern France, Belgium and 

Germany [69]. However, low rates of biting humans and 

autogeny (the ability to produce eggs without a bloodmeal) 

cast doubt on its epidemiological importance [2]. 

Based on distribution and vectorial competence elsewhere, 

P. sergenti sensu lato is likely to be the main 

vector of L. tropica in southern Europe [3]. 

Importation or dispersal of 

vectors and reservoir hosts 

The importation or inter-continental dispersal of vectors 

is unlikely because sandflies are not as robust as some 

mosquitoes and are not known to be wind-dispersed 

[3]. Any importations are unlikely to be significant 

for several reasons: The natural vectors of Old World 

leishmaniasis are already abundant in Mediterranean 

Europe (Table 2, Figure 2); most American sandflies are 

believed to be poor vectors of Old World Leishmania 

species [3]; and Leishmania species native to the 

Americas have hosts that do not occur in Europe [56]. 

Table 2 

European distributions of sandfly vectors of human leishmaniasis up to 2009 (unproven role throughout range) 

Leishmania species 

Human disease 


L. tropica species complex 

- Greece only 

(Diffuse and muco-) 

cutaneous leishmaniasis 

L. major 

EU biome Mediterranean forests Absent 

The vector of L. major in North Africa and the Middle 

East is P. papatasi, which is locally abundant in southern 

Europe. However, the natural reservoir hosts of 

this parasite are usually gerbil species not present in 

EU countries [18] and the risks of them dispersing into 

southern Europe or surviving accidental/deliberate 

release by humans have not been assessed. 

Importance of travel within Europe (mainland 

and overseas territories) and internationally 

Travel has led to increasing numbers of HumL cases that 

need to be treated, e.g. in France [70], Germany [25], 

Italy [71] and the United Kingdom [72]. Leishmaniasis 

in Guyana (overseas region of France) is a major source 

of exotic cases imported to mainland France, and 

L. infantum has travelled in the reverse direction in a 

dog [73]. Isoenzyme [12] and molecular markers [32,34] 

can sometimes identify the origins of Leishmania 

strains. 

Travel poses the risk of the emergence in southern 

Europe of anthroponotic L. donovani [74] and L. tropica 

(see above), and the introduction to northern Europe of 

L. infantum in dogs taken to the Mediterranean region 

on holiday or rescued from there as strays [7]. 

L. infantum (= L. chagasi in 

Neotropics) - Mediterranean 

region only 

L. infantum (= L. chagasi in 

Neotropics) - Mediterranean 

region only 

Cutaneous leishmaniasis Cutaneous leishmaniasis Visceral leishmaniasis 

Mediterranean forests, Temperate 


Mediterranean forests, Temperate 


EU: Cyprus P. sergenti s.l. P. papatasi P. perfiliewi s.l., P. tobbi P. perfiliewi s.l., P. tobbi 

EU: France P. sergenti s.l. P. papatasi 

P. ariasi, P.perniciosus, 

P. perfiliewi ? 

P. ariasi, P. perniciosus 

EU: Germany No vectors No vectors P. perniciosus P. perniciosus 

EU: Greece P. sergenti s.l. P. papatasi P. perfiliewi s.l., P. tobbi 

P. perfiliewi, P. tobbi, 

P. neglectus 

EU: Hungary No vectors? No vectors? P. neglectus, P. perfiliewi ? P. neglectus, P. perfiliewi ? 

EU: Italy P. sergenti s.l. P. papatasi 

P. ariasi P. perfiliewi, 

P. perniciosus, P. neglectus 

P. ariasi, P. perfiliewi, 

P. perniciosus,P. neglectus 

EU: Malta P. sergenti s.l. P. papatasi 

P. perfiliewi, P. perniciosus, 

P. neglectus 

P. perfiliewi, P. perniciosus, 

P. neglectus 

EU: Portugal P. sergenti s.l. P. papatasi P. ariasi, P. perniciosus P. ariasi, P. perniciosus 

EU: Romania No vectors? P. papatasi P. perfiliewi, P. neglectus P. perfiliewi 

EU: Spain P. sergenti s.l. P. papatasi P. ariasi, P. perniciosus P. ariasi, P. perniciosus 

EU candidate: Former Yugoslav 

Republic of Macedonia 

P. sergenti s.l. P. papatasi 


P. neglectus 


P. neglectus 

EU candidate: European 

Turkey (Asiatic Turkey) 

(P. sergenti s.l.) (P. papatasi) 

(P. perfiliewi s.l., P. tobbi, 

P. neglectus) 

(P. perfiliewi s.l., P. tobbi, 

P. neglectus) 

Other Europe: Albania P. sergenti s.l. P. papatasi 


P. neglectus 


P. neglectus 

Other Europe: Switzerland 

No vectors No vectors P. perniciosus P. perniciosus 

Adapted from a contribution [2] published in the OIE (World Organisation for Animal Health) Scientific and Technical Review: In Climate 

change: the impact on the epidemiology and control of animal diseases (S. de la Roque, ed.). Rev Sci Tech Off Int Epiz. 2008;27(2):399-412. 

47

Changes in environments (e.g. urbanisation, 

deforestation) and socio-economic patterns 

Deforestation and urbanisation are known to affect 

leishmaniasis worldwide [6] because of the associations 

of many vectors and reservoirs with natural or 

rural areas. Based on the EDEN partners’ findings, 

most Mediterranean regions have at least one vector 

associated more closely with rural or peri-urban zones 

[64]. From 1945, most of the socio-economic changes 

favoured a reduction in ‘infantile visceral leishmaniasis’ 

(caused by L. infantum) in southern Europe, including 

better nutrition, widespread insecticide spraying 

(against malaria-transmitting mosquitoes), better 

housing and a reduction in the rural population. The 

last 20 years have seen changes that have increased 

contact with the Mediterranean vectors, including more 

holidays and second homes for northern Europeans, 

unforeseen modes of transmission (among intravenous 

drug users), and immunosuppression (HIV/Leishmania 

co-infections). The latter is highest in south-western 

Europe [15]. 

Risk prediction models 

The logic of visceral leishmaniasis control 

Based on compartmental mathematical (R 0 ) models, 

Dye [75] concluded that insecticides can be expected 

to reduce the incidence of HumL caused by L. infantum 

even more effectively than they reduce the incidence of 

CanL, but only where transmission occurs peridomestically 

and the sandfly vectors are accessible to treatment, 

as in parts of Latin America. For control of HumL 

and CanL in Europe, Dye [75] concluded that a dog vaccine 

is highly desirable, because sandfly vectors here 

are less accessible to insecticide treatment. In Europe, 

CanL is a veterinary problem with socio-economic 

importance and a vaccine is more likely to be afforded 

than elsewhere. 

Risk assessment of introduction, 

establishment and spread in the European 

Union (EU) for the short term (2-3 years) 

‘Oriental sore’ caused by L. tropica is usually anthroponotic, 

and it is sporadically endemic in Greece and 

endemic in neighbouring countries to the EU. The 

principal vector (P. sergenti s.l.) is locally abundant in 

southern Europe, where new foci could be initiated by 

people infected in North Africa and the Middle East, 

including members of the European armed forces based 

in Iraq and Afghanistan [76,77]. Recently, L. donovani 

has been introduced to Cyprus [74]. Good surveillance, 

followed by prompt diagnosis and treatment should be 

extended to all areas of high risk, in order to help prevent 

the emergence of anthroponotic leishmaniasis. 

Cutaneous leishmaniasis caused by the Old World parasite 

L. major has a low risk of emergence as a sandflyborne 

disease in southern Europe in the short and long 

term, even though its principal vector (P. papatasi) is 

locally abundant, because its main gerbil reservoir 

hosts are absent. 

Cutaneous leishmaniases caused by the American parasites 

of the L. braziliensis and L. mexicana complexes 

have low risks of emergence as sandfly-borne diseases 

in southern Europe in the short and long term because 

of the absence of their exotic vectors and mammalian 

reservoir hosts. 

However, all these parasites pose a significant risk 

of introduction to Europe by intravenous drug users 

(iVDUs) and the establishment of local transmission 

by syringe needles, especially if these patients have 

HIV co-infections. This is based on the experience with 

endemic visceral leishmaniasis caused by L. infantum 

[15,42]. 

Risk assessment of introduction, 

establishment and spread in the European 

Union for the long term (15-20 years) 

Leishmania infantum is currently the only significant 

causative agent of visceral and cutaneous HumL 

endemic in Europe. Its high prevalence in asymptomatic 

humans and in the widespread reservoir host 

(the domestic dog) means there is a high risk of emergence 

in parts of Europe further north, as demonstrated 

in northern Italy [65]. In addition to risk factors 

[78] and statistical models [64] with associated risk 

maps, EDEN is producing R0 mathematical models as 

part of research to explain why large regions of temperate 

Europe have sandflies without HumL in the 

presence of imported CanL. Some of the key data come 

from questionnaires to veterinary clinics, validated by 

prospective serological surveys of CanL, from northern 

and southern areas with a wide range of disease 

prevalence. 

Increasing dog travel poses a significant risk of introduction 

of L. infantum into northern Europe from the 

Mediterranean region. There is also a risk of establishment 

of non-vector transmission and spread as 

has been observed in North America [24]. Non-vector 

transmission might explain the autochthonous cases 

of CanL in Germany [25, 26]. 

L. tropica has been isolated from both the domestic 

dog and the black rat [5,8], and so the risk of introduction 

and spread of CL caused by this parasite in the 

EU should be re-assessed if either these mammals or 

related synanthropic species were found to be reservoir 

hosts (rather than dead-end hosts) in the disease 

foci in North Africa and southwest Asia [35]. 

Assessment of whether the existing 

data sources are adequate and, if not, 

identification of missing key data needed 

for conducting risk assessment studies 

Research data about leishmaniasis and its spatial 

distribution in Europe and the Mediterranean region 

are being enhanced [79] and made accessible online 

by EDEN and another EU-funded project, LeishRisk, 

which has collaborated with the WHO to produce an 


E-compendium, a compilation of peer-reviewed literature 

on leishmaniasis epidemiology [1,80]. 

However, public health and veterinary surveillance data 

are more fragmentary, which undoubtedly caused the 

public health impact of leishmaniasis to be underestimated 

for many years in Europe as well as worldwide 

[1]. The WHO has concluded that more surveillance is 

necessary in Europe to assess an emergence of leishmaniasis 

[9], but the partners of the EDEN leishmaniasis 

sub-project have stressed the need for better coordination 

of existing surveillance, including linking human 

health and veterinary data for the zoonotic disease. 

Currently (EDEN partners, personal communications), 

HumL is notifiable in Greece, Italy, Portugal and Turkey, 

and in endemic autonomous regions in Spain. CanL is 

notifiable in Greece and at municipality level in the 

endemic regions of the four other countries mentioned 

above. Neither disease is notifiable in France. At international 

level, WHO organised a meeting of Eurasian 

countries in 2009 (J. Alvar, personal communication), 

aimed at standardising surveillance and reporting, and 

CanL is reported as a listed disease (‘other diseases’) 

of the World Organisation for Animal Health (OIE) [81]. 

The monitoring of dog travel [7] should continue to be 

improved and standardised. 


This review is based on an extensive literature study conducted 

as part of the European Centre for Disease Prevention 

and Control funded V-borne project: “Assessment of the 

magnitude and impact of vector-borne diseases in Europe”, 

tender n°o. OJ/2007/04/13-PROC/2007/003. It was compiled 

using EDEN funding, EU grant GOCE-2003-010284 EDEN. 

The paper is catalogued by the EDEN Steering Committee as 

EDEN0199 (http://www.eden-fp6project.net/). The contents 

of this publication are the responsibility of the author and 

do not necessarily reflect the views either of the ECDC or of 

the European Commission. I thank all partners of the EDEN 

leishmaniasis sub-project for information and support. 

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en_classification2007.htm?e1d7 


51


Arthropod-borne viruses transmitted by Phlebotomine 

sandflies in Europe: a review 

J Depaquit (jerome.depaquit@univ-reims.fr) 1 , M Grandadam 2,3 , F Fouque 4 , PE Andry1, C Peyrefitte 5 

1. Université de Reims Champagne Ardenne, AFSSA, JE 2533-USC “VECPAR”, Reims, France 

2. Laboratoire associé au Centre national de référence des arbovirus, Unité de virologie tropicale, Institut de médecine tropicale 

du Service de santé des armées, Marseille, France 

3. Centre national de référence des arbovirus, Institut Pasteur, Paris, France 

4. Cellule d’Intervention Biologique d’Urgence (CIBU), Institut Pasteur, Paris, France 

5. Unité de virologie, Département de Biologie des Agents transmissibles, Centre de Recherche du Service de Santé des Armées, 

Grenoble, France 


Citation style for this article: Depaquit J, Grandadam M, Fouque F, Andry P, Peyrefitte C. Arthropod-borne viruses transmitted by Phlebotomine sandflies in Europe: 

a review . Euro Surveill. 2010;15(10):pii=19507. Available online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19507 

Phlebotomine sandflies are known to transmit leishmaniases, 

bacteria and viruses that affect humans 

and animals in many countries worldwide. These 

sandfly-borne viruses are mainly the Phlebovirus, the 

Vesiculovirus and the Orbivirus. Some of these viruses 

are associated with outbreaks or human cases in the 

Mediterranean Europe. In this paper, the viruses transmitted 

by Phlebotomine sandflies in Europe (Toscana 

virus, Sicilian virus, sandfly fever Naples virus) are 

reviewed and their medical importance, geographical 

distribution, epidemiology and potential spreading 

discussed. Data on vertebrate reservoirs is sparse for 

sandfly fever viruses. The factor currently known to 

limit the spread of diseases is mainly the distribution 

areas of potential vectors. The distribution areas of 

the disease may not be restricted to the areas where 

they have been recorded but could be as wide as 

those of their vectors, that is to say Larroussius and 

P. papatasi mainly but not exclusively. Consequently, 

field work in form of viral isolation from sandflies 

and possible reservoirs as well as laboratory work to 

establish vectorial competence of colonised sandflies 

need to be encouraged in a near future, and epidemiological 

surveillance should be undertaken throughout 

the European Union. 

Introduction 

During the last decade, several cases of infections due 

to Toscana virus have been recorded in Europe (Italy, 

France, Spain, and Portugal). A few studies focusing 

on the viruses transmitted by Phlebotomine sandflies 

have been carried out. This review summarises the 

data related to arthropod-borne viruses transmitted by 

Phlebotomine sandflies in Europe. 

Phlebotomine sandflies are the vectors of the 

Leishmania, pathogens that cause diseases called 

leishmaniases in more than 80 countries in the Old and 

New World. Sandflies are also vectors of other human 

pathogens such as Bartonella and viruses belonging 


to three different genera: (i) the Phlebovirus (family 

Bunyaviridae) including sandfly fever Sicilian virus, 

sandfly fever Naples virus, Toscana virus and Punta 

Toro virus; (ii) the Vesiculovirus (family Rhabdoviridae) 

including Chandipura virus [2-3] and (iii) the Orbivirus 

(family Reoviridae) including Changuinola virus [1]. 

The latter viruses have been associated with several 

outbreaks in humans. Further less important 

viruses have also been found in Europe. Chios virus 

was isolated from a human case of severe encephalitis 

(Papa and Pavlidou, personal communication) 

in Greece and additionally three other viruses were 

isolated from Phlebotomine sandflies: Corfou virus 

from Phlebotomus (Larroussius) neglectus, in Greece 

[4], Massilia virus from P. (L.) perniciosus, in France 

[5] and Arbia virus from P. (L.) perniciosus and from 

P. (L.) perfiliewi in Italy. However, so far there are no 

reports of human disease from these viruses. 

Little is known about the viruses transmitted by 

Phlebotomine sandflies and they can be, to our opinion, 

considered as neglected pathogens. However, the 

Toscana virus, sandfly fever Naples and Sicilian viruses 

are endemic in the Mediterranean region and could 

spread to more temperate areas in Europe where vectors 

are abundant. Moreover, other viruses transmitted 

by sandflies and circulating in India may be imported 

into Europe by introduction of viremic patients emphasising 

the need to consider these viruses relevant from 

a European public health perspective. 

Clinical picture and geographical 

distribution 

Sandfly fever Sicilian and Naples 

virus infections 

Sandfly fever Sicilian and Naples virus and other 

related viruses cause the “three-day fever” or “papatacci 

fever”. Patients present with influenza-like 

symptoms including fever, retro-orbital pain, myalgia 

and malaise and usually recover fully within a week. 


However, infections with sandfly fever Naples and 

Sicilian viruses, even when mild, have shown to be 

highly incapacitating for the time patients are affected. 

The human cases and some virus isolation from sandflies 

were reported around the Mediterranean Sea 

(Figure 1) in Algeria [6], Cyprus [7, 8], Egypt [9], Iran 

[9-11], Israel [12], Italy [13], Jordan [14] and Portugal 

[15]. An earlier review based on serological data, 

without virus isolation and characterisation, indicated 

that sandfly fever Sicilian or Naples viruses 

have been recorded in Bangladesh, Djibouti, Ethiopia, 

Iraq, Morocco, Saudi Arabia, Somalia, Sudan, Tunisia, 

southern and central Asian republics of the former 

Soviet Union, and the former Republic of Yugoslavia 

[16]. The same study showed the absence of neutralising 

antibodies in humans in Algeria, central Africa and 

eastern Asia [16]. 

Sandfly fevers were first described in Italy, in 1943- 

1944 during outbreaks of influenza-like illness among 

United States (US) soldiers due to Sicilian and Naples 

viruses [16]. Human cases are often found in people 

visiting Mediterranean countries. A total of 37 cases of 

sandfly fever Sicilian virus infections and one case of 

sandfly fever Naples virus infection were recorded in 

Swedish tourists returning from Cyprus between 1986 

and 1989. In 1985, the incidence was low (0.3%) among 

members of Swedish troops stationed in Cyprus [17]. 

More recently, a 2002 outbreak affecting 256 among 

581 Greek soldiers stationed in Cyprus showed increasing 

incidence (44%) for infections with sandfly fever 

Sicilian virus [8]. 

Toscana virus infections 

Many infections with the Toscana virus are asymptomatic. 

Reported clinical cases mostly present with 

influenza-like symptoms, but the virus displays a 

strong neurotropism. Outbreaks of acute meningitis or 

meningo-encephalitis due to infections with Toscana 

virus have been reported in several European countries 

bordering the Mediterranean Sea: (Italy, France [18-25], 

Spain [26-30] and Portugal [31]). 


Seroprevalence studies in Italy, show large variations 

ranging from 3% in northern Italy (Torino) [32], to 

16% in Umbria [33] and 22% in central Italy. The virus 

is widespread in several regions including Tuscany, 

Piedmont, the Marches, Umbria and Emilia-Romagna. 

In Spain, the seroprevalence rate is higher and ranges 

from 5% [27] to 26% [26]. However, the large difference 

in prevalence observed between the two surveys 

might be related to the fact that the authors did 

not use the same serological tests [21]. In France, the 

seroprevalence observed recently was 12% in a survey 

using blood from donors in south-eastern France [21]. 

In Turkey [34], a pilot study reports also positive serologies 

for sandfly fever Toscana, Naples and Sicilian 

viruses. 

In Italy, from May to October, Toscana virus is a major 

cause of meningitis and meningo-encephalitis with 

a peak of incidence in August. During this period it 

causes 80% of cases in children and 50% of cases in 

adults [32, 33]. Toscana virus is among the three most 

prevalent viruses associated with meningitis during the 

warm season. Therefore, Toscana virus must be considered 

as an emerging pathogen in the Mediterranean 

basin [19] and significant public health issue in Europe. 

Chandipura virus infections 

Outside of Europe, epidemics of acute encephalitis 

characterised by rapid onset of fever and central nervous 

system involvement with high case fatality rate 

were reported in Asia [35, 36]. These outbreaks were 

caused by the highly pathogenic Chandipura virus, 

a Vesiculovirus of the Rhabdoviridae family originally 

isolated in India from a patient [2]. To date, no 

human cases have been reported in Europe and Africa, 

although Chandipura virus has been isolated in Nigeria 

from hedgehogs (Atelerix spiculus) [37]. The fact that 

no human cases have been reported from there so far 

may reflect a lack of specific testing for Chandipura 

virus. 

Figure 1 

Distribution of (a) Toscana, (b) Sicilian, and (c) Naples viruses in the European Union and neighbouring countries around 

the Mediterranean Sea up to 2009 

A B C 

Countries with confirmed cases are depicted in mid grey, the estimated distribution limits are depicted with a dark grey line. 

Source: V-borne project; reproduced with permission from the European Centre for Disease Prevention and Control. 

53

Among the nine species of the genus Vesiculovirus, 

Chandipura virus should be considered of great public 

health importance. Conducting surveys on Chandipura 

virus in the south of Europe and along the south-eastern 

European borders is necessary to anticipate an 

introduction of the virus into Europe from Asia and/or 

Africa. 

Figure 2 

Neighbour-joining tree based on nucleotide sequences 

of the large segment encoding the viral polymerase with 

bootstrap values (%) calculated with 500 replicates. 

Transmission 

Genus Phlebovirus 

This genus contains the majority of known sandflyborne 

viruses. Many serotypes have been characterised 

in the Americas from sandflies belonging to the 

genera Lutzomyia sensu lato, and in Africa, Europe and 

Central Asia mainly from Phlebotomus and also from 

Sergentomyia. 

According to the eighth Report of the International 

Committee on Taxonomy of Viruses [37], the genus 

Phlebovirus can be divided into nine antigenic complexes 

and includes 37 classified viruses. Further 16 

virus serotypes are unclassified and are considered 

to be tentative members of the genus. Current knowledge 

suggests that many of the phleboviruses are 

maintained in their arthropod vectors by vertical (transovarial) 

transmission and that vertebrate hosts play 

little or no role in the basic maintenance cycle of these 

agents [1]. This maintenance mechanism has important 

ecological implications for the phleboviruses, as it 

allows them to persist during periods when adult vectors 

are absent or when susceptible vertebrate hosts 

are not available. 

Sandfly fever Sicilian virus 

This virus has been isolated in natura [39] and in 

vitro [40, 41] from P. papatasi captured from the 

Mediterranean basin to Central Asia. It has also 

recently been isolated in natural conditions from 

P. (L.) ariasi in Algeria [6]. In some parts of its distribution 

area such as Cyprus where a local strain, Cyprus virus, 

has been isolated from Swedish troop members [7], 

P. papatasi is an abundant species [42] and could be a 

Figure 3 

Distribution of main vectors in the European Union and neighbouring countries around the Mediterranean Sea up to 2009 

Phlebotomus papatasi P. perniciosus 

P. ariasi P. perfiliewi s. st 

Countries with confirmed presence are depicted in mid grey, estimated distribution limits are depicted in dark grey. 

Source: V-borne project; reproduced with permission from the European Centre for Disease Prevention and Control. 


suspected vector. However, in Italy P. papatasi is now 

scarce whereas it was abundant before DDT was used 

in the 1940s and cannot be a candidate for the transmission 

of sandfly fever Sicilian virus. Autochtonous 

Phlebotomus belonging to the subgenus Larroussius 

(P. perniciosus, P. perfiliewi and P. neglectus) seem to 

be better candidates for its transmission. In Greece, a 

closely related virus called Corfou virus has been isolated 

from P. (Larroussius) neglectus [4]. 

Different vertebrate species including rodents 

(Apodemus spp., Mus musculus, Rattus rattus, 

Clethrionomys glareolus, Meriones libycus, Gerbillus 

aureus), insectivora (Soricidae and Talpidae) and carnivora 

(Mustela nivalis) may participate to the maintenance 

of Sandfly Sicilian virus life cycle [43-46]. 

The virus is endemic in Europe and currently there 

are no known reasons why it would not extend over 

the entire distribution range of the vectors. Its further 

spread could follow a wider distribution of the vector 

and/or the reservoirs taking into account the unknown 

potential impact of climatic shifts on the development 

of the virus in the vector. Future studies will have to 

determine (i) the distribution and prevalence of the disease 

according to serological studies in the European 

Mediterranean region, (ii) the vector competences and 

capacities of local sandfly species, (iii) the temperatures 

required for viral replication in infected sandflies 

in order to evaluate the risk of development in local 

vectors, and field work will have to be performed in 

foci where human cases, infected animal reservoirs 

and infected sandflies occur. 

Sandfly fever Naples virus 

The virus has been isolated in Italy from P. perniciosus 

[47], in Serbia from P. perfiliewi [48] and in Egypt from 

P. papatasi [49]. The area in which a stable focus is 

recorded has been delimited to Serbia [50]. Reservoirs 

for Sandfly fever Naples virus are unknown. An important 

seroprevalence rate of 30% has been recorded in 

Jordan [14]. Because the identity of the virus cannot 

be assessed with certainty, the virus could circulate 

in Turkey [51]. Future investigations similar to those 

developed for Toscana virus need to be carried out to 

gain better understanding of the potential spread of 

the virus. 

Toscana virus 

The distribution of Toscana virus includes Spain, 

France, Italy, Greece, Cyprus [19], Portugal [30], and 

Turkey [38] and it has been isolated several times from 

P. perniciosus and P. perfiliewi belonging to the subgenus 

Larroussius. Transovarial transmission has been 

demonstrated in laboratory conditions and by viral isolation 

from male Phlebotomus spp. Venereal transmission 

from infected males to uninfected females has also 

been demonstrated [52]. It is suggested that the reservoir 

of Toscana virus is most likely the vector itself. 

However, a progressive decline of vector infected rates 

from generation to generation, suggests that this virus 


cannot be maintained indefinitely by vertical transmission 

[53-55]. Consequently, the existence of reservoirs 

has to be considered. Serological data have shown no 

evidence of infection among domestic or wild animals. 

However, a Toscana virus strain was isolated from the 

brain of the bat Pipistrellus kuhli [56]. The viral genome 

detection of Toscana virus in Sergentomyia minuta [57], 

a species considered as feeding exclusively on lizards 

and geckos, points towards the possible existence of 

unknown reservoirs. The short duration of viraemia, 

and the lack of evidence for a persistent infection in 

humans, compromises the participation of humans in 

the maintenance of the virus. 

The geographical extension potential of Toscana virus 

is high in Europe. At this time numbers of endemic foci 

of the virus have been identified in different neighbouring 

countries (Spain, France, Italy) and potential 

vectors are widely dispersed. However, a rapid spreading 

of the virus is unlikely due to the lack of evidence 

of animal reservoir. Humans may favour viral transportation 

but the shortness of viraemia may limit an 

efficient transmission to naïve vectors. Moreover, the 

potential impact of climatic shifts on the vector competence 

is unknown. Similarly as for the two viruses 

mentioned above, future studies - both field work and 

experimental - will have to determine the distribution 

and prevalence of the disease caused by Toscana 

virus based on serological investigations around the 

European Mediterranean region, the vector competences 

and capacities of local sandfly species (in particular 

P. ariasi and P. perniciosus), the temperatures 

required for viral replication in infected sandflies and 

the possible impact of climate change on the potential 

spread in Europe. 

Massilia virus, phlebovirus isolate 

and Punta Toro virus 

The recent isolation of Massilia virus - a new 

Phlebovirus - from P. (L.) perniciosus in south-eastern 

France [5], emphasised the necessity of performing 

field studies to anticipate the possible eruption in 

humans of this new virus. Furthermore, the isolation of 

a probable new Phlebovirus from a sandfly (Figure 2) in 

southern France during the summer 2007 increases the 

number of phleboviruses and the potential pathogens 

for humans. This highlights the need to carry out new 

investigations in Europe taking into account the variability 

of phleboviruses [58]. 

The distribution area of Punta Toro virus is limited to 

Central America where it is transmitted by Lutzomyia 

(Nyssomyia) trapidoi and L. (Ny.) ylephiletor. The taxonomic 

status of these vectors has to be clarified in the 

light of an entomological revision. Even if the subgenus 

Nyssomyia has never been recorded in the West Indies, 

some species belonging to it have been recorded in 

French Guyana such as L. anduzei, L. flaviscutellata, 

L. umbratilis, L. yuilli pajoti [59]. These sandfly species 

could be considered as possible candidates for 

native transmission in the overseas territories of the 

55

European Union (EU) which are important leisure destinations 

during local dry seasons. Whereas importation 

of Punta Toro virus in European countries is unlikely, 

the possible emergence of the virus will highlight the 

importance to have the capacity to diagnose etiologically 

any imported febrile syndromes in tourists returning 

from these areas. 

The main natural Phlebotomus vectors seem to belong 

to the subgenus Larroussius. The vectors of the main 

phleboviruses in the eastern part of the Mediterranean 

basin are not known: in Turkey, P. perniciosus and 

P. ariasi are not recorded (Figure 3). However, it 

appears difficult to assess a co-evolution between 

viruses and sandflies within the subgenus Larroussius: 

the isolation of viruses (or viral RNA) in P. papatasi or 

in Sergentomyia spp. strongly suggests the capture of 

the viruses by Phlebotomine sandflies. 

Genus Vesiculovirus 

Chandipura virus 

Under laboratory conditions, P. papatasi is an efficient 

reservoir for the virus, showing growth, and 

venereal and transovarial transmission [60, 61]. The 

experimental transmission of Chandipura virus by 

P. (Euphlebotomus) argentipes has been recently demonstrated 

[62]. In natural conditions, it has been isolated 

from a pool of 253 unidentified Phlebotomine 

sandflies (Phlebotomus spp.) in the Maharashtra State 

of India [63] and from unidentified Sergentomyia in 

the Karimnagar district in Andhra Pradesh, India [64]. 

Four strains have also been isolated from batches of 

sandflies from Senegal, belonging probably to the 

genus Sergentomyia [65, 66]. These data show a wide 

distribution of the virus and the capacity of two genera 

of sandflies namely Phlebotomus (subgenera 

Phlebotomus and Euphlebotomus) and Sergentomyia 

to transmit the virus. 

Chandipura virus is currently endemic only in India and 

its introduction to Europe by an infected Phlebotomine 

sandfly is unlikely to occur, due to the fact that no settlement 

of Phlebotomine Chandipura virus vector has 

been documented yet. However, the importation of 

Chandipura virus through an infected individual with 

or without clinical symptoms cannot be excluded. This 

could be the main risk of introduction in European 

areas where P. papatasi is an abundant species. To 

assess the transmission risk, it is necessary to carry 

out studies on the duration of the viraemia in infected 

humans and the vector competence of autochtonous 

Phlebotomine species in European countries where 

P. papatasi is scarce or not recorded. The recent introduction 

in Cyprus of Leishmania donovani, an Asiatic 

and African parasite transmitted by local Phlebotomine 

sandflies highlights the risk of introduction diseases 

potentially transmitted by European Phlebotomine 

sandflies [67-69]. 

The virus Isfahan has been isolated only in Iran 

in P. papatasi, rodents and patients [70]. The Jug 

Bogdanovac virus has been isolated in P. (L.) perfiliewi 

in Serbia [71]. 

Genus Orbivirus 

Orbiviruses transmitted by sandfly bites are restricted 

to the 12 species from the Americas belonging to the 

Changuinola virus group. Human infection caused by 

this group is not well documented and until now has 

presented with mild influenza-like symptoms and does 

not show major clinical importance [72]. 

Laboratory diagnosis 

Direct viral diagnosis, such as isolation, RT-PCR, in 

blood or cerebrospinal fluid is only possible in early 

stages of infection i.e. the first two days after symptom 

onset and before the IgM sero-conversion. In most 

cases the diagnosis is based on serological investigation 

of acute and early convalescent sera. In-house 

enzyme-linked immunosorbent assay (ELISA) methods 

(MAC-ELISA and IgG sandwich) are developed in 

reference laboratories. To date, only one commercial 

kit is registered in Italy for Toscana virus diagnosis. 

Serological cross reactions exist within the sandfly 

fever Naples virus and sandfly fever Sicilian virus antigenic 

complex. Seroneutralisation assays using early 

convalescent sera remain the reference method to specifically 

identify the viruses or to assess the antibody 

response specificity. Reference tools, reagents and 

quality control are not widely available. However, the 

collaborative working group of the European Network 

for the diagnosis of imported viral diseases (ENIVD 

www.enivd.de/index.htm) is able to provide some of 

these reagents. 

Treatment and prevention 

The treatment of phlebovirus infections is symptomatic. 

Treatment with hepatotoxic medication as well 

as aspirin and other NSAIDs such as ibuprofen and 

ketoprofen are not recommended. 

No human vaccine against Phlebotomus-borne virus 

is available. The prevention of phlebovirus infection 

relies on the control of vector proliferation in limited 

areas where people are highly exposed. Individual protective 

measures such as insect repellents and insecticide 

impregnated mosquito bednets are recommended 

in these areas. 

In most cases, phlebovirus infections are self-resolving 

pathologies. Only two complicated forms of Toscana 

virus infections have been reported in the literature. If 

a vaccine were available, the implementation of mass 

vaccination programmes would not seem to be relevant 

for the prevention for sandfly fever Naples and sandfly 

fever Sicilian viruses. 

Risk for the future 

Data on vertebrate reservoirs is sparse for sandfly 

fever viruses. The factor currently known to limit the 

spread of diseases is the distribution areas of potential 

vectors. The distribution areas of the disease may 


not be restricted to the areas where they have been 

recorded but could be as wide as those of their vectors, 

that is to say Larroussius and P. papatasi mainly 

but not exclusively (Figure 3). Consequently, field work 

in form of viral isolation from sandflies and possible 

reservoirs as well as laboratory work to establish vectorial 

competence of colonised sandflies need to be 

encouraged in a near future for three main reasons:(i) 

phleboviruses already endemic in the southern part of 

Europe have a potential to spread to other areas where 

their vectors are circulating, (ii) new phleboviruses of 

unknown pathogenicity such as the Massilia virus, that 

circulate among Phlebotomine sandflies may emerge in 

humans, (iii) the highly pathogenic Chandipura virus is 

paradigmatic of arthropod-borne viruses transmitted 

by Phlebotomine sandflies that may be introduced to 

Europe. At the present time, Rift Valley Fever virus has 

not been isolated from Phlebotomine sandflies under 

natural conditions. However, sandfly infections have 

been demonstrated under laboratory conditions for 

P. (P.) papatasi, P. (P.) duboscqi, P. (Paraphlebotomus) 

sergenti, Sergentomyia schwetzi and Lutzomyia longipalpis 

[73-75]. A vector competence has been demonstrated 

after oral infection for P. papatasi and 

P. duboscqi whereas P. sergenti, S. schwetzi and 

L. longipalpis do not seem to be able to transmit Rift 

Valley Fever virus after oral infection [73-75]. The lack 

of isolates of Rift Valley Fever virus from field-collected 

Phlebotomine sandflies may could be a consequence of 

the low rates of capture of sandflies in arthropod field 

collections for virus isolation assays. Their geographical 

range coinciding with that of Rift Valley Fever virus 

in sub-Saharan Africa, and nearly all known phleboviruses 

seem primarily associated with sandflies. Thus, 

additional studies are needed to evaluate the role of 

sandflies as maintenance and epizootic vectors for Rift 

Valley Fever virus [75]. An epidemiological surveillance 

is also required in the EU. 

Phleboviruses as a potential 

means of biological warfare 

Efficient arboviruses transmission mainly depends on 

vectors. Except for RVFV, no other way of Phlebovirus 

transmission has been reported. Breeding of 

Phlebotomine species and artificial infection difficulties 

are limiting factors for the use of phleboviruses 

as efficient biological weapons. Moreover, most phleboviruses 

are associated with asymptomatic or mild 

self-resolving infections in humans. Direct inter-human 

transmission has never been demonstrated. These criteria 

make phleboviruses bad candidates for the development 

of biological weapons. 

Phleboviruses are characterised by their tripartite RNA 

genome. Genetic exchanges between phleboviruses 

are possible with unpredictable effects. Compared to 

RVFV, only less pathogenic phleboviruses have been 

identified so far. The possible genesis of a new, highly 

virulent Phlebovirus by this genetic-exchange mechanism 

seems unlikely. 



This review is based on an extensive literature study conducted 

as part of the European Centre for Disease Prevention 

and Control funded V-borne project “Assessment of the magnitude 

and impact of vector-borne diseases in Europe”, tender 

n° OJ/2007/04/13-PROC/2007/003. 

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59

Perspectives 

Crimean-Congo hemorrhagic fever in Europe: current 

situation calls for preparedness 

H C Maltezou (helen-maltezou@ath.forthnet.gr) 1 , L Andonova 2 , R Andraghetti 3 , M Bouloy 4 , O Ergonul 5 , F Jongejan 6 , 

N Kalvatchev 7 , S Nichol 8 , M Niedrig 9 , A Platonov 10 , G Thomson 11 , K Leitmeyer 12 , H Zeller 12 

1. Hellenic Center for Diseases Control and Prevention, Athens, Greece 

2. Medical University, Sofia, Bulgaria 

3. World Health Organization, Copenhagen, Denmark 

4. Institut Pasteur, Paris, France 

5. Marmara University, Istanbul, Turkey 

6. Utrecht Centre for Tick-borne Diseases, Utrecht University, Utrecht, the Netherlands 

7. National Centre of Infectious and Parasitic Diseases, Sofia, Bulgaria 

8. Centers of Disease Control and Prevention, Atlanta, United States 

9. Robert Koch Institute, Berlin, Germany 

10. Central Research Institute of Epidemiology, Moscow, Russian Federation 

11. Health Protection Agency, London, United Kingdom 

12. European Centre for Disease Control and Prevention, Stockholm, Sweden 


Citation style for this article: Maltezou HC, Andonova L, Andraghetti R, Bouloy M, Ergonul O, Jongejan F, Kalvatchev N, Nichol S, Niedrig M, Platonov A, Thomson G, 

Leitmeyer K, Zeller H. Crimean-Congo hemorrhagic fever in Europe: current situation calls for preparedness. Euro Surveill. 2010;15(10):pii=19504. Available online: 

http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19504 

During the last decade Crimean-Congo hemorrhagic 

fever (CCHF) emerged and/or re-emerged in several 

Balkan countries, Turkey, southwestern regions of the 

Russian Federation, and the Ukraine, with considerable 

high fatality rates. Reasons for re-emergence of 

CCHF include climate and anthropogenic factors such 

as changes in land use, agricultural practices or hunting 

activities, movement of livestock that may influence 

host-tick-virus dynamics. In order to be able to 

design prevention and control measures targeted 

at the disease, mapping of endemic areas and risk 

assessment for CCHF in Europe should be completed. 

Furthermore, areas at risk for further CCHF expansion 

should be identified and human, vector and animal 

surveillance be strengthened. 

Introduction 

Crimean-Congo hemorrhagic fever (CCHF) is an acute, 

highly-contagious viral zoonosis transmitted to 

humans mainly by ticks of the genus Hyalomma, but 

also through direct contact with blood or tissues of 

viraemic hosts. In humans CCHF typically presents 

with high fever of sudden onset, malaise, severe headache 

and gastrointestinal symptoms. Prominent hemorrhages 

may occur in late stages of the disease with 

published fatality rates ranging from 10% to 50% [1,2]. 

The disease is endemic in parts of Africa, Asia, the 

Middle East and eastern Europe. Main animal hosts 

include a number of domestic animals such as cattle, 

sheep, goats, and hares. CCHF has the potential 

to cause community and nosocomial outbreaks. Due 

to the high case fatality rates and difficulties in treatment, 

prevention, and control, CCHF is a disease which 

should be notified immediately to public health authorities 

in the European Union (EU). CCHF virus is also in 


the list of agents for which the Revised International 

Health Regulations of 2005 call for implementation of 

the decision algorithm for risk assessment and possible 

notification to the World Health Organization 

(WHO) [3]. 

In Europe, CCHF is currently only endemic in Bulgaria, 

however during the last decade an increased number 

of CCHF cases and outbreaks have been recorded in 

other countries in the region such as Albania, Kosovo, 

Turkey, and the Ukraine as well as south-western 

regions of the Russian Federation [4-9]. In June 2008, 

the first case was registered in Greece [10]. In response 

to this situation, the European Centre for Disease 

Prevention and Control (ECDC) invited a group of CCHF 

experts to review the situation of CCHF in Europe and 

to consult on interventions necessary to strengthen 

preparedness and response at the European level [11]. 

This article provides an update on the current situation 

of CCHF in Europe, and emphasises existing prevention 

and control capacities within the EU. Aspects 

relevant to strengthen preparedness for CCHF are also 

discussed. 

CCHF situation in Europe 

CCHF is endemic in Bulgaria since the 1950’s, when a 

large outbreak occurred from 1954 to 1955 with 487 

notified cases mainly in the Shumen area in north-east 

Bulgaria. In total, 1,568 CCHF cases were notified in 

Bulgaria from 1953 to 2008, with an overall case fatality 

rate of 17% [4]. Endemic areas are confined to the 

vicinity of Shumen, Razgrad, Veliko Tarnovo, Plovdiv, 

Pazardjik, Haskovo, Kardjali, and Bourgas, however 

in April 2008 a cluster of six probable cases occurred 

in Gotse Delchev in the south-western province 


Blagoevgrad near the border with Greece, an area 

considered of low CCHF endemicity until recently [5]. 

During the last decade, CCHF outbreaks have also been 

noted in Albania in 2001 and 2003, and in Kosovo in 

2001 [6,7]. 

In Turkey, the first symptomatic human CCHF cases 

were noted in 2002, however, serologic evidence of 

enzootic CCHF virus circulation as well as limited evidence 

of CCHF infections among humans (2.4% among 

1,100 tested humans) has been found since the 1970’s 

[4]. Starting in 2003, Turkey has experienced an 

expanding outbreak with increasing numbers of notified 

cases and associated fatalities (2002: 17/0; 2003: 

133/6; 2004: 249/13; 2005: 266/13; 2006: 438/27; 

2007: 713/33; 2008: 1,315/63; 2009: 1,300/62) [4,12]. 

Overall, there are more than 4,400 recorded laboratory 

confirmed CCHF cases in this country, mainly among 

residents in rural areas in north-central and northeast 

Anatolia [4,8,12]. Within the CCHF endemic areas, 

there are hyperendemic areas where one out of every 

five residents and one out of every two residents with 

a history of tick bite has antibodies against CCHF virus 

[13]. A predictive map model using satellite-based climate 

data and high-resolution vegetation images from 

Turkey from 2003 to 2006 revealed that areas with 

higher CCHF reporting were significantly associated 

with zones of high climate suitability for Hyalomma 

ticks and high rate of fragmentation of agricultural 

land [13]. 

In Greece, a serosurvey conducted between 1981 and 

1988 among 3,388 rural residents from across the 

country showed 1% seroprevalence rate against CCHF 

virus [4]. More than 400 cases with a CCHF compatible 

clinical syndrome have tested negative for CCHF virus 

in this country since 1982, therefore, the seroprevalence 

rate of 1% was attributed to the non-pathogenic 

AP-92 strain and not to the pathogenic Balkan CCHF 

virus strain. A number of the cases tested for CCHF 

were finally diagnosed as hemorrhagic fever with renal 

syndrome (HFRS), leptospirosis and ricketsial infections. 

Other diagnoses were meningococcal meningitis 

and unspecified bacterial sepsis. The first CCHF 

case was recorded in June 2008 in a woman with a tick 

bite working in agriculture near the city of Komotini 

in north-eastern Greece [10]. This town is situated 

within a few kilometres distance from there where the 

Bulgarian cluster occurred in 2008 [5]. A seroepidemiological 

study for CCHF virus among local population 

and animals are underway in northern Greece. 

After nearly 27 years without any human cases, CCHF 

re-emerged in the south-western regions of the Russian 

Federation in 1999. Outbreaks have been reported in 

Astrakhan, Rostov and Volgograd Provinces, Krasnodar 

and Stavropol Territories, Kalmykia, Dagestan and 

Ingushetia Republics. Between 2000 and 2009 more 

than 1,300 clinical cases were diagnosed in the 

Russian Federation with an overall fatality rate of 3.2% 

for the period from 2002-2007 [4]. Most cases occurred 


among residents of rural areas in the Southern Federal 

Distinct. The largest number of cases was registered 

in Stavropol Territory, Kalmykia Republic and Rostov 

Province, where the mean annual CCHF incidence 

rate was 1.7, 10.1, and 0.7 cases per 100,000 population, 

respectively. During 2008 alone, the incidence in 

Stavropol Territory increased by 1.3 times, and was the 

highest recorded in this region during the last decade 

[4,9,14]. In 2009, CCHF cases were also reported from 

Georgia, Kazakhstan, Tajikistan, Iran, and Pakistan 

[15]. 

CCHF emergence and/or re-emergence in south-eastern 

Europe and neighboring countries is attributed to climate 

and ecologic changes and anthropogenic factors 

such as changes in land use, agricultural practices, 

hunting activities, and movement of livestock, that 

may have an impact on ticks and hosts and accordingly 

on CCHF epidemiology [1,2]. The geographic distribution 

of CCHF coincides with that of Hyalomma ticks. 

H. marginatum, the main CCHF virus vector in Europe, 

is found in Albania, Bulgaria, Cyprus, France, Greece, 

Italy, Kosovo, Moldavia, Portugal, Romania, Russia, 

Serbia, Spain, Turkey, and the Ukraine. In 2006 it was 

detected for the first time in the Netherlands and in 

southern Germany [16,17]. Given the wide distribution 

of its vector, the numerous animals that can serve as 

hosts, and the favorable climate and ecologic conditions 

in several European countries bordering the 

Mediterranean Sea, it is possible that the occurrence 

of CCHF will expand in the future. A model that studied 

various climate scenarios on the habitat areas of 

different ticks, showed that a rise in temperature and 

a decrease in rainfall in the Mediterranean region will 

result in a sharp increase in the suitable habitat areas 

for H. marginatum and its expansion towards the north, 

with the highest impact noted at the margins of its current 

geographic range [18]. 

Current prevention and control in Europe 

Several elements relating to laboratory diagnosis, surveillance 

and therapy of CCHF should be addressed in 

order to increase preparedness capacity in Europe and 

to design appropriate prevention and control measures. 

Laboratory diagnosis 

In 2008 there were 20 laboratories with diagnostic 

capacities for CCHF virus in Europe: 14 in EU Member 

States, eight in the endemic regions of the Russian 

Federation, and one in Turkey. Most of them used 

immunofluorescence assays (IFA), ELISA, and/or molecular 

methods to diagnose CCHF whereas eight among 

them were also able to isolate CCHF virus [11], a BSL-4 

containment agent. Limitations for diagnosing CCHF 

concern both the limited diagnostic capacities in several 

endemic areas as well as difficulties in the international 

transfer of samples for logistic and economic 

reasons. However, rapid and easy tests are needed to 

guide initial therapeutic decisions for the patient. 

61

Surveillance 

Currently, there are no standardised case definitions 

for CCHF notification and contact tracing within 

European countries [19]. Recent cases of nosocomial 

acquisition of CCHF in health care workers were well 

documented [6,8,20]. These cases underline the need 

for educating health-care workers about the modes of 

getting infected with CCHF virus and for strict implementation 

of infection control measures within health 

care facilities, and the importance of providing adequate 

resources to do so [1,2]. 

Therapy 

The World Health Organization (WHO) recommends ribavirin 

for the treatment of CCHF cases [21,22]. Ribavirin 

appears to be more effective when introduced early 

in the course of illness [23]. Evidence of its efficacy is 

based on in vitro data and on limited observations in 

humans [24-26]. Randomised controlled trials have not 

been conducted so far, and ethical issues concerning 

the use of a control group remain a major obstacle for 

this [27]. Severity of infection, duration of illness prior 

to initiation of therapy, and route of administration may 

impact the clinical outcome of CCHF cases. On individual 

country level, recommendations for treatment 

of CCHF cases with ribavirin existed in 2008 in Turkey, 

Russia, Bulgaria, and Greece. In Bulgaria, in addition, 

specific hyperimmune globulin collected from convalescent 

CCHF cases is used for prophylaxis and treatment 

and an inactivated suckling mouse brain vaccine 

is in use since the 1970’s for high-risk groups living in 

CCHF endemic regions [28]. There is no vaccine against 

CCHF licensed in any other EU Member State. 

Conclusions 

CCHF is a disease with a high fatality rate and the 

potential to cause outbreaks. The vector for CCHF, 

the Hyalomma tick is present in southeastern and 

southern Europe. Climate factors may contribute to 

a further spread of the vector and to a consecutive 

extension of the geographic range of CCHF, which may 

further expand to European countries bordering the 

Mediterranean Sea, with the highest risk in neighbouring 

areas with already established endemicity. This 

highlights the need for strengthening human, vector, 

and veterinary surveillance, especially in areas where 

CCHF is expected to occur in the future. Together with 

the implementation of standardised case definitions 

for CCHF this will allow an estimate of the CCHF burden 

and of epidemiologic trends in various areas and 

countries. Guidance for contact tracing and the establishment 

of early detection and response systems will 

allow prompt interventions at patient, community, and 

hospital level. To enable early detection, laboratory 

capacities are crucial to rapidly confirm the suspected 

clinical diagnosis and besides being available, tests 

need to be reliable and affordable. Overall, laboratory 

capacities for CCHF should increase. Considering the 

high case fatality rate of CCHF, development of a vaccine 

and new drugs against CCHF are of major importance. 

Ribavirin efficacy should be assessed through 

well-designed clinical protocols and in endemic areas 

general public and health-care workers should be 

aware about modes of CCHF transmission and prophylactic 

measures. Climate and environmental factors 

and human behavior that may influence CCHF epidemiology 

and spread should be further studied. Mapping 

of endemic areas and risk assessment for CCHF in 

Europe should be completed and areas at risk for CCHF 

expansion should be identified and finally, appropriate 

tick-control strategies including public education 

should be implemented. All these measures should be 

undertaken as part of a multidisciplinary collaboration 

at interregional and international level and link with 

initiatives such as the International network for capacity 

building for the control of emerging viral vectorborne 

zoonotic diseases: ARBO-ZOONET [29]. 

In accordance with an ECDC-initiated assessment on 

the importance of vector-borne diseases in 2008, CCHF 

has been identified as a priority disease for the EU [12]. 

In order to strengthen preparedness and response for 

CCHF and build capacity for its prevention and control, 

it is necessary to identify relevant gaps and work in an 

integrated fashion. 

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63


Chikungunya infection in a French traveller returning 

from the Maldives, October, 2009 

M Receveur 1 , K Ezzedine (khaled.ezzedine@chu-bordeaux.fr) 1 , T Pistone 1 , D Malvy 1 

1. Travel Clinics and Unit for Tropical Medicine and Imported Diseases, Department of Internal Medicine and Tropical Diseases, 

Hôpital St-André, University Hospital Centre, Bordeaux, France 


Citation style for this article: Receveur M, Ezzedine K, Pistone T, Malvy D. Chikungunya infection in a French traveller returning from the Maldives, October, 2009. 

Euro Surveill. 2010;15(8):pii=19494. Available online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19494 

In the last years, cases of chikungunya fever have been 

reported in international travellers returning from 

the Indian Ocean region. The cases have been linked 

to the re-emergence of chikungunya fever on Indian 

Ocean islands in 2006. We describe the first case of 

chikungunya fever in a French traveller returning from 

Malé, an island of the Maldives islands, confirming the 

permanence of virus circulation by the end of 2009. 

Introduction 

Chikungunya virus is a mosquito-borne alphavirus 

found in the tropical regions of Africa and Asia where 

it causes endemic and epidemic chikungunya fever, 

an acute self-limiting febrile algo-eruptive illness [1]. 

Chikungunya fever has been increasingly reported in 

international travellers following its re-emergence on 

Indian Ocean islands and its spread to southern Asia 

thereafter [2-4]. Moreover, some African and southeast 

Asian countries show an endemic circulation of 

the virus [5] which may contribute to occurrence of the 

disease among travellers. The illness was suspected 

to have emerged in the Maldives archipelago in 2007 

[6], following the sweeping succession of outbreaks 

that occurred in the Indian Ocean region where it first 

affected Kenya in 2004, Réunion Island in 2005 and 

southern India in 2006 [1,7]. Here we report a confirmed 

case of chikungunya fever in a French traveller 

returning from Malé island, the Maldives, where an 

outbreak of chikungunya fever was reported starting in 

January 2009. 

Case report 

A French male in his thirties presented at the posttravel 

clinic of the Department of Internal Medicine and 

Tropical Diseases of the University Hospital Centre, 

Bordeaux, France in October 2009 with symptoms of 

recurrent high-grade fever (up to 40°C), headache, 

generalised muscle aches and severe joint pain mainly 

affecting fingers, wrists, knees and ankles, and an 

itching skin rash, since three days. Two days before, 

he had returned directly from a holiday trip to the 

Maldives where he had stayed for 14 days exclusively 

in the northern part of Malé island. 

This article has been published on 25 February 2010 

In our centre, the patient presented with a slight macular 

skin rash on the trunk and limbs, a slightly swollen 

right knee and small joints of hands and feet. 

Laboratory tests at the time of presentation showed a 

leucocyte cell count of 4,600 white blood cells (WBC)/ 

µL, a thrombocyte count of 178,000 platelets/µL and 

an elevated C-reactive protein level (27 mg/L; normal 

Maldives. To the best of our knowledge, this case is 

reportedly the third confirmed chikungunya fever case 

imported from the Maldives since the first documented 

outbreak of chikungunya in Malé and other islands of 

the Maldives that lasted from December 2006 to April 

2007 [6], followed by a suspected cluster on the Laamu 

Atoll from December 2008 to January 2009 [10] and the 

report of two confirmed cases in German travellers, 

a father and son returning from a 10-day visit to the 

Maldives mid-September 2009 [11]. 

The region is probably one of the most popular travel 

destinations in the Indian Ocean area. This may result 

in an increase of symptomatic travellers returning from 

this area and seeking medical advice at travel or primary 

care clinics. Hence, chikungunya together with 

dengue fever should be considered as an important 

differential diagnosis in those patients, assuming that 

both diseases are endemic in certain regions of India 

and the Indian Ocean area and may present with similar 

symptoms. 

For more than 10 years, dengue fever was the only 

vector-borne viral disease reported in the Maldives. 

Aedes aegypti and A. albopictus, the dengue virus vectors 

which can also transmit the chikungunya virus, 

have been identified in the Maldives, with A. aegypti 

identified as the predominant vector in Malé [6]. The first 

chikungunya fever outbreak occurred from December 

2006 to April 2007 with abrupt onset and high attack 

rates due to the lack of herd immunity. Epidemics may 

occur following an interval of 20-30 years of the virus 

not circulating as has been the case in western Africa 

and Malaysia [5, 6]. Confirmed imported cases among 

travellers support the assumption of endemic circulation 

of the virus which is consistent with the prevailing 

chikungunya epidemic in the Indian Ocean region. 

This report highlights the need for surveillance in 

countries where emerging infections may be introduced 

by returning travellers as in the case with the 

Italian chikungunya fever epidemic which occurred in 

the province of Ravenna in 2007 [12]. It illustrates how 

travellers can serve as sentinel population providing 

information regarding the emergence or re-emergence 

of an infectious pathogen in a source region. Travellers 

can thus act as carriers who inadvertently ferry pathogens 

that can be used to map the location, dynamics 

and movement of pathogenic strains [9]. Thus, with the 

increase in intercontinental travel, travellers can provide 

insights into the level of the risk of transmission 

of infections in other geographical regions. 

Conclusion 

We report a case of dengue-like illness diagnosed as 

chikungunya in a tourist returning from the Maldives, a 

popular tourist spot. Despite the clinical similarity with 

dengue fever, chikungunya should be recognised early 

in returning travellers because of its specific protracted 

morbidity and its potential for causing local outbreaks 

in European countries, where local transmission is 


possible through the presence of the receptive vector 

in southern European countries. 

References 

1. Renault P, Solet JL, Sissoko D, Balleydier E, Larrieu S, Filleul 

L, et al. A major epidemic of chikungunya virus infection 

on Reunion Island, France, 2005-2006. Am J Trop Med Hyg. 

2007;77(4):727-31. 

2. Taubitz W, Cramer JP, Kapaun A, Pfeffer M, Drosten C, Dobler G, 

et al. Chikungunya fever in travellers: clinical presentation and 

course. Clin Infect Dis. 2007;45(1):e1-4. 

3. Larrieu S, Pouderoux N, Pistone T, Filleul L, Receveur MC, 

Sissoko D, et al. Factors associated with persistence of 

arthralgia among chikungunya virus-infected travellers: report 

of 42 French cases. J Clin Virol. 2010;47(1):85-8. 

4. Simon F, Parola P, Grandadam M, Fourcade S, Oliver M, Brouqui 

P, et al. Chikungunya infection: an emerging rheumatism 

among travellers returned from Indian Ocean islands. Report of 

47 cases. Medicine (Baltimore). 2007;86(3):123-37. 

5. AbuBakar S, Sam IC, Wong PF, MatRahim N, Hooi PS, Roslan N. 

Reemergence of endemic Chikungunya, Malaysia. Emerg Infect 

Dis. 2007;13(1):147-9. 

6. Yoosuf AA, Shiham I, Mohamed AJ, Ali G, Luna JM, Pandav R, et 

al. First report of chikungunya from the Maldives. Trans R Soc 

Trop Med Hyg. 2009;103(2):192-6. 

7. Sissoko D, Malvy D, Ezzedine K, Renault P, Moscetti F, 

Ledrans M, et al. Post-epidemic Chikungunya disease on 

Reunion Island: course of rheumatic manifestations and 

associated factors over a 15-month period. PLoS Negl Trop Dis. 

2009;3(3):e389. 

8. Pastorino B, Bessaud M, Grandadam M, Murri S, Tolou 

H, Peyrefitte CN. Development of a TaqMan RT-PCR 

assay without RNA extraction step for the detection and 

quantification of African Chikungunya viruses. J Virol Methods. 

2005;124(1-2):65-71. 

9. Pistone T, Ezzedine K, Schuffenecker I, Receveur MC, Malvy 

D. An imported case of Chikungunya fever from Madagascar: 

use of the sentinel traveller for detecting emerging arboviral 

infections in tropical and European countries. Travel Med Infect 

Dis. 2009;7(1):52-4. 

10. Acute febrile disease - Maldives: (Laamu Atoll). In: ProMEDmail 


Diseases; 14 January 2009. Archive no. 20090114.0150. 

Available from: http://www.promedmail.org 

11. Chikungunya (34): Germany ex Maldives. In: ProMED-mail 


Diseases; 22 September 2009. Archive no. 200900922.3337. 

Available from: http://www.promedmail.org 

12. Angelini R, Farinelli AC, Angelini P, Po C, Petropulacos K, 

Macini P, et al. An outbreak of chikungunya fever in the 

province of Ravenna, Italy. Euro Surveill. 2007;12(36):pii=3260. 



65


Chikungunya fever in two German tourists returning 

from the Maldives, September, 2009 

M Pfeffer (pfeffer@vetmed.uni-leipzig.de) 1 , I Hanus 2 , T Löscher 2 , T Homeier 1 , G Dobler 3 

1. Institute of Animal Hygiene and Veterinary Public Health, Veterinary Faculty, University of Leipzig, Germany 

2. Department of Infectious Diseases and Tropical Medicine, University of Munich, Munich, Germany 

3. Bundeswehr Institute of Microbiology, Munich, Germany 


Citation style for this article: Pfeffer M, Hanus I, Löscher T, Homeier T, Dobler G. Chikungunya fever in two German tourists returning from the Maldives, September, 

2009. Euro Surveill. 2010;15(13):pii=19531. Available online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19531 

This report describes the first isolation and molecular 

characterisation of a chikungunya virus from 

two German tourists who became ill after a visit 

to the Maldives in September 2009. The virus contained 

the E1 A226V mutation, shown to be responsible 

for an adaptation to the Asian tiger mosquito 

Aedes albopictus. The E1 coding sequence was identical 

to chikungunya virus isolates from Sri Lanka and 

showed three nt-mismatches to the only available E1 

nt sequence from the Maldives. 

Introduction 

Since the start of the current chikungunya fever pandemic 

on the east coast of Africa in 2005, many cases 

have been reported in countries in Asia and south-east 

Asia [1,2]. These cases were attributed to a particular 

chikungunya virus (CHIKV) strain that has adapted to 

very efficient transmission to humans via the Asian 

tiger mosquito (Aedes albopictus) due to a A226V 

mutation in the E1 envelope protein [3,4]. The Maldives 

were first hit by the chikungunya virus pandemic in 

late 2006 after the wet season which usually lasts 

until September. Based on almost 12,000 suspected 

cases of chikungunya fever the disease was reported 

on 121 of the 197 inhabited islands with incidence rates 

between 82 and 722 per 1,000 population [5]. A small 

set of blood samples from febrile patients with symptoms 

meeting the chikungunya fever case definition 

at that time confirmed CHIKV as causative agent in 

64 of 67 cases by reverse-transcription PCR (RT-PCR) 

[5]. However, no further characterisation of the virus 

strain responsible for the 2006-7 outbreak was performed. 

One case of a traveller returning to Singapore 

in January 2007 was confirmed by RT-PCR and the nt 

sequence of the E1 gene was determined [6]. In early 

2009, an outbreak of a viral fever with symptoms 

including myalgia or arthralgia and rash occurred on 

several islands of the Laamu Atoll about 400 km south 

of Malé [7], but no further virological investigation was 

carried out to determine whether this was due to dengue 

or chikungunya fever. 


Case report and laboratory findings 

Between 1 and 10 September 2009, a German couple 

visited the Dhiffushi Holiday Island resort at the southern 

tip of the Ari Atoll, the Maldives (Figure 1), together 

with their seven year-old son. They flew directly from 

Munich to Malé with a stopover in Dubai, United Arab 

Emirates. 

Two and three days respectively after the family 

had returned to Munich, the son and the 35 year-old 

father developed symptoms compatible with either 

dengue or chikungunya fever (Table) while the wife 

stayed healthy. A test for dengue virus showed neither 

virus RNA nor anti-dengue virus (DENV) IgM for both 

patients, but the father had IgG antibodies reactive 

against DENV indicating an earlier anamnestic dengue 

fever or a cross-reaction with an earlier flavivirus 

vaccination. CHIKV-specific real-time RT-PCR yielded 

ct-values of 23 (son) and 22.5 (father) in the respective 

acute serum samples obtained on 14 September, 

indicating high-level viremia [8,9]. Chikungunya virus 

was isolated in Vero B4 cells from both sera and the 

entire nucleotide sequence of the isolate from the 

father was determined. The viral genome was 11,811 

nucleotides in length and showed high levels of identity 

with the pandemic CHIKV that is circulating in 

many parts of the Indian subcontinent and other parts 

of Asia since 2006. Most interestingly the CHIKV isolate 

from the Maldives contained the A226V change 

in the E1 glycoprotein which has been shown to be 

responsible for shorter extrinsic incubation periods in 

Aedes albopictus mosquitoes [4]. While the son made 

an uneventful recovery after one week of symptoms, 

the father developed persisting arthralgias with limited 

mobility in the affected extremities and still requires 

analgesic treatment (Table). 

Discussion 

Together with a very recent report on chikungunya 

fever in a French traveller returning from the northern 

part of Malé Island, Maldives, in October 2009 [10], 

our findings suggest a continuous circulation of CHIKV 

also in other parts of the Maldives. The family stayed 


on Dhiffushi Holiday Island throughout their holidays 

with a daytrip to the neighboring Sun Island. Malé with 

its international airport was only visited for the intercontinental 

flight connection, leaving not much time 

to become exposed to mosquito bites. We cannot rule 


out that both infections were acquired while waiting at 

the airport, because this would fit well with both the 

incubation period of the disease and with the previous 

case report of the French traveller, who became 

infected while staying at the Malé Atoll. However, given 

Figure 1 

Location of Holiday and Sun Islands on the southernmost rim of the south Ari Atoll, about 100 km away from Male 

International Airport 

Ihavandhippolhu 

Atoll 

Maamakunudhoo 

Atoll 

Maalhosmadulu 

Atoll 

Goidhoo 

Atoll 

Ari Atoll 

Holiday Island/ 

Sun Island 

Nilandhoo 

Atoll 

Kolhumadulu 

Atoll 

100 km 

Huvadhoo 

Atoll 

MALDIVES 

Thiadhunmathee 

Atoll 

Miladhunmadulu 

Atoll 

Faadhippolhu 

Atoll 

Male 

Atoll 

Felidhoo 

Atoll 

Malaku 

Atoll 

Hadhdhunmathee 

Atoll 

Equator 

Table 

Clinical and laboratory data of patients diagnosed with chikungunya fever, Germany, September 2009 

Patient Son (7 years) Father (35 years) 

Travel schedule 

Munich-Dubai-Male and back on 1-10 September, 

2009 

ASIA 

Munich-Dubai-Male and back on 1-10 September, 

2009 

Onset of disease 12 September 2009 13 September 2009 

Clinical presentation Fever 39.5°C Fever 39.0°C 

Headache 

Ague, retrobulbar pressure and pain, arthritis of 

both wrists and ankles 

Macular and partially confluent exanthema Erythema, macular and partially confluent exan- 

(mainly on face and torso) 

thema (mainly on torso and arms) 

Laboratory findings 

Leucocytes 2,700/µl 

CRP 2.5 mg/dl 

Leucocytes 5,300/µl 

CRP 13 mg/dl 

Creatinine 1.4 mg/dl 

CHIKV RT-PCR positive CHIKV RT-PCR positive 

DENV RT-PCR negative DENV-PCR negative; anti-DENV IgG 15E 

Therapy Paracetamol, Ibuprofen Paracetamol, Ibuprofen 

Since 16 September fever-free and creatinin 

Since 16 September fever-free, exanthema back to normal (1.1 mg/dl), exanthema gone, but 

Further course 

gone on 17 September, no further complica- arthralgias of ankles, wrists, and digital joints 

tions since then 

persist for more than six months including limited 

mobility and requiring NSAID treatment 

CRP: C-reactive protein; CHIKV: Chikungunya virus; DENV: Dengue virus; RT-PCR: reverse transcription-polymerase chain reaction; NSAID: non 

steroidal anti-inflammatory drugs 

67

the high incidence rates of 65.2 per 1,000 population 

previously reported for the Ari Atoll [5], both infections 

could likewise have been acquired on Holiday Island. 

Further, a considerable number of people travel constantly 

between India and Sri Lanka and the tourist 

resorts on the Atolls’ islands of the Maldives where 

they are employed. This frequent exchange may argue 

for a repeated and renewed introduction of CHIKV from 

India or Sri Lanka via viraemic workers or tourists 

and limited local transmission through aedine mosquitoes 

at the respective islands. Analyses of the E1 

gene revealed three nt-mismatches when compared to 

the 2007 case that was analysed in Singapore [6], but 

identical nt sequences to a series of CHIKV strains from 

Sri Lanka (Figure 2) [6,11]. 

Figure 2 

Phylogenetic relationship generated by Maximum 

Parsimony method as implemented in MEGA4 based on 

the complete E1 protein coding sequence (1314 nt) of a set 

of CHIKV of different geographic origin 

99 

AF369024 S27 

10 

FJ513629 Sri 2008 

Mal 2009 

FJ513628 Sri 2008 

GU013528 Sri 2008 

GU013530 Sri 2008 

50 

EU244823 Ita 2007 

FJ513635 Sri 2008 

GU013529 Sri 2008 

65 

FJ513637 Sri 2008 

FJ51379 Sri 2008 

FJ513657 Sri 2008 

63 AB455493 Sri 2006 

59 FJ445427 Sri 2006 

AB455494 Sri 2006 

EF027138 Ind 2006 

EF027134 Ind 2006 

35 

FJ445428 Sri 2007 

EF027137 Ind 2006 

FJ000065 Ind 2006 

EU564335 Ind 2006 

99 

FJ445485 Mal 2007 

EF027136 Ind 2006 

AM258990 Reu 2005 

AM258991 Sey 2005 

EF027135 Ind 2006 

32 AM258995 Com 2005 

AM258992 Reu 2005 

DQ443544 Reu 2006 

49 EF012359 Mau 2006 

EU037962 Mau 2006 

EF027139 Ind 2000 

99 

„African prototype strain“ 

Sri Lanka 

Cluster B 

Sri Lanka 

Cluster A 

„India 2006“ 

Indian Ocean 

Islands at the 

beginning of 

the pandemic 

98 

EF027140 Ind 1963 

EF027141 Ind 1973 

73 EU703761 My 2006 

EU703762 My 2006 

FJ807887 Indo 2007 

99 

FJ807888 Indo 2008 

90 FJ807889 Indo 2008 

68 

96 

EU192143 Indo 2007 

FJ807897 Indo 2007 

Sequence data are provided with their accession number, country 

and year. The term Cluster A and B for sequence data from Sri 

Lanka is adapted from [10]. Please note that the only available 

CHIKV E1 sequence from the Maldives from 2007 clusters together 

with CHIKV from India from 2006, while the CHIKV reported here 

is part of the Cluster B from Sri Lanka in 2008. Sri = Sri Lanka, 

Mal = Maldives, Ita = Italy (imported from India in 2007), Ind = 

India, Reu = Reunion, Sey = Seychelles, Com = Comores, Mau = 

Mauritius, My = Malaysia, Indo = Indonesia. The percentage of 

replicate trees in which the associated taxa clustered together in 

the bootstrap test (100 replicates) are shown next to the branches. 

The branch lengths are informative (bar length corresponds to 10 

nt- differences) 

pandemic CHIKV type that evolved since 2005 

traditional Asian type 

It will be seen in the near future whether more 

cases of chikungunya fever will be reported for the 

Maldives, but we feel that this is already an issue in 

travel medicine although the German Robert Koch 

Institute reported only three chikungunya fever cases 

in returning travellers from the Maldives in 2009 (two 

of which we describe here). A crucial question concerning 

the current global situation on chikungunya fever 

is the adaptation of the pandemic CHIKV strain to 

Ae. albopictus. Aedes aegypti has been long known to 

occur on several islands of the Maldives and seems 

to be the predominant vector on Malé itself while 

Ae. albopictus has established foci on other islands 

where it seems to be the main mosquito vector species 

[5]. We do not know which Aedes species has infected 

the German tourists, but we do know that the A226V 

mutation is suggestive for Ae. albopictus as the vector. 

This particular mosquito is present in many areas 

around the Mediterranean Sea and was responsible 

for a CHIKV outbreak in Italy in 2007 resulting in more 

than 300 cases [12,13]. With a continuing circulation of 

CHIKV in major tourist destinations in Asia and Africa, 

imported cases of chikungunya fever will also be 

seen in Europe and North America. In countries were 

Ae. albopictus is abundant, returning viraemic tourists 

could cause smaller outbreaks. 


Part of this work was supported by the Federal Ministry of 

Education and Research (BMBF) grant 01KI 0712 to MP and 

GD as part of the network “Emerging arthropod-borne viral 

infections in Germany”. The technical assistance of Nadine 

Roßner is gratefully acknowledged. 

References 

1. Powers AM, Logue CH. Changing patterns of Chi¬kungunya 

virus: re-emergence of a zoonotic arbovirus. J Gen Virol. 

2007;88(9):2363–77. 

2. Staples JE, Breiman RF, Powers AM. Chikungunya fever: an 

epidemiological review of a re-emerging infectious disease. 

Clin Infect Dis. 2009;49(6):942-8. 

3. Schuffenecker I, Iteman I, Michault A, Murri S, Frangeul L, 

Vaney MC, et al. Genome micro evolution of chikungunya 

viruses causing the Indian Ocean outreak. PLoS Med. 

2006;3(7):e263. 

4. Tsetsarkin KA, Vanlandingham DL, McGee CE, Higgs S. A single 

mutation in chikungunya virus affects vector specificity and 

epidemic potential. PloS Pathog. 2007;3(12):e201. 

5. Yoosuf AA, Shiham I, Mohamed AJ, Ali G, Luna JM, Pandav R, et 

al. First report of chikungunya from the Maldives. Trans R Soc 

Trop Med Hyg. 2009;103(2):192-6. 

6. Ng LC, Tan LK, Tan CH, Tan SS, Hapuarachchi HC, Pok KY, et 

al. Entomologic and virologic investigation of chikungunya, 

Singapore. Emerg Infect Dis. 2009;15(8):1243-9. 

7. Acute febrile disease – Maldives: (Laamu Atoll) Request for 

information, 13 Jan 2009. In:ProMED-Mail-mail [online]. Boston 

US: International Society for Infectious Diseases: 13 January 

2009. Archive no. 20090114.0150:. Available from: http://www. 

promedmail.org/pls/apex/f?p=2400:1202:60109559749468 

20::NO::F2400_P1202_CHECK_DISPLAY,F2400_P1202_PUB_ 

MAIL_ID:X,75589 

8. Panning M, Grywna K, van Esbroeck M, Emmerich P, Drosten 

C. Chikungunya fever in travelers returning to Europe 

from the Indian Ocean region, 2006. Emerg Infect Dis. 

2008;14(3):416-22. 


9. Panning M, Hess M, Fischer W, Grywna K, Pfeffer M, 

Drosten C. Performance of the RealStar Chikungunya Virus 

Real-Time Reverse Transcription-PCR Kit. J Clin Microbiol. 

2009;47(9):3014-6. 

10. Receveur M, Ezzedine K, Pistone T, Malvy D. Chikungunya 

infection in a French traveler returning from the Maldives, 

October, 2009. Euro Surveill. 2010; 15(8):pii=19494. 



11. Hapuarachchi HC, Bandara KB, Sumanadasa SD, Hapugoda 

MD, Lai YL, Lee KS, et al. Re-emergence of chikungunya virus 

in South-east Asia: virological evidence from Sri Lanka and 

Singapore. J Gen Virol. 2010;91(Pf 4):1067-76. 

12. Angelini P, Macini P, Finarelli AC, Pol C, Venturelli C, Bellini R, 

Dottori M. Chikungunya epidemic outbreak in Emilia-Romagna 

(Italy) during summer 2007. Parassitologia. 2008;50(1-2):97-8. 

13. Bonilauri P, Bellini R, Calzolari M, Angelini R, Venturi L, 

Fallacara F, et al. Chikungunya virus in Aedes albopictus, Italy. 



69


A case of dengue type 3 virus infection imported from 

Africa to Italy, October 2009 

C Nisii (carla.nisii@inmi.it) 1 , F Carletti 1 , C Castilletti 1 , L Bordi 1 , S Meschi 1 , M Selleri 1 , R Chiappini 1 , D Travaglini 1 , M Antonini 1 , S 

Castorina 1 , F N Lauria 1 , P Narciso 1 , M Gentile 1 , L Martini 1 , G Di Perri 2 , S Audagnotto 2 , R Biselli 3 , M Lastilla 3 , A Di Caro 1 , 

M Capobianchi 1 , G Ippolito 1 

1. WHO Collaborating Center for clinical care, diagnosis, response and training on Highly Infectious Diseases, National Institute 

for Infectious Diseases Lazzaro Spallanzani, Rome, Italy 

2. Department of Infectious Diseases, ‘Amedeo di Savoia’ Hospital, Turin, Italy 

3. Italian Air Force, Aeromedical Isolation Unit, Rome, Italy 


Citation style for this article: Nisii C, Carletti F, Castilletti C, Bordi L, Meschi S, Selleri M, Chiappini R, Travaglini D, Antonini M, Castorina S, Lauria FN, Narciso P, 

Gentile M, Martini L, Di Perri G, Audagnotto S, Biselli R, Lastilla M, Di Caro A, Capobianchi M, Ippolito G. A case of dengue type 3 virus infection imported from 

Africa to Italy, October 2009. Euro Surveill. 2010;15(7):pii=19487. Available online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19487 

In October 2009, a traveller returning from Africa to 

Italy was hospitalised with symptoms suggestive of a 

haemorrhagic fever of unknown origin. The patient was 

immediately placed in a special biocontainment unit 

until laboratory investigations confirmed the infection 

to be caused by a dengue serotype 3 virus. This 

case reasserts the importance of returning travellers 

as sentinels of unknown outbreaks occurring in other 

countries, and highlights how the initial symptoms of 

dengue fever resemble those of other haemorrhagic 

fevers, hence the importance of prompt isolation of 

patients until a final diagnosis is reached. 

Case description 

In October 2009, a Senegalese man in his forties, 

who had been living in Italy for 20 years, presented 

to Hospital A in the northern Italian city of Turin three 

days after returning (via Madrid) from a four-month 

visit to his home village in Senegal. During his stay, 

he had never left the village and had not visited any 

healthcare centres. The initial symptoms of disease, a 

persistent fever (>38 °C) accompanied by an unremitting 

headache, started during the flight from Madrid 

to Turin worsened over the previous two days despite 

treatment with paracetamol. 

On the day after hospitalisation, when laboratory test 

results showed a platelet count of 5,000 cells/mm3 as 

well as evidence of altered liver function (aspartate aminotransferases 

(AST): 3,539 U/L; alanine aminotransferases 

(ALT): 815 U/L; lactate dehydrogenase (LDH) 

3,609 U/L; gamma-glutamyl transferases (gamma- 

GT): 112 U/L), the patient was first transferred to an 

infectious diseases hospital in Turin (Hospital B), and 

two days later to the National Institute for Infectious 

Diseases in Rome (Hospital C), which is the Italian 

national reference centre for emerging infections and 

bioterrorism, on suspicion of a viral haemorrhagic 

fever. The patient was admitted to a special biocontainment 

unit during the same night. Over the following 


days, the patient’s clinical condition improved and he 

was discharged nine days after admission. 

Laboratory investigations 

After the patient’s admission, series of tests were 

run in parallel on samples taken at Hospital C and in 

Turin, before the transfer to Rome. The tests included 

in the differential diagnosis ruled out malaria (which 

had already been excluded in Turin), as well as infection 

with the major agents of viral hepatitis, herpes 

simplex virus and viral haemorrhagic fever viruses. 

Serological investigation using an in-house immunofluorescence 

assay (slides prepared with a mix of 

uninfected and dengue type 2-infected Vero cells) 

revealed a high IgG titre against dengue virus soon 

after the onset of symptoms (in a sample taken on 10 

October), with the presence of IgM antibodies at a low 

titre, and reverse-transcription polymerase chain reaction 

(RT-PCR) followed by sequencing of the NS5 region 

confirmed the infectious agent to be a serotype 3 dengue 

virus. In addition, a 224 bp fragment of the E gene 

was amplified and sequenced. The phylogenetic analysis 

of this fragment is shown in the Figure. Our patient 

was infected with a strain belonging to dengue virus 

serotype 3, genotype III closely related to other strains 

found in Africa, but never reported from Senegal before 

the case described here [1]. 

The tests carried out the day before the patient left 

the hospital showed that his IgG titre was still increasing. 

Overall, the immune response was consistent with 

a re-infection with a dengue serotype 3, genotype III 

virus which can lead to a severe form of the disease. 

Case management 

At the time the patient reported to the first hospital in 

Turin, there was no information on any outbreaks of 

dengue virus taking place in Senegal [1]. Therefore, the 

fact that the patient developed symptoms suggestive 

of a haemorrhagic disease led the clinicians to suspect 

also other, more dangerous viral infections, namely 


Lassa virus or Crimean-Congo haemorrhagic fever 

virus, both categorised as biosafety level 4 (BSL4) 

agents [2-3]. As these infections could not be ruled out 

on clinical and epidemiological criteria, it was decided 

to transfer the patient to a high-security isolation 

facility. 

When contacted by Hospital B in Turin, the National 

Institute for Infectious Diseases immediately requested 

the shipping of clinical samples, but because of 

Figure 

Phylogenetic relationship of the strain isolated in October 

2009 in Rome (identified by the black dot) with other 

dengue serotype 3 virus strains described elsewhere [10] 

Sequence identification is as follows: strain name, country and 

year of isolation. 

The scale bar indicates nucleotide substitutions per site. 

Multiple alignment of our sequence and other dengue serotype 3 

virus sequences available in GenBank was generated with 

ClustalW1.7 software included in the Bioedit package. The 

phylogenetic tree was constructed by using nucleotide alignment, 

the Kimura-2-parameters algorithm, and the neighbourjoining 

method implemented in the MEGA 4.1 software. The 

robustness of branching patterns was tested by 1,000 bootstrap 

pseudoreplications. 


logistical difficulties the transport could not be organised. 

It was then decided by the Italian Ministry of 

Health, in consultation with the institute, to make use 

of the procedure already in place in Italy for the transport 

of highly infectious patients under high isolation 

conditions, which included the use of military aircraft 

and equipment of the Italian Air Force and Hospital 

C (stretcher isolators and a high containment ambulance). 

On arrival, the patient was immediately transferred 

to the biocontainment unit and was attended to 

by specially trained, dedicated staff employing all necessary 

biosafety precautions. 

Conclusions 

We have described a case of dengue virus infection 

imported into Italy from Africa, which to date has been 

a rare occurrence as most cases seen in European 

travel clinics have been imported either from Asia or 

the Americas. Dengue virus has been known to circulate 

in parts of Africa for decades, and Senegal in particular 

has experienced several outbreaks, mainly of 

dengue serotype 2 virus [1]. Following our report [4], 

which was the first for a dengue serotype 3 virus in 

Senegal, the country’s health authorities were alerted 

to a possible epidemic and since then, over 50 cases 

have been registered in the country [5-6], confirming 

the importance of returning travellers as sentinels of 

as yet unreported outbreaks occurring abroad [7]. 

From a clinical point of view, the patient did not meet 

the World Health Organization’s (WHO) criteria for 

severe dengue fever [8], but presented a set of warning 

signs indicative of haemorrhagic fever such as 

liver enlargement (>2 cm), altered liver enzymes (20fold) 

and a rapid decrease in platelet count. Therefore, 

and because the information available at the time did 

not allow excluding the involvement of a BSL4 agent, 

the decision to treat the case as a potential viral 

haemorrhagic fever was made early in the management 

process. The recent report of a case of Marburg 

haemorrhagic fever imported from Uganda into the 

United States [9] that was diagnosed retrospectively 

in a serum sample archived six months earlier after it 

had tested negative for agents of viral haemorrhagic 

fever, should teach us that in similar cases, patients 

should immediately be placed in an adequate containment 

facility until a final diagnosis is reached. Our recommendation 

is to perform all diagnostic procedures 

safely at the appropriate biosafety level, and as a general 

rule any activities involving patient samples at a 

level lower than BSL3 should be kept to a minimum. 

It remains a problem that the transport of samples is 

still an unsolved issue in Italy as well as in Europe and 

worldwide. The timely shipping of biological samples 

in advance of the patient’s arrival would have shortened 

the time to reach a diagnosis and avoided the 

extra costs of unnecessary biosafety measures. 

71

References 

1. World Health Organization. Dengue in Africa: emergence of 

DENV-3, Côte d’Ivoire, 2008. Wkly Epidemiol Rec. 2009;84(11- 

12):85-8. Available from: http://www.who.int/wer/2009/ 

wer8411_12.pdf 

2. Laboratory Biosafety Manual. Second edition (revised). 

Geneva: World Health Organisation; 2003. WHO/CDS/ 

CSR/LYO/2003.4. Available from: http://www.who.int/csr/ 

resources/publications/biosafety/Labbiosafety.pdf 

3. JY Richmond, RW McKinney, editors. Biosafety in 

Microbiological and Biomedical Laboratories. Fourth edition. 

U.S. Department of Health and Human Services, Public Health 

Service, Centers for Disease Control and Prevention and 

National Institutes of Health; 1999. Available from: http:// 

www.cdc.gov/od/ohs/pdffiles/4th%20BMBL.pdf 

4. Hemorrhagic fever – Italy ex Senegal (02): dengue. In: ProMEDmail 


Diseases; 16 October 2009. Archive no. 20091016.3559. 

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f?p=2400:1001:::NO::F2400_P1001_BACK_PAGE,F2400_ 

P1001_PUB_MAIL_ID:1000%2C79642 

5. Dengue/DHF Update 2009 (47). In: ProMED-mail 


Diseases; 15 November 2009. Archive no. 20091115.3944. 



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6. Dengue/DHF Update 2009 (48). In: ProMED-mail [online]. 

Boston US: International Society for Infectious Diseases; 

23 November 2009. Archive no. 20091123.4016. 



P1001_PUB_MAIL_ID:1000%2C80198 

7. Schwartz E, Weld LH, Wilder-Smith A, von Sonnenburg F, 

Keystones JS, Kain KC, et al. Seasonality, annual trends, and 

characteristics of dengue among ill returned travellers, 1997- 

2006. Emerg Infect Dis. 2008;14(7):1081-8. 

8. Dengue: Guidelines for diagnosis, treatment, prevention 

and control. New edition. Geneva: World Health 

Organization, 2009. ISBN 978 92 4 154787 1. WHO/HTM/ 

NTD/DEN/2009.1. Available from: http://whqlibdoc.who.int/ 

publications/2009/9789241547871_eng.pdf 

9. Marburg hemorrhagic fever – USA ex Uganda 2008. In: 

ProMED-mail [online]. Boston US: International Society 

for Infectious Diseases; 31 January 2009. Archive no. 

20090131.0423. Available from: http://www.promedmail. 

org/pls/apex/f?p=2400:1001:::NO::F2400_P1001_BACK_ 

PAGE,F2400_P1001_PUB_MAIL_ID:1000%2C75888 

10. Weaver SC, Vasilakis N. Molecular evolution of Dengue viruses: 

contributions of phylogenetics to understanding the history 

and epidemiology of the preeminent arboviral disease. Infect 

Genet Evol. 2009;9(4):523-40. 



Recent expansion of dengue virus serotype 3 in 

West Africa 

L Franco (francolet@isciii.es) 1,2 , A Di Caro 3,2 , F Carletti 3 , O Vapalahti 4,2 , C Renaudat 5,2 , H Zeller 6 , A Tenorio 1,2 

1. National Centre for Microbiology, Instituto Carlos III, Madrid, Spain 

2. European Network for Imported Viral Disease - Collaborative Laboratory Response Network (ENIVD-CLRN). www.enivd.org 

3. National Institute for Infectious Diseases ‘Lazzaro Spallanzani’, Rome, Italy 

4. Haartman Institute and Dept of Veterinary Biosciences, University of Helsinki, Finland 

5. National Reference Centre for Arboviruses, Institut Pasteur, Paris, France 

6. European Centre for Disease Prevention and Control, Stockholm, Sweden 


Citation style for this article: Franco L, Di Caro A, Carletti F, Vapalahti O, Renaudat C, Zeller H, Tenorio A. Recent expansion of dengue virus serotype 3 in West Africa. 


Due to non-existing or limited surveillance in Africa, 

little is known about the epidemiology of dengue illness 

in the continent. Serological and virological 

data obtained from returning European travellers is a 

key complement to this often flawed information. In 

the past years, dengue 3 virus has emerged in West 

Africa and has been detected in travellers returning to 

Europe. The first dengue epidemic in Cape Verde with 

more than 17,000 cases from September to December 

2009 demonstrated that dengue virus is still expanding 

worldwide to new territories. 

Introduction 

Dengue virus is widely distributed in tropical and subtropical 

countries and is transmitted by day-biting mosquitoes 

of the genus Aedes. It often goes unrecognised 

in African countries, where the lack of surveillance 

systems, or their poor implementation, is the cause of 

missing information on dengue virus activity [1]. 

Laboratory-based surveillance of dengue virus infection 

in febrile travellers could provide useful information 

about the different dengue virus serotypes 

circulating worldwide and in particular those circulating 

in areas where limited surveillance is available. 

To this end, the European Network for Imported Viral 

Disease - Collaborative Laboratory Response Network 

(ENIVD-CLRN network) provides outbreak support, in 

particular related to laboratory diagnostics, to assist 

European Union (EU) Member States, candidate countries 

and members of the European Economic Area 

and European Free Trade Association (EEA/EFTA) in 

detecting, investigating and responding to outbreakprone 

diseases, imported, rare or unknown infectious 

agents, or outbreaks related to the intentional release 

of pathogens. 

A large outbreak of dengue illness with more than 

17,000 cases occurred in the Cape Verde archipelago 

at the end of 2009 [2]. It was the first time that dengue 

virus was detected in the archipelago. Concomitant 



detection of dengue virus in Senegal and identification 

of several imported cases among travellers returning 

from West Africa were reported. This article provides a 

brief review of historic reports of dengue virus in Africa 

focused on West Africa and summarises the recent outbreaks 

and the links to imported cases of dengue virus 

infection in Europe. 

Dengue virus in West Africa 

The burden of dengue virus infection in Africa has not 

been estimated yet. Outbreaks of dengue fever and 

dengue haemorrhagic fever are poorly documented, 

however, we cannot conclude that mild and severe 

dengue infection is infrequent in African countries. The 

circulation of different dengue virus serotypes is also 

poorly documented. Nevertheless some information is 

provided in publications on outbreaks and serosurvey 

studies in Africa and reports involving dengue virus 

infection in travellers. 

A retrospective serological study in 1956 [3] suggests 

that dengue virus caused an epidemic in Durban, South 

Africa in 1927. This report is the first documented dengue 

virus epidemic in Africa. It was not until the end 

of the 1960s, however, that the virus responsible for 

dengue fever outbreaks in Africa could be isolated. 

The first dengue virus (DENV) isolate was DENV-1, 

detected in Nigeria in 1964 [4]. Since then DENV-1, 2 

and 4 have been circulating in West Africa although the 

main serotype reported has been DENV-2 [1]. Viral isolates 

have been predominantly detected in wild-caught 

mosquitoes (Aedes luteocephalus, Ae. taylori and/or 

Ae. furcifer) involved in sylvatic transmission cycles in 

Senegal and Nigeria, and from a few cases in humans 

who were in contact with forest cycles [5,6]. DENV-2 

from sylvatic cycles have also been isolated in Côte 

d’Ivoire, Burkina Faso and Guinea [5,7,8]. More recently 

in 2005, DENV-2 was identified in a traveller returning 

from Ghana [9]. The last detection of DENV-4 in West 

Africa was in the 1980s in two inhabitants of Dakar, 

Senegal [10]. 

73

The first description of DENV-3 activity in Africa was 

related to outbreaks detected during 1984 and 1985 in 

Pemba, Mozambique, with two deaths due to dengue 

haemorrhagic fever [11]. DENV-3 was then detected in 

1993 in Somalia and areas around the Persian Gulf [12]. 

Phylogenetic studies suggested that these outbreaks 

were caused by a virus imported from the Indian subcontinent 

[13]. DENV-3 circulation in West Africa was 

first identified in a traveller returning to Spain from 

Cameroon in 2006 and subsequently in a traveller 

returning to Spain from Senegal in 2007 (C. Domingo 

et al., unpublished results). However the first article 

on DENV-3 in West Africa was published in 2008, when 

DENV-3 was detected co-circulating with yellow fever in 

Côte d’Ivoire [14]. 

Dengue virus importation from Africa into 

Europe 

Reports on the importation of dengue virus to Europe 

have been increasing since the 1990s. Some of these 

are publications or reports from single countries or networks 

and show that the frequency of travel-acquired 

Figure 1 

West African countries where dengue serotypes have been 

identified in recent years (2006-2009) 

DENV-1 Burkina Faso 2007 

DENV-2 Côte d’Ivoire 2007 

DENV-2 Gabon 2007 

DENV-2 Senegal 2008 

DENV-2 Mali 2008 

DENV-3 Cameroon 2006 

DENV-3 Côte d’Ivoire 2008 

DENV-3 Senegal 2007, 2009 

DENV-3 Cape Verde 2009 

DENV: dengue virus. 

Sources: ENIVD-CLRN, INVS (France) ,WHO, ECDC. 

dengue virus infections in Africa is low compared to 

south-east Asia and the Americas [15,16]. This distribution 

is due to two main factors: worldwide dengue 

virus activity and the popularity of certain countries 

as tourist destinations. In a study by the European 

Network on Imported Infectious Disease Surveillance 

(TropNetEurop) covering 481 European travellers 

between 1999 and 2002, 8% of dengue fever cases 

were imported from Africa [17], a proportion similar 

to that found in other European studies. In France, 

between 2002 and 2005, 14% of imported dengue 

fever cases originated in Africa [18]. Ten per cent of 

the cases in Austrian and Finnish travellers were also 

acquired in Africa, as analysed over a 15-year (1990- 

2005) and 10-year (1999-2009) period, respectively 

[19,20]. In 2008, dengue virus cases imported from 

Africa reported by TropNetEurop dropped to 4% [21]. 

Dengue virus importation from 

West Africa (2006-2008) 

In recent years, dengue fever has been documented 

in travellers returning from several West African 

countries, caused in particular by DENV-3 which was 

recently identified in that region (Figure 1). 

From January 2006 to August 2008, 19 imported cases 

of dengue virus infection were reported in travellers 

from West Africa in France: 11 cases from Côte d’Ivoire 

(one in 2006, three in 2007 and seven in 2008), four 

cases in Burkina Faso (one in 2006 and three in 2007), 

two cases in Benin in 2006, one case in Senegal in 2007 

and one case in Mali in 2008 [22]. Dengue serotypes 

detected during this period were DENV-1 in Burkina 

Faso [22], DENV-2 in Côte d’Ivoire [22] and DENV-2 in 

a simultaneous outbreak of chikungunya and dengue 

viruses in Gabon [23], all in 2007. In 2008, DENV-2 

serotype was identified in Mali [14] and in Senegal 

[24]. Moreover, DENV-3 was detected in a Japanese 

tourist and in a French expatriate returning from Côte 

d’Ivoire between May and July 2008 [25]. DENV-3 activity 

was also detected in East Africa in 2008 in a Finnish 

traveller returning from Eritrea (O. Vapalahti, personal 

communication). 

Recent DENV-3 activity in West Africa 

In the beginning of October 2009, a case of dengue 

fever was reported in a Senegalese returning 

to Italy after a holiday in his home country. DENV-3 

infection was diagnosed at the National Institute for 

Infectious Diseases in Rome [26]. At the same time 

France reported DENV-3 in travellers returning from 

Senegal (C. Renaudat, personal communication). Also, 

ProMED posted several archives describing dengue 

virus outbreaks in Senegal: in the Kedoungo region 

[27] and Dakar [28]. The Pasteur Institute in Dakar 

identified DENV-3 in febrile patients (A. Sall, personal 

communication). 

Meanwhile, an unprecedented outbreak has been 

detected in the Cape Verde archipelago in the beginning 

of September 2009 (week 40) [29]. This is the first 


eport of dengue virus activity in that country (Figure 

1). The highest number of cases were reported during 

week 45 (5,512 cases), decreasing in week 47 to 

1,447 cases and finally five cases in week 53. A total of 

17,224 cases including six deaths were reported from 

18 of the 22 municipalities in Cape Verde by the end 

of 2009. The municipality of Praia on Santiago island, 

notified the highest number of cases (13,000 cases) 

followed by Sao Felipe in Fogo Island (3,000 cases) 

[2]. The first samples tested at the Pasteur Institute in 

Dakar confirmed DENV-3 circulation [29]. 

Recently imported DENV-3 cases in Europe and the 

identified outbreaks in West Africa suggest that this 

serotype is spreading in the region. 

We used a phylogenetic approach in order to determine 

genotype association among the recent DENV-3 

circulating in the region, using sequences provided by 

ENIVD-CLRN laboratories. DENV-3 viruses are divided 

into four geographically different genotypes (I, II, III, 

and IV) [13]. The emergence of a virulent lineage of genotype 

III in Sri Lanka at the end of 1980s was largely 

associated with a high incidence of the disease and the 

emergence of Dengue haemorrhagic fever in Asia and 

Figure 2 

Phylogenetic tree of DENV-3 sequences 


the Americas. All DENV-3 detected in East Africa from 

1984 to 1993 [13] belonged to this lineage of genotype 

III (Figure 2). 

Also, isolates from geographically distant outbreaks, 

strains ENIVD Spain ex Cameroon 2006, Japan ex Ivory 

Coast 2008 and Saudi Arabia 2004 (Figure 2), and from 

Eritrea in 2008 (not shown in Figure 2; E. Huhtamo 

et al, unpublished results) belong to DENV-3 genotype 

III. The DENV-3 strain that circulated in Senegal in 2009 

and was isolated from a traveller in Italy [26], also 

belonged to genotype III and was closely related to 

the DENV-3 strain circulating in Côte d’Ivoire in 2008. 

Therefore, the DENV-3 that has emerged in the Cape 

Verde archipelago is likely to have been introduced 

from a West African country due to the geographical 

proximity with strong trade and travel activities. 

Future outlook 

The recent DENV-3 expansion in West Africa was first 

detected in European travellers returning from this 

area, which triggered the alert for active surveillance 

in the exporting countries. 

This tree is based on a 139 nt fragment of the E gene. Sequence identification is as follows: country of origin, year of identification. The 

sequences from imported cases are labelled as ENIVD/country of detection/country of exportation/year of detection. Phylogenetic analysis 

was conducting using MEGA 4.0 (Tamura, Dudley, Nei, and Kumar 2007). Genetic distance was calculated with the Tamura Nei algorithm. 

Phylogentic tree was constructed using Neighbor-Joining model and the resultant tree was tested by Bootstrap (1,000 Replicates). Only 

bootstrap probabilities over 60% are shown. 

75

However, although most of the African countries are 

prepared for surveillance of yellow fever and human 

immunodeficiency virus infections, most of them lack 

specific methods for dengue virus diagnostics and 

require new diagnostic tools. As a recent example of 

such technology transfer, the Pasteur Institute laboratories 

in Paris and Dakar have implemented differential 

diagnostics for dengue at the Pasteur Institute in 

Abidjan [14]. This model of cooperation is required also 

in other African countries. 

As long as active dengue virus surveillance is poorly 

implemented in Africa, the study of febrile travellers 

returning to Europe could help to detect viral activity 

on the African continent. 

As part of the ENIVD-CLRN, a collaborative study on 

imported chikungunya and dengue virus infections in 

European travellers will start in 2010. The aim of the 

study is to complete the global map of chikungunya 

and dengue virus circulation, including the global distribution 

of viral genotypes of those viruses. It will permit 

clinicians to compare the clinical symptoms, signs 

and analytical data of imported cases in Europe. The 

surveillance of travellers returning to Europe will continue 

to improve our knowledge about dengue virus 

distribution in Africa. 

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6. Vasilakis N, Tesh RB, Weaver SC. Sylvatic dengue type 

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considerations. Emerg Infect Dis. 2003;9(3):362-7. 

9. Huhtamo E, Uzcátegui N, Siikamäki H, Saarinen A, Piiparinen 

H, Vaheri A, et al. Molecular epidemiology of dengue 

virus strains from Finnish Travelers. Emerg Infect Dis. 

2008;14(1):80-3. 

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of dengue 2 and dengue 4 viruses from patients in Senegal. 

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Arthur RR, et al. Dengue fever in US troops during Operation 

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Emergence and global spread of dengue serotype 3, Subtype 

III virus. Emerg Infec Dis. 2003; 9(7):800-9. 

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Dengue type 3 virus infections in European travellers 

returning from the Comoros and Zanzibar, February- 

April 2010 

P Gautret (surveillance@eurotravnet.eu) 1,2 , F. Simon 2 , H Hervius Askling 3 , O Bouchaud 4 , I Leparc-Goffart 5 , L Ninove 6 , P Parola 1 , 

for EuroTravNet 1 

1. Infectious and Tropical Diseases Unit, Hospital Nord, Marseille, France 

2. Infectious and Tropical Diseases Unit, Military Hospital Lavéran, Marseille, France 

3. Department of Infectious Diseases, Karolinska University Hospital, Stockholm, Sweden 

4. Infectious and Tropical Diseases Unit, Avicenne Hospital, Bobigny, France 

5. Associated National Reference Center for Arboviruses, IRBA-IMTSSA, Marseille, France 

6. Virology Laboratory, AP-HM Timone, Marseille, France 


Citation style for this article: Gautret P, Simon F, Hervius Askling H, Bouchaud O, Leparc-Goffart I, Ninove L, Parola P, for EuroTravNet. Dengue type 3 virus 

infections in European travellers returning from the Comoros and Zanzibar, February-April 2010. Euro Surveill. 2010;15(15):pii=19541. Available online: http://www. 


In late February-early April 2010, five cases of dengue 

fever were diagnosed in returning travellers in Europe 

in EurotravNet sites in Sweden and France in patients 

with travel history to the Comoros and/or Zanzibar, 

Tanzania. Four cases were non-complicated dengue 

fever and one case dengue hemorrhagic fever. Three 

patients were viraemic at the time of diagnosis and 

infected with Dengue type 3 virus. 

An estimated 100 million cases of dengue fever and 

250,000 cases of dengue haemorrhagic fever occur 

annually worldwide [1]. The past 20 years have seen 

a dramatic geographic expansion of epidemic dengue 

fever from Southeast Asia to the South Pacific Islands, 

the Caribbean, and the Americans. An increasing 

number of reports of dengue fever and associated illness 

among travellers to dengue virus–infected areas 

paralleled the changing epidemiology of dengue in 

local populations [1]. 

In 2010 (until 14 April), five cases of dengue fever 

including one case of dengue haemorrhagic fever, have 

been reported from EurotravNet sites in France and 

Sweden, in four travellers returning from the Comoros 

and one traveller returning from Zanzibar, Tanzania. 

EurotravNet, the Network for travel medicine and tropical 

diseases of the European Centre for Disease Control 

consists of 14 core sites in nine European countries and 

participants monitor travel related infectious diseases 

in Europe (www.eurotravnet.eu). 

Case reports 

Cases were diagnosed in Paris (one case) and Marseille 

(three cases), France and Stockholm, Sweden (one 

case). The age of cases ranged from 41 to 69 years, 

three were females, two males. All travellers to the 

Comoros had visited friends and relatives where they 

had stayed between 15 and 93 days in the period from 

December to March. The case returning from Zanzibar 



who had travelled as a tourist, had stayed for two days 

in Stone Town and seven days in Nungwy. All cases 

were non-complicated dengue except for one case of 

dengue hemorrhagic fever. Detailed clinical presentations 

and onset of symptoms after return and laboratory 

findings are displayed in Table 1. 

Cases were confirmed by serology and four were positive 

for IgM and IgG and once case positive for IgM 

only. Three cases were confirmed as dengue type 3 

virus (DENV-3) by PCR. 

Discussion and conclusion 

Six autochthonous cases of dengue fever were recently 

identified in the Comoros (March 2010). Additional 

cases were identified in individuals with travel history 

from the Comoros, in Madagascar (1 case), Mayotte (3 

cases) and Reunion Island (1 case) [1,2]. In addition, two 

cases were potentially imported from Tanzania to Japan 

[2,3]. DENV-3 was identified in the cases in Madagascar 

and Japan. These results indicate that DENV-3 is currently 

circulating in the Comoros and Zanzibar, and 

given that the last outbreak in the Comoros took place 

in 1993 and involved DENV-1,[4] we may face a situation 

with the possibility for the emergence of a new 

outbreak including possible severe cases, similarly to 

what was recently observed in Sri-Lanka, East Africa 

and Latin America [5]. 

In order to protect themselves, travellers to areas 

where vector-borne diseases such as dengue fever 

and malaria are present should be advised to adopt 

some protective measures to avoid mosquito bites. 

Moreover, physicians should be prepared to diagnose 

and manage imported cases of dengue fever in 

travellers returning from the Comoros and East Africa 

early. Viraemic patients may spread the infection to 

regions where competent vectors are present, including 

the Mediterranean area and the south of Europe. 

77

In metropolitan France, dengue fever is a mandatory 

notifiable disease since Aedes albopictus has become 

established in the Mediterranean French littoral in 

2004 and in Corsica in 2006 [6]. 

A. albopictus were found in August 2009, in the centre 

of Marseille [7]. Given the intensity of population 

flows between the Comoros and Marseille, especially 

during summer, early detection of viraemic travellers 

and entomological surveillance are critical. The establishment 

of A. albopictus, the vector for dengue and 

chikungunya viruses, in the south of Europe and the 

presence of viraemic imported cases of dengue fever 

in these regions could lead to autochthonous transmission 

[8]. In 2007, a viraemic patient infected with 

chikungunya virus was the source of an outbreak of 

chikungunya in Emilia-Romagna, Italy, with 205 cases 

occurring between 4 July and 27 September 2007 [9]. 

So far no sustained outbreaks from imported dengue 

fever or chickungunya have occurred but vigilance is 

needed. 

Our report confirms that returning travelers may serve 

as sentinels for local outbreaks of dengue fever in 

endemic areas [10]. Finally, the case presenting exclusively 

with fever and without additional symptoms 

commonly associated with dengue fever, illustrates 

that dengue fever should be included early in the differential 

diagnosis in febrile travellers particularly 

when returning from areas with potential transmission 

of the disease [11]. 

References 


Keystone, JS, Kain KC, et al., for the GeoSentinel Surveillance 

Network. Seasonality, annual trends, and characteristics of 

dengue among ill returned travelers, 1997–2006. Emerg Infect 

Dis. 2008; 14(7):1081-8. 

2. Dengue/DHF update 2010 (15). In: Promed-mail [online]. Boston 

US: International Society for Infectious Diseases: 23 March 

2010. Archive no. 20100323.0922. Available from: http://www. 

promedmail.org/pls/apex/f?p=2400:1202:51849094839886 

40::NO::F2400_P1202_CHECK_DISPLAY,F2400_P1202_PUB_ 

MAIL_ID:X,81876 

3. Institut de Veille Sanitaire (InVS). [Alerte dengue dans le 

sud-ouest de l’Océan Indien. Point épidémiologique N°07 

au 25 mars 2010]. [French]. Available from: http://www. 

invs.sante.fr/surveillance/dengue/asie_se_ocean_indien/ 

pe_dengue_250310.pdf 

4. Boisier P, Morvan JM, Laventure S, Charrier N, Martin E, Ouledi 

A, et al. [Dengue 1 epidemic in the Grand Comoro Island 

(Federal Islamic Republic of the Comoros). March-May 1993]. 

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5. Messer WB, Gubler DJ, Harris E, Sivananthan K, de Silva AM. 

Emergence and global spread of dengue serotype S, subtype III 

virus. Emerg Infect Dis. 2003;9(7):800-9. 

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Mediterranéen (EID) Méditerranée. [Enquête entomologique 

dans le cadre de la surveillance d’Aedes albopictus]. 

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Internal document. [French]. 

8. Soumahoro MK, Fontenille D, Turbelin C, Pelat C, Boyd A, 

Flahault A, et al. Imported chikungunya virus infection. Emerg 

Infect Dis. 2010;16(1):162-3. 

9. Rezza G, Nicoletti L, Angelini R, Romi R, Finarelli AC, Panning 

M, et al. ; CHIKV study group. Infection with chikungunya 

virus in Italy: an outbreak in a temperate region. Lancet. 

2007;370(9602):1840-6. 

Table 

Clinical characteristics dengue fever in travellers returning from the Comoros and Zanzibar (Tanzania), February-April 

2010 (n=5) 

Clinical symptoms 

Case 1 Case 2 Case 3 Case 4 Case 5 

Fever, shivering, 

myalgias 

Fever, arthralgias, myalgias, 

diarrhoea, headache, seizures, 

bleeding (haematemesis, ulorrhagia, 

metrorrhagia) 


Fever 

Fever, shivering, arthralgias, 

myalgias, 

headaches, diffused 

non-petechial rash 


anorexia, cough, 

diarrhoea 

Onset after return 

(in days) 

7 4 (before return) 1 0 0 

Leucocyte count/µL 5,280 2,300 3,200 4,900 7,500 

Platelet count/µL 83,000 34,000 15,500 13,000 56,000 

SGOT (U/L) 214 (norm 4.6) 257 (norm 6.5) Not available 506 (norm 10.1) 214 (norm 4.6) 

SGPT (U/L) 125 (norm 1.9) 183 (norm 4.5) Not available 191 (norm 3.2) 101 (norm 1.5) 

GGT (U/L) 54 (norm 1.5) 70 (norm 2.3) Not available 278 (norm 4.6) 47 (norm 1.3) 

LDH (U/L) 785 (norm 1.6) 1,221 (norm 3.2) Not available 2,199 (norm 3.5) 765 (norm 1.6) 

Serology IgM + IgGa IgM + IgGb IgM + IgGc IgMd IgM + IgGa PCR Negative DENV-3e Not available DENV-3e DENV-3e DENV-3: Dengue virus type 3; GGT: Gamma-glutamyl transpeptidase; LDH: Lactate dehydrogenase; norm: normal upper value; SGOT: Serum 

glutamic oxaloacetic transaminase; SGPT: Serum glutamic pyruvic transaminase. 

a Detected by in house enzyme-linked immunosorbent assay (Associated National Reference Center for arboviruses IRBA-IMTSSA, Marseille). 

b Panbio ELISA IgM and IgG (rapid test dengue duo Ig M and Ig G Panbio negative); EIA Biotrin Ig M (1.7; N< 1.5) and Ig G (3.6; N< 1.5). 

c Dengue IgM specific for dengue virus was detected (54 PBU; ≥11 PBU positive) by ELISA (Dengue IgM PanBio) and high levels of dengue IgG 

was detected with IFI. 

d Detected by indirect immunofluorescence (IFI) test (Standard Diagnostics Dengue Duo) and confirmed by ELISA (Panbio Dengue DUO Test). 

e DENV-3 RNA was demonstrated in serum using 4 real time reverse-transcription-PCR (Taqman RT-PCR)-based assays [12] .

10. Ninove L, Parola P, Baronti C, De Lamballerie X, Gautret 

P, Doudier B, et al. Dengue virus type 3 infection in 

traveler returning from west Africa. Emerg Infect Dis. 

2009;15(11):1871-2. 

11. Askling HH, Lesko B, Vene S, Berndtson A, Björkman P, 

Bläckberg J, et al. Serologic analysis of returned travelers with 

fever, Sweden. Emerg Infect Dis. 2009;15(11):1805-8. 

12. Leparc-Goffart I, Baragatti M, Temmam S, Tuiskunen A, 

Moureau G, Charrel R, et al. Development and validation of 

real-time one-step reverse transcription-PCR for the detection 

and typing of dengue viruses. J Clin Virol. 2009; 45(1):61-6. 


79


Fatal and mild primary dengue virus infections 

imported to Norway from Africa and south-east Asia, 

2008-2010 

K Vainio 1 , S Noraas 2 , M Holmberg 3 , H Fremstad 1 , M Wahlstrøm 4 , G Ånestad 1 , S Dudman (susannegjeruldsen.dudman@fhi.no) 1 

1. Norwegian Institute of Public Health, Department of Virology, Oslo, Norway 

2. Sørlandet Hospital, Kristiansand, Norway 

3. Vestfold Hospital, Tønsberg, Norway 

4. Swedish Institute for Infectious Disease Control, Solna, Sweden 


Vainio K, Noraas S, Holmberg M, Fremstad H, Wahlstrøm M, Ånestad G, Dudman S. Fatal and mild primary dengue virus infections imported to Norway from Africa 

and south-east Asia, 2008-2010. Euro Surveill. 2010;15(38):pii=19666. Available online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19666 

Between 2008 and 2010, eight cases of viraemic dengue 

fever in travellers were diagnosed in Norway. They 

had returned from Eritrea, Thailand and Indonesia. 

All cases were primary dengue infections, seven noncomplicated 

dengue fever and one dengue shock 

syndrome with a fatal outcome. Four patients were 

infected with dengue virus serotype 1, one with type 

2 and three with type 3. Two cases from Thailand, 

the fatal case and the two imported from Eritrea were 

infected with type 1. 

Introduction 

Global incidence of dengue fever has increased 

strongly in recent decades, and dengue infections are 

now endemic in more than 120 countries throughout 

the world [1-3]. South-east Asia is the most important 

region of origin for the import of dengue fever 

into Europe [4]. In recent years, dengue virus has 

become a more prevalent cause of imported fever in 

Norwegian patients than malaria. Due to this increase, 

the Norwegian Institute of Public Health (NIPH) has 

recently proposed to the health authorities to make 

dengue fever a notifiable disease. Most cases diagnosed 

in Norway have been mild, but there have also 

been several cases with complicated dengue infections, 

including one fatal case in 2005 [5]. 

Of all dengue cases confirmed at the NIPH, we describe 

here the eight viraemic cases imported to Norway 

between 2008 and 2010. 

Case descriptions 

The eight viraemic cases reported in Norway between 

2008 (n=1), 2009 (n=1) and 2010 (n=6), were imported 

from Eritrea (n=2), Thailand (n=4) and Indonesia (n=2). 

The patient’s ages ranged from 19 to 65 years, five 

females and three males. None of the cases had evidence 

of previous dengue virus infection based on 

their medical history and serological evidence. Seven 

of the cases had non-complicated dengue fever, but 

Article published on 23 September 2010 

one patient suffered from dengue shock syndrome with 

a fatal outcome. 

The fatal case first presented to the local health centre 

with a febrile viral influenza-like illness four days after 

returning from Thailand [6]. Nine days after returning, 

the patient visited the emergency centre as no relief 

was obtained from using paracetamol and ibuprofen, 

but returned home to continue ibuprofen treatment. 

Twelve hours later the patient was admitted to the 

intensive care unit, but was then suffering from circulatory 

collapse and died within a few hours. During the 

resuscitation attempts there was abnormal bleeding 

from the endotracheal tube and needle injection sites. 

Laboratory results showed a fall in haemoglobin from 

15 to 7 g/dL and thrombocytopenia. 

Another patient returning with dengue fever from Bali 

was examined for airway infections due to hoarseness 

and nasal congestion. Mycoplasma pneumoniae was 

detected by PCR in nasopharyngeal secretions and 

erythromycin tablets were prescribed. Clinical characteristics 

of all patients and their laboratory results are 

displayed in the Table. 

Laboratory methods 

Acute phase sera were obtained from the eight patients. 

Cases with no previous history of dengue virus infection 

and acute serum negative for anti-dengue IgG 

were defined as primary infections. Convalescent 

sera were available from only three patients and were 

taken 18-22 days after the acute sera. The acute samples 

were initially tested at the local laboratory and 

the positive samples were then referred to the virology 

laboratory at the NIPH for confirmation, except for 

one sample which was analysed directly at the NIPH. 

Infection with dengue virus was initially diagnosed in 

seven of the travellers by Panbio Dengue Duo IgM and 

IgG Rapid Strip Test (Inverness Medical Innovations, 

Australia) or SD Bioline Dengue NS1 Antigen and IgG/ 

IgM tests (Standard Diagnostics, South Korea). All 


Table 

Characteristics of the viraemic cases of dengue fever in Norway, 2008–2010 (n=8) 

Case 1 2008 Case 2 2009 Case 3 2010 Case 4 2010 Case 5 2010 Case 6 2010 Case 7 2010 Case 8 2010 



Fever, 

Fever, 

Fever, headache, vomiting, nuchal rigidity, Fever, headache, hoarseness, 

Fever, Fever, cough, shivering, headache, 

Clinical symptoms 

dyspnoea, cyanosis, hypothermia, fatigue, muscle aches, nasal congestion, 

Fever 

headache headache headache, muscle aches, 

confusion, hypotension, cardiac arrest non-petechial rash epistaxis, non- 

arthralgias arthralgias 

petechial rash 

Onset of symptoms after 

Symptoms started 

4 

1 2 3 2 1 1 

return (days) 

before return 

Travel destination Thailand Bali, Indonesia Bali, Indonesia Eritrea Thailand Eritrea Thailand Thailand 

Initial laboratory findings DENV IgM+ a Dengue IgM(+) a Mycoplasma DENV NS1 

pneumoniae PCR+ antigen+ b 

DENV NS1 

antigen+ b 

DENV NS1 

antigen+ b 

DENV NS1 

antigen+ b 

DENV NS1 

antigen+/IgM+ b 

DENV serology IgG- / IgM+ c 

Acute sample: 

IgG- / IgM+ c 

Convalescent sample: 

IgG+ / IgM+ c 

Acute sample: 

IgG- / IgM- c 

Convalescent 

sample: 

IgG+ / IgM+ c 

IgG- / IgM- c IgG- / IgM- c IgG- / IgM- c IgG- / IgM+ c 

Acute sample: 

IgG- / IgM+ c 

Convalescent 

sample: 

IgG+ / IgM+ c 

DENV PCR+ (day of 

sampling after symptom 

PCR+ (day 9) 

debut) 

d 

PCR+ (day of sampling 

after symptom debut 

unknown) d 

PCR+ (day 1) d PCR+ (day 3) d PCR+ (day 4) d PCR+ (day 5) d PCR+ (day 4) d PCR+ (day 6) d 

DENV serotype DENV-1 DENV-2 DENV-3 DENV-1 DENV-3 DENV-1 DENV-3 DENV-1 

DENV-1: dengue virus serotype 1; DENV-2: dengue virus serotype 2; DENV-3: dengue virus serotype 3 

a Detected by Panbio Dengue Duo IgM and IgG rapid test 

b Detected by SD Bioline Dengue NS1 Antigen, IgG and IgM test 

c Detected by Euroimmun Indirect Immunofluorescens anti-dengue virus IgM and IgG test 

d Detected by RT-PCR assays [7,8] 

81

samples were further analysed at the NIPH for the 

presence of dengue IgG and IgM antibodies using a 

commercial indirect immunofluorescence assay (IFA) 

(Euroimmun AG, Germany) and by a reverse transcription 

(RT) PCR detecting the four dengue serotypes [7,8]. 

The virus strains were characterised by direct sequencing 

of the PCR-products and a phylogenetic tree was 

obtained by comparing these strains with other dengue 

virus strains available in the NCBI GenBank sequence 

database. 


We have described eight cases of viraemic dengue 

virus primary infection imported from endemic areas 

to Norway in 2008 to 2010, one of them with a fatal 

outcome. All patients developed fever within four days 

after returning to Norway, and the serological analyses 

demonstrated that they suffered from primary dengue 

infections. Anti-dengue IgM antibodies were detected 

in acute samples from the fatal case, in addition to 

the dengue virus serotype 2-positive case from 2009 

and two cases from 2010. During primary infection, 

IgM and IgG antibodies are usually detectable from 

respectively five and 14 days after onset of symptoms 

Figure 

Phylogenetic tree comparing published dengue viruses sequences with those from viruses isolated in Norway, 2008-2010 

(n=8) 

The tree is based on an approximately 300 nt fragment of the E glycoprotein gene. Sequence identification of selected dengue virus 

sequences is as follows: country of origin, year of isolation and NCBI GenBank accession number. The sequences determined in our study are 

identified by case number and country of origin. Phylogenetic analysis was conducted using MEGA 4 [12], and the tree was constructed using 

neighbour-joining method. 


[3,9]. The fatal case highlights that fatal or severe dengue 

fever can also be caused by a primary infection. 

Severe or fatal dengue fever cases are more frequent 

in secondary than primary infections, but fatal primary 

dengue virus infection has been described in earlier 

reports [3,5,9]. 

For the reported cases, the rapid NS1 antigen tests 

were helpful for the initial diagnosis of dengue fever in 

the early phase of the disease. Our results show that 

dengue virus can be detected by NS1 antigen tests in 

patients who are negative for anti-dengue IgG and IgM. 

This report shows the importance of performing dengue 

virus diagnostics in febrile patients returning from 

endemic areas even if other pathogens have been 

detected. Awareness of the different causes of travelrelated 

infections and early inclusion of these in the 

differential diagnosis is particularly important in the 

context of destinations with a risk of such transmission. 

Other studies have shown that most dengue virus 

infections diagnosed in European countries have 

been imported from Asia or the Americas, and in 

these regions all four dengue virus types have been 

shown to circulate [9,10]. Six of our study cases had 

returned from south-east Asia with dengue virus infection 

caused by virus serotype 1, 2 or 3. Import of dengue 

virus serotype 4 into Norway has so far not been 

reported. 

Two of the cases in this study were imported from 

Eritrea, where only one dengue virus serotype 3 isolate 

has been reported earlier [10]. This country is 

not a specifically popular destination for Norwegian 

travellers and we are not aware of a concurrent outbreak 

in Eritrea. To date, there have been few reports 

of viraemic dengue fever cases imported into Europe 

from Africa. This may be due to underreporting in some 

African countries, as well as lack of adequate diagnostic 

tools [11]. Dengue surveillance is poorly implemented 

in Africa and surveillance of febrile travellers 

returning to Europe will add new knowledge on dengue 

virus distribution throughout Africa. 

A comparison of sequences obtained in this study and 

from studies published elsewhere, are shown in the 

phylogenetic tree (Figure). In general, the sequence 

similarity between isolates of one dengue serovirus 

type was greater than 95%. The dengue virus serotype 

1 isolates from Eritrea were closely related to dengue 

virus serotype 1 isolated in Kenya in 2004-5. Similarly, 

the dengue virus serotypes 1, 2 and 3 imported from 

south-east Asia in our study clustered together with 

the respective serotypes reported from this area earlier. 

To our knowledge, only few reports of dengue 

virus serotype 1 isolates from East Africa have been 

published [10], and this study provides evidence that 

this serotype 1 is circulating in this area. 


Our report confirms that returning travellers may serve 

as sentinels for local outbreaks of dengue fever in 

endemic areas. The worldwide surveillance of dengue 

virus requires simple and accurate methods for the 

identification of virus types and is especially important 

since air travellers move quickly between endemic and 

non-endemic regions, allowing introductions of dengue 

virus to new areas that already are populated with 

Aedes mosquitoes. 

References 

1. Kyle JL, Harris E. Global spread and persistence of dengue. 

Annu Rev Microbiol. 2008;62:71-92. 

2. World Health Organization. Dengue and dengue haemorrhagic 

fever. Fact sheet N°117. Geneva: World Health Organization; 

May 2008. Available from: http://www.who.int/mediacentre/ 

factsheets/fs117/en/ 

3. Halstead SB. Dengue. Lancet. 2007;370(9599):1644-52. 

4. Jelinek T. Trends in the epidemiology of dengue fever and 

their relevance for importation to Europe. Euro Surveill. 



5. Jensenius M. Berild D. Ormaasen V. Maehlen J. Lindegren G. 

Falk KI. Fatal subarachnoidal haemorrhage in a Norwegian 

traveller with dengue virus infection. Scand J Infect Dis. 

2007;39(3):272-4. 

6. Blystad H, Borgen K. Dødelig denguevirus-infeksjon etter 

turistopphold i Thailand. [Fatal dengue virus infection after 

tourist visit to Thailand]. MSIS-rapport 2008;9. [Accessed 29 

August 2010]. Oslo: Nasjonalt folkehelseinstitutt; 30 April 

2008. Available from: http://www.fhi.no/dav/6c111c7181.pdf 

7. Lindegren G, Vene S, Lundkvist A, Falk KI. Optimized Diagnosis 

of Acute Dengue Fever in Swedish Travelers by a Combination 

of Reverse Transcription-PCR and Immunoglobulin M Detection. 

J Clin Microbiol. 43(6):2850-5, 2005 

8. Domingo C, Palacios G, Niedrig M, Cabrerizo M, Jabado O, 

Reyes N, et al. A New Tool for the Diagnosis and Molecular 

Surveillance of Dengue Infections in Clinical Samples. Dengue 

Bulletin. 2004;28. New Delhi: WHO Regional Office for South- 

East Asia; 2010. Available from: http://www.searo.who.int/en/ 

Section10/Section332/Section1985_9804.htm 

9. Gubler DJ, Kuno G, Markoff L. Flaviviruses. In Fields Virology, 

Knipe DM, Howley PM. 5th ed. Philadelphia: Lippincott 

Williams & Wilkins; 2007. p.1155-252. 

10. Franco L, Di Caro A, Carletti F, Vapalahti O, Renaudat C, 

Zeller H, et al. Recent expansion of dengue virus serotype 

3 in West Africa. Euro Surveill. 2010;15(7):pii=19490. 



11. Sang RC. Dengue in Africa. Nairobi: Arbovirology/Viral 

Haemorrhagic Fever Laboratory, Centre for Virus Research, 

Kenya Medical Research Institute; 2007. [Accessed 29 August 

2010]. Available from: http://www.tropika.net/review/061001- 

Dengue_in_Africa/article.pdf 

12. Tamura K, Dudley J, Nei M, Kumar S. MEGA4: Molecular 

Evolutionary Genetics Analysis (MEGA) software version 4.0. 

Mol Biol Evol. 2007;24(8):1596-9. 

83


First two autochthonous dengue virus infections in 

metropolitan France, September 2010 

G La Ruche (g.laruche@invs.sante.fr) 1 , Y Souarès 1 , A Armengaud 2 , F Peloux-Petiot 3 , P Delaunay 4 , P Desprès 5 , A Lenglet 6 , 

F Jourdain 7 , I Leparc-Goffart 8 , F Charlet 3 , L Ollier 4 , K Mantey 6 , T Mollet 6 , J P Fournier 4 , R Torrents 2 , K Leitmeyer 6 , P Hilairet 4 , 

H Zeller 6 , W Van Bortel 6 , D Dejour-Salamanca 1 , M Grandadam 5 , M Gastellu-Etchegorry 1 

1. French Institute for Public Health Surveillance (Institut de Veille Sanitaire, InVS), Saint-Maurice, France 

2. Regional office of the French Institute for Public Health Surveillance (Cire Sud), Marseille, France 

3. Regional Health Agency of Provence-Alpes-Côte d’Azur, Marseille and Nice, France 

4. Entomology-Parasitology, Virology and Emergency Medicine and Internal Medicine Departments, University Hospital of Nice, 

Nice, France 

5. Institut Pasteur, National Reference Centre for arboviruses, Paris, France 

6. European Centre for Disease Prevention and Control (ECDC), Stockholm, Sweden 

7. Directorate General for Health, Ministry of Health, Paris, France 

8. Institut de recherche biomédicale des armées, National Reference Centre for arboviruses associated laboratory, Marseille, 

France 


La Ruche G, Souarès Y, Armengaud A, Peloux-Petiot F, Delaunay P, Desprès P, Lenglet A, Jourdain F, Leparc-Goffart I, Charlet F, Ollier L, Mantey K, Mollet T, Fournier 

JP, Torrents R, Leitmeyer K, Hilairet P, Zeller H, Van Bortel W, Dejour-Salamanca D, Grandadam M, Gastellu-Etchegorry M. First two autochthonous dengue virus 

infections in metropolitan France, September 2010. 


In September 2010, two cases of autochthonous dengue 

fever were diagnosed in metropolitan France for 

the first time. The cases occurring in Nice, southeast 

France, where Aedes albopictus is established, 

are evidence of dengue virus circulation in this area. 

This local transmission of dengue calls for further 

enhanced surveillance, active case finding and vector 

control measures to reduce the spread of the virus and 

the risk of an epidemic. 

Dengue fever is the most important mosquito-borne 

viral disease in the world and is endemic in Africa, 

Asia, Caribbean and Latin America. According to the 

World Health Organization, there are annually more 

than 50 million cases and 22,000 deaths [1]. Dengue 

fever is caused by viruses of the Flaviviridae family and 

transmitted by mosquito vectors of the Aedes genus, 

mainly Ae. aegypti and Ae. albopictus [2]. 

In Europe, the last dengue epidemic was reported 

from 1927 to 1928 in Greece with high mortality and 

Ae. aegypti was implicated as the vector [3]. Since 

the 1970s, mainly through global trade of car tyres, 

Ae. albopictus has become increasingly established 

in European Union Member States, including France, 

Greece, Italy, the Netherlands (though only in greenhouses), 

Slovenia and Spain [4]. This mosquito species 

is also established in neighbouring countries such as 

Albania, Bosnia and Herzegovina, Croatia, Monaco, 

Montenegro, San Marino, Switzerland and Vatican 

City [2,5]. Imported cases of dengue fever in travellers 

returning from countries where dengue is endemic or 

where dengue epidemics are taking place have been 

frequently reported in European countries in recent 

years [6-10]. 

Article published on 30 September 2010 

In metropolitan France, sporadic Ae. albopictus mosquitoes 

were first detected in Normandy in 1999 [11], 

but the mosquito is known to have been established 

since 2004 in south-east France [12]. Since 2006, and 

the widespread epidemic of chikungunya in Réunion 

which had posed an increased risk of importation of 

cases, enhanced surveillance is implemented each 

year from May to November in the departments where 

Ae. albopictus is established, as part of the national 

plan against the spread of chikungunya and dengue 

viruses in metropolitan France [13]. Enhanced surveillance, 

compared with routine surveillance, allows 

the reporting and confirmation of suspected cases to 

be accelerated. The laboratory network surveillance 

system, the most sensitive routine system in France, 

detected around 350–400 imported dengue cases per 

year between 2006 and 2009 in metropolitan France 

[14,15]. During the same four-year period, enhanced 

surveillance reported a total of 33 imported dengue 

cases (including 11 cases in 2009). Between 1 May and 

17 September 2010 (i.e. the first 4.5 months of surveillance), 

120 imported cases of dengue have been 

reported by the enhanced surveillance system [16], 

which represents an 11-fold increase when compared 

with the entire 2009 season. This increase in imported 

cases is mostly related to the ongoing epidemics in the 

French West Indies, Martinique and Guadeloupe, since 

the beginning of 2010. Here we report on the two first 

cases of autochthonous dengue virus infection ever 

diagnosed in metropolitan France and the public health 

measures subsequently implemented. 

Case 1 

The first case was detected through the routine 

enhanced surveillance system. The patient was a man 


in his 60s, resident in Nice, Alpes-Maritimes department, 

who developed fever, myalgia and asthenia on 

23 August 2010. He was hospitalised on 27 August 

2010, but his clinical condition remained stable. A temporary 

thrombocytopenia with a minimal platelet count 

of 48,000/µl (norm: 150,000–400,000) on day five 

of the illness resolved without complications and he 

recovered within a few days after disease onset. 

Laboratory findings 

A panel of sera obtained during the acute and recovery 

phases on days five, seven, 11 and 25 of the illness was 

investigated by serological tests (in-house MAC-ELISA 

and direct IgG ELISA) and real-time RT-PCR. Moreover, 

a serum sample collected during a previous medical 

examination in May 2010 was tested retrospectively. 

Presence of IgM and IgG against dengue virus antigens 

was documented in all samples except for the serum 

sampled in May 2010. Antibody titration revealed sharp 

increases in IgM titres from 1:800 to 1:12,800 and 

in IgG titres from 1:32,000 to >1:128,000 over the 25 

days follow-up. Anti-dengue virus IgA (Assure Dengue 

IgA rapid test, MP Biomedicals) were also detected on 

days five and seven. The dengue NS1 antigenic test 

(Dengue NS1 strip, Biorad) was positive on days five 

and seven but negative on day 11, demonstrating the 

active replication of a dengue virus during the symptomatic 

period. RT-PCR for dengue virus was positive 

on day five and negative thereafter. Molecular typing 

identified a dengue virus serotype 1. 

It is of interest to note that high levels of specific antidengue 

IgG were detected during the acute phase of 

disease. Our hypothesis is that these IgG might result 

from activation of memory B cells (original antigenic 

sin) related to an ancient primary infection with a heterologous 

serotype of dengue virus. Seroneutralisation 

tests will be informative on the immunological status 

of the patient regarding a possible previous infection 

with a dengue virus of another serotype. Virus isolation 

and sequencing are also ongoing. No serum crossreactions 

were observed with tick-borne encephalitis 

and West Nile viruses and no markers of chikungunya 

virus infection were found (absence of IgM and IgG 

antibodies, negative RT-PCR). The patient had been 

vaccinated against yellow fever 28 years ago. 

Friends from the French West Indies had stayed with 

him since April 2010. He had no recent history of international 

travel or blood transfusion. Consequently, the 

patient was considered a confirmed autochthonous 

case of dengue virus infection. 

Control measures 

This classification prompted an immediate reaction of 

public health authorities to reduce the risk of further 

spread of the virus. Various measures were undertaken 

by health authorities as laid out in the national plan 

against the spread of dengue in France (level 2 of the 

plan) [13]: (i) 200 metres perifocal vector control activities 

centred on the case’s residence, including spraying 


for adult mosquitoes and destruction of breeding 

sites; (ii) active case finding in the neighbourhood of 

the case’s residence and in other areas visited by the 

case; (iii) providing information about dengue virus to 

health professionals, including incitation for screening 

suspected dengue cases and information to the public. 

The active case finding conducted by physicians and 

laboratories will be continued on a weekly basis up to 

45 days after the onset of symptoms of the last autochthonous 

case. 

The routine laboratory network surveillance system 

noticed that six recently imported confirmed dengue 

cases, including four with a RT-PCR positive for 

dengue, had been detected in Nice between 24 July 

and 23 August 2010. One of them had returned from 

Martinique and lives about 200 metres from the autochthonous 

case. This imported case was reported too 

late to implement vector control measures which routinely 

follow imported viraemic dengue cases in those 

departments where the vector is present, and could 

therefore be a potential source of infection of local 

Ae. albopictus. As of 24 September 2010, the active 

case finding has detected nine new suspected autochthonous 

cases of dengue fever in the neighbourhood 

of the index case. In four of them, no markers of dengue 

virus infection were found (absence of IgM and IgG 

antibodies, negative RT-PCR), results from epidemiological 

and laboratory investigations for further four 

patients are still pending. One case was confirmed to 

be infected by dengue virus; the latter patient is the 

second autochthonous dengue fever case ever diagnosed 

in metropolitan France. 

Case 2 

This second case is an 18 year-old man who had no 

recent history of international travel. He lives approximately 

70 metres from the first autochthonous case. He 

developed fever, myalgia, headache and asthenia on 11 

September 2010. He was hospitalised briefly because 

of fever of unknown origin and thrombocytopenia with 

a mild clinical disease. The thrombocytopenia (platelet 

count 53,000/µL on day seven of the illness) was temporary 

and moderate, and he has recovered fully. 

Laboratory investigations 

Laboratory tests conducted on an early serum sample 

on day three of illness indicate negative serology for 

IgG and IgM antibodies but strongly positive RT-PCR 

for dengue virus. Molecular typing identified a dengue 

virus serotype 1. The strain appears to be quite similar 

to those which currently circulate in Martinique; more 

detailed analyses are ongoing. 

Discussion 

The identification of two autochthonous cases of dengue 

fever which are clustered in space and time is 

strongly suggestive that a local transmission of dengue 

virus is ongoing. Therefore level 3 of the national 

plan against the spread of dengue virus has been activated 

[13]. It entails additional measures to those taken 

85

at level 2: (i) active case finding of autochthonous 

cases in hospital emergency wards, at present in Nice 

and surrounding towns, (ii) implementation of vector 

control measures in hospitals, together with protection 

of potential viraemic patients against mosquito 

bites using electric light traps, electric diffusers for 

insecticides, and repellents, and vector control measures 

around the port and the international airport of 

Nice including enhanced entomological surveillance, 

and (iii) toxicovigilance related to the wide use of 

insecticides. 

Based on the currently available information, these 

are the first confirmed cases of autochthonous transmission 

of dengue fever in metropolitan France and 

Europe, since the epidemic in Greece in the late 1920s 

and apart from one nosocomial case of dengue infection 

reported from Germany in 2004 [17]. The event is 

not entirely unexpected, as reflected in a specific preparedness 

plan and taking into account the increase in 

imported cases from the French West Indies and other 

endemo-epidemic areas. It is known that France, as 

well as other countries in Europe, has competent vectors 

for transmitting this flavivirus. The chikungunya 

outbreak in Italy that occurred in 2007, with over 300 

cases reported, has shown that non-endemic arboviruses 

can be efficiently transmitted in continental 

Europe [18]. 

Whether the transmission of dengue virus in France followed 

a bite from an infectious mosquito imported to 

the area via airplanes or boats, or one already present 

in the area after biting a viraemic person residing or 

visiting Nice, remains to be determined. However, with 

the second confirmed case, the latter scenario is the 

most likely one. Therefore, taking into consideration 

the longest possible incubation period for dengue 

fever, 15 days, it can be considered that the conditions 

for successful transmission of dengue virus to humans 

existed in Nice during August 2010. To date, only 

two autochthonous cases of dengue fever have been 

detected in Nice, but the identification of new dengue 

cases in the near future cannot be excluded. The 

enhanced surveillance and strict vector control measures 

are expected to limit the risk for further spread as 

much as possible. 

At this stage, the risk for further spread to humans in 

Europe, as well as the possibility of the establishment 

of dengue virus transmission in Nice or in neighbouring 

areas in France, may appear limited but needs to 

be closely monitored. Recent evidence demonstrates 

that compared with Ae. aegypti, which has been implicated 

in the majority of large dengue outbreaks worldwide, 

Ae. albopictus is a less efficient vector of this 

virus [2]. Nevertheless, it was involved in outbreaks 

in Japan from 1942 to 1945 [19], the Seychelles in 1977 

[20], Hawaii from 2001 to 2002 [21] and Réunion island 

in 2004 [22]. Vertical transmission of dengue virus 

from mosquitoes to their offspring does not seem very 

efficient, and therefore overwintering of the virus in 

continental European Ae. albopictus populations is 

unlikely [2] but cannot be excluded [23,24]. The public 

health consequences of the presence of Ae. albopictus, 

in this context, appear to be more important for the 

transmission of chikungunya for example, for which 

experimentally better competence has been demonstrated, 

although the competence of local Ae. albopictus 

for dengue virus is far from being negligible [25]. It 

should also be noted, that the currently affected area 

of France as well as other countries in Europe is faced 

with a high number of imported dengue cases every 

year. However, despite this and established mosquito 

populations being potentially able to transmit arboviral 

diseases, local transmission of the dengue virus with 

Ae. albopictus as the vector in mainland Europe has 

never been observed before this reported emergence 

in the south-east of metropolitan France. The high 

vector density in Nice and the increase in the number 

of imported cases in this area in 2010, mainly due to 

intense epidemics in the French West Indies, are two 

major factors to explain this emergence and highlight 

the need to maintain an appropriate active surveillance. 

In terms of blood safety, reported dengue infection 

following blood transfusion in dengue endemic areas 

is rare [26-28] but is also difficult to detect as a large 

proportion of the population would already have antibodies 

against the virus. However, as dengue infection 

is mild or asymptomatic in 40–80% of infected persons, 

depending on the area and the epidemiological 

context [29-31], it does pose a risk to blood safety. The 

two identified cases in Nice are suggestive that other 

infected persons may have lived in the city during the 

same period of exposure, without showing any symptoms. 

Asymptomatic carriers of dengue virus could 

pose a potential risk to blood safety if they donate 

blood while being viraemic. It is possible however, 

that the duration of viraemia in mild or asymptomatic 

cases is shorter and the virus titre is lower than in 

symptomatic persons, but this hypothesis is far from 

proven. Moreover, the limited extend of current virus 

dissemination, as shown by the actual clustering of 

confirmed autochthonous cases, does not indicate 

that such asymptomatic infections could have been 

spread around the whole city of Nice. At present, it is 

difficult to quantify this risk, and only a retrospective 

survey of blood supplies from Nice between July and 

September 2010 would allow to estimate it better. In 

France, authorities in charge of blood routinely exclude 

all febrile donors from donation. No additional exclusion 

measures have been implemented after the two 

neighbouring cases as the risk for dengue transmission 

has been considered very low. 

Further investigations to identify the likely source of 

exposure of the two cases, as well as extensive comparison 

of the dengue virus genotypes between the locally 

identified viruses and the strains currently circulating 

in the French West Indies, will hopefully allow a better 

understanding of this event. The reactive surveillance 

in addition to the routine enhanced surveillance 


is likely to identify new symptomatic cases in the area, 

determining also the potential geographic extension 

of the risk. Finally, better understanding is needed on 

how the vector abundance, activity and competence of 

Ae. albopictus for dengue transmission influence the 

risk for further transmission in the region [25,32]. 

Conclusion 

The current clustering of cases of locally transmitted 

dengue fever in Nice is a significant public health 

event, but is not unexpected and more cases can be 

predicted. Such transmission was anticipated by the 

development of a national plan. Although this plan 

should be adjusted in the light of this experience, this 

event shows the advantage of such preparedness in 

order to implement rapid and proportionate measures 

of surveillance and control. Previous events, including 

a mosquito-borne arbovirus outbreak in Italy, the 

occurrence of vector-borne diseases around airports 

and other ports of entry and a previous risk assessment 

on dengue virus introduction in European Union 

countries [4] indicate that autochthonous transmission 

in continental Europe is possible, as confirmed by 

the present event. However, according to the available 

epidemiological information, the risk for establishment 

of dengue transmission in south-eastern France 

or further spread in Europe currently appears limited. 

Further data available in the near future will allow us to 

re-assess this likelihood. 


We thank the biomedical laboratories involved in the enhanced 

surveillance: Cerba (Saint-Ouen l’Aumone), Biomnis 

(Lyon), La Timone (Marseille) and Avicenne hospital 

(Bobigny), part of the national laboratory surveillance system, 

and the private laboratories in south-east France. 

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Dengue virus infection in a traveller returning from 

Croatia to Germany 

J Schmidt-Chanasit (jonassi@gmx.de) 1 , M Haditsch 2 , I Schöneberg 3 , S Günther 1 , K Stark 3 , C Frank 3 

1. Bernhard Nocht Institute for Tropical Medicine, Department of Virology, Hamburg, Germany 

2. Labor Hannover MVZ GmbH, Hannover, Germany 

3. Robert Koch Institute, Department for Infectious Disease Epidemiology, Berlin, Germany 


Schmidt-Chanasit J, Haditsch M, Schöneberg I, Günther S, Stark K, Frank C. Dengue virus infection in a traveller returning from Croatia to Germany. 


Dengue virus (DENV) is endemic in south-east Asia 

and Central to South America. In August 2010, a DENV 

infection was diagnosed in a German traveller returning 

from a trip to Croatia in south-east Europe. The 

patient presented with fever and other typical symptoms 

of DENV-infection. Virological investigation 

revealed the presence of DENV-specific IgM, a rise in 

DENV-specific IgG and the presence of DENV NS1 antigen 

in the patient’s blood. 

Dengue virus (DENV) is an arthropod-borne RNA virus 

of the Flaviviridae family causing dengue fever in 

humans. Since 2001 dengue fever has been mandatorily 

reported to the German public health authorities, 

in accordance with the Federal Protection against 

Infection Act [1]. According to German notification data, 

between 60 and 387 imported DENV infections are 

reported annually (Table). 

The DENV infections in imported cases are mainly 

acquired in south-east Asia as well as South and 

Central America. Very recently, autochthonous DENV 

Table 

Imported cases of dengue fever per year, Germany, 

2001–2010 a 

Year Number of recorded cases 

2001 60 

2002 213 

2003 131 

2004 121 

2005 144 

2006 175 

2007 264 

2008 273 

2009 298 

2010 387 

Source: Robert Koch Institute, SurvStat 

(http://www3.rki.de/SurvStat). 

a As of 4 October 2010. 


Article published on 7 October 2010 

infections were reported in southern France, diagnosed 

for the first time ever in Europe [2]. Here we report on a 

case of DENV infection that was apparently acquired in 

Croatia and imported to Germany by a traveller. 

Case report 

A 72-year-old man from Germany visited Croatia in 

August 2010: he left on 1 August and returned on 15 

August. He was accompanied by seven family members, 

including grandchildren. The family travelled by 

car from Germany via Austria and Slovenia to Croatia 

without overnight stops. The group stayed the entire 

time around Podobuce close to Orebić on the Peljesac 

peninsula and on the isle of Korčula in the south of 

Croatia. Podobuce and Korčula are located approximately 

100 km north-west of the city of Dubrovnik, 

which was also visited. Temperatures were reported 

to be very high (approximately 30 °C at night). After 

returning to Germany, on 16 August, the patient developed 

a febrile illness with a temperature of up to 

39 °C, chills, arthralgia, headache, and retro-orbital 

pain. Following a short period of improvement, his 

temperature rose again to 39 °C on 21 August, and he 

continued to have arthralgia, myalgia, weakness and 

dyspnoea. Among several other diseases, dengue fever 

was suspected by the general practitioner, because of 

the clinical picture. 

Laboratory results 

Serum samples were taken from the patient for virological 

investigation on 23 and 30 August and on 2 

September. The sample from 23 August was positive 

for DENV-specific IgM, but negative for IgG in an 

enzyme-linked immunosorbent assay (ELISA). On 30 

August, DENV-specific IgM and IgG was positive with 

a titre of 1:2,560 (cut-off 1:20) and 1:80 (cut-off 1:20), 

respectively, in an indirect immunofluorescence assay 

based on DENV-infected cells. In addition, the serum 

sample tested positive for DENV NS1 antigen (Dengue 

Early ELISA, Panbio). Real-time reverse transcriptionpolymerase 

chain reaction (RT-PCR) for DENV [3] was 

negative. The detection of DENV NS1 antigen and 

the simultaneous absence of DENV RNA during this 

89

phase of dengue fever are in line with previous studies 

demonstrating an acute DENV infection [4]. The 

sample taken on 2 September showed an increase 

in the DENV-specific IgG titre (1:1,280), while the IgM 

titre remained unchanged and the ELISA for NS1 antigen 

was negative. In this sample, immunofluorescence 

assay titres for related flaviviruses were lower than for 

DENV: West Nile virus (IgM negative, IgG 1:160) and 

tick-borne encephalitis virus (IgM 1:80, IgG 1:80). The 

patient did not report vaccination against tick-born 

encephalitis or yellow fever. A temporary thrombocytopenia 

with a minimal platelet count of 97,000/µl (norm: 

150,000–440,000/µl) on the eighth day of the illness 

resolved without complications and the patient recovered 

within two weeks after disease onset. 

Conclusions 

The clinical suspicion of dengue fever was confirmed 

by the laboratory tests. As the incubation period for 

dengue fever ranges from three to 14 days , the infection 

was probably acquired in southern Croatia and not 

en route. The Croatian authorities were given all available 

information about the case, enabling them to investigate 

this further at local level. To our knowledge, this 

is the second report on an autochthonous DENV transmission 

in Europe after France. Antibodies against 

DENV have been previously detected in Croatian individuals 

in the context of international travel; however, 

the specificity of the assay is questionable [5]. The 

presence of Aedes albopictus as a potential DENV vector 

in Croatia [6] and the importation of confirmed dengue 

fever cases from endemic areas into Croatia [7,8] 

allow autochthonous DENV transmission within this 

country. The mosquito season in parts of the northern 

Mediterranean coast may last from May to November. 

Therefore, dengue fever should be considered in 

patients with fever of unknown origin and relevant 

clinical symptoms who stayed in areas in Europe where 

Ae. albopictus occurs. 


We thank M. Reichmann, B. Noack, S. Kunstmann, P. 

Emmerich, B. Braunsdorf and B.Hiller for preparation of additional 

clinical, laboratory and epidemiological data related 

to this case. 

References 

1. Dreesman J, Benzler J. [Infectious disease surveillance based 

on the Protection against Infection Act in the German public 

health sector]. Bundesgesundheitsblatt Gesundheitsforschung 

Gesundheitsschutz. 2005;48(9):979-89. German 

2. La Ruche G, Souarès Y, Armengaud A, Peloux-Petiot F, 

Delaunay P, Desprès P, et al. First two autochthonous dengue 

virus infections in metropolitan France, September 2010. Euro 



3. Drosten C, Göttig S, Schilling S, Asper M, Panning M, Schmitz 

H, Günther S. Rapid detection and quantification of RNA of 

Ebola and Marburg viruses, Lassa virus, Crimean-Congo 

hemorrhagic fever virus, Rift Valley fever virus, dengue virus, 

and yellow fever virus by real-time reverse transcription-PCR. J 

Clin Microbiol. 2002;40(7):2323-30. 

4. Schilling S, Emmerich P, Günther S, Schmidt-Chanasit J. 

Dengue and Chikungunya virus co-infection in a German 

traveller. J Clin Virol. 2009;45(2):163-4. 

5. Ropac D, Gould E, Punda V, Vesenjak-Hirjan J. [Dengue viruses 

in northeastern Croatia]. Lijec Vjesn. 1988;110(6-7):177-80. 

Croatian. 

6. Klobucar A, Merdic E, Benic N, Baklaic Z, Krcmar S. First record 

of Aedes albopictus in Croatia. J Am Mosq Control Assoc. 

2006;22(1):147-8. 

7. Markotic A, Betica Radic L, Maretic T. [Viral tourism: dengue 

virus]. Croatian Journal of Infection. 2007; 27(4):181-4. 

Croatian. 

8. Pinazo MJ, Muñoz J, Betica L, Maretic T, Zekan S, Avsic-Zupanc 

T, et al. Imported dengue hemorrhagic fever, Europe. Emerg 

Infect Dis. 2008;14(8):1329-30. 



Relapsing vivax malaria cluster in Eritrean refugees, 

Israel, June 2010 

E Kopel (eran.kopel@mail.huji.ac.il) 1 , E Schwartz 2 , Z Amitai 1 , I Volovik 1 

1. Tel Aviv District Health Office, Ministry of Health, Tel Aviv, Israel 

2. Centre for Geographic Medicine and Tropical Diseases, Sheba Medical Centre, Tel Hashomer, Ramat Gan, Israel 


Kopel E, Schwartz E, Amitai Z, Volovik I. Relapsing vivax malaria cluster in Eritrean refugees, Israel, June 2010. Euro Surveill. 2010;15(26):pii=19601. Available 

online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19601 

We report on a cluster of relapsing vivax malaria 

among Eritrean refugees residing in Israel. Since the 

beginning of 2010, 15 cases have been identified. Five 

of the six patients who had complete medical and epidemiological 

histories, reported Sudan as the place 

of primary infection during their journey to Israel, and 

having had the first relapse in Israel, six months later 

(median). Suggested place of exposure is the region of 

the Eritrean refugee camps in Sudan. 

Introduction 

Malaria, once endemic in Israel, was eradicated almost 

50 years ago, although its vectors, several malariatransmitting 

species of Anopheles mosquitoes, still 

exist in various parts of the country [1,2]. Every year 

between 60 and 100 imported cases of malaria are 

reported to the Ministry of Health. Most of these cases 

are travellers returning from endemic countries to 

Israel and only few of them are immigrants from Sub- 

Saharan Africa [3]. 

Currently, there is a cluster of relapsing Plasmodium 

vivax malaria among Eritrean refugees in Israel. The 

epidemiological investigation, which is summarised 

here, was conducted by the local health office in Tel 

Aviv and is therefore limited to the Tel Aviv district. 

Methods 

On 7 June 2010, a cluster of five malaria cases was 

reported to the Tel Aviv District Health Office. These 

cases were Eritrean refugees under treatment in the 

same hospital during the first week of June. An epidemiological 

investigation was initiated. The case 

definition was laboratory-confirmed malaria excluding 

returning travellers. 

The following investigation measures were taken. 

• Species were identified by thick and thin smears 

with confirmation by real-time reverse-transcriptase 

PCR [4]. 

• Medical records of cases were obtained from 

hospitals. 

• Oral interviews were conducted with four cases by 

a native speaker of Tigrinya (one of the two working 


Article published on 1 July 2010 

languages in Eritrea). The interviews were based 

on a short epidemiological questionnaire that contained 

questions on demographics, the route of 

the journey to Israel, current and past malaria illness 

history and possible exposures (time, place, 

mosquito bites) in Israel or abroad. 

• The local health office provided records on past 

epidemiological data regarding non-traveller cases 

of malaria in the district since 2006. 

• An alert was issued to the hospitals and laboratories 

in the district, which were requested to report 

all cases of diagnosed malaria and to confirm them 

at the national reference laboratory. 

• With the assistance of an expert malaria advisor, 

concise clinical guidelines for proper management 

[5,6] of cases were promptly issued to local infectious 

diseases units at all hospitals in the district. 

Results 

Five cases of non-traveller malaria were reported in the 

district from 2006 to 2009 (Figure). All of these cases 

had been imported, three of whom were refugees from 

Eritrea. Since the beginning of 2010, 15 cases, all of 

them Eritrean refugees, have been reported. Most of 

them (nine of 15) were diagnosed in the first two weeks 

of June 2010. No other cases of imported malaria in 

other migrants were documented during this period of 

2010 in the district. 

Twelve cases were male (80%) and the median age was 

25 years (interquartile range (IQR): 21.5–29 years). All 

the cases had laboratory-confirmed P. vivax infection. 

Low parasitaemia, ranging from

(median: 2.8 months, IQR: 0.7–4.4 months). Ten of the 

15 cases had information on previous malaria attacks 

and reported having had one during the journey to 

Israel. For six of these 10 it was known that this previous 

attack occurred in Sudan, within the two-month 

period before arriving in Israel, of whom five recalled 

it was the primary infection. A further patient of these 

10 reported a one-month period between the previous 

attack on the journey and arrival to Israel and another 

patient had the previous attack in Ethiopia, 12 months 

before arrival to Israel. An additional of the 15 patients, 

a 2.5-year-old female baby, had the first attack in Israel, 

approximately one month after her arrival through 

Sudan. No data on any previous attacks and their location 

were available for the last four of the 15 patients. 

The median time interval between attacks was 6.1 

months (IQR: 4.0-8.1 months, n=9). Five cases had 

received partial, inadequate or no treatment at the 

time of their previous attack, whereas no complete 

past medical history was available for the rest of the 

15 patients. 

The probable place of exposure of four patients who 

have been interviewed to date, was a complex with 

three refugee camps in Shagrab, operated by the United 

Nations’ Refugee Agency, in southeastern Sudan, near 

the Eritrean border. In addition, the reported duration 

of stay in these camps was between one and two 

months before continuing directly to the Israeli border, 

by organised smuggling [7], apparently without significant 

stations of stay in neighbouring Egypt. 

Discussion 

We describe a cluster of relapsing vivax malaria among 

Eritrean refugees who had recently arrived in Israel. 

The fact that this cluster is mostly limited to young 

men probably reflects the overall current composition 

of Eritrean refugee population arriving in Israel. 

During the last several years until the end of 2009, 

9,517 Eritrean refugees, who constituted 48.6% of 

all asylum seekers, entered Israel. Since 2009, the 

Figure 

Laboratory-confirmed malaria cases (excluding returning 

travellers), Tel Aviv, Israel, 1 January 2006-15 June 2010 

(n=18) 


14 

12 

10 

8 

6 

4 

2 

0 

2006 2007 2008 2009 2010 a 

Year 

a Number of cases until 15 June 2010. Two additional cases 

registered outside the district of Tel Aviv are not shown in the 

figure. 

Eritrean refugees have gradually become the predominant 

group of asylum seekers who enter Israel from the 

Sinai desert (Egypt). In the first four months of 2010, 

3,793 Eritrean refugees entered the country and constitute 

82% of all asylum seekers who entered in this 

period [8]. 

These figures represent a 40% increase in the Eritrean 

refugee population in Israel during the first four 

months of 2010 alone. In addition, a more than fourfold 

increase in the crude incidence rate of non-traveller 

malaria in Israel was observed in the same period: 

in 2009, the crude incidence rate was 0.77 cases per 

1,000 migrants, while in the first half of 2010, it was 

3.58 cases per 1,000 migrants*. These estimated 

calculations are based on data from the National 

Department of Epidemiology in the Israeli Ministry 

of Health and from an analysis report of the Knesset 

Research and Information Centre [8]. Consequently, an 

overall increase in malaria activity along the refugee 

route in Africa must have played a significant role in 

this cluster. 

Therefore, it is reasonable to assume that a place on 

the journey is a recent common place of exposure. This 

place was most probably in Sudan rather than Eritrea 

as the lowland of Eritrea is mostly affected by P. falciparum 

and not by P. vivax [9]. Furthermore, a dramatic 

decline in the incidence rate of malaria was observed 

in the recent years in Eritrea, due to successful eradication 

programmes [10]. In fact, almost all cases who 

had complete medical and epidemiological history, 

namely a third of the total number of cases, specifically 

recalled having had the first malaria attack during 

their stay in Sudan, which was usually a period of two 

months before arriving to Israel. 

Moreover, this cluster may actually reflect the expanding 

prevalence of a previously reported [11] small focus 

of vivax malaria in Sudan: 10 of 83 blood samples were 

PCR-positive for P. vivax (on the background of predominant 

P. falciparum in this area) in some of the Eritrean 

refugee camps in eastern Sudan, a frequently flooded 

low plain region in which malaria may have remained 

the leading cause of morbidity and mortality. 

Conclusions 

The evident ongoing rise of human reservoirs of 

malaria in the region may potentiate the risk for the 

re-emergence of locally acquired mosquito-transmitted 

malaria in Israel and neighbouring countries. This warrants 

tight national surveillance for new cases, proper 

clinical management and follow-up of current cases, 

and effective control measures of the local Anopheles 

vectors. In addition, it highlights the need for increased 

malaria surveillance in the refugee camps of eastern 

Sudan. 



We thank Anat Scheffer, Sofia Katser and Sara Germay 

for their assistance in the epidemiological investigation. 

We also thank the epidemiology unit of Tel Aviv Sourasky 

Medical Centre for the initial notification and the National 

Reference Centre for Parasitology for the cooperation. 

*Authors’correction 

The following correction was made on 5 July 2010 on request of 

the authors: In the Discussion section, the second sentence of the 

second paragraph: ‘in 2009, the crude incidence rate was 0.77 per 

1,000 cases, while in the first half of 2010, it was 3.58 per 1,000 

cases.’ was replaced with the following sentence: ‘in 2009, the 

crude incidence rate was 0.77 cases per 1,000 migrants, while in 

the first half of 2010, it was 3.58 cases per 1,000 migrants.’ 

References 

1. Pener H, Kitron U. Distribution of mosquitoes (Diptera: 

Culicidae) in northern Israel: a historical perspective. I. 

Anopheline mosquitoes. J Med Entomol. 1985;22(5):536-3. 

2. Kitron U, Pener H, Costin C, Orshan L, Greenberg Z, Shalom 

U. Geographic information system in malaria surveillance: 

mosquito breeding and imported cases in Israel, 1992. Am J 

Trop Med Hyg. 1994;50(5):550-6. 

3. Anis E, Pener H, Goldmann D, Leventhal A. [The reemergence 

of malaria in Israel?]. Harefuah. 2004;143(11):815-19, 838, 837. 

[Article in Hebrew]. 

4. Shokoples SE, Ndao M, Kowalewska-Grochowska K, Yanow 

SK. Multiplexed real-time PCR assay for discrimination of 

Plasmodium species with improved sensitivity for mixed 

infections. J Clin Microbiol. 2009;47(4):975-80. 

5. Schwartz E, Regev-Yochay G, Kurnik D. Short report: a 

consideration of primaquine dose adjustment for radical 

cure of Plasmodium vivax malaria. Am J Trop Med Hyg. 

2000;62(3):393-5. 

6. Hill DR, Baird JK, Parise ME, Lewis LS, Ryan ET, Magill AJ. 

Primaquine: report from CDC expert meeting on malaria 

chemoprophylaxis I. Am J Trop Med Hyg. 2006;75(3):402-15. 

7. Hartman B. UNHCR: Eritreans by far largest refugee group in 

Israel. Jerusalem Post. 10 March 2010. Available from: http:// 

www.jpost.com/Israel/Article.aspx?id=170593. 

8. The Knesset Research and Information Center. [The 

management of infiltrators from the Egyptian border]. 26 May 

2010. Available from: http://www.knesset.gov.il/mmm/data/ 

pdf/m02524.pdf. [In Hebrew]. 

9. Sintasath DM, Ghebremeskel T, Lynch M, Kleinau E, Bretas G, 

Shililu J, et al. Malaria prevalence and associated risk factors 

in Eritrea. Am J Trop Med Hyg. 2005;72(6):682-7. 

10. Nyarango PM, Gebremeskel T, Mebrahtu G, Mufunda J, 

Abdulmumini U, Ogbamariam A, et al. A steep decline of 

malaria morbidity and mortality trends in Eritrea between 

2000 and 2004: the effect of combination of control methods. 

Malar J. 2006;5:33. 

11. Charlwood JD, Qassim M, Elnsur EI, Donnelly M, Petrarca V, 

Billingsley PF, et al. The impact of indoor residual spraying 

with malathion on malaria in refugee camps in eastern Sudan. 

Acta Trop. 2001;80(1):1-8. 


93


First autochthonous malaria case due to Plasmodium 

vivax since eradication, Spain, October 2010 

P Santa-Olalla Peralta (psantaolalla@msps.es) 1 , M C Vazquez-Torres 1 , E Latorre-Fandós 2 , P Mairal-Claver 3 , P Cortina-Solano 4 , 

A Puy-Azón 4 , B Adiego Sancho 5 , K Leitmeyer 6 , J Lucientes-Curdi 7 , M J Sierra-Moros 1 

1. Coordinating Centre for Health Alerts and Emergencies, Ministry of Health and Social Policy, Madrid, Spain 

2. Haematology Department, Hospital San Jorge, Huesca, Spain 

3. Microbiology Department, Hospital San Jorge, Huesca, Spain 

4. Sub-directorate General of Public Health of Huesca, Regional Health Service of Aragón, Huesca, Spain 

5. Service of Public Health Surveillance, Directorate General of Public Health of Aragón, Regional Health Service of Aragón, 

Zaragoza, Spain 

6. European Centre for Disease Prevention and Control (ECDC), Stockholm, Sweden 

7. Animal Pathology Department, Veterinary School, Universidad de Zaragoza, Zaragoza, Spain 


Santa-Olalla Peralta P, Vazquez-Torres MC, Latorre-Fandós E, Mairal-Claver P, Cortina-Solano P, Puy-Azón A, Adiego Sancho B, Leitmeyer K, Lucientes-Curdi 

J, Sierra-Moros MJ. First autochthonous malaria case due to Plasmodium vivax since eradication, Spain, October 2010. Euro Surveill. 2010;15(41):pii=19684. 

Available online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19684 

In October 2010, one case of autochthonous malaria 

due to Plasmodium vivax was diagnosed in Spain. The 

case occurred in Aragon, north-eastern Spain, where 

the vector Anopheles atroparvus is present. Although 

the source of infection could not be identified, this 

event highlights that sporadic autochthonous transmission 

of vector-borne diseases in continental Europe 

is possible and calls for enhanced surveillance and 

vector control measures. 

Background 

Malaria is a mosquito-borne parasitaemic disease 

caused by parasites of the Plasmodium genus and 

endemic in Africa, Asia, Central and South America. 

According to the World Health Organization (WHO), 

there were 247 million cases of malaria and nearly one 

million deaths worldwide in 2008, mostly among children 

living in Africa [1]. Four species of Plasmodium 

have long been recognised to infect humans in nature: 

Plasmodium falciparum, P. vivax, P. malariae and 

P. ovale. Recently, the simian parasite P. knowlesi has 

been found as a cause of human malaria in some areas 

of south-east Asia [2]. Worldwide, P. falciparum and 

P. vivax are the most common causes of malaria. The 

malaria parasites are transmitted by female Anopheles 

mosquito vectors. Of the approximately 430 Anopheles 

species, only 20 species are important for transmission. 

Infection with malaria parasites may result in a wide 

variety of symptoms, ranging from absent or very mild 

symptoms to severe disease and even death in the 

case of P. falciparum malaria. The main symptoms of 

malaria include episodes of cyclical or irregular fever, 

chills, headache, weakness, vomiting and diarrhoea. 

The incubation period in most cases varies from seven 

to thirty days after the infective mosquito bite. 

Article published on 14 October 2010 

In P. vivax malaria, the incubation period usually 

ranges from 10 to 21 days and sometimes up to a year. 

Unlike P. falciparum malaria, P. vivax malaria is rarely 

fatal. However, for P. vivax, clinical relapses may occur 

weeks to months after the first infection. These new 

episodes arise from dormant forms in the liver, and 

special treatment with primaquine – targeted at these 

liver stages – is mandatory for a complete cure. 

Situation in Europe and Spain 

Within the WHO European Region, six countries 

reported autochthonous malaria infections in 2008 

caused by P. vivax: Azerbaijan, Georgia, Kyrgyzstan, 

Tajikistan (the only country in the Region reporting 

P. falciparum malaria), Turkey and Uzbekistan [3]. 

In the European Union (EU) and European Economic 

Area (EEA) countries, malaria has been eradicated 

since 1975 and nearly all reported malaria cases are 

imported. In 2008, 5,848 malaria cases were reported; 

the vast majority of cases for which the species was 

known were caused by P. falciparum (78%) while less 

than 10% were caused by P. vivax [4]. During the last 10 

years, less than 20 cases of autochthonous transmission 

of malaria have been reported in the EU/EEA [3,5]. 

Despite the presence of potential anopheline vectors in 

some countries, sustained local transmission has not 

been identified in continental EU countries [5]. 

In Spain, the last autochthonous case of malaria was 

reported in 1961 [6] and malaria was officially declared 

eradicated in 1964. According to the Spanish National 

Surveillance Network, an average of 400 imported 

malaria cases are reported each year (with less than 

5% due to P. vivax). The Spanish population is susceptible 

to malaria infection given the absence or disappearance 

of the immunity acquired in the past by 

contact with the parasite. 


The principal potential anopheline vector of malaria in 

Spain is Anopheles atroparvus which is widely distributed 

throughout Spain (Figure) and can transmit Asiatic 

strains of P. vivax but is refractory to African strains of 

P. falciparum [7]. 

Case report 

On 5 October 2010, the Regional Health Authorities 

of Aragon reported to the Coordinating Centre for 

Health Alerts and Emergencies at the Spanish Ministry 

of Health one laboratory-confirmed case of P. vivax 

malaria in a patient in their 40s living in the province of 

Huesca (Region of Aragon). The patient had developed 

fever on 20 September 2010 and was diagnosed on 

25 September with acute tonsillitis and started treatment 

with amoxicillin and ibuprofen. Four days later, 

the patient was hospitalised because of clinical deterioration 

with fever and jaundice. On the same day, 

Plasmodium spp. parasites were detected in the blood 

smear, and antimalarial treatment with chloroquine 

and primaquine was initiated. On 1 October the patient 

was dismissed in good clinical condition. 

Laboratory results 

Detection of macrocytosis on the first blood sample 

taken upon hospital admission led to a Giemsa staining 

where Plasmodium spp. parasites were unexpectedly 

identified. Further tests (Rapid Test Binax, chromatography) 

diagnosed Plasmodium spp. (non-falciparum). 

On 4 October, the National Centre for Microbiology in 

Madrid (National Reference Laboratory) confirmed the 

presence of P. vivax by microscopy and multiplex PCR. 

Genomic analysis of the parasite is still ongoing. 

Epidemiological investigation 

According to the epidemiological investigation, the 

patient did not have any travel history to an endemic/ 

epidemic area ever, or contact to persons visiting or 

Figure 

Distribution of Anopheles atroparvus in Spain (dots 

indicate presence) 


residing in such areas. There was no history of surgeries, 

invasive examinations or diagnostics, or blood 

transfusions. The patient was never an injecting drug 

user or had any treatments involving injections. The 

patient reported two visits to airports, Barcelona airport 

in summer 2008 and Zaragoza airport in summer 

2009. 

In the vicinity of the patient’s residence there were 

swine exploitations and was frequently exposed to 

mosquito bites. Furthermore, the patient lives in an 

area of the province of Huesca where An. atroparvus is 

present in several nearby localities. No malaria cases 

have been reported amongst the case’s contacts or 

residents in the locality. There have been no reports of 

imported malaria cases from this area in recent years, 

including 2010. 

Control measures 

The implemented control measures included testing 

household members for malaria, active case finding 

in the neighbourhood of the case and through alerting 

healthcare centres (including hospitals) in the area, 

as well as entomological survey and vector control. 

The entomological survey carried out so far has not 

proven the presence of Plasmodium parasites in local 

mosquitoes. 

Risk assessment for Spain 

Although the investigation was very detailed, we 

have not been able to identify the source of infection. 

Ongoing genetic analysis of the parasite may help to 

specify its possible origin. Transmission may have 

occurred through local Anopheles species after infection 

from people coming from endemic areas carrying 

gametocytes in their blood. Airport malaria caused 

by infected mosquitoes imported from endemic areas 

seems improbable due to the distance to the next international 

airport (approx. 100 km) and the limited flight 

range of local anophelines (4.5 km). 

The possibility of a secondary case originating from 

the reported case is unlikely as the patient has been 

treated, comprehensive control measures have been 

implemented, and the person had never donated blood. 

In Spain, the situation following the eradication of 

malaria in 1964 is defined as ‘anophelism without 

malaria’ with the presence of potential vectors for the 

parasite (mainly An. atroparvus, which is a species 

refractory to P. falciparum) and environmental conditions 

favourable for the breeding, development and 

permanence of the vector [7]. The risk for local transmission 

of malaria will depend on the presence of parasitaemic 

individuals and competent vectors at a given 

time and place. This risk is reduced by early and appropriate 

detection and treatment of cases and vector 

control activities in place. However, it is still possible 

that other sporadic autochthonous cases could still be 

identified. 

95

Conclusions 

Given the described conditions in Spain, an autochthonous 

case of malaria is not unexpected. Previous 

events, including the occurrence of several emerging 

vector-borne disease outbreaks in different countries 

in Europe, indicate that sporadic autochthonous transmission 

of vector-borne diseases in continental Europe 

is possible [9-11]. 

The available epidemiological information does not 

suggest that there is a risk for human health in the 

area. The epidemiological investigation suggested that 

this was a sporadic case with no evidence of further 

local transmission. With the current information, this 

event does not pose a significant risk to EU/EEA citizens. 

Despite the fact that autochthonous cases have 

been reported sporadically in the EU in the past, such 

cases never resulted in established local transmission 

involving more than a few cases. 

Given the presence of competent vectors for malaria 

in the EU, we cannot exclude similar events in the 

future. Continued monitoring of the situation in areas 

where Anopheles mosquito populations are present is 

needed, including increased awareness among clinicians, 

to rapidly identify and report suspected malaria 

cases to respective authorities, and ensure an appropriate 

public health response. 

References 

1. World Health Organisation (WHO). Fact sheet No 94. April 2010. 

Malaria. Geneva: World Health Organization; 2010. Available 

from: http://www.who.int/mediacentre/factsheets/fs094/en/ 

index.html. 

2. Luchavez J, Espino F, Curameng P, Espina R, Bell D, Chiodini 

P, et al. Human Infections with Plasmodium knowlesi, the 

Philippines. Emerg Infect Dis. 2008 May;14(5):811-3. 

3. World Health Organisation (WHO), Regional Office for Europe. 

[Internet]. Centralized information system for infectious 

diseases (CISID) database. Malaria. Copenhagen: World Health 

Organization; 2010. Available from: http://data.euro.who.int/ 

cisid/ 


The European Surveillance System (TESSy). TESSy database. 

Data source 2008. Stockholm: World Health Organisation; 

2010. 

5. Alten B, Kampen CH, Fontenille D, eds. Malaria in Southern 

Europe: resurgence from the past? Emerging Pests and 

Vector-Borne Diseases in Europe, ed. W. Takken and B. Knols. 

2007, Wageningen Academic Publishers, Wageningen, The 

Netherlands. 35-58. 

6. Clavero G. [The eradication of malaria in Spain]. Rev San Hig 

Publ. 1961. 35: p. 265-92. Spanish. 

7. Bueno Marí R, Jiménez Peydró R. [Malaria in Spain: 

entomological aspects and future outlook]. Rev Esp Salud 

Pública. 2008;82(5):467-79. Spanish. 

8. Delacour S, Melero-Alcibar R, Aranda C, Cortés M, Eritja R, 

Escosa R et al. Detailed maps of the geographical distribution 

of the mosquitoes of Spain based on a literature review. Part 

II: Genus Anopheles. The 5th European Mosquito Control 

Association Workshop. Turin Italy (2009). 

9. Schmidt-Chanasit J, Haditsch M, Schöneberg I, Günther S, 

Stark K, Frank C. Dengue virus infection in a traveller returning 

from Croatia to Germany. Euro Surveill. 2010;15(40):pii=19677. 



10. La Ruche G, Souarès Y, Armengaud A, Peloux-Petiot F, 

Delaunay P, Desprès P, et al. First two autochthonous dengue 

virus infections in metropolitan France, September 2010. Euro 



11. Institut de Veille Sanitaire (InVS, French Institute for Public 

Health Surveillance). [Autochtnous cases of chikungunya 

infection in the Var] 27 September 2010. Available from: http:// 

www.invs.sante.fr/regions/sud/pe_paca_corse_011010.pdf 



Infection with Mayaro virus in a French traveller 

returning from the Amazon region, Brazil, 

January, 2010 

M C Receveur 1 , M Grandadam 2 , T Pistone 1 , D Malvy (denis.malvy@chu-bordeaux.fr) 1 

1. Travel clinics and Division of Tropical Medicine and Imported Diseases, Department of Internal Medicine and Tropical 

Diseases, Hôpital St-André, University Hospital Centre, Bordeaux, France 

2. Centre national de référence des arbovirus, Institut Pasteur, Paris, France 


Citation style for this article: Receveur MC, Grandadam M, Pistone T, Malvy D. Infection with Mayaro virus in a French traveller returning from the Amazon region, 

Brazil, January, 2010. Euro Surveill. 2010;15(18):pii=19563. Available online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19563 

Mayaro virus (MAYV) disease is a mosquito-borne 

zoonosis endemic in humid forests of tropical South 

America. MAYV is closely related to other alphaviruses 

that produce a dengue-like illness accompanied 

by long-lasting arthralgia. A French tourist developed 

high-grade fever and severe joint manifestations following 

a 15-day trip in the Amazon basin, Brazil, and 

was diagnosed with MAYV infection in January 2010. 

This case is the first reported in a traveller returning 

from an endemic South American country to Europe. 

Introduction 

Mayaro virus (MAYV) (family Togaviridae, genus 

Alphavirus) is an arthropod-borne zoonotic pathogen 

circulating only in tropical South America [1]. The 

transmission cycle of MAYV in the wild is nearly similar 

to the continuous sylvatic cycle of yellow fever and 

is believed to involve wild primates (monkeys) as the 

reservoir and the tree-canopy-dwelling Haemagogus 

mosquito as the vector. Thus, human infections are 

strongly associated with recent exposure to humid 


This article has been published on 6 May 2010 

tropical forest environments [1,2]. MAYV disease is an 

acute, self-limited dengue-like illness of three to five 

days’ duration. Moreover, MAYV is closely related to 

chikungunya virus and produces a nearly indistinguishable, 

highly debilitating arthralgic disease [1-3]. 

Here we report the case of MAYV disease that recently 

occurred in a French citizen who presented with severe 

rheumatologic disorders after visiting the Brazilian 

Amazon. This report illustrates that with increasing 

travel to remote areas, travellers are at risk of acquiring 

and importing rare diseases that are not indigenous 

to Europe. 

Case report 

The patient, a man in his late 20s, came to the travel 

clinic of the Department of Internal Medicine and 

Tropical Diseases of the University Hospital Centre, 

Bordeaux, France on 4 January 2010 with persistent 

incapacitating arthralgia for a two-month period and 

predominating in his knees and joints of the hands. 

Figure 

Timeline for travel history and symptoms in a French traveller with Mayaro virus disease, October 2009 - January 2010 

W0 W1 

W2 W10 W15 W23 

23/10/09 25/10/09 27/10/09 05/11/09 05/01/10 08/02/10 10/04/10 

Manaus Barcelos Rio Negro 

area, 

Forest 

Amazon 

Barcelos 

Return 

Febrile 

polyarthritis 

Admission 

Severe 

arthralgia and 

incapacitation 

1 st serum sample 

MAYV IgM+ 

2nd serum sample 

MAYV IgM+ 

Clinical 

recovery 

97

The patient had travelled in the Amazon forest region 

for two weeks in October and November 2009 for the 

purpose of fishing and butterfly hunting. He stayed for 

two days in Manaus, Amazonas, Brazil, and for a further 

two days at Barcelos, Amazonas, north-western 

Brazil, before travelling in a dugout canoe along the 

Rio Negro River for ten days to the confluence area of 

the Demini River with the Araca River, a forest place 

situated 70 miles north of Barcelos. After another two 

days in Barcelos, he returned to France via Manaus and 

Sao Paulo (Figure). 

During his second stay in Barcelos, in early November, 

he developed symptoms assumed to be related to dengue 

virus infection, with high-grade fever, headache, 

generalised myalgia and diffuse arthralgia. Macular 

and partially confluent transient exanthema mainly on 

his arms appeared around the fifth day of illness. After 

his return to France, the patient had increasingly difficulty 

walking and was severely impaired in daily activities 

because of severe recurrent joint pains. 

The patient had received yellow fever vaccine 10 years 

before. During the trip to the Amazon, he had taken 

doxycyclin as prophylaxis for malaria. 

When he presented to our centre on 5 January 2010 

(two months after onset of symptoms), the patient complained 

of persistent headache, myalgia and severe 

symmetrical joint pains (wrists and ankles). At the time 

of presentation, laboratory tests showed a leukocyte 

cell count of 6,600 cells/µL and a thrombocyte count of 

177,000 platelets/µL. No markers of autoimmunity were 

found, notably anti-citrullin peptide antibodies or antinuclear 

antibodies. He was negative for the major histocompatibility 

complex HLA B27 gene. Concurrently, 

serologic status for dengue, chikungunya and yellow 

fever viruses as well as MAYV was evaluated using 

IgM capture and IgG sandwich ELISA at the National 

Reference Centre for Arboviruses, Institut Pasteur, 

Paris. Serology for MAYV revealed positive results for 

specific IgM (optical density [OD]=0.34; serum control 

OD=0.122). OD values for specific IgG were negative 

(OD=0.082; serum control OD=0.092). The other 

serological results were negative, as well as tests for 

leptospirosis, rickettsiosis, Q fever, cytomegalovirus 

and Plasmodium falciparum malaria. Five weeks later, 

on 8 February 2010, MAYV antibody serology showed 

persistence of specific IgM (OD=0.494; serum control 

OD=0.116) and a lack of immunoglobulin switching 

from IgM to IgG (OD for IgG=0.076; serum control 

OD=0.084). 

The patient recovered completely, although severe 

joint pain persisted for eight further weeks until 10 

April despite symptomatic treatment. The diagnosis 

of a presumptive case of MAYV infection diagnosed by 

serology was established. 

Discussion 

To the best of our knowledge, this case is the first published 

report of MAYV disease in a traveller returning 

to Europe. The presenting symptoms and signs were 

almost identical to those reported in previous clinical 

descriptions of the disease [2,4,5]. In this case, the 

decision to test for a rather exotic virus such as MAYV 

was based on several factors: the patient’s detailed 

travel history in tropical South America, which allowed 

risk factors to be identified such as potential exposure 

to vectors carrying diseases endemic in that area; the 

clinical presentation with incapacitating arthralgia following 

acute febrile illness; and finally, the expertise 

and technical tools available in the specialist clinic for 

tropical medicine where the patient was treated. Other 

viral infections with similar clinical presentation and 

geographical distribution were ruled out by laboratory 

tests. 

The case illustrates the challenge of clinically differentiating 

MAYV disease from classical dengue fever and 

other febrile exanthematous diseases that also circulate 

in South America, as well as the role of laboratory 

confirmation in establishing a correct diagnosis. 

Indeed, dengue fever was initially suspected considering 

its occurrence in most cities and places on tropical 

America, including the Amazon basin. The pathogenesis 

of debilitating symptoms in MAYV disease is still a 

poorly understood phenomenon [5], although persistent 

infection of synovial macrophages has been documented 

for other closely related and also arthritogenic 

alphaviruses [6]. The results of serological studies of 

the two consecutive convalescent-phase serum samples 

showed that the patient did not seroconvert with 

a switch from IgM to IgG. In most acute arboviral infections, 

IgM class-specific antibodies are generally no 

longer detectable after a period of 6-12 months post 

infection [7,8]. Considering the period for seroconversion 

in MAYV infection, we can therefore assume that 

the time between disease onset and the last late-phase 

blood sampling in this patient was not long enough for 

to allow Ig class switching. 

Interestingly, this report highlights the need for 

increased awareness MAYV disease as a differential 

diagnosis in travellers or migrants returning from 

endemic areas of tropical South America with febrile 

illnesses involving peripheral rheumatism and persistent 

arthralgia. Finally, it illustrates how travellers can 

act as signals for alert that can provide insights into 

the risk of transmission of infections in certain geographical 

areas. 

References 

1. Figueirido LTM. Emergent arboviruses in Brazil. Rev Soc Bras 

Med Trop. 2007;40(2):224-9. 

2. Tesh RB, Watts DM, Russell KL, Damodaran C, Calampa C, 

Cabezas C, et al. Mayaro virus disease: an emerging mosquitoborne 

zoonosis in tropical South America. Clin Infect Dis. 

1999;28(1):67-73. 


3. Sissoko D, Malvy D, Ezzedine K, Renault P, Moscetti F, Ledrans 

M, et al. Post-epidemic chikungunya disease in Reunion Island: 

course of rheumatic manifestations and associated factors 

over a 15-month period. PLoS Negl Trop Dis. 2009;3(3):e389. 

4. Azevedo RSS, Silva EVP, Carvalho VL, Rodrigues SG, Nunes 

Neto JP, et al. Mayaro fever virus, Brazilian Amazon. Emerg 

Infect Dis. 2009;15(11):1830-2. 

5. Taylor SF, Patel PR, Herold TJ. Recurrent arthralgias in a 

patient with previous Mayaro fever infection. South Med J. 

2005;98(4):484-5. 

6. Jaffar-Bandjee MC, Das T, Hoarau JJ, Krejbich Trotot P, 

Denizot M, Ribera A, et al. Chikungunya virus takes centre 

stage in virally induced arthritis: possible cellular and 

molecular mechanisms to pathogenesis. Microbes Infect. 

2009;11(14-15):1206-18. 

7. Malvy D, Ezzedine K, Mamani-Matsuda M, Autran B, Tolou 

H, Receveur MC, et al. Destructive arthritis in a patient with 

chikungunya virus infection with persistent specific IgM 

antibodies. BMC Infect Dis.2009;9:200. 

8. Weaver SC, Reisen WK. Present and future arboviral threats. 

Antiviral Res. 2010;85(2):328-45. 


99


Epidemiological analysis of mosquito-borne Pogosta 

disease in Finland, 2009 

J Sane (jussi.sane@helsinki.fi) 1 , S Guedes 2,3 , S Kurkela 1,4 , O Lyytikäinen 2 , O Vapalahti 1,3,5 

1. Department of Virology, Haartman Institute, Faculty of Medicine, University of Helsinki, Finland 

2. National Institute for Health and Welfare (THL), Department of Infectious Disease Surveillance and Control, Helsinki, Finland 

3. European Programme for Intervention Epidemiology Training (EPIET), European Centre for Disease Prevention and Control, 

Stockholm, Sweden 

4. Department of Virology, Helsinki University Central Hospital Laboratory, Finland 

5. Division of Microbiology and Epidemiology, Department of Basic Veterinary Sciences, Faculty of Veterinary Medicine, 

University of Helsinki, Finland 


Citation style for this article: Sane J, Guedes S, Kurkela S, Lyytikäinen O, Vapalahti O. Epidemiological analysis of mosquito-borne Pogosta disease in Finland, 

2009. Euro Surveill. 2010;15(2):pii=19462. Available online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19462 

Pogosta disease is a viral disease caused by a mosquito-borne 

alphavirus, Sindbis virus (SINV), and large 

human outbreaks of SINV infection have emerged in 

Finland every seven years. After a major outbreak in 

2002 an epidemic was expected to take place in 2009. 

Data from the National Infectious Disease Registry 

showed a small outbreak in humans in 2009 with a 

total of 105 reported cases but the seven-year cycle 

did not recur as anticipated. 

Introduction 

Sindbis virus (SINV) is a mosquito-borne alphavirus 

(of the family Togaviridae), present in Eurasia, Africa 

and Oceania [1,2]. Antibodies to SINV are detected in 

humans in various geographical areas but clinical infections 

caused by SINV are reported mostly from Finland 

where SINV is associated with fever, rash and arthritis, 

known as Pogosta disease [3]. The treatment is symptomatic. 

Clinically similar diseases are found in Sweden 

(Ockelbo disease) and in Russia (Karelian fever) [4,5]. 

The majority of clinical cases occur in Finland during 

August and September when the primary vectors, the 

ornitophilic late summer mosquito species Culex and 

Culiseta, are abundant. The incidence of Pogosta disease 

has been highest in the eastern parts of Finland 

in recent decades.[3,6]. 

Remarkably, outbreaks of Pogosta disease have thus 

far emerged every seven years since the first outbreak 

was noted in 1974, and the cause for this phenomenon 

is yet to be discovered. Tetraonid birds such as 

grouse, might contribute to this pattern [3]. Grouse 

have previously shown population cycles with population 

“crashes” coinciding with SINV outbreaks [7]. 

Antibodies to SINV have been detected in grouse and 

migratory birds [3,6] 

The last major epidemic in Finland took place in 2002 

with almost 600 reported cases and it was anticipated 

that an outbreak would occur again in 2009. This paper 

This article has been published on 14 January 2010 

describes the characteristics of the Pogosta disease in 

Finland from June through October 2009 and discusses 

the findings in relation with the previous epidemic. 

Methods 

Since 1995, all confirmed diagnoses of SINV infection 

have been reported to the National Infectious Disease 

Registry (NIDR) at the National Institute for Health and 

Welfare (THL). Notifications include information on 

date of sample collection, date of birth, sex, and on 

place of treatment. Multiple notifications of persons 

with the same date of birth, sex and place of treatment 

received within a 12-month period were combined as 

one case. The place of treatment refers to the health 

care center or hospital (in particular hospital district) 

where the diagnosis has been made. Data were analysed 

by sex, age, week and month of disease onset 

Figure 1 

Number and incidence rates of laboratory confirmed 

Sindbis virus cases, Finland, 1995-2009 (n=3,041) 



1,400 

1,200 

1,000 

800 

600 

400 

200 

0 

1,310 

40 

1995 

1996 

264 

1997 

1998 

135 123 

77 

27 

1999 

2000 

Sindbis virus cases 

597 

2001 

2002 

211 

105 

40 31 21 30 30 

2003 

2004 

2005 

2006 

2007 

2008 

2009 

Incidence rate = number of cases per 100,000 inhabitants 

30 

25 

20 

15 

10 

5 

0 

Incidence

and by hospital district of treatment. Finland has a 

population of 5.3 million and is divided into 20 hospital 

districts. Laboratory diagnosis is based on enzyme 

immunoassays (EIA) and/or in some cases a haemagglutination 

inhibition test (HI) (5). 

Results 

From June through October 2009, a total of 105 laboratory 

confirmed cases were reported to the NIDR and the 


incidence of SINV infection was two cases per 100,000 

inhabitants per year (Figure 1). Most of the cases 

occurred in September (n=60) followed by August 

(n=33). Sixty percent (n=63) of the cases were females. 

The highest incidence (4.6/100,000/year) was among 

persons aged 50-59 years. Only two of the cases were 

aged under 18 years. 

Figure 2 

Number and incidence rates of laboratory confirmed Sindbis virus cases, by health care districts, Finland 2009 (n=105) and 

2002 (n=597) 

Figure 3 

Sindbis virus cases in north Karelia and central Finland, 2009 (n=29) and 2002 (n=212) 


10 

9 

8 

7 

6 

5 

4 

3 

2 

1 

0 

North Karelia 

Central Finland 

June July Aug Sep Oct Nov Dec 


80 

70 

60 

50 

40 

30 

20 

10 

0 

North Karelia 

Central Finland 

June July Aug Sep Oct Nov Dec 

101

The incidence was highest in north Karelia, followed 

by east Savo, central Ostrobothnia and central Finland 

together with southern Ostrobothnia (Figure 2). The 

incidence rates in 2009 were considerably lower than 

those in 2002. 

The number of cases was highest in central Finland 

(n=15). The majority of the cases (n=10) in central 

Finland occurred in July-August whereas only one case 

was reported from north Karelia during this time period 

(Figure 3). On the contrary, 13 cases were reported from 

north Karelia and five from central Finland during the 

months of September-October. In 2002, cases peaked 

in September in both of the hospital districts. 

Discussion 

A major Pogosta disease outbreak has occurred every 

seven years in Finland since 1974, with hundreds or 

even thousands of patients. Following this pattern, 

another outbreak was expected for 2009. However, the 

number of cases was substantially lower than in previous 

epidemics in 1995 and 2002 when 1301 and 597 

cases were reported respectively [6]. The 105 cases 

reported in 2009 exceed the average number of cases 

(n=57) in the non-epidemic years during 1995-2009. 

However,in some intermediate years, 1997-1998, 2000 

and 2003, the number of cases exceeded the number 

reported in 2009. In comparison, five SINV infections 

were reported in Sweden in 2009 (Sirkka Vene, personal 

communication, 1 December 2009). 

The factors behind the puzzling cycles in the epidemiology 

of Pogosta disease are unclear but recently attention 

has focused on tetraonid birds. In the epidemic 

years of 1974 and 1981, grouse population crashed in 

north Karelia [4]. Further, detection of SINV antibodies 

in one quarter of grouse examinated in the year following 

the 2002 epidemic, indicated vast exposure of 

grouse to SINV, and suggested that the virus may have 

an endemic cycle in tetraonid birds [6]. The density of 

grouse was above average in 2007 but the population 

crashed in 2008. The recovery of grouse population 

can be rapid and was anticipated to occur in 2009. 

However, the density of their population continued 

to decline to an all time low (since the measurements 

from 1980s) [8]. Hence, it is plausible that grouse play 

a significant role in the human epidemiology of SINV. 

It is possible that the continuing decline in the grouse 

population in 2009 diminished the role of grouse as 

amplifying hosts and that therefore, a milder outbreak 

than expected was observed. 

Similar to previous findings, the incidence of SINV was 

highest in the hospital district of north Karelia. The difference 

in incidence of this hyperendemic region with 

other hospital districts was not, however, as prominent 

as previously. In 2009 many of the hospital districts 

with high incidence were located in central and northwestern 

parts of Finland. These observations may point 

towards a geographical shift in the incidence of SINV 

virus infection. The relatively low incidence in north 

Karelia compared to the epidemic in 2002 may also 

reflect the increased human seroprevalence towards 

SINV which indicates immunity. The district of Kainuu 

had no cases, which was surprising since previous 

studies showed high seroprevalence in that area [6]. 

This could be attributable to significant underdiagnosis 

or acquired immunity, which may result from high 

number of cases during the 1970s and 1980s. 

Most cases in central Finland occurred in August 

whereas in north Karelia the number of cases peaked 

in September. This could reflect the variations in mosquito 

activity and population size due to differences 

in weather conditions in these areas. The month of 

May was drier than normally in Joensuu (the largest 

city in north Karelia) but the rainfall in June-July was 

considerably higher than on average [9]. Perhaps the 

dry May in Joensuu contributed to fewer cases in July- 

August but the high rainfalls in June and July created 

better environmental conditions for mosquito development 

and thus, more human cases of SINV occurred in 

September-October. Weather conditions are also likely 

to influence human outdoor activities, and thereby 

exposure to SINV. 

In summary, a limited outbreak of SINV in humans took 

place in Finland in late summer and autumn of 2009 

but the expected seven-year cycle did not recur. The 

data suggest a geographical shift in disease incidence. 

It is likely that fluctuations in grouse populations play 

a major role in the occurrence of SINV epidemics. To 

further elucidate the role of tetraonid birds and other 

factors, such as weather and climate variations, more 

epidemiological studies and proper mathematical modeling 

of SINV epidemics is needed. 

References 

1. Taylor RM, Hurlbut HS, Work TH, Kingston JR, Frothingham TE. 

Sindbis virus: a newly recognized arthropodtransmitted virus. 

Am J Trop Med Hyg. 1955 Sep;4(5):844-62. 

2. Hubalek Z. Mosquito-borne viruses in Europe. Parasitol Res. 

2008;103 Suppl 1:S29-43. 

3. Brummer-Korvenkontio M, Vapalahti O, Kuusisto P, Saikku 

P, Manni T, Koskela P, et al. Epidemiology of Sindbis virus 

infections in Finland 1981-96: possible factors explaining a 

peculiar disease pattern. Epidemiol Infect. 2002;129(2):335-45. 

4. L’vov DK, Skvortsova TM, Gromashevskii VL, Berezina LK, 

Iakovlev VI. [Isolation of the causative agent of Karelian fever 

from Aedes sp. mosquitoes].[Russian]. Vopr Virusol. 1985 

May-Jun;30(3):311-3. 

5. Skogh M, Espmark A. Ockelbo disease: epidemic arthritisexanthema 

syndrome in Sweden caused by Sindbis-virus like 

agent. Lancet. 1982;1(8275):795-6. 

6. Kurkela S, Ratti O, Huhtamo E, Uzcategui NY, Nuorti JP, 

Laakkonen J, et al. Sindbis virus infection in resident birds, 

migratory birds, and humans, Finland. Emerg Infect Dis. 2008 

Jan;14(1):41-7. 

7. Lindén H. Characteristics of tetranoid cycles in Finland. Finnish 

Game Research. 1989;46:34-42. 

8. Finnish Game and Fisheries Research Institute. [Internet]. 

Helsinki: Metsäkanalinnut 2009. [Finnish]. Available from: 

http://www.rktl.fi/riista/riistavarat/metsakanalinnut_2009/. 

9. Finnish Meteorological Institute. [Internet]. Helsinki: Kuinka 

tilaan säätilastoja tai säähavaintosarjoja. [Finnish]. Available 

from: www.fmi.fi. Accessed 6 November 2009. 



Ongoing outbreak of West Nile virus infections in 

humans in Greece, July – August 2010 

A Papa 1 , K Danis (daniscostas@yahoo.com) 2 , A Baka 2 , A Bakas 3 , G Dougas 2 , T Lytras 2 , G Theocharopoulos 2 , D Chrysagis 3 , 

E Vassiliadou 3 , F Kamaria 3 , A Liona 2 , K Mellou 2 , G Saroglou 2 , T Panagiotopoulos 2,4 

1. Reference Laboratory for Arboviruses, First Department of Microbiology, Medical School, Aristotle University of Thessaloniki, 

Thessaloniki, Greece 

2. Hellenic Centre for Disease Control and Prevention (KEELPNO), Athens, Greece 

3. Department of Internal Medicine, Infectious Disease Hospital, Thessaloniki, Greece 

4. National School of Public Health, Athens, Greece 


Papa A, Danis K, Baka A, Bakas A, Dougas G, Lytras T, Theocharopoulos G, Chrysagis D, Vassiliadou E, Kamaria F, Liona A, Mellou K, Saroglou G, Panagiotopoulos 

T. Ongoing outbreak of West Nile virus infections in humans in Greece, July – August 2010. Euro Surveill. 2010;15(34):pii=19644. Available online: http://www. 


Between early July and 22 August 2010, 81 cases of 

West Nile neuroinvasive disease were reported in 

the region of Central Macedonia, northern Greece. 

The median age of cases was 70 years. Encephalitis, 

meningoencephalitis or aseptic meningitis occurred 

mainly in patients aged 50 years or older. This is the 

first time that West Nile virus (WNV) infection has 

been documented in humans in Greece. Enhanced surveillance 

and mosquito control measures have been 

implemented. 

Introduction 

On 4 August 2010, physicians from the Infectious 

Disease Hospital in Thessaloniki, northern Greece, 

informed the Hellenic Centre for Disease Control and 

Prevention (KEELPNO) about an increase in the number 

of hospitalised cases with encephalitis in the previous 

month (13 patients with encephalitis were hospitalised 

in July 2010, compared with a mean of five hospitalised 

cases in the same month of the previous three years). 

Despite several laboratory tests, no aetiological factor 

had been identified. Most patients were elderly (over 

65 years of age) and resided in the region of Central 

Macedonia, northern Greece. On the same day, 11 

serum and three cerebrospinal fluid (CSF) specimens 

from 11 patients with encephalitis and/or aseptic meningitis 

were sent for further testing to the Reference 

Laboratory for Arboviruses at the Aristotle University 

of Thessaloniki. The following day, the results showed 

that IgM antibodies against West Nile virus (WNV) had 

been detected in 10 of the 11 serum specimens and in 

all three CSF specimens. WNV infection in humans had 

not been previously documented in Greece. 

WNV is a positive-sense RNA virus of the Flaviviridae 

family, belonging to the Japanese encephalitis antigen 

group of viruses [1]. WNV is maintained in an enzootic 

cycle between birds and mosquitoes, mainly Culex 

species, while humans, horses and other mammals 

are incidental or dead-end hosts. Most human WNV 

infections are subclinical, and approximately 20% of 



infected individuals develop a febrile illness, while in 

less than 1%, the disease progresses to neuroinvasive 

disease, with the most severe form seen among elderly 

and immunocompromised individuals [2]. 

Although the virus was first isolated in 1937, interest in 

its impact on humans increased in 1996, when a large 

outbreak of West Nile neuroinvasive disease (WNND) 

was observed in Romania and in 1999, when WNV was 

introduced into the United States [3,4]. Several cases 

of WNV infection have been reported in horses and 

humans in Mediterranean countries [5-8], while WNND 

has been recently reported in humans in Hungary and 

Italy [7-9]. 

Methods 

Surveillance 

Following an alert issued by the Ministry of Health and 

KEELPNO on 6 August 2010 about 11 reported WNV 

infection cases, physicians in Greece were asked to 

notify KEELPNO of all confirmed or probable cases of 

WNV infection using a standardised reporting form, 

which included information on the demographic characteristics, 

clinical manifestations, underlying chronic 

medical conditions, potential risk factors and laboratory 

results of cases. The exact address of cases’ place 

of residence was obtained from hospital registries. In 

addition, active surveillance to identify cases included 

daily telephone inquiries to the hospitals of the region 

of Central Macedonia, from where the cases had been 

reported. 

Case definition 

The 2008 European Union case definition of WNV infection 

[10] was used, with slight modifications. A confirmed 

case was defined as a person meeting any of 

the following clinical criteria: encephalitis, meningitis, 

fever without specific diagnosis and at least one of the 

four laboratory criteria: (i) isolation of WNV from blood 

or CSF, (ii) detection of WNV nucleic acid in blood or 

CSF, (iii) WNV-specific antibody response (IgM) in CSF, 

103

and (iv) WNV IgM high titre, and detection of WNV IgG, 

and confirmation by neutralisation. 

A case was considered probable if the patient met the 

above clinical criteria and a WNV-specific antibody 

response was demonstrated in his or her serum sample. 

Epidemiological criteria were not used in the case 

definition due to the absence of recent surveillance 

data in animals. 

Laboratory methods 

Serum and CSF specimens were tested for the presence 

of WNV-specific IgM and IgG antibodies using commercial 

ELISA kits (Focus Technologies, Cypress, CA, 

USA). Reverse transcription-polymerase chain reaction 

(RT-PCR) was performed on RNA from 15 of 99 specimens, 

because the remaining samples had been taken 

between three and 15 days after the onset of illness, 

when viraemia is usually over. Primer sets specific for 

WNV and degenerate primers (able to detect flavivirus 

RNA) were used [11,12]. 

Data analysis 

Data were entered in a database designed using 

Epidata software (Epidata association, Denmark, version 

3.1) and were analysed using the GNU R software 

environment. Incidence was calculated using the 

2007 mid-year estimated population from the Hellenic 

Statistical Authority as denominator [13]. 

Results 

By 22 August 2010, 99 cases of WNV infection had been 

notified to KEELPNO. Of these, 81 had central nervous 

system manifestations (West Nile neuroinvasive disease, 

WNND) and 18 (eight probable and 10 confirmed 

cases) had only mild symptoms of fever and headache. 

We analyse here the 81 cases of WNND. Of these, 39 

were confirmed and 42 were probable cases. The overall 

incidence of WNND was 0.72 cases per 100,000 

population. 

In total, 77 serum and 47 CSF specimens were available; 

for 45 of the 81 WNND patients both CSF and serum 

specimens were provided, while for four patients only 

CSF was available. WNV-specific IgM antibodies were 

Figure 1 

Reported cases of West Nile neuroinvasive disease by date of symptom onset, Greece, 1 July – 22 August 2010 (n=81) 


6 

5 

4 

3 

2 

1 

1 2 3 4 5 

Probable case 

Confirmed case 

6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 

July August 

Date of symptom onset, 2010 

Table 1 

Characteristics of reported cases of West Nile neuroinvasive disease, Greece, 1 July – 22 August 2010 (n=81) 

Characteristic Number of cases Incidence (per 100,000 population) Risk ratio (95% CI) 

Age group (years) 

detected in all 77 serum and in 39 of the 47 CSF specimens, 

while WNV-specific IgG antibodies were detected 

in 42 of the 77 serum and 17 of the 47 CSF specimens. 

In 39 of the 45 patients for whom both types of specimen 

were available, the presence of IgM in both CSF 

and serum was seen, proving autochthonous antibody 

production; for IgG this was not tested. As crossreactions 

are common among flaviviruses, specimens 

were also tested for tick-borne encephalitis (TBE) virus 

(although TBE is not prevalent in the area and none of 


the patients reported tick bites): all were negative. Low 

cross-reactivity was seen with dengue virus; however, 

when a positive result was obtained for dengue virus, 

the titres were very low compared with the high titres 

seen for WNV. None of the patients had been vaccinated 

for yellow fever. RT-nested PCR was negative in 

all specimens tested. 

The first cases of WNND had onset of symptoms in 

early July (Figure 1). 

Figure 2 

Place of residence of reported cases of West Nile neuroinvasive disease, Greece, 1 July – 22 August 2010 (n=80) a 

a For one of the 81 cases of West Nile neuroinvasive disease, the place of residence has not been confirmed and is not included on the map. 

Each dot represents one case; the grey areas represent towns or cities; the blue lines represent rivers. 

The district of Larissa belongs to the region of Thessalia. All other districts from where cases have been reported belong to the region of 

Central Macedonia. 

105

The median age of the WNND cases was 70 years 

(range: 12–86 years), with most (n=71) aged 50 years 

or older (Table 1). Of all WNND cases, 45 (56%) were 

males. 

The incidence, 1.7 per 100,000 population, of WNND 

among those aged 50 years or older was almost 12 

times higher (risk ratio: 12.2; 95% confidence interval: 

4.9 to 28.5) than that of individuals aged under 50 

years. The risk of WNND was 27% higher among males 

compared with females (Table 1). 

The place of residence of the WNND cases is presented 

in Figure 2: 30 resided in an urban area and 51 in a rural 

setting. The vast majority (n=79) lived in districts (prefectures) 

of the region of Central Macedonia and only 

two were reported from the region of Thessalia (district 

of Larissa). It is of note that a large number of cases 

(n=58) lived near rivers and/or on the irrigated plains 

between and surrounding the Aliakmonas and Axios 

rivers (Figure 2). 

None of the notified cases reported travel to a known 

WNV-endemic area during the two weeks before onset 

of symptoms. Information on outdoor activities was 

gathered from 55 of the cases: 27 cases reported 

spending many hours outdoors in the countryside 

every day. None of the cases had a history of blood 

transfusion or tissue/organ transplant during the two 

weeks before the onset of symptoms. 

All notified cases with central nervous system manifestations 

were hospitalised. Of those, 65 had encephalitis 

or meningoencephalitis, and 16 had aseptic meningitis 

(Table 2). 

Of the 65 WNND cases with encephalitis and/or 

meningoencephalitis, 60 were aged 50 years or older. 

Information on underlying chronic medical conditions 

was available for 60 of the 81 patients. These included 

hypertension (n=26), a history of immunosuppression 

(n=17), coronary artery disease (n=11), and diabetes 

Table 2 

Clinical manifestations of reported cases of West Nile 

neuroinvasive disease by age group, Greece, 1 July – 22 

August 2010 (n=81) 

Age group (years) 


with encephalitis or 

meningoencephalitis 

Number of cases with 

aseptic meningitis 

Public health measures 

After the initial notification of the cluster of WNV infection 

cases, the Hellenic Centre for Disease Control and 

Prevention alerted physicians throughout the country 

and prepared a set of guidelines for health professionals, 

with the necessary instructions for laboratory 

diagnosis. Surveillance of human WNV infection cases 

has been established and the public has been given 

guidance about personal protective measures. 

The Hellenic Centre for Blood Transfusion prepared 

specific guidelines and measures for blood and blood 

products safety, including a 28-day deferral policy for 

all donors residing in the specific districts (prefectures) 

where WNV infection cases were detected, as 

well as any blood donor who had visited the same districts 

(prefectures) for one or more days. Blood units 

that had been collected in the same region since the 

beginning of July were quarantined until tested for 

WNV by PCR. They were all found negative for WNV 

and were released for use. Finally, specific advice was 

provided to all blood donors asking them to inform 

the blood donation department they attended, if they 

develop a fever within 15 days after their donation. The 

National Organisation for Transplantations has also 

been informed and is requesting WNV testing of donors 

depending on residence area or travel history. 

The Ministry of Health and Social Solidarity coordinates 

the intensification of mosquito control programmes at 

district level, which are already being implemented. 

The Ministry also undertook the coordination of the 

health sector preparedness and the collaboration with 

the veterinary services of the Ministry of Agriculture. 

Surveillance for WNV in mosquitoes (using bait traps) 

has been put in place. 

Updates on reported WNV infection cases and deaths 

in Greece are published in the daily epidemiological 

surveillance reports (available in English from http:// 

www.keelpno.gr/eng/wnv). 


We would like to thank all hospital physicians and local public 

health authorities who contributed to the surveillance 

of WNV infections in Greece. The technical support of the 

personnel in the Reference Laboratory for Arboviruses in 

Thessaloniki is highly appreciated. 

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107


Retrospective screening of solid organ donors in Italy, 

2009, reveals unpredicted circulation of West Nile virus 

M R Capobianchi (maria.capobianchi@inmi.it) 1,2 , V Sambri 3,2 , C Castilletti 1 , A M Pierro 3 , G Rossini 3 , P Gaibani 3 , F Cavrini 3 , M 

Selleri 1 , S Meschi 1 , D Lapa 1 , A Di Caro 1 , P Grossi 4 , C De Cillia 5 , S Venettoni 5 , M P Landini 3 , G Ippolito 1 , A Nanni Costa 5 , on behalf of 

the Italian Transplant Network 6 

1. National Institute for Infectious Diseases (INMI) “L. Spallanzani”, Rome, Italy 

2. These authors contributed equally to the work and share first authorship 

3. Regional Reference Centre for Microbiological Emergencies (CRREM), Unit of Clinical Microbiology, St Orsola University 

Hospital, University of Bologna, Bologna, Italy 

4. Department of Infectious Diseases, University of Insubria, Varese, Italy 

5. National Transplant Centre, Rome, Italy 

6. Members are listed at the end of the article 


Capobianchi M, Sambri V, Castilletti C, Pierro AM, Rossini G, Gaibani P, Cavrini F, Selleri M, Meschi S, Lapa D, Di Caro A, Grossi P, De Cillia C, Venettoni S, Landini 

MP, Ippolito G, Nanni Costa A, on behalf of the Italian Transplant Network. Retrospective screening of solid organ donors in Italy, 2009, reveals unpredicted 

circulation of West Nile virus. Euro Surveill. 2010;15(34):pii=19648. Available online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19648 

Since the occurrence of West Nile virus (WNV) infection 

in humans in 2008 in Italy, concerns have been 

raised about the potential risks associated with solid 

organ transplantation (SOT). A nationwide retrospective 

survey showed that 1.2% of SOT donors in 2009 

were WNV-seropositive and demonstrated that human 

WNV infection is distributed throughout several Italian 

regions. Transmission of WNV or other arboviruses 

through SOT is a possibility and risk assessment 

should be carried out before SOT to avoid infection 

through transplantation. 

Background 

In 1998, when the first cases of equine West Nile virus 

(WNV) infection in Italy were detected in Tuscany, no 

human cases were reported [1]. WNV re-emerged in 

Italy in 2008, and viral circulation was identified among 

different vectors and different animal species, including 

horses and wild birds [2]. The first cases of human 

WNV neuroinvasive infections in Italy were identified in 

September 2008 in the Emilia-Romagna region [3]. 

In the summer of 2009, additional human cases were 

reported in the same and in other neighbouring Italian 

regions. In one of the most affected areas in the province 

of Ferrara, a WNV seroprevalence of 0.68% was 

observed in healthy blood donors, raising concerns 

about the potential risks of WNV transmission by blood 

transfusion and organ transplantation [4,5]. 

In 2009, as a precautionary measure, the National 

Transplant Centre issued guidance that potential donations 

from all donors of solid organs, tissues and cells 

from the Bologna, Ferrara, Modena and Reggio-Emilia 

provinces who were screened by nucleic acid amplification 

tests (NAAT) for the presence of WNV viraemia, 


and tested positive, had to be rejected. Donors from 

other regions who had spent at least one overnight 

stay in the above-listed provinces during the 28 days 

before donation should not be considered eligible for 

donation [6]. 

Two days before screening of donors was implemented, 

in Bologna, Emilia-Romagna province, transmission 

of WNV infection through liver transplantation was 

detected by NAAT before clinical symptoms appeared 

in the recipient. The transmission of WNV was managed, 

post-transplant, by administration of hyperimmune 

serum and viraemia-guided adjustment of the 

immunosuppressive drug regimen, accompanied by 

supportive care [7]. The patient recovered from the 

transplantation and did not develop symptoms of WNV 

infection or sequelae. Before this case, all reported 

SOT donor-derived WNV infections were identified 

retrospectively in symptomatic transplanted patients 

with severe outcomes in the United States (US) [8]. 

This post-transplant detection of WNV infection in SOT 

recipients demonstrates that the absence of a universal 

screening policy in the immediate pre-transplant 

period makes it almost impossible to accurately quantify 

the transmission rate and the subsequent clinical 

impact of WNV transmission to SOT recipients. In the 

US, a recent serosurvey carried out among SOT recipients 

suggested that asymptomatic WNV infection may 

be common in areas of WNV activity and that the severe 

clinical presentations of WNV infections are equally 

frequent in both immunocompromised and immunocompetent 

subjects [9]. 

On the basis of this evidence, re-defining the risks for 

WNV transmission by organ, tissue and cell transplantation 

was identified by the National Transplant Centre 


as a necessary safety measure before the start of the 

next WNV season, from late spring to early autumn. 

The Italian Transplant Network considered this to be 

a priority in order to establish the factual basis for 

implementing future strategies for preventing WNV 

transmission. 

A nationwide retrospective survey of WNV seroprevalence 

was therefore undertaken in all SOT donors 

recruited by the National Transplant Centre during 

2009. 

Methods 

Serum samples from the Italian SOT donors in 2009 

stored in the biorepository facilities of the Italian 

Transplant Network, were analysed by the two reference 

laboratories identified by the National Transplant 

Centre for assessment of WNV infection: the Laboratory 

of Virology at the National Institute for Infectious 

Diseases ’L. Spallanzani‘ in Rome and the Regional 

Reference Centre for Microbiological Emergencies of 

the Microbiology Unit, St Orsola–Malpighi University 

Hospital, in Bologna. 

The presence of WNV-specific IgG and IgM was investigated 

using a two-step approach: first, a screening 

test was performed using a commercial enzyme-linked 

immunosorbent assay (ELISA) method (Euroimmun, 

AG, Lübeck, Germany); all samples that were positive 

Table 1 

Region of residence of solid organ donors tested for West Nile virus, Italy, 2009 (n=15) 


for either IgG or IgM were then confirmed by an immunofluorescent 

antibody assay (IFA, Euroimmun). Second, 

the IgG-positive sera were further evaluated by ELISA 

to measure the IgG avidity, using the method of Fox 

et al. [10]. All the IgG- and IgM-positive samples identified 

in the previous steps were further characterised 

by microneutralisation assay (MNTA) against WNV 

as previously described [11], and to rule out possible 

cross-reactions of the test, serum samples were also 

tested by MNTA against Usutu virus. All IgM-positive 

sera samples were retrospectively tested by NAAT 

using Procleix-WNV assay performed on the TIGRIS 

system, Novartis (for donors from Emilia Romagna) or 

Artus Real Art WNV LC RT RCRt kit, QIAGEN (for donors 

from Piedmont and Tuscany) in order to evaluate the 

presence of WNV viraemia. During the screening activity 

for WNV in 2009 no WNV-positive donor was identified 

before SOT. 

Results 

A total of 1,248 serum samples from SOT donors in 2009 

were analysed, accounting for 98.1% of SOT donors 

recruited during that year. Table 1 lists the number of 

SOT donors evaluated in this study, by region of residence. 

WNV-specific antibodies were detected in 15 

samples from individual donors at the time of organ 

donation, thus giving an overall positivity rate of 1.2%. 

Data from MNTA indicate that seven of 15 samples had 

Region Number of donors tested Number of donors positive WNVa Abruzzo-Molise 12 0 

Basilicata 11 1 

Calabria 12 0 

Campania 74 0 

Emilia Romagna 120 4 

Friuli-Venezia Giulia 45 1 

Lazio 103 1 

Liguria 33 0 

Lombardy 226 0 

Marche 51 0 

Piedmont-Aosta Valley 120 2 

Bolzano-Bozen independent provincial administration 10 0 

Trento independent provincial administration 17 0 

Apulia 44 0 

Sardinia 24 0 

Sicily 59 0 

Tuscany 167 6 

Umbria 11 0 

Veneto 109 0 

Total 1,248 15 

WNV: West Nile virus 

a IgG - and/or IgM-positive, by serological screening (see Table 2). 

109

neutralisation activity against WNV. However, eight 

samples did not show appreciable neutralisation titre 

against WNV. Among these, two samples were reverse 

transcription-polymerase chain reaction (RT-PCR) positive 

and two other samples probably showed neutralisation 

activity against the related Usutu virus (Table 2). 

The inability to confirm by MNTA all the samples testing 

positive by ELISA and IFA might be due to the 

cross-reactivity to closely related flaviviruses or might 

be consistent with the notion that the neutralising 

response to WNV in humans is variable and that only 

a subset of infected individuals generate antibodies 

against high-neutralising epitopes [12]. Furthermore, 

from a technical point of view, maturation state of WNV 

particles used in the MNTA might have been important 

for determining whether antibodies in a given 

serum samples were judged WNV-specific by MNTA , as 

already reported by Nelson et al. [13]. 

The highest number of WNV-seropositive donors were 

from the regions of Emilia-Romagna and Tuscany (Table 

1 and Figure). 

Table 2 shows the place of residence, demographic 

information and laboratory results obtained for these 

15 patients. 

Figure 

Distribution of solid organ donors shown to be positive 

for West Nile virus a , Italy, 2009 (n=15) 

a Shown to have West Nile virus-specific antibodies (IgG and/ 

or IgM) by enzyme-linked immunosorbent assay (ELISA) and 

immunofluorescent antibody assay (IFA). 

Two donors (from Piedmont and Emilia-Romagna) were 

identified as IgG- and IgM-positive. One donor, from 

Tuscany, was positive only for IgM. All these three 

IgM-positive samples were also positive for viral RNA, 

showing quite a low viral load (equivalent to a genome 

copy number of ≤3 log copies/ml). We speculate that 

such a low level of viral concentration in blood is likely 

to pose a limited risk of viral transmission through 

SOT. Four IgG-positive specimens showed an antibody 

avidity of 80% to 90% suggesting that exposure to 

WNV probably occurred more than six months before 

the organ donation; another four IgG-positive samples 

(all from donors who donated organs after June 2009) 

showed an antibody avidity lower than 40 %, suggesting 

that the infection was probably acquired within six 

months before the organ donation, a time frame consistent 

with WNV exposure during the 2009 mosquito 

season (June to October). 

In addition, recipients of a solid organ from donors who 

were shown to have been positive for WNV genome 

or to have had IgM-specific antibodies at the time of 

organ donation were also included in this study. Two 

recipients from the Piedmont IgM-positive donor (one 

kidney recipient and one liver recipient) did not show 

any seroconversion to WNV. The remaining recipient 

(who received a kidney) was not available for testing. 

All three recipients are well and have never shown 

signs consistent with WNV infection. The IgM-positive 

donor from Tuscany did not in the end donate organs 

for reasons not related to WNV infection. 

Discussion and conclusions 

The occurrence of an antibody response in some donors 

from the Piedmont, Friuli-Venezia Giulia, Marche and 

Basilicata regions is rather unexpected and shows 

evidence of WNV infection in humans in several Italian 

regions. However, it is possible that WNV activity has 

hitherto been underestimated in some regions, due to 

an insufficient veterinarian and entomological surveillance 

system, which may have generated inconsistent 

data. Evidence from the present study – i.e. the low IgG 

avidity index, the presence of an IgM-specific response 

and/or WNV RNA in blood samples – is consistent with 

the notion that recrudescence of WNV activity in Italy 

occurred in the last two years, following the report 

of the first human cases of neuroinvasive disease 

[3-5]. These findings highlights the need for an accurate 

nationwide approach to risk assessment related 

to transplantation, in order to implement appropriate 

prevention strategies and limit the potential burden of 

severe neurological complications in the immunocompromised 

recipients. During the 2009 season, only 

one case of WNV transmission by SOT was observed 

in the Emilia-Romagna region [7]. No additional cases 

of WNV transmission from infected donors were documented 

retrospectively in our study, suggesting that 

the transmission of WNV to recipients of SOT from 

viraemic donors does not always occur. Furthermore, 


the positivity in MNTA against Usutu virus in two samples 

additionally confirms that this virus has to be 

added to the list of those that can be transmitted to 

humans [14,15] and its possible transmission through 

SOT deserves particular attention. 

These results indicate that, due to the well-known circulation 

of WNV in many different areas in Italy, transmission 

of WNV or other arboviruses through SOT is 

possible, and that the risk assessment process related 

to transplantation is a challenging issue that requires a 

systematic approach [7]. 

Regional Coordinators of the Italian Transplant 

Network: 

M Scalamogna (Milan); A Famulari (L’Aquila); A Saracino 

(Matera); B Giacon (Bolzano); P Mancini (Reggio 

Calabria); E Di Florio (Naples); L Ridolfi (Bologna); 

R Peressutti (Udine); D Adorno (Roma); A Gianelli 

Castiglione (Genoa); S Vesconi (Milan); D Testasecca 


(Ancona); A Amoroso (Turin); F Paolo Schena (Bari); 

C Carcassi (Cagliari); V Sparacino (Palermo); R Lippi 

(Florence); E Gabardi (Trento); C Gambelunghe 

(Perugia); F Calabrò (Padua). 


The work was supported in part by grants to the National 

Transplant Center and to the National Institute for Infectious 

Diseases (INMI) (Ricerca Corrente, Ricerca Finalizzata/ 

Progetto ordinario 2007-9AEF Grant 28C5/3 ), and to CRREM 

(Regione Emilia Romagna). 

Table 2 

Detailed information of solid organ donors who tested positive by serological screening for West Nile virus 

Region 

Sampling 

date, 2009 

Cause of death 

Age at 

death 

(years) 

Place of 

residence 

(province) 

Possible place of exposure a 

(if different from 

place of residence) 

WNV- specific 

antibody titres 

IgG IgM 

IgG 

avidity 

(%) b 

MNTA 

titre 

against 

WNV d 

Basilicata 22 Sep Stroke 64 Matera Lake Garda ≥1:160

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eurosurveillance.org/Public/Articles/Archives.aspx 



Case report: West-Nile virus infection in two Dutch 

travellers returning from Israel 

N Aboutaleb 1 , M FC Beersma 2 , H F Wunderink 3 , A CTM Vossen 3 , L G Visser (L.G.Visser@lumc.nl) 1 

1. Department of Infectious Diseases, Leiden University Medical Centre, Leiden, Netherlands 

2. Department of Virology, Erasmus University Medical Centre, Rotterdam, Netherlands 

3. Department of Virology, Leiden University Medical Centre, Leiden, Netherlands 


Aboutaleb N, Beersma MF, Wunderink HF, Vossen AC, Visser LG. Case report: West-Nile virus infection in two Dutch travellers returning from Israel. Euro Surveill. 


We report about West Nile virus (WNV) infections in 

a symptomatic traveller returning from Israel and in 

her asymptomatic travel companion. Knowledge of the 

current epidemiological situation in Israel from where 

WNV cases were reported recently enabled a rapid 

diagnosis. The described cases serve as a reminder 

for physicians to consider WNV in the diagnosis of 

patients returning from areas with potential circulation 

of the virus. 

At the end of July 2010, a Dutch woman in her early 

thirties presented to our first aid department with 

fever, retro-orbital headache and a macular rash. The 

day before she had returned from a 10-day holiday to 

Israel where she noticed several mosquito bites during 

a camping trip at the Sea of Galilee. There was no history 

of tick bites. Five days before presentation, she 

had suddenly fallen ill with extreme fatigue, myalgia, 

fever and an increasingly severe headache. Movement 

of the eyes had been painful. The next day she had 

developed a burning sensation of the skin that was followed 

by a skin eruption. The rash started on the trunk 

and spread to arms and legs. 

Upon presentation in the first aid department she 

did not appear ill. Body temperature was 36.4°C. 

There was a generalised macular rash, sparing hands 

and feet, and no petechiae or eschars were present. 

Neurological examination revealed no abnormalities. 

Potential infection with West-Nile virus was suspected 

because of the clinical picture and recent reports of 

West Nile virus cases in Israel [1]. 

Laboratory examination showed a leukopenia of 

1.8x10 9 /L with a predominance of large granular lymphocytes; 

thrombocyte count was 92x10 9 /L. Infection 

with Epstein Barr virus, cytomegalovirus, dengue 

virus, enterovirus and parechovirus was excluded by 

serology and antigen test or PCR. A lumbar punction 

was not performed due to missing neurological symptoms. 

West Nile virus RNA could not be detected in the 

EDTA plasma sample taken on day five after onset of 

disease by a TaqMan reverse transcriptase-PCR assay, 

using a probe for the WN3’NC [2]. However, in a second 

blood sample obtained fifteen days later, seroconversion 

for both serum IgG and IgM antibodies against 



West Nile virus was detected with an indirect qualitative 

enzyme-linked immunosorbent assay (ELISA) 

(Focus Diagnostics, Cypress, California). This assay 

uses antibody capture technique for the detection of 

IgM antibodies. 

Once the diagnosis was confirmed, serology was performed 

in the patient’s travel companion who reported 

having had similar complaints, but who had already 

recovered by the time he returned to the Netherlands. 

The laboratory results showed that he was seropositive 

for WNV IgG and IgM. Recovery was uneventful in 

both patients. 

West-Nile virus is endemic in Israel. The largest recent 

outbreak in humans in Israel occurred in the year 2000 

with more than 400 reported cases [3]. Recently, 12 

cases of West Nile fever have been reported, centered 

around the Tel Aviv area [1]. Culex perexiguus, Cx. pipiens, 

and Aedes caspius are the vectors of West Nile 

virus in Israel where most cases of West Nile fever 

occur between August and October. The seasonal 

occurrence of human cases reaches a peak one month 

after the mosquito peak [4]. 

Measures to avoid mosquito bites such as wearing protective 

clothes and using repellents are recommended 

during the whole transmission season and this case 

report serves as a reminder to physicians to consider 

West Nile fever in patients with fever returning from 

Israel. 

References 

1. West Nile fever hits 12 people in Israel, leaving one dead. 

Haaretz.com [online]. 2 August 2010. Available from: http:// 

www.haaretz.com/print-edition/news/west-nile-fever-hits-12people-in-israel-leaving-one-dead-1.305379 

2. Lanciotti RS, Kerst AJ, Nasci RS, Godsey MS, Mitchell CJ, 

Savage HM, et al. Rapid detection of West Nile virus from 

human clinical specimens, Field-collected mosquitoes, and 

avian samples by a TaqMan reverse transcriptase-PCR assay J 

Clin Microbiol. 2000 Nov;38(11):4066-71. 

3. Chowers MY, Lang R, Nassar F, Ben-David D, Giladi M, 

Rubinshtein E et al. Clinical characteristics of the West 

Nile fever outbreak, Israel, 2000. Emerg Infect Dis. 

2001;7(4):675-8. 

4. Orshan L, Bin H, Schnur H, Kaufman A, Valinsky A, Shulman L, 

Mosquito vectors of West Nile fever in Israel. J Med Entomol. 

2008;45:939-47. 

113


Introduction and control of three invasive mosquito 

species in the Netherlands, July-October 2010 

E J Scholte (e.j.scholte@minlnv.nl) 1 , W Den Hartog 1 , M Dik 1 , B Schoelitsz 2 , M Brooks 2 , F Schaffner 3,4 , R Foussadier 5 , M Braks 6 , 

J Beeuwkes 1 

1. National Centre for Monitoring of Vectors (CMV), New Food and Consumer Product Safety Authority (nVWA), Dutch Ministry of 

Economic Affairs, Agriculture and Innovation, Wageningen, the Netherlands 

2. Kenniscentrum Dierplagen (KAD), Wageningen, the Netherlands 

3. Agriculture and Veterinary Information and Analysis (Avia-GIS), Zoersel, Belgium 

4. Institute of Parasitology, University of Zurich, Switzerland 

5. Entente Interdépartementale (EID) Rhone-Alpes, Chindrieux, France 

6. Laboratory for Zoonoses and Environmental Microbiology, Centre for Infectious Disease Control (RIVM), the Netherlands, 

Bilthoven, the Netherlands 


Scholte EJ, Den Hartog W, Dik M, Schoelitsz B, Brooks M, Schaffner F, Foussadier R, Braks M, Beeuwkes J. Introduction and control of three invasive mosquito 

species in the Netherlands, July-October 2010. Euro Surveill. 2010;15(45):pii=19710. Available online: http://www.eurosurveillance.org/ViewArticle. 


In July 2010, during routine mosquito surveillance 

inspections at companies that import used tires, 

three invasive species were found at five locations 

in the Netherlands: the yellow fever mosquito (Aedes 

aegypti), the Asian tiger mosquito (Ae. albopictus), 

and the American rock-pool mosquito (Ae. atropalpus). 

This is the first time that Ae. aegypti is reported from 

the Netherlands. Mosquito control was initiated one 

week after the first invasive mosquito was found, 

using adulticides and larvicides. The available data 

suggest that the implemented control measures have 

been effective for this season. 

Introduction 

Following the discovery of Aedes albopictus in the 

Netherlands in 2005 related to companies that import 

Lucky bamboo [1], continuous surveillance at these 

companies was started in 2006. Gradually, other 

national surveillance activities for this mosquito species 

were initiated, including passive surveillance 

(since 2007), active surveillance at parking lots along 

main highways entering the country from the south and 

east (since 2008), and at companies that import used 

tires (since 2009). In 2009, during routine surveillance 

activities, the exotic mosquito species Ae. atropalpus, 

a North American species that had been encountered 

several times in Europe [2], but had never established 

here, was found for the first time in the Netherlands 

[3]. 

These surveillance activities are meant to identify 

as early as possible the presence of exotic mosquito 

species with the aim to prevent the establishment of 

invasive exotic mosquito species, especially those that 

are known to be vectors of pathogens of public health 

importance such as dengue- and chikungunya virus. 

Here we report the finding and the successive con- 

Article published on 11 November 2010 

trol of three invasive mosquito species, Ae. aegypti, 

Ae. albopictus and Ae. atropalpus in the Netherlands. 

Methods 

A total of 34 companies that import used tires into 

the Netherlands were included in the invasive mosquito 

survey. Routine inspections were carried out 

from April to the last week of October [2]. A qualitative 

risk assessment on the introduction of invasive 

mosquito species was performed to determine the 

frequency of inspection of a company. Parameters 

in the risk assessment were (i) the type of tires that 

are imported, (ii) the countries from which tires are 

imported, and (iii) whether the tire storage is in- or 

outdoors. Collected larvae and adult mosquitoes were 

diagnosed either morphologically by using the diagnostic 

keys from Schaffner et al. [4], or molecularly 

by PCR sequencing the mitochondrial cytochrome 

oxidase subunit 1 (CO1) gene [5]. A week after the 

first finding, infested locations were treated by spraying 

Bacillus thuringiensis israelensis (B.t.i.) serotype 

H14 or Bacillus sphaericus (B.s.) against larvae and/ 

or deltamethrin (aqua K-Othrine, Bayer Environmental 

Sciences) against adult mosquitoes. Larval control of 

the surrounding area (predefined perimeter of 500 

m) consisted of removal of potential larval habitats 

for container-breeding Aedes spp. when possible, or 

treatment with either B.t.i. space spray (VectoBac WG, 

Valent BioSciences), or with B.t.i./Bacillus sphaericus 

(B.s.) granules (Vectomax, Valent BioSciences). It was 

decided to perform larvicidal treatment once every two 

to three weeks, until the first week of November. 

Following the discovery of an exotic species at a 

location, surveillance was intensified to assess the 

potential spread of the invasive species and the effectiveness 

of the control activities by placing traps for 

adult mosquitoes (BG-sentinel, Biogents) and oviposi- 


tion traps [6] in the 500 m perimeter surrounding the 

company site. 

Results 

Three exotic mosquito species (Ae. aegypti, 

Ae. albopictus, and Ae. atropalpus) were found in five 

locations in the Netherlands. The first two mosquito 

larvae, Ae. atropalpus, were found on 21 July 2010, 

during a routine inspection at Location 1 (Heijningen) 

(Figure, Table 1). 

On the next day, during an intensified inspection, one 

adult Ae. albopictus and one adult Ae. aegypti were collected, 

in addition to the two initial Ae. atropalpus larvae. 

The infestation level for Ae. atropalpus (in terms 

of percentage of infested tires and total number of larvae) 

at this company was relatively high, but less so for 

Ae. albopictus and Ae. aegypti, of which no larvae and/ 

or pupae were found. Results of intensified inspection 

suggest that Ae. atropalpus and Ae. albopictus (but not 

Ae. aegypti) had spread to the surrounding areas of 

Location 1. On 3 September 2010, the last exotic species 

was collected from Location 1 and its surroundings 

(Table 2). 

At Location 2 (Oosterhout), several male Ae. aegypti 

specimens were collected starting with 26 July. The 

last invasive species were found at this location on 6 

August, when two adult Ae. atropalpus were collected. 

Despite intensive surveillance, no immature forms of 

invasive species were found at the company’s premises 

or in the surrounding areas. 

Figure 

Locations of tire companies that were positive (n=5) and 

negative (n=29) for at least one of the invasive mosquito 

species, the Netherlands, 2010 

Negative locations 

Positive locations 

Amsterdam 

Den Haag Utrecht 

1 

2 


5 

0 15 30 60 90 120 Km 

3 

Eindhoven 

4 

Groningen 

On 26 July, three adult specimens (but no larvae) of 

Ae. atropalpus were collected from Location 3 (Oss). 

In addition, another three adult Ae. atropalpus were 

found on 5 August at this location. In the surrounding 

area, one Ae. albopictus was collected in a BG-sentinel 

trap placed approximately 50 m from the tire platform 

on 9 August, but no larvae of exotic species were found 

in the surrounding area. The last specimen (larva) was 

found at this location on 23 August. 

On 24 August, the first larvae (six specimens) of Ae. 

atropalpus were collected from Location 4 (Weert). On 

13 September, high numbers of this species (larvae and 

adults) were found at this location and several larvae 

were found in the surrounding area, including at the 

premises of a neighbouring tire-importing company. 

The two companies are considered as one location 

(Location 4). In addition, one Ae. albopictus specimen 

was collected from the tire platform on 13 September 

and one Ae. albopictus specimen was found in a BG 

sentinel trap at approximately 25 m from the infested 

companies, one week later. The last specimen was 

found on 27 September. 

On 28 September, two Ae. albopictus larvae were collected 

from Location 5 (Montfoort). The third (and last) 

specimen was collected in an adult trap on the tire 

platform on 5 October. No specimens were found in the 

surrounding area. 

All infested companies described here belong to the 

‘high risk’-category for importing exotic mosquito species, 

based on the type, origin and storage of the tires 

that are imported, and are therefore inspected every 

two weeks. No invasive mosquito species were found 

at any of the other companies that were included in the 

survey. 


The discovery of Ae. aegypti in the Netherlands was 

unexpected, mostly because, unlike Ae. albopictus 

[3], Ae. aegypti is not directly associated with the 

international trade in used tires [7]. Even without control 

measures, the tropical Ae. aegypti will probably 

not survive the winter in temperate areas such as the 

Netherlands and consequently does not pose a direct 

health risk for the country. This is in contrast with the 

public health risks related to re-introduction of Ae. 

aegypti into southern Europe [8,9]. 

In addition, this report describes the discovery of an 

Ae. albopictus for the first time in the outdoor environment 

in the Netherlands. Although the species is still 

regularly found in glasshouses as hitchhikers in importation 

of Lucky bamboo [10], preventive and curative 

indoor control measures in these glasshouses appear 

to be effective to prevent indoor or outdoor establishment, 

since a location never stays positive for Ae. 

albopictus longer than 1,5 month (Scholte, unpublished 

data). 

115

Back-tracing data of the company at Location 1 suggests 

introduction of Ae. albopictus and Ae. aegypti by a 

shipment of used airplane tires at the end of May 2010, 

originating from southern Florida, an area inhabited by 

both species. On 24 July, part of the same shipment 

was transported to Location 2 (belonging to the same 

company), and on 4 August to Location 3. Back-tracing 

information of the companies at Location 4 showed 

recent tire import from Italy. Ae. albopictus from Italy 

[11] and the United States [12] are considered to display 

diapause and potentially to survive temperate climates 

[13,14]. Ae. atropalpus had already been found at two 

sites in the Netherlands in 2009 [2] which indicates 

that the first introduction of Ae. atropalpus was in or 

before 2009, although more recent introductions are 

not excluded either. This species had a relatively large 

population at Locations 1 and 4, and colonised larval 

habitats in the surrounding areas, other than tires. 

The fact that relatively few adults and no other lifestages 

of Ae. aegypti and Ae. atropalpus were found at 

Location 2, indicates a low level of infestation. 

The available data for this season (Table 2) suggest 

that the implemented control measures have been 

effective, although it is too early at this moment in 

time to assess if eradication has been achieved. Per 

location, it took between one and three treatments 

and a maximum time span of seven weeks between the 

first treatment and the day when the last exotic species 

was found. It will be crucial in the years to come to 

monitor the locations (including the surrounding areas) 

that had been infested with one or more of the exotic 

species in 2009 and 2010, in order to restart mosquito 

control as early as possible. 

Having witnessed these introductions of exotic invasive 

mosquito species that pose a potential threat to 

public health in Europe, international collaboration 

and action of medical entomologists, public health 

experts, policy makers, and the tire-business industry 

is critical to address this. 


Roel Coutinho (RIVM), Marleen Kraaij-Dirkzwager (Ministry 

of VWS), Corien Swaan (RIVM), André Jacobi (RIVM), Peter de 

Vries (VROM), Martin van den Homberg (nVWA), Ad Dilven 

(GGD West Brabant), Roel Vincken (nVWA), Nicole Bouwer 

(nVWA), Arie Verhoef (VACO), Robert Luttik (RIVM), Heiko 

Kotter (Sumitomo). 

Table 1 

Summary of the results of the invasive mosquito survey at used tire companies by location, the Netherlands, July-October 2010 

Location 

Ae. aegypti 

Adults collected 

Ae. albopictus Ae. atropalpus Total Ae. aegypti 

Larvae collected 

Ae. albopictus Ae. atropalpus Total 

1 5 11 68 84 0 0 80 80 

2 8 0 2 10 0 0 0 0 

3 0 1 6 7 0 0 1 1 

4 0 2 45 47 0 6 122 128 

5 0 1 0 1 0 2 0 2 

Total 13 15 121 149 0 8 203 211 

Table 2 

Inspections, mosquito control, and findings of at least one of the three exotic mosquito species for each location per week, 

the Netherlands, July-October 2010 

Location 

15 16 17 18 19 20 21 22 23 24 25 26 27 

Week (2010) 

28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 

1 x x x x x x x 

2 x x x x x 

3 x x x x x 

4 x x x x 

5 x x 

No inspection 

No exotic species found (negative) 

Larvae and/or adults found of one of the three exotic mosquito species 

X Control measures 


References 

1. Scholte EJ, Jacobs F, Linton YM, Dijkstra E, Fransen J, Takken 

W. 2007. First record of Aedes (Stegomyia) albopictus in the 

Netherlands. Euro Mosq Bull. 2007;22:5-9. 

2. Schaffner F, Van Bortel W. Current status of invasive 

mosquitoes in Europe. European Centre for Disease Prevention 

and Control; 31 Jan 2010. Available from: http://www.ecdc. 

europa.eu/en/activities/sciadvice/Lists/ECDC%20Reviews/ 

ECDC_DispForm.aspx?List=512ff74f-77d4-4ad8-b6d6bf0f23083f30&ID=758&RootFolder=/en/activities/sciadvice/ 

Lists/ECDC%20Reviews&MasterPage=1 

3. Scholte EJ, Den Hartog W, Braks M, Reusken C, Dik M, Hessels 

A. First report of a North American invasive mosquito species 

Ochlerotatus atropalpus (Coquillett) in the Netherlands, 2009. 

Euro Surveill. 2009;14(45):pii=19400. Available from: http:// 

www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19400 

4. Schaffner F, Angel G, Geoffrey B, Hervy J-P, Rhaiem A, 

Brunhes J. The mosquitoes of Europe. CD-ROM. Montpellier: 

Institut de Recherche pour le Développement/Entente 

interdépartementale pour la démoustication du littoral (EID) 

Méditerrannée ; 2001. 

5. Simon C, Frati F, Beckenbach A, Crespi B, Liu H, Flook P. 

Evolution, weighting, and phylogenetic utility of mitochondrial 

gene sequences and a compilation of conserved polymerase 

chain reaction primers. Ann Entomol Soc America. 

1994;87(6):651-701. 

6. Fay RW, Eliason DA. A preferred oviposition site as a 

surveillance method for Aedes aegypti. Mosq News. 

1966;26:531-5. 

7. Schaffner F. Mosquitoes in used tyres in Europe: species list 

and larval key. Eur Mosq Bull. 2003;16:7-12. 

8. Theiler M, Casals J, Moutousses C. Etiology of the 1927- 

28 epidemic of dengue in Greece. Proc Soc Exp Biol Med. 

1960;103:244-6. 

9. Reiter P. Yellow fever and dengue: a threat to Europe?. Euro 



10. Scholte EJ, Dijkstra E, Blok H, De Vries A, Takken W, Hofhuis A, 

et al. Accidental importation of the mosquito Aedes albopictus 

into the Netherlands: a survey of mosquito distribution and the 

presence of dengue virus. Med Vet Entomol. 2008;22(4):352-8. 

11. Romi R, Severini F, Toma L. Cold acclimation and overwintering 

of female Aedes albopictus in Roma. J Am Mosq Control Assoc. 

2006;22(1):149-51. 

12. Lounibos LP, Escher RL, Lourenço-de-Oliveira R. Asymmetric 

evolution of photoperiodic diapause in temperate and tropical 

invasive populations of Aedes albopictus (Diptera: Culicidae). 

Ann Entomol Soc Am. 96(4):512-18. 


Development of Aedes albopictus risk maps. Technical report. 

Stockholm:ECDC. 2009. Available from: http://ecdc.europa. 

eu/en/publications/Publications/0905_TER_Development_of_ 

Aedes_Albopictus_Risk_Maps.pdf 

14. Takumi K, Scholte EJ, Braks M, Reusken C, Avenell D, Medlock 

JM. Introduction, scenarios for establishment and seasonal 

activity of Aedes albopictus in the Netherlands. Vector Borne 

Zoonotic Dis. 2009;9(2):191-6. 


117

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