ORIGINAL ARTICLE
In-vivo evaluation of the contamination
of Super Slick elastomeric rings by
Streptococcus mutans in orthodontic patients
Amanda Fahning Ferreira Magno,a Carla Enoki,b Izabel Yoko Ito,c Mirian Aiko Nakane Matsumoto,d
Gisele Faria,e and Paulo Nelson-Filhof
São Paulo, Brazil
Introduction: We investigated in vivo the contamination by Streptococcus mutans of Super Slick elastomeric
rings (TP Orthodontics, LaPorte, Ind), manufactured with Metafasix technology (TP Orthodontics), using
microbial culture and scanning electron microscopy (SEM). Methods: Twenty patients undergoing fixed
orthodontic appliance therapy were selected. Super Slick elastomeric rings (n = 160) were tied to brackets on
the right maxillary premolars or molars and left mandibular premolars or molars. Conventional elastomeric
rings (n = 160) were tied to brackets on the contralateral premolars or molars with the same split-mouth
design. After a 15-day intraoral period, 75 elastomeric rings of each type were retrieved, submitted to
microbiologic processing, and cultured in bacitracin sucrose broth-selective enrichment broth culture
media. The number of S mutans colonies or biofilms on the surface of the electrometric rings was
counted by using a stereomicroscope. Data were analyzed statistically with the Wilcoxon nonparametric
test at the 5% significance level. Four representative rings of each type were chosen for SEM analysis.
Results: Statistical analysis by the Wilcoxon nonparametric test showed that the Super Slick elastomeric
rings had statistically significant greater S mutans contamination than the conventional elastomeric rings
(P <.0001). No formation of S mutans colonies or biofilms was observed in the elastomeric rings removed
directly from their original packages. SEM micrographs showed fissures on the surface of Super Slick
elastomeric rings. No fissures were found on conventional elastomeric rings. When the microbiologic
culture was positive, S mutans bacterial biofilm was observed on both types of ligatures. Conclusions:
There was no clinical evidence that Super Slick elastomeric rings are effective in reducing bacterial biofilm
formation on their surfaces, and a recommendation for their use in orthodontic therapy for that purpose is
not justifiable. (Am J Orthod Dentofacial Orthop 2008:133:S104-9)
P
atients receiving orthodontic treatment have
alterations in the oral cavity, such as drops in pH
and the creation of additional retentive sites for
From the University of São Paulo, Ribeirão Preto, Brazil.
a
Orthodontist, research fellow, Department of Pediatric Clinics, Preventive and
Social Dentistry, Faculty of Dentistry.
b
Orthodontist, research fellow, Department of Ophthalmology, Otorhinolaryngology and Head and Neck Surgery, Faculty of Medicine and Department of
Pediatric Clinics, Preventive and Social Dentistry, Faculty of Dentistry.
c
Professor, Department of Clinical Analysis, Toxicology and Bromatology,
Faculty of Pharmaceutical Sciences.
d
Professor (orthodontics), Department of Pediatric Clinics, Preventive and
Social Dentistry, Faculty of Dentistry.
e
Pediatric dentist, Department of Pediatric Clinics, Preventive and Social
Dentistry, Faculty of Dentistry; doctoral student, Department of Pathology,
Faculty of Medicine.
f
Professor (pediatric dentistry), Department of Pediatric Clinics, Preventive and
Social Dentistry, Faculty of Dentistry.
Reprint requests to: Paulo Nelson-Filho. Departamento de Clínica Infantil,
Odontologia Preventiva e Social, Faculdade de Odontologia de Ribeirão Preto,
USP. Avenida do Café, S/Nº, Ribeirão Preto, SP, Brasil, 14040-904; e-mail,
nelson@forp.usp.br.
Submitted, January 2006; revised and accepted, April 2006.
0889-5406/$34.00
Copyright © 2008 by the American Association of Orthodontists.
doi:10.1016/j.ajodo.2006.04.054
S104
food particles and Streptococcus mutans; these changes
increase the levels of these microorganisms in saliva and
dental bioilm.1-3
It has been shown that placement of ixed orthodontic
appliances leads to continuous accumulation and
retention of bacterial bioilm, which might increase
the incidence of enamel demineralization4-7 and
gingival inlammation.8-10 According to Gorelick et
al,11 an increase in the number of white spot lesions
during ixed orthodontic appliance therapy occurs in
approximately 50% of patients. In spite of regular use
of luoridated dentifrices, enamel dissolution occurs
rapidly around brackets and orthodontic bands,12 and
white spot lesions can develop 1 month after placement
of the appliances.5,13
Preventive measures, including luoridated or chlorhexidine-based mouth-rinses as well as bonding the
brackets with luoride-containing composites or glass
ionomer cements, have been advised for orthodontic
patients to reduce cariogenic bioilm accumulation and
enamel demineralization.14
Magno et al
American Journal of Orthodontics and Dentofacial Orthopedics
Volume 133, Number 4, Supplement 1
The method of ligation of orthodontic archwires is
a relevant additional factor that accounts for dental bioilm retention. Currently, in the search for more practical and eficient orthodontic accessories, elastomeric
rings have been suggested as the materials of choice to
connect stainless steel archwires to brackets instead of
metallic ligatures.15,16 The recently introduced luoridereleasing elastomeric rings have been reported to reduce
dental bioilm formation and improve enamel remineralization in areas adjacent to the bracket base, which are
dificult to clean.17-19
Metafasix technology, recently developed by the
medical industry, was adopted by the orthodontic industry to improve the quality of elastomeric products.
This is a water-insoluble, hydrogel-polymer coating that
transforms the polyurethane-based elastomeric surface,
when moistened, into a highly smooth surface, thus providing low-friction products.
Studies in the medical ield have shown that this technology can signiicantly reduce bacterial adherence due
to a more lubricated and polished surface.20 In view of
this, a new elastomeric module system also manufactured
with Metafasix technology, Super Slick rings (TP Orthodontics, LaPorte, Ind), was recently introduced. These
new ligatures might be effective for controlling bacterial
colony-forming units, thus reducing the risk of iatrogenic
enamel demineralization during orthodontic therapy.
Because of the lack of data in the literature, the purpose of this study was to investigate in vivo the contamination of Super Slick elastomeric rings by S mutans by
using microbial cultures and scanning electron microscopy (SEM).
S105
Maxilla
Super Slick elastomeric rings
Conventional elastomeric rings
(teeth 14, 15, 16, and 17)
(teeth 24, 25, 26, and 27)
Right
Left
Conventional elastomeric rings
Super Slick elastomeric rings (teeth
side
(teeth 44, 45, 46, and 47)
side
34, 35, 36, and 37)
Mandible
Fig 1. Distribution of Super Slick and conventional elastomeric rings tied to the brackets bonded the patients’
teeth.
MATERIAL AND METHODS
Twenty middle-class patients of both sexes, aged
12 to 25 years, who were undergoing ixed orthodontic
appliance treatment were selected from the orthodontics postgraduate clinic at the Faculty of Dentistry of
Ribeirão Preto, University of São Paulo, Brazil. Patients
who were using antimicrobial mouthwashes, had any
systemic disease, had used antibiotics within the previous 3 months, or had brackets bonded with glass ionomer cements were excluded from the trial. This research
project was approved by the local Ethics in Research
Committee (process # 2004.1.750.58.7), and written informed consent was obtained from the patients or their
parents (or legal representatives).
In each volunteer, Super Slick elastomeric rings were
tied to brackets bonded to the maxillary premolars or
molars on the right side of the dental arch and mandibular
premolars or molars on the left side, for a total of 160
ligatures (Fig 1). Conventional elastomeric rings (size,
.120 inches; grey color; code 383-020; TP Orthodontics)
Fig 2. Metallic support used for hanging the
elastomeric rings during microbiologic procedures.
(control group) were tied to the contralateral teeth, with
the same design and also totaling 160 ligatures (Fig 1).
The volunteers were not asked to follow any speciic
dietary, toothbrushing, and mouth-rinsing protocols, but
they were asked to maintain the eating habits and the oral
hygiene measures recommended by the orthodontics
postgraduate clinic for patients with ixed orthodontic
appliances.
S106
Magno et al
American Journal of Orthodontics and Dentofacial Orthopedics
April 2008
Table. Positive cases according to the scores based on
the number of S mutans colonies or bioilms on the surfaces of Super Slick and conventional elastomeric rings
Score
0
1
2
3
4
5
Super Slick
0
2
1
6
2
64
Conventional
5
12
9
22
9
18
After 15 days in the mouth, the Super Slick and
conventional rings were aseptically and carefully
removed with a ligature applicator (code 75.01.002;
Dental Morelli, Sorocaba, SP, Brazil) by the same
professional (A.F.F.M.) who placed them and taken
to a metallic support for adequate stabilization (Fig 2).
Additional sticks with 24 Super Slick elastomeric rings
and 24 conventional elastomeric rings were removed from
their original packages and submitted to microbiologic
processing without use to conirm the absence of
S mutans contamination during ligature manufacturing
and packaging. This would ensure that the elastomeric
rings were mutans-free before being tested.
The metallic supports carrying the elastomeric rings
were individually and vertically placed into 25 × 115 mm
test tubes containing 25 mL bacitracin sucrose broth-selective enrichment broth for 3 to 4 days at 37°C. The culture medium was prepared according to the modiication
of Jensen and Bratthall21 and was speciic for S mutans
without trypan blue, according to Cesco et al.22 Care was
taken to prevent contact of the rings with the test tube
walls. The metallic supports were withdrawn and rinsed
in the broth with gentle shaking to remove planktonic
microbiota and leaving sessile bacteria adhered as spikes
or mushroom-like colonies or bioilms. The elastomeric
rings were carefully analyzed on all sides, and sessile S
mutans colonies or bioilms, based on colony morphology, were counted under aseptic conditions with a stereomicroscope (Nikon, Tokyo, Japan) with relected light.
The number of colonies on the surface of the elastomeric rings was expressed according to a ranked scale:
score 0, no colonies or bioilms, indicating no microorganisms on the ligature surface; scores 1 to 5, colonies
or bioilms on 5%, 25%, 50%, 75%, and 100% of the
ligature surfaces, respectively.
Conirmation that the adhering microorganisms
were S mutans was obtained by a sequence of steps: 4
or 5 colonies or bioilms representing the bacterial development were collected from 3 or 4 rings of each type
and transferred to tubes containing 2 mL of phosphate
buffered solution and glass beads; the colonies were
vortexed for 2 minutes; the resulting suspension was
seeded on SB20 agar (tryptone soy yeast agar plus 20%
sucrose and 0.2 U/mL bacitracin (Sigma, St Louis, Mo)
and incubated in microaerophilia at 37°C for 48 hours.
The SB20 agar medium, which is selective for S mutans,
was prepared according to the method of Davey and
Rogers23 and modiied by replacing sucrose with cane
sugar, as proposed by Torres et al.24 The formed colonies were submitted to the following tests: fermentation
of mannitol, sorbitol, rafinose, and melibiose; hydrolysis of arginine and esculin; production of hydrogen
peroxide; and sensitivity to 2.0 IU of bacitracin.25
A total of 6 elastomeric rings were analyzed by SEM:
1 ligature scored 1 and another scored 5 from each type
(obtained from those placed intraorally) and 2 unused
elastomeric rings (1 of each type) were taken directly
from the package. The specimens were processed for
SEM analysis, with the sequence proposed by Adriaens
et al26 to evaluate S mutans bioilm formation on their
surfaces. The elastomeric rings were submitted to critical point drying, mounted on stubs, sputter-coated with
gold under vacuum (Denton Vacuum Desk II, Mooretown, NJ) for 60 seconds, and examined with a SEM
(JSM 5410, JEOL, Tokyo, Japan) at 15 kV.
All microbiologic and SEM procedures were performed by an investigator (I.Y.I.) who was blinded to the
type of elastomeric ring. The researcher was also blinded
to the type of elastomeric ring when analyzing the data.
Statistical analysis
The results of colonies and bioilm counting were
analyzed statistically with the Wilcoxon nonparametric
test by using GraphPad Prism statistical software for
Windows (version 4.0, GraphPad Software, San Diego,
Calif) at the 5% signiicance level.
RESULTS
After 15 days of use, 75 Super Slick and 75 conventional elastomeric rings were collected from the 20
volunteers in the study, corresponding to 47% of the 160
elastomeric rings of each type initially tied to the patients’ brackets. Several specimens were lost due to detachment of the rings from the brackets and debonding
of brackets between appointments or problems during
laboratory procedures (some rings fell from the metallic support during introduction into the test tubes containing the culture medium and remained on the bottom
of the tube; these were discarded). However, the large
number of ligatures initially tied to the patient’s brackets (n = 160) made an adequate sample size possible in
spite of the high specimen loss rate.
The numbers of S mutans colonies or bioilms on the
surfaces of the Super Slick and the conventional elastomeric rings are shown in the Table.
Magno et al
American Journal of Orthodontics and Dentofacial Orthopedics
Volume 133, Number 4, Supplement 1
S107
Fig 3. S mutans colonies or biofilms on Super Slick elastomeric rings scored 1 to 5 (left to right) after 15 days in
the mouth and incubation in bacitracin sucrose brothselective enrichment broth selective for S mutans.
Fig 5. Scatter plot showing the distribution and the median of the scores referring to the number of S mutans
colonies or biofilms on Super Slick and conventional
elastomeric rings. There was a statistically significant
difference at P <.0001 (Wilcoxon test) between the
types of elastomeric rings.
Fig 4. S mutans colonies or biofilms on conventional
elastomeric rings scored 1 to 5 (left to right) after 15 days
in the mouth and incubation in bacitracin sucrose brothselective enrichment broth selective for S mutans.
When the microbiologic culture was positive,
S mutans bacterial bioilm was observed on both Super
Slick and conventional elastomeric rings on SEM analysis (Figs 6 and 7).
Fissures were found on the surface of the Super Slick
elastomeric rings after 15 days of intraoral placement
(Fig 6). No issures were observed on the unused Super
Slick rings taken directly from their original packages.
No issures were observed on the surface of the conventional elastomeric rings at any time.
DISCUSSION
S mutans were observed on all 75 Super Slick elastomeric rings. Eleven ligatures were scored 1 to 4 (15%),
and 64 ligatures were scored 5 (85%) (Fig 3).
Seventy conventional elastomeric rings were contaminated by S mutans. Of these, 52 ligatures were
scored 1 to 4 (74%), and 18 ligatures were scored 5
(26%) (Fig 4). Five elastomeric rings were not contaminated by S mutans.
Statistical analysis by the Wilcoxon nonparametric
test showed that the Super Slick elastomeric rings had
statistically signiicant greater S mutans contamination
than did the conventional elastomeric rings (P <.0001)
(Fig 5).
No formation of S mutans colonies or bioilms was
observed in the elastomeric rings removed directly from
their original packages
In view of current concepts of health promotion and
biosecurity, the adoption of strict measures for maintenance of oral health during orthodontic therapy has
become even more justiiable. Patients with systemic
pathologies also undergo orthodontic treatment and demand additional care because of the possibility of developing undesirable bacteremias or infections.
Orthodontic materials with modiied properties
that are claimed to provide greater eficacy in controlling bacterial development on their surfaces have been
introduced.
There has been interest in investigating the role of
materials used for ligation of orthodontic archwires on
the increase of bacterial contamination in the areas adjacent to the brackets.16,27,28
Forsberg et al16 observed greater microbial colonization
on teeth ligated to the archwire with elastomeric rings
S108
Magno et al
Fig 6. SEM micrograph of a Super Slick elastomeric ring
scored 5.
Fig 7. SEM micrograph of a conventional elastomeric
ring scored 5.
compared with teeth ligated with steel wires. On the
other hand, Sukontapatipark et al27 and Türkkahraman
et al28 found no signiicant differences between both
materials regarding microbial contamination. Brêtas et
al29 evaluated the inluence of topical application of 0.4%
stannous luoride gel on the development of bioilm and
S mutans colony-forming units on stainless steel ligatures
and elastomeric rings and found no difference between
those methods of orthodontic archwire ligation.
Several studies have investigated the performance
of luoride-releasing elastomers on decreasing both the
formation of S mutans colonies or bioilms and the susceptibility for development of carious lesions around
orthodontic brackets.18,19,30,31 Generally, the indings of
these studies have shown that luoride-releasing elastomeric rings were not effective for that purpose.
The Metafasix technology, used in the Super Slick ligatures, has brought improvements to the characteristics of
elastomeric rings by decreasing friction during orthodontic
movement. This is a result of the hydrogel-polymer coat-
American Journal of Orthodontics and Dentofacial Orthopedics
April 2008
ing that transforms the elastomeric surface into a smooth
sliding surface when moistened.20 As a consequence, less
accumulation of bacterial bioilm on the ligature surface
during orthodontic therapy is expected. Our results showed
that, after a 15-day intraoral period, S mutans colonies or
bioilms were observed on the entire surface of 85% of the
Super Slick elastomeric rings. The contamination of Super
Slick ligatures was signiicantly greater than that of conventional ligatures of the same brand.
Although the reason for these results is not clear, it can
be speculated that these indings are due to the formation
of issures on the surface of the elastomeric chain of Super
Slick rings when they are tied to the orthodontic brackets.
To be connected to the brackets, elastomeric rings, either
conventional or modiied, should be tractioned. It is possible that the polymer coating added to the elastomeric
surface with the Metafasix technology detached in some
areas during tractioning of the ligature, thus causing issures. We observed this during the SEM examination of
the Super Slick rings placed on the patients’ orthodontic
brackets. The formation of additional retentive sites with
the creation of issures might have contributed to increase
the accumulation of S mutans colonies or bioilms on the
surface of Super Slick elastomeric rings. The polymer
coating used to produce a more lubricated and smooth
surface was affected during tractioning of the elastomeric
ring for placement on the bracket, thus altering the quality of the elastomeric ligature surface. In view of these
results, the use of Metafasix technology for elastomeric
rings should be reevaluated because, in addition to increased bacterial contamination, the issures created on
polymer coating might increase friction, which can interfere with orthodontic movement.
Comparison of these results to those other studies
was impossible because there are no published studies
assessing bacterial contamination on the surface of conventional elastomeric rings and elastomers modiied by
Metafasix technology.
Our indings suggest that further investigations with
different scopes are needed to provide detailed information about the several commercial brands of conventional and modiied elastomeric rings available before they
can be advised for clinical use as orthodontic ligation
accessories with lesser bacterial bioilm formation.
CONCLUSIONS
According to our methodology and based on these
outcomes, the following conclusions can be drawn:
1. Super Slick elastomeric rings had signiicantly
greater contamination by S mutans than conventional
elastomeric rings.
2. There was no clinical evidence that Super Slick
elastomeric rings are effective in reducing bacterial bio-
American Journal of Orthodontics and Dentofacial Orthopedics
Volume 133, Number 4, Supplement 1
ilm formation on their surface, and a recommendation
for their use in orthodontic therapy for that purpose is
not justiiable.
REFERENCES
1. Balenseifen JW, Madonia JV. Study of dental plaque in orthodontic patients. J Dent Res 1970;49:320-4.
2. Scheie AA, Arneberg P, Krogstad O. Effect of orthodontic treatment on prevalence of streptococcus mutans in plaque and saliva.
Scand J Dent Res 1984;92:211-7.
3. Sinclair PM, Berry CW, Bennett CL, Israelson H. Changes in gingiva and gingival lora with bonding and banding. Angle Orthod
1987;57:271-8.
4. Mizrahi E. Enamel demineralization following orthodontic treatment. Am J Orthod 1982;82:62-7.
5. Ögaard B, Rølla G, Arends J. Orthodontic appliances and enamel
demineralization. Part 1. Lesion development. Am J Orthod Dentofacial Orthop 1988;94:68-73.
6. Ögaard B. Prevalence of white spot lesions in 19-years-olds: a
study on untreated and orthodontically treated persons 5 years
after treatment. Am J Orthod Dentofacial Orthop 1989;96:423-7.
7. Mitchell L. Decalciication during orthodontic treatment with
ixed appliances—an overview. Br J Orthod 1992;19:199-205.
8. Zachrisson S, Zachrisson BU. Gingival condition associated with
orthodontic treatment. Angle Orthod 1972;42:26-34.
9. Pender N. Aspects of oral health in orthodontic patients. Br J Orthod 1986;13:95-103.
10. Huser MC, Baehni PC, Lang R. Effects of orthodontic bands on
microbiologic and clinical parameters. Am J Orthod Dentofacial
Orthop 1990;97:213-8.
11. Gorelick L, Geiger AM, Gwinnett AJ. Incidence of white spot formation after bonding and banding. Am J Orthod 1982;81:93-8.
12. O’Reilly MM, Featherstone JDB. Demineralization and remineralization around orthodontic appliances: an in vivo study. Am J
Orthod Dentofacial Orthop 1987;92:33-40.
13. Glatz EGM, Featherstone JDB. Demineralization related to
orthodontic bands and brackets—a clinical study. Am J Orthod
1987;87:87.
14. Ögaard B, Rølla G, Arends J, ten Cate JM. Orthodontic appliances
and enamel demineralization. Part 2—prevention and treatment
of lesions. Am J Orthod Dentofacial Orthop 1988;94:123-8.
15. Echols MP. Elastic ligatures, binding forces and anchorage taxation. Am J Orthod Dentofac Orthop 1975;67:219-220.
16. Forsberg CM, Brattstrom V, Malmberg E, Nord CE. Ligature
wires and elastomeric rings: two methods of ligation, and their
association with microbial colonization of streptococcus mutans
and lactobacilli. Eur J Orthod 1991;13:416-20.
17. McNeill CJ, Wiltshire WA, Dawes C, Lavelle CLB. Fluoride
release from new light-cured orthodontic bonding agents. Am J
Orthod Dentofacial Orthop 2001;120:392-7.
Magno et al
S109
18. Mattick CR, Mitchell L, Chadwick SM, Wright J. Fluoride-releasing elastomeric modules reduce decalciication: a randomized
controlled trial. J Orthod 2001;28:217-9.
19. Doherty UB, Benson PE, Higham SM. Fluoride-releasing elastomeric ligatures assessed with the in situ caries model. Eur J
Orthod 2002;24:371-8.
20. MetafãsixTM technology. Assorted low friction products: products transformed by technology. Produzido por TP Orthodontics, Inc. [folder]. Available at: http://www.tportho.com/products/metafasix.html. Accessed on November 16, 2003.
21. Jensen B, Bratthall D. A new method for the estimation of mutans streptococci in human saliva. J Dent Res 1989;68:468-71.
22. Cesco RT, Bignelli P, Santos CP, Ito YI. Toothbrushes: evaluation of contamination level by streptococci of mutans group.
Proceedings of the 5th World Congress on Preventive Dentistry; 1995 Apr 27-30; São Paulo, Brazil. ABOP; 1995. p. 103.
23. Davey AL, Rogers AH. Multiple types of the bacterium streptococcus mutans in the human mouth and their intra family transmission. Arch Oral Biol 1984;29:453-60.
24. Torres AS, Pizzolitto AC, Ellias AM, Ito IY. Estreptococos do
grupo mutans: avaliação do agar SB20 e MSB na determinação
de ufc na saliva e na placa dental de adolescentes. Rev Bras
Odontol 1993;50:18-21.
25. Shklair IL, Keene J. A biochemical scheme for the separation of 5 varieties of Streptococcus mutans. Arch Oral Biol
1974;19:1079-81.
26. Adriaens PA, Seynhaeve TM, Be Boever JA. A morphologic
and SEM investigation of 58 toothbrushes. Clin Prev Dent
1985;7:8-16.
27. Sukontapatipark W, El-Agroudi MA, Selliseth NJ, Thunold K,
Selving K. Bacterial colonization associated with ixed orthodontic appliances. A scanning electron microscopy study. Eur
J Orthod 2001;23:475-84.
28. Türkkahraman H, Sayin MÖ, Bozkurt FY, Yetkin Z, Kaya S,
Önal S. Archwire ligation techniques, microbial colonization,
and periodontal status in orthodontically treated patients. Angle
Orthod 2005;75:231-6.
29. Brêtas SM, Macari S, Elias AM, Ito IY, Matsumoto MAN. Effect of 0.4% stannous luoride gel on streptococci mutans in
relation to elastomeric rings and steel ligatures in orthodontic
patients. Am J Orthod Dentofacial Orthop 2005;127:428-33.
30. Banks PA, Chadwick SM, Asher-Mcdade C, Wright JL. Fluoride-releasing elastomerics—a prospective controlled clinical
trial. Eur J Orthod 2000;22:401-7.
31. Benson PE, Ian Douglas CW, Martin MV. Fluoridated elastomers: effect on the microbiology of plaque. Am J Orthod
Dentofacial Orthop 2004;126:325-30.