S C IE N T IF IC

S E C T IO N

Journal o f Orthodontics, V o l. 41, 2014, 110-117

An assessment of the effectiveness of mechanical and

chemical cleaning of Essix orthodontic retainer
Chiew Sinn Chang1, Sarah Al-Awadi1, Derren Ready2 and Joseph Noar1
O rth o d o n tic D e p a rtm e n t, Eastman D e n ta l In s titu te , U niversity C ollege London, Lo ndon, UK; 2M ic ro b io lo g y D e p a rtm e n t, Eastman D ental
In s titu te , U niversity C ollege Lo nd on , Lo ndon, UK

Objectives: To determine the effectiveness of mechanical and chemical cleaning on the removal of microorganisms
from Essix orthodontic retainers. Design: In vitro laboratory study. Setting: Department of Orthodontics and
Microbiology, Eastman Dental Institute, University College London, UK. Methods: Study 1: 120 Essix retainers were
divided into four cleaning groups. The effectiveness of each cleaning method to remove a single species biofilm of
Streptococcus mutans from the retainer was assessed. Study 2: 140 Essix retainers were divided into four study
groups (brushing w ith fluoride toothpaste, chlorhexidine gel, immersion in chlorhexidine solution only and a
control) to investigate the chemical and mechanical cleaning of the multispecies biolfilm of (Streptococcus sanguis,
Actinomyces naeslundii, methicillin-resistant Staphylococcus aureus and Candida albicans). Relevant results: In
study 1, brushing with toothpaste resulted in 99% reduction of Streptococcus mutans. In study 2, all three cleaning
methods recorded similarly statistically significant reductions in colony forming units per millilitre compared to the
control. There were no statistically significant differences between any of the cleaning groups for any of the
microorganisms except MRSA-16. For MRSA-16, chlorhexidine mouthwash and gel were significantly more potent in
eliminating the microorganism than the fluoride toothpaste. Conclusions: All three cleaning methods effectively
removed 99% of microorganisms from the Essix retainers. Brushing w ith fluoride toothpaste can therefore be
confirmed as an effective method for cleaning retainers in most circumstances. The use of chlorhexidine gel or
mouthwash is recommended in patients where bacterial infection has to be avoided due to immunosuppression or
other reasons.
Key words: Biofilm, chlorhexidine, cleaning, thermoplastic retainers, tooth brushing
Received 7 A pril2013: accepted 12 November 2013

In tro d u c tio n

The Essix orthodontic retainer is the most popular form of

post-orthodontic retention in the UK (Hichens et al,
2007). Its ease of fabrication and good aesthetics means
that it is cost effective and has increased patient
compliance. Orthodontists generally advise their patients
to wear their retainers long-term to prevent relapse, which
can be associated with a tendency to build-up of bacteria
on the fitting surface. Indeed, opportunistic pathogens,
such as Candida albicans and methicillin-resistant
Staphylococcus aureus (MRSA), can be found on retainers,
which can theoretically lead to local or systemic infection
(Al Groosh et al., 2011). A cleaning regimen to remove a
biofilm from the retainer can be either mechanical or
chemical, or a combination of both approaches.
Mechanical cleaning can be achieved by brushing alone,
or using an additional compound such as soap, dentifrices
or by using an ultrasonic device. Chemical cleaning is
Address for correspondence: J. Noar, Orthodontic Unit, Division
of Craniofacial & Development Sciences, Eastman Dental Hospital/
Institute, 256 Grays Inn Road, London WC1X 8LD, UK.
Email: J.Noar@ucl.ac.uk
2014 British Orthodontic Society

usually achieved by soaking the retainer in an antimicro

bial agent such as chlorhexidine (Nikawa et al., 1999). To
date, no studies have investigated the efficacy of mechan
ical or chemical removal of multi-species biofilms from
Essix orthodontic retainers. In order to consider the best
cleaning methods, we have considered relevant studies
assessing the efficacy of mechanical and chemical removal
of the biofilm from removable prostheses such as dentures
and removable acrylic orthodontic appliances.
Tooth brushing with a dentifrice or tap water is the most
common method of mechanical cleaning and this method
has been shown to be simple and cost effective (Smith,
1966; Neill, 1968; Peracini et al., 2010). Brushing with
dentifrices can be considered as a combination method as
most dentifrices contain antimicrobials to enhance the
brushing effect (Tarbet et al., 1984; Kulak-Ozkan et al.,
2002; Dikbas et al., 2006). Peixoto et al. (2011) investi
gated the efficacy of 0.12% chlorhexidine gluconate spray

DOI 10.1179/1465313313Y.0000000088

JO June 2014

Scientific Section

(Periogard) to remove Streptococcus mutans from acrylic

baseplates of removable orthodontic appliances and they
showed that this method significantly reduced the
bacterial count when used either once or twice a week.
The aim of the present study was to assess the
effectiveness of mechanical and chemical cleaning in
eliminating single and multi-species microorganisms on
Essix orthodontic retainers.
Study 1

The sample size was calculated using nQuery Advisor

software. The data used for the calculation were taken
from the prosthodontics literature. It was calculated that
a total sample size of 30 retainers per group would give a
power of 90% to detect a difference in means of 1.000,
assuming that the common standard deviation is 1.140
using a two-sample f-test with a 0.050 two-sided
significant level.
M a te ria ls a nd m e th o d s f o r s tu d y 1

This was a laboratory based in vitro study carried out in

the Departments of Orthodontics and Microbiology at
the Eastman Dental Institute.
A total of 120 pressure-formed retainers were fabri
cated from Essix ACE (Dentsply) thermoplastic
copolyester sheets with a thickness of 0.030 inch by
the same operator on duplicates of a standard maxillary
cast.
All retainers were disinfected by immersion in 1%
NaOCl (sodium hypochlorite) for 30 min. After this
time, the retainers were removed with sterile forceps and
placed in the neutralizing broth for 30 min, before being
cleaned by three rinses in sterile distilled water.
A plaque-forming solution which consisted of 15 ml of
sterile sucrose, 30 ml of Brain heart infusion broth and
5 ml of Streptococcus mutans (NCTC 10449) culture was
prepared. The retainers were then placed into this
solution and incubated for 72 h in air supplemented
with 5% carbon dioxide. At the end of the 72-h period,
retainers were taken at random and assigned to their
respective group for investigation. Each retainer was
tested once with each cleaning method.
1.

2.
3.
4.

Group 1: Brushing with a toothbrush (Colgate 360

whole-mouth clean medium toothbrush) and tooth
paste (Colgate Total toothpaste) for 30 s. A mean
amount of 0.54 g of toothpaste was used;
Group 2: Brushing with sterile distilled water for
30 s;
Group 3: Rinsing with 50 ml sterile distilled water
for 30 s;
Group 4: No cleaning (control group).

Methods of cleaning Essix orthodontic retainer

111

For all groups, the retainers were swabbed using calcium

alginate swab after the regimen and was placed immedi
ately into 10 ml of calgon ringers solution (Oxoid Ltd) in
a sterile bijou tube (Starstedt, Leicester, UK) containing
three sterile glass beads. The sample was vortex-mixed for
1 min to dissolve the alginate swab before processing.
Spread-plating of each sample was carried out. Ten-fold
serial dilutions were prepared in phosphate buffered saline
(Oxoid Ltd) to a dilution 10-6. Samples from the 10~2,
10-3, 10~4 and 10-5 dilutions were plated onto plates of
Columbia blood agar (CBA; Oxoid Ltd) containing 5%
defibrinated horse blood (E&O Laboratories, Bonnybridge, UK) to determine the total number of cultivable
bacteria present in the specimen. The CBA plates were
incubated for 2 days in air supplemented with 5% carbon
dioxide.
After incubation, the plates were placed on a commer
cial colony counter with an illuminated background and
a x 5 magnifying lens. The colonies were manually
counted and recorded. The total viable number of
bacteria was determined by multiplying the number of
colonies, by the dilution factor.
A single species microorganism does not represent the
oral condition, which is comprised of multi-species
microorganisms. In order to address this and based on
the data from the single species study, study 2 was
carried out.
Study 2

The nQuery Advisor software (version 4) was used for

sample size calculation. The data used for the calculation
was taken from study 1. It was calculated that a total
sample size of 30 retainers per group would give a
power of 95% to detect a difference in means of 1.000
log difference of bacteria, assuming that the common
standard deviation is 1.050 using a two sample t-test. The
statistical significance level was set at P< 0.05. Owing to
the concern of the normality of the data, we decided to
estimate the sample size using non-parametric statistics
inflated the sample size by 15% (Lehmann, 2006). It was
therefore decided that, 35 retainers would be fabricated
for each test or control group.
M a te ria ls and m e th o d s f o r s tu d y 2

A total of 140 Essix retainers made from the same

thermoplastic copolyester sheets with a thickness of
0.030 inch were fabricated in the orthodontic laboratory
by the same operator on the standard maxillary casts.
Four strains of microorganism Methicillin-resistant
Staphylococcus aureus-16 (MRSA-16), Streptococcus san
guinis, Candida albicans and Actinomyces naeslundii were
grown on Columbia blood agar (Oxoid) individually to

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Chang et al.

Scientific Section

produce pure cultures. A fresh colony of each micro

organism was inoculated into 10 ml of artificial saliva
(Coulter and Russell, 1976) at the temperature of 37C for
24 h. The microorganism concentration was determined
by optical density (OD) using a spectrophotometer. The
growth curve of each microorganism was observed for 6 h
and the OD was recorded hourly. Spread plating of each
sample on CBA was carried out every 2 h.
The Essix orthodontic retainers were divided into
seven batches. Each batch consisted of 20 retainers and
the retainers were divided into four groups. The
retainers were disinfected by immersion in 1% sodium
hypochlorite (NaOCl) for 10 min before the inoculation.
After this, the retainers were then removed from the
NaOCl solution with sterile forceps and placed in the
neutralizing broth for 10 min.
The retainers were placed into a sterile plastic
container containing 300 ml of artificial saliva for 2 h
at 37C to precipitate a layer of salivary protein on the
retainer material to act as a substrate pellicle for the
attachment and growth of plaque. The retainers were
first inoculated with 1 ml of Streptococcus sanguinis
(OD^oo 0.4 + 0.02) and 1 ml of Actinomyces naeslundii
(OD6oo 0.4 + 0.02). After 24 h, 1 ml of Candida albicans
(OD600 0.4 + 0.02) and 1 ml of MRSA-16 (OD600
0. 05.+ 0.005) were added. The Candida albicans and
MRSA-16 were added later as they have a faster growth
rate compare to Streptococcus sanguinis and Actinomyces
naeslundii.
The inoculated retainers were then incubated for a
further 24 h at 37C in an anaerobic cabinet. At the end
of this, the retainers were removed from the culture and
assigned to each of the respective cleaning regimes under
investigation.
1.

2.

3.

4.

Group 1: Brushing with a toothbrush (Colgate 360

whole-mouth clean medium toothbrush) and tooth
paste (Colgate cavity protection fluoride tooth
paste) for 30 s;
Group 2: Brushing with a toothbrush (Colgate 360
whole-mouth clean medium toothbrush) and chlorhexidine gluconate gel (Corsodyl Dental Gel) for
30 s;
Group 3: Immersion in chlorhexidine solution
(Corsodyl Alcohol Free Mint Mouthwash) for
10 min.
Group 4: No cleaning (control group); immersion
in phosphate-buffered saline for 10 min.

The retainers were removed from the culture by using a

sterile forcep and placed onto a petri dish. Each retainer
was then brushed with toothpaste (Group 1) or gel
(Group 2) for 30 seconds and then rinsed with sterile
distilled water. A new toothbrush was used to clean each

JO June 2014

retainer. Approximately 0.55 g (range: 0.50-0.59 g) of

toothpaste or gel was used for each procedure.
All the retainers that were soaked in the chlorhexidine
(Group 3) or phosphate-buffered saline (Group 4) for
10 min were then rinsed with sterile distilled water. For
all groups, once they had been cleaned, the whole
retainer was swabbed with a cotton swab for 30 s. The
swab was then placed into 2 ml of neutralizing broth
solution in a sterile tube. The sample was vortexed for
30 s to dissolve the bacteria from the swab before
processing. A serial dilution process was carried out to
determine the countable numbers of bacteria colonies on
the agar plates using a multichannel pipetting tray. Ten
fold serial dilutions of the sample from dilution factor
10 1 to 10-4 were prepared in neutralizing broth. The
dilutions were transferred onto the agar plates using a
pipette and spread uniformly using a sterile spreader.
Once this had been done, the agar plates were incubated
for 48 h at the temperature of 37C in an anaerobic
cabinet. After incubation for 48 h, all the agar plates
were examined and the colonies were manually counted
and recorded. The total viable number of bacteria was
determined by multiplying the number of colonies and
the dilution factor. An overview of the method is shown
in Figure 1.
Statistical methods
Data was analysed statistically with Kruskal-Wallis test
to compare the differences in the median values of
colony forming units (cfu) between groups. Figure 2
shows the numbers of viable bacteria present (cfu/ml) on
retainers after exposure to three different mechanical
cleansing methods.

Results

Study 1
Of the 120 retainers, four were excluded from brushing
with toothpaste and another four from rinsing with
sterilized distilled water as they were partially contami
nated and additional readings were added to the control
and brushing alone from the extra measurements that
were undertaken during the study.
The data show that using different hygiene methods
promotes changes in the microbial load present on the
surface of the retainer. Rinsing with sterile distilled water
was shown to be less efficacious in removing Streptococcus
mutans than the other methods and brushing with
toothpaste the most effective. On average, 5.25 x 10234cfu/
ml of Streptococuus mutans remained after brushing with
toothpaste, compared to brushing alone (5.7 x 105) and
rinsing with sterile distilled water (8.37 x 106).

JO June 2014

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Methods of cleaning Essix orthodontic retainer

113

Figure 1 The flo w ch a rt o f th e m ethod

Brushing with a toothbrush and toothpaste for 30 s

resulted in 99.9% reduction in the microbial load. In
decreasing order of effectiveness, brushing with a tooth
brush alone produced a 92.8% microbial reduction
followed by rinsing with sterile distilled water, which
resulted in only 19.61% mirobial reduction. The mean,
median, minimum, maximum and standard deviation of
Streptococcus mutans counts found on the internal
surface of the evaluated Essix retainers are presented in
Table 1. The data were compared using Kruskal Wallis

test to detect significant differences among the groups

CP<0.05) (Table 2). An inspection of the mean ranks for
the groups shown in Table 2 suggests that rinsing with
sterile distilled water had the highest numbers of bacterial
counts, with brushing with toothpaste reporting the
lowest.
In summary, the results showed that brushing with
toothpaste resulted in 99.9% reduction of Streptococcus
mutans. Brushing with distilled water was less effective
than combining tooth brushing with toothpaste. There

Percentage of the microorganisms removed from the retainer after

cleaning

Brushing with toothpaste

Brushing with distilled water

Rinsing with distilled water

C le a n in g m e t h o d s

Figure 2

Percentage o f m icroorganism s rem oved a fte r exposure to th re e d iffe re n t cleaning m ethods

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Chang et al.

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Table 1 The mean, median, minimum, maximum and standard deviation o f Streptococcus m utans counts after
exposure to three d iffe re n t mechanical cleansing method
Method

A/= 120

Mean

Minimum

Maximum

Standard deviation

Median

Control
Brushing
Brushing with toothpaste
Rinsing

35
33
26
26

1.35 x 107
9.79 x 1 0 5
1.09 x 1 0 3
1.08 x 1 0 7

8.0 x 1 0 5
8.8 x 104
1.3 x 102
5.0 x 102

6.3 x 1 0 7
5.4 x 106
3.4 x 1 0 3
5.0 xIO 7

1.56 x 107
1.06 x 106
1.02 x 103
1.36 x 1 0 7

5.25 x 1 0 6
5.7 x 10s
5.25 x 1 0 2
8.37 x 106

was no statistically significant difference between the

control group and rinsing with sterile distilled water.
This suggests that it is not effective in biofilm removal.

Study 2
The three cleaning regimes resulted in reduction of
99.9% of the microbial load except MRSA-16 (brushing
with fluoride toothpaste) which showed 99.8% of
microbial reduction. 100% of Candida albicans were
successfully removed from the retainer no matter what
cleaning methods used (Figure 3).
Box plots were used to look at the distribution of the
microorganism present on each Essix orthodontic
retainer after different cleaning regimens. All of the
distribution of the data was skewed to the right. As the
data were not normally distributed, the non-parametric
Kruskal-Wallis test was performed.
Kruskal-Wallis test showed statistically significant
differences between the groups (P<0.001). It indicated
that at least two groups differed from each other, but did
not identify the specific groups that differ. Therefore, the
results were analysed further using the pairwise compar
ison test to identify the pattern of differences in the
results.
Pairwise comparison of groups demonstrated that all
three cleaning methods were statistically significant
different (P<0.001) compared with the control. There
were no statistically differences, however, between the
three test cleaning groups (Table 3).
For MRSA-16, all of the cleaning groups showed stati
stically significant differences (P<0.001) except between
the cleaning group using chlorhexidine mouthwash and
chlorhexidine gel (Table 4).
Table 2 Kruskal-Wallis test to compare the differences
in the median values o f colony form ing units between
the groups

D is c u s s io n

In study 1, a single species was used to test the proposed

method and confirm that it was the best approach to
carry out this project with multi-species microorganisms
which represent the oral condition better. Actinomyces
naeslundii and Streptococcus sanguis were chosen
because they are commonly present in the oral cavity
and they are easily cultured in the laboratory. Candida
albicans and MRSA-16 are not commonly found on
orthodontic appliances. However, they are opportunis
tic microorganisms which can be dangerous to the
susceptible host and have been found on orthodontic
appliances and denture surfaces in previous studies.
Human saliva would be the ideal medium of selection for
the growth of the bacteria. However, large quantities of
saliva medium are required for this study and human
saliva composition, viscosity and levels of electrolytes and
enzymes vary between individuals (Bjorklund, 2011). In
addition, the quantities and types of bacteria contained in
the saliva will vary between different individuals and
therefore, it would not have been possible to provide a
standardized approach to this study, which would
introduce bias into the results. Owing to this, artificial
saliva was chosen as a substitute in this study. The cleaning
agents used were Corsodyl mouthwash (0.2% chlorhex
idine gluconate), Corsodyl dental gel (T.0% w/w chlorhex
idine gluconate) and regular Colgate cavity protection
toothpaste (0.76% sodium monofluorophosphate). The
amount of the cleaning agents used for each group was
controlled to ensure the consistency of the results.
To the best of our knowledge, there are no studies in
the literature that have examined the effectiveness of
mechanical and chemical cleaning of the Essix ortho
dontic retainer, although investigations relating to
debonding burs have been previously carried out in
vitro (Sheriteh et al., 2010). The results in this study
using multi-species microorganisms demonstrated that
all the cleaning methods were effective in removing at
least 99.8% of the microorganisms from the Essix
orthodontic retainers. This finding was similar to the
results from the single species study.
In study 2, both types of toothpastes (fluoride based
and chlorhexidine based) were equally good for removal

JO June 2014

Scientific Section

Methods of cleaning Essix orthodontic retainer

115

% of microorganism s rem oved fro m retain er a fte r 3 differen t regimes

Actinom yces
naeslundii

Streptococcus
sanguinis

MRSA-16

Candida albicans Total viable count

Microorganism

Soaking into chlorhexidine mouthwash

Brushing with fluoride toothpaste
Brushing with chlorhexidine gel
Figure 3 Percentage o f th e m icroorganism rem oved fro m th e re ta in e r a fte r cleaning

of the biofilm, despite being very different. This might

be due to the fluoride toothpaste containing abrasive
particles (silica), which permitted a more effective
mechanical cleaning action, while the chlorhexidine-based
dental gel has the antimicrobial agent, chlorhexidine
gluconate, which promoted better chemical removal of the
microorganisms. Since there was no significant differences
between either toothpastes, fluoride toothpaste should be
advised for cleaning the retainers because most of the
people use fluoride toothpaste for brushing their teeth and
it is more sensible to use the same toothpaste to clean the

retainers. There were, however, statistically significant

differences between using a fluoride based toothpaste and
a chlorhexidine toothpaste or mouthwash for the removal
of MRSA-16. Chlorhexidine gel and mouthwash was
much more effective in reducing the colony forming units
for MRSA-16 compared with the toothpaste. MRSA-16
although rarely found on retainers or dentures (Tawara
et al., 1996; A1 Groosh et al., 2011) is a dangerous
microorganism, which exhibits resistance towards most
antimicrobial agents. As fluoride toothpaste used in this
study does not contain any specific antimicrobial agent,
we know that microorganisms have been removed from

Table 3

Pairwise com parison o f d iffe re n t cleaning

g ro u p fo r rem oving Actinomyces naeslundii, Candida
albicans and Streptococcus sanguinis fro m th e retainers

Table 4 Pairwise com parison o f d iffe re n t cleaning

g ro u p fo r rem oving MRSA-16 fro m th e retainers

Group 1-Group 2

Adj. Sig. (P)

Group 1-Group 2

Adj. Sig. (P)

CHX gel-CHX mouthwash

CHX g e l-flu o rid e toothpaste
CHX m o u th w a sh -flu o rid e toothpaste
CHX g e l-control
CHX m outhw ash-control
Fluoride too th p a ste -co n tro l

>0.999
>0.999
>0.999
<0.001
<0.001
<0.001

CHX gel-CHX mouthwash

CHX g e l-flu o rid e toothpaste
CHX m o uthw ash -fluo ride toothpaste
CHX g e l-control
CHX m outhw ash-control
Fluoride too thpa ste-co ntro l

>0.999
<0.001
<0.001
<0.001
<0.001
<0.001

116

Chang et at.

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the retainers mainly by the mechanical effort. In normal

healthy individuals, it would appear that it does not
matter if normal toothpaste is used for cleaning the
retainer as toothpaste alone removes almost all micro
organisms. However, in critical conditions such as an
immunocompromised patient, a patient taking immuno
suppressant drugs or a patient in hospital, the use of a
chlorhexidine based gel or mouthwash in cleaning the
retainer should be advised because even single bacteria
might cause a life-threatening condition to the patient.
Our findings showed that chemical cleaning by
soaking the Essix retainers into 0.2% of chlorhexidine
gluconate mouthwash for 10 min showed no significant
difference in the reduction of the colony-forming units
compared with brushing with 1.0% (w/w) of chlorhex
idine dental gel. Surprisingly, the use of chlorhexidine
mouthwash with less concentration of chlorhexidine and
without and mechanical scrubbing of the retainer has
the same percentage of microorganisms left on the
retainer compared with the use of higher concentration
of chlorhexidine and mechanical cleaning in brushing
group. A possible explanation of this might be that
chlorhexidine is a very powerful antimicrobial agent,
which can easily kill the microorganisms at low
concentrations even without mechanical cleaning.
Corsodyl, which contains 0.2% chlorhexidine gluco
nate, may alter the properties of the Essix retainer and
compromise its lifespan. The changes of the properties
of the retainer might cause distortion, cracks or even
fractures and thus, compromise the tooth position.
These surface imperfections might not only weaken the
retainer but also aid the retention and adherence of the
microorganisms. Budtz-Jorgensen (1979) showed that
daily soaking of the dentures into 1% or 2% of
chlorhexidine gluconate causes staining of acrylic
dentures. For these reasons, chlorhexidine-based anti
microbial agents are not recommended for daily use.
Future work to investigate the effect of different
concentrations of the chemical solution on the proper
ties of retainers would be a good idea for those patients
where removal of all microorganisms is important.
It should be highlighted that all three cleaning
methods were good in removing most of the micro
organisms from the retainers, but there were still
microorganisms left on the retainer, which could be
dangerous to a susceptible host. Therefore, these
methods cannot be described as completely effective
methods of removing microorganisms. At present,
orthodontists and general dentists recommend various
approaches of cleaning regimes for retainers to patients
without any scientific evidence. Some clinicians might
not agree with the recommended approaches, because
they believed that brushing the retainer with toothpaste

JO June 2014

might scratch the surfaces of the retainer and lead to an

increase surface area which allows further microbial
build-up. Another reason would be that brushing might
deteriorate the cosmetic appearance of the retainer and
affect the longevity of the retainer. To date, limited
studies had been conducted to look into these matters.
It is clear that work is required to investigate the
effects of abrading the surface, but until that work is
done, it would seem better to err on the side of caution
and use the most effective cleaning regime available.

C o n c lu s io n s

All three cleaning methods removed 99% of micro

organisms from the Essix orthodontic retainers.
Chlorhexidine gel or mouthwash was more effective in
eliminating MRSA-16 compared with fluoride tooth
paste.
Brushing with fluoride toothpaste can be employed as
an efficacious method for cleaning Essix orthodontic
retainers in most circumstances.
Chlorhexidine gel or mouthwash should be used as a
supplement to tooth brushing if there is any concern
that infection caused by intraoral opportunistic
microorganisms would compromise the patients
health.

A c k n o w le d g e m e n ts

The authors thank Mr Dheaa Al-Groosh for helping

with the microbiology culturing and Ms Aviva Petrie for
providing all the statistical support for this study.

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128.

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