22124403_s2212440312x00257_s2212440313004586_main

8102019 22124403_S2212440312X00257_S2212440313004586_main

httpslidepdfcomreaderfull22124403s2212440312x00257s2212440313004586main 16

Association between glycemic status and oral Candida carriagein patients with prediabetesFawad Javed BDS PhDa Hameeda Bashir Ahmed BDS CORTHb Abid Mehmood BDS MCPS FACSc

Anwar Saeed BDS MPHd Khalid Al-Hezaimi BDS MSc FRCD(C)ae and

Lakshman P Samaranayake BDS DDS FRCPathf

King Saud University and Al-Farabi Dental College Riyadh Saudi Arabia Jinnah Postgraduate Medical Center and Liaquat College of Medicine

and Dentistry Karachi Pakistan and Prince Philip Dental Hospital University of Hong Kong Hong Kong

Objective This study assessed the association between glycemic status and oral Candida carriage among patients with

prediabetes

Study Design This was a comparative study of oral Candida carriage among individuals with prediabetes Oral yeast samples

were collected from 150 individuals group A was 43 patients with prediabetes (fasting blood glucose levels and hemoglobin

A1c 100 to 125 mgdL and 5 respectively) group B was 37 individuals previously considered prediabetic but having

fasting blood glucose levels lt100 mgdL and hemoglobin A1c lt5 and group C was 70 medically healthy individuals Oral

yeasts were identified using standard techniques Unstimulated whole salivary flow rate and number of missing teeth were

recorded

Results Oral Candida was isolated from 100 of patients with prediabetes and from 657 of control participants Candida

albicans carriage was higher among patients with prediabetes (487) (P lt 01) and patients in group A (512) (P lt 01) than

among controls (257) Candida carriage unstimulated whole salivary flow rate and number of missing teeth were similar ingroups A and B

Conclusions Oral Candida carriage was higher in patients with prediabetes than in controls and was independent of glycemic

status in patients with prediabetes (Oral Surg Oral Med Oral Pathol Oral Radiol 201411753-58)

It is well known that immunosuppression in1047298uences

oral Candida carriage1-4 Studies1-3 have reported that

oral carriage of Candida species predominantly

Candida albicans (C albicans) is higher in patients

with poorly controlled diabetes compared with healthy

controls An explanation may be derived from the fact

that xerostomia (due to reduced unstimulated wholesalivary 1047298ow rate [UWSFR]) in patients with poorly

controlled prediabetes provides a platform for Candida

stagnation and growth on oral tissues primarily the

dorsum of the tongue15 In addition a high prevalence

of Candida species has also been reported in peri-

odontal sites among patients with prediabetes wit h

chronic periodontitis compared with healthy controls6

It is pertinent to mention however that previous

studies1256 in which oral Candida carriage was

investigated in hyperglycemic patients were performed

in patients with poorly controlled diabetes mellitusPrediabetes a state of impaired glucose tolerance

(IGT) is characterized by IGT (140 to 199 mgdL)

impaired fasting glucose (100 to 125 mgdL) or both7

In addition a hemoglobin A1c (HbA1c) test is a useful

diagnostic test that correlates with t he average blood

glucose levels over the past 3 months8 Individuals with

HbA1c levels between 55 and 64 are categorized

as individuals with prediabetes8

Periodontal in1047298ammation has been reported to be

worse in patients with prediabetes compared with

healthy controls9-12 it has been hypothesized that

The authors thank the College of Dentistry Research Center and

Deanship of Scienti1047297c Research at King Saud University Saudi

Arabia for funding this research project (Project FR 0072)a Engineer Abdullah Bugshan Research Chair for Growth Factors and

Bone Regeneration 3D Imaging and Biomechanical Laboratory

College of Applied Medical Sciences King Saud University Riyadh

Saudi Arabiab

Department of Dentistry Al-Farabi Dental College Riyadh SaudiArabiacDepartment of Dentistry Jinnah Postgraduate Medical Center Kar-

achi PakistandDepartment of Public Health Dentistry Liaquat College of Medicine

and Dentistry Karachi PakistaneDepartment of Periodontics and Community Dentistry College of

Dentistry King Saud University Riyadh Saudi Arabif Oral Biosciences Faculty of Dentistry Prince Philip Dental Hospital

University of Hong Kong Hong Kong

Received for publication Jul 8 2013 returned for revision Aug 16

2013 accepted for publication Aug 24 2013

2014 Elsevier Inc All rights reserved

2212-4403$ - see front matter

httpdxdoiorg101016joooo201308018

Statement of Clinical Relevance

Oral Candida carriage is similar in patients with

prediabetes and in individuals previously diagnosed

with prediabetes but having normal blood glucose

levels due to glycemic control This indicates that

individuals in either group are equally susceptible to

oral Candida infections and that their susceptibility

is higher than that of controls

53

Vol 117 No 1 January 2014

8102019 22124403_S2212440312X00257_S2212440313004586_main


besides the oxidative stress induced by chronic hyper-

glycemia a reduced UWSFR in patients with prediabetes

may also have contributed in worsening periodontal

status Because xerostomia is a com mon manifestation in

patients with chronic hyperglycemia15 it is assumed that

oral Candida carriage is also high in patients with

prediabetes compared with healthy controls However toour knowledge from indexed literature oral Candida

carriage in patients with prediabetes has not yet been

investigated Furthermore studies61314 have shown that

glycemic control reduces the severity of periodontal

in1047298ammation in patients with poorly controlled diabetes

and prediabetes We therefore hypothesized that oral

Candida carriage would be reduced in patients previ-

ously diagnosed with prediabetes but who maintain their

fasting blood glucose level (FBGL) within normal limits

(70 to lt100 mgdL) as compared with patients with

poorly controlled prediabetes (100 to 125 mgdL) To

our knowledge from indexed literature this hypothesishad not been tested before our study

METHODSEthical approvalThe study was approved by the research ethics reviewcommittee of the Jinnah Postgraduate Medical Center

Karachi Pakistan The study was performed in accor-

dance with the Declaration of Helsinki as revised in

2000 It was mandatory for all study participants to

have read and signed the consent form before being

included in this study

Inclusion and exclusion criteriaOnly individuals with medically diagnosed prediabetes

(FBGL 100-125 mgdL [56-69 mmolL] hemoglobin

A1c [HbA1c] 57-64) were included Exclusion

criteria wer e (1) tobacco smoking13 (2) alcohol

consum ption15 (3) exclusive areca nut and gutka

chewing1617 (4) use of antibiotics antifungal agents

steroids or nonsteroidal anti-in1047298ammatory drugs within

the past 3 months131819 (5) self-reported systemic

diseases including type 1 and type 2 diabetes mellitus

hepat itis B hepatitis C and infection with HIV or

AIDS1132021

and (6) wearing partial or completedentures2223

Study participantsPatients with prediabetes were recruited from the dia-

betes care unit of a local hospital in Karachi Pakistan

Medical records of these patients were explored to

con1047297rm the diagnosis of prediabetes Control participants

self-reporting as not prediabetic were recruited from

a residential area near the hospital All participants were

invited to an oral health care center in the early morning

hours (in a fasting state) for FBGL measurement and

collection of oral yeast and unstimulated whole saliva

(UWS) samples

Hemoglobin A1c levels and fasting blood glucoselevelsHospital records of patients with prediabetes were

searched to determine the most recent HbA1c levels A

digital glucometer (Accu-Chek Activ Roche Diagnos-

tics Mannheim Germany) was used to measure the

FBGL Depending on the glycemic levels patients with

prediabetes were divided into 2 subgroups as follows

group A patients with prediabetes with FBGL between

100 and 125 mgdL (HbA1c 5) and group B

patients with prediabetes with FBGL lt100 mgdL

(HbA1c lt5) Self-reported systemically healthyindividuals (FBGL 70 to lt100 mgdL) were catego-

rized as controls (group C)

Collection of UWS samplesTo collect the UWS samples participants were seated

comfortably in a chair in a ldquocoachmanrdquo position and

requested to spit (without swallowing) into a gauged

measuring cylinder for 1047297ve continuous minutes UWSFR

was recorded in milliliters per minute (mLmin)

Collection of oral yeast samplesOral Cand ida samples were collected as described

previously1 In summary each sample was collected by

scraping the dorsum of the tongue and bilateral buccal

mucosa with a sterile cotton swab (bioMeacuterieux SAMontalieu-Vercieu France)1 Immediately after

sampling the swabs were returned to the containment

tube to avoid contamination At 37C Candida strains

were cultured in Sabouraud dextrose agar (Becton

Dickinson and Company Sparks MD USA) to quan-

tify the colony-forming unit s in the or al ca vities of

individuals with and without prediabetes2425 After 24

hours all cultures were inspected and monitoringcontinued until 7 days of incubation for yeast growth

following which they were subjected to speciation

Identification of oral yeast samplesA yeast identi1047297cation system (API 32-C System yeast

identi1047297cation programme bioMeacuterieux) was used to

identify the oral yeast species1 Yeast isolates that could

not be identi1047297ed with the oral yeast identi1047297cation

system were subjected to molecular identi1047297cation

Molecular identi1047297cation was performed as described

elsewhere26 Brie1047298y for DNA isolation yeast cells

were suspended in 200 mL sterile polymerase chain

reaction (PCR)egrade water and genomic DNA was

prepared using a DNA preparation robot (Roche

Diagnostics GmbH Mannheim Germany)27 Using

ORAL MEDICINE OOOO

54 Javed et al January 2014

8102019 22124403_S2212440312X00257_S2212440313004586_main


universal primers and ampliTaq Gold DNA polymerase

for DNA sequencing and PCR analysis (Applied Bio-

systems Foster City CA) a region of about 500 base

pairs (bp) of 18S ribosomal ribonucleic acid gene was

ampli1047297ed by PCR Primers and free nucleotides from the

PCR products were removed using the QIAquick PCR

puri1047297cation kit (250) (Qiagen GmbH HildenGermany) The puri1047297ed PCR products were processed

for DNA sequencing by BigDye Terminator Cycle

Sequencing using capillary electrophoresis technology

in a genetic analyzer (ABI 310 Applied Biosystems

Foster City CA USA) Both strands of PCR ampli1047297ed

DNA fragm ents were sequenced in order to shun error of

sequencing28 The DNA sequence was analyzed and

searched in the Bla st DNA database for yeast identi1047297-

cation and typing29

Questionnaire

A standardized questionnaire was administered to allparticipants by one investigator (AS) The question-

naire comprised the following questions ldquo(1) What is

your age (in years) (2) What is your gender (male

female) (3) Do you have prediabetes (yesno) (3a) If

yes since when do you have prediabetes (3b) What type of treatment has your doctor recommended for the

management of prediabetes (allopathic herbal dietary

control others) (4) Do you brush your teeth (yesno)

(4a) If yes then how many times do you brush your

teeth ([i] once a day [ii] two times a day (iii) three

times a day (iv) more than three times a day) (5) If

you do not brush your teeth every day then how oftendo your brush your teeth (specify) (6) Do you brush

your tongue (yesno) (6a) If yes then how many times

do you brush your tongue every day ([i] once a day [ii]

two times a day (iii) three times a day (iv) more than

three times a day) (7) Do you rinse your mouth with an

oral rinse or mouthwash (yesno) (7a) If yes then how

many times do you rinse your mouth with an oral rinse

or mouthwash ([i] once a day [ii] two times a day (iii)

three times a day (iv) more than three times a day)rdquo

Oral lesions lesions on the tongue and number of missing teethIn all groups one examiner (AM) performed clinical

diagnosis of lesions on the buccal vestibule and tongue

(coated tongue 1047297ssured tongue hairy tongue

geographic tongue and median rhomboid glossitis

[MRG]) using standardized World Health Organization

criteria30-34 In all groups the number of missing teeth

(MT) (excluding third molars) were counted by the

same investigator (AM)

Statistical analysisData were statistically analyzed using SPSS software

(version 18 SPSS Inc Chicago IL USA) Level of

signi1047297cance between the groups (groups A B and C)

was assessed using Mann-Whitney U test For multiple

comparisons the Bonferroni post hoc test was used

Level of signi1047297cance was set at P lt 05 A multiple

logistic regression model was applied to adjust for

confounding variables (age gender number of MT

oral hygiene measures UWSFR and culturePCRresults)

RESULTSCharacteristics of the study cohortEighty patients with prediabetes (43 patients [38 males

and 5 females] in group A and 37 patients [35 males and

2 females] in group B) and 70 controls (61 males and 9

females) were included for study There was no signi1047297-

cant difference in age among participants in groups A

(412 16 years) B (431 21 years) and C (406

15 years) The mean duration of prediabetes among

participants in groups A and B was 11

22 months and132 14 months respectively (Table I)

The mean FBGL was signi1047297cantly higher among the

population with prediabetes (1093 42 mgL) and

patients in group A (1193 35 mgdL) than among

individuals in the control group (786 07 mgdL)

(P lt 05) respectively Mean FBGL was signi1047297cantly

higher among patients with prediabetes in group A

(1193 35 mgdL) than in group B (886 22 mgdL)

(P lt 05) Among patients with prediabetes mean HbA1c

levels were signi1047297cantly high in group A (62 05)

than in group B (49 03) (P lt 01) (see Table I)

HbA1c levels among patients with prediabetes in group Aand group B were measured 445 46 days and 113

24 days respectively prior to the present investigation

On clinical examination none of the participants

displayed tongue lesions and there was no signi1047297cant

difference in the number of MT and UWSFR in indi-

viduals with and without prediabetes (see Table I)

Oral Candida carriageOral C albicans carriage was signi1047297cantly higher in the

population with prediabetes (n frac14 80) (487) and the

patients with prediabetes in group A (512) compared

with group C (n frac14 70) (257) (P lt 01) There was no

difference in carriage of Candida tropicalis (C tropi-

calis) Candida parapsilosis (C parapsilosis) and C

albicans thorn C tropicalis (as mixed species) among

patients with prediabetes and individuals in group C

Among patients in groups A and B there was no

signi1047297cant difference in oral carriage of C tropicalis C

parapsilosis and C albicans thorn C tropicalis C

albicans thorn C parapsilosis as mixed species and

Candida krusei were isolated from 29 and 14 of

individuals in group C (Table II)

Our multiple logistic regression model showed no

signi1047297cant association between oral Candida carriage

OOOO ORIGINAL ARTICLE

Volume 117 Number 1 Javed et al 55

8102019 22124403_S2212440312X00257_S2212440313004586_main


and number of MT daily oral hygiene maintenance

regimens and UWSFR (data not shown)

Oral and tongue lesionsOral and tongue lesions (such as coated tongue 1047297ssured

tongue hairy tongue geographic tongue and MRG)

were not detected in any group clinically examined inthis study

QuestionnaireIn group A 86 (n frac14 3743) individuals reported

brushing their teeth once daily whereas in groups B

and C 837 (n frac14 3137) and 843 (n frac14 5970)

individuals respectively reported brushing their teeth

once daily None of the individuals in the study pop-

ulation reported brushing their tongue or using oral

rinses or mouthwashes as a component of their oral

hygiene maintenance regimens

DISCUSSIONTo our knowledge from indexed literature this is the

1047297rst study in which oral Candida carriage was

investigated in patients with prediabetes with particular

emphasis on glycemic status In general the population

with prediabetes investigated in the present study was

hyperglycemic (FBGL 1093 42 mgdL HbA1c

58 02) which is a possible explanation for the

increased oral C albicans carriage in patients with

prediabetes (n frac14 80) compared with healthy controls

(70 individuals in group C) Our 1047297ndings are in

accordance with those of earlier studies1235 in which

oral Candida carriage was reported to be increased in

patients with poorly controlled type 2 diabetes as

compared with controls

Glycemic control has been reported to enhance

healing and reduce periodontal in1047298amm ation in patients

with diabetes mellitus and prediabetes9121336 In the

present study we hypothesized that glycemic control

reduces oral Candida carriage in patients with predia-

betes Interestingly the present results showed no

signi1047297cant difference in oral Candida carriage among

individuals with prediabetes in Group A and individuals

previously prediabetic but now having normal glycemic

levels due to dietary control (group B) Various

explanations may be proposed to explain these results

Table II Oral Candida species isolated from individuals with and without prediabetes

Oral Candida species

All patients with

prediabetes (N frac14 80) n ()

Patients in group A

(N frac14 43) n ()

Patients in group B

(N frac14 37) n ()

Individuals in control

group (N frac14 70) n ()

Candida albicans 39 (487)y 22 (512)z 17 (46) 18 (257)yz

Candida tropicalis 26 (325) 14 (325) 12 (324) 17 (243)

Candida albicans thorn Candida tropicalis 13 (163) 6 (14) 7 (189) 7 (10)

Candida parapsilosis 2 (25) 1 (23) 1 (27) 1 (14)

Candida albicans thorn Candida parapsilosis d d d 2 (29)

Candida lusitaniae d d d d

Candida glabr at ad d d d

Candida krusei d d d 1 (14)

Candida guilliermondii

d d d d

No Candida species isolated d d d 24 (343)

These Candida species were identi1047297ed using polymerase chain reactionyP lt 01zP lt 01

Table I Characteristics of the study cohort

Parameters

All patients with

prediabetes (n frac14 80)

Patients in

group A (n frac14 43)

Patients in

group B (n frac14 37)


group (n frac14 70)

Gender 73M 7F 38M 5F 35M 2F 61M 9F

Mean age (y) 414 21 412 16 431 21 406 15

Duration of prediabetes (mo) 118 15 11 22 132 14 d

Fasting blood glucose level (mgdL) 1093 42z

1193 35y

886 22

786 07yz

Hemoglobin A1c () 58 02ǁ 62 05x 49 03xǁd

Mean No of missing teeth 38 12 47 33 44 14 21 02

Unstimulated whole salivary 1047298ow rate (mLmin) 042 03 041 01 043 02 052 04

F female M male

P lt 05yP lt 05zP lt 05xP lt 01ǁP lt 01

ORAL MEDICINE OOOO


8102019 22124403_S2212440312X00257_S2212440313004586_main


It is known that aging increased number of MT poor

oral hygiene maintenance and xerostomia are signi1047297-

cant risk factors for Candida growt h on oral tissues

particularly the dorsum of the tongue537 In the present

study participants in group A and group B were nearly

40 years old performed similar daily oral hygiene

maintenance protocols and had no signi1047297cant differ-ences in UWSFR and number of MT In addition the

short duration of prediabetes among patients in groups

A and B (nearly 1 year) may have been unable to

induce signi1047297cant changes in the periodontal status as

well as salivary 1047298ow rate in these individuals

Furthermore it is pertinent to mention that the most

recent HbA1c levels among participants in group B

were measured nearly 2 weeks before the present

investigation It is tempting to speculate that individuals

in group B could have been maintaining glycemic

levels since merely 2 weeks which may have been an

insuf 1047297cient time duration to reduce oral Candidacarriage in these individuals as compared with those in

group A (in which HbA1c levels were measured nearly

40 days before the present investigation) It is probable

that long-term control of hyperglycemia may reduce

oral Candida carriage in patients with diabetes andprediabetes however further longitudinal studies are

warranted in this regard

A direct association between tongue lesions

(including MRG) and oral candidiasis tobacco

smoking denture wearing and systemic conditions

(such as diabetes mellitus and AIDS) has been re-

ported

38-40

Lesions in the oral cavity (particularly thoseon the buccal mucosae) and tongue lesions (such as

hairy tongue 1047297ssured tongue coated tongue and

MRG) were not detected in any group clinically

examined in this study Although none of the study

participants reported brushing the dorsum of the tongue

as an adjunct to the regular oral hygiene maintenance

regimen the normal UWSFR that existed in all study

groups could have prevented oral Candida species from

accumulating and multiplying on the dorsum of the

tongue thereby preventing the occurrence of tongue

lesions Since tobacco smokers and betel nut chewers

were excluded from the present study it is possible that

oral and tongue lesions are more common in patients

with prediabetes who habitually smoke or chew tobacco

products than in those who do not use tobacco in any

form

There are a few limitations of the present study that

we address First quanti1047297cation of the oral Candida

species was not performed and this would have been

useful for better understanding these data Second

categorization of the individuals with prediabetes into

groups A and B was based on measurement of HbA1c

and FBGL levels whereas glycemic levels in self-

reported controls were determined using FBGL alone It

is known that the oral glucose tolerance test (OGTT) is

a valua ble and reliable tool for monitoring hypergly-

cemia 41 therefore it is highly recommended that

OGTT should be considered as a critical parameter in

future studies dealing with glycemic status in patients

with diabetes and in undiagnosed individuals Third

tobacco users were excluded from this study andtobacco smoking is a signi1047297ca nt risk factor for an

increased oral Candida carriage4243 It is tempting to

speculate that smokers with prediabetes are more

susceptible to oral fungal infections (due to an

increased oral Candida carriage) as compared with

nonsmokers with prediabetes and nondiabetic smokers

and nonsmokers Fourth most of our study participants

were men It has been reported that oral Candida

carriage is signi1047297cantly higher in women with type 2

diabetes compared with men with type 2 diabetes1

Thus further studies are needed to assess the limitations

of the present studyWithin the limits of the present investigation it is

concluded that oral Candida carriage is higher in

patients with prediabetes than in controls and may be

independent of glycemic status in patients with

prediabetes

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with and without prediabetes Am J Med Sci 201334594-98

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Engstrom PE Comparison of periodontal and socioeconomic

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diabetes J Oral Rehabil 20093639-44

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ethanol against Candida albicans and Staphylococcus aureus

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16 Javed F Tenenbaum HC Nogueira-Filho G et al Oral Candidacarriage and species prevalence amongst habitual gutka-chewers

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23 Yasui M Ryu M Sakurai K Ishihara K Colonisation of the oral

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24 Reichart PA Samaranayake LP Samaranayake YH Grote M

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29 Jonasson J Olofsson M Monstein HJ Classi1047297cation identi1047297ca-

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33 van der Wal N van der Waal I Candida albicans in median

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34 van der Wal N van der Kwast WA van der Waal I Medianrhomboid glossitis a follow-up study of 16 patients J Oral Med

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35 Martinez RF Jaimes-Aveldanez A Hernandez-Perez F

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control on the osseointegration of dental implants a systematic

literature review J Periodontol 2009801719-1730

37 Wang J Ohshima T Yasunari U et al The carriage of Candida

species on the dorsal surface of the tongue the correlation with

the dental periodontal and prosthetic status in elderly subjects

Gerodontology 200623157-163

38 Goregen M Miloglu O Buyukkurt MC Caglayan F Aktas AE

Median rhomboid glossitis a clinical and microbiological study

Eur J Dent 20115367-372

39 Arendorf TM Walker DM Tobacco smoking and denture

wearing as local aetiological factors in median rhomboid glossitis

Int J Oral Surg 198413411-415

40 Flaitz CM Nichols CM Hicks MJ An overview of the oral

manifestations of AIDS-related Kaposirsquos sarcoma Compend

Contin Educ Dent 199516136-138 140 142 passim quiz 148

41 Ouchi M Suzuki T Hashimoto M et al Urinary N-acetyl-beta-

D-glucosaminidase levels are positively correlated with 2-hr

plasma glucose levels during oral glucose tolerance testing in

prediabetes J Clin Lab Anal 201226473-480

42 Muzurovic S Hukic M Babajic E Smajic R The relationship

between cigarette smoking and oral colonization with Candida

species in healthy adult subjects Med Glas (Zenica) 201310397-399

43 Baboni FB Barp D Izidoro AC Samaranayake LP Rosa EA

Enhancement of Candida albicans virulence after exposition to

cigarette mainstream smoke Mycopathologia 2009168227-235

Reprint requests

Fawad Javed BDS PhD

3D Imaging and Biomechanical Laboratory

College of Applied Medical Sciences

King Saud University Riyadh Saudi Arabia

fawjavgmailcom

ORAL MEDICINE OOOO


8102019 22124403_S2212440312X00257_S2212440313004586_main


besides the oxidative stress induced by chronic hyper-

glycemia a reduced UWSFR in patients with prediabetes

may also have contributed in worsening periodontal

status Because xerostomia is a com mon manifestation in

patients with chronic hyperglycemia15 it is assumed that

oral Candida carriage is also high in patients with

prediabetes compared with healthy controls However toour knowledge from indexed literature oral Candida

carriage in patients with prediabetes has not yet been

investigated Furthermore studies61314 have shown that

glycemic control reduces the severity of periodontal

in1047298ammation in patients with poorly controlled diabetes

and prediabetes We therefore hypothesized that oral

Candida carriage would be reduced in patients previ-

ously diagnosed with prediabetes but who maintain their

fasting blood glucose level (FBGL) within normal limits

(70 to lt100 mgdL) as compared with patients with

poorly controlled prediabetes (100 to 125 mgdL) To

our knowledge from indexed literature this hypothesishad not been tested before our study

METHODSEthical approvalThe study was approved by the research ethics reviewcommittee of the Jinnah Postgraduate Medical Center

Karachi Pakistan The study was performed in accor-

dance with the Declaration of Helsinki as revised in

2000 It was mandatory for all study participants to

have read and signed the consent form before being

included in this study

Inclusion and exclusion criteriaOnly individuals with medically diagnosed prediabetes

(FBGL 100-125 mgdL [56-69 mmolL] hemoglobin

A1c [HbA1c] 57-64) were included Exclusion

criteria wer e (1) tobacco smoking13 (2) alcohol

consum ption15 (3) exclusive areca nut and gutka

chewing1617 (4) use of antibiotics antifungal agents

steroids or nonsteroidal anti-in1047298ammatory drugs within

the past 3 months131819 (5) self-reported systemic

diseases including type 1 and type 2 diabetes mellitus

hepat itis B hepatitis C and infection with HIV or

AIDS1132021

and (6) wearing partial or completedentures2223

Study participantsPatients with prediabetes were recruited from the dia-

betes care unit of a local hospital in Karachi Pakistan

Medical records of these patients were explored to

con1047297rm the diagnosis of prediabetes Control participants

self-reporting as not prediabetic were recruited from

a residential area near the hospital All participants were

invited to an oral health care center in the early morning

hours (in a fasting state) for FBGL measurement and

collection of oral yeast and unstimulated whole saliva

(UWS) samples

Hemoglobin A1c levels and fasting blood glucoselevelsHospital records of patients with prediabetes were

searched to determine the most recent HbA1c levels A

digital glucometer (Accu-Chek Activ Roche Diagnos-

tics Mannheim Germany) was used to measure the

FBGL Depending on the glycemic levels patients with

prediabetes were divided into 2 subgroups as follows

group A patients with prediabetes with FBGL between

100 and 125 mgdL (HbA1c 5) and group B

patients with prediabetes with FBGL lt100 mgdL

(HbA1c lt5) Self-reported systemically healthyindividuals (FBGL 70 to lt100 mgdL) were catego-

rized as controls (group C)

Collection of UWS samplesTo collect the UWS samples participants were seated

comfortably in a chair in a ldquocoachmanrdquo position and

requested to spit (without swallowing) into a gauged

measuring cylinder for 1047297ve continuous minutes UWSFR

was recorded in milliliters per minute (mLmin)

Collection of oral yeast samplesOral Cand ida samples were collected as described

previously1 In summary each sample was collected by

scraping the dorsum of the tongue and bilateral buccal

mucosa with a sterile cotton swab (bioMeacuterieux SAMontalieu-Vercieu France)1 Immediately after

sampling the swabs were returned to the containment

tube to avoid contamination At 37C Candida strains

were cultured in Sabouraud dextrose agar (Becton

Dickinson and Company Sparks MD USA) to quan-

tify the colony-forming unit s in the or al ca vities of

individuals with and without prediabetes2425 After 24

hours all cultures were inspected and monitoringcontinued until 7 days of incubation for yeast growth

following which they were subjected to speciation

Identification of oral yeast samplesA yeast identi1047297cation system (API 32-C System yeast

identi1047297cation programme bioMeacuterieux) was used to

identify the oral yeast species1 Yeast isolates that could

not be identi1047297ed with the oral yeast identi1047297cation

system were subjected to molecular identi1047297cation

Molecular identi1047297cation was performed as described

elsewhere26 Brie1047298y for DNA isolation yeast cells

were suspended in 200 mL sterile polymerase chain

reaction (PCR)egrade water and genomic DNA was

prepared using a DNA preparation robot (Roche

Diagnostics GmbH Mannheim Germany)27 Using

ORAL MEDICINE OOOO


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cation and typing29

Questionnaire
















































































8102019 22124403_S2212440312X00257_S2212440313004586_main










































All patients with


Patients in group A

(N frac14 43) n ()

Patients in group B

(N frac14 37) n ()












d d d d




Parameters

All patients with


Patients in


Patients in



group (n frac14 70)


Mean age (y) 414 21 412 16 431 21 406 15



1193 35y

886 22

786 07yz




F female M male


ORAL MEDICINE OOOO


8102019 22124403_S2212440312X00257_S2212440313004586_main





























ported

38-40
















form






























prediabetes











Pathol 198817354-357















2184-2189










8102019 22124403_S2212440312X00257_S2212440313004586_main











2012344100-104











74-82













201221442-446







2006476-10










4479-4485








ahead of print]




3536




Chemother 200044710-712



263-272




demiol 198081-26





Case Rep 2011564-68



Surg 198615322-325


198641117-120




201388










Eur J Dent 20115367-372

















Reprint requests

Fawad Javed BDS PhD




fawjavgmailcom

ORAL MEDICINE OOOO


8102019 22124403_S2212440312X00257_S2212440313004586_main
















cation and typing29

Questionnaire
















































































8102019 22124403_S2212440312X00257_S2212440313004586_main










































All patients with


Patients in group A

(N frac14 43) n ()

Patients in group B

(N frac14 37) n ()












d d d d




Parameters

All patients with


Patients in


Patients in



group (n frac14 70)


Mean age (y) 414 21 412 16 431 21 406 15



1193 35y

886 22

786 07yz




F female M male


ORAL MEDICINE OOOO


8102019 22124403_S2212440312X00257_S2212440313004586_main





























ported

38-40
















form






























prediabetes











Pathol 198817354-357















2184-2189










8102019 22124403_S2212440312X00257_S2212440313004586_main











2012344100-104











74-82













201221442-446







2006476-10










4479-4485








ahead of print]




3536




Chemother 200044710-712



263-272




demiol 198081-26





Case Rep 2011564-68



Surg 198615322-325


198641117-120




201388










Eur J Dent 20115367-372

















Reprint requests

Fawad Javed BDS PhD




fawjavgmailcom

ORAL MEDICINE OOOO


8102019 22124403_S2212440312X00257_S2212440313004586_main










































All patients with


Patients in group A

(N frac14 43) n ()

Patients in group B

(N frac14 37) n ()












d d d d




Parameters

All patients with


Patients in


Patients in



group (n frac14 70)


Mean age (y) 414 21 412 16 431 21 406 15



1193 35y

886 22

786 07yz




F female M male


ORAL MEDICINE OOOO


8102019 22124403_S2212440312X00257_S2212440313004586_main





























ported

38-40
















form






























prediabetes











Pathol 198817354-357















2184-2189










8102019 22124403_S2212440312X00257_S2212440313004586_main











2012344100-104











74-82













201221442-446







2006476-10










4479-4485








ahead of print]




3536




Chemother 200044710-712



263-272




demiol 198081-26





Case Rep 2011564-68



Surg 198615322-325


198641117-120




201388










Eur J Dent 20115367-372

















Reprint requests

Fawad Javed BDS PhD




fawjavgmailcom

ORAL MEDICINE OOOO


8102019 22124403_S2212440312X00257_S2212440313004586_main





























ported

38-40
















form






























prediabetes











Pathol 198817354-357















2184-2189










8102019 22124403_S2212440312X00257_S2212440313004586_main











2012344100-104











74-82













201221442-446







2006476-10










4479-4485








ahead of print]




3536




Chemother 200044710-712



263-272




demiol 198081-26





Case Rep 2011564-68



Surg 198615322-325


198641117-120




201388










Eur J Dent 20115367-372

















Reprint requests

Fawad Javed BDS PhD




fawjavgmailcom

ORAL MEDICINE OOOO


8102019 22124403_S2212440312X00257_S2212440313004586_main











2012344100-104











74-82













201221442-446







2006476-10










4479-4485








ahead of print]




3536




Chemother 200044710-712



263-272




demiol 198081-26





Case Rep 2011564-68



Surg 198615322-325


198641117-120




201388










Eur J Dent 20115367-372

















Reprint requests

Fawad Javed BDS PhD




fawjavgmailcom

ORAL MEDICINE OOOO


22124403_s2212440312x00257_s2212440313004586_main

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