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Indian J Med Res 126, September 2007, pp 216-219 Prevalence & susceptibility to fluconazole of Candida speciescausing vulvovaginitis Srujana Mohanty, Immaculata Xess, Fahmi Hasan, Arti Kapil, Suneeta Mittal* & Jorge E. Tolosa** Departments of Microbiology & *Obstetrics & Gynaecology, All India Institute of Medical SciencesNew Delhi, India & **Global Network for Perinatal & Reproductive Health, Thomas Jefferson UniversityPhiladelphia, PA, USA Background & objectives: Vulvovaginal candidiasis is an important cause of morbidity in women of
reproductive age. This study was carried out to determine the species prevalence and susceptibility
pattern to fluconazole of yeasts isolated from the vagina of symptomatic women.

Methods: This prospective study was conducted in a rural primary health care center of north India
from May 2003 to April 2004 and included 601 married, sexually active women (18-49 yr) with the
self reported symptoms of vaginal discharge and/or genital itching and/or genital burning. Specific
aetiology of the genitourinary symptoms including candidal infection were determined. Specimens
from the lateral wall of vagina were subjected to direct wet mount microscopy and fungal culture
on Sabouraud’s dextrose agar. Susceptibility testing to fluconazole was carried out using broth
microdilution method.

Results: Yeasts were isolated in 111 (18.5%) women and these consisted of Candida glabrata (56,
50.4%), C. albicans
(39, 35.1%), C. tropicalis (12, 10.8%), C. krusei (3, 2.7%) and C. parapsilosis (1,
0.9%). Susceptibility testing carried out on 30 representative isolates (15 C. glabrata
, 10 C. albicans,
4 C. tropicalis
and 1 C. parapsilosis) revealed that 21 isolates (70%) were susceptible (MIC, < 8 mg/
ml) to fluconazole while 9 (30%) were susceptible-dose dependent (S-DD, MIC 16-32
Interpretation & conclusion
: Our findings suggest a low prevalence of fluconazole resistance in
vaginal candida isolates in our population. However, a high prevalence of non-albicans candida
species and increased dose-dependent resistance in these isolates necessitates vigilance since this
may warrant a change in the optimal therapy of non-albicans candida vaginitis.

Key words Antifungal susceptibility - Candida - candida vaginitis - fluconazole - non-albicans species
Approximately three-quarters of all women and refractory episodes1,2. The majority of cases of experience at least one episode of vulvovaginal vulvovaginal candidiasis are caused by Candida candidiasis during their lifetime and nearly half of them albicans; however, episodes due to non-albicans species suffer from multiple episode1. In about 5 per cent of of Candida appear to be increasing1,3. Most non- cases, the disease has a chronic course, showing frequent albicans Candida species have higher minimum MOHANTY et al: PREVALENCE & SUSCEPTIBILITY OF CANDIDA SPECIES TO FLUCONAZOLE inhibitory concentrations (MICs) to the azole antifungal Media Mumbai, India), assimilation of various sugars, agents, and infections they cause are often difficult to and growth in presence of actidione5,6.
treat3,4. This phenomenon emphasizes the importance Antifungal susceptibility testing: Susceptibility testing of identification and surveillance of the Candida species to fluconazole was performed using a broth microdilution in the clinical settings. This study was carried out to method (M27-A2) according to the Clinical Laboratory determine the species prevalence and fluconazole Standards Institute (CLSI) guidelines7. The microtitre susceptibility among yeast isolates from women with plates were incubated at 35°C for 24-48 h. The amount of growth in a well containing the antifungal agent was Material & Methods
compared with the amount of growth in an antifungal-free growth control well. The minimum inhibitory This prospective study was conducted from May concentration (MIC) was read as the lowest concentration 2003 to April 2004 at a rural primary health care center of antifungal that inhibited 50 per cent growth of the at Ballabhgarh, Haryana, a rural field practice area in organism detected visually. Quality control was ensured north India, under the All India Institute of Medical by testing the CLSI recommended quality control strains Sciences (AIIMS), New Delhi. It was done as part of a Candida parapsilopsis ATCC 22019 (MIC range 2-8 mg/ larger study of evaluation of the use of syndromic ml) and Candida krusei ATCC 6258 (MIC range 16-64 management of reproductive tract infections. The ethical review committees of AIIMS, New Delhi, ThomasJefferson University, Global Network for Perinatal and Isolates with MIC < 8 mg/ml were considered to be Reproductive Health (GNPRH), USA, and Population susceptible to fluconazole, whereas isolates with MIC > 64 mg/ml were considered to be resistant7. Isolateswith MICs between 16-32 mg/ml were fluconazole Patient population: The inclusion criteria for the study were married and sexually active women between 18-49 yr of age who presented to the health care center Results & Discussion
with self reported symptoms of vaginal discharge and/ A total of 710 women were screened, of whom only or genital itching and/or genital burning during the study 611 were found eligible for enrollment. Ten women period. Pregnant women, women with severe medical declined internal examination. Thus, a total of 601 disorders requiring immediate referral to higher level women were included in the study and yeasts were of healthcare, women who were currently menstruating, isolated in 111 (18.5%) of them. Candida glabrata was never been sexually active, who had a hysterectomy, the most common species among the isolates (56, had taken a course of antibiotics within preceding three 50.4%) followed by C. albicans (39, 35.1%). Other weeks and who had been previously enrolled in this Candida species isolated were C. tropicalis (12, 10.8%), study were excluded. Written informed consent was C. krusei (3, 2.7%) and C. parapsilosis (1, 0.9%). Thus, obtained from each participant woman.
the overall prevalence of non-albicans candida species An attempt was made to determine the specific was 64.8 per cent (72/111). Susceptibility testing to aetiology of the genitourinary symptoms and fluconazole was carried out on 30 representative isolates determination of candidal infection was one of them.
(15 C. glabrata, 10 C. albicans, 4 C. tropicalis and 1 Two sterile, cotton tipped swabs were used to collect C. parapsilosis). Of these, 21 isolates (70%) were specimens from lateral wall of vagina of each woman.
susceptible to fluconazole while 9 (30%) were One of the two swabs was used to determine the susceptible-dose dependent (Table). Complete presence of yeast by direct wet-mount microscopy using resistance was not detected in any of the isolates tested.
a drop of 10 per cent potassium hydroxide solution.
Of the 9 isolates with elevated MICs, 7 were C. glabrata, The other swab was used for fungal culture on 1 was C. tropicalis and 1 was C. albicans.
Sabouraud’s dextrose agar (Hi-Media, Mumbai, India) In this study, the overall prevalence of vulvovaginal supplemented with 0.06 mg/ml gentamicin, with and candidiasis in a community setting was found to be 18.5 per cent which is similar to studies from India8,9 and Identification: Species identification of yeast isolates elsewhere10,11 with rates ranging from 20.8 - 23 per cent.
was done by standard procedures including morphology, However, the overall percentage of non-albicans germ tube test, cornmeal agar test (Hi-Media, Mumbai, vaginitis (64.8%) was much higher than in previous India), triphenyl tetrazolium chloride reduction (Hi- reports3,4,10,11. In a study conducted on 1498 women who Thus in recent years, there has been a significant Table. Susceptibility to fluconazole of vaginal Candida isolates
increase in infections caused by non-albicans species of Candida, particularly, C. glabrata and C. tropicalis.
We speculate this increasing detection of non-albicans Candida species is probably related to the widespread and inappropriate use of antimycotic treatments (self- medication, long-term maintenance treatments and repeated treatments for candidosis episodes)1. C. albicans eradication by these means causes a selection of species (such as C. glabrata) that are resistant to In conclusion, our study provides information on antifungal susceptibility of vaginal yeast isolates in a rural community in India. Since the majority of C.
isolates were susceptible to fluconazole, its attended the Edinburgh Family Planning Centre over a use may be continued for empirical therapy of 18 month period10, fungi other than C. albicans uncomplicated candidal vulvovaginitis in the comprised 27.7 per cent of all fungal species obtained.
community. Use of alternative agents (like boric acid, An Australian study found vulvovaginal yeast carriage flucytosine)16 may be considered when treating among 21 per cent of 5,802 women in a primary care vulvovaginitis caused by non-albicans species setting; non-albicans species were isolated in only 11 (especially C. glabrata and C. krusei). As only a limited per cent of the positive cultures11. Spinillo et al3 reported number of Candida isolates could be tested in this study, that 17 per cent of 209 isolates from symptomatic further clinical studies need to be performed involving patients referred to an Italian vulvovaginitis clinic were more number of isolates to confirm the findings.
non-albicans species. Richter et al4 reported an overallpercentage of 24 per cent of non-albicans vaginitis Acknowledgment
among 564 vaginal yeast cultures collected from 429 This work was supported by the European Commission symptomatic candidal vuvovaginitis patients.
through a grant, provided to the HIV/STI Prevention and CareResearch Programme of the Population Council India and with Antifungal susceptibility testing in our study support from the Global Network for Perinatal and Reproductive revealed that none of the Candida isolates tested were Health and its donors, INCLEN, the Rockefeller Foundation.
resistant to fluconazole, though 9 of 30 (30%) isolates Authors thank Dr Sarah Hawkes from the London School ofHygiene and Tropical Medicine; Dr Christopher Elias from were fluconazole S-DD and maximum number of these Population Council (Thailand) for technical support; Dr Heiner S-DD isolates were C. glabrata. However, we could Grosskurth, Dr Sabine Flessenkaemper and Dr Gurumurthy not perform susceptibility testing for C. krusei which Rangaiyan from Population Council, India for technical support is intrinsically resistant to fluconazole. Fluconazole resistance in vaginal C. albicans isolates is an References
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recovered from women with recurrent vulvovaginitis12.
Sobel JD. Candida vulvovaginitis. Sem Dermatol 1996; 15 : No fluconazole resistance was identified among 75 C. albicans vaginal isolates from symptomatic women in Spinillo A, Capuzzo E, Gulminetti R, Marone P, Colonna L, England13. A study in Brazil also reported no fluconazole Piazzi G. Prevalence of and risk factors for fungal vaginitis resistance among 56 C. albicans vaginal isolates14.
caused by non-albicans species. Am J Obstet Gynecol 1997; However, in another US study, fluconazole resistance has been reported in 14 of 393 (3.6 %) C. albicans Richter SS, Galask RP, Messer SA, Hollis RJ, Diekema DJ, isolates collected from complicated vaginitis patients Pfaller MA. Antifungal susceptibilities of Candida species prior to 200115. As regards the antifungal susceptibility causing vulvovaginitis and epidemiology of recurrent cases.
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Haricharan RN, et al. Species-specific prevalence of vaginal 15. Sobel JD, Zervos M, Reed BD, Hooton T, Soper D, Nyirjesy candidiasis among patients with diabetes mellitus and its P, et al. Fluconazole susceptibility of vaginal isolates obtained relation to their glycaemic status. J Infect 2000; 41 : 162-6.
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Reprint requests: Dr Immaculata Xess, Associate Professor, Department of Microbiology, All India Institute of Medical Sciences New Delhi 110029, Indiae-mail: i_xess@yahoo.com

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Biographical sketch

PROFESSIONAL BIOGRAPHICAL PROFILE OF CHARLES W. KING EDUCATION Charles W. King received his bachelor's and master's degrees from the University of Texas in 1956 and 1957 respectively. From 1957 until 1960, King was a marketing research analyst and section head with Humble Oil and Refining Company (now Exxon). In 1960, King initiated doctoral study and subsequently received his d

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