SPECIATION AND ANTIFUNGAL SUSCEPTIBILITY PATTERN OF CANDIDA ISOLATES FROM VARIOUS CLINICAL SPECIMENS AT A TERTIARY CARE HOSPITAL, MAHARASHTRA.

Dr. Ishan Pandita 1 , Dr. A. S. Wyawahare 2, , Dr. M.V. Mulay 3 , Dr. Tazammul Hussain 4 and Dr. Smita Kulkarni 5 . 1. PG Resident, Department of Microbiology, MGM Medical College and Hospital, Aurangabad, Maharashtra. 2. Professor, Department of Microbiology, MGM Medical College and Hospital, Aurangabad, Maharashtra. 3. Professor and Head, Department of Microbiology, MGM Medical College and Hospital, Aurangabad, Maharashtra. 4. PG Resident, Department of Microbiology, MGM Medical College and Hospital, Aurangabad, Maharashtra. 5. Assistant Professor, Department of Microbiology, MGM Medical College and Hospital, Aurangabad, Maharashtra. ...................................................................................................................... Manuscript Info Abstract ......................... ........................................................................ Manuscript History Received: 06 August 2019 Final Accepted: 08 September 2019 Published: October 2019


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antibiotics, AIDS and other immunosuppressive conditions have led to significant rise in Candida and other fungal infections. 4 In the recent years emergence of anti-fungal resistance has further complicated the situation. 5, 6, 7 Emergence of multi-drug resistant non-albicans Candida (NAC) as important cause of infections is also a matter of concern. 8 It is well known fact that the conventional methods of identification are time consuming and Vitek-2 system is an acceptable alternative method for rapid speciation and sensitivity testing of Candida species. 9, 10, 11 The rising burden of Candida infections and ever increasing anti-fungal resistance among Candida species necessitates accurate and rapid identification of Candida isolates for better patient outcome. 12 This study was undertaken to find out institutional anti-fungal susceptibility pattern of Candida isolates for an effective anti-fungal drug therapy for Candida infection.

Material and methods:-
This study was undertaken only after approval of institutional ethical committee. It was a prospective study undertaken at MGM Medical College, Aurangabad for 18 Months (1 st January, 2018 to 30 th June, 2019). All clinical samples which were received in Microbiology department for routine culture and sensitivity during the study period from which Candida species were isolated were included in the study; repeat isolates from same patient from same source were excluded.
Presumptive identification of the Candida isolates was based on colony morphology on blood agar (small, 1-2 mm, creamy white, smooth, pasty, non-hemolytic colonies with yeasty odor after 24 hours incubation) 13 , Gram Stain (Gram positive oval budding yeast like cells) 13 and Germ tube test (Isolate giving positive result presumptively identified as Candida albicans). 13 Final identification and antifungal susceptibility testing of Candida isolates was done using Vitek 2 compact system using Yeast card ID (VITEK 2 YST) and Yeast card AST (AST-YS07) respectively. For inoculum preparation pure sub-cultured colonies were suspended in aqueous 0.45% (wt/vol) NaCl to achieve a turbidity equivalent to a McFarland 1.8-2.2 McFarland standard according to the manufacturer's recommendations and measured on the DensiChek turbidity meter (Biomerieux, India). The cassettes were loaded into the Vitek-2 compact system. Quality control was done using the following strains as controls for the evaluation: C. albicans ATCC14053 and Candida parapsilosis ATCC22019. Yeast AST card (AST-YS07) contains these 6 anti-fungal agents: Amphotericin B, Caspofungin, Flucytosine, Fluconazole, Micafungin, Voriconazole.

Results:-
A total of 164 Candida isolates were analysed. Out of 164 Candida isolates, Candida albicans with 67 (40.8%) isolates was most common species. Among non-albicans Candida, Candida tropicalis was the most predominant species (29.3%) ( Table 1). The age and sex distribution of Candida species is described in Table 2. About 51% of the Candida were isolated from adult age group  and 54% of the isolates were from male patients.  Table 3). Out of 67 isolates of Candida albicans, 31 (46.2%) isolates were from urine followed by 18 (26.8%) isolates from sputum. Candida parapsilosis was the most common isolate from blood (8 isolates) while Candida tropicalis was the most common isolate from Endo-Tracheal secretions (2 isolates).

Table 3:-Specimen distribution of Candida Species
Among 164 Candida isolates, 82 were isolated from patients admitted in various wards and an equal number i.e 82 were isolated from ICU patients.
Percentage susceptibility of Candida albicans and major isolates of non-albicans Candida to various anti-fungal agents has been described in Table 4 and Table 5.
All the isolates of Candida albicans (100%) were susceptible to Amphotericin B, while least susceptibility was seen for Voriconazole (85%). In isolates of Candida tropicalis maximum susceptibility was seen to Fluconazole (100%) while least susceptibility was seen to Voriconazole (93.7%). Among other non-albicans Candida species susceptibility results were as follows: Among 6 isolates of Candida ciferrii 3 isolates (50%) were susceptible to all the anti-fungal agents, 2 isolates (33.3%) were resistant to both Amphotericin B and Voriconazole and 1 isolate (16.7%) was resistant to Amphotericin B. Among 5 isolates of Candida lusitaniae 4 isolates (80%) were susceptible to all anti-fungal agents while 1 isolate (20%) was resistant to Voriconazole and 1 isolate (100%) of Candida krusei was resistant to Flucytosine.   Recent studies have shown that Candida species could cause severe infections with high morbidity and mortality in hospitalized patients. 14, 15, 16 Among various Candida species isolated in routine clinical laboratory samples C. albicans, C. glabrata, C. tropicalis, C. parapsilosis, and C. krusei are the most common species. 17 In our study Candida albicans (40.8%) was the most common isolate followed by Candida tropicalis (29.3%) and Candida parapsilosis (9.7%) as major isolates which is consistent with the study of Kaur R et al. 11 Candida albicans has been reported as the most isolated species from patients in various other studies also done in Asian, European, and American countries. Most Candida isolated were from urine 78 (47.6%) isolates, out of which 31 (46.2%) isolates were of Candida albicans. About 33 (20.1%) isolates were from sputum. When demographically distributed, about 51% of the Candida were isolated from adult age group  and 54% of the isolates were from male patients.
Our study showed an increase in the antifungal drug resistance, especially for the azole group of drugs, both in C. albicans and non-albicans Candida species which is in agreement with study by

Conclusion:-
Candida infections in hospitalized patients is a significant problem worldwide. Resistance among Candida species is increasing and various species already have inherent resistance to some commonly used antifungals. The rising burden of Candida infections and ever increasing anti-fungal resistance among Candida species necessitates accurate and rapid identification of Candida isolates for better patient outcome as any delay can result in increased mortality and morbidity in patients. Non-albicans Candida species are on rise, especially C. tropicalis which was the most frequent non-albicans Candida isolated in our tertiary care center. Resistance among Candida species to various antifungal agents especially to azole group of drugs is on rise as is evident from our study also which can increase further in future if steps are not taken to control it. Based on the present results, it is evident that routine identification of Candida isolates to the species level, and the detection of resistant strains by antifungal susceptibility test is essential. Furthermore, there is a continued need for surveillance of Candida infections to monitor changes in the epidemiological features and antifungal susceptibility and also to develop and evaluate prevention strategies.