Objectives:
To isolate C. albicans & C. tropicalis from hospital environment and clinical samples.To determine biofilm, phospholipase and protease production of clinical and environmental isolates of C. albicans & C. tropicalis.To compare the virulence profile of clinical and environmental C. albicans & C. tropicalis isolates.




Methods:
Yeasts were isolated from high touch surfaces in the hospital (medical & surgical intensive care units, operation theatre, casualty) by swabbing sterile swabs soaked in antibiotics solution and inoculating them into brain heart infusion broth with chloramphenicol, followed by incubation at 37℃ for 5 days. Turbid broths were then sub cultured onto CHROM agar and Candida identified as per standard microbiological protocol. Clinically significant yeasts submitted to diagnostic laboratory for routine culture and sensitivity during the study period were also included in this study. Ten each of C. albicans & C. tropicalis from hospital environment and clinical infection were subjected to assays for biofilm formation, phospholipase, and proteinase production. Assays were performed in triplicate.

For detecting biofilm production active cultures were inoculated to 96-well plate and the wells were rinsed with phosphate buffered saline (PBS). Following incubation, the wells were rinsed with PBS, stained with 0.4% crystal violet and air dried. Wells were then rinsed with distilled water and the stain extracted using 95% ethanol. Absorbance of the extracted crystal violet was measured at 595nm.

For detecting the proteinase and phospholipase production 48 hours old cultures were adjusted to 106 cells and 10 µl of this was seeded to milk agar & egg yolk agar. Test was interpreted using proteinase and phospholipase index.

Results:
Fifty-three Candida (26.5%,53/200) were isolated from 200 environmental samples and 43 clinically significant isolates were obtained from urine (34) and blood samples (9) during the study period. Non-albicans Candida spp. (NAC) predominated (81.1%) among environmental isolates; C. tropicalis (24.52%), and C. lusitaniae (24.52%) followed by C. albicans (18.9%), C. parapsilosis (15.1%), C. krusei (11.3%), C. guilliermondii (5.7%). NAC also predominated (55.8%) in clinical isolates; C. albicans (44.2%) followed by C. tropicalis (39.5%), C. parapsilosis (11.6%), C. krusei (4.7%). All isolates produced biofilm. Mean A595 for clinical and environment C. albicans isolates was 0.138±0.06 and 0.128±0.08. Environmental and clinical C. tropicalis isolates had A595 of 0.147±0.08 & 0.160±0.02. All clinical & 70% of environmental C. albicans, & 20% clinical & 60 % of environmental C. tropicalis produced phospholipase. No environmental & 40% of clinical C. albicans, & 10% of the environmental & 40% of clinical C. tropicalis produced proteinase. No statistically significant difference in biofilm, phospholipase and proteinase production was observed between environmental & clinical isolates of C. albicans & C. tropicalis except more clinical C. albicans isolates (40%) produced protease(P=0.09).

Conclusion:
No statistically significant difference in biofilm, phospholipase and proteinase production was observed between environmental & clinical isolates ofC. albicans &C. tropicalis except more clinicalC. albicans isolates (40%) produced protease(P=0.09).
Low sample size was the biggest limitation of the study and the findings need to be confirmed using more isolates.

Comparison of virulence profile of C. albicans and C. tropicalis isolated from patients with Candida infection and hospital environment