Advanced approach for antifungal susceptibility and characterization of resistance properties in clinical and environmental isolates of Cryptococcus species complex

Background: Meningitis due to Cryptococcus neoformans/gattii is a fatal infection affecting immunocompromised population worldwide. Amphotericin B (AmB), fluconazole (FLC) and 5-flucytosine are the drugs of choice to treat the infection. We studied antifungal susceptibility pattern of clinical and environmental cryptococcal species using newer approach and analyze their resistant characteristics. Methods: Eighty clinical (54 C. neoformans and 26 C. gattii) ) and 18 environmental (14 C. neoformans and 4 C. gattii) ) isolates were subjected to antifungal susceptibility testing by automated (VITEK2C) method. Minimum inhibitory concentrations (MIC) were analyzed statistically. Genomic DNA of FLC resistant isolates was extracted and amplified to detect presence of CnAFR1 gene. Results: C. neoformans showed 1.85% and 21.4% AmB resistance, and 1.85% and 28.5% FLC- resistance, whereas C. gattii showed 25% and 50% FLC-resistance among clinical and environmental isolates respectively. MIC values were significantly ( p < 0.05) different for the isolates from 2 sources. CnAFR1 gene sequence analysis revealed phylogenetic relationship among the resistant isolates. Conclusions: This pioneering study provides an insight into the sensitivity patterns of clinical and environmental cryptococcal isolates from south India. The recent emergence of AmB-resistance may transpire as a challenge for the clinicians. As the clinical and environmental isolates are phylogenetically evolved from CnAFR1 gene of Filobasidiella neoformans, , the resistance is most probably an inherent attribute. This study emphasizes the need for speciation and antifungal susceptibility testing of cryptococcal isolates from clinical sources to institute appropriate antifungal therapy and to reduce the mortality and morbidity.