Molecular epidemiology and antifungal susceptibility of Serbian Cryptococcus neoformans isolates

Molecular typing and antifungal susceptibility testing of 34 clinical Serbian Cryptococcus neoformans isolates from 25 patients was retrospectively performed. Amplified fragment length polymorphism (AFLP) fingerprinting was used for genotyping, whereas a novel real-time PCR was used to determine the mating- and serotype. The antifungals amphotericin B, 5-fluorocytosine, fluconazole, voriconazole, itraconazole and posaconazole were used to determine the antifungal susceptibility profiles. The majority of isolates belonged to genotype AFLP1/VNI (n=20; 58.8%), followed by AFLP2/VNIV (n=10; 29.4%), AFLP3/VNIII (n=3; 8.8%) and AFLP1B/VNII (n=1; 2.9%). All AFLP1/VNI isolates were mating-serotype A, the sole AFLP1B/VNII isolate was found to be aA, whereas AFLP2/VNIV harboured serotype D isolates with either the a (n=2; 5.9%) or (n=8; 23.5%) mating-type allele. The isolates (n=3; 8.8%) that were found to be genotype AFLP3/VNIII had the hybrid mating- and serotype combination aA-D. In vitro antifungal susceptibility testing showed that all isolates were susceptible to amphotericin B, voriconazole and posaconazole. Low resistance level was observed for fluconazole (n=1; 2.9%) and 5-fluorocytosine. (n=2; 5.8%). A large percentage of isolates was found to be susceptible dose dependent to itraconazole (n=16; 47.1%). AFLP1/VNI was the most common genotype among clinical C. neoformans isolates from immunocompromised patients in Serbia. C. neoformans from HIV-negative patients were significantly less susceptible to 5-fluorocytosine (P<0.01). Correlation between genotypes and antifungal susceptibility was not observed.