Evaluation of the CHROMagar Candida Plus medium for presumptive identification of yeasts and MALDI-TOF MS identification.

Rapid and accurate yeasts species identification in clinical laboratories is important for appropriate and timely antifungal treatment. We evaluate the performance of the new medium CHROMagar™ Candida Plus for presumptive identification of yeasts species and MALDI-TOF identification. We identify 303 strains belonging to 60 clinically relevant yeasts species by using the new medium. Presumptive identification was correct at the Candida albicans complex, Candida tropicalis and Pichia kudriavzevii (Candida krusei) species. However, although this medium was able to identify all Candida auris and Candida glabrata strains, other species were misidentified as C. auris or C. glabrata. A total of 215 strains were identified by using MALDI-TOF and evaluated two incubation temperatures (30°C and 37°C) and two incubation times (24 h and 72 h). Most strains (94%; 202/215) were correctly identified at the species (n:190) or complex level (n:12) at both temperatures and incubation times. However, we observed that the time of incubation (24 h vs. 72 h) affects the score values when yeasts are incubated at 37°C, but does not affect score values when yeasts are incubated at 30°C. In conclusion, the new medium has a good performance in the presumptive identification of the C. albicans complex, C. tropicalis and P. kudriavzevii (C. krusei). In addition, this medium is useful for the screening of C. auris and C. glabrata isolates, but identification should be confirmed by other more specific techniques, like MALDI-TOF.

Genotypic Diversity and Antifungal Susceptibility of Clinical Isolates of Cryptococcus Gattii Species Complex from Argentina.

The aim of this study was to determine the genotypic diversity of 22 Cryptococcus gattii species complex clinical isolates from Argentina and to place these genotypes within the diversity of clinical, veterinary and environmental isolates from Latin America. Mating type and antifungal susceptibility of the isolates were also determined. By URA5-RFLP, nine isolates were identified as molecular type VGI, 10 as VGII, one as VGIII and two as VGIV. Multilocus sequence typing (MSLT), following the International Society for Human and Animal Mycology (ISHAM) consensus MLST scheme, was used to determine the genotypic diversity. Our results suggest that, in Argentina, VGI isolates have low genetic diversity, while VGII isolates have high genetic diversity. Both isolates identified as VGIV by URA5-RFLP were genotyped by MLST as belonging to the currently named VGVI clade. From all isolates, eight sequence types (STs) were unique for Argentina, while five STs have been reported already in other countries, being of high interest the genotypes ST20 and ST7 since they belong to the subtypes VGIIa and VGIIb, respectively, which are associated with hypervirulent strains responsible for outbreaks in North America. To note, geographical analysis showed that some genotypes may be associated with some regions in Argentina. Most isolates were MATα, but we are reporting one isolate MATa for the first time in the country. Antifungal susceptibility tests showed that itraconazole, voriconazole and posaconazole had high activity against all isolates, while amphotericin B, fluconazole and 5-fluorocytosine were the least active drugs against all studied isolates.

Frequency and geographical distribution of genotypes and mating types of Cryptococcus neoformans and Cryptococcus gattii species complexes in Argentina / Frecuencia y distribución geográfica de genotipos y tipos sexuales de los complejos de especies Cryptococcus neoformans y Cryptococcus gattii en Argentina

Abstract The aim of this work was to know the frequency and geographical distribution of genotypes and mating types of Cryptococcus neoformans and Cryptococcus gattii species complexes isolated from human infections in Argentina during the period from April 2009 to April 2011. A multicenter study was conducted, in which 372 isolates were obtained from 61 laboratories throughout the country. Of those, 98.8% of the isolates belonged to the C. neoformans species complex and 1.1% to the C. gattii species complex. Genotype VNI (MATa) was the most frequently isolated (n = 326, 87.6%), followed by VNII (MATa) (n = 22, 5.9%), the recently described VNII-VNIV (aADa) hybrid (n = 14, 3.8%), VNIV (MATa) (n=4, 1.1%), VNIII (aADa) hybrid (n = 1, 0.3%), and VNIII (aADa) hybrid (n = 1, 0.3%). The Argentine Central region showed the greatest number of cases and genotype diversity. Interestingly, a relative high frequency was observed in genotype VNII (MATa) in the Cuyo, Northeast and Northwest regions and, also in VNII-VNIV (aADa) hybrids in the Northwest region. C. gattii species complex was isolated at a low rate; 3 VGI (MATa) and 1 VGII (MATa) isolates were obtained from the Northwest and Central regions. In conclusion, this study shows that genotype frequencies seem to vary among regions in Argentina and reveals a relatively high frequency of rare hybrids in the Northwest region. Further regional clinical and environmental studies may help to elucidate if those varia-tions in frequencies are associated with the existence of regional ecological niches or any other regional factors.

Resumen El objetivo de! trabajo fue conocer la frecuencia y la distribución geográfica de genotipos y tipos sexuales de aislados pertenecientes a los complejos de especies Cryptococcus neoformansy Cryptococcus gattii obtenidos de infecciones humanas en Argentina. Entre abril de 2009 y abril de 2011 se realizó un estudio multicéntrico del que se obtuvieron 372 aislados de 61 laboratorios de diferentes zonas del país. El 98,8% de los aislados pertenecieron al complejo C. neoformansy el 1,1% al complejo C. gattii. El genotipo VNI (MATa) fue el más frecuente (n = 326; 87,6%), le siguieron VNII (MATa) (n = 22; 5,9%), el híbrido VNII-VNIV (aADa) (n = 14; 3,8%), VNIV (MATa) (n =4; 1,1%) y los híbridos VNIII (aADa) (n = 1; 0,3%) y VNIII (aADa) (n = 1; 0,3%). La región Centro mostró el mayor número de casos y la mayor diversidad de genotipos. Cabe destacar que el genotipo VNII (MATa) tuvo una frecuencia relativamente alta en las regiones de Cuyo, Noreste y Noroeste. En esta última región, también fue alta la frecuencia del híbrido VNII-VNIV (aADa). La frecuencia de aislamiento de miembros del complejo C. gattii fue baja: se obtuvieron 3 aislados VGI (MATa) y 1 VGII (MATa) de las regiones Centro y Noroeste. En conclusión, este estudio muestra que las frecuencias de genotipos varían entre las distintas regiones de Argentina y señala la presencia de híbridos poco comunes en una frecuencia relativamente alta dentro de la región Noroeste. Contar con mayor número de estudios clínicos y ambientales regionales podría ayudar a elucidar si tales variaciones están asociadas a la existencia de nichos ecológicos particulares o a algún otro factor regional.

Frequency and geographical distribution of genotypes and mating types of Cryptococcus neoformans and Cryptococcus gattii species complexes in Argentina.

The aim of this work was to know the frequency and geographical distribution of genotypes and mating types of Cryptococcus neoformans and Cryptococcus gattii species complexes isolated from human infections in Argentina during the period from April 2009 to April 2011. A multicenter study was conducted, in which 372 isolates were obtained from 61 laboratories throughout the country. Of those, 98.8% of the isolates belonged to the C. neoformans species complex and 1.1% to the C. gattii species complex. Genotype VNI (MATα) was the most frequently isolated (n=326, 87.6%), followed by VNII (MATα) (n=22, 5.9%), the recently described VNII-VNIV (aADα) hybrid (n=14, 3.8%), VNIV (MATα) (n=4, 1.1%), VNIII (αADa) hybrid (n=1, 0.3%), and VNIII (αADα) hybrid (n=1, 0.3%). The Argentine Central region showed the greatest number of cases and genotype diversity. Interestingly, a relative high frequency was observed in genotype VNII (MATα) in the Cuyo, Northeast and Northwest regions and, also in VNII-VNIV (aADα) hybrids in the Northwest region. C. gattii species complex was isolated at a low rate; 3 VGI (MATα) and 1 VGII (MATα) isolates were obtained from the Northwest and Central regions. In conclusion, this study shows that genotype frequencies seem to vary among regions in Argentina and reveals a relatively high frequency of rare hybrids in the Northwest region. Further regional clinical and environmental studies may help to elucidate if those variations in frequencies are associated with the existence of regional ecological niches or any other regional factors.