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Abstract We report the first case of cryptococcosis due to Cryptococcus decagattii in an immunocompetent pediatric patient from an indigenous community in Argentina with a successful outcome. Two isolates (blood, cerebrospinal fluid) were genotyped by restriction fragment length polymorphism of the orotidine monophosphate pyrophosphorylase (URA5) gene as VGIV and identified by multi-locus sequence typing as C. decagattii. Matrix-assisted laser desorption/ionization time of flight mass spectrometry identification indicated genotype VGIII. The minimum inhibitory concentration of amphotericin B, fluconazole, itraconazole, and voriconazole was determined (cerebrospinal fluid: 0.25, 16, 0.12, and 0.12, blood: 0.25, 4, 0.12, and 0.06, respectively, all in mg/L).

As the distribution of Candida species and their susceptibility to antifungal agents have changed, a new means of accurately and rapidly identifying these species is necessary for the successful early resolution of infection and the subsequent reduction of morbidity and mortality. The current work aimed to evaluate ribosomal RNA gene sequencing for the identification of medically relevant Candida species in comparison with a standard phenotypic method. Eighteen reference strains (RSs), 69 phenotypically identified isolates and 20 inconclusively identified isolates were examined. Internal transcribed spaces (ITSs) and D1/D2 of the 26S ribosomal RNA gene regions were used as targets for sequencing. Additionally, the sequences of the ITS regions were used to establish evolutionary relationships. The sequencing of the ITS regions was successful for 88% (94/107) of the RS and isolates, whereas 100% of the remaining 12% (13/107) of the samples were successfully analysed by sequencing the D1/D2 region. Similarly, genotypic analysis identified all of the RS and isolates, including the 20 isolates that were not phenotypically identified. Phenotypic analysis, however, misidentified 10% (7/69) of the isolates. Phylogenetic analysis allowed the confirmation of the relationships between evolutionarily close species. Currently, the use of genotypic methods is necessary for the correct identification of Candida species.

As the distribution of Candida species and their susceptibility to antifungal agents have changed, a new means of accurately and rapidly identifying these species is necessary for the successful early resolution of infection and the subsequent reduction of morbidity and mortality. The current work aimed to evaluate ribosomal RNA gene sequencing for the identification of medically relevant Candida species in comparison with a standard phenotypic method. Eighteen reference strains (RSs), 69 phenotypically identified isolates and 20 inconclusively identified isolates were examined. Internal transcribed spaces (ITSs) and D1/D2 of the 26S ribosomal RNA gene regions were used as targets for sequencing. Additionally, the sequences of the ITS regions were used to establish evolutionary relationships. The sequencing of the ITS regions was successful for 88% (94/107) of the RS and isolates, whereas 100% of the remaining 12% (13/107) of the samples were successfully analysed by sequencing the D1/D2 region. Similarly, genotypic analysis identified all of the RS and isolates, including the 20 isolates that were not phenotypically identified. Phenotypic analysis, however, misidentified 10% (7/69) of the isolates. Phylogenetic analysis allowed the confirmation of the relationships between evolutionarily close species. Currently, the use of genotypic methods is necessary for the correct identification of Candida species.

The Mycology Department of the Instituto Nacional de Enfermedades Infecciosas "Dr. C. Malbrán", conducted the Second National Multicenter Survey on Fungemia due to Yeasts in Argentina. The aim was to obtain updated data of the frequency of the causative species encountered and their in vitro susceptibility to seven antifungal agents. Yeast species were identified by micromorphological and biochemical studies. Antifungal susceptibility testing was performed by the reference microdilution method E.Def 7.1 of the European Committee on Antibiotic Susceptibility Testing (EUCAST). A total of 461 viable yeasts were identified. The most frequent species were: Candida albicans (38.4 %), Candida parapsilosis (26 %), Candida tropicalis (15.4 %) and Candida glabrata (4.3 %). Other uncommon species, such as Candida viswanathii (0.6 %), Candida haemulonii (0.4 %), Candida inconspicua (0.2 %) and Candida fermentati (0.2 %) were also isolated. Among the Candida spp., 5.4 % and 1.6 % were resistant to fluconazole and voriconazole, respectively. Itraconazole and caspofungin were the most efficient agents against all Candida spp. tested (MIC < 1 mg/l). For anidulafungin, 21.6 % of C. parapsilosis showed a MIC value of 4 mg/l. Fluconazole was less active against 53.1 % of Cryptococcus neoformans (MIC > 8 mg/l), 75 % of Trichosporon spp., and 100 % of Rhodotorula spp., Geotrichum candidum, Saccharomyces cerevisiae. The global percentage of mortality was 20 %. The presence of uncommon species reinforces the need for performing continuous laboratory surveillance in order to monitor possible changes, not only in the epidemiological distribution of species, but also in the resistance to antifungal drugs.Distribución de especies y perfil de sensibilidad de levaduras aisladas de hemocultivos: resultados de un estudio multicéntrico de vigilancia de laboratorio en Argentina. El Departamento Micología del Instituto Nacional de Enfermedades Infecciosas "Dr. Carlos G. Malbrán" condujo el segundo estudio multicéntrico nacional sobre funge- mias debidas a levaduras. El objetivo fue obtener datos actualizados sobre la distribución de especies y la sensibilidad in vitro frente a siete antifúngicos. Las levaduras fueron identificadas mediante el estudio de la micromorfología y la realización de pruebas bioquímicas. La determinación de la sensibilidad se realizó según el método de referencia E.Def 7.1 del European Committee on Antibiotic Susceptibility Testing (EUCAST). Se identificaron 461 levaduras. Las especies más frecuentes fueron Candida albicans (38,4 %), Candida parapsilosis (26 %), Candida tropicalis (15,4 %) y Candida glabrata (4,3 %). Se aislaron otras especies menos comunes, como Candida viswanathii (0,6 %), Candida haemulonii (0,4 %), Candida inconspicua (0,2 %) y Candida fermentati (0,2 %). Entre las especies del género Candida, el 5,4 % y el 1,6 % fueron resistentes al fluconazol y al voriconazol, respectivamente. El itraconazol y la caspofungina fueron los antifúngicos más eficaces in vitro frente a las especies de Candida evaluadas (CIM < 1 mg/l). Para la anidulafungina, el 21,6 % de los aislamientos de C. parapsilosis mostraron una CIM de 4 mg/l. El fluconazol fue menos activo para el 53,1 % de los aislamientos de Cryptococcus neoformans (CIM > 8 mg/l), el 75 % de los aislamientos de Trichosporon spp. y el 100 % de los aislamientos de Rhodotorula spp., Geotrichum candidum y Saccharomyces cerevisiae. El porcentaje de mortalidad fue del 20 %. La presencia de especies infrecuentes refuerza la necesidad de realizar la continua vigilancia de laboratorio con el fin de monitorear posibles cambios, no solo en la epidemiología de las especies causantes de fungemia, sino también en la resistencia a los antifúngicos.

Los métodos de referencia E. Def 7.1 y M27-A3, que detectan resistencia in vitro a los antifúngicos, son onerosos y muy laboriosos, por lo que su implementación en los laboratorios hospitalarios es limitada. Existen técnicas comerciales de simple realización, que permitirían obtener resultados comparables a los que se obtienen con los métodos estándares. Los objetivos de esta investigación fueron: a) comparar los resultados de concentración inhibitoria mínima obtenidos según el método de referencia E.Def 7.1 con los obtenidos mediante el empleo del equipo comercial ATB® Fungus 3 en un conjunto de 82 aislamientos clínicos de Candida spp. frente a los siguientes antifúngicos: anfotericina B, 5-fluorocitosina, fluconazol e itraconazol; b) comparar en ese mismo conjunto de aislamientos los resultados del estudio de sensibilidad al fluconazol por difusión en agar empleando tabletas Neo-SensitabsTM o discos Malbrán con los que se obtienen por el método de referencia. La concordancia general entre el método de referencia y el ATB® Fungus 3 fue del 90,2 %, mientras que la concordancia del método de referencia con los métodos por difusión con discos y con tabletas alcanzó el 96,3% y el 92,7 %, respectivamente. El ATB® Fungus 3 fue eficaz para determinar la sensibilidad a la anfotericina B y a la 5-fluorocitosina, pero se observaron discrepancias al evaluar la sensibilidad a los azoles. Los métodos por difusión resultaron útiles para determinar la sensibilidad al fluconazol; sin embargo, observamos 3 discrepancias muy mayores, 1 mayor y 2 menores con el método de difusión con tabletas, mientras que con los discos solo se produjeron 3 discrepancias menores.Reference methods E.Def 7.1 and M27-A3 detect in vitro resistance; however, they are expensive and very laborious. Thus, their actual use in hospital laboratories is limited. There are commercial techniques available, having easier accessibility and development, which would yield results comparable to those of the reference methods. The objectives of this study were: a) to compare the results of minimal inhibitory concentration of 82 Candida spp. clinic isolates according to reference method E.Def 7.1 and ATB® Fungus 3; b) to compare the results of fluconazole susceptibility testing by disk diffusion in agar with Neo-SensitabsTM tablets and Malbrán disks with those of the reference methods. Minimal inhibitory concentration for amphotericin B, 5-flucytosine, fluconazole and itraconazole was performed according to the E.Def 7.1 and the ATB Fungus 3 methods and diffusion was carried out with fluconazole disks and tablets. General concordance between the reference method and ATB Fungus 3 was 90.2 % and 96.3 and 92.7% for diffusion with disks and tablets. The ATB Fungus 3 method was effective to determine susceptibility against amphotericin B and 5-flucytosine; however, discrepancies were observed with azole drugs. Disk diffusion methods are useful to determine susceptibility to fluconazole; however, 3 very major, 1 major and 2 minor errors were observed with the tablets, whereas only 3 minor errors were observed with the disks.