[CAP59 gene amplification in Cryptococcus neoformans and Cryptococcus gattii directly from a yeast suspension]. / Amplificación del gen CAP59 en Cryptococcus neoformans y Cryptococcus gattii directamente a partir de una suspensión de levaduras.

Cryptococcus is an encapsulated yeast of class Basidiomycetes, etiologic agent of cryptococcosis. Cryptococcosis is one of the most common opportunistic infections in immunosuppressed patients, although it can affect immunocompetent individuals. In recent years, the identification of medically important fungal species has been achieved through the amplification of specific regions or genes of fungal DNA by polymerase chain reaction (PCR). The CAP59 gene is involved in the synthesis of the capsule in Cryptococcus neoformans and Cryptococcus gattii, and is useful in the molecular identification of serotypes. In this research, we use yeasts of different serotypes from collection strains and C. neoformans isolates recovered from patients with cryptococcosis. A standardized yeast suspension from different Cryptococcus isolates as template allowed CAP59 gene amplification. This procedure was quick, simple, and inexpensive and required no PCR steps. This is important for taxonomic studies in laboratories with implemented molecular biology tools.

Phenotypic and genotypic identification of Candida dubliniensis from subgingival sites in immunocompetent subjects in Argentina.

INTRODUCTION: It is generally recognized that Candida dubliniensis is commonly found in immunocompromised patients, such as those with advanced human immunodeficiency virus infection, at sites of periodontal disease. Since there are no data available for Argentina, the aim of this study was to determine the prevalence of and to identify C. dubliniensis in periodontal pockets from immunocompetent subjects living in Buenos Aires, Argentina, through a comparison of phenotypic and molecular assays. METHODS: Yeasts recovered from subgingival plaque samples were studied for 180 immunocompetent non-smoking patients with periodontal disease. Yeasts were identified by conventional mycological methods and by specific polymerase chain reaction (PCR) assay. Fluconazole and voriconazole susceptibility studies were performed in keeping with the Clinical and Laboratory Standards Institute. RESULTS: Among 76 yeasts isolated, C. dubliniensis comprised 10.5% (n = 8; 95% confidence interval 4.7-19.7), which corresponded to 4.4% of patients studied (8/180). C. albicans was the most frequently isolated species of yeast. A great majority of C. dubliniensis isolates was susceptible with only one isolate resistant to both antifungals. CONCLUSION: Micromorphology on Staib agar was the phenotypic method that was most concordant with PCR and it was useful for selecting presumptive C. dubliniensis. This is the first report to use PCR to identify C. dubliniensis in subgingival fluid from immunocompetent individuals with periodontal disease in Argentina. On the basis of the findings presented here, we confirm that C. dubliniensis can colonize periodontal pockets of immunocompetent patients with periodontal disease.

[Comparison of different methods of DNA extraction from blood to detect fungal DNA by PCR]. / Comparación de métodos de extracción de ADN de sangre para detectar ADN fúngico por PCR.

Comparison of different methods of DNA extraction from blood to detect fungal DNA by PCR. Invasive fungal infections (IFI) are associated with high mortality by reaching levels of 50%, and also with a significant failure in antifungical treatments. This fact mostly obeys to difficulties in obtaining a fast and accurate mycologic diagnosis due to the low sensitivity of conventional methods, mainly in neutropenic and AIDS patients. Various methods based on fungal DNA study are currently being used for the diagnosis of mycotic infections. We herein evaluated two procedures of extraction and purification of fungal DNA in blood for their use in PCR detection. Both of them showed equal efficiency in obtaining high performance DNA with universal primers ITS land ITS 4 as target.

Comparación de métodos de extracción de ADN de sangre para detectar ADN fúngico por PCR / Comparison of different methods of DNA extraction from blood to detect fungal DNA by PCR

La infección fúngica invasora (IFI) está asociada a un alto índice de mortalidad, que alcanza el 50% debido a la frecuente falla en el tratamiento antifúngico. Existen dificultades para realizar un diagnóstico micológico rápido y certero dada la baja sensibilidad de los métodos convencionales, especialmente en pacientes neutropénicos y con SIDA. Numerosos métodos para diagnosticar infecciones micóticas basados en el estudio del ADN fúngico están actualmente en desarrollo. Nosotros evaluamos la utilidad de dos procedimientos de extracción y purificación del ADN fúngico presente en sangre para su posterior detección por PCR. Ambos métodos resultaron igualmente eficientes para obtener ADNs de óptima calidad y para realizar la técnica de PCR con los iniciadores universales para hongos ITS 1 e ITS 4.

Invasive fungal infections (IFI) are associated with high mortality by reaching levels of 50%, and also with a significant failure in antifungical treatments. This fact mostly obeys to difficulties in obtaining a fast and accurate mycologic diagnosis due to the low sensitivity of conventional methods, mainly in neutropenic and AIDS patients. Various methods based on fungal DNA study are currently being used for the diagnosis of mycotic infections. We herein evaluated two procedures of extraction and purification of fungal DNA in blood for their use in PCR detection. Both of them showed equal efficiency in obtaining high performance DNA with universal primers ITS 1and ITS 4 as target.