Genetic diversity of polysporic isolates of Moniliophthora perniciosa (Tricholomataceae).

The causal agent of witches' broom disease, Moniliophthora perniciosa is a hemibiotrophic and endemic fungus of the Amazon basin and the most important cocoa disease in Brazil. The purpose of this study was to analyze the genetic diversity of polysporic isolates of M. perniciosa to evaluate the adaptation of the pathogen from different Brazilian regions and its association with different hosts. Polysporic isolates obtained previously in potato dextrose agar cultures of M. perniciosa from different Brazilian states and different hosts (Theobroma cacao, Solanum cernuum, S. paniculatum, S. lycocarpum, Solanum sp, and others) were analyzed by somatic compatibility grouping where the mycelium interactions were distinguished after 4-8 weeks of confrontation between the different isolates of M. perniciosa based on the precipitation line in the transition zone and by protein electrophoresis through SDS-PAGE. The diversity of polysporic isolates of M. perniciosa was grouped according to geographic proximity and respective hosts. The great genetic diversity of M. perniciosa strains from different Brazilian states and hosts favored adaptation in unusual environments and dissemination at long distances generating new biotypes.

Evaluation of biochemical and serological methods to identify and clustering yeast cells of oral Candida species by CHROMagar test, SDS-PAGE and ELISA.

The purpose of this work was to evaluate biochemical and serological methods to characterize and identify Candida species from the oral cavity. The strains used were five Candida species previously identified: C. albicans, C. guilliermondii, C. parapsilosis, C. krusei, C. tropicalis, and Kluyveromyces marxianus, as a negative control. The analyses were conducted through the SDS-PAGE associated with statistical analysis using software, chromogenic medium, and CHROMagar Candida (CA), as a differential medium for the isolation and presumptive identification of clinically important yeasts and an enzyme-linked immunoabsorbent assay (ELISA), using antisera produced against antigens from two C. albicans strains. This method enabled the screening of the three Candida species: C. albicans, C. tropicalis, and C. krusei, with 100% of specificity. The ELISA using purified immunoglobulin G showed a high level of cross-reaction against protein extracts of Candida species. The SDS-PAGE method allowed the clustering of species-specific isolates using the Simple Matching coefficient, S(SM) = 1.0. The protein profile analysis by SDS-PAGE increases what is known about the taxonomic relationships among oral yeasts. This methodology showed good reproducibility and allows collection of useful information for numerical analysis on information relevant to clinical application, and epidemiological and systematical studies.

Evaluation of biochemical and serological methods to identify and clustering yeast cells of oral Candida species by CHROMagar test, SDS-PAGE and ELISA

Este trabalho teve o propósito de avaliar métodos bioquímicos e sorológicos para serem aplicados na caracterização e identificação de linhagens do gênero Candida isoladas da cavidade bucal. As cepas empregadas representam cinco espécies de Candida previamente identificadas: C. albicans, C. guilliermondii, C. parapsilosis, C. krusei e C. tropicalis, utilizando como controle negativo Kluyveromyces marxianus. Foram empregadas as técnicas de gel de poliacrilamida (SDS-PAGE) associado à análise estatística em software, CHROMagar Candida (CA), meio cromogênico diferencial descrito para o isolamento e identificação presuntiva de leveduras de importância clínica e um ensaio de imunoabsorção ligado a enzima (ELISA), utilizando antissoro produzido contra extratos protéicos de uma linhagem-padrão de Candida e um isolado de cavidade oral de C. albicans. O método mostrou-se adequado para a identificação presuntiva de C. albicans, C. tropicalis e C. krusei, com 100% de sensibilidade e especificidade, com base na coloração e textura das colônias. O método de ELISA utilizando imunoglobulinas G purificadas apresentou alto teor de reação cruzada com as outras espécies de Candida estudadas. A análise do perfil protéico por SDS-PAGE permitiu agrupar os isolados da cavidade oral por intermédio do coeficiente "Simple Matching", SSM = 1,0. Os perfis protéicos analisados por SDS-PAGE ampliam os conhecimentos sobre as relaçäes taxonômicas de leveduras isoladas da cavidade oral. Esta metodologia demonstra boa reprodutibilidade e origina informaçäes úteis para aplicação clínica e estudos que envolvem a sistemática e a epidemiologia.