RESUMO
Using a Tn7 transposon library of Candida albicans, we have identified a mutant that exhibited sensitivity in drop plate assays to oxidants such as menadione and hydrogen peroxide. To verify the role of the mutated gene in stress adaptation, null mutants were constructed and phenotypically characterized. Because of its apparent functions in growth and oxidant adaptation, we have named the gene GOA1. Goa1p appears to be unique to the CTG subclade of the Saccharomycotina, including C. albicans. Mutants of C. albicans lacking goa1 (strain GOA31) were more sensitive to 6 mM H(2)O(2) and 0.125 mM menadione than the wild type (wt) or a gene-reconstituted (GOA32) strain. The sensitivity to oxidants correlated with reduced survival of the GOA31 mutant in human neutrophils and avirulence compared to control strains. Other phenotypes of GOA31 include reduced growth and filamentation in 10% serum, Spider, and SLAD agar media and an inability to form chlamydospores. Since Goa1p has an N-terminal mitochondrion localization site, we also show that green fluorescent protein-tagged Goa1p shows a mitochondrionlike distribution during oxidant or osmotic stress. Further, the inability of GOA31 to grow in medium containing lactate, ethanol, or glycerol as the sole carbon source indicates that the mitochondria are defective in the mutant. To determine how Goa1p contributes to mitochondrial function, we compared the wt, GOA32, and GOA31 strains for mitochondrial electrical membrane potential, respiration, and oxidative phosphorylation. We now show that GOA31, but not the wt or GOA32, had decreased respiration and mitochondrial membrane potential such that mutant cells are unable to drive oxidative phosphorylation. This is the first report in C. albicans of a respiratory defect caused by a loss of mitochondrial membrane potential.
Assuntos
Candida albicans/fisiologia , Candida albicans/patogenicidade , Candidíase/microbiologia , Proteínas Fúngicas/metabolismo , Mitocôndrias/metabolismo , Animais , Candida albicans/genética , Candidíase/imunologia , Células Cultivadas , Feminino , Proteínas Fúngicas/genética , Humanos , Potencial da Membrana Mitocondrial , Camundongos , Camundongos Endogâmicos BALB C , Mitocôndrias/genética , Neutrófilos/imunologia , Neutrófilos/microbiologia , Transporte Proteico , Estresse Fisiológico , VirulênciaRESUMO
The relative performance of 2 widely used reverse transcription polymerase chain reaction (RT-PCR) assays, the Focus diagnostics Simplexa™ Flu A/B & RSV kit (nucleic acid extraction-dependent assay) and the Prodessa Proflu+™ assay, was evaluated using 735 prospectively and retrospectively collected nasopharyngeal swab specimens. Overall, the assays showed positive and negative agreements of 100% and 99.7% for influenza A, 98.1% and 99.9% for influenza B, and 99.3% and 99.5% for respiratory syncytial virus. The relative analytical sensitivity of the 2 assays was also similar.
Assuntos
Vírus da Influenza A/genética , Vírus da Influenza B/genética , Influenza Humana/virologia , Técnicas de Diagnóstico Molecular/métodos , Infecções por Vírus Respiratório Sincicial/virologia , Vírus Sinciciais Respiratórios/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Influenza Humana/diagnóstico , Masculino , Pessoa de Meia-Idade , Nasofaringe/virologia , Infecções por Vírus Respiratório Sincicial/diagnóstico , Estudos Retrospectivos , Sensibilidade e Especificidade , Adulto JovemRESUMO
Regulation of hyphal morphogenesis in Candida albicans can occur through quorum sensing (QS). A QS signal, farnesol, is produced during high-density growth and inhibits morphogenesis. However, the signal transduction pathway that regulates QS is unknown. Here, we show that a C. albicans mutant lacking Chk1p but not either the Sln1p or the Nik1p histidine kinase is refractory to the inhibitory effect of farnesol both in cell suspension and during the formation of a biofilm. This study is the first to demonstrate a role for a two-component signal transduction protein in QS by a eukaryotic organism.