RESUMO
Cryptococcus gattii causes life-threatening infection of the pulmonary and central nervous systems in hosts with normal immunity and traditionally has been considered to be restricted geographically to tropical and subtropical climates. The recent outbreak of C. gattii in the temperate climate of Vancouver Island, BC, Canada, led to a collaborative investigation. The objectives of the current study were to ascertain the environmental source of the outbreak infections, survey the molecular types of the outbreak and environmental cryptococcal isolates, and determine the extent of genetic diversity among the isolates. PCR-fingerprinting and amplified fragment length polymorphism (AFLP) were used to examine the genotypes, and mating assays were performed to determine the mating type of the isolates. All outbreak and environmental isolates belonged to C. gattii. Concordant results were obtained by using PCR-fingerprinting and AFLP analysis. The vast majority of clinical and veterinary infections were caused by isolates of the molecular type VGII/AFLP6, but two were caused by molecular type VGI/AFLP4. All environmental isolates belonged to molecular type VGII/AFLP6. Two or three subtypes were observed within VGII/AFLP6 among outbreak and environmental isolates. All mating-competent isolates were of the alpha-mating type. The emergence of this usually tropical pathogen on Vancouver Island highlights the changing distribution of this genotype and emphasizes the importance of an ongoing collaborative effort to monitor the global epidemiology of this yeast.
Assuntos
Criptococose/epidemiologia , Criptococose/microbiologia , Cryptococcus/genética , Surtos de Doenças , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Colúmbia Britânica , Criança , Cryptococcus/isolamento & purificação , Exposição Ambiental , Feminino , Variação Genética , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Epidemiologia MolecularRESUMO
The STE12alpha gene of Cryptococcus neoformans encodes a protein containing both homeodomain and zinc finger regions. As homeodomains and zinc finger regions are important domains for the function of many transcription factors, we used site-specific mutagenesis to delineate the roles of these two domains. The homeodomain and zinc finger regions are each important for the function of Ste12alphap. DNA binding ability, mating frequency, and haploid fruiting capability were reduced in strains with mutations in the homeodomain, whereas virulence and capsule size in the mouse brain were increased. In contrast, mutations in the zinc fingers region resulted in decreased virulence, reduced capsule size in the mouse brain and decreased gene expression of capsule associated genes. In addition, phospholipase activity was increased in the zinc finger mutants. Taken together, most of the phenotypes previously observed in the ste12alpha deletion strains were reproduced in these two types of mutants. However, unlike mutations in the homeodomain/zinc finger region, complete deletion of STE12alpha caused a severe reduction in virulence and a decrease in phospholipase activity. These data suggest that region(s) other than the homeodomain and zinc finger regions of Ste12alphap contribute to the variable influences on the different phenotypes observed in C. neoformans.
Assuntos
Cryptococcus neoformans/metabolismo , Proteínas Fúngicas , Proteínas de Homeodomínio , Fatores de Transcrição , Dedos de Zinco , Animais , Encéfalo/metabolismo , Cryptococcus neoformans/genética , Cryptococcus neoformans/crescimento & desenvolvimento , Cryptococcus neoformans/patogenicidade , Feminino , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Regulação Fúngica da Expressão Gênica , Proteínas de Homeodomínio/química , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Mutagênese Sítio-Dirigida , Fenótipo , Taxa de Sobrevida , Fatores de Transcrição/química , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Técnicas do Sistema de Duplo-HíbridoRESUMO
Under appropriate conditions, haploid Cryptococcus neoformans cells can undergo a morphological switch from a budding yeast form to develop hyphae and viable basidiospores, which resemble those produced by mating. This process, known as haploid fruiting, was previously thought to occur only in MATalpha strains. We identified two new strains of C. neoformans var. neoformans serotype D that are MATa type and are able to haploid fruit. Further, a MATa reference strain, B-3502, also produced hyphae and fruited after prolonged incubation on filament agar. Over-expression of STE12a dramatically enhanced the ability of all MATa strains tested to filament. Segregation analysis of haploid fruiting ability confirmed that haploid fruiting is not MATalpha-specific. Our results indicate that MATa cells are intrinsically able to haploid fruit and previous observations that they do not were probably biased by the examination of a small number of genetically related isolates that have been maintained in the laboratory for many years.
