Project description:An array analysis of C. gattii (C. bacillosporus), intended to identify loci associated with the hypervirulence of the Vancouver Island Outbreak (VIO). 23 C. gattii isolates, representing both VIO strains and control strains, were grown for 24 hours in mammalian macrophages. RNA was isolated and gene expression for each strain quantified relative to pooled RNA from all 23 samples. Linear regression was used to identify loci showing positive or negative correlation with "intracellular proliferation rate", a proxy measure for virulence. Data from this analysis is included in Ma et al, 2009, PNAS 106(31) 12980-12985. The abstract is included below. In 1999, the population of Vancouver Island, Canada, began to experience an outbreak of a fatal fungal disease caused by a highly virulent lineage of Cryptococcus gattii. This organism has recently spread to the Canadian mainland and Pacific Northwest, but the molecular cause of the outbreak remains unknown. Here we show that the Vancouver Island outbreak (VIO) isolates have dramatically increased their ability to replicate within macrophages of the mammalian immune system in comparison with other C. gattii strains. We further demonstrate that such enhanced intracellular parasitism is directly linked to virulence in a murine model of cryptococcosis, suggesting that this phenotype may be the cause of the outbreak. Finally, microarray studies on 24 C. gattii strains reveals that the hypervirulence of the VIO isolates is characterized by the up-regulation of a large group of genes, many of which are encoded by mitochondrial genome or associated with mitochondrial activities. This expression profile correlates with an unusual mitochondrial morphology exhibited by the VIO strains after phagocytosis. Our data thus demonstrate that the intracellular parasitism of macrophages is a key driver of a human disease outbreak, a finding that has significant implications for a wide range of other human pathogens. Dual-colour hybridization (each sample was hybridized against pooled RNA), one array per isolate, 23 biological samples, no technical replicates.
Project description:An array analysis of C. gattii (C. bacillosporus), intended to identify loci associated with the hypervirulence of the Vancouver Island Outbreak (VIO). 23 C. gattii isolates, representing both VIO strains and control strains, were grown for 24 hours in mammalian macrophages. RNA was isolated and gene expression for each strain quantified relative to pooled RNA from all 23 samples. Linear regression was used to identify loci showing positive or negative correlation with "intracellular proliferation rate", a proxy measure for virulence. Data from this analysis is included in Ma et al, 2009, PNAS 106(31) 12980-12985. The abstract is included below. In 1999, the population of Vancouver Island, Canada, began to experience an outbreak of a fatal fungal disease caused by a highly virulent lineage of Cryptococcus gattii. This organism has recently spread to the Canadian mainland and Pacific Northwest, but the molecular cause of the outbreak remains unknown. Here we show that the Vancouver Island outbreak (VIO) isolates have dramatically increased their ability to replicate within macrophages of the mammalian immune system in comparison with other C. gattii strains. We further demonstrate that such enhanced intracellular parasitism is directly linked to virulence in a murine model of cryptococcosis, suggesting that this phenotype may be the cause of the outbreak. Finally, microarray studies on 24 C. gattii strains reveals that the hypervirulence of the VIO isolates is characterized by the up-regulation of a large group of genes, many of which are encoded by mitochondrial genome or associated with mitochondrial activities. This expression profile correlates with an unusual mitochondrial morphology exhibited by the VIO strains after phagocytosis. Our data thus demonstrate that the intracellular parasitism of macrophages is a key driver of a human disease outbreak, a finding that has significant implications for a wide range of other human pathogens.
Project description:Proper mitochondrial function is critical for the ability of the fungal pathogen Cryptococcus neoformans to cause disease. Mitochondrial dysfunction of WT cells treated with antimycin A or L-DOPA, or on cells lacking Cir1 is evident from this dataset. These experiments revealed influences on transcript levels of genes encoding Fe-S cluster assembly components and the response to oxidative stress in Cryptococcus neoformans.
Project description:Opportunistic infections of the respiratory tract often succeed under a weakened immune response caused by an underlying illness or hospitalization. The human fungal pathogen, Cryptococcus neoformans, and the bacterial pathogen, Klebsiella pneumoniae, are both well-characterized microbes that cause severe infections within immunocompromised individuals. In this study, we simulate a concentration-dependent pulmonary coinfection of a bacterial and fungal pathogen, and profile the proteomic changes by DDA vs. DIA. Dual perspective profiling provides new insights into host defense regulation of infection and pathogenic mechanisms of invasion.
Project description:CircRNAs are a recently well-known regulator that mediates a variety of biological processes. Cryptococcus neoformans is an environmental fungal pathogen that can cause fatal cryptococcal meningitis in immunocompromised individuals. However, the involvement of circRNA in cryptococcal infection remains unclear. In this study, high‐throughput microarray was performed to identify the circRNA expression profile in cryptococcal meningitis patients.
Project description:Host gene expression responses can be used to determine the etiology of acute infection. PBMCs were stimulated with bacterial, viral, and fungal stimuli and then analyzed for differential gene expression utilizing microarrays to derive pathogen class-specific gene expression classifiers of infection. Discovery Cohort: In vitro human PBMC challenges were performed with bacteria (gram postive and gram negative), viruses (4 strains of influenza), and fungi (Cryptococcus and Candida)
Project description:Sexual reproduction facilitates infections and adaptations in Cryptococcus pathogens that cause fatal meningoencephalitis and have two distinct sexual cycles (bisexual and unisexual). Here, by constructing a gene-deletion strains library for transcription factor (TF) genes in Cryptococcus deneoformans, we explore spatiotemporal TF networks in sequential stages of mating community development and identified key regulators that determine processes specific to different reproductive modes. We show that the TFs crucial for bisex-specific syngamy coordinately induce the expression of a specific group of genes centered on the ancient mating determinants, which includes FMP1, a previously unidentified component of the conserved fungal mating pathway. Furthermore, we identify that a recently evolved regulatory cascade mediates the pre-meiotic autodiploidization process that defines Cryptococcus unisex, supporting that unisex is a recent evolutionary innovation. Our findings indicate that genetic circuits with different evolutionary ages govern the hallmark events distinguishing unisex and bisex, enabling reproductive flexibility that benefits Cryptococcus pathogenicity.
Project description:Mass spectrometry-based proteomics supports investigation into diverse biological systems, providing insights into regulation, signaling, modifications, and interactions. Optimized sample preparation is critical to ensure robust and reproducible analyses balanced with time and cost. In this article, we evaluated trypsin digestion parameters for the human fungal pathogen, Cryptococcus neoformans, and determined conditions for optimal peptide identification