Project description:Clavibacter michiganensis subsp. michiganensis is an important Gram-positive phytopathogenic bacteria that causes bacterial wilt and canker in tomato. The genome of the type strain, NCPPB382, has been sequenced and annotated, however comparative genomics suggests that certain regions are under- or misannotated. In order to improve the genome annotation, we have undertaken a proteogenomic study of this important pathogen. Samples were grown in culture and the proteome of the pellet and supernatant were analyzed separately using shotgun HPLC-MS/MS. These proteomics datasets were analyzed and a number of missing gene were found and a number of existing gene calls were modified.
Project description:The aims of this study were to present modifications to the annotations of the genome of C. posadasii, one of two closely related species of Coccidioides, a dimorphic fungal pathogen that causes coccidioidomycosis, also called Valley Fever. Proteins present in lysates and filtrates of in vitro grown mycelia and parasitic phase spherules from C. posadasii strain Silveira were analyzed using a GeLC-MS/MS method.
Project description:The proteome of the anaerobic bacterium Dehalococcoides mccartyi strain CBDB1 from the phylum Chloroflexi was investigated. D. mccartyi strain CBDB1 is a model organism for organohalide respiration where halogenated organic compounds serve as terminal electron acceptors. A wide range of halogenated organic compounds have been shown to be dehalogenated by the strain CBDB1. Therefore, D. mccartyi strain CBDB1 is a promising candidate for bioremediation application. Proteomic analysis of cultures grown with hexachlorobenzene as only electron acceptor resulted in identification of 8,491 distinct peptides which represents 1,023 proteins. A coverage of 70% of the 1,458 annotated proteins for strain CBDB1 was achieved. In addition, a spectral library was created from the annotated spectra. By using proteogenomics, 18 previously not annotated peptides were identified which contribute to four proteins previously not annotated and corrections in length of eight protein coding sequences.
Project description:Clostridium difficile is a gram-positive, spore-forming enteric anaerobe which can infect humans and a wide variety of animal species. Recently, the incidence and severity of human C. difficile infection has markedly increased. In this study, we evaluated the genomic content of 73 C. difficile strains isolated from humans, horses, cattle, and pigs by comparative genomic hybridization with microarrays containing coding sequences from C. difficile strains 630 and QCD-32g58. The sequenced genome of C. difficile strain 630 was used as a reference to define a candidate core genome of C. difficile and to explore correlations between host origins and genetic diversity. Approximately 16% of the genes in strain 630 were highly conserved among all strains, representing the core complement of functional genes defining C. difficile. Absent or divergent genes in the tested strains were distributed across the entire C. difficile 630 genome and across all the predicted functional categories. Interestingly, certain genes were conserved among strains from a specific host species, but divergent in isolates with other host origins. This information provides insight into the genomic changes which might contribute to host adaptation. Due to a high degree of divergence among C. difficile strains, a core gene list from this study offers the first step toward the construction of diagnostic arrays for C. difficile. Overall design: Dye-swap experiments with genomic DNA of tested and reference strains
Project description:Investigation of whole genome gene expression level changes in an Frh-deficient mutant, Frh overexpression mutant and TON_0282 deletion mutant, compared to the wild-type strain. In this study, as a first step to assess the role of the frhAGB-encoding hydrogenases and TON_0282 gene, we constructed three mutants by deleting each gene in a T. onnurineus NA1. Overall design: A custom microarray was manufactured by Roche NimbleGen. Unique six 60-mer oligonucleotides for all of the predicted 1,986 open reading frames (ORFs) in the annotated genome of T. onnurineus NA1 were designed and synthesized. Examined Wildtype, Frh-deficient mutant, Frh overexpression mutant and TON_0282 deletion mutant in T. onnurineus NA1 strain.