Project description:The goal of this study is the discovery of (a) meaningful phylogenomic relationships among members of this B. cereus/B. anthracis group, and (b) reliable gene-phenotype associations, e.g. recognition of links between genomic traits and the ability of certain strains to cause various forms of disease. We also tried to elucidate genome evolution aspects that may lead to the emergence of variants that are capable (or have the potential) of causing anthrax-like disease. This large-scale comparative genomics approach is unprecedented for this taxonomic group. Dr. A. Hoffmaster (CDC) provided the PFGRC with 73 B. cereus and B. anthracis isolates from the CDC culture collection. Of these, 27 were isolated from patients with severe or systemic disease; ten isolates of this group were obtained from patients (welding factory workers) with anthrax-like disease or from the environment near their workplace. Another set of 26 represented isolates from food-born illnesses. Of the 26 gastrointestinal disease isolates (GIDI), 10 were obtained from patients with diarrhea, whereas another set of 10 had been shown to harbor the emetic (vomit) toxin gene by PCR. The rest of the group consisted of 20 isolates with various phenotypes. All strains were screened for their genomic content using the B. cereus/B. anthracis species microarray.
Project description:Bacillus cereus is the second leading cause of collective food poisoning in France. B. cereus is also associated with severe clinical infections leading to patient death in 10% of the cases. The emergence of B. cereus as a foodborne and opportunistic pathogen has intensified the need to distinguish strains of public health concern. In this work, by performing a screen on a large collection of B. cereus strains of varying pathogenic potential, we identified genetic determinants capable of discriminating B. cereus strains inducing negative clinical outcomes. The combination of 4 biomarkers is sufficient to accurately discern clinical strains from harmless strains. Three of the biomarkers are located on the chromosome, with a fourth one identifying a plasmid carried by most pathogenic strains. A 50 kbp region of this plasmid promotes the virulence potential of these strains and could thus be defined as a new pathogenicity island of B. cereus. These new findings help in the understanding of B. cereus pathogenic potential and complexity and may provide tools for a better assessment of the risks associated with B. cereus contamination to improve patient health and food safety.
Project description:The aim of the study was to carry out a CGH study utilizing a set of 39 diverse Bacillus isolates. Thirty four B. cereus and five B. anthracis strains and isolates were chosen so as to represent different lineages based on previous characterizations, including MLEE and MLST (Helgason, Okstad et al. 2000; Helgason, Tourasse et al. 2004). They represent the spectrum of B. cereus phenotypic diversity by including soil, dairy and periodontal isolates in addition to virulent B. anthracis strains.
Project description:The goal of this study is the discovery of (a) meaningful phylogenomic relationships among members of this B. cereus/B. anthracis group, and (b) reliable gene-phenotype associations, e.g. recognition of links between genomic traits and the ability of certain strains to cause various forms of disease. We also tried to elucidate genome evolution aspects that may lead to the emergence of variants that are capable (or have the potential) of causing anthrax-like disease. This large-scale comparative genomics approach is unprecedented for this taxonomic group. Dr. A. Hoffmaster (CDC) provided the PFGRC with 73 B. cereus and B. anthracis isolates from the CDC culture collection. Of these, 27 were isolated from patients with severe or systemic disease; ten isolates of this group were obtained from patients (welding factory workers) with anthrax-like disease or from the environment near their workplace. Another set of 26 represented isolates from food-born illnesses. Of the 26 gastrointestinal disease isolates (GIDI), 10 were obtained from patients with diarrhea, whereas another set of 10 had been shown to harbor the emetic (vomit) toxin gene by PCR. The rest of the group consisted of 20 isolates with various phenotypes. All strains were screened for their genomic content using the B. cereus/B. anthracis species microarray. Seventy-three query strains were investigated in this study, with each query strain hybridized against the reference strain, Sterne. Dye-swap experiments were performed with all the 73 strains on both chipA and chipB of the microarray, for a total of four or more hybridizations per query strain. Each 70mer oligo spotted on the B. cereus species microarray is spotted once. Positive controls on the array consist of oligos designed from the sequenced reference genome, Sterne, and negative controls on the array consist of oligos designed from the thale cress plant, Arabidopsis thaliana.
Project description:Bacillus cereus Hemolysin BL is a tripartite toxin responsible for a diarrheal type of food poisoning.We found distinct complexes of up to 600 kDa on native gels, whose structure and size were primarily defined by Hbl B. Hbl L1 and L2 were also identified in these complexes using Western blotting and an LC-MS approach. LC-MS also revealed that many other proteins secreted by B. cereus exist in complexes.
Project description:The alternative sigma factor sigmaB has an important role in the acquisition of stress-resistance in many gram-positive bacteria. In the foodborne pathogen Bacillus cereus sigmaB is activated strongly upon a heat shock and other stress conditions. Here we describe the identification of the set of sigmaB-regulated genes in B. cereus by DNA microarray analysis of the transcriptome upon a mild heat shock. A total of 24 genes could be identified as being sigmaB-dependent as witnessed by (i) significantly lower expression-levels of these genes in mutants deleted for sigB and rsbY (encoding the alternative sigma factor sigmaB and a PP2C phosphatase that acts as a crucial positive regulator of sigmaB-activity, respectively) than in the parental strain B. cereus ATCC 14579, and (ii) increased expression of these genes upon a heat shock. Newly identified members of the sigmaB-regulon of B. cereus include an ECF-sigma factor (sigmaZ), a histidine kinase and two genes that have predicted functions in spore germination. Our data indicate that the sigmaB-regulon of B. cereus is considerably smaller than that of other gram-positive bacteria. This appears to be in line with phylogenetic analyses of sigmaB in which sigmaB in the B. cereus group was suggested to be close to the ancestral form of sigmaB in gram-positive bacteria. Keywords: stress response, comparative genomics
Project description:Pathogenic species belonging to Bacillus cereus sensu lato group possess a high evolutionary advantage in the environment and in food matrices thanks to their capacity to survive as silent spores to harsh environmental insults and grow at relatively low temperatures. Ready to re-heat products are at severe risk for contamination by members of Bacillus cereus s.l. group if not stored at proper conditions. In this work, the goal was to assess, by means of a genome-wide transcriptional assay, the isolated strain Bacillus cereus UC10070 gene expression behind the process of spore germination and consequent outgrowth in an artificially contaminated vegetable-based food model. A vegetable food model subjected to a heat treatment was determined to present favourable conditions for spores germination. Microscopic analyses together with OD measurements were applied to select the key steps of B. cereus cell cycle to be used for the microarray analysis. Using this approach we found a total of 1,646 probe sets differentially expressed and modulated during the entire B. cereus life cycle in the vegetable foodstuff.