Dataset Information

Identification and classification of bcl genes and proteins of Bacillus cereus group organisms and their application in Bacillus anthracis detection and fingerprinting.

ABSTRACT: The Bacillus cereus group includes three closely related species, B. anthracis, B. cereus, and B. thuringiensis, which form a highly homogeneous subdivision of the genus Bacillus. One of these species, B. anthracis, has been identified as one of the most probable bacterial biowarfare agents. Here, we evaluate the sequence and length polymorphisms of the Bacillus collagen-like protein bcl genes as a basis for B. anthracis detection and fingerprinting. Five genes, designated bclA to bclE, are present in B. anthracis strains. Examination of bclABCDE sequences identified polymorphisms in bclB alleles of the B. cereus group organisms. These sequence polymorphisms allowed specific detection of B. anthracis strains by PCR using both genomic DNA and purified Bacillus spores in reactions. By exploiting the length variation of the bcl alleles it was demonstrated that the combined bclABCDE PCR products generate markedly different fingerprints for the B. anthracis Ames and Sterne strains. Moreover, we predict that bclABCDE length polymorphism creates unique signatures for B. anthracis strains, which facilitates identification of strains with specificity and confidence. Thus, we present a new diagnostic concept for B. anthracis detection and fingerprinting, which can be used alone or in combination with previously established typing platforms.

SUBMITTER: Leski TA

PROVIDER: S-EPMC2786505 | biostudies-literature | 2009 Nov

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Identification and classification of bcl genes and proteins of Bacillus cereus group organisms and their application in Bacillus anthracis detection and fingerprinting.

Leski Tomasz A TA Caswell Clayton C CC Pawlowski Marcin M Klinke David J DJ Bujnicki Janusz M JM Hart Sean J SJ Lukomski Slawomir S

Applied and environmental microbiology 20090918 22

The Bacillus cereus group includes three closely related species, B. anthracis, B. cereus, and B. thuringiensis, which form a highly homogeneous subdivision of the genus Bacillus. One of these species, B. anthracis, has been identified as one of the most probable bacterial biowarfare agents. Here, we evaluate the sequence and length polymorphisms of the Bacillus collagen-like protein bcl genes as a basis for B. anthracis detection and fingerprinting. Five genes, designated bclA to bclE, are pres ...[more]

PMID: 19767469

Similar Datasets

Project description:BACKGROUND:Bacillus cereus biovar anthracis (Bcbva) is an emergent bacterium closely related to Bacillus anthracis, the etiological agent of anthrax. The latter has a worldwide distribution and usually causes infectious disease in mammals associated with savanna ecosystems. Bcbva was identified in humid tropical forests of Côte d'Ivoire in 2001. Here, we characterize the potential geographic distributions of Bcbva in West Africa and B. anthracis in sub-Saharan Africa using an ecological niche modeling approach. METHODOLOGY/PRINCIPAL FINDINGS:Georeferenced occurrence data for B. anthracis and Bcbva were obtained from public data repositories and the scientific literature. Combinations of temperature, humidity, vegetation greenness, and soils values served as environmental variables in model calibrations. To predict the potential distribution of suitable environments for each pathogen across the study region, parameter values derived from the median of 10 replicates of the best-performing model for each pathogen were used. We found suitable environments predicted for B. anthracis across areas of confirmed and suspected anthrax activity in sub-Saharan Africa, including an east-west corridor from Ethiopia to Sierra Leone in the Sahel region and multiple areas in eastern, central, and southern Africa. The study area for Bcbva was restricted to West and Central Africa to reflect areas that have likely been accessible to Bcbva by dispersal. Model predicted values indicated potential suitable environments within humid forested environments. Background similarity tests in geographic space indicated statistical support to reject the null hypothesis of similarity when comparing environments associated with B. anthracis to those of Bcbva and when comparing humidity values and soils values individually. We failed to reject the null hypothesis of similarity when comparing environments associated with Bcbva to those of B. anthracis, suggesting that additional investigation is needed to provide a more robust characterization of the Bcbva niche. CONCLUSIONS/SIGNIFICANCE:This study represents the first time that the environmental and geographic distribution of Bcbva has been mapped. We document likely differences in ecological niche-and consequently in geographic distribution-between Bcbva and typical B. anthracis, and areas of possible co-occurrence between the two. We provide information crucial to guiding and improving monitoring efforts focused on these pathogens.

Project description:The conversion of ketomethiobutyrate to methionine has been previously examined in a number of organisms, wherein the aminotransferases responsible for the reaction have been found to be members of the Ia subfamily (L. C. Berger, J. Wilson, P. Wood, and B. J. Berger, J. Bacteriol. 183:4421-4434, 2001). The genome of Bacillus subtilis has been found to contain no subfamily Ia aminotransferase sequences. Instead, the analogous enzymes in B. subtilis were found to be members of the If subfamily. These putative aspartate aminotransferases, the yugH, ywfG, ykrV, aspB, and patA gene products, have been cloned, expressed, and characterized for methionine regeneration activity. Only YkrV was able to convert ketomethiobutyrate to methionine, and it catalyzed the reaction only when glutamine was used as amino donor. In contrast, subcellular homogenates of B. subtilis and Bacillus cereus utilized leucine, isoleucine, valine, alanine, phenylalanine, and tyrosine as effective amino donors. The two putative branched-chain aminotransferase genes in B. subtilis, ybgE and ywaA, were also cloned, expressed, and characterized. Both gene products effectively transaminated branched-chain amino acids and ketoglutarate, but only YbgE converted ketomethiobutyrate to methionine. The amino donor preference for methionine regeneration by YbgE was found to be leucine, isoleucine, valine, phenylalanine, and tyrosine. The B. subtilis ybgE gene is a member of the family III of aminotransferases and falls in a subfamily designated here IIIa. Examination of B. cereus and Bacillus anthracis genome data found that there were no subfamily IIIa homologues in these organisms. In both B. cereus and B. anthracis, two putative branched-chain aminotransferases and two putative D-amino acid aminotransferases were discovered as members of subfamily IIIb. These four sequences were cloned from B. cereus, expressed, and characterized. Only the gene product from the sequence designated Bc-BCAT2 was found to convert ketomethiobutyrate to methionine, with an amino donor preference of leucine, isoleucine, valine, phenylalanine, and tyrosine. The B. anthracis homologue of Bc-BCAT2 was also cloned, expressed, and characterized and was found to be identical in activity. The aminooxy compound canaline was found to be an uncompetitive inhibitor of B. subtilis YbgE and also inhibited growth of B. subtilis and B. cereus in culture.

Project description:BackgroundIt is possible to detect bacterial species in shotgun metagenome datasets through the presence of only a few sequence reads. However, false positive results can arise, as was the case in the initial findings of a recent New York City subway metagenome project. False positives are especially likely when two closely related are present in the same sample. Bacillus anthracis, the etiologic agent of anthrax, is a high-consequence pathogen that shares >99% average nucleotide identity with Bacillus cereus group (BCerG) genomes. Our goal was to create an analysis tool that used k-mers to detect B. anthracis, incorporating information about the coverage of BCerG in the metagenome sample.MethodsUsing public complete genome sequence datasets, we identified a set of 31-mer signatures that differentiated B. anthracis from other members of the B. cereus group (BCerG), and another set which differentiated BCerG genomes (including B. anthracis) from other Bacillus strains. We also created a set of 31-mers for detecting the lethal factor gene, the key genetic diagnostic of the presence of anthrax-causing bacteria. We created synthetic sequence datasets based on existing genomes to test the accuracy of a k-mer based detection model.ResultsWe found 239,503 B. anthracis-specific 31-mers (the Ba31 set), 10,183 BCerG 31-mers (the BCerG31 set), and 2,617 lethal factor k-mers (the lef31 set). We showed that false positive B. anthracis k-mers-which arise from random sequencing errors-are observable at high genome coverages of B. cereus. We also showed that there is a "gray zone" below 0.184× coverage of the B. anthracis genome sequence, in which we cannot expect with high probability to identify lethal factor k-mers. We created a linear regression model to differentiate the presence of B. anthracis-like chromosomes from sequencing errors given the BCerG background coverage. We showed that while shotgun datasets from the New York City subway metagenome project had no matches to lef31 k-mers and hence were negative for B. anthracis, some samples showed evidence of strains very closely related to the pathogen.DiscussionThis work shows how extensive libraries of complete genomes can be used to create organism-specific signatures to help interpret metagenomes. We contrast "specialist" approaches to metagenome analysis such as this work to "generalist" software that seeks to classify all organisms present in the sample and note the more general utility of a k-mer filter approach when taxonomic boundaries lack clarity or high levels of precision are required.

Dataset Information

Identification and classification of bcl genes and proteins of Bacillus cereus group organisms and their application in Bacillus anthracis detection and fingerprinting.

Publications

Identification and classification of bcl genes and proteins of Bacillus cereus group organisms and their application in Bacillus anthracis detection and fingerprinting.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets