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A randomly amplified polymorphic DNA marker specific for the Bacillus cereus group is diagnostic for Bacillus anthracis.

ABSTRACT: Aiming to develop a DNA marker specific for Bacillus anthracis and able to discriminate this species from Bacillus cereus, Bacillus thuringiensis, and Bacillus mycoides, we applied the randomly amplified polymorphic DNA (RAPD) fingerprinting technique to a collection of 101 strains of the genus Bacillus, including 61 strains of the B. cereus group. An 838-bp RAPD marker (SG-850) specific for B. cereus, B. thuringiensis, B. anthracis, and B. mycoides was identified. This fragment included a putative (366-nucleotide) open reading frame highly homologous to the ypuA gene of Bacillus subtilis. The restriction analysis of the SG-850 fragment with AluI distinguished B. anthracis from the other species of the B. cereus group.

SUBMITTER: Daffonchio D

PROVIDER: S-EPMC91177 | biostudies-literature | 1999 Mar

REPOSITORIES: biostudies-literature

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A randomly amplified polymorphic DNA marker specific for the Bacillus cereus group is diagnostic for Bacillus anthracis.

Daffonchio D D Borin S S Frova G G Gallo R R Mori E E Fani R R Sorlini C C

Applied and environmental microbiology 19990301 3

Aiming to develop a DNA marker specific for Bacillus anthracis and able to discriminate this species from Bacillus cereus, Bacillus thuringiensis, and Bacillus mycoides, we applied the randomly amplified polymorphic DNA (RAPD) fingerprinting technique to a collection of 101 strains of the genus Bacillus, including 61 strains of the B. cereus group. An 838-bp RAPD marker (SG-850) specific for B. cereus, B. thuringiensis, B. anthracis, and B. mycoides was identified. This fragment included a putat ...[more]

PMID: 10049896

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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.

Dataset Information

A randomly amplified polymorphic DNA marker specific for the Bacillus cereus group is diagnostic for Bacillus anthracis.

Publications

A randomly amplified polymorphic DNA marker specific for the Bacillus cereus group is diagnostic for Bacillus anthracis.

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