To provide new insights into the population and genomic structure of the Bacillus cereus group of bacteria. The genetic relatedness among B. cereus group strains was assessed by multilocus sequence ...typing (MLST) using an optimized scheme based on seven chromosomal housekeeping genes. A set of 48 strains from different clinical sources was included, and six clonal complexes containing several genetically similar isolates from unrelated patients were identified. Interestingly, several clonal groups contained strains that were isolated from similar human sources. Furthermore, comparative whole genome sequence analysis of 16 strains led to the discovery of novel ubiquitous genome features of the B. cereus group, such as atypical group II introns, IStrons, and hitherto uncharacterized repeated elements. The B. cereus group constitutes a coherent population unified by the presence of ubiquitous and specific genetic elements which do not show any pattern, either in their sequences or genomic locations, which allows to differentiate between the member species of the group. Nevertheless, the population is very dynamic, as particular lineages of clinical origin can evolve to form clonal complexes. At the genome level, the dynamic behaviour is indicated by the presence of numerous mobile and repeated elements. The B. cereus group of bacteria comprises species that are of medical and economic importance. The MLST data, along with the primers and protocols used, will be available in a public, web-accessible database ( http://mlstoslo.uio.no).
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Members of the Bacillus cereus group (B. anthracis, B. cereus, B. mycoides and B. thuringiensis ) are well‐known pathogens of mammals (B. anthracis and B. cereus ) and insects (B. thuringiensis ). ...The specific diseases they cause depend on their capacity to produce specific virulence factors, such as the lethal toxin of B. anthracis and the Cry toxins of B. thuringiensis. However, these Bacillus spp. also produce a variety of proteins, such as phospholipases C, which are known to act as virulence factors in various pathogenic bacteria. Few genes encoding these virulence factors have been characterized in pathogenic Bacillus spp. and little is known about the regulation of their expression. We had previously reported that in B. thuringiensis expression of the phosphatidylinositol‐specific phospholipase C gene is regulated by the transcriptional activator PlcR. Here we report the identification of several extracellular virulence factor genes by the virtue of their PlcR‐regulated expression. These PlcR‐regulated genes encode degradative enzymes, cell‐surface proteins and enterotoxins. The PlcR‐regulated genes are widely dispersed on the chromosome and therefore do not constitute a pathogenic island. Analysis of the promoter region of the PlcR‐regulated genes revealed the presence of a highly conserved palindromic region (TATGNAN4TNCATA), which is presumably the specific recognition target for PlcR activation. We found that the plcR gene is also present in and probably restricted to all the members of the B. cereus group. However, although the polypeptide encoded by the B. cereus plcR gene is functionally equivalent to the B. thuringiensis regulator, the polypeptide encoded by the B. anthracis gene is truncated and not active as a transcriptional activator. PlcR is the first example described of a pleiotropic regulator involved in the control of extracellular virulence factor expression in pathogenic Bacillus spp. These results have implications for the taxonomic relationships among members of the B. cereus group, the virulence properties of these bacteria and the safety of B. thuringiensis‐based biopesticides.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Aims: The aim of this research was to isolate and characterize an antimicrobial substance from the Bacillus cereus type strain ATCC 14579.
Methods and Results: A substance with antimicrobial ...activity was isolated from B. cereus ATCC 14579. The substance was produced during late exponential growth and well into the stationary phase with a maximum 9 h after inoculation. The inhibitory substance was purified by reverse‐phase HPLC and shown to be highly active against closely related Bacillus spp. Clinically relevant species such as Staphylococcus aureus and Micrococcus luteus were also inhibited. The substance was characterized as a bacteriocin‐like inhibitory substance (BLIS) with a molecular mass of ca 3·4 kDa. The BLIS was very heat stable, and sensitive only to pronase E and proteinase K. Antimicrobial activity was stable and high in the pH range of 2·0–9·0, and relatively unaffected by organic chemicals.
Conclusions: An antimicrobial substance produced by the B. cereus type strain ATCC 14579 was characterized, with a wide spectrum of activity and the potential to be applied as a control agent against pathogenic bacteria.
Significance and Impact of the Study: The present study is the first report of a substance with antimicrobial activity from the B. cereus type strain.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
To isolate and identify DNA-binding protein(s) with affinity for the mobile chromosomal repeat element bcr1 in Bacillus cereus group bacteria. A biotinylated bcr1 element was immobilized to ...streptavidin-coated magnetic beads and used to pull out a 20 kDa DNA-binding protein from a whole cell protein extract of B. cereus ATCC 14579. The protein was identified as the product of ORF 2 encoded by the bacteriophage-related autonomously replicating linear genetic element pBClin15 carried by the strain. DNA binding was not bcr1-specific. By Northern blotting ORF 2 was co-transcribed with ORF 1, and also in certain instances with ORF 3 by transcriptional readthrough of the terminator located between ORF 2 and ORF 3. ORF 2 from pBClin15 encodes a DNA-binding protein. ORF 2 is co-transcribed with its upstream gene ORF 1, and in a subset of the transcripts also with the downstream gene ORF 3 through alternative transcription termination. The B. cereus group contains bacterial species of medical and economic importance. Bacteriophages or phage-encoded proteins from these bacteria have been suggested as potential therapeutic agents. Understanding the biology of bacteriophage-related genetic elements through functional characterization of their genes is of high relevance.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
We sequenced the complete genome of Bacillus cereus ATCC 10987, a non‐lethal dairy isolate in the same genetic subgroup as Bacillus anthracis. Comparison of the chromosomes demonstrated that B.cereus ...ATCC 10987 was more similar to B.anthracis Ames than B.cereus ATCC 14579, while containing a number of unique metabolic capabilities such as urease and xylose utilization and lacking the ability to utilize nitrate and nitrite. Additionally, genetic mechanisms for variation of capsule carbohydrate and flagella surface structures were identified. Bacillus cereus ATCC 10987 contains a single large plasmid (pBc10987), of ∼208 kb, that is similar in gene content and organization to B.anthracis pXO1 but is lacking the pathogenicity‐associated island containing the anthrax lethal and edema toxin complex genes. The chromosomal similarity of B.cereus ATCC 10987 to B.anthracis Ames, as well as the fact that it contains a large pXO1‐like plasmid, may make it a possible model for studying B.anthracis plasmid biology and regulatory cross‐talk.
DNA from over 300 Bacillus thuringiensis, Bacillus cereus, and Bacillus anthracis isolates was analyzed by fluorescent amplified fragment length polymorphism (AFLP). B. thuringiensis and B. cereus ...isolates were from diverse sources and locations, including soil, clinical isolates and food products causing diarrheal and emetic outbreaks, and type strains from the American Type Culture Collection, and over 200 B. thuringiensis isolates representing 36 serovars or subspecies were from the U.S. Department of Agriculture collection. Twenty-four diverse B. anthracis isolates were also included. Phylogenetic analysis of AFLP data revealed extensive diversity within B. thuringiensis and B. cereus compared to the monomorphic nature of B. anthracis. All of the B. anthracis strains were more closely related to each other than to any other Bacillus isolate, while B. cereus and B. thuringiensis strains populated the entire tree. Ten distinct branches were defined, with many branches containing both B. cereus and B. thuringiensis isolates. A single branch contained all the B. anthracis isolates plus an unusual B. thuringiensis isolate that is pathogenic in mice. In contrast, B. thuringiensis subsp. kurstaki (ATCC 33679) and other isolates used to prepare insecticides mapped distal to the B. anthracis isolates. The interspersion of B. cereus and B. thuringiensis isolates within the phylogenetic tree suggests that phenotypic traits used to distinguish between these two species do not reflect the genomic content of the different isolates and that horizontal gene transfer plays an important role in establishing the phenotype of each of these microbes. B. thuringiensis isolates of a particular subspecies tended to cluster together.
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