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The Bacillus anthracis chromosome contains four conserved, excision-proficient, putative prophages.

ABSTRACT: BACKGROUND: Bacillus anthracis is considered to be a recently emerged clone within the Bacillus cereus sensu lato group. The B. anthracis genome sequence contains four putative lambdoid prophages. We undertook this study in order to understand whether the four prophages are unique to B. anthracis and whether they produce active phages. RESULTS: More than 300 geographically and temporally divergent isolates of B. anthracis and its near neighbors were screened by PCR for the presence of specific DNA sequences from each prophage region. Every isolate of B. anthracis screened by PCR was found to produce all four phage-specific amplicons whereas none of the non-B. anthracis isolates, produced more than one phage-specific amplicon. Excision of prophages could be detected by a PCR based assay for attP sites on extra-chromosomal phage circles and for attB sites on phage-excised chromosomes. SYBR-green real-time PCR assays indicated that prophage excision occurs at very low frequencies (2 x 10(-5) - 8 x 10(-8)/cell). Induction with mitomycin C increased the frequency of excision of one of the prophages by approximately 250 fold. All four prophages appear to be defective since, mitomycin C induced culture did not release any viable phage particle or lyse the cells or reveal any phage particle under electron microscopic examination. CONCLUSION: The retention of all four putative prophage regions across all tested strains of B. anthracis is further evidence of the very recent emergence of this lineage and the prophage regions may be useful for differentiating the B. anthracis chromosome from that of its neighbors. All four prophages can excise at low frequencies, but are apparently defective in phage production.

SUBMITTER: Sozhamannan S

PROVIDER: S-EPMC1475869 | biostudies-literature | 2006

REPOSITORIES: biostudies-literature

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The Bacillus anthracis chromosome contains four conserved, excision-proficient, putative prophages.

Sozhamannan Shanmuga S Chute Michael D MD McAfee Farrell D FD Fouts Derrick E DE Akmal Arya A Galloway Darrell R DR Mateczun Alfred A Baillie Leslie W LW Read Timothy D TD

BMC microbiology 20060406

<h4>Background</h4>Bacillus anthracis is considered to be a recently emerged clone within the Bacillus cereus sensu lato group. The B. anthracis genome sequence contains four putative lambdoid prophages. We undertook this study in order to understand whether the four prophages are unique to B. anthracis and whether they produce active phages.<h4>Results</h4>More than 300 geographically and temporally divergent isolates of B. anthracis and its near neighbors were screened by PCR for the presence ...[more]

PMID: 16600039

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Project description:Purpose: Small regulatory RNAs (sRNAs) are short transcripts that base-pair to mRNA targets or interact with regulatory proteins. sRNA function has been extensively studied in Gram-negative bacteria; comparatively less is known about sRNAs in Firmicutes. Here we investigate two sRNAs encoded by the virulence plasmid pXO1 of Bacillus anthracis, the causative agent of anthrax. We designated the sRNAs as as “XrrA” and “XrrB” (for pXO1-encoded regulatory RNA). We constructed deletion mutants ∆xrrA, ∆xrrB, and ∆xrrA∆xrrB in an Ames strain background and compared gene expression of these mutants to that of the Ames-derived parent strain using RNA-seq. Methods: Total RNA from Ames strains UTA37 (Parent), UTA38 (∆xrrB), UTA39 (∆xrrA), UTA41 (∆xrrA∆xrrB) was isolated from cultures grown in CA-CO2 in triplicate, using saturated phenol: chloroform extraction. One microgram of pure and quality-checked RNA from each sample was subjected to Illumina sequencing by synthesis using a NextSeq550 sequencer. Sequencing by synthesis was performed to generate 75bp paired-end reads. Two 130M-read sequencing runs (Run1 and Run2) were performed and an average of 31M reads per sample were obtained. Results: All bioinformatic analysis was performed using the publicly available Galaxy web resource (https://usegalaxy.org/). Comparison of the transcriptomes of a virulent Ames-derived strain to isogenic sRNA-null mutants revealed multiple 4.0- to >100-fold differences in gene expression. Deletion of xrrA resulted in 50 transcripts showing a ≥ 4.0-fold change in expression compared to the parent strain. In contrast, deletion of xrrB led to one transcript having a ≥ 4.0-fold change in expression compared to the parent strain. In the ∆xrrA∆xrrB mutant, levels of 116 transcripts were affected with a fold-change of ≥ 4.0. Many sRNA-regulated targets were chromosome genes associated with branched-chain amino acid metabolism, proteolysis, and transmembrane transport. Conclusions: In this work, we provide experimental evidence for sRNA-mediated regulation in the mammalian pathogen B. anthracis. XrrA and XrrB are the first regulatory RNAs examined in B. anthracis and are shown here to have multiple effects on gene expression in this pathogen. The data suggest distinct regulatory roles for XrrA, as well as some functional overlap between XrrA and XrrB.

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The Bacillus anthracis chromosome contains four conserved, excision-proficient, putative prophages.

Publications

The Bacillus anthracis chromosome contains four conserved, excision-proficient, putative prophages.

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