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


CONCLUSIONS
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.
Sozhamannan
Shanmuga
S
Biological Defense Research Directorate, Naval Medical Research Center, 503 Robert Grant Avenue, Silver Spring, Maryland 20852, USA. sozhamannans@nmrc.navy.mil
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
D001287:Attachment Sites, Microbiological / Q000235:genetics; D001407:Bacillus / Q000145:classification / Q000235:genetics; D017103:Bacillus Phages / Q000235:genetics* / Q000502:physiology; D001408:Bacillus anthracis / Q000145:classification / Q000235:genetics*; D001483:Base Sequence; D002876:Chromosomes, Bacterial / Q000235:genetics*; D017931:DNA Primers / Q000737:chemistry; D004279:DNA, Viral / Q000737:chemistry; D004587:Electrophoresis, Agar Gel / Q000379:methods; D023061:Gene Order; D046529:Microscopy, Electron, Transmission / Q000379:methods; D016685:Mitomycin / Q000494:pharmacology; D016133:Polymerase Chain Reaction / Q000379:methods; D039002:Prophages / Q000235:genetics* / Q000502:physiology; D014775:Virus Activation / Q000187:drug effects / Q000502:physiology*
D017931:DNA Primers; D004279:DNA, Viral; D016685:Mitomycin