Abstract
The Bacillus anthracis spore constitutes the infectious form of the bacterium, and sporulation is an important process in the organism’s life cycle. Herein, we show that disruption of SpoVG resulted in defective B. anthracis sporulation. Confocal microscopy demonstrated that a ΔspoVG mutant could not form an asymmetric septum, the first morphological change observed during sporulation. Moreover, levels of spoIIE mRNA were reduced in the spoVG mutant, as demonstrated using β-galactosidase activity assays. The effects on sporulation of the ΔspoVG mutation differed in B. anthracis from those in B. subtilis because of the redundant functions of SpoVG and SpoIIB in B. subtilis. SpoVG is highly conserved between B. anthracis and B. subtilis. Conversely, BA4688 (the protein tentatively assigned as SpoIIB in B. anthracis) and B. subtilis SpoIIB (SpoIIBBs) share only 27.9% sequence identity. On complementation of the B. anthracis ΔspoVG strain with spoIIBBs, the resulting strain pBspoIIBBs/ΔspoVG could not form resistant spores, but partially completed the prespore engulfment stage. In agreement with this finding, mRNA levels of the prespore engulfment gene spoIIM were significantly increased in strain pBspoIIBBs/ΔspoVG compared with the ΔspoVG strain. Transcription of the coat development gene cotE was similar in the pBspoIIBBs/ΔspoVG and ΔspoVG strains. Thus, unlike in B. subtilis, SpoVG appears to be required for sporulation in B. anthracis, which provides further insight into the sporulation mechanisms of this pathogen.
Highlights
Bacillus anthracis, the causative agent of anthrax, mostly exists in nature in the form of spores, which play an important role in anthrax infection
The sporulation efficiency of the ∆spoVG strain was assessed via its heat-resistance (70 ◦C for 30 min) after cells were cultured in Difco sporulation medium (DSM) for 24 h
Levels of spoIIQ and cotE mRNA had no significant change in the pBspoIIBBs/∆spoVG strain compared with those in strain ∆spoVG. These results indicated that SpoIIBBs could partially restore sporulation of the B. anthracis ∆spoVG strain at the eMngicurolofrmgaennismt sst2a0g20e,.8, x FOR PEER REVIEW
Summary
The causative agent of anthrax, mostly exists in nature in the form of spores, which play an important role in anthrax infection. Sporulation in B. subtilis has been divided into so-called “stages” 0 to VII using electron microscopy [2,3]—vegetative cells (stage 0), axial filamentation (stage I), asymmetric division (stage II), engulfment (stage III), formation of the cortex and coat (stages IV and V), and spore maturation and mother cell lysis (stages VI and VII) [2,4,5,6]. SpoIIE, an integral membrane protein [9], is involved in asymmetric septum formation in B. subtilis and plays a crucial role in sporulation in B. anthracis. The cotE gene is involved in assembly of the outer coat structure in stage V of sporulation [12,13]
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