Bundle-Forming Pilus Locus of Aeromonas veronii bv. Sobria

Abstract

ABSTRACTLittle is known about the colonization mechanisms ofAeromonasspp. Previous work has suggested that the type IV bundle-forming pilus (Bfp) is an aeromonad intestinal colonization factor. This study provides the first genetic characterization of this structure. To define the role of Bfp inAeromonas veroniibv. Sobria adherence, a 22-kb locus encoding the bundle-forming pilus was isolated; this contained 17 pilus-related genes similar to the mannose-sensitive hemagglutinin (MSHA) ofVibrio cholerae. Reverse transcriptase PCR (RT-PCR) demonstrated that the locus had two major transcriptional units,mshItomshFandmshBtomshQ. Transcriptional fusion experiments demonstrated the presence of two strong promoters upstream ofmshIandmshB. The locus encoded four putative prepilin proteins, one of which (MshA) corresponded to the N-terminal sequence of the previously isolated major pilin protein. All the pilin genes were inactivated, mutation of each minor or major pilin gene greatly reduced the bacterium's ability to adhere and form biofilms, and complementation of each mutant intransrescued this phenotype. Mutation of the major pilin MshA and MshB, a minor pilin, resulted in their loss. The position of themshHgene is conserved within a number of bacteria, and we have shown it is not transcriptionally linked to the othermshgenes; moreover, its mutation did not have a dramatic effect on either adhesion or biofilm formation. We conclude that the bundle-forming pilus is required forA. veroniibv. Sobria adherence and biofilm formation; furthermore, both the major and minor pilin proteins are essential for this process.