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Infect. Immun. doi:10.1128/IAI.02014-06
Copyright (c) 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Virulence Plasmid Diversity In Clostridium perfringens Type D Isolates

University of Pittsburgh School of Medicine, Department of Molecular Genetics and Biochemistry, Pittsburgh, PA 15261, USA

^* To whom correspondence should be addressed. Email: bamcc{at}pitt.edu.

	Abstract

Clostridium perfringens Type D isolates have biodefense importance and also cause natural enterotoxaemias in sheep, goats and, occasionally, cattle. In these isolates, the gene (etx) encoding -toxin is thought to reside on poorly-characterized large plasmids. Type D isolates sometimes also produce other potentially plasmid-encoded toxins, i.e. C. perfringens enterotoxin and beta2 toxin, encoded by the cpe and cbp2 genes, respectively. The current study demonstrated plasmid carriage of the etx, cpe and cpb2 genes in Type D isolates and characterized those toxin-encoding plasmids to gain insights into their genetic organization, potential transferability and diversity. Southern blotting of pulsed-field gels showed the etx gene of Type D isolates can be present on at least five different plasmids, ranging in size from 48-110 kb. The etx plasmids also typically carried IS1151 and tcp ORFs known to mediate conjugative transfer of C. perfringens plasmid pCW3. PCR studies revealed that, beyond their tcp ORFs, etx plasmids of Type D isolates do not carry substantial portions of the conserved or variable regions in the cpe plasmids of type A isolates. Southern blots also demonstrated that, in Type D isolates, the cpe and cpb2 genes are sometimes present on the etx plasmid. Collectively, these studies confirm that the virulence of type D isolates is heavily plasmid-dependent and further indicate that, i) a single Type D isolate can carry multiple virulence plasmids, ii) a single Type D virulence plasmid can carry up to three different toxin genes, and iii) many etx plasmids should be capable of conjugative transfer.

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