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Infect. Immun. doi:10.1128/IAI.01981-06
Copyright (c) 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Comparison of Virulence Plasmids Among Clostridium perfringens Type E Isolates

Department of Molecular Genetics and Biochemistry, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA; Department of Microbiology, Wakayama University School of Medicine, 811-1 Kimiidera, Wakayama, 641-0012, Japan; Australian Research Council Centre of Excellence in Structural and Functional Microbial Genomics, Monash University, Victoria, Australia

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

	Abstract

Clostridium perfringens type E isolates produce iota toxin, which is encoded by iap and ibp genes. Using Southern blot analyses, the current study identified iap/ibp plasmids of 97 or 135 kb among eight type E isolates. For most of these isolates, their iap/ibp plasmid also encoded urease and lambda toxin. However, the beta2 toxin gene, if present, was on a different plasmid from the iap/ibp plasmid. For all isolates, the iap/ibp plasmid carried a tcp locus, strongly suggesting these plasmids are conjugative. Overlapping PCR analyses demonstrated some similarity between the iap/ibp plasmids and enterotoxin-encoding plasmids of type A isolates. Additional PCR analyses demonstrated the iap/ibp locus is located near dcm sequences, an apparent plasmid hot-spot for toxin gene insertion, and that two IS1151-related sequences are present in the iap/ibp locus. To begin testing whether those IS1151-like sequences can mobilize iap/ibp genes, a PCR assay was performed that only amplifies a product from circular DNA forms that could represent transposition intermediates. This PCR assay detected circular forms containing iap/ibp genes and silent enterotoxin gene sequences, with or without an IS1151-like sequence. Collectively, these results suggest that a mobile genetic element carrying iap/ibp has inserted onto a tcp-carrying enterotoxin plasmid in a type A isolate, creating a progenitor iap/ibp plasmid. That plasmid then spread via conjugation to other isolates, converting them to type E. Further iap/ibp plasmid diversity occurred when either the iap/ibp genes later re-mobilized and inserted onto other conjugative plasmids or some iap/ibp plasmids acquired additional DNA sequences.

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