Toxin plasmids of Clostridium perfringens

Abstract

In both humans and animals, Clostridium perfringens is an important cause of histotoxic infections and diseases originating in the intestines, such as enteritis and enterotoxemia. The virulence of this Gram-positive, anaerobic bacterium is heavily dependent upon its prolific toxin-producing ability. Many of the ∼16 toxins produced by C. perfringens are encoded by large plasmids that range in size from ∼45 kb to ∼140 kb. These plasmid-encoded toxins are often closely associated with mobile elements. A C. perfringens strain can carry up to three different toxin plasmids, with a single plasmid carrying up to three distinct toxin genes. Molecular Koch's postulate analyses have established the importance of several plasmid-encoded toxins when C. perfringens disease strains cause enteritis or enterotoxemias. Many toxin plasmids are closely related, suggesting a common evolutionary origin. In particular, most toxin plasmids and some antibiotic resistance plasmids of C. perfringens share an ∼35-kb region containing a Tn916-related conjugation locus named tcp (transfer of clostridial plasmids). This tcp locus can mediate highly efficient conjugative transfer of these toxin or resistance plasmids. For example, conjugative transfer of a toxin plasmid from an infecting strain to C. perfringens normal intestinal flora strains may help to amplify and prolong an infection. Therefore, the presence of toxin genes on conjugative plasmids, particularly in association with insertion sequences that may mobilize these toxin genes, likely provides C. perfringens with considerable virulence plasticity and adaptability when it causes diseases originating in the gastrointestinal tract.

Fig 1

Size diversity of C. perfringens plasmids encoding key toxins. Shared colors (other than black) indicate a similar (if not identical) plasmid; e.g., the 65-kb etx- and cpb2-carrying plasmid of type B strains is also apparently present in some type D strains. “seq” denotes a sequenced plasmid.

Fig 2

Comparative alignment of sequenced C. perfringens plasmids. Shown are sequence alignments for pCW3 (236); pJIR3537 (tet⁺), pJIR3844 (cpb2⁺), and pJIR3535 (netB⁺) (19); pCP8533etx (etx⁺ cpb2⁺) (14); pCPF5603 (cpe⁺ cpb2⁺) (13); pCPPB-1 (cpe⁺ iota⁺) (23); and pCPF4969 (cpe⁺) (13). Each arrow represents an ORF; ORF arrows shown are as follows: red arrows, the conserved tcp locus (note the adjacent dcm ORF); dark blue arrows, other conserved ORFs shared by these plasmids; light purple arrows, tetracycline resistance gene; green arrows, the cpb2 toxin gene; purple arrows, the netB toxin gene; pink arrows, the etx gene; gray arrows, the cpe gene; dark gray arrows, the iota-toxin gene; yellow arrows, plasmid replication region; light blue arrows, regions unique to each plasmid. Asterisks denote a toxin gene. The GenBank accession numbers for the plasmid sequences are DQ366035 for pCW3, JN689220 for pJIR3537, JN689217 for pJIR3844, JN689219 for pJIR3536, AB444205 for pCP8533etx, AB236337 for pCPF5603, AB604032 for pCPPB-1, and AB236336 for pCPF4969. RR refers to response regulator, and SHK refers to sensor histidine kinase.

Fig 3

Organization of toxin (cpe, etx, and cpb) loci in type A, B, C, D, and E strains of C. perfringens. (A) Organization of plasmid-borne cpe loci in type A, E, C, and D strains. (B) Organization of the type A chromosome cep locus. (C) Organization of plasmid-borne etx loci in type B and D strains. (D) Organization of plasmid-borne cpb loci in type B and C isolates. Each arrow represents an ORF. Asterisks indicate a region with similarity to sequences present downstream of the cpe gene in F4969, except for the absence of an IS1470-like gene. (Panels A and B adapted from reference ; panel C adapted from reference ; panel D adapted from references and .)

Fig 4

Model for evolution of characterized C. perfringens toxin plasmids. See the text for discussion of the possible evolution of pIP401 (230), pCW3 (236), pJIR3535 (netB⁺) (19), pCP8533etx (etx⁺ cpb2⁺) (14), pCPF5603 (cpe⁺ cpb2⁺) (13), pCPPB-1 (cpe⁺ iota⁺) (23), and pCPF4969 (cpe⁺) (13). Each box color depicts a different region of importance on the toxin plasmids, as indicated.

Fig 5

The pCW3 tcp locus. The genetic organization and protein products of the tcp locus are shown. (Adapted from reference with permission of the publisher [copyright 2012 Blackwell Publishing Ltd.].)

Fig 6

Putative model of the pCW3 conjugation apparatus. The relative locations and known protein-protein interactions of the TcpA (A) (orange), TcpH (H) (brown), TcpC (C) (green), TcpG (purple), and TcpF (F) (red) proteins are based on data from previous studies (236, 242, 244, 245, 248). (Based on a model prepared by Jessica Wisniewski, Monash University.)