Molecular characterization of Clostridium perfringens isolates from humans with sporadic diarrhea: evidence for transcriptional regulation of the beta2-toxin-encoding gene.
Clostridium perfringens type A food poisoning is caused by C. perfringens isolates carrying a chromosomal enterotoxin gene (cpe), while non-food-borne gastrointestinal (GI) diseases, such as antibiotic-associated diarrhea (AAD) and sporadic diarrhea (SD), are caused by C. perfringens plasmid cpe isolates. A recent study reported the association of beta2 toxin (CPB2) with human GI diseases, and particularly AAD/SD, by demonstrating that a large percentage of AAD/SD isolates, in contrast to a small percentage of food poisoning isolates, carry the beta2-toxin gene (cpb2). This putative relationship was further tested in the current study by characterizing 14 cpe+ C. perfringens fecal isolates associated with recent cases of human SD in England (referred to hereafter as SD isolates). These SD isolates were all classified as cpe+ type A, and 12 of the 14 cpe+ isolates carry their cpe gene on the plasmid and 2 carry it on the chromosome. Interestingly, cpb2 is present in only 12 plasmid cpe isolates; 11 isolates carry cpe and cpb2 on different plasmids, but cpe and cpb2 are located on the same plasmid in one isolate. C. perfringens enterotoxin is produced by all 14 cpe+ SD isolates. However, only 10 of the 12 cpe+/cpb2+ SD isolates produced CPB2, with significant variation in amounts. The levels of cpb2 mRNA in low- to high-CPB2-producing SD isolates differed to such an extent (30-fold) that this difference could be considered a major cause of the differential level of CPB2 production in vitro by SD isolates. Furthermore, no silent or atypical cpb2 was found in a CPB2 Western blot-negative isolate, 5422/94, suggesting that the lack of CPB2 production in 5422/94 was due to low expression of cpb2 mRNA. This received support from our observation that the recombinant plasmid carrying 5422/94 cpb2, which overexpressed cpb2 mRNA, restored CPB2 production in F4969 (a cpb2-negative isolate). Collectively, our present results suggest that CPB2 merits further study as an accessory toxin in C. perfringens-associated SD.
Harrison
Ben
B
Department of Microbiology, Oregon State University, 220 Nash Hall, Corvallis, OR 97331, USA.
Raju
Deepa
D
Garmory
Helen S
HS
Brett
Moira M
MM
Titball
Richard W
RW
Sarker
Mahfuzur R
MR
D001427:Bacterial Toxins / Q000235:genetics*; D015139:Blotting, Southern; D002874:Chromosome Mapping; D002876:Chromosomes, Bacterial; D003016:Clostridium perfringens / Q000235:genetics* / Q000302:isolation & purification; D003967:Diarrhea / Q000382:microbiology*; D016521:Electrophoresis, Gel, Pulsed-Field; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D012333:RNA, Messenger / Q000235:genetics; D014158:Transcription, Genetic
D001427:Bacterial Toxins; D012333:RNA, Messenger; C109896:cpb2 protein, Clostridium perfringens