Both Epsilon-Toxin and Beta-Toxin Are Important for the Lethal Properties of Clostridium perfringens Type B Isolates in the Mouse Intravenous Injection Model

Abstract

Clostridium perfringens is capable of producing up to 15 toxins, including alpha-toxin (CPA), beta-toxin (CPB), epsilon-toxin (ETX), enterotoxin, beta2-toxin (CPB2), and perfringolysin O. Type B isolates, which must produce CPA, CPB, and ETX, are associated with animal illnesses characterized by sudden death or acute neurological signs, with or without intestinal damage. Type B pathogenesis in ruminants is poorly understood, with some animals showing lesions and clinical signs similar to those caused by either type C or type D infections. It is unknown whether host or environmental conditions are dominant for determining the outcome of type B disease or if disease outcomes are determined by variable characteristics of type B isolates. To help clarify this issue, 19 type B isolates were evaluated for toxin production during late-log-phase growth via quantitative Western blotting and by biological activity assays. Most type B isolates produced CPB levels similar to those produced by type C isolates in vitro and have the potential to produce genotype C-like disease. The lethality of type B isolate supernatants administered intravenously to mice was evaluated with or without prior trypsin treatment, and monoclonal antibody neutralization studies also were performed. Correlation analyses comparing toxin levels in type B supernatants versus lethality and neutralization studies both found that the main contributor to lethality without pretreatment with trypsin was CPB, whereas neutralization studies indicated that CPB and ETX were both important after trypsin pretreatment. At least part of the CPB produced by type B isolates remained active after trypsin treatment. However, the overall lethalities of most supernatants were lower after trypsin pretreatment. Also, there was a significant association between ETX, CPB2, and CPA production in vitro among type B isolates. However, our results suggest that both CPB and ETX are likely the most important contributors to the pathogenesis of C. perfringens type B infections in domestic animals.

Toxin Plasmids of Clostridium perfringens

“…These opposing effects of trypsin on ETX and CPB activity suggest that when both toxins are present together in the intestine, such as during type B-associated infections, variations in intestinal conditions select for the predominant activity of ETX over CPB or vice versa. In animal model studies, at least some CPB produced by type B isolates remained active after trypsin treatment, but the overall lethality of most type B supernatants was lower after trypsin pretreatment (52).…”

“…For example, without pretreatment with trypsin, CPB was found to be the main contributor to the lethal properties of type B supernatants using a mouse intravenous (i.v.) injection model, whereas seroneutralization studies with this model indicated that CPB and ETX are both important after trypsin pretreatment of type B supernatants (52). CPB is very sensitive to trypsin digestion, so animals with low levels of intestinal trypsin (such as neonates) are usually the most susceptible to infection by type B or C isolates (3,6,210).…”

“…Preliminary results suggest that both CPB and ETX are the most important toxins for the pathogenesis of type B infections in domestic animals (52). For example, without pretreatment with trypsin, CPB was found to be the main contributor to the lethal properties of type B supernatants using a mouse intravenous (i.v.)…”

Batı Cezayir’in Tiaret Bölgesinde Nekrotik Enteritli Broiler Tavuklardan İzole Edilen Clostridium perfringens’in İdentifikasyonu ve Karakterizasyonu

“…0.3 mL from the clear supernatant fluid was I/V inoculated in the tail vein of each Swiss mouse (25-40 g), and injected mice were observed over a period of three days for nervous symptoms or death [23] . One mouse was injected with broth culture without bacteria as a control.…”

Characterization of Clostridium perfringens TpeL Toxin Gene Carriage, Production, Cytotoxic Contributions, and Trypsin Sensitivity

“…Some toxins produced by C. perfringens in the intestines are exceptionally sensitive to trypsin, while other toxins made by this bacterium must be activated by trypsin or other proteases (2,(16)(17)(18). CPB, produced by C. perfringens type B and type C strains, is very sensitive to endogenous trypsin degradation in the intestines of natural host animals (18,19).…”