Naturally occurring Clostridium perfringens nontoxic alpha-toxin variant as a potential vaccine candidate against alpha-toxin-associated diseases

Abstract

Clostridium perfringens mutant strain 121A/91 shows neither enzymatic (phospholipase C) nor hemolytic activity. Nevertheless, the cpa gene and the corresponding alpha-toxin variant are detectable. Vaccination with this genetically constructed alpha-toxin variant, rAT121/91, induces antibodies capable of significantly reducing activities induced by wild-type toxin. Thus, rAT121/91 could be a useful vaccine candidate.

FIG. 1

Comparison of the alpha-toxin-related phenotypic characteristics of C. perfringens type A reference strain ATCC 13124 and strain 121A/91. (A) Bacterial growth of C. perfringens strains ATCC 13124 (left) and 121A/91 (right) on EYSA. (B) Bacterial growth of C. perfringens strains ATCC 13124 (left) and 121A/91 (right) on mouse blood agar.

FIG. 2

Detection of the cpa gene (775 bp) encoding C. perfringens alpha-toxin by PCR. Lanes: 1, DNA size marker (1- kb DNA ladder); 2, negative control (no template DNA); 3, C. perfringens ATCC 13124 (positive control); 4, C. perfringens strain 121A/91.

FIG. 3

Immunoreactivity against alpha-toxin-specific MAb 3B4 (1 μg/ml of PBS-T20) of protein extracts from the culture supernatants of C. perfringens strains ATCC 13124 and 121A/91, as well as genetically constructed rAT121A/91, by immunoblot analysis. Lanes contained the following culture supernatants: 1, C. perfringens strain ATCC 13124 (20 μg of protein extract/slot); 2, C. perfringens strain 121A/91 (20 μg of protein extract/slot); 3, affinity-purified rAT121A/91 (3 μg/slot).