Enterotoxemia, a disease that affects domestic ruminants, is caused mainly by the epsilon toxin from Clostridium perfringens type D. Its eradication is virtually impossible, control and prophylaxis are based on systematic vaccination of herds with epsilon toxoids that are efficient in inducing protective antibody production. The use of recombinant toxins is one of the most promising of these strategies. This work evaluates the potency of a Cl. perfringens type D epsilon toxoid expressed by Escherichia coli administered to goats, sheep, and cattle. The etx gene was cloned into the pET-11a plasmid of E. coli strain BL21 to produce the recombinant toxin. Rabbits (n=8), goats, sheep, and cattle (n=5 for each species) were immunized with 0.2mg of the insoluble recombinant protein fraction to evaluate vaccine potency of the epsilon toxoid studied. Antibody titers were 40, 14.3, 26, and 13.1 IU/mL in the rabbit, goat, sheep, and cattle serum pools, respectively. The epsilon toxoid produced and tested in this work is adequate for immunization of ruminants against enterotoxemia.

Clostridium perfringens is an anaerobic bacterium that produces several toxins. Of these, the alpha, beta, and epsilon toxins are responsible for causing the most severe C. perfringens-related diseases in farm animals. The best way to control these diseases is through vaccination. However, commercially available vaccines are based on inactivated toxins and have many production drawbacks, which can be overcome through the use of recombinant antigens. In this study, we produced recombinant alpha, beta, and epsilon toxins in Escherichia coli to formulate a trivalent vaccine. Its effectiveness was evaluated through a potency test in rabbits, in which the vaccine generated 9.6, 24.4, and 25.0 IU/mL of neutralizing antibodies against the respective toxins. Following this, cattle, sheep, and goats received the same formulation, generating, respectively, 5.19 ± 0.48, 4.34 ± 0.43, and 4.70 ± 0.58 IU/mL against alpha toxin, 13.71 ± 1.17 IU/mL (for all three species) against beta toxin, and 12.74 ± 1.70, 7.66 ± 1.69, and 8.91 ± 2.14 IU/mL against epsilon toxin. These levels were above the minimum recommended by international protocols. As such, our vaccine was effective in generating protective antibodies and, thus, may represent an interesting alternative for the prevention of C. perfringens-related intoxications in farm animals.

Due to the increasingly restricted use of antimicrobials in animal production systems, the prevention and control of Clostridium perfringens type A- and C-induced diarrhea in piglets should be based on passive immunization via the prepartum vaccination of sows. Given the current obstacles in the production of conventional clostridial vaccines, the use of recombinant proteins has been considered to represent a promising alternative. In the present study, the neutralizing antibody response of immunized sows and their litters to a bivalent vaccine containing the C. perfringens recombinant toxoids alpha (rTA) and beta (rTB) produced in Escherichia coli was assessed. Rabbits (n=8) and pregnant sows (n=7) were immunized with 200μg of each recombinant antigen using Al(OH)3 as adjuvant. The alpha and beta antitoxin titer detected in the rabbits' serum pool was 9.6 and 20.4IU/mL, respectively. The mean alpha and beta antitoxin titers in the sows' sera were 6.0±0.9IU/mL and 14.5±2.2IU/mL, and the corresponding individual coefficients of variation (CV) were 16.04% and 14.91%, respectively. The mean alpha and beta antitoxin titers in the litters' serum pools were 4.2±0.4IU/mL and 10.9±1.7IU/mL, and the CV between litters was 9.23% and 9.85%, respectively. The results showed that the rTA and rTB proteins produced and tested in the present study induced an immune response and can be regarded as candidates for the development of a commercial vaccine against C. perfringens type A- and C-induced diarrhea in pigs.

Clostridium perfringens is a normal inhabitant of the intestinal tract of chickens as well as a potential pathogen that causes necrotic enteritis and colangio hepatitis. The minimum inhibitory concentration (MIC) of seven different compounds used for therapy, growth promotion or prevention of coccidiosis was determined by agar dilution method for 55 C. perfringens strains isolated from the intestines of broiler chickens. All strains showed high susceptibility to penicillin, avilamycin, monensin and narasin. Only 7.3% of the strains showed an intermediated sensitivity to lincomycin, and 49 (89.1%) were considered susceptible. For tetracycline and bacitracin, 41.8% and 47.3% of strains, respectively, were considered resistant.

Clostridium difficile has emerged as a major cause of neonatal colitis in piglets, displacing classic bacterial pathogens. However, there is no information regarding the distribution of this microorganism in pig farms in

Clostridium perfringens is a spore-forming, commensal, ubiquitous bacterium that is present in the gastrointestinal tract of healthy humans and animals. This bacterium produces up to 18 toxins. The species is classified into five toxinotypes (A–E) according to the toxins that the bacterium produces: alpha, beta, epsilon, or iota. Each of these toxinotypes is associated with myriad different, frequently fatal, illnesses that affect a range of farm animals and humans. Alpha, beta, and epsilon toxins are the main causes of disease. Vaccinations that generate neutralizing antibodies are the most common prophylactic measures that are currently in use. These vaccines consist of toxoids that are obtained from C. perfringens cultures. Recombinant vaccines offer several advantages over conventional toxoids, especially in terms of the production process. As such, they are steadily gaining ground as a promising vaccination solution. This review discusses the main strategies that are currently used to produce recombinant vaccines containing alpha, beta, and epsilon toxins of C. perfringens, as well as the potential application of these molecules as vaccines for mammalian livestock animals.

No abstract

. Epsilon toxin produced by Clostridium perfringens types B and D causes enterotoxemia in sheep, goats and calves. Enterotoxemia can cause acute or superacute disease, with sudden death of the affected animal. It provokes huge economic losses when large numbers of livestock are affected. Therapeutic intervention is challenging, because the disease progresses very rapidly. However, it can be prevented by immunization with specific immunogenic vaccines. We cloned the etx gene, encoding epsilon toxin, into vector pET-11a; recombinant epsilon toxin (rec-ε) was expressed in inclusion bodies and was used for animal immunization. Serum protection was evaluated and cross-serum neutralization tests were used to characterize the recombinant toxin. To analyze the potency of the 267 ©FUNPEC-RP www.funpecrp.com.br Genetics and Molecular Research 9 (1): 266-276 (2010) Immunization using recombinant epsilon toxin from C. perfringens toxin (as an antigen), rabbits were immunized with 50, 100 or 200 µg recombinant toxin, using aluminum hydroxide gel as an adjuvant. Titers of 10, 30 and 40 IU/mL were obtained, respectively. These titers were higher than the minimum level required by the European Pharmacopoeia (5 IU/mL) and by the USA Code of Federal Regulation (2 IU/mL). This rec-ε is a good candidate for vaccine production against enterotoxemia caused by epsilon toxin of C. perfringens type D.