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ALPHA AND BETA TOXINS ACTIVITIES IN REGIONAL STRAINS OF CLOSTRIDIUM CHAUVOEI.

Cortiñas, T.I.*, Mattar, M.A. and Guzmán A.M.S.
Area Microbiología, Universidad Nacional de San Luis, San Luis, ARGENTINA

Clostridium chauvoei is the ethiological agent of blackleg, a high mortality rate disease affecting sheep an cattle. The microorganism produces at least five soluble antigens (toxins) including an oxygen-stable hemolysin (alpha), a DNase (beta), a hyaluronidase (gamma), an oxygen-labile hemolysin (delta) and a neuroaminidase. These toxins enhance the ability of C. chauvoei to invade and multiply in the tissue of a susceptible host and they exhibit immunological activity. It has been informed that the toxin is produced in variable amount by different strains of C. chauvoei and there is very little information about the culture conditions affecting their production. The aim of this work was to determine the effect of culture conditions on the activities of alpha and beta toxins from two regional and one reference strains. The strains were cultured in batch system using glucose at 0.5%, 1% and at limiting concentrations and incubated at 37ºC and 41ºC. The activity of the alpha toxin was assessed by HU₅₀/ml determination and that of the beta toxin by DNA-methyl green assay as modified by Miyakawa et al. No significant differences were found on the hemolytic activity of regional and reference strains at both temperatures and at the glucose concentrations analyzed (p less than/equal 0.05). On the other hand, there was a marked influence of temperature and glucose concentrations on DNase activity which varied with the strains from 2.5% to 27% using glucose at 0.5% and 1% concentrations to 63% to 93% under limitation of the carbon source at 37ºC while at 41ºC no DNase activity was detected. The protein profiles obtained in SDS-PAGE varied markedly with the temperature used. The protein production in supernatants, indicated by the number and intensity of the bands, was enhanced in cultures developed under limitation of the carbon source. Our results indicate that DNase is mainly produced as an active form while hemolysin is liberated as an inactive form. No significant differences (p<-0.05) were found in DNase and hemolytic activities between the different strains assayed. These results would show that the toxin activities were determined primarily by the culture conditions