Isolation and characterization of a haemolysin from Trichophyton mentagrophytes

Haemolytic activities of Trichophyton (T.) mentagrophytes were detected and characterized by qualitative and quantitative assays. On Columbia agar supplemented with blood from horses, cattle or sheep, T. mentagrophytes expressed a strong zone of complete haemolysis. No haemolytic activities could be detected in the closely related T. verrucosum var. ochraceum. The same results were obtained after cultivation of the fungi on sterile cellulose acetate filters placed on the surface on Columbia blood agar. After removal of the filter, complete haemolysis was detected below the colony of T. mentagrophytes. A soluble haemolysin from culture supernatant of this strain was isolated and partially purified. Specific haemolytic activity per mg protein was enriched 2.6-fold in filtrate F(1), a fraction obtained as filtrate after filtration through 3kDa cut-off membranes. The partially purified haemolysin was neither affected by proteinase K treatment, nor by high and low temperatures, suggesting that it represents a small peptide haemolysin. Accordingly, in a commercial enzymatic activity test only the crude culture filtrate, but none of the subsequent purification fractions showed reactivity. Evaluation of the specificity of the haemolysin using erythrocytes from different mammalian species revealed that sensitivity was highest to those of equines, followed by erythrocytes from sheep, cattle, swine, dogs and humans. None of the erythrocytes was lysed by filtrate F(1) from T. verrucosum var. ochraceum. Furthermore, different eukaryotic cell lines from different species were tested in their sensitivity to cytolytic activities of the haemolysin, but no membrane damage could be detected.