Extracellular Proteinases of Miamiensis avidus Causing Scuticociliatosis are Potential Virulence Factors

Miamiensis avidus is the causative agent of scuticociliatosis in various marine fish species. The virulence factors of the parasite have not been identified, so far. In this study, we examined M. avidus extracellular proteinases (ECPs) as potential virulence factors, using culture supernatants as an ECPs source. We investigated the substrate specificity of ECPs using artificial peptides, and the cytotoxicity of the ECPs was examined using CHSE-214 cells. To elucidate the role of ECPs in ciliate growth, M. avidus was cultured on CHSE-214 cells in the presence of proteinase inhibitors. We detected proteinase activities from the supernatant of M. avidus. Viable CHSE-214 cells decreased significantly in number, when incubated in a medium supplemented with the culture supernatant of M. avidus. The growth of ciliates on CHSE-214 cells was delayed in the presence of PMSF (serine proteinase inhibitor) and E-64 (cysteine proteinase inhibitor). These results suggested that the culture supernatant contained ECPs showing cytotoxicity, and the proteinases facilitated nutrient uptake by the ciliates. Thus, ECPs may be responsible for virulence factors of M. avidus.