DAO 67:201-207 (2005)  -  doi:10.3354/dao067201

ABSTRACT: The pathogenicity of Vibrio penaeicida Strains KH-1 and AM101, their culture-free supernatant (CFS), and their protein fraction obtained by 40% of ammonium sulfate precipitation (PFs40) were assessed in experimental challenges against juvenile Litopenaeus vannamei. Live Vibrio cells, CFS, and PFs40 from the AM101 strain produced a significantly higher mortality (p < 0.05) compared to the KH-1 strain. Toxicity and median lethal doses (LD₅₀) of Fast Protein Liquid Chromatography (FPLC) products were evaluated on L. vannamei. The first FPLC fraction sample (A) from PFs40 of the AM101 strain displayed LD₅₀ values of 1.68 and 5.61 µg protein ind.^–1, respectively. The second FPLC process from Fraction A showed a peak (A1) also with toxic effects to shrimp. PFs40, Fraction A, and Peak A1 showed a 38.5 kDa molecular band (SDS-PAGE), with activity on a gelatin protease zymogram. The lethal effect of PFs40 and Fraction A was inhibited by Proteinase K, CuCl₂, E-64, and heat (60 and 100°C) treatments, but was not inhibited by EDTA-Na₂, aprotinin, and soy trypsin treatments. These results and the zymogram inhibition test suggest the presence of a cysteine protease-like proteinaceous exotoxin as a dominant protease, secreted by V. penaeicida Strain AM101.

KEY WORDS: Vibrio penaeicida · Cysteine protease · Litopenaeus vannamei · Shrimp · Exotoxin · Pathogenicity