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

Pathogenicity of Vibrio penaeicida for white shrimp Litopenaeus vannamei: a cysteine protease-like exotoxin as a virulence factor

Gabriel Aguirre-Guzmán1,3,*, Felipe Ascencio1, Denis Saulnier2

1Centro de Investigaciones Biológicas del Noroeste, Apdo. Postal 128, Unidad de Patología Marina, La Paz,Baja California Sur, Mexico
2Laboratory of Genetic and Pathology, French Research Institute for Exploitation of the Sea (IFREMER),La Tremblade 17390, France
3Present address: Fac. de Medicina Veterinaria y Zootecnia, Universidad Autónoma de Tamaulipas, Km 5 Carreter Cd.Victoria—Mante, Cd. Victoria, Tamps 87000, Mexico

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

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