Identification and characterization of a zinc metalloprotease associated with invasion by the fish pathogen Vibrio anguillarum

Abstract

An invasiveness-defective mutant of the fish-pathogenic bacterium Vibrio anguillarum was isolated. Compared with the wild type, this mutant had a 1,000-fold higher 50% lethal dose after inmersion infection of rainbow trout, Oncorhynchus mykiss, while after intraperitoneal infection, the mutant had only a 10-fold higher 50% lethal dose. In addition, the mutant showed a lower level of protease activity. Two forms of the protease (Pa and Pb) were found after sodium dodecyl sulfate-polyacrylamide gel electrophoresis of nonheated samples. Pa was found predominantly in protease preparations of the wild type, while Pb was the predominant form in the mutant. Conversion of Pb to Pa was observed in protease preparations after incubation at 4°C. Characterization of the protease showed that it was an elastolytic enzyme which required Zn2+ for activity and Ca2+ for stability. The molecular mass of the protease was 36 kilodaltons. N-terminal amino acid sequence analysis of the protease of V. anguilarum revealed homology to the elastase of Pseudomonas aeruginosa and the

NaCl concentration in the medium modulates the secretion of active EmpA protease in Vibrio anguillarum at post-transcriptional level

“…2b). The protease activity of the mutant strain was strongly decreased in all conditions tested, confirming that the empA gene mutation results in decreased extracellular proteolytic activity (Weber et al 2008;Norqvist et al 1990) (Fig. 2b).…”

Isolation and characterisation of two extracellular metalloproteases from Aeromonas salmonicida ssp. salmonicida

“…In other fish-pathogenic bacteria, single metalloproteases seem to be important for host invasion and establishment of infection. This is strongly suggested in infection of fish with V. anguillarum and Aeromonas hydrophila (Leung & Stevenson 1988;Norqvist, Norrman & Wolf-Watz 1990). Metalloproteases may also be related to virulence by activation of other extracellular toxins.…”

Mutation ofrpoS gene decreased resistance to environmental stresses, synthesis of extracellular products and virulence ofVibrio anguillarum

“…Vibrio anguillarum is a halophilic, gram-negative commashaped rod bacterium that causes vibriosis, a lethal hemorrhagic septicemia affecting a variety of fish species and other aquatic animals, and causing important economic losses throughout the world (Austin & Austin, 1999). Several virulence-related factors have been identified in V. anguillarum, including an iron-uptake system (Crosa, 1980;Crosa et al, 1980;Stork et al, 2007), extracellular metalloprotease (Norqvist et al, 1990;Milton et al, 1992;Yang et al, 2007), hemolysin (Munn, 1980;Hirono et al, 1996;Rock & Nelson, 2006), dermatotoxin, hemagglutinin and cytotoxin (Toranzo et al, 1983;Toranzo & Barja, 1993), genes affecting chemotaxis and motility (McGee et al, 1996;Milton et al, 1996;O'Toole et al, 1996). Recently, a repeat-in-toxin (rtxA) is also found to be a major virulence factor for V. anguillarum (Li et al, 2008).…”