The A-layer influences the susceptibility of Aeromonas salmonicida to antibacterial peptides

Abstract

*Corresponding author. E-mail: c.secombes@abdn.ac.uk †Present address: The Randall Centre, King’s College London, New Hunts House, Guy’s Campus, St Thomas Street, London SE1 9RT. Bacterial diseases in fish are widespread in farmed as well as in wild stocks. In aquaculture, these diseases can sometimes be treated with antibiotics. However, bacteria can become resistant to some antibiotics by producing -lactamase that destroys them (Coghlan, 1996). It is not surprising then that increasing observations of multiresistance in areas where salmonid culture is intensive are reported (Richards et al., 1992; Inglis et al., 1993b). Host non-specific antimicrobial molecules may be a good alternative to antibiotics, since the known mechanisms by which bacteria become resistant to antibiotics would not operate against them (Cannon, 1987; Coghlan, 1996). They have been particularly well studied in insects, amphibians and mammals, as exemplified by cecropins, magainins and defensins. They are usually cationic peptides that interact with the bacterial membranes, disturbing their integrity and membrane potential (Nicolas & Mor, 1995; Hancock & Lehrer, 1998). Whilst some large enzymatic proteins with antimicrobial activity are well known in fish (e.g. lysozyme, Grinde et al., 1988), only recently have small non-enzymatic antibacterial peptides that can inflict physical damage to bacteria been reported (Lemaitre et al., 1996; Oren & Shai, 1996; Cole et al., 1997). Indeed, partial sequences for several antibacterial peptides have been reported in salmonids (Henry, 2000) suggesting that common bacterial pathogens of salmonids, such as those that cause vibriosis, furunculosis, enteric redmouth disease (all caused by Gram-negative bacteria) and bacterial kidney disease (a Gram-positive bacterium) (Inglis et al., 1993a), are likely to be sensitive to such molecules. In this study the lethal concentration of a number of commercially available synthetic antibacterial peptides have been determined for these bacterial pathogens, and analysed with respect to phenotypes that influence virulence in the cases of Aeromonas salmonicida, the causative agent of furunculosis. Yersinia ruckeri (strain MT252), responsible for enteric redmouth disease, and strains of A. salmonicida (Table 1) were cultured in Tryptic Soy Broth (TSB, Fluka) at 20 1 C. The broth containing Vibrio anguillarum (strain MT 1746) was grown in TSB supplemented with 2% NaCl. Renibacterium salmoninarum (strain MT426), the pathogenic agent of bacterial kidney disease, was cultured in Mueller Hinton Broth (Fluka) at 16 1 C.