The development of probiotics for the control of multiple bacterial diseases of rainbow trout, Oncorhynchus mykiss (Walbaum) J Brunt, A Newaj-Fyzul and B Austin School of Life Sciences, Heriot-Watt University, Edinburgh, UK Abstract JB-1 and GC2, which were equated with Bacillus sp. and Aeromonas sobria respectively, were recov- ered from the digestive tract of rainbow trout, Oncorhynchus mykiss and ghost carp, Cyprinus sp. respectively, and demonstrated effectiveness as probiotics for the control of infections caused by Aeromonas salmonicida, Lactococcus garvieae, Strep- tococcus iniae, Vibrio anguillarum, Vibrio ordalii and Yersinia ruckeri. When administered to rainbow trout (average weight = 12 g) for 14 days in feed dosed at 2 · 10 8 cells g )1 of feed, JB-1 led to a reduction in mortalities to 0–13% after challenge with a range of bacterial pathogens compared to 80–100% mortalities of the controls. Similarly, use of GC2 reduced mortalities to 0–16% following the challenge compared to 80–100% mortalities of the controls. The mode of action reflected nutrition, production of inhibitory substances and stimulation of the innate immune responses. Specifically, JB-1 and especially GC2 were positive for siderophore and chitinase production, and increased lysozyme, phagocytic and respiratory burst activity. Keywords: Aeromonas, Bacillus, control, disease, probiotic, rainbow trout. Introduction Probiotics are becoming increasingly accepted as a valuable means of controlling bacterial diseases of aquatic vertebrates and invertebrates (e.g. Irianto & Austin 2002). However, a wider range of probiotics, including Gram-positive and -negative bacteria (Irianto & Austin 2002), have been used in aquaculture compared with the lactic acid bacteria often used for humans and terrestrial animals. To date, the development of probiotics for use in aquaculture has focused on single diseases, such as enteric redmouth disease (Kim & Austin 2006), which is caused by Yersinia ruckeri (Austin & Austin 1999). The research described here has sought to determine whether or not probiotics could be effective against a range of bacterial fish pathogens. Materials and methods Fish Rainbow trout, Oncorhynchus mykiss (Walbaum), (average weight = 12 g) were obtained from a commercial fish farm in the UK, and maintained in aerated-free flowing de-chlorinated fresh water at 12 °C. The health status was examined immedi- ately upon arrival and at 14 day intervals thereafter (after Austin & Austin 1989). Bacterial cultures Lactococcus garvieae 29–99 and Streptococcus iniae 00–318 were obtained from the Institute of Aquaculture, University of Stirling, and Louisiana State University, Baton Rouge, respectively. Aero- monas salmonicida (Hooke), Vibrio anguillarum (VIB1), Vibrio ordalii (VIB2), and Y. ruckeri (Pri10) were obtained from the School of Life Sciences, Heriot-Watt University. Authenticity was verified after Austin & Austin (1999). The two Journal of Fish Diseases 2007, 30, 573–579 Correspondence Jason Brunt, School of Life Sciences, John Muir Building, Heriot-Watt University, Riccarton, Edinburgh EH14 4AS, UK (e-mail: jasonbrunt@hotmail.com) 573 Ó 2007 The Authors. Journal compilation Ó 2007 Blackwell Publishing Ltd