Infectious diseases have been estimated to cost the global aquaculture industry billions of dollars annually 1,2 . With concerns over emerging resistance and residues of antibiotics in food 3 many such chemicals are now being banned and environmentally friendly alternatives are being sought.Probiotics influence the composition of the gut microbiota and confer health benefits to their host 4,5 and are one of several alternative approaches gaining significant popularity in aquaculture. Whilst primarily used to manage bacterial disease, there is also some evidence that probiotics can provide protection against parasites 4 and viruses 6 . Probiotics can inhibit the growth of pathogens in the gut through the excretion of antagonistic substances including bacterocins 6,7 ; prevent pathogen adhesion in the gut through competition of space and nutrients 3 and by modulating the immune system 5 . Some probiotics have been reported to improve growth and feed utilisation efficiency 5 and others can also improve water quality 2,8,9 , which confers indirect benefits to host health 6 .It has been estimated that 50 000 tonnes of probiotics are used annually in the aquaculture industry 10 yet analysis of the literature reveals a great deal of equivocal data on their efficacy. This is likely due, at least in part, to the wide diversity of both hosts and probiotic species within this industry and the fact that probiotic efficacy against particular pathogens is often both host specific and probiotic strain specific 5 . These factors demonstrate the need for testing to ensure probiotics are fit for purpose. Aquaculture species span many phyla and their probionts are far more diverse than the typical lactic acid bacteria (LABS) used in terrestrial animals 6 and which do not dominate in the normal gut flora of aquatic animals 2 .The diversity of probiotics used in aquaculture is highlighted by Newaj-Fyzul et al. who reviewed 18 genera of Gram-negative and 19 genera of Gram-positive bacteria that have been used in aquaculture 4 . This diversity reflects that of the aquatic habits where aquaculture species live and closely interact with these microbes 3,7 .Despite this diversity, Bacillus subtilis and B. licheniformis remain the most commonly used commercial probiotics in aquaculture 5 , due to their spore forming nature and subsequent proclivity for long term storage and stability in formulated feeds 6,11 . Evidence exists, however, that probiotics isolated from host gut or their environment are more likely to outperform commercial products because they are more likely to colonise the host gut and effectively compete with pathogens 6,12 . The first stage of screening potential probiotics usually occurs in vitro and seeks to measure antagonism against pathogens of interest using methods including well diffusion, cross streaking and disc diffusion and the co-culture method 3,13 . These methods can also be used to test antagonism of putative probiotics against pathogens of interest. Whilst such methods allow cost effective, simultaneous s...