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In the present study, two N-acyl homoserine lactone degrading bacteria (Lactobacillus plantarum QQ8 and Lactobacillus casie QQ10) isolated from the gastrointestinal tract of Oreochromis niloticus were tested for their probiotic effect on goldfish. The isolates were able to degrade short chain synthetic C6-HSL in vitro and hampered violacein production by Chromobacterium violaceum. They were susceptible to all the five antibiotics tried. The isolates grew well at pH 3.0–7.0, were resistant to high level of bile salts (0–0.9%) and 0.5% of phenol. They also exhibited high degree of auto-aggregation and co-aggregation, confirming that they possessed good probiotic attributes. Disease resistance in goldfish against Aeromonas hydrophila infection was assessed with 28 different combinations of probiotic mixtures of the four bacterial strains:L. plantarum QQ8, L. casei QQ10, Enterococcus faecium QQ12 and Bacillus thuringiensis QQ17 (QQ12 and QQ17 were previously isolated in our laboratory) by using the statistical approach of response surface methodology (RSM). The ability of goldfish to resist A. hydrophila infection was higher in probiotic combination fed treatments than that fed with commercial feed alone. The highest survival was observed in gold fish fed diet incorporated with the combination dose of 103 CFU/g feed of L. plantarum QQ8, 102 CFU/g feed of L. casei QQ10, 102 CFU/g feed of E. faecium QQ12 and 103 CFU/g feed of B. thuringiensis QQ17. The corresponding desirability score was almost 1. Goldfish showed 91.7% survival when challenged with A. hydrophila. This study demonstrated the potential of multi-species quorum quenching probiotics as feed additive in aquaculture to control bacterial diseases and use of response surface methodology in determining the optimum combination of probiotic mixture.