Evaluation of the presence and efficiency of potential probiotic bacteria in the gut of tilapia (Oreochromis niloticus) using the fluorescent in situ hybridization technique

Abstract

The Fluorescent in situ Hybridization (FISH) technique was employed to enumerate potential probiotic and putative pathogenic bacteria in the gut of tilapia (Oreochromis niloticus). Bacteria used in the study were isolated from water, sediment and intestines of tilapia (Oreochromis niloticus) raised in an aquaculture system. These isolates were tested in vitro on antagonism tests against putative pathogenic bacteria (Aeromonas hydrofila, Enterococcus faecalis, Edwardsiella tarda, Pseudomonas fluorescens and Pseudomonas putida), also isolated fromthe same aquaculture system. Two isolates that inhibited largest number of pathogenic bacteria were identified by sequencing as Bacillus sp. and Enterococcus sp. and were added to the commercial feed (106 cells g−1) for in vivo tests. Treatments of the in vivo experiment were: 1) Control — fish fed with no added bacteria, 2) Bacil. — fish fed diets containing Bacillus sp.; 3) Enter. — fish fed diets containing Enterococcus sp., and 4) Bacil.+Enter. — fish fed diets containing Bacillus sp. and Enterococcus sp. (1:1). Each treatment consisted of four replicates with 15 juveniles of tilapia (O. niloticus — 16.74± 4.35 g e 9.82±0.85 cm). The experiment lasted for 30 days and at the end of this period, three fish from each tank were killed, and the intestines were taken for microbiological analysis by FISH technique, where Bacillus and Enterococcus, as well as two putative pathogenic bacteria (Aeromonas and Pseudomonas sp.) were quantified. Enterococcus sp. and Bacillus sp. were present in high number in the gut microbiota of fish. However, Bacillus sp., showed an increase in its abundance, indicating a successful incorporation of this potential probiotic bacteria into the tilapia gut microbiota. Furthermore, in the Bacil. treatment it was observed a significant reduction of Aeromonas and Pseudomonas sp. abundances compared with the other treatments. These results indicate that the FISH technique is a potential tool to characterize the dynamics of potential probiotic bacteria and their efficiency in the control of pathogenic bacteria.