Antagonism against fish pathogens by cellular components and verification of probiotic properties in autochthonous bacteria isolated from the gut of an Indian major carp,Catla catla(Hamilton)

Abstract

Probiotic potential of the autochthonous bacteria in catla, Catla catla has been evaluated through determination of antagonistic activity (in vitro) of the cellular components of gut bacteria against seven fish pathogens. Altogether 208 strains were isolated, inhibitory activity of the isolates was evaluated through cross-streaking and 16 primarily selected antagonistic strains were confirmed using the double-layer method. Four bacteria that showed antagonism against ≥4 pathogens were selected as putative probiotics. The intracellular, extracellular, whole-cell and heat-killed cell components exhibited bactericidal activity against the pathogens. In addition, the selected strains were capable of producing different extracellular enzymes, competent to grow in intestinal mucus and could tolerate diluted bile juice. Analysis of 16SrRNA partial gene sequence revealed that both the strains CC1FG2 and CC1FG4 were Bacillus methylotrophicus (KF559344 and KF559345), while the isolates CC1HG5 and CC2HG7 were Bacillus subtilis subsp. spizizenii (KF559346) and Enterobacter hormaechei (KF559347) respectively. Bio-safety evaluation through intra-peritoneal injection of the isolates did not induce any pathological signs or mortalities in C. catla. The study confirmed probiotic properties of autochthonous gut bacteria in C. catla and demonstrated potential for using them as biocontrol agents. However, in vivo studies are essential to explore their efficacy in the commercial aquaculture.

Screening of Autochthonous Intestinal Microbiota as Candidate Probiotics Isolated from Four Freshwater Teleosts

“…According to Ramesh et al 40 , the bile tolerance ability of probiotic candidate is an important characteristic, as it has to work in the gut environment (high bile concentration). Similar observations have also been reported by probiotic bacteria 19,39 . The evaluation of haemolytic activity is another important selection criterion of probiotic strain, as haemolytic bacteria may cause malfunction in the defence system 41,42 .…”

“…The putative probiotic candidates were grown in TSB (pH 7.0), biomass was collected using centrifugation (3000 g for 20 min) and suspended in 0.1 mM PSB. Then 100 l solution (approximately 10 9 CFU/ml) was intraperitoneally injected to experimental fish, whereas 100 l PBS solution (without bacteria) was injected to control fish 19 . The swimming behaviour and health status were monitored up to 14 days.…”

“…During the last decade, attempts have been made by researchers to isolate probiotic bacteria from indigenous or exogenous microbiota of aquatic animals 13 . Although several studies have reported the efficiency of probiotics in aquaculture isolated from marine fish and shellfish [14][15][16][17][18] , only a few reports have been published regarding probiotic bacteria isolated from the gut of freshwater teleosts 19,20 . In this study, four freshwater teleosts were selected for isolation, screening and molecular identification of autochthonous gut bacteria having favourable probiotic properties such as antagonistic characteristics, production of extracellular digestive enzymes, bile tolerance, nonhaemolytic nature and bio-safety aspects.…”

Evaluation of Gut Associated Extracellular Enzyme-producing and Pathogen Inhibitory Microbial Community as Potential Probiotics in Nile Tilapia, <i>Oreochromis niloticus</i>

“…Thus, the study might suggest that extracellular enzyme-producing gut bacteria and yeast may restrain the growth of some fish pathogens and tolerate conditions within the GI tract. The present observation was in agreement to the previous reports establishing probiotic attributes of fish gut bacteria (Geraylou et al, 2014;Mukherjee and Ghosh, 2016). Although to the author's knowledge, such attempt has never been made to characterize yeasts as likely probiotics.…”

“…Apart from nutritional benefits, the enzyme-producing gut bacteria in fish have been assumed to compete continuously with pathogens through the competitive exclusion or the production of antimicrobial compounds (Ray et al, 2012). Although several strains of Bacilli were demonstrated as probiotics for fish, antagonism of pathogens by the Bacilli has been seldom indicated (Lalloo et al, 2010;Geraylou et al, 2014;Mukherjee and Ghosh, 2016). In the present study, most of the promising exo-enzyme producing bacteria and yeast strains displayed antagonism against at least two of the tested fish pathogens.…”