Isolation and Characterization of Gut-derived Probiotic Bacteria from Clarias batrachus with Antagonistic Activity against Aquaculture Pathogens

Abstract

Diseases caused by opportunistic bacteria such as Aeromonas hydrophila and Vibrio alginolyticus remain a major constraint in intensive aquaculture, while the use of antibiotics raises concerns about resistance and environmental risks.This research aimed to isolate and characterize gut-derived probiotic bacteria from Clarias batrachus and to evaluate their antagonistic activity against major aquaculture pathogens.Intestinal bacterial isolates were obtained through selective culturing, followed by biochemical and phenotypic characterization, antagonistic testing, agar-diffusion antibacterial assays, pH tolerance evaluation, and pathogenicity testing using the brine shrimp lethality test (BSLT).Six isolates were obtained, of which four (BP-1, BP-3, BP-4, and BP-6) exhibited significant inhibitory activity (P<0.05) against both pathogens, whereas two (BP-2 and BP-5) were inactive.The BP-3 isolate showed the best antibacterial activity against A. hydrophila and V. alginolyticus, with inhibition zone diameters of 14.33 mm and 13.67 mm, respectively.The active isolates were predominantly Gram-positive, catalase-variable, and tolerant of acidic conditions (pH 2-6), indicating suitability for survival in the gastrointestinal tract.Pathogenicity testing confirmed all active isolates as non-toxic (LC50>3510 3 ppm).These findings demonstrate that the intestine of C. batrachus harbors host-associated probiotic candidates with strong antibacterial potential and promising biosafety attributes.Based on biochemical and microscopic characterization, all four isolates were suspected to be Lactobacillus, Bacillus, and Eubacterium.Further molecular identification and in vivo validation are recommended to confirm functional stability and the feasibility of application.

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