Isolation and characterization of indigenous probiotic bacteria from wild Nile tilapia (Oreochromis niloticus) in Lake Naivasha, Kenya, with first evidence of Paucilactobacillus vaccinostercus as a potential aquaculture probiotic

Background and Aim: The growing burden of infectious diseases and antimicrobial resistance (AMR) in aquaculture demands safe, host-adapted alternatives to antibiotics. Probiotics derived from the gastrointestinal tract (GIT) of the target host are considered more ecologically compatible and effective than non-host strains. This study aimed to isolate and characterize indigenous gut bacteria from wild Nile tilapia (Oreochromis niloticus) collected from Lake Naivasha, Kenya, to evaluate their probiotic attributes, assess pathogenicity, and identify promising candidates using conventional phenotypic methods and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Materials and Methods: Thirty-eight apparently healthy O. niloticus were sampled, and bacteria were isolated from the entire GIT using standard bacteriological techniques. Fifty autochthonous isolates were obtained and subjected to stepwise screening, including hemolytic activity, antibiotic susceptibility, enzymatic activity (protease and amylase), tolerance to low pH and bile salts, adhesion to stainless steel surfaces, growth kinetics, and in vivo pathogenicity in O. niloticus. Species-level identification was performed using biochemical tests and MALDI-TOF MS. Results: Of the 50 isolates, 10 (20%) were nonhemolytic and sensitive to at least eight antibiotics. Functional screening reduced these to four candidates exhibiting enzymatic activity, acid and bile tolerance, and adhesion. Three isolates, identified as Rossellomorea marisflavi, Micrococcus luteus, and Paucilactobacillus vaccinostercus, were nonpathogenic to O. niloticus. In contrast, Aeromonas ichthiosmia, despite exhibiting several probiotic-like traits in vitro, caused 80% cumulative mortality and was excluded. Among the nonpathogenic isolates, P. vaccinostercus demonstrated the strongest overall probiotic profile, including superior acid and bile tolerance, high enzymatic indices, robust adhesion (~4.7 × 10⁴ CFU/mL), and favorable growth kinetics. Conclusion: The gut microbiota of wild O. niloticus from Lake Naivasha harbors a limited but valuable pool of indigenous bacteria with probiotic potential. This study provides the first evidence that P. vaccinostercus is a promising, nonpathogenic probiotic candidate for tilapia aquaculture. These findings support the development of locally adapted, antibiotic-free probiotic strategies to enhance fish health and sustainable aquaculture in Kenya. Further in vivo feeding trials and genomic safety assessments are warranted. Keywords: aquaculture, aquaculture probiotics, fish gut microbiota, Kenya aquaculture, Lake Naivasha, MALDI-TOF MS, Nile tilapia, Paucilactobacillus vaccinostercus, sustainable aquaculture.