Deciphering the probiotic potential of Limosilactobacillus reuteri S8 in broilers through phenotypic and genomic analyses

The ban on antibiotic growth promoters in poultry has increased the need for probiotics. This study evaluates Limosilactobacillus reuteri S8, isolated from the gastrointestinal tract of broiler chickens, using in vitro assays, whole-genome sequencing and comparative pan-genomics. The strain showed a log-based survival rate of 85% at pH 3.0 (a 1.2 log₁₀ reduction in viable count over 3 h) and a relative growth of 50.5% in 0.3% bile salts, consistent with the presence of the atp operon and nhaC . It adhered to Caco-2 cells more densely than a low-adhesion reference strain, a favourable prerequisite for potential competitive exclusion that warrants further verification in vivo, and the genome encodes a sortase A homologue, a putative adhesin and exopolysaccharide biosynthesis genes. Under the in vitro conditions tested, it displayed antagonistic activity against all indicator organisms, including reference strains of Escherichia coli ATCC 25,922 and Salmonella enterica ATCC 14,028, consistent with multiple lactate dehydrogenase variants and a putative Class III bacteriocin locus. Pan-genome analysis with 62 reference genomes identified 155 strain-specific gene clusters, mainly associated with cellular biogenesis and DNA maintenance. The strain was resistant to tetracycline and ampicillin, and the genome carries the acquired antimicrobial resistance genes (ARGs) tet(W) , tet(L) and lnu(A) ; tet(L) lies next to a recombinase gene ( xerC ), which warrants investigation of its possible mobility. Therefore, while commercial application is cautioned against due to the presence of acquired ARGs, this strain remains a valuable biological model for investigating antimicrobial resistance reservoirs in avian commensals.