Integrative genomic characterization of five Pediococcus acidilactici strains reveals differing probiotic safety profiles

The increasing use of probiotics in livestock necessitates rigorous safety assessments to mitigate risks such as their inadvertent contribution to antimicrobial resistance (AMR) and horizontal gene transfer (HGT). This study employs whole-genome sequencing using both long-read (GridION, Oxford Nanopore Technologies) and short-read (Illumina, San Diego, CA, USA) platforms to assess the genomic and plasmidome profiles of five Thai strains of Pediococcus acidilactici, that previously have been evaluated for probiotic potential in livestock. Our comprehensive analysis identified genes encoding AMR, virulence factors, and probiotic-related genes. Notably, strains AF2519 and AF2019 harbored plasmid-borne tet(M) and erm(B) genes, with tet(M) embedded in a novel composite genetic arrangement flanked by mobile elements, suggesting historical recombination and altered mobility potential. Strains IAF6519, IAF5919, and P72N, free from plasmid-borne AMR genes, emerged as safer candidates, lacking virulence genes. Phenotypic tests revealed discrepancies with genomic data; for instance, AF2019 was resistant to clindamycin without detectable genes, and showed susceptibility to tetracycline despite the presence of tet(M). The absence of complete transfer machinery in AF2519 and AF2019 suggests a reduced HGT risk. These findings underscore the importance of integrating genomic and phenotypic approaches in probiotic safety evaluations. The presence of plasmid-borne AMR genes in certain strains advises caution in their use, impacting probiotic selection and regulatory compliance in agriculture. This research informs policies and best practices for safe probiotic deployment, ensuring both efficacy and safety.