Exploring W1 and W2 Strains Isolated from Khao-Mahk As Probiotic Candidates: From Phenotypic Traits to Genomic Insights

Growing interest in probiotics has spurred research into their health benefits for hosts. This study aimed to evaluate the probiotic properties, especially antibacterial activities and the safety of two strains, W1 and W2, isolated from Khao-Mahk by describing their phenotypes and genotypes through phenotypic assays and whole genome sequencing. In vitro experiments demonstrated that both strains exhibited robust survival under gastric and intestinal conditions, such as in the presence of low pH, bile salt, pepsin, and pancreatin, indicating their favorable gut colonization traits. Additionally, both strains showed auto-aggregation and strong adherence to Caco2 cells, with adhesion rates of 86.86 ± 1.94% for W1 and 94.74 ± 2.29% for W2. These high adherence rates may be attributed to the significant exopolysaccharide (EPS) production observed in both strains. Moreover, they exerted remarkable antimicrobial activities against , serotype Typhi, , and , along with an absence of hemolytic activities and antibiotic resistance, underscoring their safety for probiotic application. Genomic analysis corroborated these findings, revealing genes related to probiotic traits, including EPS clusters, stress responses, adaptive immunity, and antimicrobial activity. Importantly, no transferable antibiotic-resistance genes or virulence genes were detected. This comprehensive characterization supports the candidacy of W1 and W2 as probiotics, offering substantial potential for promoting health and combating bacterial infections.