Strain-Specific Safety Evaluation of Akkermansia muciniphila Akk11: Comprehensive Genotypic, Phenotypic, and...

Akkermansia muciniphila is increasingly regarded as a next-generation probiotic with clinical and commercial potential. This study aimed to evaluate the strain-specific safety and probiotic characteristics of Akk11, isolated from the feces of healthy infants. The genotypic profile of Akk11 was assessed via whole-genome sequencing, including antibiotic resistance, virulence factors, and metabolic pathway annotations. Phenotypic analyses included antibiotic susceptibility, mucin degradation, biogenic amine and D-/L-lactic acid production, and hemolytic activity. Stress tolerance and cytotoxicity were performed using simulated gastrointestinal conditions and Caco-2 cells. In vivo safety was further evaluated through acute oral toxicity, bacterial reverse mutation, mammalian erythrocyte micronucleus, and 90-day sub-chronic toxicity, following international guidelines. Akk11 showed 98.36% genomic similarity to the type strain adeF with no mobile genetic elements or transferable resistance. Phenotypically, Akk11 exhibited intrinsic resistance to several antibiotics while lacking harmful metabolic activities and excellent simulated gastrointestinal stress tolerance with non-cytotoxicity. No adverse effects were observed in any of the in vivo toxicity studies, even under long-term oral administration of bacterial powder preparations containing both viable and non-viable cells. The NOAEL was 9 × 10¹¹ AFU/kg/day, the highest dose tested in both sexes, using a lyophilized preparation that contained approximately 20%-30% non-viable Akk11 cells (n-AFU). Akk11 possesses a robust, strain-specific safety profile and probiotic potential, providing a scientific basis for its application in next-generation probiotic products.