PHENOTYPIC AND GENOTYPIC DIVERSITY AMONG POTENTIAL NEXT-GENERATION PROBIOTICS

The gut microbiota constitutes a complex and dynamic community of microorganisms that plays a central role in host physiology, including digestion, metabolic regulation, immune function, and maintenance of intestinal barrier integrity. In the gut of healthy individuals, major bacterial phyla such as Firmicutes, Bacteroidetes, Actinobacteria, and Verrucomicrobia co-exist in a balanced state that supports homeostasis and protects against pathogenic colonization. Among these, Akkermansia muciniphila, a Gram-negative anaerobe localized in the colonic mucus layer, has gained increasing attention due to its functional relevance. This bacterium degrades mucin glycoproteins, thereby contributing to mucus turnover, strengthening barrier function, and limiting systemic exposure to pro-inflammatory molecules. In addition, A. muciniphila participates in metabolic cross-talk within the microbiota through short-chain fatty acid production and modulation of host immune responses, making it a key indicator of gut health. In this study, multiple A. muciniphila isolates were comparatively evaluated against the reference strain MucT to investigate their peculiar functional properties. The results highlighted strain-specific activities, with pasteurized forms consistently outperforming their live counterparts in several in vitro assays. In particular, pasteurized cells promoted enhanced epithelial repair, increased ATP and NAD⁺ production, and stronger activation of hTLR2 and SIRT1 signaling pathways. These findings were supported by in vivo experiments in Caenorhabditis elegans, where nematodes fed with A. muciniphila revealed an extended lifespan, together with improved healthspan indicators, including locomotion and neuromuscular function. Notably, these effects were highly strain-dependent and associated with distinct metabolic profiles, especially in terms of short-chain fatty acid production. Safety assessment confirmed the absence of antimicrobial resistance genes and virulence-related traits, supporting the suitability of A. muciniphila for biotechnological applications. Overall, this work reinforces the concept of A. muciniphila as a next-generation probiotic (NGP) with potential applications beyond metabolic health. The use of pasteurized formulations represents a stable and safe alternative to live microorganisms, with promising systemic effects. However, further validation will be essential to translate these findings into effective nutritional strategies.