Akkermansia muciniphila-Derived Outer Membrane Vesicles as a Novel Therapeutic Approach for Mastitis: Insights From In Vitro and Vivo Studies.

Mastitis is a major disease affecting the dairy industry. Although antibiotics are the most prevalent treatment for mastitis, their overuse presents significant risks to public health. Probiotic therapy has emerged as a promising alternative for controlling mastitis, but there is a lack of comprehensive studies on the efficacy and mechanisms of specific probiotics in treating this condition. Here, we investigate the potential of Akkermansia muciniphila (A. muciniphila) and its derived outer membrane vesicles (AOMVs) as therapeutic agents for mastitis. Significant differences in microbial communities were identified through an analysis of the microbiota composition in milk from healthy and mastitis cows. Spearman's correlation analysis revealed a significant negative correlation between the relative abundances of Verrucomicrobia and Akkermansia and the levels of inflammatory cytokines in mastitis milk. Furthermore, we evaluated the roles of live and pasteurized A. muciniphila in lipopolysaccharide (LPS)-induced inflammatory responses in bovine mammary epithelial cells (MAC-T). Compared to pasteurized A. muciniphila, live A. muciniphila exhibited a stronger anti-inflammatory effect, notably inhibiting the TLR4 and NF-κB signaling pathways. We also demonstrated that these effects of A. muciniphila may be mediated through AOMVs. Subsequently, the therapeutic potential of AOMVs was evaluated in vivo using an LPS-induced mastitis mouse model. The results demonstrated that AOMVs targeted the mammary glands, alleviated LPS-induced damage, and reduced inflammatory cytokine levels and signaling pathway activation. Finally, we conducted a preliminary investigation to assess the therapeutic effects of A. muciniphila on mastitis in dairy cows. Administration of A. muciniphila significantly reduced somatic cell count (SCC) in the milk of cows with mastitis. Notably, four of the five treated cows showed a decrease in SCC to below 2 × 10(5) cells/mL, with negative results in the California Mastitis Test. Our findings provide compelling evidence for the therapeutic potential of A. muciniphila and AOMVs in the treatment of mastitis. These results not only highlight the anti-inflammatory properties of A. muciniphila as a potential probiotic, proposing novel strategies for developing alternatives to antibiotic therapies, but also provide a theoretical basis for exploring the role of outer membrane vesicles in the host-microbe interface.