In situ expression of yak DEFB127 via plasmid-mediated delivery alleviates bovine Staphylococcus aureus-induced mastitis through anti-inflammatory mechanisms

Abstract

Mastitis, a disease affecting milk production and quality, poses a significant challenge to agricultural practitioners. Conventional antibiotic treatments present potential issues such as drug resistance, disruption of animal intestinal microbiota, and milk safety issues, necessitating the development of novel alternative therapies for bovine mastitis. Yak β-defensin 127 ( DEFB127 ), a natural antimicrobial peptide, represents a promising candidate for alternative to antibiotics, yet remains underexplored as a therapy for mastitis. In MAC-T cell, qRT-PCR analysis revealed that expression of IL-6 and IL-1β in the pLF- DEFB127 treatment group were significantly lower than the infection group ( P < 0.01). In vivo , histopathological examination showed that degree of inflammatory cell infiltration in the mammary gland tissue of the treatment group was significantly reduced compared to the infection group, accompanied by marked alleviation of alveolar dilation and epithelial cell necrosis. ELISA and qRT-PCR results demonstrated that the expression of TNF- α, IL-6 , and IL-1 β in the mammary gland tissue of mice from the pLF- DEFB127 treatment group were significantly downregulated than those in the infection group ( P < 0.01). Furthermore, 16S rRNA gene sequencing revealed that pLF- DEFB127 treatment altered the structure of the mouse intestinal microbiota, increasing the relative abundance of Bacteroidia and Bacilli, while decreasing that of Gammaproteobacteria. Finally, functional prediction of the intestinal microbiota based on 16S rRNA correlation analysis indicated that yak DEFB127 might exert anti-inflammatory effects by mechanisms involving glycosphingolipid biosynthesis and immune signaling, though this requires experimental validation.