Mastitis is one of the most prevalent diseases in dairy farming worldwide. The gut microbiota plays an important role in the regulation of systemic and local inflammatory diseases, such as mastitis. However, the regulatory mechanism of the gut microbiota on mastitis is still unclear. Thus, the aim of this study was to investigate the function and regulatory mechanisms of the gut microbiota in host defense against mastitis caused by Staphylococcus aureus (S. aureus) infection. Increased blood-milk barrier permeability, and S. aureus-induced mastitis severity were observed gut microbiota-dysbiosis mice compared with those in control mice. Moreover, feces microbiota transplantation (FMT) to microbbiota-dysbiosis mice reversed these effects. Furthermore, established disruption of commensal homeostasis results in significantly increased abundance of pathogenic Enterobacter bacteria, while the relative abundance of short-chain fatty acid (SCFAs)-producing bacterial phyla (Firmicutes and Bacteroidetes) was significantly reduced. However, FMT to gut microbiota-dysbiosis mice reversed these changes. In addition, dysbiosis reduced the levels of SCFAs, and administration of sodium propionate, sodium butyrate, and probiotics (butyrate-producing bacteria) reversed the changes in the blood-milk barrier and reduced the severity of mastitis induced by S. aureus. In conclusion, this new finding demonstrated that the gut microbiota acts as a protective factor in host defense against mastitis and that targeting the gut-mammary gland axis represents a promising therapeutic approach for mastitis treatment.

Thymol is a natural monoterpene phenol primarily found in thyme, oregano, and tangerine peel. It has been shown to possess anti-inflammatory property both in vivo and in vitro. In the present paper, we studied the anti-inflammatory effect of thymol in lipopolysaccharide (LPS)-stimulated mouse mammary epithelial cells (mMECs). The mMECs were stimulated with LPS in the presence or absence of thymol (10, 20, 40 μg/mL). The concentrations of tumor necrosis factor α (TNF-α), interleukin (IL)-6, and IL-1β in the supernatants of culture were determined using enzyme-linked immunosorbent assay. Cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), extracellular signal-regulated protein kinase (ERK), c-Jun N-terminal kinase (JNK), nuclear factor-κB (NF-κB), and inhibitor protein of NF-κB (IκBα) were measured using western blot. The results showed that thymol markedly inhibited the production of TNF-α and IL-6 in LPS-stimulated mMECs. The expression of iNOS and COX-2 was also suppressed by thymol in a dose-dependent manner. Furthermore, thymol blocked the phosphorylation of IκBα, NF-κB p65, ERK, JNK, and p38 mitogen-activated protein kinases (MAPKs) in LPS-stimulated mMECs. These results indicate that thymol exerted anti-inflammatory property in LPS-stimulated mMECs by interfering the activation of NF-κB and MAPK signaling pathways. Thereby, thymol may be a potential therapeutic agent against mastitis.

The intestinal microbiota has been associated with the occurrence and development of mastitis, which is one of the most serious diseases of lactating women and female animals, but the underlying mechanism has not yet been elucidated. Aryl hydrocarbon receptor (AhR) activation by microbiota tryptophan metabolism-derived ligands is involved in maintaining host homeostasis and resisting diseases. We investigated whether AhR activation by microbiota-metabolic ligands could influence mastitis development in mice. In this study, we found that AhR activation using Ficz ameliorated mastitis symptoms, which were related to limiting NF-κB activation and enhancing barrier function. Impaired AhR activation by disturbing the intestinal microbiota initiated mastitis, and processed Escherichia coli (E. coli)-induced mastitis in mice. Supplementation with dietary tryptophan attenuated the mastitis, but attenuation was inhibited by the intestinal microbiota abrogation, while administering tryptophan metabolites including IAld and indole but not IPA, rescued the tryptophan effects in dysbiotic mice. Supplementation with a Lactobacillus reuteri (L. reuteri) strain with the capacity to produce AhR ligands also improved E. coli-induced mastitis in an AhR-dependent manner. These findings provide evidence for novel therapeutic strategies for treating mastitis, and support the role of metabolites derived from the intestinal microbiota in improving distal disease.

Mastitis is a common and frequently occurring disease of humans and animals, especially in dairy farming, which has caused huge economic losses and brought harmful substance residues, drug-resistant bacteria, and other public health risks. The traditional viewpoint indicates that mastitis is mainly caused by exogenous pathogenic bacteria infecting the mammary gland.

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