# Lactic Acid Bacteria Metabolites Modulate Immune Response Against Staphylococcus haemolyticus-Infected RAW264.7 Murine Macrophage: A Novel Approach for Bovine Mastitis

## TL;DR

This study explores how natural compounds from lactic acid bacteria can boost cow immunity against a common mastitis-causing bacteria, offering a potential antibiotic-free treatment.

## Contribution

The study identifies lactic acid bacteria metabolites, especially from Enterococcus faecalis, as novel immunostimulatory agents for bovine mastitis.

## Key findings

- LAB metabolites increased nitric oxide production and activated immune signals in infected macrophages.
- Enterococcus faecalis metabolites upregulated inflammatory genes like iNOS, COX-2, TNF-α, and IL-6.
- Strain-specific metabolic profiles of LAB were linked to varying immunostimulatory effects.

## Abstract

Bovine mastitis, a painful udder infection in dairy cows, is a major economic burden for farmers. Although antibiotics are commonly used, their overuse of the antibiotics leads to resistance and milk safety concerns. Natural compounds from beneficial lactic acid bacteria that might help cows fight Staphylococcus haemolyticus, a common mastitis pathogen, were investigated in this study. In lab tests using mouse immune cells, these compounds boosted defenses by increasing nitric oxide (a natural antibacterial agent) and activating inflammation-related immune signals. Metabolites from Enterococcus faecalis were especially effective. These results suggested that such natural compounds could become antibiotic-free treatments for mastitis. This approach would benefit dairy farmers by lowering costs, improvement of cow welfare, and providing consumers with safer milk by reducing antibiotic use. Therefore, the compounds from the beneficial bacteria have potential as a sustainable solution for mastitis control.

Bovine mastitis, an inflammation of the mammary gland, is recognized as the most prevalent disease in dairy cattle, leading to significant economic losses due to the reduction in quality and yield of milk. While antibiotic treatment remains to be the primary control method but their use of antibiotics contributes to the emergence of antibiotic-resistant bacteria and possesses potential health risks to consumers. Enhancement of host immune responses represents a promising alternative strategy for combating pathogenic bacteria. This study aimed to evaluate the immunomodulatory potential of metabolites derived from lactic acid bacteria (LAB) in murine macrophage RAW264.7 cells infected with Staphylococcus haemolyticus, a causative agent of bovine mastitis. Notably, LAB-derived metabolites demonstrated significant, strain-specific immunostimulatory activity. A comparative metabolomic analysis confirmed that each strain possessed a unique metabolic profile, providing a chemical basis for these various responses. The most pronounced effects were observed with metabolites from the isolated strain Enterococcus faecalis, which markedly increased NO production. Furthermore, these metabolites upregulated the expression of key inflammatory genes, e.g., iNOS, COX-2, TNF-α, and IL-6 and enhanced the protein levels of iNOS and COX-2. These findings suggest that LAB metabolites, particularly those from E. faecalis, may offer a novel therapeutic approach for enhancing immune defenses against mastitis-causing pathogens.