Postbiotics secreted by Lactobacillus sakei EIR/CM-1 isolated from cow milk microbiota, display antibacterial and antibiofilm activity against ruminant mastitis-causing pathogens

Mastitis is one of the most important and multi-factorial diseases affecting dairy cows. This study was conducted to evaluate the effect of postbiotics secreted by lactic acid bacteria on mastitis. Within this aim, a total of 10 raw milk samples of healthy dairy cows were processed for the isolation of microbiota-derived bacteria. Following the isolation protocol, 24 isolates showing phenotypic characters similar to Lactobacillus species on specific MRS agar were selected and screened for their antibacterial activity against important mastitis-causing pathogens using the agar well diffusion method. According to the screening of the isolates, only one strain was found effective against all tested pathogens and selected for further assays. The isolate that exhibited a remarkable inhibitory effect was then identified as Lactobacillus sakei by 16S rRNA sequence analysis. The minimum inhibitory concentration of the postbiotics secreted by L. sakei into the culture medium was defined between the range of 40–50 mg/L. The antibacterial activity was also determined to be an acid-dependent effect, and HPLC analysis confirmed the presence of lactic acid (21.42 mg/mL) as the most common acid in the postbiotics. Among the 37 fatty acids, only oleic acid, palmitic acid, and stearic acid were detected. In addition to its antibacterial activity, postbiotics significantly reduced the biofilm formation of pathogens following the co-incubation, pre-treatment, and eradication assays, confirmed by confocal-laser and scanning electron microscopy. In conclusion, postbiotics with antibacterial and antibiofilm activity against important mastitis-causing pathogens can be used as a promising and natural agent in the prevention of mastitis.HIGHLIGHTS Lactobacillus sakei EIR/CM-1 was isolated from Holstein cow’s milk microbiota. Postbiotics from the strain EIR/CM-1 displayed the highest antibacterial activity against ruminant mastitis-causing pathogens. Co-incubation with postbiotics at 5 mg/mL and above concentrations reduced the biofilm formation of ruminant mastitis-causing pathogens by more than 70%. Lactic acid was found to be the highest organic acid in postbiotics