In vitro evaluation of synergistic antimicrobial effect of lactoferrin and nisin along with penicillin and gentamicin, and potential application in mastitis treatment

Mastitis is a leading challenge to the production and welfare of dairy animals, caused by a variety of pathogens. Existing antimicrobial strategies have failed to treat mastitis while contributing to the development and dissemination of antimicrobial resistance. This situation demands investigation of the antimicrobial properties of new candidates. Lactoferrin and nisin have been reported to have strong antimicrobial effects in previous studies. This study aimed to assess the synergistic antimicrobial activities of lactoferrin, nisin, and antibiotics (penicillin, gentamicin) alone and in combination against major pathogens, including Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae, and Streptococcus uberis. 100 dairy cows were selected for milk sampling from different herds in the district of Bahawalnagar, Pakistan. The California mastitis test was performed to screen for cases of mastitis. Bovine lactoferrin was purchased, the MIC of the mentioned bacteria was determined, and the effect of lactoferrin concentrations on the growth of these bacteria was assessed. After isolating the bacteria through colony culture, different combinations were applied to these bacterial colonies. The minimum inhibitory concentration of lactoferrin for L. monocytogenes, S. uberis, and E. coli was recorded 0.1%, 0.5%, and 1%, respectively. The application of nisin for the treatment of subclinical mastitis is advantageous 93% of the time. Sensitivity analysis reveals that treatment with nisin is beneficial for 73% of primiparous cows. Significant synergistic effects against resistant microbes were observed with different combinations, especially the combination of lactoferrin and nisin with penicillin. This in vitro study may be useful in establishing formulations to combat antimicrobial resistance issues at the commercial level. These findings can also help design further effective research plans to set the dose rate of these animal-origin compounds in combination with existing antibiotics to mitigate mastitis at the herd level.