title: Antibacterial activity and immunomodulatory effects on a bovine mammary epithelial cell line exerted by nisin A-producing Lactococcus lactis strains.
abstract: Twenty-nine strains of mastitis pathogens were used to study the antibacterial activity of the cell-free supernatants (CFS) of 25 strains of Lactococcus lactis ssp. lactis. Out of the tested strains, only the CFS of L. lactis LL11 and SL153 were active, inhibiting and killing most of the pathogens. By means of ultra-performance liquid chromatography/high resolution mass spectrometry, they were shown to produce nisin A, a class I bacteriocin. A variable sensitivity to nisin A-containing CFS was observed among Streptococcus uberis and Enterococcus faecalis strains. Nonetheless, Streptococcus agalactiae, Strep. uberis, and E. faecalis displayed high minimum inhibitory concentration values, reaching 384 arbitrary units/mL. Interestingly, the minimum inhibitory values and the bactericidal concentrations were almost identical among them for each of the 2 stains, LL11 and SL153. Staphylococci were, on average, less sensitive than streptococci, but the 2 CFS inhibited and killed, at different dilutions, strains of methicillin-resistant Staphylococcus aureus. The immune response to nisin A-containing CFS was tested using the bovine mammary epithelial cell line BME-UV1. Application of CFS did not damage epithelial integrity, as demonstrated by the higher activity of N-acetyl-β-d-glucosaminidase (NAGase) and lysozyme inside the cells, in both treated and control samples. On the other hand, the increase of released NAGase after 15 to 24h of treatment with LL11 or SL153 live cultures demonstrated an inflammatory response of epithelial cells. Similarly, a significantly higher lysozyme activity was detected in the cells treated with LL11 live culture confirming the stimulation of lysosomal activity. The treatment of epithelial cells with SL153 live culture induced a significant tumor necrosis factor-α downregulation in the cells, but did not influence IL-8 expression. The control of tumor necrosis factor-α release could be an interesting approach to reduce the symptoms linked to clinical intramammary infections. Due to their antibacterial activity and to the stimulation of lysosomal activity of mammary epithelial cells, the L. lactis strains SL153 and LL11 could be of interest for the development of alternative intramammary treatments to control cow mastitis.
authors.affiliation: Department of Veterinary Science and Public Health, University of Milan, via Celoria 10, 20133 Milan, Italy.
authors.forename: M
authors.initials: M
authors.lastname: Malvisi
authors.affiliation: Department of Food, Environmental and Nutritional Sciences (DeFENS), University of Milan, via Celoria 2, 20133 Milan, Italy.
authors.forename: M
authors.initials: M
authors.lastname: Stuknytė
authors.affiliation: Department of Veterinary Science and Public Health, University of Milan, via Celoria 10, 20133 Milan, Italy.
authors.forename: G
authors.initials: G
authors.lastname: Magro
authors.affiliation: Department of Veterinary Science and Public Health, University of Milan, via Celoria 10, 20133 Milan, Italy.
authors.forename: G
authors.initials: G
authors.lastname: Minozzi
authors.affiliation: Clerici-Sacco Group, Via Manzoni, 29/A, 22071 Cadorago, Italy.
authors.forename: A
authors.initials: A
authors.lastname: Giardini
authors.affiliation: Department of Food, Environmental and Nutritional Sciences (DeFENS), University of Milan, via Celoria 2, 20133 Milan, Italy.
authors.forename: I
authors.initials: I
authors.lastname: De Noni
authors.affiliation: Department of Veterinary Science and Public Health, University of Milan, via Celoria 10, 20133 Milan, Italy. Electronic address: renata.piccinini@unimi.it.
authors.forename: R
authors.initials: R
authors.lastname: Piccinini
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001419:Bacteria / Q000187:drug effects*; D001430:Bacteriocins / Q000494:pharmacology*; D002417:Cattle; D018929:Cell Culture Techniques; D002474:Cell-Free System; D002851:Chromatography, High Pressure Liquid / Q000662:veterinary; D004847:Epithelial Cells / Q000187:drug effects; D005260:Female; D007113:Immunity, Innate / Q000187:drug effects; D056747:Immunomodulation / Q000187:drug effects*; D013294:Lactococcus lactis / Q000737:chemistry*; D013058:Mass Spectrometry / Q000662:veterinary; D008414:Mastitis, Bovine / Q000276:immunology* / Q000382:microbiology
chemical_list: D000900:Anti-Bacterial Agents; D001430:Bacteriocins
keywords: Lactococcus lactis; dairy cow mastitis; mammary epithelial cells; nisin A; ultra-performance liquid chromatography/high resolution mass spectrometry