Bovine mastitis is the most important infectious disease on dairy farms. Conventional antibiotic therapy is often unsatisfactory and alternative treatments are continually under investigation. Lactobacillus (Lb.) perolens CRL 1724 and Lactobacillus plantarum CRL 1716 were previously isolated from milk of dairy cows and selected according to their potential probiotic properties. In the present work the in-vitro capacity of Lactobacillus strains to adhere to bovine teat canal epithelial cells (BTCEC) and to inhibit and co-aggregate 14 mastitis-causing pathogens (MCPs) was investigated. The effect of Lb. perolens CRL 1724 after intramammary inoculation in lactating cows was evaluated through determination of clinical signs of mastitis, milk appearance, somatic cell counts and Lb. perolens CRL 1724 recovery from milk. Lb. perolens CRL 1724 was able to inhibit 12 of 14 MCPs (85·7%) in vitro, especially those considered to be major pathogens. In addition, Lb. perolens CRL 1724 co-aggregated with all of them. Lb. plantarum CRL 1716 was able to inhibit 7 of 14 MCPs (50%) in vitro and showed co-aggregation ability similar to Lb. perolens CRL 1724. Lb. perolens CRL 1724 showed a higher efficacy of adhesion to BTCEC (values of percentage of adhesion and adhesion index of 75% and 14·4, respectively) than Lb. plantarum CRL 1716 (37% and 7·4, respectively). Lb. perolens CRL 1724 was recovered from all mammary quarters and no clinical signs or teat damage were observed after the inoculation of 106 cfu/ml. The udders presented a normal aspect and there were no changes in the appearance of the milk. The results obtained will serve as the basis for further trials to evaluate the potential of Lb. perolens CRL 1724 to be included in a non-antibiotic formulation for the prevention of bovine mastitis.

A vaccine was developed against bovine mastitis based on inactivated, highly encapsulated Staphylococcus aureus cells; a crude extract of Staph. aureus exopolysaccharides; and inactivated, unencapsulated Staph, aureus and Streptococcus spp. cells. This vaccine was tested on 30 heifers during a 7-mo period. The 30 heifers were randomly assigned to three groups of 10 heifers each. The prepartum group received two injections of the vaccine at 8 and 4 wk before calving, and the postpartum group received two injections at 1 and 5 wk after calving. The control group received two injections of a placebo at 8 and 4 wk before calving. The vaccine or the placebo was administered subcutaneously in the brachiocephalicus muscle of the neck. The frequencies of intramammary infections caused by Staph. aureus were reduced from 18.8% for heifers in the control group to 6.7 and 6.0% for heifers in the prepartum and postpartum groups, respectively. This protective effect was maintained for at least 6 mo. The relative risk of mastitis caused by Staph. aureus was 0.31 and 0.28 for heifers in the prepartum and postpartum groups, respectively, compared with that for heifers in the control group. The results of the trial indicated the effectiveness of the vaccine in decreasing the incidence of intrammammary infections caused by Staph. aureus. A slight but nonsignificant increase occurred in fat production in the milk of vaccinated cows. The vaccine had no observable effect on somatic cell count or streptococcal infections.

The use of lactic acid bacteria (LAB) in animal feed, constitute an alternative tool for bovine mastitis prevention. Previously, two LAB strains were isolated from bovine milk and selected for their probiotics properties. So far, immune response of inoculating LAB in bovine udders at dry-off period has not been investigated. The immunoglobulin isotype levels and memory cell proliferation in blood and milk of animals inoculated with Lactobacillus lactis subsp. lactis CRL1655 and Lactobacillus perolens CRL1724 at dry-off period was studied. Ten animals were inoculated intramammarily with 10 cells of each LAB (IG) and 2 animals used as control (NIG). Milk and blood samples were taken before inoculation and 1, 2, 4, 6, 12 and 24 h and 7 and 14 days after inoculation. Somatic cell count (SCC) in milk, the presence of bovine mastitis pathogens, the levels of antibodies and lymphocyte proliferation were determined. In the IG, the SCC was <250,000 cells/ml up to 4 h after intramammary inoculation. Six and 12 h after inoculation, the SCC increased up to 600,000 and 2,000,000 cells/ml, respectively. In the NIG, the SCC reached the maximum value 7 days after inoculation. Microbiological analysis showed that all samples were negative for major bovine mastitis pathogens after 24-48 h of incubation. In general, LAB inoculation increased the amount of IgG isotypes in blood and milk, and these antibodies were able to recognise Staphylococcus aureus epitopes. Lymphocytes proliferation was significantly higher in the IG at all time points assayed, following LAB or S. aureus stimulation. The lymphocytes of animals inoculated with LAB do not react in vitro to the presence of S. aureus antigen.. The results showed that probiotic microorganisms could be a natural and effective alternative in the prevention of bovine mastitis at dry-off period and act as immunomodulatory stimulating local and systemic defence lines.

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