In the present study, we aimed to determine the antimicrobial resistance and molecular typing of Staphylococcus aureus recovered from transient and persistent intramammary infections and nares/muzzles in dairy cows. We investigated the antimicrobial resistance of 189 S. aureus strains using a broad antimicrobial susceptibility profile. Furthermore, 107 S. aureus isolates were strain-typed using staphylococcal protein-A (spa) typing. A large proportion of strains exhibited multidrug resistance to antimicrobials, including resistance to critically important antimicrobials, although no methicillin-resistant S. aureus strains were found. Our study did not strengthen the idea that extramammary niches (i.e., nares/muzzles) are an important source of S. aureus for bovine mastitis. A discrepancy in the antimicrobial resistance between S. aureus strains isolated from nares/muzzles and milk samples was observed. Furthermore, S. aureus isolates from transient and persistent intramammary infections (IMIs) did not differ by spa typing, suggesting that the persistence of bovine IMIs was determined by cow factors. Thus, the high level of multidrug-resistant S. aureus found in the two herds, considered together with the predominance of a well udder-adapted S. aureus strain, may contribute to our knowledge of the history of the high prevalence of mastitis caused by S. aureus, which is of great concern for animal and public health.

Staphylococcus aureus is one of the pathogens most frequently isolated from cases of mastitis worldwide. To decrease the effect of S. aureus mastitis in dairy farming, alternative strategies for controlling mastitis are needed that depend on a better knowledge of cowto-cow variations in S. aureus antibody production. The present study sought to explore the diversity of S. aureus antibodies produced by dairy cows with a distinct mastitis history and vaccinated with a polyvalent mastitis vaccine. We obtained protein extracts from S. aureus isolates derived from persistent subclinical mastitis. Proteins were fractionated using 2-dimensional gel electrophoresis and Western blotting. Then, Western blotting membranes were exposed to sera from 24 dairy cows that had been divided into the following groups: vaccinated dairy cows that were infected with S. aureus, further subdivided according to whether they (a) remained infected by S. aureus or (b) recovered from the intramammary infection; unvaccinated dairy cows infected with S. aureus; and vaccinated healthy dairy cows with no history of S. aureus mastitis. Proteins found to be reactive by Western blot were identified by mass spectrometry (MALDI/TOF-TOF). Our most important finding was that F0F1 ATP synthase subunit a, succinyl-diaminopimelate desuccinylase, and cysteinyl-tRNA synthetase were potential candidate proteins for the prevention of S. aureus mastitis. This study strengthens the notion that variations among animals should not be ignored and shows that the heterogeneity of antibody production against anti-staphylococcal antigens in animals may enable the identification of new immunotherapy targets.

The somatic cell count (SCC) of refrigerated raw milk reflects the occurrence of mastitis in the herd and the management of rural property. Therefore, this study aimed to evaluate the SCC of the refrigerated raw milk and its determinant factors in dairy properties of Viçosa, State of Minas Gerais. Monthly, milk samples were taken from 44 cooling tanks during 2012. A structured questionnaire was applied to evaluate the production and management characteristics of the herds. There was a significant variation (p < 0.05) in the mean SCC of the milk, which was lower from June to August. Higher mean SCC and percentages of samples above the limit set by the legislation were observed in months of higher rainfall and ambient temperature (p < 0.05). Mechanical milking, lower milk yield, productivity, inadequate milking procedures, equipment hygiene, and water quality were all factors that influenced (p < 0.05) the SCC of raw refrigerated milk. Milker training is required for production of low SCC milk.

SUMMARY Anthelmintics are used to combat nematodes. The misuse of anthelmintics can raise the cost of milk production. The objective of this research was to determine the presence of anthelmintics in goat milk. Twenty goats were used, divided into four groups of five animals: I- animals treated with an ivermectin-based anthelmintic; II- animals treated with moxidectin; III- animals treated with levamisole hydrochloride; and IV: animals treated with albendazole. Milk samples were collected individually: before, and 1, 2, 3, 15 and 21 days after administration of the anthelmintics. Determination of anthelmintic residues was performed by liquid chromatography-mass spectrometry (LC-MS). According to the results, there was an exponential effect (P<0.05) for ivermectin and moxidectin. Moxidectin was the anthelmintic that left a residue in the milk for the longest time, up to 21 days. However, with all the anthelmintics researched, residues were below the maximum limit recommended by the inspecting agencies.

New approaches are needed to quickly indicate possible contamination of UHT milk, among them the technique of ATP-Bioluminescence. Therefore, the aim of this study was to compare the results of culture methods with the results of ATP-Bioluminescence technique of 102 UHT whole milk samples incubated at 48, 72, and 168 hours. UHT milk samples were analyzed for the presence of mesophilic and psychrotrophic aerobic microorganisms using Plate Count Agar (PCA), Brain-Heart Infusion (BHI) media and Petrifilm^TM Aerobic Count (AC) plates. The ATP-Bioluminescence technique was applied through the Microbial Luminescent Screening (MLS) system. Significant correlations were found between counts of aerobic mesophilic microorganisms on PCA, Petrifilm^TM AC, BHI and results of ATP bioluminescence technique (P≤0.05). The ATP-Bioluminescence technique had higher correlation with counting method in PCA than BHI media. At lower pass/fail limits of Relative Light Units (60, 50, 45 and 40 RLU), the number of samples identified as positive increased and statistically agreed with aerobic mesophilic microorganism counts (P>0.05). For the dairy industry, the ATP-Bioluminescence technique may become an important tool that assists the official methods to quickly monitor the microbiological quality of UHT milk though this will likely require a threshold below 150 RLU.

Staphylococcus aureus mastitis remains a major challenge for dairy farming. Here, 24 mice were immunized and divided into four groups: G1: control; G2: Granulocyte Macrophage Colony-Stimulating Factor (GM-CSF) DNA vaccine; G3: F0F1 ATP synthase subunit α (SAS), succinyl-diaminopimelate (SDD), and cysteinyl-tRNA synthetase (CTS) recombinant proteins; and G4: SAS+SDD+CTS plus GM-CSF DNA vaccine. The lymphocyte subpopulations, and the intracellular interleukin-17A (IL-17A) and interferon-γ production in the draining lymph node cells were immunophenotyped by flow cytometry. The immunophenotyping and lymphocyte proliferation was determined in spleen cells cultured with and without S. aureus stimulus. Immunization with S. aureus recombinant proteins generated memory cells in draining lymph nodes. Immunization with the three recombinant proteins plus GM-CSF DNA led to an increase in the percentage of IL-17A⁺ cells among overall CD44⁺ (memory), T CD4⁺, CD4⁺ T CD44⁺ CD27⁻, γδ TCR, γδ TCR⁺ CD44⁺ CD27⁺, and TCRVγ4⁺ cells. Vaccination with S. aureus recombinant proteins associated with GM-CSF DNA vaccine downregulated T_H2 immunity. Immunization with the three recombinant proteins plus the GM-CSF DNA led to a proliferation of overall memory T, CD4⁺, and CD4⁺ TEM cells upon S. aureus stimulus. This approach fostered type 3 immunity, suggesting the development of a protective immune response against S. aureus.