Background: Staphylococcus aureus is one of the main causative agents of mastitis in small ruminants. Antimicrobial use is the major treatment, but there are many flaws linked to resistance, tolerance or persistence. This study aimed to verify changes in resistance, virulence and clonal profiles of S. aureus isolated from persistent mastitis goat milk before and after enrofloxacin treatment. Results: MIC increased to at least one antimicrobial in S. aureus isolates after enrofloxacin treatment compared to before. The most detected resistance genes before and after treatment were tetK, tetM, and blaZ, with more resistance genes detected after enrofloxacin treatment (p < 0.05). Occasional variations in efflux system gene detection were observed before and after treatment. Nine virulence genes (hla, fnbA, fnbB, eta, etb, sea, sec, seh, and sej) were detected at both times, and between these, the hla and eta genes were detected more in isolates after treatment. All isolates of S. aureus belonged to the same sequence type (ST) 133, except for two S. aureus isolates prior to enrofloxacin treatment which were classified as ST5 and the other as a new one, ST4966. Isolates of S. aureus 4, 8, and 100 from before and after treatment had identical pulse types, while others obtained from other animals before and after treatment were classified into distinct pulse types. Conclusion: There were occasional changes in the studied profiles of S. aureus isolated before and after treatment of animals with enrofloxacin, which may have contributed to the permanence of bacteria in the mammary gland, even when using traditional treatment, resulting in persistent mastitis.

Mastitis is an inflammation of the mammary gland that causes major losses in the dairy industry. Streptococcus spp. are among the main agents of this disease. Increased resistance to antibiotics is one of the causes of therapeutic failure. Plants, due to their broad chemodiversity, are an interesting source of new molecules with antibacterial activity. Using these compounds along with traditional antibiotics is a possible method for reversing resistance. The objective of this work was to determine the interactions between the activities of guttiferone-A and 7-epiclusianone, two active substances isolated from the fruits of Garcinia brasiliensis, and traditional antibiotics against Streptococcus spp. isolated from bovine mastitis and known to be resistant to them. First, the MIC for the antibiotics and bioactive compounds was determined, followed by their activities, alone and in combination. Then, their cytotoxicity was measured in bovine mammary epithelial cells. Finally, molecular docking simulations were performed to elucidate molecular details of the interactions between β-lactamase and the compounds binding to it (clavulanic acid, ampicillin, 7-epiclusianone, and guttiferone-A). The bacterial isolates were resistant to ampicillin and gentamicin. Both antibiotics showed predominantly synergistic antibacterial activities in combination with guttiferone-A or 7-epiclusianone. These two active substances were not cytotoxic at synergistic concentrations and both showed strong binding to β-lactamase, which may explain the reversal of ampicillin resistance. These substances are promising for the treatment of bovine mastitis.

Mastitis is an inflammation of mammary gland parenchyma that adversely affects bovine health and dairy production worldwide despite significant efforts to eradicate it. The aim of this work was to characterize the antimicrobial activity of 7-epiclusianone (7-epi), a compound extracted from the Rheedia brasiliensis fruit, its complex with copper against Streptococcus spp. isolated from bovine mastitis, and to assess their cytotoxicity to bovine mammary alveolar cells (MAC-T). The complex 7-epiclusianone-Cu (7-epi-Cu) was an amorphous green solid with optical activity. Its vibrational spectrum in the infrared region showed absorption bands in the high-frequency region, as well as bands that can be attributed to the unconjugated and conjugated stretching of the free ligand. The complex was anhydrous. One of the tested bacterial strains was not sensitive to the compounds, while the other three had MIC values of 7.8 µg mL−1 and minimum bactericidal concentration (MBC) values between 15.6 and 31.3 µg mL−1. These two compounds are bacteriostatic, did not cause damage to the cell wall and, at sub-inhibitory concentrations, did not induce bacterial adhesion. The compounds were not cytotoxic. Based on these results, 7-epi and 7-epi-Cu exhibited desirable antimicrobial properties and could potentially be used in bovine mastitis treatment.

This study aimed to investigate the antimicrobial, antibiofilm, and cytotoxic effects of biosurfactant lipopeptides synthesized by Bacillus subtilis TR47II. For this purpose, the lipopeptides were partially purified using a three-step process and characterized. In the first step, the crude extract obtained from acid precipitation exhibited strong antibacterial activity against the Gram-negative opportunistic pathogens Alcaligenes faecalis ATCC 8750, Achromobacter xylosoxidans ATCC 13138, Pseudomonas alcaligenes ATCC 14909, and Pseudomonas putida ATCC 15175. Moreover, partial inhibition was observed against Klebsiella aerogenes ATCC 13048 (42%), Escherichia coli ATCC 25922 (16%), and Pseudomonas aeruginosa ATCC 27853 (47%). The lipopeptides in the crude extract were extracted with methanol and fractioned on a silica gel chromatography column, rendering four TLC-pooled chromatographic fractions, named F1, F2, F3, and F4. The chromatographic fraction F4 was the most bioactive, with MIC values between 300 and 600 µg mL −1. Besides, F4 at sub-MIC doses dislodged the biofilms of A. faecalis, A. xylosoxidans, and P. alcaligenes by about 100, 85, and 81%, respectively. No cytotoxic effect was observed in mammalian cells at MIC. MALDI-TOF-MS analysis revealed that F4 contained cyclic lipopeptides belonging to two families: iturins (m/z 1004 to 1087) and fengycins (m/z 1424 to 1545). The dual effect of F4 on planktonic and sessile growth could suggest that the synergistic application of these biosurfactants could be efficient in the control of these opportunistic pathogens.

Goat farming is a low-cost alternative to dairy production in developing countries. In Brazil, goat production has increased in recent years due in part to the implementation of programs encouraging this activity. Mycobacterium avium ssp. paratuberculosis (MAP) is the causative agent of paratuberculosis, a disease that causes chronic granulomatous enteritis in ruminants, but MAP transmission dynamics are still poorly understood in goats. In a previously published study of our research group, 10 dairy goat farms (467 animals) from Minas Gerais state were analyzed for MAP detection; 2 fecal cultures and 11 milk samples tested positive for MAP by conventional PCR and were confirmed by sequencing. Because no clinical signs were observed over 1 yr of monitoring, we hypothesized that these MAP-positive goats could be passive shedders. Thus, in the present study, 4 positive goats (4/13) from the previous study were purchased and feces and milk samples were collected for evaluation (twice, with an interval of 3 mo between tests) by culture of MAP, IS900 PCR, or both. All analyses were negative for MAP. At the last time point, blood samples were collected for ELISA, the animals were killed, and tissues collected for tissue culture and histopathology. At necropsy, no macroscopic lesions related to paratuberculosis were observed. Similarly, no histological changes were observed and MAP in samples stained by Ziehl-Neelsen was not detected. These animals were characterized as potential passive shedders with upward contamination of the teat canal by MAP. This is the first report of the passive shedding phenomenon in goats in Brazil and it highlights the importance of identifying these animals for control programs and to ensure the quality of dairy products.