Enterococcus spp., including E. faecalis and E. faecium, pose risks to dairy farms as opportunistic pathogens. The study evaluates antimicrobial resistance (AMR) and virulence characteristics of Enterococcus spp. isolated from bovine milk. Bile esculin agar was used to assess 1471 milk samples, followed by colony identification, gram staining, catalase tests, and 45 °C incubation. PCR analysis targeted E. faecalis and E. faecium in characteristic Enterococcus spp. colonies, with MALDI-TOF used for negative samples. Multiple tests, including disk diffusion, chromogenic VRE agar for vancomycin resistance, Vancomycin Etest^® for MIC determination, and PCR for virulence factors (cylA, esp, efaA, ace, asa1, gelE, and hyl genes), were performed. Out of 100 identified strains, E. durans (30.66%), E. faecium (26.28%), and E. faecalis (18.25%) were predominant. AMR in Enterococcus spp. varied, with the highest rates against rifampicin (27%), tetracycline (20%), and erythromycin (18%). Linezolid (5%), vancomycin, ciprofloxacin, and teicoplanin (3% each) had lower prevalence. E. faecium and E. faecalis showed high AMR to rifampicin, erythromycin, and tetracycline. Thirty-two strains (18.98%) grew on VRE Chromoselect agar, while 4 (2 E. faecalis and 2 E. faecium) showed vancomycin resistance by MIC values. E. faecalis carried gelE (45.5%) and asa1 (36%), and E. gallinarum had 9.1% with the asa1 gene. Detecting resistant Enterococcus in bovine milk supports control strategies for enterococci on dairy farms, highlighting AMR concerns in the food chain.

Staphylococcus aureus is a major cause of subclinical mastitis in dairy cows, and its development of antibiotic resistance has limited treatment efficacy. Essential oils (EOs) are natural products with a wide range of antimicrobial properties that could be used to treat bovine mastitis. This study aims to investigate the antimicrobial activity of EOs against S. aureus isolated from subclinical bovine mastitis cases in the State of São Paulo—Brazil. A total of 14 S. aureus isolates were selected, based on the presence of biofilm-forming genes (icaA, icaD, and bap), and were cultured to a final concentration of 10³ CFU.mL⁻¹ for the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) analysis of five EOs (Citrus aurantium bergamia—bergamot, Copaifera reticulata—copaiba, Foeniculum vulgare—fennel, Zingiber officinale—ginger, and Ocimum basilicum—basil). The chemical compositions of the EOs were characterized using gas chromatography coupled with a mass-selective detector (GC/MSD). Basil and bergamot EOs exhibited the highest antimicrobial activity against S. aureus strains, with mean MIC/MBC values of 1.561 ± 0.223/2.806 ± 0.255 mg.mL⁻¹ and 2.782 ± 0.228/4.396 ± 0.198 mg.mL⁻¹, respectively. The primary compounds in basil EO were methyl-chavicol, linalool, and α-humulene, while bergamot EO predominantly contained linalyl acetate, limonene, and linalool. This research highlights the potential of basil and bergamot EOs as natural antimicrobial agents for treating bovine mastitis caused by S. aureus, offering a potential alternative to traditional antibiotics and contributing to animal welfare and public health. In addition, it emphasizes the need for further studies to validate the long-term effects, optimal dosages, and application methods.

Staphylococcus aureus is one of the most common pathogens associated with mastitis in dairy herds worldwide. This study evaluated the profile of virulence and antimicrobial resistance genes of spa type t605 methicillin-susceptible Staphylococcus aureus isolated from subclinical bovine mastitis in São Paulo, Brazil. Fifty-seven S. aureus strains were screened by conventional PCR for 49 virulence genes. The most prevalent virulence genes detected were icaD (94.7%), fib (93%), fnbA (82.5%), clfA (80.7%), bap (78.9%), clfB (73.7%), icaA (66.7%), see (64.9%), and sed (61.4%). The blaZ (94.7%), aac6’aph2’ (15.8%), and ant4 (12.3%) genes were the most common antimicrobial resistance genes; however, mecA and mecC genes were not found. All methicillin-susceptible S. aureus (MSSA) strains were characterised through spa and agr typing. The spa type t605 was found in all isolates. By agr typing, the most prevalent were type II (56.1%). Antimicrobial resistance was determined by the disk diffusion method, and 93% showed resistance to at least one antibiotic. Penicillin resistance was the most prevalent (87.7%), followed by tetracycline (12.3%), oxacillin (10.5%), and gentamicin (10.5%) resistance. Our study confirmed the spa type t605 as endemic, carrying a wide variety of virulence factors and high-level penicillin resistance. The profile seems to be associated with the colonisation of MSSA and its persistence in subclinical mastitis.