On-farm molecular epidemiology and therapeutic insights into multidrug-resistant Staphylococcus aureus isolated from bubaline mastitis

Staphylococcus aureus (S. aureus) is a pathogen of veterinary and public health concern; majorly responsible for bubaline mastitis and huge farm economic losses. The current study, aimed to investigate the on-farm epidemiology, associated antibiotic resistance and possible resistance modulation strategies against S. aureus isolated from bubaline mastitis. S. aureus methicillin and vancomycin-resistant (MRSA and VRSA) from buffalo (n = 384) herds of district Bahawalpur and Rahim yar khan, Pakistan were identified based on bacteriological, biochemical, and molecular confirmation of nuc, mecA, and vanb genes, respectively. The antibiotic susceptibility profiling of S. aureus isolates was done by Kirby-Bauer’s disc diffusion method. The resistant antibiotics were combined with non- antibiotics (NSAIDs and ivermectin) using well diffusion method and checkerboard assay to evaluate resistance modulation. The risk factor analysis was done by chi-square and logistic regression analysis to find out the potential risk factors associated with mastitis in buffalo herds. The results found 50.26% and 38.02% samples positive for subclinical mastitis and S. aureus, respectively. Among local isolates, 62.5% and 20.83% isolates were confirmed MRSA and VRSA, respectively. The final logistic regression analysis revealed that host-related risk factors like (udder depth, teat pathology) and farm management risk factors (udder hygiene during milking, hock score, post-milking disinfection, and dry cow therapy) were significantly associated with the occurrence of sub-clinical mastitis in buffaloes. Antibiotic resistance profiling of local isolates showed significantly (p<0.05) higher resistance to gentamicin, oxytetracycline, and cotrimoxazole. The antimicrobial effects of non-antibiotics against study isolates showed higher zones of inhibition for meloxicam followed by flunixin, ketoprofen, and ivermectin. The combinations of resistant antibiotics with non-antibiotics showed that the combinations of amoxicillin/ meloxicam, cotrimoxazole/flunixin, cotrimoxazole/ketoprofen, and gentamicin/flunixin on well diffusion method and cotrimoxazole/flunixin, amoxicillin/ketoprofen, and gentamicin/flunixin on checkerboard assay revealed synergistic interactions. The study concluded that S. aureus, MRSA, and VRSA is anemerging and prevailing causes of bubaline mastitis in dairy farms in Pakistan. The increasing antibiotic resistance in S. aureus can be modulated by combining the resistant antibiotics with NSAIDs, especially flunixin and ketoprofen.