Genomic signatures of beta-lactam resistance and reversion in Staphylococcus aureus from bovine mastitis

This study uses whole genome sequencing (WGS) to identify beta-lactam resistance-associated mutations in in vitro selected Staphylococcus aureus subsp. aureus ATCC 25923 strains, and correlates the findings with isolates collected from mastitis-infected dairy cows. Resistance was induced in a susceptible strain of S. aureus subsp. aureus ATCC 25923 through serial in vitro exposure to ampicillin sodium salt, cefapirin, cefuroxime sodium salt, and cefquinome. The resulting resistant isolates exhibited thousands of fold increases in minimum inhibitory concentrations (MICs), compared to the susceptible strain. In the absence of antimicrobial selection pressure, the MIC decreased by up to 32-fold, indicating a significant restoration of antimicrobial susceptibility. WGS identified resistance-associated mutations in the penicillin-binding proteins, ABC transporter ATP-binding protein, cell wall-active antibiotics response protein, cyclic-di-AMP phosphodiesterase, and two-component system sensor histidine kinase. Additionally, these mutations were investigated in the S. aureus isolates collected from mastitis-infected dairy cows. These isolates shared the same resistance-associated mutations as in vitro-selected strains. These findings demonstrate that resistance mutations identified through in vitro selection are also present in clinical isolates, highlighting the clinical applicability of in vitro selection for understanding antimicrobial resistance in mastitis-associated S. aureus.