Abstract

Cefquinome (CEQ) is a novel β-lactam antibiotic that exhibits excellent antibacterial activity against Staphylococcus aureus. However, the bacterial protein targets of CEQ are unclear. To evaluate the relationship between the pharmacokinetic/ pharmacodynamic (PK/PD) parameters of CEQ and strains with varying degrees of resistance and to elucidate bacterial protein responses to CEQ treatment, label-free quantitative proteomics analysis was conducted. The sensitive S. aureus ATCC6538 and the resistant 2MIC and 8MIC were tested for differentially expressed proteins. An in vitro model was treated with different concentrations of CEQ (3, 5, or 10 μg/ml) with different terminal half-lives (2.5 or 5 h) at different intervals (12 or 24 h). Differentially expressed proteins were evaluated using Gene Ontology analysis followed by KEGG pathway enrichment analysis and STRING network analysis. RT-qPCR was performed to validate the differentially expressed proteins at the molecular level. The results showed that the degree of resistance increased in a cumulative manner and increased gradually with the extension of administration time. The resistant strain would not have appeared in the model only if %T > mutant prevention concentration ≥ 50%. The expression of 45 proteins significantly changed following CEQ treatment, among which 42 proteins were obviously upregulated and 3 were downregulated. GO analysis revealed that the differentially expressed proteins were mainly present on cells and the cell membrane, participated in metabolic and intracellular processes, and had catalytic and binding activities. The RPSO, SDHB, CITZ, ADK, and SAOUHSC 00113 genes in S. aureus may play important roles in the development of resistance to CEQ. These results provided important reference candidate proteins as targets for overcoming S. aureus resistance to CEQ.

• Keywords: cefquinome, quantitative proteomic, Staphylococcus aureus, antibacterial activity, PK/PD, in vitro model