Subinhibitory antibiotic concentrations promote the horizontal transfer of plasmid-borne resistance genes from Klebsiellae pneumoniae to Escherichia coli

Horizontal gene transfer plays an important role in the spread of antibiotic resistance, in which plasmid-mediated conjugation transfer is the most important mechanism. While sub-minimal inhibitory concentrations (sub-MIC) of antibiotics could promote conjugation frequency, the mechanism by which sub-MIC levels of antibiotics affect conjugation frequency is not clear. Here, we used Klebsiella pneumoniae SW1780 carrying the multi-drug resistance plasmid pSW1780-KPC as the donor strain, to investigate the effects of sub-MICs of meropenem (MEM), ciprofloxacin (CIP), cefotaxime (CTX), and amikacin (AK) on conjugational transfer of pSW1780-KPC from SW1780 to Escherichia coli J53. Our results showed that the transfer frequencies increased significantly by treating SW1780 strain with sub-MIC levels of MEM, CIP, CTX and AK. Transfer frequencies at sub-MIC conditions in a Galleria mellonella were significantly higher than in vitro. To investigate gene expression and metabolic effects, RT-qPCR and LC-MS-based metabolome sequencing were performed. Transcript levels of T4SS genes virB1, virB2, virB4, virB8, and conjugation-related genes traB, traK, traE, and traL were significantly upregulated by exposure to sub-MICs of MEM, CIP, CTX, and AK. Metabolome sequencing revealed nine differentially regulated metabolites. Our findings are an early warning for a wide assessment of the roles of sub-MIC levels of antibiotics in the spread of antibiotic resistance.

Automated summary

Horizontal gene transfer, especially plasmid-mediated conjugation transfer, plays a crucial role in the spread of antibiotic resistance. It is found that sub-minimal inhibitory concentrations (sub-MICs) of antibiotics can promote conjugation frequency, although the underlying mechanism remains unclear. In this study, the effects of sub-MICs of meropenem, ciprofloxacin, cefotaxime, and amikacin on conjugational transfer were investigated using Klebsiella pneumoniae as the donor strain. The results showed that sub-MIC levels of antibiotics significantly increased the transfer frequencies. Furthermore, the study also revealed changes in gene expression and metabolome under sub-MIC conditions. These findings highlight the importance of assessing the roles of sub-MIC levels of antibiotics in the spread of antibiotic resistance.