Inter-plasmid transfer of antibiotic resistance genes accelerates antibiotic resistance in bacterial pathogens

Antimicrobial resistance is a major threat for public health. Plasmids play a critical role in the spread of antimicrobial resistance via horizontal gene transfer between bacterial species. However, it remains unclear how plasmids originally recruit and assemble various antibiotic resistance genes (ARGs). Here, we track ARG recruitment and assembly in clinically relevant plasmids by combining a systematic analysis of 2420 complete plasmid genomes and experimental validation. Results showed that ARG transfer across plasmids is prevalent, and 87% ARGs were observed to potentially transfer among various plasmids among 8229 plasmid-borne ARGs. Interestingly, recruitment and assembly of ARGs occur mostly among compatible plasmids within the same bacterial cell, with over 88% of ARG transfers occurring between compatible plasmids. Integron and insertion sequences drive the ongoing ARG acquisition by plasmids, especially in which IS26 facilitates 63.1% of ARG transfer events among plasmids. In vitro experiment validated the important role of IS26 involved in transferring gentamicin resistance gene aacC1 between compatible plasmids. Network analysis showed four beta-lactam genes (blaTEM-1, blaNDM-4, blaKPC-2, and blaSHV-1) shuffling among 1029 plasmids and 45 clinical pathogens, suggesting that clinically alarming ARGs transferred accelerate the propagation of antibiotic resistance in clinical pathogens. ARGs in plasmids are also able to transmit across clinical and environmental boundaries, in terms of the high-sequence similarities of plasmid-borne ARGs between clinical and environmental plasmids. This study demonstrated that inter-plasmid ARG transfer is a universal mechanism for plasmid to recruit various ARGs, thus advancing our understanding of the emergence of multidrug-resistant plasmids.