Pooled Plasmid Sequencing Reveals the Relationship Between Mobile Genetic Elements and Antimicrobial Resistance Genes in Clinically Isolated Klebsiella pneumoniae |
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Authors: | Yan Jiang Yanfei Wang Xiaoting Hua Yue Qu Anton Y Peleg Yunsong Yu |
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Institution: | Department of Infectious Diseases,Sir Run Run Shaw Hospital,Zhejiang University School of Medicine,Hangzhou 310016,China;Key Laboratory of Microbial Technology and Bioinformatics of Zhejiang Province,Hangzhou 310016,China;Biomedicine Discovery Institute,Department of Microbiology,Faculty of Medicine,Nursing and Health Sciences,Monash University,Melbourne 3800,Australia;Department of Infectious Diseases,The Alfred Hospital and Central Clinical School,Monash University,Melbourne 3004,Australia |
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Abstract: | Plasmids remain important microbial components mediating the horizontal gene transfer (HGT) and dissemination of antimicrobial resistance. To systematically explore the relationship between mobile genetic elements (MGEs) and antimicrobial resistance genes (ARGs), a novel strategy using single-molecule real-time (SMRT) sequencing was developed. This approach was applied to pooled conjugative plasmids from clinically isolated multidrug-resistant (MDR) Klebsiella pneumoniae from a tertiary referral hospital over a 9-month period. The conjugative plasmid pool was obtained from transconjugants that acquired antimicrobial resistance after plasmid conjugation with 53 clinical isolates. The plasmid pool was then subjected to SMRT sequencing, and 82 assembled plasmid fragments were obtained. In total, 124 ARGs (responsible for resistance to β-lactam, fluoroquinolone, and aminoglycoside, among others) and 317 MGEs including transposons (Tns), insertion sequences (ISs), and integrons] were derived from these fragments. Most of these ARGs were linked to MGEs, allowing for the establishment of a relationship network between MGEs and/or ARGs that can be used to describe the dissemination of resistance by mobile elements. Key elements involved in resistance transposition were identified, including IS26, Tn3, IS903B, ISEcp1, and ISKpn19. As the most predominant IS in the network, a typical IS26-mediated multicopy composite transposition event was illustrated by tracing its flanking 8-bp target site duplications (TSDs). The landscape of the pooled plasmid sequences highlights the diversity and complexity of the relationship between MGEs and ARGs, underpinning the clinical value of dominant HGT profiles. |
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Keywords: | Horizontal transfer profile Transconjugant Single-molecule real time sequencing Insertion sequence Plasmid |
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