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Objectives: To characterize the complete sequences of four plasmids in MCR-1-producing clinical Escherichia coli strain D72, and to depict the formation mechanism and characteristics of the cointegrate plasmid derived from the pD72-mcr1 and pD72-F33 plasmids., Methods: The genetic profiles of plasmids in strain D72 and its transconjugant were determined by conjugation, S1-PFGE, Southern hybridization, WGS analysis and PCR. Plasmid sequences were analysed with bioinformatic tools. The traits of the fusion plasmid were characterized by cointegration, stability and conjugation assays., Results: Strain D72, belonging to ST1114, contained four plasmids, including mcr-1-carrying pD72-mcr1, blaCTX-M-55-carrying pD72-F33, blaTEM-238-bearing pD72-IncP and pD72-IncX1 carrying aph(3')-Ia, qnrS2 and floR. A single plasmid, pD72C, in the transconjugant was found to be larger than any plasmid in the original strain D72. Sequence analysis showed that pD72C was the fusion product of pD72-mcr1 and pD72-F33, and the recombinant event involved an intermolecular replicative mechanism. Plasmid fusion occurred at a frequency of 1.75 × 10-4 cointegrates per transconjugant. The fusion plasmid presented a high stability and conjugation frequency of 8.00 × 10-3., Conclusions: To our knowledge, this is the first report of the IS26-mediated fusion of an IncN1-F33:A-:B- plasmid and an mcr-1-carrying phage-like plasmid, providing evidence for the important role of IS26 in the recombination of plasmids. The biological advantages of the fusion plasmid indicated that the fusion event presumably plays a potential role in the dissemination of mcr-1., (© The Author(s) 2019. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please email: journals.permissions@oup.com.)