title: Identification of IncA/C Plasmid Replication and Maintenance Genes and Development of a Plasmid Multilocus Sequence Typing Scheme.
abstract: Plasmids of incompatibility group A/C (IncA/C) are becoming increasingly prevalent within pathogenic Enterobacteriaceae They are associated with the dissemination of multiple clinically relevant resistance genes, including blaCMYand blaNDMCurrent typing methods for IncA/C plasmids offer limited resolution. In this study, we present the complete sequence of a blaNDM-1-positive IncA/C plasmid, pMS6198A, isolated from a multidrug-resistant uropathogenic Escherichia coli strain. Hypersaturated transposon mutagenesis, coupled with transposon-directed insertion site sequencing (TraDIS), was employed to identify conserved genetic elements required for replication and maintenance of pMS6198A. Our analysis of TraDIS data identified roles for the replicon, including repA, a toxin-antitoxin system; two putative partitioning genes, parAB; and a putative gene, 053 Construction of mini-IncA/C plasmids and examination of their stability within E. coli confirmed that the region encompassing 053 contributes to the stable maintenance of IncA/C plasmids. Subsequently, the four major maintenance genes (repA, parAB, and 053) were used to construct a new plasmid multilocus sequence typing (PMLST) scheme for IncA/C plasmids. Application of this scheme to a database of 82 IncA/C plasmids identified 11 unique sequence types (STs), with two dominant STs. The majority of blaNDM-positive plasmids examined (15/17; 88%) fall into ST1, suggesting acquisition and subsequent expansion of this blaNDM-containing plasmid lineage. The IncA/C PMLST scheme represents a standardized tool to identify, track, and analyze the dissemination of important IncA/C plasmid lineages, particularly in the context of epidemiological studies.
authors.affiliation: Australian Infectious Diseases Research Centre, University of Queensland, Brisbane, Australia.|School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Australia.
authors.forename: Steven J
authors.identifier: 0000-0001-7203-2640
authors.initials: SJ
authors.lastname: Hancock
authors.affiliation: Australian Infectious Diseases Research Centre, University of Queensland, Brisbane, Australia m.phan1@uq.edu.au m.schembri@uq.edu.au.|School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Australia.
authors.forename: Minh-Duy
authors.identifier: 0000-0002-3426-1044
authors.initials: MD
authors.lastname: Phan
authors.affiliation: Australian Infectious Diseases Research Centre, University of Queensland, Brisbane, Australia.|School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Australia.
authors.forename: Kate M
authors.initials: KM
authors.lastname: Peters
authors.affiliation: Australian Infectious Diseases Research Centre, University of Queensland, Brisbane, Australia.|School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Australia.
authors.forename: Brian M
authors.initials: BM
authors.lastname: Forde
authors.affiliation: Faculty of Science, Division of Genetics and Molecular Biology, Institute of Biological Sciences, University of Malaya, Kuala Lumpur, Malaysia.
authors.forename: Teik Min
authors.initials: TM
authors.lastname: Chong
authors.affiliation: Faculty of Science, Division of Genetics and Molecular Biology, Institute of Biological Sciences, University of Malaya, Kuala Lumpur, Malaysia.
authors.forename: Wai-Fong
authors.initials: WF
authors.lastname: Yin
authors.affiliation: Faculty of Science, Division of Genetics and Molecular Biology, Institute of Biological Sciences, University of Malaya, Kuala Lumpur, Malaysia.
authors.forename: Kok-Gan
authors.identifier: 0000-0002-1883-1115
authors.initials: KG
authors.lastname: Chan
authors.affiliation: University of Queensland Centre for Clinical Research, Brisbane, Australia.
authors.forename: David L
authors.initials: DL
authors.lastname: Paterson
authors.affiliation: Department of Medical Microbiology and Infectious Disease, Cardiff University, Cardiff, United Kingdom.
authors.forename: Timothy R
authors.initials: TR
authors.lastname: Walsh
authors.affiliation: Australian Infectious Diseases Research Centre, University of Queensland, Brisbane, Australia.|School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Australia.
authors.forename: Scott A
authors.identifier: 0000-0002-1806-3283
authors.initials: SA
authors.lastname: Beatson
authors.affiliation: Australian Infectious Diseases Research Centre, University of Queensland, Brisbane, Australia m.phan1@uq.edu.au m.schembri@uq.edu.au.|School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Australia.
authors.forename: Mark A
authors.identifier: 0000-0003-4863-9260
authors.initials: MA
authors.lastname: Schembri
mesh_terms: D015780:Carbapenems / Q000494:pharmacology; D004261:DNA Replication; D004251:DNA Transposable Elements; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics; D058885:Multilocus Sequence Typing / Q000379:methods*; D010957:Plasmids / Q000235:genetics*; D056689:Uropathogenic Escherichia coli / Q000145:classification / Q000235:genetics*; D001618:beta-Lactamases / Q000235:genetics
chemical_list: D015780:Carbapenems; D004251:DNA Transposable Elements; D029968:Escherichia coli Proteins; D001618:beta-Lactamases; C552864:beta-lactamase NDM-1
keywords: IncA/C plasmid; New Delhi metallo-beta-lactamase; functional genomics; plasmid multilocus sequence typing; uropathogenic E. coli