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      Genomic analyses of an Escherichia coli and Klebsiella pneumoniae urinary tract co‐infection using long‐read nanopore sequencing

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          Abstract

          Escherichia coli and Klebsiella pneumoniae isolates presenting with the same antimicrobial susceptibility profile were recovered from the same catheter sample of urine (CSU). Both strains were recovered from a patient with a long‐standing indwelling urinary catheter. Each isolate had its DNA extracted following culture. Nanopore long‐read sequencing was used to build the plasmids and chromosomes from each strain to closure to discern the potential horizontal propagation of resistance‐encoding plasmids and the relationship between resistance genes and insertion sequences. Plasmids derived from resistance strains in the urinary microbiota remain poorly characterized. The same 11 antimicrobial resistance (AMR) genes were found in plasmids from each strain. The 185,239‐bp FIB(K) pKBM1, from the K. pneumoniae strain, additionally encoded the five AMR genes: sul2, strA, strB, bla TEM‐1B, and bla CTX‐M‐15. A multimeric array of AMR genes and IS 26 insertion sequences were found in the plasmids from both isolates. Both plasmids from each isolate were similar. Horizontal transfer of plasmids, followed by subsequent plasmid rearrangement, is likely to have occurred during infection. Furthermore, the resistance region in the plasmids shared similarity against the internationally prevalent plasmid, pKPN3‐307_typeA, commonly identified in K. pneumoniae ST307. Biofilm formation in catheterized patients may allow close cell contact between strains. Horizontal propagation of resistance genes may occur, leading to polymicrobial infections.

          Abstract

          Two bacterial strains were recovered from the same catheter specimen of urine sample. The Escherichia coli strain encoded 11 resistance genes on a plasmid, including the ESBL gene, bla CTX‐M‐15. The Klebsiella pneumoniae strain encoded an additional 5 resistance genes on a plasmid, including 2 copies of bla CTX‐M‐15. Resistance genes were flanked between IS 26 in a multimeric array. Both plasmids were similar, suggesting horizontal transfer occurred during the course of infection.

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          Most cited references31

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          Prokka: rapid prokaryotic genome annotation.

          T Seemann (2014)
          The multiplex capability and high yield of current day DNA-sequencing instruments has made bacterial whole genome sequencing a routine affair. The subsequent de novo assembly of reads into contigs has been well addressed. The final step of annotating all relevant genomic features on those contigs can be achieved slowly using existing web- and email-based systems, but these are not applicable for sensitive data or integrating into computational pipelines. Here we introduce Prokka, a command line software tool to fully annotate a draft bacterial genome in about 10 min on a typical desktop computer. It produces standards-compliant output files for further analysis or viewing in genome browsers. Prokka is implemented in Perl and is freely available under an open source GPLv2 license from http://vicbioinformatics.com/. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.
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            Multidrug-resistant, extensively drug-resistant and pandrug-resistant bacteria: an international expert proposal for interim standard definitions for acquired resistance.

            Many different definitions for multidrug-resistant (MDR), extensively drug-resistant (XDR) and pandrug-resistant (PDR) bacteria are being used in the medical literature to characterize the different patterns of resistance found in healthcare-associated, antimicrobial-resistant bacteria. A group of international experts came together through a joint initiative by the European Centre for Disease Prevention and Control (ECDC) and the Centers for Disease Control and Prevention (CDC), to create a standardized international terminology with which to describe acquired resistance profiles in Staphylococcus aureus, Enterococcus spp., Enterobacteriaceae (other than Salmonella and Shigella), Pseudomonas aeruginosa and Acinetobacter spp., all bacteria often responsible for healthcare-associated infections and prone to multidrug resistance. Epidemiologically significant antimicrobial categories were constructed for each bacterium. Lists of antimicrobial categories proposed for antimicrobial susceptibility testing were created using documents and breakpoints from the Clinical Laboratory Standards Institute (CLSI), the European Committee on Antimicrobial Susceptibility Testing (EUCAST) and the United States Food and Drug Administration (FDA). MDR was defined as acquired non-susceptibility to at least one agent in three or more antimicrobial categories, XDR was defined as non-susceptibility to at least one agent in all but two or fewer antimicrobial categories (i.e. bacterial isolates remain susceptible to only one or two categories) and PDR was defined as non-susceptibility to all agents in all antimicrobial categories. To ensure correct application of these definitions, bacterial isolates should be tested against all or nearly all of the antimicrobial agents within the antimicrobial categories and selective reporting and suppression of results should be avoided. © 2011 European Society of Clinical Microbiology and Infectious Diseases. No claim to original US government works.
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              Assembly of long, error-prone reads using repeat graphs

              Accurate genome assembly is hampered by repetitive regions. Although long single molecule sequencing reads are better able to resolve genomic repeats than short-read data, most long-read assembly algorithms do not provide the repeat characterization necessary for producing optimal assemblies. Here, we present Flye, a long-read assembly algorithm that generates arbitrary paths in an unknown repeat graph, called disjointigs, and constructs an accurate repeat graph from these error-riddled disjointigs. We benchmark Flye against five state-of-the-art assemblers and show that it generates better or comparable assemblies, while being an order of magnitude faster. Flye nearly doubled the contiguity of the human genome assembly (as measured by the NGA50 assembly quality metric) compared with existing assemblers.
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                Author and article information

                Contributors
                sfordham@bournemouth.ac.uk
                Journal
                Microbiologyopen
                Microbiologyopen
                10.1002/(ISSN)2045-8827
                MBO3
                MicrobiologyOpen
                John Wiley and Sons Inc. (Hoboken )
                2045-8827
                17 January 2024
                February 2024
                : 13
                : 1 ( doiID: 10.1002/mbo3.v13.1 )
                : e1396
                Affiliations
                [ 1 ] Department of Life & Environmental Sciences, Talbot Campus Fern Barrow Bournemouth University Poole UK
                [ 2 ] Department of Medical Microbiology, Poole Hospital University Hospitals Dorset NHS Foundation Trust Poole England
                Author notes
                [*] [* ] Correspondence Stephen Mark Edward Fordham, Bournemouth University, Department of Life & Environmental Sciences, Talbot Campus Fern Barrow, Poole BH12 5BB, UK.

                Email: sfordham@ 123456bournemouth.ac.uk

                Author information
                http://orcid.org/0000-0002-7814-2880
                Article
                MBO31396
                10.1002/mbo3.1396
                10794504
                116f1287-3573-4d4e-93fe-3be35f20a30b
                © 2024 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

                This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.

                History
                : 22 December 2023
                : 22 August 2023
                : 08 January 2024
                Page count
                Figures: 7, Tables: 3, Pages: 11, Words: 6481
                Funding
                Funded by: Pfizer , doi 10.13039/100004319;
                Award ID: 68198087
                Categories
                Commentary
                Commentary
                Custom metadata
                2.0
                February 2024
                Converter:WILEY_ML3GV2_TO_JATSPMC version:6.3.6 mode:remove_FC converted:17.01.2024

                Microbiology & Virology
                is26 ,plasmid,resistance,urinary tract infection
                Microbiology & Virology
                is26 , plasmid, resistance, urinary tract infection

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