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      Evidence of structural rearrangements in ESBL-positive pESI(like) megaplasmids of S.Infantis

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          Abstract

          The increasing prevalence of pESI(like)-positive, multidrug-resistant (MDR) S. Infantis in Europe is a cause of major concern. As previously demonstrated, the pESI(like) megaplasmid is not only a carrier of antimicrobial resistant (AMR) genes (at least tet, dfr, and sul genes), but also harbours several virulence and fitness genes, and toxin/antitoxin systems that enhance its persistence in the S. Infantis host. In this study, five prototype pESI(like) plasmids, of either CTX-M-1 or CTX-M-65 ESBL-producing strains, were long-read sequenced using Oxford Nanopore Technology (ONT), and their complete sequences were resolved. Comparison of the structure and gene content of the five sequenced plasmids, and further comparison with previously published pESI(like) sequences, indicated that although the sequence of such pESI(like) ‘mosaic’ plasmids remains almost identical, their structures appear different and composed of regions inserted or transposed after different events. The results obtained in this study are essential to better understand the plasticity and the evolution of the pESI(like) megaplasmid, and therefore to better address risk management options and policy decisions to fight against AMR and MDR in Salmonella and other food-borne pathogens.

          Graphical representation of the pESI-like plasmid complete sequence (ID 12037823/11). Block colours indicate the function of the genes: red: repB gene; pink: class I integrons (IntI); yellow; mobile elements; blue: resistance genes; green: toxin/anti-toxin systems; grey: mer operon; light green: genes involve in conjugation.

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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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            Unicycler: Resolving bacterial genome assemblies from short and long sequencing reads

            The Illumina DNA sequencing platform generates accurate but short reads, which can be used to produce accurate but fragmented genome assemblies. Pacific Biosciences and Oxford Nanopore Technologies DNA sequencing platforms generate long reads that can produce complete genome assemblies, but the sequencing is more expensive and error-prone. There is significant interest in combining data from these complementary sequencing technologies to generate more accurate “hybrid” assemblies. However, few tools exist that truly leverage the benefits of both types of data, namely the accuracy of short reads and the structural resolving power of long reads. Here we present Unicycler, a new tool for assembling bacterial genomes from a combination of short and long reads, which produces assemblies that are accurate, complete and cost-effective. Unicycler builds an initial assembly graph from short reads using the de novo assembler SPAdes and then simplifies the graph using information from short and long reads. Unicycler uses a novel semi-global aligner to align long reads to the assembly graph. Tests on both synthetic and real reads show Unicycler can assemble larger contigs with fewer misassemblies than other hybrid assemblers, even when long-read depth and accuracy are low. Unicycler is open source (GPLv3) and available at github.com/rrwick/Unicycler.
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              The RAST Server: Rapid Annotations using Subsystems Technology

              Background The number of prokaryotic genome sequences becoming available is growing steadily and is growing faster than our ability to accurately annotate them. Description We describe a fully automated service for annotating bacterial and archaeal genomes. The service identifies protein-encoding, rRNA and tRNA genes, assigns functions to the genes, predicts which subsystems are represented in the genome, uses this information to reconstruct the metabolic network and makes the output easily downloadable for the user. In addition, the annotated genome can be browsed in an environment that supports comparative analysis with the annotated genomes maintained in the SEED environment. The service normally makes the annotated genome available within 12–24 hours of submission, but ultimately the quality of such a service will be judged in terms of accuracy, consistency, and completeness of the produced annotations. We summarize our attempts to address these issues and discuss plans for incrementally enhancing the service. Conclusion By providing accurate, rapid annotation freely to the community we have created an important community resource. The service has now been utilized by over 120 external users annotating over 350 distinct genomes.
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                Author and article information

                Contributors
                Journal
                FEMS Microbiol Lett
                FEMS Microbiol Lett
                femsle
                FEMS Microbiology Letters
                Oxford University Press
                0378-1097
                1574-6968
                2023
                20 February 2023
                20 February 2023
                : 370
                : fnad014
                Affiliations
                National Reference Laboratory for Antimicrobial Resistance, Istituto Zooprofilattico Sperimentale del Lazio e della Toscana “M. Aleandri,” General Diagnostics Department , Rome 00178, Italy
                National Reference Laboratory for Antimicrobial Resistance, Istituto Zooprofilattico Sperimentale del Lazio e della Toscana “M. Aleandri,” General Diagnostics Department , Rome 00178, Italy
                National Reference Laboratory for Antimicrobial Resistance, Istituto Zooprofilattico Sperimentale del Lazio e della Toscana “M. Aleandri,” General Diagnostics Department , Rome 00178, Italy
                National Reference Laboratory for Antimicrobial Resistance, Istituto Zooprofilattico Sperimentale del Lazio e della Toscana “M. Aleandri,” General Diagnostics Department , Rome 00178, Italy
                National Reference Laboratory for Antimicrobial Resistance, Istituto Zooprofilattico Sperimentale del Lazio e della Toscana “M. Aleandri,” General Diagnostics Department , Rome 00178, Italy
                National Reference Laboratory for Antimicrobial Resistance, Istituto Zooprofilattico Sperimentale del Lazio e della Toscana “M. Aleandri,” General Diagnostics Department , Rome 00178, Italy
                National Reference Laboratory for Antimicrobial Resistance, Istituto Zooprofilattico Sperimentale del Lazio e della Toscana “M. Aleandri,” General Diagnostics Department , Rome 00178, Italy
                National Reference Laboratory for Antimicrobial Resistance, Istituto Zooprofilattico Sperimentale del Lazio e della Toscana “M. Aleandri,” General Diagnostics Department , Rome 00178, Italy
                National Reference Laboratory for Antimicrobial Resistance, Istituto Zooprofilattico Sperimentale del Lazio e della Toscana “M. Aleandri,” General Diagnostics Department , Rome 00178, Italy
                National Reference Laboratory for Antimicrobial Resistance, Istituto Zooprofilattico Sperimentale del Lazio e della Toscana “M. Aleandri,” General Diagnostics Department , Rome 00178, Italy
                National Reference Laboratory for Antimicrobial Resistance, Istituto Zooprofilattico Sperimentale del Lazio e della Toscana “M. Aleandri,” General Diagnostics Department , Rome 00178, Italy
                Author notes
                Corresponding author. National Reference Laboratory for Antimicrobial Resistance, Istituto Zooprofilattico Sperimentale del Lazio e della Toscana “M. Aleandri,” General Diagnostics Department, Rome 000178, Italy. E-mail: patricia.alba@ 123456izslt.it
                Author information
                https://orcid.org/0000-0002-9621-0955
                https://orcid.org/0000-0002-3835-4723
                https://orcid.org/0000-0003-1308-199X
                Article
                fnad014
                10.1093/femsle/fnad014
                9990980
                36806934
                f091f9f0-2aaf-48dc-917e-9faecf2b8485
                © The Author(s) 2023. Published by Oxford University Press on behalf of FEMS.

                This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License ( https://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@ 123456oup.com

                History
                : 29 July 2022
                : 19 January 2023
                : 17 February 2023
                : 06 March 2023
                Page count
                Pages: 8
                Funding
                Funded by: European Union, DOI 10.13039/501100000780;
                Funded by: Horizon 2020, DOI 10.13039/100010661;
                Award ID: 773830
                Categories
                Research Letter
                Clinical & Clinical Veterinary Microbiology
                AcademicSubjects/SCI01150

                Microbiology & Virology
                salmonella infantis,megaplasmid,long read sequencing,rearrangements,antibiotic resistance,esbl-producing salmonella

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