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      Whole genome sequencing and characteristics of Escherichia coli with co-existence of ESBL and mcr genes from pigs

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          Abstract

          This study aimed to analyze three ESBL-producing E. coli co-harboring mcr and ESBL genes from a healthy fattening pig (E. 431) and two sick pigs (ECP.81 and ECP.82) in Thailand using Whole Genome Sequencing (WGS) using either Illumina MiSeq or HiSeq PE150 platforms to determine their genome and transmissible plasmids. E. 431 carrying mcr-2. 1 and mcr-3. 1 belonged to serotype O142:H31 with ST29 sequence type. ECP.81 and ECP.82 from sick pigs harboring mcr-1. 1 and mcr-3. 1 were serotype O9:H9 with ST10. Two mcr-1. 1 gene cassettes from ECP.81 and ECP.82 were located on IncI2 plasmid with 98% identity to plasmid pHNSHP45. The mcr-2. 1-carrying contig in E. 431 showed 100% identity to plasmid pKP37-BE with the upstream flanking sequence of IS 1595. All three mcr-3. 1-carrying contigs contained the ΔTn As2- mcr-3. 1- dgkA core segment and had high nucleotide similarity (85–100%) to mcr-3. 1-carrying plasmid, pWJ1. The mobile elements i.e. IS 4321, ΔTn As2, IS Kpn40 and IS 3 were identified in the flanking regions of mcr-3. Several genes conferring resistance to aminoglycosides ( aac(3)-IIa, aadA1, aadA2b, aph(3’’)-Ib, aph(3’)-IIa and aph(6)-Id), macrolides ( mdf( A)), phenicols ( cmlA1), sulphonamide ( sul3) and tetracycline ( tet(A) and tet(M)) were located on plasmids, of which their presence was well corresponded to the host’s resistance phenotype. Amino acid substitutions S83L and D87G in GyrA and S80I and E62K in ParC were observed. The bla CTX-M-14 and bla CTX-M-55 genes were identified among these isolates additionally harbored bla TEM-1B. Co-transfer of mcr-1. 1/ bla TEM-1B and mcr-3. 1/ bla CTX-M-55 was observed in ECP.81 and ECP.82 but not located on the same plasmid. The results highlighted that application of advanced innovation technology of WGS in AMR monitoring and surveillance provide comprehensive information of AMR genotype that could yield invaluable benefits to development of control and prevention strategic actions plan for AMR.

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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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            Ror2 signaling regulates Golgi structure and transport through IFT20 for tumor invasiveness

            Signaling through the Ror2 receptor tyrosine kinase promotes invadopodia formation for tumor invasion. Here, we identify intraflagellar transport 20 (IFT20) as a new target of this signaling in tumors that lack primary cilia, and find that IFT20 mediates the ability of Ror2 signaling to induce the invasiveness of these tumors. We also find that IFT20 regulates the nucleation of Golgi-derived microtubules by affecting the GM130-AKAP450 complex, which promotes Golgi ribbon formation in achieving polarized secretion for cell migration and invasion. Furthermore, IFT20 promotes the efficiency of transport through the Golgi complex. These findings shed new insights into how Ror2 signaling promotes tumor invasiveness, and also advance the understanding of how Golgi structure and transport can be regulated.
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              In silico detection and typing of plasmids using PlasmidFinder and plasmid multilocus sequence typing.

              In the work presented here, we designed and developed two easy-to-use Web tools for in silico detection and characterization of whole-genome sequence (WGS) and whole-plasmid sequence data from members of the family Enterobacteriaceae. These tools will facilitate bacterial typing based on draft genomes of multidrug-resistant Enterobacteriaceae species by the rapid detection of known plasmid types. Replicon sequences from 559 fully sequenced plasmids associated with the family Enterobacteriaceae in the NCBI nucleotide database were collected to build a consensus database for integration into a Web tool called PlasmidFinder that can be used for replicon sequence analysis of raw, contig group, or completely assembled and closed plasmid sequencing data. The PlasmidFinder database currently consists of 116 replicon sequences that match with at least at 80% nucleotide identity all replicon sequences identified in the 559 fully sequenced plasmids. For plasmid multilocus sequence typing (pMLST) analysis, a database that is updated weekly was generated from www.pubmlst.org and integrated into a Web tool called pMLST. Both databases were evaluated using draft genomes from a collection of Salmonella enterica serovar Typhimurium isolates. PlasmidFinder identified a total of 103 replicons and between zero and five different plasmid replicons within each of 49 S. Typhimurium draft genomes tested. The pMLST Web tool was able to subtype genomic sequencing data of plasmids, revealing both known plasmid sequence types (STs) and new alleles and ST variants. In conclusion, testing of the two Web tools using both fully assembled plasmid sequences and WGS-generated draft genomes showed them to be able to detect a broad variety of plasmids that are often associated with antimicrobial resistance in clinically relevant bacterial pathogens. Copyright © 2014, American Society for Microbiology. All Rights Reserved.
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                Author and article information

                Contributors
                Role: Data curationRole: Formal analysisRole: InvestigationRole: VisualizationRole: Writing – original draftRole: Writing – review & editing
                Role: ConceptualizationRole: Funding acquisitionRole: MethodologyRole: ResourcesRole: SupervisionRole: ValidationRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS One
                PLoS One
                plos
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                16 November 2021
                2021
                : 16
                : 11
                : e0260011
                Affiliations
                [001] Research Unit in Microbial Food Safety and Antimicrobial Resistance, Department of Veterinary Public Health, Faculty of Veterinary Science, Chulalongkorn University, Bangkok, Thailand
                Zhejiang University, CHINA
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Author information
                https://orcid.org/0000-0002-9714-9199
                Article
                PONE-D-21-23097
                10.1371/journal.pone.0260011
                8594834
                34784400
                7bb667df-3d6d-46df-b648-d570b6cd4627
                © 2021 Trongjit, Chuanchuen

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 16 July 2021
                : 30 October 2021
                Page count
                Figures: 4, Tables: 3, Pages: 17
                Funding
                Funded by: funder-id http://dx.doi.org/10.13039/501100004396, thailand research fund;
                Award ID: BRG6080014
                Award Recipient :
                Funded by: the 90th anniversary of chulalongkorn university fund (ratchadaphisek somphot endowment fund)
                Award Recipient :
                Funded by: chulalongkorn university one healthy research cluster
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/501100012309, royal golden jubilee (rgj) ph.d. programme;
                Award ID: PHD/ 0158/2556
                Award Recipient :
                Funded by: the overseas academic presentation scholarship for graduate student from chulalongkorn university
                Award Recipient :
                This work was supported by TRF Basic Research Grant BRG6080014 co-funded by Thailand Research Fund (TRF), the Faculty of Veterinary Science and Chulalongkorn University. It was also partially supported by the 90th Anniversary of Chulalongkorn University Fund (Ratchadaphisek Somphot Endowment Fund) and Chulalongkorn University One Healthy Research Cluster. ST is a recipient of the Royal Golden Jubilee Ph.D. Program (PHD/ 0158/2556), co-funded by TRF and Chulalongkorn University. We also thank the Overseas Academic Presentation Scholarship for Graduate Student from Chulalongkorn University to Suthathip Trongjit. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
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