We present single-contig assemblies for Escherichia coli strain KV7 (serotype O27, phylogenetic group D) and its six plasmids, isolated from a healthy pig, as determined by PacBio RS II and Illumina MiSeq sequencing. The chromosome of 4,997,475 bp and G+C content of 50.75% harbored 4,540 protein-encoding genes

Bateman, Michael D

Cavaco, Lina M

de Vries, Stefan PW

Grant, Andrew J

Guardabassi, Luca

Gupta, Srishti

Holmes, Mark A

English

We present single-contig assemblies for Escherichia coli strain KV7 (serotype O27, phylogenetic group D) and its six plasmids, isolated from a healthy pig, as determined by PacBio RS II and Illumina MiSeq sequencing. The chromosome of 4,997,475 bp and G+C content of 50.75% harbored 4,540 protein-encoding genes

Bateman, Michael D.

de Vries, Stefan P.W.

Cavaco, Lina M.

Grant, Andrew J.

Holmes, Mark A.

NARCIS 

Genome and plasmid sequences of Escherichia coli KV7, an extended- spectrum β-Lactamase isolate derived from feces of a healthy pig

ABSTRACT
          
            We present single-contig assemblies for
            Escherichia coli
            strain KV7 (serotype O27, phylogenetic group D) and its six plasmids, isolated from a healthy pig, as determined by PacBio RS II and Illumina MiSeq sequencing. The chromosome of 4,997,475 bp and G+C content of 50.75% harbored 4,540 protein-encoding genes.
          </jats:p

Michael D. Bateman

Stefan P. W. de Vries

Srishti Gupta

Luca Guardabassi

Lina M. Cavaco

Andrew J. Grant

Mark A. Holmes

Crossref

Genome Announcements

Genome and Plasmid Sequences of Escherichia coli KV7, an Extended-Spectrum β-Lactamase Isolate Derived from Feces of a Healthy Pig

de Vries, Stefan P W

Cavaco, Lina

Online Research Database In Technology

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Downloaded from orbit.dtu.dk on: Dec 16, 2017Genome and Plasmid Sequences of Escherichia coli KV7, an Extended-Spectrum -Lactamase Isolate Derived from Feces of a Healthy PigBateman, Michael D; de Vries, Stefan P W; Gupta, Srishti; Guardabassi, Luca; Cavaco, Lina; Grant,Andrew J; Holmes, Mark APublished in:Genome AnnouncementsLink to article, DOI:10.1128/genomeA.00595-17Publication date:2017Document VersionPublisher's PDF, also known as Version of recordLink back to DTU OrbitCitation (APA):Bateman, M. D., de Vries, S. P. W., Gupta, S., Guardabassi, L., Cavaco, L., Grant, A. J., & Holmes, M. A.(2017). Genome and Plasmid Sequences of Escherichia coli KV7, an Extended-Spectrum -Lactamase IsolateDerived from Feces of a Healthy Pig. Genome Announcements, 5(38), [e00595-17]. DOI:10.1128/genomeA.00595-17Genome and Plasmid Sequences ofEscherichia coli KV7, an Extended-Spectrum -Lactamase Isolate Derivedfrom Feces of a Healthy PigMichael D. Bateman,a Stefan P. W. de Vries,a* Srishti Gupta,a Luca Guardabassi,bLina M. Cavaco,c Andrew J. Grant,a Mark A. HolmesaDepartment of Veterinary Medicine, University of Cambridge, Cambridge, United Kingdoma; Ross UniversitySchool of Veterinary Medicine, Basseterre, St. Kitts, West Indiesb; National Food Institute, Technical University ofDenmark, Kongens Lyngby, DenmarkcABSTRACT We present single-contig assemblies for Escherichia coli strain KV7 (sero-type O27, phylogenetic group D) and its six plasmids, isolated from a healthy pig, asdetermined by PacBio RS II and Illumina MiSeq sequencing. The chromosome of4,997,475 bp and GC content of 50.75% harbored 4,540 protein-encoding genes.A 2007 study (1) of healthy Danish pigs under prophylactic ceftiofur treatmentuncovered a number of distinct strains of Escherichia coli carrying the blaCTX-M-1gene and suggested that CTX-M-1 resistance spreads horizontally rather than clonally.A spontaneous nalidixic acid-resistant derivative of one of these strains that producesextended-spectrum -lactamase (serotype 27, phylogenetic group D), named KV7, wasinvestigated in a further study (2), showing that such strains thrive in pigs undertreatment with several -lactams. Sequencing of this isolate was performed in order toprovide more detailed genetic data of a strain used for experimental infections of pigs.The genomic and plasmid DNA was isolated from early exponential-phase cultures(LB medium) with Genomic-tip 100/G columns (Qiagen) and the plasmid midikit(Qiagen), respectively. Sequencing was performed using Illumina MiSeq and PaciﬁcBiosciences technologies. Prior to PacBio RS II sequencing, the DNA was cleaned witha phenol extraction, and each preparation was sequenced on a single single-moleculereal-time (SMRT) cell (Earlham Institute, Norwich, United Kingdom). For Illumina se-quencing, DNA was sheared in an M220-focused ultrasonicator (Covaris). Sequencinglibraries were prepared using the NEBNext Ultra II DNA library prep kit (New EnglandBiolabs). Sequencing was performed using 250-bp paired-end sequencing on the MiSeqplatform (V3 chemistry).Assembly of genomic and plasmid DNA PacBio sequence reads was carried out atthe Earlham Institute using the Hierarchical Genome Assembly Process (HGAP.3).Combining results from PacBio assemblies yielded single contigs for the chromosomeand for ﬁve plasmids. A sixth plasmid was identiﬁed manually using a combination ofraw reads and assembly fragments from both PacBio and Illumina sequencing assem-blies; assembly of the Illumina reads was done with plasmidSPAdes version 3.9.0 (3, 4).This plasmid (P6) harbored a large repetitive region spanning approximately 2 kb.Polishing at the single-nucleotide polymorphism level was performed using Illuminareads with Snippy version 3.1. We did not ﬁnd evidence of further plasmids.The strain has multilocus sequence types (MLST) ST57 and ST533 using theoriginal and enhanced E. coli MLST schemes, respectively. The KV7 chromosomewas 4,997,475 bp with a GC content of 50.75%. The plasmid sizes were 223,014 bp (P1),111,092 bp (P2), 71,546 bp (P3), 56,562 bp (P4), 32,688 bp (P5), and 39,510 bp (P6).Automated annotation was obtained with Prokka version 1.11 at Galaxy QueenslandReceived 8 May 2017 Accepted 10 August2017 Published 21 September 2017Citation Bateman MD, de Vries SPW, Gupta S,Guardabassi L, Cavaco LM, Grant AJ, HolmesMA. 2017. Genome and plasmid sequences ofEscherichia coli KV7, an extended-spectrumβ-lactamase isolate derived from feces of ahealthy pig. Genome Announc 5:e00595-17.https://doi.org/10.1128/genomeA.00595-17.Copyright © 2017 Bateman et al. This is anopen-access article distributed under the termsof the Creative Commons Attribution 4.0International license.Address correspondence to Mark A. Holmes,mah1@cam.ac.uk.* Present address: Stefan P. W. de Vries, Host-Microbe Interactomics, Department of AnimalSciences, Wageningen University and Research,Wageningen, the Netherlands.M.D.B. and S.P.W.D.V contributed equally to thiswork.PROKARYOTEScrossmVolume 5 Issue 38 e00595-17 genomea.asm.org 1 on October 10, 2017 by DTU Libraryhttp://genomea.asm.org/Downloaded from (5), revealing the presence of 4,540 chromosomal protein-encoding genes. In additionto blaCTX-M-1, a variety of antimicrobial resistance genes were identiﬁed on both thechromosome and several of the plasmids using BLAST together with the ARG-ANNOTdatabase (6). These included genes predicted to encode resistance to aminoglycosides(strA, strB, and aadA), -lactams (blaCTX-M-1, blaTEM-1D, ampC1, ampC2, and ampH andgenes encoding penicillin-binding proteins), chloramphenicol (catA1 and cmlA), amacrolide (mphA), sulfonamides (sulI and sulII), tetracycline (tetB), and trimethoprim(dfrA).Accession number(s). The complete genome sequences of E. coli KV7 and plasmidshave been deposited in the European Nucleotide Archive (ENA) (http://www.ebi.ac.uk/ena) and GenBank under accession no. LT795502 for the KV7 genome and LT795503,LT795504, LT795505, LT795506, LT795507, and LT795508 for the plasmids (study no.PRJEB19461).ACKNOWLEDGMENTThis work was supported by Medical Research Council (UK) grants MR/N002660/1and G1001787/1.REFERENCES1. Jørgensen CJ, Cavaco LM, Hasman H, Emborg HD, Guardabassi L. 2007.Occurrence of CTX-M-1-producing Escherichia coli in pigs treated withceftiofur. J Antimicrob Chemother 59:1040–1042. https://doi.org/10.1093/jac/dkm075.2. Cavaco LM, Abatih E, Aarestrup FM, Guardabassi L. 2008. Selection andpersistence of CTX-M-producing Escherichia coli in the intestinal ﬂora ofpigs treated with amoxicillin, ceftiofur, or cefquinome. Antimicrob AgentsChemother 52:3612–3616. https://doi.org/10.1128/AAC.00354-08.3. Bankevich A, Nurk S, Antipov D, Gurevich AA, Dvorkin M, Kulikov AS, LesinVM, Nikolenko SI, Pham S, Prjibelski AD, Pyshkin AV, Sirotkin AV, Vyahhi N,Tesler G, Alekseyev MA, Pevzner PA. 2012. SPAdes: a new genome as-sembly algorithm and its applications to single-cell sequencing. J ComputBiol 19:455–477. https://doi.org/10.1089/cmb.2012.0021.4. Antipov D, Hartwick N, Shen M, Raiko M, Lapidus A, Pevzner PA. 2016.plasmidSPAdes: assembling plasmids from whole genome sequencing data.Bioinformatics 32:3380–3387. https://doi.org/10.1093/bioinformatics/btw493.5. Seemann T. 2014. Prokka: rapid prokaryotic genome annotation. Bio-informatics 30:2068–2069. https://doi.org/10.1093/bioinformatics/btu153.6. Gupta SK, Padmanabhan BR, Diene SM, Lopez-Rojas R, Kempf M, Lan-draud L, Rolain JM. 2014. ARG-ANNOT, a new bioinformatic tool todiscover antibiotic resistance genes in bacterial genomes. AntimicrobAgents Chemother 58:212–220. https://doi.org/10.1128/AAC.01310-13.Bateman et al.Volume 5 Issue 38 e00595-17 genomea.asm.org 2 on October 10, 2017 by DTU Libraryhttp://genomea.asm.org/Downloaded from 

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Genome and Plasmid Sequences ofEscherichia coli KV7, an Extended-Spectrum -Lactamase Isolate Derivedfrom Feces of a Healthy PigMichael D. Bateman,a Stefan P. W. de Vries,a* Srishti Gupta,a Luca Guardabassi,bLina M. Cavaco,c Andrew J. Grant,a Mark A. HolmesaDepartment of Veterinary Medicine, University of Cambridge, Cambridge, United Kingdoma; Ross UniversitySchool of Veterinary Medicine, Basseterre, St. Kitts, West Indiesb; National Food Institute, Technical University ofDenmark, Kongens Lyngby, DenmarkcABSTRACT We present single-contig assemblies for Escherichia coli strain KV7 (sero-type O27, phylogenetic group D) and its six plasmids, isolated from a healthy pig, asdetermined by PacBio RS II and Illumina MiSeq sequencing. The chromosome of4,997,475 bp and GC content of 50.75% harbored 4,540 protein-encoding genes.A 2007 study (1) of healthy Danish pigs under prophylactic ceftiofur treatmentuncovered a number of distinct strains of Escherichia coli carrying the blaCTX-M-1gene and suggested that CTX-M-1 resistance spreads horizontally rather than clonally.A spontaneous nalidixic acid-resistant derivative of one of these strains that producesextended-spectrum -lactamase (serotype 27, phylogenetic group D), named KV7, wasinvestigated in a further study (2), showing that such strains thrive in pigs undertreatment with several -lactams. Sequencing of this isolate was performed in order toprovide more detailed genetic data of a strain used for experimental infections of pigs.The genomic and plasmid DNA was isolated from early exponential-phase cultures(LB medium) with Genomic-tip 100/G columns (Qiagen) and the plasmid midikit(Qiagen), respectively. Sequencing was performed using Illumina MiSeq and PacificBiosciences technologies. Prior to PacBio RS II sequencing, the DNA was cleaned witha phenol extraction, and each preparation was sequenced on a single single-moleculereal-time (SMRT) cell (Earlham Institute, Norwich, United Kingdom). For Illumina se-quencing, DNA was sheared in an M220-focused ultrasonicator (Covaris). Sequencinglibraries were prepared using the NEBNext Ultra II DNA library prep kit (New EnglandBiolabs). Sequencing was performed using 250-bp paired-end sequencing on the MiSeqplatform (V3 chemistry).Assembly of genomic and plasmid DNA PacBio sequence reads was carried out atthe Earlham Institute using the Hierarchical Genome Assembly Process (HGAP.3).Combining results from PacBio assemblies yielded single contigs for the chromosomeand for five plasmids. A sixth plasmid was identified manually using a combination ofraw reads and assembly fragments from both PacBio and Illumina sequencing assem-blies; assembly of the Illumina reads was done with plasmidSPAdes version 3.9.0 (3, 4).This plasmid (P6) harbored a large repetitive region spanning approximately 2 kb.Polishing at the single-nucleotide polymorphism level was performed using Illuminareads with Snippy version 3.1. We did not find evidence of further plasmids.The strain has multilocus sequence types (MLST) ST57 and ST533 using theoriginal and enhanced E. coli MLST schemes, respectively. The KV7 chromosomewas 4,997,475 bp with a GC content of 50.75%. The plasmid sizes were 223,014 bp (P1),111,092 bp (P2), 71,546 bp (P3), 56,562 bp (P4), 32,688 bp (P5), and 39,510 bp (P6).Automated annotation was obtained with Prokka version 1.11 at Galaxy QueenslandReceived 8 May 2017 Accepted 10 August2017 Published 21 September 2017Citation Bateman MD, de Vries SPW, Gupta S,Guardabassi L, Cavaco LM, Grant AJ, HolmesMA. 2017. Genome and plasmid sequences ofEscherichia coli KV7, an extended-spectrumβ-lactamase isolate derived from feces of ahealthy pig. Genome Announc 5:e00595-17.https://doi.org/10.1128/genomeA.00595-17.Copyright © 2017 Bateman et al. This is anopen-access article distributed under the termsof the Creative Commons Attribution 4.0International license.Address correspondence to Mark A. Holmes,mah1@cam.ac.uk.* Present address: Stefan P. W. de Vries, Host-Microbe Interactomics, Department of AnimalSciences, Wageningen University and Research,Wageningen, the Netherlands.M.D.B. and S.P.W.D.V contributed equally to thiswork.PROKARYOTEScrossmVolume 5 Issue 38 e00595-17 genomea.asm.org 1 on October 2, 2017 by CAMBRIDGE UNIVERSITY LIBRARYhttp://genomea.asm.org/Downloaded from (5), revealing the presence of 4,540 chromosomal protein-encoding genes. In additionto blaCTX-M-1, a variety of antimicrobial resistance genes were identified on both thechromosome and several of the plasmids using BLAST together with the ARG-ANNOTdatabase (6). These included genes predicted to encode resistance to aminoglycosides(strA, strB, and aadA), -lactams (blaCTX-M-1, blaTEM-1D, ampC1, ampC2, and ampH andgenes encoding penicillin-binding proteins), chloramphenicol (catA1 and cmlA), amacrolide (mphA), sulfonamides (sulI and sulII), tetracycline (tetB), and trimethoprim(dfrA).Accession number(s). The complete genome sequences of E. coli KV7 and plasmidshave been deposited in the European Nucleotide Archive (ENA) (http://www.ebi.ac.uk/ena) and GenBank under accession no. LT795502 for the KV7 genome and LT795503,LT795504, LT795505, LT795506, LT795507, and LT795508 for the plasmids (study no.PRJEB19461).ACKNOWLEDGMENTThis work was supported by Medical Research Council (UK) grants MR/N002660/1and G1001787/1.REFERENCES1. Jørgensen CJ, Cavaco LM, Hasman H, Emborg HD, Guardabassi L. 2007.Occurrence of CTX-M-1-producing Escherichia coli in pigs treated withceftiofur. J Antimicrob Chemother 59:1040–1042. https://doi.org/10.1093/jac/dkm075.2. Cavaco LM, Abatih E, Aarestrup FM, Guardabassi L. 2008. Selection andpersistence of CTX-M-producing Escherichia coli in the intestinal flora ofpigs treated with amoxicillin, ceftiofur, or cefquinome. Antimicrob AgentsChemother 52:3612–3616. https://doi.org/10.1128/AAC.00354-08.3. Bankevich A, Nurk S, Antipov D, Gurevich AA, Dvorkin M, Kulikov AS, LesinVM, Nikolenko SI, Pham S, Prjibelski AD, Pyshkin AV, Sirotkin AV, Vyahhi N,Tesler G, Alekseyev MA, Pevzner PA. 2012. SPAdes: a new genome as-sembly algorithm and its applications to single-cell sequencing. J ComputBiol 19:455–477. https://doi.org/10.1089/cmb.2012.0021.4. Antipov D, Hartwick N, Shen M, Raiko M, Lapidus A, Pevzner PA. 2016.plasmidSPAdes: assembling plasmids from whole genome sequencing data.Bioinformatics 32:3380–3387. https://doi.org/10.1093/bioinformatics/btw493.5. Seemann T. 2014. Prokka: rapid prokaryotic genome annotation. Bio-informatics 30:2068–2069. https://doi.org/10.1093/bioinformatics/btu153.6. Gupta SK, Padmanabhan BR, Diene SM, Lopez-Rojas R, Kempf M, Lan-draud L, Rolain JM. 2014. ARG-ANNOT, a new bioinformatic tool todiscover antibiotic resistance genes in bacterial genomes. AntimicrobAgents Chemother 58:212–220. https://doi.org/10.1128/AAC.01310-13.Bateman et al.Volume 5 Issue 38 e00595-17 genomea.asm.org 2 on October 2, 2017 by CAMBRIDGE UNIVERSITY LIBRARYhttp://genomea.asm.org/Downloaded from 

Genome and Plasmid Sequences of Escherichia coli KV7, an Extended-Spectrum β-Lactamase Isolate Derived from Feces of a Healthy Pig.

https://www.repository.cam.ac.uk/bitstream/1810/286050/1/Genome%20and%20Plasmid%20Sequences%20of%20Escherichia%20coli%20KV7%2c%20an%20Extended-Spectrum%20%ce%b2-Lactamase%20Isolate%20Derived%20from%20Feces%20of%20a%20Healthy%20Pig.pdf

Genome and Plasmid Sequences of Escherichia coli KV7, an Extended-Spectrum β-Lactamase Isolate Derived from Feces of a Healthy Pig.

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