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Prokaryotes

Whole-Genome Sequence of CMY-2 β-Lactamase-Producing Salmonella enterica Serovar Typhimurium Strain L-3553

Tsuyoshi Sekizuka, Ken-ichi Lee, Makoto Kuroda, Masahiro Kusumoto, Taketoshi Iwata, Ikuo Uchida, Kiyoshi Tanaka, Yukino Tamamura, Masato Akiba
Tsuyoshi Sekizuka
aPathogen Genomics Center, National Institute of Infectious Diseases, Shinjuku, Tokyo, Japan
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Ken-ichi Lee
bBacterial and Parasitic Disease Research Division, National Institute of Animal Health, National Agriculture and Food Research Organization, Tsukuba, Ibaraki, Japan
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Makoto Kuroda
aPathogen Genomics Center, National Institute of Infectious Diseases, Shinjuku, Tokyo, Japan
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Masahiro Kusumoto
bBacterial and Parasitic Disease Research Division, National Institute of Animal Health, National Agriculture and Food Research Organization, Tsukuba, Ibaraki, Japan
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Taketoshi Iwata
bBacterial and Parasitic Disease Research Division, National Institute of Animal Health, National Agriculture and Food Research Organization, Tsukuba, Ibaraki, Japan
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Ikuo Uchida
cHokkaido Research Station, National Institute of Animal Health, National Agriculture and Food Research Organization, Sapporo, Hokkaido, Japan
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Kiyoshi Tanaka
cHokkaido Research Station, National Institute of Animal Health, National Agriculture and Food Research Organization, Sapporo, Hokkaido, Japan
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Yukino Tamamura
cHokkaido Research Station, National Institute of Animal Health, National Agriculture and Food Research Organization, Sapporo, Hokkaido, Japan
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Masato Akiba
bBacterial and Parasitic Disease Research Division, National Institute of Animal Health, National Agriculture and Food Research Organization, Tsukuba, Ibaraki, Japan
dGraduate School of Life and Environmental Sciences, Osaka Prefecture University, Izumisano, Osaka, Japan
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DOI: 10.1128/genomeA.00711-14
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ABSTRACT

Salmonella enterica serovar Typhimurium pulsed-field gel electrophoresis cluster VII has been isolated from cattle populations in Japan since the mid-2000s. Some cluster VII isolates exhibited extended-spectrum cephalosporin resistance defined by the blaCMY-2 gene located in a chromosomal genomic island, GI-VII-6. We determined the whole-genome sequence of strain L-3553 as the reference strain.

GENOME ANNOUNCEMENT

Salmonella enterica serovar Typhimurium strains belonging to pulsed-field gel electrophoresis cluster VII and having no recognized phage type have been dominant among cattle populations in Hokkaido, Japan, since the mid-2000s (1). Some of the cluster VII isolates exhibited resistance to extended-spectrum cephalosporin and harbored a resistance island, GI-VII-6, in their chromosome (2). GI-VII-6 contains multiple antimicrobial resistance genes, including blaCMY-2. We determined the whole-genome sequence of GI-VII-6-harboring S. Typhimurium strain L-3553, which was isolated in 2004, as the reference strain.

The whole-genome sequencing DNA library of this strain was constructed by a Nextera XT DNA sample prep kit (Illumina, San Diego, CA) using an Illumina MiSeq (Illumina) and a MiSeq 500 cycle kit v. 2 (Illumina). The trimmed and filtered short reads were assembled with the De Novo Assembler program in CLC Genomics Workbench v. 6.5 (CLC Bio, Aarhus, Denmark). The predicted gaps were amplified using specific PCR primer pairs, followed by Sanger DNA sequencing using a BigDye Terminator v. 3.1 cycle sequencing kit (Applied Biosystems, Foster City, CA). To validate the gap-closed sequences, the short reads were aligned with tentative whole L-3553 genome sequences using BWA-SW (v. 0.6.1) (3) and SAMtools (v. 0.1.18) (4). Gene prediction was performed with the RAST annotation server for the whole-genome sequence (5).

The chromosome of L-3553 comprised 5,051,841 bp (G+C content, 53.18%), and the plasmid denominated as pST3553 comprised 132,611 bp (G+C content, 54.42%). In total, 4,947 and 159 open reading frames were identified in the chromosome and pST3553, respectively. Six prophages were identified in the chromosome, including three common prophages, i.e., Gifsy-1, Gifsy-2, and ST64B, which are present in other S. Typhimurium strains. However, the common prophages Fels-1 and Fels-2 were not found. The effector protein-encoding genes gogB in Gifsy-1, sodC and sseI in Gifsy-2, and sseK3 in ST64B were all intact. Three additional prophage regions were identified using Phage_Finder (6), which were similar to bacteriophages P22 (coverage, 46%; identity, 97%), PsP3 (coverage, 62%; identity, 92%), and P2 (coverage, 72%; identity, 97%). GI-VII-6 contained nine antimicrobial resistance genes, aadA, strA, strB, blaCMY-2, floR, sul1, sul2, tet(A), and dfrA12, and was flanked by directly repeated IS26 copies. No other horizontally acquired antimicrobial resistance genes were observed in the chromosome. The sequence of pST3553 comprised the serovar-specific virulence plasmid pSLT (7) and a single horizontally acquired region containing five antimicrobial resistance genes, including aadA1, aph(3′)-Ic, blaTEM-1, sul1, and tet(A); thus, pST3553 is regarded as a virulence-resistance plasmid. The horizontally acquired region was flanked by IS1294 copies. The whole sequence of pST3553 was highly similar to that of the pYT2 plasmid (coverage, 100%; identity, 99%), which is harbored by S. Typhimurium strain KT262 (8).

Nucleotide sequence accession numbers.The whole-genome sequence of S. Typhimurium L-3553 has been deposited in the DNA Data Bank of Japan under accession no. AP014565 for the chromosome and AP014566 for the plasmid pST3553.

ACKNOWLEDGMENTS

This research was supported by the Japan Society for the Promotion of Science, a Grant-in-Aid for Scientific Research (C), 2013 to 2015 (25460553), and a Grant-in-Aid from the Ministry of Health, Labour, and Welfare of Japan (H24-Shokuhin-Ippan-008).

FOOTNOTES

    • Received 21 June 2014.
    • Accepted 3 July 2014.
    • Published 24 July 2014.
  • Copyright © 2014 Sekizuka et al.

This is an open-access article distributed under the terms of the Creative Commons Attribution 3.0 Unported license.

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Whole-Genome Sequence of CMY-2 β-Lactamase-Producing Salmonella enterica Serovar Typhimurium Strain L-3553
Tsuyoshi Sekizuka, Ken-ichi Lee, Makoto Kuroda, Masahiro Kusumoto, Taketoshi Iwata, Ikuo Uchida, Kiyoshi Tanaka, Yukino Tamamura, Masato Akiba
Genome Announcements Jul 2014, 2 (4) e00711-14; DOI: 10.1128/genomeA.00711-14

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Whole-Genome Sequence of CMY-2 β-Lactamase-Producing Salmonella enterica Serovar Typhimurium Strain L-3553
Tsuyoshi Sekizuka, Ken-ichi Lee, Makoto Kuroda, Masahiro Kusumoto, Taketoshi Iwata, Ikuo Uchida, Kiyoshi Tanaka, Yukino Tamamura, Masato Akiba
Genome Announcements Jul 2014, 2 (4) e00711-14; DOI: 10.1128/genomeA.00711-14
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