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

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 bla_CMY-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, bla_CMY-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, bla_TEM-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).