Draft Genome Sequence of a Hypermucoviscous Extended-Spectrum-β-Lactamase-Producing Klebsiella quasipneumoniae subsp. similipneumoniae Clinical Isolate.

A clinical isolate of extended-spectrum-β-lactamase-producing Klebsiella quasipneumoniae subsp. similipneumoniae 06-219 with hypermucoviscosity phenotypes obtained from a urine culture of an adult patient was used for whole-genome sequencing. Here, we report the draft genome sequences of this strain, consisting of 53 contigs with an ~5.6-Mb genome size and an average G+C content of 57.36%. The annotation revealed 6,622 coding DNA sequences and 77 tRNA genes.

GENOME ANNOUNCEMENT

Klebsiella quasipneumoniae is a new bacterial species that has been recently described. It is broadly related to K. pneumoniae and K. variicola (1), and includes two subspecies: K. quasipneumoniae subsp. quasipneumoniae and K. quasipneumoniae subsp. similipneumoniae. Both have been exclusively isolated from human infections (2). K. quasipneumoniae subsp. quasipneumoniae and K. variicola with hypermucoviscosity phenotypes have been described (3, 4). A collection of 220 extended-spectrum-β-lactamase (ESBL)-producing K. pneumoniae clinical isolates (2005 and 2014) were retrospectively screened for hypermucoviscosity phenotype, using the semi-quantitatively string test (5). All isolates were previously identified by means of the MicrosScan Walkaway system (Dade Behring, West Sacramento, CA), and the determination of the ESBL-producing isolates was carried out by double-disc synergy test (6).

As a result of screening, the K. pneumoniae 06-219 isolate was identified with the hypermucoviscosity phenotype. This isolate was obtained from a urine culture of a 32-year-old man at the University Hospital in Monterrey, Nuevo Leon, Mexico in 2006. It was analyzed by multiplex-PCR (M-PCR-1) (7), showing a negative result for the molecular markers of K. variicola, K. pneumoniae, and for the mtnC gene that corresponds to Klebsiella spp. genera. Then, this isolate was used for the whole-genome sequencing.

A total genomic sample was purified using a DNeasy kit (Qiagen, Germany). The genome sequence was obtained using the Illumina (MiSeq) platform and a total of 4,705,070 pair-end reads with a length of 300-bp were obtained. Quality-based trimming was performed with the SolexaQA software and de novo assembly was done with SPAdes v3.1.1. In total, 57 contigs with an N50 of 1,041,587-bp were obtained. The estimated genome size was 5,645,875 bp with a 150× coverage. Gene prediction and annotation were carried out using the bioinformatic MicroScope platform (8). A total of 5,622 coding DNA sequences (CDS) and 77 tRNA genes were identified. The rmpA and rmpA2 genes described in hypervirulent-K. pneumoniae turned out to be absent in the 06-219 genome. This bacteria contains the following virulence-associated determinants: allABCDRS, entB, iroN, iutA, kfuABC, mrkABCDFHIJ, uge, ureA, and wabG. The chromosomal OKP-B-2 and the plasmid-encoded TEM-1 and ESBL SHV-12 genes, which encode the β-lactamase genes, were identified. The operons conferring heavy metal resistance were identified: merRTPCAPE (mercury), pcoABCDERS (copper), silCERS (silver), pbrACR (lead), and terABCDEWYZ (tellurium). The latter has been associated with hypervirulent-K. pneumoniae clones (9).

To determine the bacterial specie to which isolate 06-219 belongs, an average nucleotide identity (ANI) (10) and rpoB gene analysis were carried out using the K. quasipneumoniae subsp. quasipneumoniae 18A069 (CBZM000000000), K. quasipneumoniae subsp. similipneumoniae 07A044 (CBZR000000000), K. variicola At-22 (CP001891), and K. pneumoniae MGH78578 (CP000647) reference genomes. A value of 99.23% for K. quasipneumoniae subsp. similipneumoniae, was obtained in comparison with a value of 96.47% obtained for K. quasipneumoniae subsp. quasipneumoniae. A value lower to 93.27% was obtained for K. variicola At-22 and K. pneumoniae MGH78578 genomes. In addition, the phylogenetic analysis of the complete rpoB gene showed the same results (data not shown). The above-mentioned analyses indicate that isolate 06-219 corresponds to hypermucoviscous ESBL-producing K. quasipneumoniae subsp. similipneumoniae.

Nucleotide sequence accession numbers.

The annotated genome sequence is available at the European Nucleotide Archive under the accession numbers FKLR01000001 to FKLR01000057.