Draft Genome Sequence of a Multidrug-Resistant Klebsiella pneumoniae Isolate from a Prosthetic Joint Infection.

Extended-spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae strain 9120005127 was isolated from a wound infection. We describe the draft genome sequence and antibiotic susceptibility of this strain.

ANNOUNCEMENT

Klebsiella pneumoniae is a Gram-negative, nonmotile, facultative anaerobic, lactose-fermenting bacillus. This bacterium is a human commensal, opportunistic pathogen and leading agent of health care-associated infection (1, 2). We sequenced the draft genome of the extended-spectrum β-lactamase (ESBL)-producing, multidrug-resistant strain K. pneumoniae 9120005127, isolated from an acute prosthetic joint skin infection at the Central Texas Veterans Health Care System.

A wound swab was cultured on tryptic soy agar (TSA) with sheep blood agar (Remel, Inc., San Diego, CA) at 35°C overnight. The isolate was identified as K. pneumoniae by matrix-assisted laser desorption ionization–time of flight mass spectrometry (3), and antimicrobial susceptibility testing was performed using the Vitek 2 system (bioMérieux, France) according to the manufacturer’s instructions. E-tests (AB Biodisk, Solna, Sweden) were used to determine the MIC to colistin according to the packet insert instructions, using Mueller-Hinton agar with an inoculum determined by a turbidity of 0.5 McFarland standard in a 0.45% NaCl suspension.

Bacteria were grown on the TSA with sheep blood agar plate. Total DNA was extracted using the QIAamp DNA microkit (Qiagen, Hilden, Germany). The DNA was quantified using a Qubit 4 fluorometer (Invitrogen, Carlsbad, CA) according to the manufacturer’s instructions. The library was constructed using the Nextera DNA Flex library prep kit (Illumina, San Diego, CA). The library was sequenced using an Illumina NextSeq 550 instrument. The de novo assembly was completed using the SPAdes version 3.9.0 genome assembler with default parameters (4) on the BaseSpace platform (Illumina). The Phred quality score was 30.56, and a total of 497.2 Mbp of reads was generated. In the assembled genome, 61 contigs with an N50 value of 222,418 bp were generated. The final genome length was comprised of 5,362,693 bp with a 93-fold average coverage and 57.41% G+C content.

Default parameters were used for all software unless otherwise specified. Average nucleotide identity (ANI) analysis (http://enve-omics.ce.gatech.edu/ani/) (5) with the K. pneumoniae type strain genome (HS11286; GenBank accession number NC_016845.1) showed the mean nucleotide identity as 99.26%. Sequence type (ST) 37 was determined by multilocus sequence typing (MLST) analysis (http://pubmlst.org) (6, 7).

Table 1 shows the MIC(s) of antibiotics for the isolate and the antibiotic resistance genes found in the genome. The resistance profile was determined using the Clinical and Laboratory Standards Institute (CLSI) MIC breakpoints for extended-spectrum β-lactamase in Klebsiella pneumonia (CLSI M100, 29th edition, Table 3A) (8). This strain is a multidrug-resistant organism showing resistance against various classes of antimicrobials, including penicillin, cephalosporins, monobactams, quinolones, aminoglycosides, tetracyclines, and sulfonamides.

TABLE 1

MICs of Klebsiella pneumoniae DN2020

Antibiotic(s)	MIC	Interpretationb	Putative resistance gene(s)
Amikacin	≤2	S	aac(6′)-Ib-cr
Gentamicin	≥16	R	aac(3)-IIa
Tobramycin	≥16	R	aac(6′)-Ib-cr, aac(3)-Iia
Cefazolin	≥64	R
Cephalothin	≥64	R	blaSHV-79, blaSHV-85, blaSHV-89, blaTEM-1B
Cefotetan	≤4	S
Cefoxitin	≤4	S
Cefuroxime-axetil	≥64	R
Cefuroxime-sodium	≥64	R
Cefotaxime	≥64	R	blaSHV-40, blaCTX-M-15
Ceftizoxime	≤1	R
Ceftriaxone	≥64	R	blaSHV-40, blaCTX-M-15
Ceftazidime	4	R	blaSHV-40, blaCTX-M-15
Cefepime	4	R	blaSHV-40, blaCTX-M-15, blaOXA-1
Amoxicillin-clavulanic acid	16	I	blaSHV-56, blaOXA-1
Ampicillin	≥32	R	blaSHV-40, blaSHV-56, blaSHV-79, blaSHV-85, blaSHV-89, blaTEM-1B, blaCTX-M-15, blaOXA-1
Ampicillin-sulbactam	≥32	R
Piperacillin	≥128	R	blaSHV-40, blaSHV-56, blaSHV-79, blaSHV-85, blaSHV-89, blaTEM-1B, blaCTX-M-15, blaOXA-1
Piperacillin-tazobactam	32	I	blaSHV-56, blaOXA-1
Aztreonam	8	R	blaSHV-40, blaCTX-M-15
Ertapenem	≤05	S
Imipenem	≤025	S
Meropenem	≤025	S
Ciprofloxacin	2	R	aac(6′)-Ib-cr, qnrB1, oqxA, oqxB
Levofloxacin	1	I
Moxifloxacin	4	I
Norfloxacin	2	S
Trimethoprim-sulfamethoxazole	≥320	R	sul2, dfrA14
Tetracycline	≥16	R	tet(A)
Omadacycline	>8	R
Tigecycline	2	S
Colistin	0.125	S

Bold text indicates results that are Advanced Expert System (AES) modified.

S, susceptible; R, resistant; I, intermediate.

The NCBI Prokaryotic Genome Annotation Pipeline (PGAP) version 4.13 (9) predicted 5,119 protein-coding sequences (CDSs), 9 copies of the complete rRNA, and 82 tRNAs. ResFinder version 4.0 analysis (https://cge.cbs.dtu.dk/services/ResFinder/) (10) identified the following putative antibiotic resistance genes: blaTEM (broad spectrum β-lactamase); blaSHV, blaCTX, and blaOXA (ESBL); aac(3)-Ila and aac(6′)-lb-cr (aminoglycoside); qnrB1, oqxA, and oqxB (quinolone); sul2 and dfrA14 (trimethoprim-sulfamethoxazole); and tet(A) (tetracycline) (11). A summary of MICs and resistance genes is given in Table 1.

Data availability.

This sequence has been deposited at DDBJ/ENA/GenBank under the accession number JADNRR000000000. The raw sequence reads are available under the SRA accession number SRR13037727.