Using Whole Genome Sequencing to Trace, Control and Characterize a Hospital Infection of IMP-4-Producing Klebsiella pneumoniae ST2253 in a Neonatal Unit in a Tertiary Hospital, China

Abstract

Background: The purpose of this study is to use whole genome sequencing (WGS) combined with epidemiological data to track a hospital infection of the carbapenem-resistant Klebsiella pneumoniae (CRKP), which affected 3 neonatal patients in the neonatal intensive care unit (NICU). Methods: The minimum inhibitory concentrations for the antimicrobial agents were determined according to the guidelines of the Clinical and Laboratory Standards Institute. Beta-lactamases were investigated using the polymerase chain reaction and DNA sequencing. The transferability of the plasmid was investigated by a conjugation experiment. The clonal relationships were evaluated using multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). WGS and single nucleotide polymorphisms (SNPs) analysis were performed on the CRKP isolates to investigate how the infection might progress. Results: Nine CRKP isolates were obtained from the NICU, seven from three patients, one from a duster cloth and one from the hand of a nurse, they all harbored blaIMP-4. Other resistance genes including blaKPC-2, blaIMP-4, blaSHV-1, blaTEM-1, blaCTX-M-15, and blaDHA-1 were also detected. PFGE analysis showed that IMP-4-producing K. pneumoniae were clonally related, and MLST assigned them to a new sequence type 2253. The SNP variations throughout the genome divided the 9 strains into three clades. Clade 1 comprised 7 strains (K1- K2 and K4-K8), whereas clade 2 and 3 consisted of only one strain each: K3 and K9, respectively.The sputum isolate K3 from patient 3 was the most distinct one differing from the other eight isolates by 239-275 SNPs. Conclusions: This is a report of using WGS to track a hospital infecion of IMP-4-producing K. pneumoniae ST2253 among neonates. Nosocomial surveillance systems are needed to limit the spread of the infection caused by these pathogens resulting from the environmental exposure in NICUs.

Keywords: Klebsiella pneumoniae; carbapenem; drug resistance; single nucleotide polymorphisms; whole genome sequencing.

Figure 1

Temporal graph of the nine isolates of ST2253 isolated from three patients and the environment of NICU (K1–K9). The used symbols are as following: Black circles, the patients and environment; P, patient; EN, environment; Red triangle, date; Yellow, detection time; Blue, admission time and discharge time; Bold black line, period of hospitalization.

Figure 2

Dendrogram analysis. Dendrogram generated using the Fingerprinting II Informatix software package (Bio-Rad Laboratories, Hercules, CA) showing the relatedness of the fingerprints (XbaI-PFGE) for the 9 K. pneumoniae strains. The phylogenetic tree was constructed using the Dice coefficient and UPGMA clustering. A genetic similarity index scale is shown to the left of the dendrogram. P, patient.

Figure 3

WGS and SNPs analysis. (A) The single nucleotide polymorphisms (SNPs) numbers between each IMP-4-producing strain and the reference strain. (B) The phylogenetic analysis of the nine CRKP isolates based on SNPs.