Characterization of a multidrug-resistant Klebsiella pneumoniae ST3330 clone responsible for a nosocomial outbreak in a neonatal intensive care unit.

Background: The incidence of Klebsiella pneumonia (Kp), which has often been found to produce, extended spectrum beta-lactamase (ESBL), is rising rapidly and poses a serious risk to neonates. To date, the mechanisms related to the spread of ESBL-Kp have not been fully elucidated. This study aimed to investigate the phenotypes, genotypes, and genetic relatedness of ESBL-KP that caused an outbreak of sepsis among neonates in an intensive care unit of a Beijing hospital. Methods: Between April 2016 and May 2018, 21 non-repetitive clinical ESBL-Kp isolates were collected from a neonatal intensive care unit (NICU) in Beijing, China and were retrospectively analyzed. Pulsed-field gel electrophoresis (PFGE) was used to analyze genetic relatedness, a VITEK 2 AST test kit was used to test antimicrobial susceptibility, sequence type (ST) was analyzed through multilocus sequence typing (MLST), and resistance genes were identified by PCR. Virulence gene profiles, biofilm formation assay, and serum killing assay were used for virulence-associated determinants. Results: All strains expressed the same antibiotype, combining ESBL production, third generation cephalosporins resistance and carbapenems sensitive. Sixteen of them produced beta-lactamases (CTX-M-3 and TEM-1B), while others possessed CTX-M-15, CTX-M-24, CTX-M-66, TEM-1C, SHV-26, SHV-172, and OXA-1. PFGE confirmed 5 types (A, B, C, D and E) and MLST identified a ST3330 clone (16 strains), a ST2791 clone (2 strains), a ST37 clone (1 strain), a ST34 clone (1 strain), and a ST2740 clone (1 strain). PFGE type A strains, which belong to ST3330, were identified as the main pathogens involved in the outbreak. All isolates contained virulence genes iutA and mrk. PFGE type A carried both mrk (type 3 fimbriae, biofilm formation) and fimH (type 1 fimbriae), and other STs possessed mrk. Isolates belonging to the endemic ST3330 lineage produced more biofilm than other ST isolates (median OD590 1.829 vs. 0.2280, respectively; P<0.0001). All five PFGE types isolates showed serum high sensitivity (grade 1). Conclusions: The dissemination and outbreak of ESBL-producing K. pneumoniae in this study seemed to be clonal, and the outbreak was mainly caused by ST3330 K. pneumoniae. The detection of genes (mrk and fimH) belonging to the biofilm formation may partly explain the epidemic strain has high colonization and diffusion potential.