Ann Lab Med.  2019 Mar;39(2):167-175. 10.3343/alm.2019.39.2.167.

Identification and Characterization of NDM-1-producing Hypervirulent (Hypermucoviscous) Klebsiella pneumoniae in China

• Liu Z
• Gu Y
• Li X
• Liu Y
• Ye Y
• Guan S
• Li J

• KMID: 2425972
• DOI: http://doi.org/10.3343/alm.2019.39.2.167

Abstract

BACKGROUND
Carbapenem-resistant hypervirulent (hypermucoviscous) Klebsiella pneumoniae (CR-HMKP) poses a significant public health challenge. We investigated its epidemiology and molecular characteristics in a tertiary care hospital in eastern China.
METHODS
CR-HMKP were identified among 106 non-duplicated carbapenem-resistant K. pneumoniae isolates (from June 2013 to September 2017) using the string test. The pulsotype (PT) and sequence type (ST) of CR-HMKP isolates were determined using pulsed-field gel electrophoresis and multilocus sequence typing. Resistance determinants, capsular serotypes, and virulence genes were detected by PCR and sequencing. Representative isolates from each PT were selected, and their virulence phenotypes were established using the serum killing and Galleria mellonella lethality assays.
RESULTS
Of the 106 isolates, 13 (12.3%) were CR-HMKP. Seven were positive for bla NDM-1 and shared the same genotype (PT5/ST1764); the others were positive for bla KPC-2, belonged to ST11, and were divided into four different PTs. The serotype of all bla NDM-1-positive isolates was K64, while that of bla KPC-2-positive isolates were K47 (N=4) and K64 (N=2). The NDM-1-producing HMKP isolates were positive for aerobactin, exhibited high serum resistance, and elicited significantly increased larval mortality compared with the other isolates. All patients had received invasive treatment prior to infection by NDM-1-producing HMKP. The infections occurred between July and August 2016 and were hospital-acquired.
CONCLUSIONS
NDM-1-producing HMKP ST1764 isolates were identified; this is the first report worldwide on an outbreak of nosocomial infection caused by these isolates. Effective surveillance and strict infection control strategies should be implemented to prevent CR-HMKP dissemination.

Keyword

Carbapenem-resistant; Hypervirulent; Hypermucoviscous; Klebsiella pneumoniae; NDM-1; Epidemiology; Molecular characteristics

MeSH Terms

China*
Cross Infection
Electrophoresis, Gel, Pulsed-Field
Epidemiology
Genotype
Homicide
Humans
Infection Control
Klebsiella pneumoniae*
Klebsiella*
Mortality
Multilocus Sequence Typing
Phenotype
Pneumonia
Polymerase Chain Reaction
Public Health
Serogroup
Tertiary Healthcare
Virulence

Figure

• 

  Fig. 1 Pulse-field gel electrophoresis (PFGE) dendrograms, genotype, serotype, resistance and virulence genes of the CR-HMKP isolates. The isolates that exhibited PFGE dendrograms with more than 90% similarity are considered one pulsotype (PT). *The isolation date is listed as month-day-year; †Only partial results are shown. Of the other virulence genes tested, entB, iucA, kfuBC, mrkD, rmpA2, ureA, wabG, ybtS, and ycfM were detected in all isolates, and the magA gene was not detected in any of them.Abbreviations: Aer, aerobactin; KPC, Klebsiella pneumoniae carbapenemase; NDM, New Delhi metallo-β-lactamase; ST, sequence type.

• 

  Fig. 2 Serum killing assays of the tested isolates. Data are presented as mean±SE, and log10-transformed values were utilized to normalize the data (N=3 for each isolate).Abbreviations: H-control, hypervirulence control; L-control, low-virulence control; Lg, log10 -transformed values.

• 

  Fig. 3 Survival curves for G. mellonella larvae inoculated with 1×107 (A), 1×106 (B), 1×105 (C), and 1×104 (D) colony-forming units of the tested isolates, and the data shown are from a single representative experiment out of three repeats. *The curves of KPN11, KPN18, KPN34, KPN100, KPN49L-control and negative control are completely overlapping.Abbreviations: H-control, hypervirulence control; L-control, low-virulence control.

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