A rapid matrix-assisted laser desorption ionization-time of flight mass spectrometry-based method for single-plasmid tracking in an outbreak of carbapenem-resistant Enterobacteriaceae.
Carbapenem-resistant Enterobacteriaceae (CRE) have spread globally and represent a serious and growing threat to public health. Rapid methods for tracking plasmids carrying carbapenemase genes could greatly benefit infection control efforts. Here, we demonstrate that real-time, direct tracking of a single plasmid in a bacterial strain responsible for an outbreak is possible using a commercial matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) system. In this case, we retrospectively tracked the bla(KPC) carbapenemase gene-bearing pKpQIL plasmid responsible for a CRE outbreak that occurred at the NIH Clinical Center in 2011. An ∼ 11,109-Da MS peak corresponding to a gene product of the bla(KPC) pKpQIL plasmid was identified and characterized using a combination of proteomics and molecular techniques. This plasmid peak was present in spectra from retrospectively analyzed K. pneumoniae outbreak isolates, concordant with results from whole-genome sequencing, and absent from a diverse control set of bla(KPC)-negative clinical Enterobacteriaceae isolates. Notably, the gene characterized here is located adjacent to the bla(KPC) Tn4401 transposon on the pKpQIL plasmid. Sequence analysis demonstrates the presence of this gene in other bla(KPC) Tn4401-containing plasmids and suggests that this signature MS peak may be useful in tracking other plasmids conferring carbapenem resistance. Plasmid identification using this MALDI-TOF MS method was accomplished in as little as 10 min from isolated colonies and 30 min from positive (spiked) blood cultures, demonstrating the potential clinical utility for real-time plasmid tracking in an outbreak.
Microbiology Service, Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA.
Anna F
AF
Lau
Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA.
Honghui
H
Wang
Microbiology Service, Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA.
Rebecca A
RA
Weingarten
Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA.
Steven K
SK
Drake
Critical Care Medicine Department, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA.
Anthony F
AF
Suffredini
Research Technologies Branch, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
Mark K
MK
Garfield
Proteomics Core Facility, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
Yong
Y
Chen
Proteomics Core Facility, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
Marjan
M
Gucek
Microbiology Service, Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA.
Jung-Ho
JH
Youn
Microbiology Service, Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA.
Frida
F
Stock
Laboratory for Informatics Development, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA.
Hanna
H
Tso
Laboratory for Informatics Development, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA.
Jim
J
DeLeo
Laboratory for Informatics Development, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA.
James J
JJ
Cimino
Microbiology Service, Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA.
Karen M
KM
Frank
Microbiology Service, Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA john.dekker@nih.gov.
John P
JP
Dekker
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000032:analysis / Q000737:chemistry; D015373:Bacterial Typing Techniques / Q000379:methods*; D015780:Carbapenems / Q000494:pharmacology; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004196:Disease Outbreaks; D004755:Enterobacteriaceae / Q000737:chemistry / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D004756:Enterobacteriaceae Infections / Q000453:epidemiology* / Q000382:microbiology; D005798:Genes, Bacterial; D006801:Humans; D017720:Molecular Epidemiology / Q000379:methods; D008970:Molecular Weight; D010957:Plasmids / Q000032:analysis*; D017422:Sequence Analysis, DNA; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D013997:Time Factors; D018440:beta-Lactam Resistance
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D015780:Carbapenems; D004269:DNA, Bacterial