TY - JOUR
T1 - Rapid identification of New Delhi metallo-β-lactamase (NDM) using tryptic peptides and LC-MS/MS
AU - Wang, Honghui
AU - Strich, Jeffrey R.
AU - Drake, Steven K.
AU - Chen, Yong
AU - Youn, Jung Ho
AU - Rosenberg, Avi Z.
AU - Gucek, Marjan
AU - Dekker, John P.
AU - Suffredini, Anthony F.
N1 - Funding Information:
This work was supported in part by the Intramural Research Programs of the National Institutes of Health Clinical Center (H.W., J.R.S., S.K.D., J.-H.Y., A.F.S., and J.P.D.); the National Institute of Allergy and Infectious Diseases (J.P.D.); the National Heart, Lung, and Blood Institute (M.G. and Y.C.); and the National Institute of Diabetes and Digestive and Kidney Diseases and Johns Hopkins University (A.Z.R.). The content is solely our responsibility and does not necessarily represent the official views of the NIH or the U.S. Government. S.K.D. has been involved in a collaborative agreement with Bruker Daltonics, Inc., to develop organism databases for MALDI-TOF MS, independently of this study. Bruker Daltonics, Inc., had no role in the work published here. We declare that we have no competing financial interests.
Funding Information:
We acknowledge the CDC and FDA Antibiotic Resistance Isolate Bank (ARISOLATEBANK), from which 22 blaNDM-containing isolates (listed in Table 4) were obtained. This work was supported in part by the Intramural Research Programs of the National Institutes of Health Clinical Center (H.W., J.R.S., S.K.D., J.-H.Y., A.F.S., and J.P.D.); the National Institute of Allergy and Infectious Diseases (J.P.D.); the National Heart, Lung, and Blood Institute (M.G. and Y.C.); and the National Institute of Diabetes and Digestive and Kidney Diseases and Johns Hopkins University (A.Z.R.). The content is solely our responsibility and does not necessarily represent the official views of the NIH or the U.S. Government. S.K.D. has been involved in a collaborative agreement with Bruker Daltonics, Inc., to develop organism databases for MALDI-TOF MS, independently of this study. Bruker Daltonics, Inc., had no role in the work published here. We declare that we have no competing financial interests. H.W., S.K.D., A.F.S., and J.P.D. conceived the project design. H.W., J.R.S., S.K.D., Y.C., and J.-H.Y. carried out the experiments. H.W., S.K.D., J.R.S., A.F.S., and J.P.D. performed primary analysis of the data, and Y.C., M.G., and A.Z.R. critically reviewed the primary analysis and provided LC-MS instrument support. H.W., J.R.S., S.K.D., A.F.S., and J.P.D. cowrote the manuscript. All of us critically evaluated and edited the manuscript. We have no conflicts to disclose.
Publisher Copyright:
Copyright © 2019 American Society for Microbiology. All Rights Reserved.
PY - 2019
Y1 - 2019
N2 - There is significant interest in the development of mass spectrometry (MS) methods for antimicrobial resistance protein detection, given the ability of these methods to confirm protein expression. In this work, we studied the performance of a liquid chromatography, tandem MS multiple-reaction monitoring (LC-MS/MS MRM) method for the direct detection of the New Delhi metallo-lactamase (NDM) carbapenemase in clinical isolates. Using a genoproteomic approach, we selected three unique peptides (SLGNLGDADTEHYAASAR, AFGAAFPK, and ASMIVMSHS APDSR) specific to NDM that were efficiently ionized and spectrally well-defined. These three peptides were used to build an assay with turnaround time of 90 min. In a blind set, the assay detected 21/24 blaNDM-containing isolates and 76/76 isolates with negative results, corresponding to a sensitivity value of 87.5% (95% confidence interval [CI], 67.6% to 97.3%) and a specificity value of 100% (95% CI, 95.3% to 100%). One of the missed identifications was determined by protein fractionation to be due to low (0.1 fm/g) NDM protein expression (below the assay limit of detection). Parallel disk diffusion susceptibility testing demonstrated this isolate to be meropenem susceptible, consistent with low NDM expression. Total proteomic analysis of the other two missed identifications did not detect NDM peptides but detected other proteins expressed from the blaNDM-containing plasmids, confirming that the plasmids were not lost. The measurement of relative NDM concentrations over the entire isolate test set demonstrated variability spanning 4 orders of magnitude, further confirming the remarkable range that may be seen in levels of NDM expression. This report highlights the sensitivity of LC-MS/MS to variations in NDM protein expression, with implications for how this technology may be used.
AB - There is significant interest in the development of mass spectrometry (MS) methods for antimicrobial resistance protein detection, given the ability of these methods to confirm protein expression. In this work, we studied the performance of a liquid chromatography, tandem MS multiple-reaction monitoring (LC-MS/MS MRM) method for the direct detection of the New Delhi metallo-lactamase (NDM) carbapenemase in clinical isolates. Using a genoproteomic approach, we selected three unique peptides (SLGNLGDADTEHYAASAR, AFGAAFPK, and ASMIVMSHS APDSR) specific to NDM that were efficiently ionized and spectrally well-defined. These three peptides were used to build an assay with turnaround time of 90 min. In a blind set, the assay detected 21/24 blaNDM-containing isolates and 76/76 isolates with negative results, corresponding to a sensitivity value of 87.5% (95% confidence interval [CI], 67.6% to 97.3%) and a specificity value of 100% (95% CI, 95.3% to 100%). One of the missed identifications was determined by protein fractionation to be due to low (0.1 fm/g) NDM protein expression (below the assay limit of detection). Parallel disk diffusion susceptibility testing demonstrated this isolate to be meropenem susceptible, consistent with low NDM expression. Total proteomic analysis of the other two missed identifications did not detect NDM peptides but detected other proteins expressed from the blaNDM-containing plasmids, confirming that the plasmids were not lost. The measurement of relative NDM concentrations over the entire isolate test set demonstrated variability spanning 4 orders of magnitude, further confirming the remarkable range that may be seen in levels of NDM expression. This report highlights the sensitivity of LC-MS/MS to variations in NDM protein expression, with implications for how this technology may be used.
KW - Mass spectrometry
KW - Multiple-reaction monitoring
KW - New Delhi metallo-lactamase
KW - Tryptic peptide
UR - http://www.scopus.com/inward/record.url?scp=85071382901&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85071382901&partnerID=8YFLogxK
U2 - 10.1128/AAC.00461-19
DO - 10.1128/AAC.00461-19
M3 - Article
C2 - 31307990
AN - SCOPUS:85071382901
SN - 0066-4804
VL - 63
JO - Antimicrobial agents and chemotherapy
JF - Antimicrobial agents and chemotherapy
IS - 9
M1 - e00461-19
ER -