Deciphering Multifactorial Resistance Phenotypes in Acinetobacter baumannii by Genomics and Targeted Label-free Proteomics
Resistance to β-lactams in Acinetobacter baumannii involves various mechanisms. To decipher them, whole genome sequencing (WGS) and real-time quantitative polymerase chain reaction (RT-qPCR) were complemented by mass spectrometry (MS) in selected reaction monitoring mode (SRM) in 39 clinical isolate...
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Published in | Molecular & cellular proteomics Vol. 17; no. 3; pp. 442 - 456 |
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Main Authors | , , , , , , , , , , |
Format | Journal Article Web Resource |
Language | English |
Published |
United States
Elsevier Inc
01.03.2018
American Society for Biochemistry and Molecular Biology American Society for Biochemistry and Molecular Biology Inc The American Society for Biochemistry and Molecular Biology |
Subjects | |
Online Access | Get full text |
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Summary: | Resistance to β-lactams in Acinetobacter baumannii involves various mechanisms. To decipher them, whole genome sequencing (WGS) and real-time quantitative polymerase chain reaction (RT-qPCR) were complemented by mass spectrometry (MS) in selected reaction monitoring mode (SRM) in 39 clinical isolates. The targeted label-free proteomic approach eNAbled, in one hour and using a single method, the quantitative detection of 16 proteins associated with antibiotic resistance: eight acquired β-lactamases (i.e. GES, NDM-1, OXA-23, OXA-24, OXA-58, PER, TEM-1, and VEB), two resident β-lactamases (i.e. ADC and OXA-51-like) and six components of the two major efflux systems (i.e. AdeABC and AdeIJK). Results were normalized using “bacterial quantotypic peptides,” i.e. peptide markers of the bacterial quantity, to obtain precise protein quantitation (on average 8.93% coefficient of variation for three biological replicates). This allowed to correlate the levels of resistance to β-lactam with those of the production of acquired as well as resident β-lactamases or of efflux systems. SRM detected enhanced ADC or OXA-51-like production and absence or increased efflux pump production. Precise protein quantitation was particularly valuable to detect resistance mechanisms mediated by regulated genes or by overexpression of chromosomal genes. CombiNAtion of WGS and MS, two orthogoNAl and complementary techniques, allows thereby interpretation of the resistance phenotypes at the molecular level. |
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Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 PMCID: PMC5836370 scopus-id:2-s2.0-85042634448 Present address: Anaquant, Villeurbanne, France. Author contributions: T.C., P.C., C.G.C., and J.P.C. conceived the SRM strategy and the research project. T.C., Y.C., C.Ba., and C.Be. developed the SRM methods. T.C. performed the SRM analysis and the de novo sequencing interpretation. E.J.Y. performed the RTqPCR experiments, determined the MICs and provided sequencing interpretations. C.G.C. and P.C. selected the A. baumannii strains. X.L. synthetized the peptides used during SRM method development. J.L., C.G.C, P.C., and J.P.C. supervised the research. T.C., E.J.Y., J.D.D., C.G.C., and J.P.C. interpreted the results. T.C., E.J.Y., C.G.C., and J.P.C. wrote the manuscript. J.D.D., J.L., C.G.C., and P.C. edited the paper. Present address: Accelerate Diagnostics S.L., Barcelona, Spain. Present address: Yonsei University College of Medicine, Seoul, South Korea. Present address: Genomic Research Laboratory, Service of Infectious Diseases, Geneva University Hospitals, Geneva, Switzerland. |
ISSN: | 1535-9476 1535-9484 1535-9484 |
DOI: | 10.1074/mcp.RA117.000107 |