Typing Pseudomonas aeruginosa Isolates with Ultrahigh Resolution MALDI-FTICR Mass Spectrometry.
The introduction of standardized matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) platforms in the medical microbiological practice has revolutionized the way microbial species identification is performed on a daily basis. To a large extent, this is due to the ease of operation. Acquired spectra are compared to profiles obtained from cultured colonies present in a reference spectra database. It is fast and reliable, and costs are low compared to previous diagnostic approaches. However, the low resolution and dynamic range of the MALDI-TOF profiles have shown limited applicability for the discrimination of different bacterial strains, as achieved with typing based on genetic markers. This is pivotal in cases where certain strains are associated with, e.g., virulence or antibiotic resistance. Ultrahigh resolution MALDI-FTICR MS allows the measurement of small proteins at isotopic resolution and can be used to analyze complex mixtures with increased dynamic range and higher precision than MALDI-TOF MS, while still generating results in a similar time frame. Here, we propose to use ultrahigh resolution 15T MALDI-Fourier transform ion cyclotron resonance (FTICR) MS to discriminate clinically relevant bacterial strains after species identification performed by MALDI-TOF MS. We used a collection of well characterized Pseudomonas aeruginosa strains, featuring distinct antibiotic resistance profiles, and isolates obtained during hospital outbreaks. Following cluster analysis based on amplification fragment length polymorphism (AFLP), these strains were grouped into three different clusters. The same clusters were obtained using protein profiles generated by MALDI-FTICR MS. Subsequent intact protein analysis by electrospray ionization (ESI)-collision-induced dissociation (CID)-FTICR MS was applied to identify protein isoforms that contribute to the separation of the different clusters, illustrating the additional advantage of this analytical platform.
Department of Medical Microbiology, Leiden University Medical Center , 2333 ZA Leiden, The Netherlands.
Frank
F
Fleurbaaij
Department of Medical Microbiology, Leiden University Medical Center , 2333 ZA Leiden, The Netherlands.
Margriet E M
ME
Kraakman
Department of Medical Microbiology, Leiden University Medical Center , 2333 ZA Leiden, The Netherlands.
Eric C J
EC
Claas
Department of Medical Microbiology, Leiden University Medical Center , 2333 ZA Leiden, The Netherlands.
Cornelis W
CW
Knetsch
Department of Medical Microbiology, Leiden University Medical Center , 2333 ZA Leiden, The Netherlands.
Hans C
HC
van Leeuwen
Center for Proteomics and Metabolomics, Leiden University Medical Center , 2333 ZA Leiden, The Netherlands.
Yuri E M
YE
van der Burgt
Department of Medical Microbiology, Leiden University Medical Center , 2333 ZA Leiden, The Netherlands.
Karin Ellen
KE
Veldkamp
Department of Medical Microbiology and Infectious Disease, Erasmus MC , 3015 CN Rotterdam, The Netherlands.
Margreet C
MC
Vos
Department of Medical Microbiology and Infectious Disease, Erasmus MC , 3015 CN Rotterdam, The Netherlands.
Wil
W
Goessens
Department of Medical Statistics, Leiden University Medical Center , 2333 ZA Leiden, The Netherlands.
Bart J
BJ
Mertens
Department of Medical Microbiology, Leiden University Medical Center , 2333 ZA Leiden, The Netherlands.
Ed J
EJ
Kuijper
Center for Proteomics and Metabolomics, Leiden University Medical Center , 2333 ZA Leiden, The Netherlands.
Paul J
PJ
Hensbergen
Center for Proteomics and Metabolomics, Leiden University Medical Center , 2333 ZA Leiden, The Netherlands.
Simone
S
Nicolardi
D054458:Amplified Fragment Length Polymorphism Analysis; D001426:Bacterial Proteins / Q000737:chemistry / Q000302:isolation & purification; D016000:Cluster Analysis; D011550:Pseudomonas aeruginosa / Q000737:chemistry / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D001426:Bacterial Proteins