Typing Pseudomonas aeruginosa Isolates with Ultrahigh Resolution MALDI-FTICR Mass Spectrometry

Abstract

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.

Metabolic Phenotyping and Strain Characterisation of Pseudomonas aeruginosa Isolates from Cystic Fibrosis Patients Using Rapid Evaporative Ionisation Mass Spectrometry

“…recently showed that the ultrahigh resolution and increased dynamic range obtained with a Fourier transform ion cyclotron resonance detector allowed the classification of strains of P . aeruginosa in accordance with cluster analysis based on amplification fragment length polymorphism (AFLP), which was not achieved with a ToF detector 19 . Unfortunately, this technology remains excessively expensive and cumbersome, making it unsuitable for clinical implementation.…”

Characterization of microbial mixtures by mass spectrometry

“…Strategies for microorganism characterization by MALDI‐FTICR MS utilize the unsurpassed resolving power (on the order of 100 000 at m/z 1000) and mass accuracy better than 5 ppm . A 15T FTICR system with a MALDI source has been successfully used for discriminating antibiotic resistant and antibiotic susceptible P. aeruginosa strains including isolates from hospital outbreaks . Using ultrahigh mass resolving power mass spectra, the isolates have been clustered in three different groups coinciding with the clustering obtained from amplification fragment length polymorphism data.…”

A decade of understanding secondary metabolism in Pseudomonas spp. for sustainable agriculture and pharmaceutical applications

“…The complementary metabolomic approach utilized by microbial IMS enabled to visualize metabolic exchange within and among microbial species capturing unique chemical information from fluorescent pseudomonads in particular (Yang et al 2012). Typing of P. aeruginosa clinical isolates using MALDI-FTICR MS allowed the identification of distinct antibiotic resistance patterns in them (Fleurbaaij et al 2016). Inter-kingdom metabolic transformation between P. aeruginosa and Aspergillus fumigatus using MALDI-IMS revealed the characteristic antifungal phenazine compounds secreted by P. aeruginosa were converted by A. fumigatus with altered properties (Moree et al 2012).…”