THE USE OF MALDI-TOF MASS SPECTROMETRY WHOLE CELL PROTEIN/PEPTIDE PROFILES FOR IDENTIFICATION OF AEROMONAS SPECIES
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| Year of publication | 2008 |
| Type | Conference abstract |
| MU Faculty or unit | |
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| Description | Matrix-Assisted Laser Desorption/Ionization - Time of Flight Mass Spectrometry (MALDI-MS) is a fast, procedurally simple and sensitive analytical technique for the determination of the molecular weight of biomolecules. In microbiology, this modern approach is nowadays widely used in chemotaxonomy offering protein/peptide profiles in a broad m/z range, which represent unique and highly reproducible fingerprints of bacteria. Proteins of cytoplasmatic membrane and cytoplasm make up 50% of bacterial cell dry weight and involve among 200 6 000 molecular species. They provide highly characteristic markers with great discriminatory power accessible by the analysis of intact bacterial cells. MALDI MS profiles can be effectively used for direct identification of microorganisms, rapid screening or selective monitoring of pathogens or their metabolic products and distinguishing drug-sensitive and drug resistant bacterial strains. Rapid screening in diagnostics is based on comparison of samples with a database of reference spectra of relevant microorganisms. The main advantage of MALDI MS approach is its rapidity (the analysis is carried out without any need of previous separation, pretreatment methods, filtering or cleavage of cells) and the possibility of bacteria identification on the genus, species and strain level. The genus Aeromonas includes opportunistic pathogens of invertebrates, cold- and warm-blooded animals including humans. The main sources of infection are freshwater environment as well as brackish, chlorinated and unchlorinated water and contaminated food. In humans, enterotoxigenic or enteroinvasive phenospecies may cause intestinal (acute gastroenteritis, chronic diarrhoea, ulcerative colitis), as well as extraintestinal (micronecrosis, cellulitis, bones, respiratory or urinary) infections. Classification is often complicated due to high percentage of nucleic acid sequence similarity and phenotypical identity of some Aeromonas species . The overall aim of our study was to develop a reliable protocol for Aeromonas MALDI MS profiling (culture conditions, sample preparation and MALDI-MS analysis) and to develop a cluster analysis software for classifying of Aeromonas species. Optimization process showed no influence of cultivation medium, prolonged cultivation and frozen sample storage on the resulting MALDI-MS profile. We obtained clear and rapid outcomes of 100 Aeromonas strains differentiation. MS data were processed with software developed at the Department of Informatics and also, in parallel, with a commercial BioTyper software (Bruker, Daltonics). Cluster analysis of Aeromonas strains showed the capability of MALDI-MS to differentiate and to classify strains at the species/sub-species level and possibility to distinguish closely related aeromonads, that are unidentified genotypically and by biotyping. We have also successfully applied this method to the identification of related and heterogenous Pseudomonas spp. yet. |
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