Rapid Characterization of Spores of Bacillus cereus Group Bacteria by Matrix-Assisted Laser Desorption-Ionization Time-of-Flight Mass Spectrometry

Abstract

Matrix-assisted laser desorption-ionization (MALDI) time-of-flight mass spectrometry was used to characterize the spores of 14 microorganisms of the Bacillus cereus group. This group includes the four Bacillus species B. anthracis, B. cereus, B. mycoides, and B. thuringiensis. MALDI mass spectra obtained from whole bacterial spores showed many similarities between the species, except for B. mycoides. At the same time, unique mass spectra could be obtained for the different B. cereus and B. thuringiensis strains, allowing for differentiation at the strain level. To increase the number of detectable biomarkers in the usually peak-poor MALDI spectra of spores, the spores were treated by corona plasma discharge (CPD) or sonicated prior to MALDI analysis. Spectra of sonicated or CPD-treated spores displayed an ensemble of biomarkers common for B. cereus group bacteria. Based on the spectra available, these biomarkers differentiate B. cereus group spores from those of Bacillus subtilis and Bacillus globigii. The effect of growth medium on MALDI spectra of spores was also explored.

MALDI-TOF-MS analysis of bacterial spores: Wet heat-treatment as a new releasing technique for biomarkers and the influence of different experimental parameters and microbiological handling

“…A standard deviation of 200 ppm was attained for all masses by So far, the detection of high-mass biomarkers directly from intact bacterial spores was only possible by treating the spores with sonication or corona plasma discharge (CPD). These techniques were introduced and successfully applied by the group of Fenselau [11,14]. These techniques have not been applied in our work.…”

“…Selected masses of similar strains measured with the CPD-technique or after acidic extraction are listed in Table 1 for comparison [12,14]. m/z Values found within a deviation of 500 ppm for different strains are given in bold.…”

“…The latest application uses bioactive glass slides to simplify the analysis of bacterial spores by specific surface absorption and lysing the spores with strong acids [16]. Most techniques so far applied damage the spore wall severely [11][12][13][14][15][16]. This enables the release of components from the coat or other interior compartments on the one hand but can cause difficulties in the subsequent analytical procedure on the other hand.…”

“…Subsequently, strong acid treatment (acidic extraction of high-mass biomarkers) was suggested by Elhanany et al [12]. Using this techniques, bacterial spores could be discriminated by means of their MALDI biomarker pattern at the strain and species level [13][14][15]. The latest application uses bioactive glass slides to simplify the analysis of bacterial spores by specific surface absorption and lysing the spores with strong acids [16].…”

Lipid fingerprinting of Bacillus spp. using online MALDI-TOF mass spectrometry

“…14,17,18 Previously, the identification of Bacillus spp. using MALDI-TOF MS was mainly based on a comparison of mass spectra of proteins, [19][20][21][22][23][24][25][26] lipopeptides, 27-29 and peptides. 30,31 The MALDI-TOF MS method relies on reproducible detection of microbial protein patterns, thus providing complementary information to genotypic or phenotypic test methods.…”