The genus Bacillus is immensely complex and is currently undergoing major taxonomic revisions based mainly upon 16S rDNA sequence analysis. Traditionally species are identified using morphological and biochemical criteria such as crystal formation on PEMBA agar and lecithinase production. However, these tests can be prolonged and may not conclusively allow species-level identification.

MALDI-TOF-MS is a rapid novel technique that allows the surface components of bacterial cells to be resolved as a characteristic mass spectrum. A small amount of growth from a culture plate is applied to a target plate and mixed with matrix solution. Irradiation of the sample with a nitrogen laser in a mass spectrometer produces a plume of ions that are determined using time of flight mass spectrometry.

Using a linear M@LDI instrument, (Waters Corporation, Manchester, UK), this technique was applied to National Collection of Type Culture (NCTC) strains representing Bacillus food pathogens frequently received for identification in the Food Safety Microbiology Laboratory (FSML), HPA, London. These included B. subtilis, B. cereus, B. megaterium, B. sphaericus, B. pumilus and B. lichenformis. Using cluster analysis MALDI-TOF-MS analysis grouped the data into species-specific clusters.

Subsequently a dedicated MALDI-TOF database (Manchester Metropolitan University, Manchester, UK) consisting of >2000 entries and representing nearly 90 strains of Bacillus was utilised in combination with database matching software (Waters Corporation, Manchester, UK) to analyse 80 well-characterised clinical isolates from FSML, HPA, London. All B. subtilis isolates were correctly identified and 75 and 79%B. cereus and B. lichenformis isolates, respectively were successfully identified with high percentage probability matches. Nine isolates of B. pumilus were incorrectly identified, but upon close inspection of the data biomarkers that enabled successful differentiation of B. pumilus from these organisms was located.

In conclusion, these preliminary findings suggest that MALDI-TOF-MS will represent a useful tool for the rapid identification of Bacillus isolates.