VALIDATION OF DIFFERENTIAL DIAGNOSIS BETWEEN BACILLUS ANTHRACIS AND BACILLUS CEREUS BY MALDI TOF MASS SPECTROMETRY

Bacillus anthracis and Bacillus cereus belong to the Bacillaceae family, genus Bacillus, and is a high bacilli, Gram-positive aerobic or facultative aerobic, catalase +, sporulated that can easily develop and survive outside the host. Bacillus anthracis causes anthrax, common infectious disease for humans and several animal species. Bacillus cereus is found in food-borne toxins, septicemia, eye infection. Diagnosis and identification of two biological agents like Bacillus spp, can be achieved via classical bacteriological techniques, morphological and cultural characters which are significant in this respect. Accurate diagnosis can be achieved via serological techniques (ELISA), molecular biology (PCR, Real Time PCR) or mass spectrometry (MALDI TOF). The cultures of Bacillus spp studied (Bacillus anthracis, Bacillus cereus) were seeded via loop prick on the simple agar and blood agar, in Petri dishes in order to obtain isolated colonies. 16S ribosomal protein extraction was performed via the "Microorganisms Profiling" extraction trifluoroacetic acid (TFA) 80% procedure. Strains of Bacillus spp genus were analyzed with the mass spectrometer Microflex LT 20. Field of the mass in which identification was carried out was set between 1,000 and 14,000 daltons. The values of characteristic peaks strain of Bacillus anthracis IC 29 are in the range of 2100-9600 Daltons and Bacillus cereus strain IC 11549 in 2500-11250 Daltons. The characteristic peak of the species Bacillus anthracis is set in the range of 4833 Daltons. Peak" protein species signature " for the biological agent Bacillus cereus, has a value of 70807 Daltons. All mass spectar obtained were identified as belonging to the tested bacterial species, with results which were confirmed by classical bacteriological techniques. By comparing the test results of identification via the MALDI TOF technologie of studied strains, we can conclude that they are similar from a structure point of view of 16S ribosomal protein, thus demonstrating a degree of similarity. The Microflex LT 20 system allows quick and accurate identification of biological agents therefore being an important confirmation/validation mean for other microbiological diagnostic methods.