Rapid Pyrosequencing and Fatty Acid Analysis for Characterization of Bacillus cereus Isolates from Food

In this study rapid pyrosequencing was used for identification and fatty acid analysis of enterotoxic and non-enterotoxic Bacillus cereus. Presumptive B. cereus strains were isolated from ready-to-eat foods on mannitol-egg yolk-polymyxin agar and the identification was made based on the characteristics of B. cereus sp. Pyrosequencing was carried out on amplicons derived from 3 different 16S gene regions of rDNA using commercial reagent kits. All 30 presumptive Bacillus cereus isolates were identified as B. cereus. At least two sequencing reads of 3 different 16S rDNR gene regions were specific for identification of individual B. cereus isolates. Among B. cereus isolates, 53.3 % were found to be enterotoxic (determined by BCET-RPLA immunoassay kit). Fatty acids produced by more than 50 % of the investigated B. cereus were assumed as typical for these bacteria. The analyzed B. cereus produced a total of 25 typical fatty acids. The strains were highly homogeneous with dominant C4:0, C14:0, C16:0, C16:1, C18:0, C18:1, C18:2 and C22:6n3 fatty acids. Enterotoxic B. cereus was differentiated from the non-enterotoxic by significantly lower amounts of C18:0 and the absence of C18:4 fatty acid. Non-enterotoxic B. cereus did not produce C21:0, C17:0, C17:1, C20:0 and C15:1 fatty acids. The observed differences of individual fatty acid amounts and similar composition of fatty acids within all investigated B. cereus strains allowed the characterization of these bacteria isolated from ready-to-eat foods.