Phenotypic and genotypic characterisation of Bacillus cereus isolates from Bangladeshi rice

We report the first phenotypic and genotypic characterisation of Bacillus cereus from Bangladeshi rice. Seven strains of B. cereus were isolated from two high-yielding rice varieties (BR5 and BRRI Dhan28) which are cultivated during different rice-growing seasons in Bangladesh. The strains were identified as B. cereus based on colonial and cellular (light microscopic) morphology and were confirmed using the API 50 CHB test system and 16S rDNA analysis. They could also be distinguished from each other based on their substrate-utilisation patterns using the API system. No psychrotrophic (cold-tolerant) isolates were obtained and all were mesophilic with a minimum growth temperature of 10 degrees C, but with an ability to grow at a higher rate than normal temperature (45 degrees C) for B. cereus. All strains had temperature optima between 35 and 45 degrees C, but two groups could be distinguished based on having either a narrow or a broad optimum range. The different strains could not be distinguished based on their phospholipid compositions including major fatty acyl chains which were typical for B. cereus. However, capillary-GC/MS analysis of trace fatty acyl components of the lipids has been used for the first time to group the isolates. Genotypic analysis of the rDNA region has also been used to distinguish the strains. Restriction fragment length polymorphism (RFLP) analysis of the spacer region between the 16S and 23S rRNA genes yielded two groups based on different base sequences, whereas sequence analysis of variable regions in the 16S rRNA gene gave four different groups by base differences at two positions. Despite their association with rice, none of the strains produced emetic toxin. The spore germination response of heat-activated spores of the Bangladeshi B. cereus strains was typical of the bacterium, but three different groups could be identified based on differences in germination rates. The use of this polyphasic approach to taxonomy has enabled all the B. cereus strains to be distinguished from each other.