Protein and deoxyribonucleic acid heterogeneity in hemolysin BL, a tripartite enterotoxin produced by Bacillus cereus

Bacillus cereus is a ubiquitous soil organism that plays a major role in food spoilage and is a common cause of food poisoning. Many B. cereus strains produce the enterotoxin, hemolysin BL (HBL), which in prototype strain F837/76 is composed of three antigenically distinct proteins designated B, L₁, and L₂, with molecular weights of 37.8, 38.5, and 43.2 kDa, respectively. All three proteins must be present to produce biological activity.; In this research, physical and antigenic variation of the HBL components was shown to be common. Forty-eight percent of the strains tested produced HBL protein profiles distinct from F837/76. Heterogeneity was observed as absence of HBL component(s) or multiple proteins reacting to specific HBL antibodies. When an HBL component(s) was missing from the protein profile it was occasionally accounted for by lack of secretion but was more often due to lack of production. Multiple proteins suggested multiple genes.; Strain S1C produces two B proteins (38 and 42 kDa), two L₁ proteins (38 and 41 kDa), and two L₂ proteins (both 43 kDa). The higher molecular weight proteins were detected in the culture supernatant one hour after, but along with, the lower molecular weight proteins. N-terminal amino acid sequences were determined for the six HBL proteins from strain S1C. The sequences were 45–100% identical to prototype HBL proteins and 50–72% identical to each other. The combined data supported the possibility of two sets of hbl genes in strain S1C.; Low stringency polymerase chain reactions (PCR) and screening of S1C genomic deoxyribonucleic acid (DNA) cosmid libraries did not provide evidence of a second set of hbl genes. Bacillus cereus DNA toxicity to the Escherichia coli hosts used in these procedures may have hindered detection efforts. One set of hbl genes, amplified by PCR, was partially sequenced. The genes were 94% and 95% identical to those sequenced from F387/76 and ATCC 14579, respectively. Although this research was not able to validate the existence of a second set of genes, heterogeneity in HBL, observed as multiple proteins, has implications for HBL detection and the pathogenicity of B. cereus.