Lactate dehydrogenase A promotes communication between carbohydrate catabolism and virulence in Bacillus cereus.

The diarrheal potential of a Bacillus cereus strain is essentially dictated by the amount of secreted nonhemolytic enterotoxin (Nhe). Expression of genes encoding Nhe is regulated by several factors, including the metabolic state of the cells. To identify metabolic sensors that could promote communication between central metabolism and nhe expression, we compared four strains of the B. cereus group in terms of metabolic and nhe expression capacities. We performed growth performance measurements, metabolite analysis, and mRNA measurements of strains F4430/73, F4810/72, F837/76, and PA cultured under anoxic and fully oxic conditions. The results showed that expression levels of nhe and ldhA, which encodes lactate dehydrogenase A (LdhA), were correlated in both aerobically and anaerobically grown cells. We examined the role of LdhA in the F4430/73 strain by constructing an ldhA mutant. The ldhA mutation was more deleterious to anaerobically grown cells than to aerobically grown cells, causing growth limitation and strong deregulation of key fermentative genes. More importantly, the ldhA mutation downregulated enterotoxin gene expression under both anaerobiosis and aerobiosis, with a more pronounced effect under anaerobiosis. Therefore, LdhA was found to exert a major control on both fermentative growth and enterotoxin expression, and it is concluded that there is a direct link between fermentative metabolism and virulence in B. cereus. The data presented also provide evidence that LdhA-dependent regulation of enterotoxin gene expression is oxygen independent. This study is the first report to describe a role of a fermentative enzyme in virulence in B. cereus.

Fnr mediates carbohydrate-dependent regulation of catabolic and enterotoxin genes in Bacillus cereus F4430/73.

We had previously demonstrated that Fnr is required for fermentative growth and oxic production of hemolysin BL (Hbl) and non-hemolytic enterotoxin (Nhe) in the food-borne pathogen Bacillus cereus F4430/73. In the present work, the regulatory impact of Fnr on microaerobic growth and enterotoxin production in response to carbohydrates was examined using glucose, fructose, sucrose or a glucose-fructose mixture as carbon and energy sources. Growth parameters, byproduct spectra and transcription levels of catabolic and enterotoxin genes were analyzed in a strain lacking Fnr in comparison to the parental F4430/73 strain. The results showed that B. cereus prefers glucose to other carbohydrates for microaerobic growth, and that lacking of Fnr less strongly affected the respiro-fermentative catabolism of glucose than fructose and, to a lesser extent than sucrose. In addition, lacking of Fnr strongly decreased expression of hbl and nhe genes, leading to the absence of Hbl and low production of Nhe independently of the carbohydrate used as carbon source. We conclude that Fnr is an important element for carbon source regulation in B. cereus F4430/73 and that the regulation of enterotoxin gene expression in response to carbohydrates may represent one aspect of overall catabolite control mediated by Fnr.