Bordetella pertussis autoregulates pertussis toxin production through the metabolism of cysteine

Pertussis toxin (Ptx) expression and secretion in Bordetella pertussis are regulated by a two-component signal transduction system encoded by the bvg regulatory locus. However, it is not known whether the metabolic pathways a nd growth state of the bacterium influence synthesis and secretion of Ptx a nd other virulence factors. We have observed a reduction in the concentrati on of Ptx per optical density unit midway in fermentation. Studies were con ducted to identify possible factors causing this reduction and to develop c ulture conditions that optimize Ptx expression. Medium reconstitution exper iments demonstrated that spent medium and a fraction of this medium contain ing components with a molecular weight of <3,000 inhibited the production o f Ptx. A complete flux analysis of the intermediate metabolism of B. pertus sis revealed that the sulfur-containing amino acids methionine and cysteine and the organic acid pyruvate accumulated in the media. In fermentation, a large amount of internal sulfate (SO42-) was observed in early stage growt h, followed by a rapid decrease as the cells entered into logarithmic growt h. This loss was later followed by the accumulation of large quantities of SO42- into the media in late-stage fermentation. Release of SO42- into the media by the cells signaled the decoupling of cell growth and Ptx productio n. Under conditions that limited cysteine, a fivefold increase in Ptx produ ction was observed. Addition of barium chloride (BaCl,) to the culture furt her increased Ptx yield. Our results suggest that B. pertussis is capable o f autoregulating the activity of the bvg regulon through its metabolism of cysteine. Reduction of the amount of cysteine in the media results in prolo nged vir expression due to the absence of the negative inhibitor SO42-. The refore, the combined presence and metabolism of cysteine may be an importan t mechanism in the pathogenesis of B. pertussis.