Effect of oxidative stress on antigen productivity in B. Pertussis cultures

Whooping cough, also known as pertussis, is an infectious respiratory disease caused by Bordetella pertussis, a type of gram-negative, aerobic, pathogenic bacteria with a coccobacillus shape and an outer capsule. The acellular vaccine for this disease contains a combination of pertussis toxin, fimbriae, filamentous hemagglutinin, and pertactin. The main constraint in the manufacturing of the vaccine relates to pertactin productivity due to its low abundance in the fermentation broth. Being the most abundant nutrient in the media, variability in initial glutamate concentration is hypothesized to be a major cause of process variability. This study proposes the combined use of cytometry and chromatography based separation to study the impact of glutamate-induced oxidative stress on the growth and productivity of pertactin antigen. Surface expression of pertactin was observed using fluorescence microscopy and flow cytometry while extracellular concentration was quantified using affinity chromatography. Oxidative stress levels and secretion of NADPH, a crucial reactant in antioxidant reactions, were monitored in flask and bioreactor experiments. The results established clear correlations between oxidative stress with growth and productivity and differentiate between the reduction in productivity to growth and to synthesis rate of pertactin. These findings are industrially relevant for improving productivity and reducing process variability.