Evaluation of a set of E. coli reporter strains as physiological tracer for estimating bioreactor hydrodynamic efficiency

A set of different green fluorescent protein (GFP) Escherichia coli reporter strains have been evaluated in mini- and stirred bioreactors operating in fed-batch mode with different degrees of perturbations in order to estimate their potential use as process-related stress biosensor. The mini-bioreactor platform comprises a set of parallel shake flasks operating in fed-batch mode. The advantage of this system is its high experimental throughput for the evaluation of the GFP synthesis capacity of our reporter strains. In the case of classical shake flask system, no significant evolution of GFP synthesis have been observed, considering the reduced microbial growth period allowed by the system, whereas in the case of fed-batch operated mini-bioreactors, evolution of GFP synthesis, as well as GFP distribution among the microbial population, has been observed for three preselected strains (prpoS, puspA and posmC::gfp). More interestingly, a binary mode of expression has been observed in the case of the cultures carried out with the reporter strains for which GFP synthesis is under the control of the rpoS promoter which is induced under carbon limitation conditions. However, the generation of controlled glucose perturbations is relatively limited in this system and, in a second step fully automated bioreactor with a sclae-down strategy has been used to correlate the response of a prpoS::gfp strains with extracellular glucose perturbations. In the case of the culture performed in perturbed bioreactor (glucose intermittent feeding or glucose addition at the level of the recycle loop of a two-compartment scale-down bioreactor), the slowdown of the GFP synthesis resulting in the observation of a binary repartition of GFP content among the microbial population, has been observed. This observation led to the conclusion that the prpoS::gfp can be used as a biosensor for the validation of a fed-batch profile in industrial-scale bioreactors.