Enabling continuous co-culture bioprocesses (C3BIO) based on oscillating environmental conditions promoting genetic and/or metabolic requirements of individual species

Continuous Co-Culture Bioprocesses (C3BIO) is one the key enabling technologies for the next generation of bioprocesses, due to increased metabolic capabilities, efficiency and modularity. However, the effective implementation of co-cultures in bioreactors is impeded by growth imbalances, leading to an unstable population. Not only does each organism have a different growth rate, but in addition each organism can have a phenotypically heterogeneous population with distributed growth rates. We have addressed the latter problem based on a previously developed technique, i.e., the segregostat, that can force a single-strain culture to adopt a homogeneous oscillatory behaviour in terms of growth and gene expression by periodically changing the environmental conditions at a rate close to the phenotypic diversification rate of the targeted population.
We want to transpose those results to co-cultures by providing oscillatory behaviours to each organism, and use these oscillations to stabilise the population. To this end, the key aspect is to synchronise the growth for each strain, and generate a phase shift between the growth phase of the different strains. We considered two approaches for providing this phase shift. In a first case study, we used the natural difference in metabolic capabilities of wild-type E. coli and S. cerevisiae to stabilise their co-culture using oscillating feeding profiles. In a second case study, we will engineer each strain of a co-culture with specific gene circuits, the toggle-switch and the repressilator, providing phase-shifted oscillatory behaiours. These behaviours will be coupled to growth by the production of essential genes.
Altogether, this approach and our current results point towards a viable strategy for the effective implementation of continuous co-culture bioprocesses.