A Microfluidic Platform for Droplet-Enabled Co-Cultivation of Microbial Communities

The majority of existing microbial species, in particular bacteria living in synergistic communities, have not been cultured in the laboratory with conventional pure-culture oriented methods. This severely limits the characterization and understanding of various microbial systems, including those closely related to the human health. To address this issue, we have developed a microfluidic co-cultivation platform to expand the repertoire of cultivable species from natural microbial communities and to characterize co-cultivated communities. We fabricated a microfluidic device that could readily encapsulate and co-cultivate subsets of a community, using highly parallel nano-liter aqueous droplets dispersed in a continuous oil phase. To demonstrate the effectiveness of this approach in discovering synergistic microbial interactions, a synthetic model system consisting of cross-feeding E. coli mutants was co-cultivated. Our device was able to detect pair-wise symbiotic relationship when one partner accounted for as low as 1% of the total population or each symbiont was about 3% of the artificial community.

Different microbial species have different level of oxygen tolerance and preference. To further enhance the likelihood of cultivating diverse species from a community, we have combined droplet co-cultivation and oxygen gradient to provide both the environment for microbial interactions and the optimal oxygen condition. Our prototype device is composed of two glass layers with fluid channels separated by a thin PDMS membrane. A linear oxygen gradient, covering the range of strictly anaerobic to fully aerobic conditions, is established and maintained via a tree-shaped channel mixing humidified nitrogen and air. The gradient is then transferred through the porous PDMS membrane to the chambers in the liquid channel incubating droplets. A murine fecal microbial sample, of which the bacteria lived with limited oxygen concentration in their native environment, was cultivated and different species were enriched in chambers featuring different oxygen conditions.