Conclusions
It has been recognized that methanotroph-photoautotroph cocultures offer a highly promising biological platform for biogas conversion. Through the interspecies metabolic coupling of methane oxidation to oxygenic photosynthesis, the coculture can simultaneously convert both CH4 and CO2 into microbial biomass without external oxygen supply. However, one key obstacle in developing methanotroph-photoautotroph based biotechnology for biogas conversion is the lack of an effective tool for fast, accurate and frequent characterization of the coculture growth dynamics. In this work, based on the organism’s growth stoichiometry, the interspecies metabolic coupling and the total mass balance, we developed an E-C protocol to characterize the coculture. The E-C protocol provides not only accurate estimates of the individual biomass concentration within the coculture, but also the individual substrate consumption and product excretion rates of each organism. To the best of our knowledge, the developed E-C protocol is the first ever approach that could obtain individual substrate consumption and product excretion rates for methanotroph-photoautotroph or any other cocultures.
The accuracy of the developed E-C protocol was validated by cell counting approach using flow cytometry. In addition, by comparing the predicted total OD from the individual biomass concentration with the measured total OD, we showed that the E-C protocol provided better accuracy than the cell counting approach through statistical testing. It is worth noting that the developed E-C protocol only requires commonly used analytical equipment to provide quick and accurate characterization of the methanotroph-photoautotroph cocultures.
Finally, we showed that it is very important to use static cocultures with known concentration to validate the cell counting method, as the cell fixation protocol could result in severely skewed cell counting results. Currently, although cell counting with flow cytometry has become increasingly common in determining the individual biomass concentration in mixed culture or microbiome, very few publications have presented validation results on their cell counting approaches.