Development of a Screening Platform and Systems Biology Tools for Enhancing Integrated Biogas Valorization and Wastewater Remediation

Abstract

Anaerobic digestion (AD) ranks among the top contributors to anthropogenic greenhouse gas (GHG) emissions in the United States. AD is a prevalent technology at wastewater treatment facilities (WWTF’s) that breaks down complex organic wastes into smaller substituents to recycle water from various waste streams. AD produces both biogas, consisting predominantly of methane and carbon dioxide, and a eutrophic liquid effluent. While some WWTF’s separate and harness the methane for the cogeneration of heat and electricity, all the carbon dioxide and produced GHG’s from burning the methane are vented to the atmosphere. In addition, the nitrogen- and phosphorous-containing ions (namely ammonium and orthophosphates) must be removed from liquid effluent before ejecting it from the WWTF. Recent studies have found that methanotroph-photoautotroph cocultures (MPCs) can serve as waste-to-value biocatalysts for the simultaneous treatment of the biogas and liquid effluents. The coculture can valorize the untreated biogas (both the methane and carbon dioxide) and liquid effluent by creating biomass for aquafeed and single-cell protein, as well as generate high titers of bioplastic precursors like formate and acetate. The MPC can also exhibit symbiotic interactions that not only increase the uptake rates of biogas and eutrophic ions but also increase the production of valuable biomass. Given the novelty of the coculture of dual biogas and wastewater remediation, much of the current literature focuses on utilizing methanotrophic communities with eukaryotic photoautotrophs or finding a single methanotrophic and photoautotrophic species that exhibits high growth rates to determine the efficacy of an MPC. However, there is very little research that screens individual pairs of methanotrophs and photoautotrophs to study metabolic cross-linking or to culture MPC’s directly into AD liquid effluent to validate their application potential. This dissertation addresses these research gaps by developing two experimental reactor designs and one systems biology toolkit to more efficiently study methanotroph-photoautotroph cocultures for biogas and wastewater remediation. The first project involves screening neutrophilic methanotrophs, microalgae, and MPC’s on diluted mesophilic AD effluent from a WWTF and synthetic biogas to actualize their application potential. To do this, a screening station consisting of nine parallel fed-batch bioreactors was constructed that regulated six key abiotic growth factors, greatly decreasing the time needed for screening. In addition, a novel Gompertzian model was developed to assess the growth performance of each monoculture and MPC systematically and unbiasedly. Using the data collected from the screening experiments, the maximum growth rate, delay time, and biomass carrying capacity of each model was used to evaluate the performance of seven methanotrophs and five microalgae previously noted in literature for their potential for wastewater remediation. The top monocultures were combined into six MPC’s, and two MPC’s produced nearly double the stationary biomass titer of their respective monocultures. This elevated biomass, along with results from the modified Gompertz model, indicates the presence of symbiotic interactions within the MPC directly grown on waste products from a WWTF. The second and third projects act serve as a platform to experimentally and computationally evaluate the synergism of Methylotuvimicrobium buryatense 5GB1 and Arthrospira platensis. Experimentally, due to the need for both shear-sensitive operations for A. platensis and high mass transfer rates for 5GB1, an airlift internal loop reactor was developed and implemented to culture both strains. Computationally, a systems biology toolkit was constructed to assist in analyzing GEMs for the coculture. The toolbox was used to evaluate two Clostridium tyrobutyricum models to demonstrate its capabilities.