Unraveling the Microbial Response to Micro-Aerobic Conditions in Biological Nutrient Removal (BNR) Ecosystem

A relevant subject in the environmental engineering field is the development of energy-efficient wastewater treatment processes. In this context, decreasing aeration during biological nutrient removal (BNR) can lead to significant reductions in carbon footprint, energy usage and operational costs of wastewater treatment. The feasibility of BNR processes using lower oxygen concentrations than conventional treatment plants has been previously demonstrated. Nevertheless, a number of major knowledge gaps remain in our understanding of the BNR microbiome metabolic potential under limited-oxygen conditions. My doctoral research project sought to fill part of these gaps by identifying and studying mechanisms of adaptation to micro-aerobiosis in key members of the BNR microbiome. The proposed approach to study this microbial community combines reactor engineering, high-throughput sequencing and molecular and bioinformatics tools. The project started with the set-up of a lab-scale reactor operated under cyclic anaerobic and micro-aerobic conditions, which effectively removed nitrogen (N) and phosphorus (P) from synthetic wastewater. The operation and performance of this bioreactor, as well as an initial assessment of its microbial community are reported in the second chapter of this thesis. Then, I focused my investigation on unraveling the metabolic shape of the microbial functional groups responsible of metabolizing N and P in the reactor's microbiome. The third chapter contains an analysis of the genomic features characterizing the lifestyle of complete ammonia oxidizer (comammox) bacteria, a novel division of microorganisms capable of oxidizing ammonia to nitrate. The fourth chapter then centers on Ca. Accumulibacter phosphatis, a keystone microorganism responsible for P cycling and removal from wastewater. In both cases, I used data generated from whole genome sequencing to assemble genomes of these uncultured bacteria and to link genomic features and regulatory mechanisms to specific phenotypes. In addition, I used RNA sequencing data to investigate the effect of environmental conditions on the expression of biochemical pathways in Accumulibacter and other members of the microbiome. The information generated in these studies improved our understanding of how self-assembled microbial communities derive energy from substrates available in wastewater while saving energy requirements during wastewater treatment.