Syntrophic acetate oxidation in the anaerobic digestion of nitrogen-rich wastes : from microbial interactions to process optimization

The anaerobic digestion (AD) consists of a cascade of syntrophic interactions between several microbial groups that result in the breakdown of biodegradable organic matter in the absence of oxygen, resulting in the production of biogas, a CH4 and CO2-rich mixture that can be valorised as renewable energy. The anaerobic digestion process has been engineered for a wide range of applications and current technology developments are aimed towards the so-called co-digestion, the treatment of complex mixtures of organic materials (e.g. slaughterhouse wastes) for an increased biogas yield. Nevertheless, the AD process might be hampered by the inhibitory effect of long chain fatty acids, ammonia and sulphate arising from the hydrolysis of proteins. The main objective of this thesis is to develop innovative techniques and methodologies to degrade protein. As described throughout the following sections, these compounds are potentially inhibitors of AD process, and consequently the production of CH4 and other value-added compounds. These wastes with high concentration of proteins emit high concentrations of NH3 when are degraded. It is reported that in this situation, the main population of methanogenic archaea are inhibited. However, the populations with the ability to carry out the hydrogenotrophic methanogenesis are active in this kind of environments. Therefore, increasing acetate oxidising bacteria populations (SAOB) is possible to achieve a syntrophy relation with some hydrogenotrophic archaea (HMA) populations, in this way is possible to avoid the inhibition caused by the NH3 concentration. Furthermore, sometimes these kind of wastes are associated with rich SO42- compounds that could be easily reduced to sulphide (H2S), molecule that is toxic, corrosive and odorous, producing several problems in the AD process and industrial facilities. The biological reaction that produced these compounds is performed by sulphate-reducing bacteria (SRB). These microbial communities, SRB, have the ability to couple the oxidation of organic matter to the reduction of SO42- outcompeting directly with methane producing archaea (MPA) and homoacetogenic bacteria for common substrates, and inhibiting the CH4 production. In order to carry out this doctoral thesis, two blocks (engineering and microbiological) have been developed that have been integrated as the work progressed. Firstly, different batch-scale studies were carried out in order to monitor the physical-chemical parameters and also the evolution of the microbial communities. In microbiology field, the main objective was to explore the evidence of acetate oxidation via syntrophic interaction between SAOB-HMA. To this end, the abundances of majorities groups were quantified using qPCR analysis based on the 16S rRNA and mcrA genes combined with isotopic compounds. Furthermore, the rapid advancement of high-throughput molecular techniques, as shotgun metagenomics sequencing and binning of scaffolds, have been allowed the implementation of genome-centric approaches that generate comprehensive databases from complex environmental samples, which includes complete or nearly-complete microbial genomes. Secondly, it was necessary that the reactors operated with the appropriate parameters to stablish this syntrophic interaction. As well as, it was also to establish protocols and different parameters concerning the configuration of the reactor based on literature and results obtained from the first set of experiments. Within this block, the hydraulic retention time (HRT), temperature ranges, organic loading rate, reactor configuration was treated. Finally, within this block, the most important point was to found the optimal reactor configuration. As it is known, the structure and the various units that make up a reactor, affect directly the efficiency and optimization of the same. Along this thesis different configurations was tested.