The Conwy River in North Wales, UK, is a large freshwater system that has been the subject of numerous studies over many years. Even so, the river’s microbiome is poorly understood. It is widely known that in this system, as with all lotic systems, dissolved organic matter (DOM) is the primary source of energy for these resident microorganisms. Therefore, understanding how these species utilize DOM can allow scientists to make better predictions concerning the river’s water quality.
However, there are two major scientific challenges that must be considered. Firstly, due to the anthropogenic inputs (from agriculture and wastewater treatment) in tandem with the variation in flow rate and weather events, make it inherently difficult to properly model this system. Secondly, the vast majority of bacteria and archaea cannot be cultured under typical in vitro conditions. Evidence also suggests that many microbial species in aquatic systems have evaded detection due to their ability to pass through ultra-small filters (<0.45 µm pore sizes), i.e. filterable microorganisms. The term filterable microorganism can refer to one of the following: (1) small-bodied cells (less than 0.1 µm3 volume), (2) shrunken cells (due to limited nutrients or senescence), and (3) large cells that squeeze through small filters (<0.45 µm pore sizes). Their exact role in freshwater systems remain largely unknown.
The purpose of this thesis was to uncover the taxonomic identity, overall function, and role in DOM cycling of filterable species residing in the Conwy River while also comparing them to the native lotic community (i.e. unfiltered population). We utilised 16S rRNA single amplicon sequencing and shotgun sequencing to conduct a phylogenetic analysis of both ultra-filtered (passed through a 0.22 µm pore size filter) and unfiltered river water to understand the phylogenies and relative phyla distributions as well as determining which clusters of orthologous groups (COGs) were present. The distribution of COGs of both microcosms were compared to other environments and bacterial genomes to (1) assess similarity, and (2) determine if organism complexity is related to environment (i.e. are more complex organisms found in nutrient rich environments, etc). Next, we examined how either microbial community utilised dissolved organic carbon (DOC) via multi-omics and 14C radio-isotope tracking in order to determine whether DOC influenced these populations or whether the residing species showed any particular preference to a DOC type.
The major findings indicated that, the dominant phyla (listed in decreasing abundance) in the whole community were Proteobacteria, Bacteriodetes, Actinobacteria, and Firmicutes. Whereas, the filtered community contained more Firmicutes than Bacteriodetes and Actinobacteria. We also we detected the presence of several candidate phyla, most notably “Candidatus Parcubacteria”. There were more COGs in the filtered community that fall under the functional categories of replication, recombination, repair, and cell wall/membrane/envelope biogenesis comparatively to the entire population. Clustering metagenomes against single genomes revealed that the filtered community’s COG distribution was closely related to COG distribution of organisms with limited/streamlined genomes. The filtered microbiome also metabolized DOC at a slower rate than the whole community and was confirmed to be a taxonomically unique subset within the greater system. Changes within each community were not influenced by the addition of DOC and neither system had a preference in DOC type.
Overall, the results obtained from this body of work demonstrate that the filtered microcosm was a unique population nestled within the general microbial community. They differ in taxonomic makeup and their usage of low-molecular weight DOC, suggested that they may have different functional roles in freshwater ecosystems. By exploring the complex microbiome of the Conwy River, researchers can gain a better understanding of water quality, ecosystem management, and the nature of filterable microorganisms.