title: Elucidating the role of long-distance electron transfer by cable bacteria in freshwater sediments and contaminated aquifers
Creators: name: Sachs, Corinna
Publisher: name: University of Bayreuth
Year: 2022
Subjects: subject: freshwater cable bacteria
subject: agar pillar columns
subject: PacBio full-length 16S rRNA gene amplicon sequencing
subject: sediment microprofiling
subject: electrogenic sulphur oxidation (e-SOx)
subject: bioremediation
subject: microbial contaminant degradation
subject: stable isotope probing (SIP)
subject: 500
subject: 550
subject: 570

description: Contamination of freshwater resources by hydrocarbons is a worldwide problem, and leads to damage of ecosystems and diminishing of drinking water supplies. Biodegradation is a possible way to remove pollutants from the environment, and certain microbes are able to metabolize and efficiently break down contaminants. In anoxic sediment, anaerobic hydrocarbon degradation predominantly occurs at redox gradients, where microbes rely on alternative electron acceptors, other than oxygen, for biodegradation, but are subject to limited electron acceptor availability. Recently discovered cable bacteria (CB) can overcome such limitation by forming filaments for long-distance electron transfer (LDET). These filaments enable CB to perform oxygen-dependent, electrogenic sulphur oxidation (e-SOX), they spatially separate redox half reactions over cm-distances, and consequentially allow them to overcome diffusion limitation and redox stratification in aquatic sediments. However, their unique geochemical requirements render CB notoriously difficult to cultivate, complicating investigations on their ecology. It is the aim of this thesis to establish a viable cultivation approach for the enrichment of freshwater cable bacteria, to investigate the occurrence and/or diversity of cable bacteria, and to elucidate the role of LDET in biodegradation of toluene. In the first part of this doctoral thesis, I focus on a viable cultivation strategy for freshwater CB and introduce a new “agar pillar” approach to selectively enrich and investigate CB populations from aquatic sediments in laboratory columns. Within sediment columns, a central agar pillar is embedded, providing a sediment-free gradient system in equilibrium with surrounding sediment. Sediments from different freshwater systems (a streambed, a meromictic sulfidic lake, a polluted aquifer, river and lake sediments from mining-impacted Fichtelgebirge region) were incubated with the agar pillar approach. Microsensor profiling during column incubation revealed multiple characteristics of e-SOx, such as a suboxic zone between oxic surface and sulfidic sediments, as well as the establishment of electric potentials. Bacterial communities in agar pillars and surrounding sediment were analysed over depth by PacBio full-length 16S rRNA gene amplicon sequencing, allowing for a very accurate phylogenetic placement of detected populations. Putative CB affiliated with Desulfobulbaceae were detected in nearly all incubations and sediment types. Indeed, the selective niche of the agar pillar was preferentially colonized by CB from surface water sediments, but not so for groundwater sediments. While aquifer sediments hosted putative CB affiliated with Desulfurivibrio spp., CB clearly within the genus Ca. Electronema, representing several potentially novel species, prevailed in surface freshwater sediment columns. CB were seemingly linked to co-enriched fermenters, hinting at a possible role of e-SOx populations as an electron sink for heterotrophic microbes. Absence of CB resulted in similar microbiomes of agar pillar and surrounding sediment, suggesting agar pillar colonization by fermenters may indeed be linked to LDET. These findings add to the current understanding of the diversity and ecology of cable bacteria in freshwater systems, highlighting a differentiation of CB from distinct surface and groundwater sediments. In conclusion, the agar pillar provides a new cultivation strategy that may facilitate enrichment of redox gradient-dependent microorganisms, including previously unrecognized CB populations. In the second part of this thesis, I focus on the role of LDET in microbial toluene biodegradation. In a 13C-toluene microcosm incubation experiment with freshwater sediment, I followed toluene degradation with a combination of biogeochemical and molecular biology methods utilizing by DNA-stable isotope probing (DNA-SIP). A subset of my incubations included filter membranes at relevant depths to prevent CB growth. The results of this multipronged approach allowed to identify toluene degraders and compare contaminant degradation between the same sediment with and without a possible involvement of LDET by CB. Depth-resolved microprofiling showed signatures of LDET in microcosms without filters, while headspace analysis revealed significantly more toluene degradation in microcosms when no filters were present. Microbial community analysis suggested that predominantly members of Gammaproteobacteria, specifically Rhodocyclaceae, Comamonodaceae, Burkholderiaceae and Moraxellaceae, and Alphaproteobacteria, specifically, Beijerinckiaceae, degraded toluene. These microbial groups were more abundant in microcosms without filters as compared to microcosms including filters, hinting at a presumed indirect impact of LDET for contaminant degradation. Concluding, this thesis provides important new insights into the diversity and ecology of cable bacteria in freshwater systems, as well as a so far mostly unrecognised impact of LDET within sedimentary microbial communities. A possible function of CB-mediated e-SOx and LDET as an electron sink for heterotrophic microbes is considered relevant for potential future remediation endeavours of contaminated sediments. Previous strategies for contaminant removal by providing electron acceptors for anaerobic biodegradation are viable, but resource-intensive. Instead, the further exploration and harnessing of the potential of LDET by CB, intrinsically present within sedimentary systems, could offer an attractive alternative.