Identification and activity of bacteria consuming key intermediates of carbon and sulfur cycling in coastal sands

Coastal and shelf sediments are hot spots for carbon remineralization and also for carbon fixation. Here, a large fraction of organic carbon is mineralized under anoxic conditions by microorganisms via fermentation or respiration of fermentation products such as molecular hydrogen (H2) and acetate. Reduced inorganic metabolites released during these anaerobic processes and inorganic carbon are used by light-independent chemolithoautotrophs for socalled secondary production. However, still little is known about the in situ relevant organisms and how they contribute to key processes like chemoautotrophy as well as H2 and acetate turnover. To understand how inorganic carbon at sediment surfaces is turned over we surveyed the diversity of candidate bacterial chemolithoautotrophs in 13 tidal and sublittoral sediments and identified ubiquitous core groups of Gammaproteobacteria mainly affiliating with sulfuroxidizing bacteria. In a novel methodological approach we quantified dark carbon fixation by scintillography of specific microbial populations extracted and flow-sorted from sediments that were short term incubated with 14C-bicarbonate. Here, we show that uncultured Gammaproteobacteria dominate dark carbon fixation in coastal sediments and three distinct gammaproteobacterial clades made up more than half of dark carbon fixation in a tidal sediment. Meta- and single cell genomics along with metatranscriptomics provided evidence for a largely sulfur-based carbon fixation. These chemolithoautotrophic gammaproteobacterial clades also accounted for a substantial fraction of the microbial community in 1,000 to 2,000 year old subsurface sediments, suggesting that burial of chemolithoautotrophic bacteria could possibly be a yet-unrecognized mechanism of carbon sequestration. Microbial scavenging of H2 is an essential process in anoxic carbon mineralization, because only low H2 levels make H2-forming fermentation thermodynamically feasible. In a sediment metagenome we identified a high diversity of genes encoding the NiFe uptake hydrogenases of numerous yet-uncultured, potentially H2-oxidizing bacteria. Metatranscriptomics together with incubation experiments suggested uncultured Desulfobacteraceae, in particular the sulfate-reducing Sva0081-clade, as important H2 oxidizers in anoxic sediments. On the contrary, Gammaproteobacteria and Flavobacteria encoding O2-tolerant hydrogenases are possibly involved in H2 oxidation in oxic sediments. In a third study, we quantified the relative contribution of single bacterial populations to total acetate assimilation. Here, we showed that acetate was assimilated by physiologically and phylogenetically distinct bacterial groups such as Gammaproteobacteria, sulfate-reducing Desulfobacteraceae and Desulfobulbaceae as well as likely lithoheterotrophic sulfur-oxidizing Roseobacter-clade bacteria. We identified uncultured Gammaproteobacteria as a major contributor to acetate assimilation under oxic and anoxic conditions accounting for 31-62% of the total acetate assimilation. In summary, this thesis contributes to our understanding how distinct bacterial populations turn over key metabolites of organic carbon degradation in marine sediments. The quantification of uptake of 14C-labeleld model compounds by defined populations is a major step forward in the identification of key organisms in element cycling in marine sediments

Uncultured Gammaproteobacteria and Desulfobacteraceae Account for Major Acetate Assimilation in a Coastal Marine Sediment

Acetate is a key intermediate in anaerobic mineralization of organic matter in marine sediments. Its turnover is central to carbon cycling, however, the relative contribution of different microbial populations to acetate assimilation in marine sediments is unknown. To quantify acetate assimilation by in situ abundant bacterial populations, we incubated coastal marine sediments with 14C-labeled acetate and flow-sorted cells that had been labeled and identified by fluorescence in situ hybridization. Subsequently, scintillography determined the amount of 14C-acetate assimilated by distinct populations. This approach fostered a high-throughput quantification of acetate assimilation by phylogenetically identified populations. Acetate uptake was highest in the oxic-suboxic surface layer for all sorted bacterial populations, including deltaproteobacterial sulfate-reducing bacteria (SRB), which accounted for up to 32% of total bacterial acetate assimilation. We show that the family Desulfobulbaceae also assimilates acetate in marine sediments, while the more abundant Desulfobacteraceae dominated acetate assimilation despite lower uptake rates. Unexpectedly, members of Gammaproteobacteria accounted for the highest relative acetate assimilation in all sediment layers with up to 31–62% of total bacterial acetate uptake. We also show that acetate is used to build up storage compounds such as polyalkanoates. Together, our findings demonstrate that not only the usual suspects SRB but a diverse bacterial community may substantially contribute to acetate assimilation in marine sediments. This study highlights the importance of quantitative approaches to reveal the roles of distinct microbial populations in acetate turnover

Genus-specific carbon fixation activity measurements reveal distinct responses to oxygen among hydrothermal vent campylobacteria

Author Posting. © American Society for Microbiology, 2022. This article is posted here by permission of American Society for Microbiology for personal use, not for redistribution. The definitive version was published in Applied and Environmental Microbiology 88(2),(2022): e02083-21, https://doi.org/10.1128/AEM.02083-21.Molecular surveys of low temperature deep-sea hydrothermal vent fluids have shown that Campylobacteria (previously Epsilonproteobacteria) often dominate the microbial community and that three genera, Arcobacter, Sulfurimonas, and Sulfurovum, frequently coexist. In this study, we used replicated radiocarbon incubations of deep-sea hydrothermal fluids to investigate activity of each genus under three experimental conditions. To quantify genus-specific radiocarbon incorporation, we used newly designed oligonucleotide probes for Arcobacter, Sulfurimonas, and Sulfurovum to quantify their activity using catalyzed-reporter deposition fluorescence in situ hybridization (CARD-FISH) combined with fluorescence-activated cell sorting. All three genera actively fixed CO2 in short-term (∼ 20 h) incubations, but responded differently to the additions of nitrate and oxygen. Oxygen additions had the largest effect on community composition, and caused a pronounced shift in community composition at the amplicon sequence variant (ASV) level after only 20 h of incubation. The effect of oxygen on carbon fixation rates appeared to depend on the initial starting community. The presented results support the hypothesis that these chemoautotrophic genera possess functionally redundant core metabolic capabilities, but also reveal finer-scale differences in growth likely reflecting adaptation of physiologically-distinct phylotypes to varying oxygen concentrations in situ. Overall, our study provides new insights into how oxygen controls community composition and total chemoautotrophic activity, and underscores how quickly deep-sea vent microbial communities respond to disturbances.This research was funded by the U.S. National Science Foundation grants OCE-1131095 (S.M.S.) and OCE-1136727 (S.M.S., J.S.S.). Further support was provided by the WHOI Investment in Science Fund (S.M.S.). Funding for J.M. was further provided by doctoral fellowships from the Natural Sciences and Engineering Research Council of Canada (PGSD3-430487-2013, PGSM-405117-2011) and the National Aeronautics and Space Administration Earth Systems Science Fellowship (PLANET14F-0075), an award from the Canadian Meteorological and Oceanographic Society, and the WHOI Academic Programs Office

Identifizierung und Aktivität von Bakterien, die am Umsatz zentraler Intermediate aus dem Kohlenstoff- und Schwefelkreislauf in Küstensedimenten beteiligt sind.

Coastal and shelf sediments are hot spots for carbon remineralization and also for carbon fixation. Here, a large fraction of organic carbon is mineralized under anoxic conditions by microorganisms via fermentation or respiration of fermentation products such as molecular hydrogen (H2) and acetate. Reduced inorganic metabolites released during these anaerobic processes and inorganic carbon are used by light-independent chemolithoautotrophs for socalled secondary production. However, still little is known about the in situ relevant organisms and how they contribute to key processes like chemoautotrophy as well as H2 and acetate turnover. To understand how inorganic carbon at sediment surfaces is turned over we surveyed the diversity of candidate bacterial chemolithoautotrophs in 13 tidal and sublittoral sediments and identified ubiquitous core groups of Gammaproteobacteria mainly affiliating with sulfuroxidizing bacteria. In a novel methodological approach we quantified dark carbon fixation by scintillography of specific microbial populations extracted and flow-sorted from sediments that were short term incubated with 14C-bicarbonate. Here, we show that uncultured Gammaproteobacteria dominate dark carbon fixation in coastal sediments and three distinct gammaproteobacterial clades made up more than half of dark carbon fixation in a tidal sediment. Meta- and single cell genomics along with metatranscriptomics provided evidence for a largely sulfur-based carbon fixation. These chemolithoautotrophic gammaproteobacterial clades also accounted for a substantial fraction of the microbial community in 1,000 to 2,000 year old subsurface sediments, suggesting that burial of chemolithoautotrophic bacteria could possibly be a yet-unrecognized mechanism of carbon sequestration. Microbial scavenging of H2 is an essential process in anoxic carbon mineralization, because only low H2 levels make H2-forming fermentation thermodynamically feasible. In a sediment metagenome we identified a high diversity of genes encoding the NiFe uptake hydrogenases of numerous yet-uncultured, potentially H2-oxidizing bacteria. Metatranscriptomics together with incubation experiments suggested uncultured Desulfobacteraceae, in particular the sulfate-reducing Sva0081-clade, as important H2 oxidizers in anoxic sediments. On the contrary, Gammaproteobacteria and Flavobacteria encoding O2-tolerant hydrogenases are possibly involved in H2 oxidation in oxic sediments. In a third study, we quantified the relative contribution of single bacterial populations to total acetate assimilation. Here, we showed that acetate was assimilated by physiologically and phylogenetically distinct bacterial groups such as Gammaproteobacteria, sulfate-reducing Desulfobacteraceae and Desulfobulbaceae as well as likely lithoheterotrophic sulfur-oxidizing Roseobacter-clade bacteria. We identified uncultured Gammaproteobacteria as a major contributor to acetate assimilation under oxic and anoxic conditions accounting for 31-62% of the total acetate assimilation. In summary, this thesis contributes to our understanding how distinct bacterial populations turn over key metabolites of organic carbon degradation in marine sediments. The quantification of uptake of 14C-labeleld model compounds by defined populations is a major step forward in the identification of key organisms in element cycling in marine sediments

Oxygen respiration and polysaccharide degradation by a sulfate-reducing acidobacterium

Abstract Sulfate-reducing microorganisms represent a globally important link between the sulfur and carbon cycles. Recent metagenomic surveys expanded the diversity of microorganisms putatively involved in sulfate reduction underscoring our incomplete understanding of this functional guild. Here, we use genome-centric metatranscriptomics to study the energy metabolism of Acidobacteriota that carry genes for dissimilation of sulfur compounds in a long-term continuous culture running under alternating anoxic and oxic conditions. Differential gene expression analysis reveals the unique metabolic flexibility of a pectin-degrading acidobacterium to switch from sulfate to oxygen reduction when shifting from anoxic to oxic conditions. The combination of facultative anaerobiosis and polysaccharide degradation expands the metabolic versatility among sulfate-reducing microorganisms. Our results highlight that sulfate reduction and aerobic respiration are not mutually exclusive in the same organism, sulfate reducers can mineralize organic polymers, and anaerobic mineralization of complex organic matter is not necessarily a multi-step process involving different microbial guilds but can be bypassed by a single microbial species

Ubiquitous Gammaproteobacteria dominate dark carbon fixation in coastal sediments

Marine sediments are the largest carbon sink on earth. Nearly half of dark carbon fixation in the oceans occurs in coastal sediments, but the microorganisms responsible are largely unknown. By integrating the 16S rRNA approach, single-cell genomics, metagenomics and transcriptomics with (14)C-carbon assimilation experiments, we show that uncultured Gammaproteobacteria account for 70-86% of dark carbon fixation in coastal sediments. First, we surveyed the bacterial 16S rRNA gene diversity of 13 tidal and sublittoral sediments across Europe and Australia to identify ubiquitous core groups of Gammaproteobacteria mainly affiliating with sulfur-oxidizing bacteria. These also accounted for a substantial fraction of the microbial community in anoxic, 490-cm-deep subsurface sediments. We then quantified dark carbon fixation by scintillography of specific microbial populations extracted and flow-sorted from sediments that were short-term incubated with (14)C-bicarbonate. We identified three distinct gammaproteobacterial clades covering diversity ranges on family to order level (the Acidiferrobacter, JTB255 and SSr clades) that made up >50% of dark carbon fixation in a tidal sediment. Consistent with these activity measurements, environmental transcripts of sulfur oxidation and carbon fixation genes mainly affiliated with those of sulfur-oxidizing Gammaproteobacteria. The co-localization of key genes of sulfur and hydrogen oxidation pathways and their expression in genomes of uncultured Gammaproteobacteria illustrates an unknown metabolic plasticity for sulfur oxidizers in marine sediments. Given their global distribution and high abundance, we propose that a stable assemblage of metabolically flexible Gammaproteobacteria drives important parts of marine carbon and sulfur cycles

Uncultured Gammaproteobacteria and Desulfobacteraceae Account for Major Acetate Assimilation in a Coastal Marine Sediment

Acetate is a key intermediate in anaerobic mineralization of organic matter in marine sediments. Its turnover is central to carbon cycling, however, the relative contribution of different microbial populations to acetate assimilation in marine sediments is unknown. To quantify acetate assimilation by in situ abundant bacterial populations, we incubated coastal marine sediments with 14C-labeled acetate and flow-sorted cells that had been labeled and identified by fluorescence in situ hybridization. Subsequently, scintillography determined the amount of 14C-acetate assimilated by distinct populations. This approach fostered a high-throughput quantification of acetate assimilation by phylogenetically identified populations. Acetate uptake was highest in the oxic-suboxic surface layer for all sorted bacterial populations, including deltaproteobacterial sulfate-reducing bacteria (SRB), which accounted for up to 32% of total bacterial acetate assimilation. We show that the family Desulfobulbaceae also assimilates acetate in marine sediments, while the more abundant Desulfobacteraceae dominated acetate assimilation despite lower uptake rates. Unexpectedly, members of Gammaproteobacteria accounted for the highest relative acetate assimilation in all sediment layers with up to 31–62% of total bacterial acetate uptake. We also show that acetate is used to build up storage compounds such as polyalkanoates. Together, our findings demonstrate that not only the usual suspects SRB but a diverse bacterial community may substantially contribute to acetate assimilation in marine sediments. This study highlights the importance of quantitative approaches to reveal the roles of distinct microbial populations in acetate turnover.© 2018 Dyksma, Lenk, Sawicka and Mußman