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The dilution hypothesis was tested by adding different concentrations of ambient DOC obtained by solid phase extraction to deep seawater samples. Microbial growth and consumption of DOC were assessed by flow cytometry, HTCO measurements of DOC and oxygen consumption measurements in 14 experiments using water collected from deep water masses of the Atlantic and Pacific Oceans.There are two kinds of experiments 14 (A-N) where prokaryotic growth was evaluated under increasing concentrations of ambient DOC and 2 additional experiments (O and P) where DOC composition and the utilization of different compounds was evaluated by means of Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS). A utilization index for each compound was derived from the FT-ICR-MS fingerprints, showing whether the relative signal for each compound remained stable (refractory or not used), decreased (was consumed) or increased (was produced). Detailed information on conditions and procedures can be found in the article. Enquiries can be sent to Jesús M. Arrieta at txetxu[at]mail.com., Access and reuse conditions: This database and its components are subject to a Creative Commons Attribution-Noncommercial-ShareAlike International licence 4.0., Experimental results on the hypothesis that deep-water DOC consists of many different, intrinsically labile compounds at concentrations too low to compensate for the metabolic costs associated to their utilization., This is a contribution to the MALASPINA Expedition 2010 project, funded by the CONSOLIDER-Ingenio 2010 program of the Spanish Ministry of Economy and Competitiveness (Ref. CSD2008-00077). J.M.A. was supported by a “Ramón y Cajal” research fellowship from the Spanish Ministry of Economy and Competitiveness. E.M. was supported by a fellowship from the JAE program of CSIC. G.J.H. and R.H. were supported by the Austrian Science Fund (FWF) projects: I486-B09 and P23234-B11 and by the European Research Council under the European Community’s Seventh Framework Programme (FP7/2007-2013) / ERC grant agreement No. 268595 (MEDEA project). We thank A. Dorsett for assistance with DOC analyses, participants in the Malaspina Expedition and the crews of the BIO Hespérides, and RV Pelagia and the personnel of the Marine Technology Unit of CSIC (UTM) for their invaluable support., Peer reviewed

© 2014 Mayol, Jiménez, Herndl, Duarte and Arrieta. Airborne transport of microbes may play a central role in microbial dispersal, the maintenance of diversity in aquatic systems and in meteorological processes such as cloud formation. Yet, there is almost no information about the abundance and fate of microbes over the oceans, which cover >70% of the Earth's surface and are the likely source and final destination of a large fraction of airborne microbes. We measured the abundance of microbes in the lower atmosphere over a transect covering 17° of latitude in the North Atlantic Ocean and derived estimates of air-sea exchange of microorganisms from meteorological data. The estimated load of microorganisms in the atmospheric boundary layer ranged between 6×104 and 1.6×107 microbes per m2 of ocean, indicating a very dynamic air-sea exchange with millions of microbes leaving and entering the ocean per m2 every day. Our results show that about 10% of the microbes detected in the boundary layer were still airborne 4 days later and that they could travel up to 11,000 km before they entered the ocean again. The size of the microbial pool hovering over the North Atlantic indicates that it could play a central role in the maintenance of microbial diversity in the surface ocean and contribute significantly to atmospheric processes., This is a contribution to the Malaspina Expedition 2010, funded by the INGENIO 2010 CONSOLIDER program (ref. CDS2008-00077) of the Spanish Ministry of Economy and Competitiveness, and projects from the Austrian Science Fund (FWF, I486-B09 and P23234-B11) and project MEDEA from the European Research Council under the European Community's Seventh Framework Programme (FP7/2007-2013, ERC grant agreement No. 268595). Eva Mayol and María A. Jiménez acknowledge the “Junta para la Ampliación de Estudios” program (JAE-predoc and JAE-doc contracts, respectively) from CSIC, supplied by the European Social Fund. Jesús M. Arrieta was supported by a “Ramón y Cajal” fellowship by the former Ministry of Science and Innovation of the Spanish Government, Peer Reviewed

Oceanic dissolved organic carbon (DOC) is the second largest reservoir of organic carbon in the biosphere. About 72% of the global DOC inventory is stored in deep oceanic layers for years to centuries, supporting the current view that it consists of materials resistant to microbial degradation. An alternative hypothesis is that deep-water DOC consists of many different, intrinsically labile compounds at concentrations too low to compensate for the metabolic costs associated to their utilization. Here, we present experimental evidence showing that low concentrations rather than recalcitrance preclude consumption of a substantial fraction of DOC, leading to slow microbial growth in the deep ocean. These findings demonstrate an alternative mechanism for the long-term storage of labile DOC in the deep ocean, which has been hitherto largely ignored., This is a contribution to the Malaspina 2010 Expedition project, funded by the CONSOLIDER-Ingenio 2010 program of the from the Spanish Ministry of Economy and Competitiveness (Ref. CSD2008-00077). J.M.A. was supported by a “Ramón y Cajal” research fellowship from the Spanish Ministry of Economy and Competitiveness. E.M. was supported by a fellowship from the Junta para la Ampliación de Estudios program of CSIC. G.J.H. and R.L.H. were supported by the Austrian Science Fund (FWF) projects I486-B09 and P23234-B11 and by the European Research Council (ERC) under the European Community’s Seventh Framework Programme (FP7/2007-2013)/ERC grant agreement 268595 (MEDEA project)., Peer Reviewed

12 pages, 4 figures, 2 tables, supplementary material https://doi.org/10.3389/fmicb.2020.00376.-- All genome data is available in the Joint Genome Institute Integrated Microbial Genome database, Bacterial candidate phylum PAUC34f was originally discovered in marine sponges and is widely considered to be composed of sponge symbionts. Here, we report 21 single amplified genomes (SAGs) of PAUC34f from a variety of environments, including the dark ocean, lake sediments, and a terrestrial aquifer. The diverse origins of the SAGs and the results of metagenome fragment recruitment suggest that some PAUC34f lineages represent relatively abundant, free-living cells in environments other than sponge microbiomes, including the deep ocean. Both phylogenetic and biogeographic patterns, as well as genome content analyses suggest that PAUC34f associations with hosts evolved independently multiple times, while free-living lineages of PAUC34f are distinct and relatively abundant in a wide range of environments, This work was funded by the United States National Science Foundation grants 1460861 (REU site at Bigelow Laboratory for Ocean Sciences), 1441717, 1335810, and 1232982 to RS, and the Simons Foundation (Life Sciences Project Award ID 510023) to RS. NR was supported by the Ministry of Science and Higher Education of Russia. GH was supported by the Austrian Science Fund (FWF) project ARTEMIS (P28781-B21) and the European Research Council under the European Community’s Seventh Framework Program (FP7/2007-2013)/ERC (Grant Agreement No. 268595). JG was supported by Spanish project RTI2018-101025-B-I00. TW and JJ were funded by the U.S. Department of Energy, Joint Genome Institute, a DOE Office of Science User Facility supported under Contract No. DE-AC02-05CH11231, With the funding support of the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2019-000928-S), of the Spanish Research Agency (AEI)

Microbes in the dark ocean are exposed to hydrostatic pressure increasing with depth. Activity rate measurements and biomass production of dark ocean microbes are, however, almost exclusively performed under atmospheric pressure conditions due to technical constraints of sampling equipment maintaining in situ pressure conditions. To evaluate the microbial activity under in situ hydrostatic pressure, we designed and thoroughly tested an in situ microbial incubator (ISMI). The ISMI allows autonomously collecting and incubating seawater at depth, injection of substrate and fixation of the samples after a preprogramed incubation time. The performance of the ISMI was tested in a high-pressure tank and in several field campaigns under ambient hydrostatic pressure by measuring prokaryotic bulk 3H-leucine incorporation rates. Overall, prokaryotic leucine incorporation rates were lower at in situ pressure conditions than under to depressurized conditions reaching only about 50% of the heterotrophic microbial activity measured under depressurized conditions in bathypelagic waters in the North Atlantic Ocean off the northwestern Iberian Peninsula. Our results show that the ISMI is a valuable tool to reliably determine the metabolic activity of deep-sea microbes at in situ hydrostatic pressure conditions. Hence, we advocate that deep-sea biogeochemical and microbial rate measurements should be performed under in situ pressure conditions to obtain a more realistic view on deep-sea biotic processes., This study was supported by JSPS KAKENHI Grant (23651004) to M.U., the Austrian Science Fund (FWF) project I486-B09 and Z194 and the European Research Council under the European Community's Seventh Framework Program (FP7/2007-2013) / ERC grant agreement No. 268595 (MEDEA project) to G.J.H., FWF project P27696-B22 to E.S., FWF project P23221-B11 to T.R., and the Axencia Galega de Innovación (GAIN, Xunta de Galicia, Spain) through IEO-GAIN Programme Contracts (Contratos Programa) and GRC grant (INGO7A 2018/2) to Plankton ecology and biogeochemistry (“Ecologia Plantónica y Biogeoquímica”, EPB) research group (https://epb-research-group.mozello.es/). C.A. was supported by JSPS Postdoctoral Fellowships for Research Abroad (H26–168) and the European Union's Horizon 2020 research and innovation program under the Marie Sklodowska-Curie No. 701324. The comments of two reviewers are gratefully acknowledged and helped to improve an earlier version of the manuscript., Peer reviewed

21 pages, 6 figures, supplementary information https://doi.org/10.1038/s41564-023-01374-2.-- Data availability: The sequence data generated in this study have been deposited in the EMBL Nucleotide Sequence Database (ENA) database under Bioproject PRJEB35712 (metagenomic and metatranscriptomic raw reads, metagenomic and metatranscriptomic assemblies, metagenomic assembled genomes and single-cell amplified genomes) and in the NCBI Sequence Read Archive (SRA) under Bioproject PRJNA593264 (16S rRNA amplicon reads).-- Code availability: Scripts available at Zenodo (https://doi.org/10.5281/zenodo.7721930.2023), The deep ocean (>200 m depth) is the largest habitat on Earth. Recent evidence suggests sulfur oxidation could be a major energy source for deep ocean microbes. However, the global relevance and the identity of the major players in sulfur oxidation in the oxygenated deep-water column remain elusive. Here we combined single-cell genomics, community metagenomics, metatranscriptomics and single-cell activity measurements on samples collected beneath the Ross Ice Shelf in Antarctica to characterize a ubiquitous mixotrophic bacterial group (UBA868) that dominates expression of RuBisCO genes and of key sulfur oxidation genes. Further analyses of the gene libraries from the ‘Tara Oceans’ and ‘Malaspina’ expeditions confirmed the ubiquitous distribution and global relevance of this enigmatic group in the expression of sulfur oxidation and dissolved inorganic carbon fixation genes across the global mesopelagic ocean. Our study also underscores the unrecognized importance of mixotrophic microbes in the biogeochemical cycles of the deep ocean, This research was facilitated by the New Zealand Antarctic Research Institute (NZARI)-funded Aotearoa New Zealand Ross Ice Shelf Program. Samples for MICRO-CARD-FISH were collected on several research cruises led by M. Simon (Sonne 248 cruise), B. Quéguiner and I. Obernosterer (MobyDick) and L. J. A. Gerringa (Geotraces-1). F.B. was supported by the Austrian Science Fund (FWF) projects OCEANIDES (P34304-B), ENIGMA (TAI534), EXEBIO (P35248) and OCEANBIOPLAST (P35619-B). G.J.H. was supported by the FWF project ARTEMIS (P28781-B21) and I486-B09 and by the ERC under the European Community’s 7th Framework Programme (FP7/2007-2013)/ERC grant agreement no. 268595 (MEDEA project). R.L. was supported by INTERACTOMICS, CTM2015-69936-P, and J.M.G. by project PID2019-110011RB-C32 (Spanish Ministry of Science and Innovation, Spanish State Research Agency, doi: 10.13039/501100011033), With the institutional support of the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2019-000928-S), Peer reviewed

Deep-sea microbial communities are exposed to high-pressure conditions, which has a variable impact on prokaryotes depending on whether they are piezophilic (that is, pressure-loving), piezotolerant or piezosensitive. While it has been suggested that elevated pressures lead to higher community-level metabolic rates, the response of these deep-sea microbial communities to the high-pressure conditions of the deep sea is poorly understood. Based on microbial activity measurements in the major oceanic basins using an in situ microbial incubator, we show that the bulk heterotrophic activity of prokaryotic communities becomes increasingly inhibited at higher hydrostatic pressure. At 4,000 m depth, the bulk heterotrophic prokaryotic activity under in situ hydrostatic pressure was about one-third of that measured in the same community at atmospheric pressure conditions. In the bathypelagic zone-between 1,000 and 4,000 m depth-~85% of the prokaryotic community was piezotolerant and ~5% of the prokaryotic community was piezophilic. Despite piezosensitive-like prokaryotes comprising only ~10% (mainly members of Bacteroidetes, Alteromonas) of the deep-sea prokaryotic community, the more than 100-fold metabolic activity increase of these piezosensitive prokaryotes upon depressurization leads to high apparent bulk metabolic activity. Overall, the heterotrophic prokaryotic activity in the deep sea is likely to be substantially lower than hitherto assumed, with major impacts on the oceanic carbon cycling., Field experiments were conducted during the research cruises: MODUPLAN (CTM-2011-24008), RADPROF201508, RADPROF201808, RADCAN201808, SO248 (BacGeoPac), M139 (MerMet 17-97), MOBYDICK and POSEIDON. This study was supported by JSPS KAKENHI Grant (23651004) to M.U., the Austrian Science Fund (FWF) project I486-B09, Z194, P28781-B21 and P35587-B to G.J.H., P27696-B22 to E.S. and P23221-B11 to T.R., and the European Research Council under the European Community’s Seventh Framework Program (FP7/2007-2013)/ERC grant agreement (MEDEA project 268595) to G.J.H. C.A. was supported by JSPS Postdoctoral Fellowships for Research Abroad (H26–168), the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie no. 701324 and ERC Advanced Grant (TACKLE project, 695192)., Peer reviewed

Bulk dark dissolved inorganic carbon (DIC) fixation rates were determined and compared to microbial heterotrophic production in subsurface, meso- and bathypelagic Atlantic waters off the Galician coast (NW Iberian margin). DIC fixation rates were slightly higher than heterotrophic production throughout the water column, however, more prominently in the bathypelagic waters. Microautoradiography combined with catalyzed reporter deposition fluorescence in situ hybridization (MICRO-CARD-FISH) allowed us to identify several microbial groups involved in dark DIC uptake. The contribution of SAR406 (Marinimicrobia), SAR324 (Deltaproteobacteria) and Alteromonas (Gammaproteobacteria) to the dark DIC fixation was significantly higher than that of SAR202 (Chloroflexi) and Thaumarchaeota, in agreement with their contribution to microbial abundance. Q-PCR on the gene encoding for the ammonia monooxygenase subunit A (amoA) from the putatively high versus low ammonia concentration ecotypes revealed their depth-stratified distribution pattern. Taken together, our results indicate that chemoautotrophy is widespread among microbes in the dark ocean, particularly in bathypelagic waters. This chemolithoautotrophic biomass production in the dark ocean, depleted in bio-available organic matter, might play a substantial role in sustaining the dark ocean's food web., Funding for the sampling and analyses was provided by the projects ‘Biodiversidade Funcional do Microplancton nas profundidades mariñas de Galicia’ (BIO-PROF, Ref. 10MMA604024PR, 2010–2012, Xunta de Galicia) and ‘Fuentes de Materia Orgánica y Diversidad Funcional del Microplancton en las aguas profundas del Atlántico Norte’ (MODUPLAN, Plan Nacional I + D+I) to MMV. Additional funding was provided by the Axencia Galega de Innovación (GAIN, Xunta de Galicia) through IEO-GAIN Programme Contracts (Contratos Programa). EGF was supported by the BIO-PROF and MODUPLAN projects. ES received funding by the Austrian Science Fund (FWF) project P27696-B22 and GJH was funded by the Austrian Science Fund (FWF) projects I486-B09 and Z194-B17 and the European Research Council under the European Community's Seventh Framework Program (FP7/2007–2013)/ERC grant agreement No. 268595 (MEDEA project). This work is in partial fulfilment of the requirements for a Ph.D. degree from the Universidade de A Coruña by E.G.-F., Peer reviewed

Background: Heterotrophic microbes inhabiting the dark ocean largely depend on the settling of organic matter from the sunlit ocean. However, this sinking of organic materials is insufficient to cover their demand for energy and alternative sources such as chemoautotrophy have been proposed. Reduced sulfur compounds, such as thiosulfate, are a potential energy source for both auto- and heterotrophic marine prokaryotes. Methods: Seawater samples were collected from Labrador Sea Water (LSW, ~ 2000 m depth) in the North Atlantic and incubated in the dark at in situ temperature unamended, amended with 1 µM thiosulfate, or with 1 µM thiosulfate plus 10 µM glucose and 10 µM acetate (thiosulfate plus dissolved organic matter, DOM). Inorganic carbon fixation was measured in the different treatments and samples for metatranscriptomic analyses were collected after 1 h and 72 h of incubation. Results: Amendment of LSW with thiosulfate and thiosulfate plus DOM enhanced prokaryotic inorganic carbon fixation. The energy generated via chemoautotrophy and heterotrophy in the amended prokaryotic communities was used for the biosynthesis of glycogen and phospholipids as storage molecules. The addition of thiosulfate stimulated unclassified bacteria, sulfur-oxidizing Deltaproteobacteria (SAR324 cluster bacteria), Epsilonproteobacteria (Sulfurimonas sp.), and Gammaproteobacteria (SUP05 cluster bacteria), whereas, the amendment with thiosulfate plus DOM stimulated typically copiotrophic Gammaproteobacteria (closely related to Vibrio sp. and Pseudoalteromonas sp.). Conclusions: The gene expression pattern of thiosulfate utilizing microbes specifically of genes involved in energy production via sulfur oxidation and coupled to CO2 fixation pathways coincided with the change in the transcriptional profile of the heterotrophic prokaryotic community (genes involved in promoting energy storage), suggesting a fine-tuned metabolic interplay between chemoautotrophic and heterotrophic microbes in the dark ocean. [MediaObject not available: see fulltext.], Open Access funding provided thanks to the CRUE-CSIC agreement with Springer Nature. ES was supported by the Austrian Science Fund (FWF) project P27696-B22 and by the AEI project PID2020-118877 GB-I00. Shiptime and laboratory work were supported by the Austrian Science Fund (FWF) projects: I486-B09 and P23234-B11 and by the European Research Council under the European Community’s Seventh Framework Program (FP7/2007–2013)/ERC grant agreement No. 268595 (MEDEA project) to GJH. Laboratory work was also supported by NSF grants OCE-1232982, OCE-1335810, and DEB-1441717 to RS. DD was supported by the Deutsche Forschungsgemeinschaft (DFG) projects CO 2218/2–1 (PN: 445462226)., Peer reviewed