Comparative genomic analysis of nickel homeostasis in cable bacteria.

BACKGROUND: Cable bacteria are filamentous members of the Desulfobulbaceae family that are capable of performing centimetre­scale electron transport in marine and freshwater sediments. This long­distance electron transport is mediated by a network of parallel conductive fibres embedded in the cell envelope. This fibre network efficiently transports electrical currents along the entire length of the centimetre­long filament. Recent analyses show that these fibres consist of metalloproteins that harbour a novel nickel­containing cofactor, which indicates that cable bacteria have evolved a unique form of biological electron transport. This nickel­dependent conduction mechanism suggests that cable bacteria are strongly dependent on nickel as a biosynthetic resource. Here, we performed a comprehensive comparative genomic analysis of the genes linked to nickel homeostasis. We compared the genome­encoded adaptation to nickel of cable bacteria to related members of the Desulfobulbaceae family and other members of the Desulfobulbales order. RESULTS: Presently, four closed genomes are available for the monophyletic cable bacteria clade that consists of the genera Candidatus Electrothrix and Candidatus Electronema. To increase the phylogenomic coverage, we additionally generated two closed genomes of cable bacteria: Candidatus Electrothrix gigas strain HY10­6 and Candidatus Electrothrix antwerpensis strain GW3­4, which are the first closed genomes of their respective species. Nickel homeostasis genes were identified in a database of 38 cable bacteria genomes (including 6 closed genomes). Gene prevalence was compared to 19 genomes of related strains, residing within the Desulfobulbales order but outside of the cable bacteria clade, revealing several genome­encoded adaptations to nickel homeostasis in cable bacteria. Phylogenetic analysis indicates that nickel importers, nickel­binding enzymes and nickel chaperones of cable bacteria are affiliated to organisms outside the Desulfobulbaceae family, with several proteins showing affiliation to organisms outside of the Desulfobacterota phylum. Conspicuously, cable bacteria encode a unique periplasmic nickel export protein RcnA, which possesses a putative cytoplasmic histidine­rich loop that has been largely expanded compared to RcnA homologs in other organisms. CONCLUSION: Cable bacteria genomes show a clear genetic adaptation for nickel utilization when compared to closely related genera. This fully aligns with the nickel­dependent conduction mechanism that is uniquely found in cable bacteria.

Cable bacteria delay euxinia and modulate phosphorus release in coastal hypoxic systems.

Cable bacteria are long, filamentous bacteria with a unique metabolism involving centimetre-scale electron transport. They are widespread in the sediment of seasonally hypoxic systems and their metabolic activity stimulates the dissolution of iron sulfides (FeS), releasing large quantities of ferrous iron (Fe2+) into the pore water. Upon contact with oxygen, Fe2+ oxidation forms a layer of iron(oxyhydr)oxides (FeOx), which in its turn can oxidize free sulfide (H2S) and trap phosphorus (P) diffusing upward. The metabolism of cable bacteria could thus prevent the release of H2S from the sediment and reduce the risk of euxinia, while at the same time modulating P release over seasonal timescales. However, experimental support for this so-called 'iron firewall hypothesis' is scarce. Here, we collected natural sediment in a seasonally hypoxic basin in three different seasons. Undisturbed sediment cores were incubated under anoxic conditions and the effluxes of H2S, dissolved iron (dFe) and phosphate (PO4 3-) were monitored for up to 140 days. Cores with recent cable bacterial activity revealed a high stock of sedimentary FeOx, which delayed the efflux of H2S for up to 102 days. Our results demonstrate that the iron firewall mechanism could exert an important control on the prevalence of euxinia and regulate the P release in coastal oceans.

Missing Value Imputation of Wireless Sensor Data for Environmental Monitoring.

Over the past few years, the scale of sensor networks has greatly expanded. This generates extended spatiotemporal datasets, which form a crucial information resource in numerous fields, ranging from sports and healthcare to environmental science and surveillance. Unfortunately, these datasets often contain missing values due to systematic or inadvertent sensor misoperation. This incompleteness hampers the subsequent data analysis, yet addressing these missing observations forms a challenging problem. This is especially the case when both the temporal correlation of timestamps within a single sensor and the spatial correlation between sensors are important. Here, we apply and evaluate 12 imputation methods to complete the missing values in a dataset originating from large-scale environmental monitoring. As part of a large citizen science project, IoT-based microclimate sensors were deployed for six months in 4400 gardens across the region of Flanders, generating 15-min recordings of temperature and soil moisture. Methods based on spatial recovery as well as time-based imputation were evaluated, including Spline Interpolation, MissForest, MICE, MCMC, M-RNN, BRITS, and others. The performance of these imputation methods was evaluated for different proportions of missing data (ranging from 10% to 50%), as well as a realistic missing value scenario. Techniques leveraging the spatial features of the data tend to outperform the time-based methods, with matrix completion techniques providing the best performance. Our results therefore provide a tool to maximize the benefit from costly, large-scale environmental monitoring efforts.

Multi-wavelength Raman microscopy of nickel-based electron transport in cable bacteria.

Cable bacteria embed a network of conductive protein fibers in their cell envelope that efficiently guides electron transport over distances spanning up to several centimeters. This form of long-distance electron transport is unique in biology and is mediated by a metalloprotein with a sulfur-coordinated nickel (Ni) cofactor. However, the molecular structure of this cofactor remains presently unknown. Here, we applied multi-wavelength Raman microscopy to identify cell compounds linked to the unique cable bacterium physiology, combined with stable isotope labeling, and orientation-dependent and ultralow-frequency Raman microscopy to gain insight into the structure and organization of this novel Ni-cofactor. Raman spectra of native cable bacterium filaments reveal vibrational modes originating from cytochromes, polyphosphate granules, proteins, as well as the Ni-cofactor. After selective extraction of the conductive fiber network from the cell envelope, the Raman spectrum becomes simpler, and primarily retains vibrational modes associated with the Ni-cofactor. These Ni-cofactor modes exhibit intense Raman scattering as well as a strong orientation-dependent response. The signal intensity is particularly elevated when the polarization of incident laser light is parallel to the direction of the conductive fibers. This orientation dependence allows to selectively identify the modes that are associated with the Ni-cofactor. We identified 13 such modes, some of which display strong Raman signals across the entire range of applied wavelengths (405-1,064 nm). Assignment of vibrational modes, supported by stable isotope labeling, suggest that the structure of the Ni-cofactor shares a resemblance with that of nickel bis(1,2-dithiolene) complexes. Overall, our results indicate that cable bacteria have evolved a unique cofactor structure that does not resemble any of the known Ni-cofactors in biology.

Closing the genome of unculturable cable bacteria using a combined metagenomic assembly of long and short sequencing reads.

Many environmentally relevant micro-organisms cannot be cultured, and even with the latest metagenomic approaches, achieving complete genomes for specific target organisms of interest remains a challenge. Cable bacteria provide a prominent example of a microbial ecosystem engineer that is currently unculturable. They occur in low abundance in natural sediments, but due to their capability for long-distance electron transport, they exert a disproportionately large impact on the biogeochemistry of their environment. Current available genomes of marine cable bacteria are highly fragmented and incomplete, hampering the elucidation of their unique electrogenic physiology. Here, we present a metagenomic pipeline that combines Nanopore long-read and Illumina short-read shotgun sequencing. Starting from a clonal enrichment of a cable bacterium, we recovered a circular metagenome-assembled genome (5.09 Mbp in size), which represents a novel cable bacterium species with the proposed name Candidatus Electrothrix scaldis. The closed genome contains 1109 novel identified genes, including key metabolic enzymes not previously described in incomplete genomes of cable bacteria. We examined in detail the factors leading to genome closure. Foremost, native, non-amplified long reads are crucial to resolve the many repetitive regions within the genome of cable bacteria, and by analysing the whole metagenomic assembly, we found that low strain diversity is key for achieving genome closure. The insights and approaches presented here could help achieve genome closure for other keystone micro-organisms present in complex environmental samples at low abundance.

The organo-metal-like nature of long-range conduction in cable bacteria.

Cable bacteria are filamentous, multicellular microorganisms that display an exceptional form of biological electron transport across centimeter-scale distances. Currents are guided through a network of nickel-containing protein fibers within the cell envelope. Still, the mechanism of long-range conduction remains unresolved. Here, we characterize the conductance of the fiber network under dry and wet, physiologically relevant, conditions. Our data reveal that the fiber conductivity is high (median value: 27 S cm-1; range: 2 to 564 S cm-1), does not show any redox signature, has a low thermal activation energy (Ea = 69 ± 23 meV), and is not affected by humidity or the presence of ions. These features set the nickel-based conduction mechanism in cable bacteria apart from other known forms of biological electron transport. As such, conduction resembles that of an organic semi-metal with a high charge carrier density. Our observation that biochemistry can synthesize an organo-metal-like structure opens the way for novel bio-based electronic technologies.

A model analysis of centimeter-long electron transport in cable bacteria.

The recent discovery of cable bacteria has greatly expanded the known length scale of biological electron transport, as these multi-cellular bacteria are capable of mediating electrical currents across centimeter-scale distances. To enable such long-range conduction, cable bacteria embed a network of regularly spaced, parallel protein fibers in their cell envelope. These fibers exhibit extraordinary electrical properties for a biological material, including an electrical conductivity that can exceed 100 S cm-1. Traditionally, long-range electron transport through proteins is described as a multi-step hopping process, in which the individual hopping steps are described by Marcus electron transport theory. Here, we investigate to what extent such a classical hopping model can explain the conductance data recorded for individual cable bacterium filaments. To this end, the conductive fiber network in cable bacteria is modelled as a set of parallel one-dimensional hopping chains. Comparison of model simulated and experimental current(I)/voltage(V) curves, reveals that the charge transport is field-driven rather than concentration-driven, and there is no significant injection barrier between electrodes and filaments. However, the observed high conductivity levels (>100 S cm-1) can only be reproduced, if we include much longer hopping distances (a > 10 nm) and lower reorganisation energies (λ < 0.2 eV) than conventionally used in electron relay models of protein structures. Overall, our model analysis suggests that the conduction mechanism in cable bacteria is markedly distinct from other known forms of long-range biological electron transport, such as in multi-heme cytochromes.

Indications for a genetic basis for big bacteria and description of the giant cable bacterium Candidatus Electrothrix gigas sp. nov.

Bacterial cells can vary greatly in size, from a few hundred nanometers to hundreds of micrometers in diameter. Filamentous cable bacteria also display substantial size differences, with filament diameters ranging from 0.4 to 8 µm. We analyzed the genomes of cable bacterium filaments from 11 coastal environments of which the resulting 23 new genomes represent 10 novel species-level clades of Candidatus Electrothrix and two clades that putatively represent novel genus-level diversity. Fluorescence in situ hybridization with a species-level probe showed that large-sized cable bacteria belong to a novel species with the proposed name Ca. Electrothrix gigas. Comparative genome analysis suggests genes that play a role in the construction or functioning of large cable bacteria cells: the genomes of Ca. Electrothrix gigas encode a novel actin-like protein as well as a species-specific gene cluster encoding four putative pilin proteins and a putative type II secretion platform protein, which are not present in other cable bacteria. The novel actin-like protein was also found in a number of other giant bacteria, suggesting there could be a genetic basis for large cell size. This actin-like protein (denoted big bacteria protein, Bbp) may have a function analogous to other actin proteins in cell structure or intracellular transport. We contend that Bbp may help overcome the challenges of diffusion limitation and/or morphological complexity presented by the large cells of Ca. Electrothrix gigas and other giant bacteria. IMPORTANCE In this study, we substantially expand the known diversity of marine cable bacteria and describe cable bacteria with a large diameter as a novel species with the proposed name Candidatus Electrothrix gigas. In the genomes of this species, we identified a gene that encodes a novel actin-like protein [denoted big bacteria protein (Bbp)]. The bbp gene was also found in a number of other giant bacteria, predominantly affiliated to Desulfobacterota and Gammaproteobacteria, indicating that there may be a genetic basis for large cell size. Thus far, mostly structural adaptations of giant bacteria, vacuoles, and other inclusions or organelles have been observed, which are employed to overcome nutrient diffusion limitation in their environment. In analogy to other actin proteins, Bbp could fulfill a structural role in the cell or potentially facilitate intracellular transport.

Cable Bacteria Activity Modulates Arsenic Release From Sediments in a Seasonally Hypoxic Marine Basin.

Eutrophication and global change are increasing the occurrence of seasonal hypoxia (bottom-water oxygen concentration <63 µM) in coastal systems worldwide. In extreme cases, the bottom water can become completely anoxic, allowing sulfide to escape from the sediments and leading to the development of bottom-water euxinia. In seasonally hypoxic coastal basins, electrogenic sulfur oxidation by long, filamentous cable bacteria has been shown to stimulate the formation of an iron oxide layer near the sediment-water interface, while the bottom waters are oxygenated. Upon the development of bottom-water anoxia, this iron oxide "firewall" prevents the sedimentary release of sulfide. Iron oxides also act as an adsorption trap for elements such as arsenic. Arsenic is a toxic trace metal, and its release from sediments can have a negative impact on marine ecosystems. Yet, it is currently unknown how electrogenic sulfur oxidation impacts arsenic cycling in seasonally hypoxic basins. In this study, we presented results from a seasonal field study of an uncontaminated marine lake, complemented with a long-term sediment core incubation experiment, which reveals that cable bacteria have a strong impact on the arsenic cycle in a seasonally hypoxic system. Electrogenic sulfur oxidation significantly modulates the arsenic fluxes over a seasonal time scale by enriching arsenic in the iron oxide layer near the sediment-water interface in the oxic period and pulse-releasing arsenic during the anoxic period. Fluxes as large as 20 µmol m-2 day-1 were measured, which are comparable to As fluxes reported from highly contaminated sediments. Since cable bacteria are recognized as active components of the microbial community in seasonally hypoxic systems worldwide, this seasonal amplification of arsenic fluxes is likely a widespread phenomenon.

Polyphosphate Dynamics in Cable Bacteria.

Cable bacteria are multicellular sulfide oxidizing bacteria that display a unique metabolism based on long-distance electron transport. Cells in deeper sediment layers perform the sulfide oxidizing half-reaction whereas cells in the surface layers of the sediment perform the oxygen-reducing half-reaction. These half-reactions are coupled via electron transport through a conductive fiber network that runs along the shared cell envelope. Remarkably, only the sulfide oxidizing half-reaction is coupled to biosynthesis and growth whereas the oxygen reducing half-reaction serves to rapidly remove electrons from the conductive fiber network and is not coupled to energy generation and growth. Cells residing in the oxic zone are believed to (temporarily) rely on storage compounds of which polyphosphate (poly-P) is prominently present in cable bacteria. Here we investigate the role of poly-P in the metabolism of cable bacteria within the different redox environments. To this end, we combined nanoscale secondary ion mass spectrometry with dual-stable isotope probing (13C-DIC and 18O-H2O) to visualize the relationship between growth in the cytoplasm (13C-enrichment) and poly-P activity (18O-enrichment). We found that poly-P was synthesized in almost all cells, as indicated by 18O enrichment of poly-P granules. Hence, poly-P must have an important function in the metabolism of cable bacteria. Within the oxic zone of the sediment, where little growth is observed, 18O enrichment in poly-P granules was significantly lower than in the suboxic zone. Thus, both growth and poly-P metabolism appear to be correlated to the redox environment. However, the poly-P metabolism is not coupled to growth in cable bacteria, as many filaments from the suboxic zone showed poly-P activity but did not grow. We hypothesize that within the oxic zone, poly-P is used to protect the cells against oxidative stress and/or as a resource to support motility, while within the suboxic zone, poly-P is involved in the metabolic regulation before cells enter a non-growing stage.

Biogeochemical impacts of fish farming on coastal sediments: Insights into the functional role of cable bacteria.

Fish farming in sea cages is a growing component of the global food industry. A prominent ecosystem impact of this industry is the increase in the downward flux of organic matter, which stimulates anaerobic mineralization and sulfide production in underlying sediments. When free sulfide is released to the overlying water, this can have a toxic effect on local marine ecosystems. The microbially-mediated process of sulfide oxidation has the potential to be an important natural mitigation and prevention strategy that has not been studied in fish farm sediments. We examined the microbial community composition (DNA-based 16S rRNA gene) underneath two active fish farms on the Southwestern coast of Iceland and performed laboratory incubations of resident sediment. Field observations confirmed the strong geochemical impact of fish farming on the sediment (up to 150 m away from cages). Sulfide accumulation was evidenced under the cages congruent with a higher supply of degradable organic matter from the cages. Phylogenetically diverse microbes capable of sulfide detoxification were present in the field sediment as well as in lab incubations, including cable bacteria (Candidatus Electrothrix), which display a unique metabolism based on long-distance electron transport. Microsensor profiling revealed that the activity of cable bacteria did not exert a dominant impact on the geochemistry of fish farm sediment at the time of sampling. However, laboratory incubations that mimic the recovery process during fallowing, revealed successful enrichment of cable bacteria within weeks, with concomitant high sulfur-oxidizing activity. Overall our results give insight into the role of microbially-mediated sulfide detoxification in aquaculture impacted sediments.

Efficient long-range conduction in cable bacteria through nickel protein wires.

Filamentous cable bacteria display long-range electron transport, generating electrical currents over centimeter distances through a highly ordered network of fibers embedded in their cell envelope. The conductivity of these periplasmic wires is exceptionally high for a biological material, but their chemical structure and underlying electron transport mechanism remain unresolved. Here, we combine high-resolution microscopy, spectroscopy, and chemical imaging on individual cable bacterium filaments to demonstrate that the periplasmic wires consist of a conductive protein core surrounded by an insulating protein shell layer. The core proteins contain a sulfur-ligated nickel cofactor, and conductivity decreases when nickel is oxidized or selectively removed. The involvement of nickel as the active metal in biological conduction is remarkable, and suggests a hitherto unknown form of electron transport that enables efficient conduction in centimeter-long protein structures.

Enhanced Laterally Resolved ToF-SIMS and AFM Imaging of the Electrically Conductive Structures in Cable Bacteria.

Cable bacteria are electroactive bacteria that form a long, linear chain of ridged cylindrical cells. These filamentous bacteria conduct centimeter-scale long-range electron transport through parallel, interconnected conductive pathways of which the detailed chemical and electrical properties are still unclear. Here, we combine time-of-flight secondary-ion mass spectrometry (ToF-SIMS) and atomic force microscopy (AFM) to investigate the structure and composition of this naturally occurring electrical network. The enhanced lateral resolution achieved allows differentiation between the cell body and the cell-cell junctions that contain a conspicuous cartwheel structure. Three ToF-SIMS modes were compared in the study of so-called fiber sheaths (i.e., the cell material that remains after the removal of cytoplasm and membranes, and which embeds the electrical network). Among these, fast imaging delayed extraction (FI-DE) was found to balance lateral and mass resolution, thus yielding the following multiple benefits in the study of structure-composition relations in cable bacteria: (i) it enables the separate study of the cell body and cell-cell junctions; (ii) by combining FI-DE with in situ AFM, the depth of Ni-containing protein-key in the electrical transport-is determined with greater precision; and (iii) this combination prevents contamination, which is possible when using an ex situ AFM. Our results imply that the interconnects in extracted fiber sheaths are either damaged during extraction, or that their composition is different from fibers, or both. From a more general analytical perspective, the proposed methodology of ToF-SIMS in the FI-DE mode combined with in situ AFM holds great promise for studying the chemical structure of other biological systems.

Cell Cycle, Filament Growth and Synchronized Cell Division in Multicellular Cable Bacteria.

Cable bacteria are multicellular, Gram-negative filamentous bacteria that display a unique division of metabolic labor between cells. Cells in deeper sediment layers are oxidizing sulfide, while cells in the surface layers of the sediment are reducing oxygen. The electrical coupling of these two redox half reactions is ensured via long-distance electron transport through a network of conductive fibers that run in the shared cell envelope of the centimeter-long filament. Here we investigate how this unique electrogenic metabolism is linked to filament growth and cell division. Combining dual-label stable isotope probing (13C and 15N), nanoscale secondary ion mass spectrometry, fluorescence microscopy and genome analysis, we find that the cell cycle of cable bacteria cells is highly comparable to that of other, single-celled Gram-negative bacteria. However, the timing of cell growth and division appears to be tightly and uniquely controlled by long-distance electron transport, as cell division within an individual filament shows a remarkable synchronicity that extends over a millimeter length scale. To explain this, we propose the "oxygen pacemaker" model in which a filament only grows when performing long-distance transport, and the latter is only possible when a filament has access to oxygen so it can discharge electrons from its internal electrical network.

Combining citizen science and deep learning for large-scale estimation of outdoor nitrogen dioxide concentrations.

Reliable estimates of outdoor air pollution concentrations are needed to support global actions to improve public health. We developed a new approach to estimating annual average outdoor nitrogen dioxide (NO2) concentrations using approximately 20,000 ground-level measurements in Flanders, Belgium combined with aerial images and deep neural networks. Our final model explained 79% of the spatial variability in NO2 (root mean square error of 10-fold cross-validation = 3.58 µg/m3) using only images as model inputs. This novel approach offers an alternative means of estimating large-scale spatial variations in ambient air quality and may be particularly useful for regions of the world without detailed emissions data or land use information typically used to estimate outdoor air pollution concentrations.

Bistability in the redox chemistry of sediments and oceans.

For most of Earth's history, the ocean's interior was pervasively anoxic and showed occasional shifts in ocean redox chemistry between iron-buffered and sulfide-buffered states. These redox transitions are most often explained by large changes in external inputs, such as a strongly altered delivery of iron and sulfate to the ocean, or major shifts in marine productivity. Here, we propose that redox shifts can also arise from small perturbations that are amplified by nonlinear positive feedbacks within the internal iron and sulfur cycling of the ocean. Combining observational evidence with biogeochemical modeling, we show that both sedimentary and aquatic systems display intrinsic iron-sulfur bistability, which is tightly linked to the formation of reduced iron-sulfide minerals. The possibility of tipping points in the redox state of sediments and oceans, which allow large and nonreversible geochemical shifts to arise from relatively small changes in organic carbon input, has important implications for the interpretation of the geological rock record and the causes and consequences of major evolutionary transitions in the history of Earth's biosphere.

Intrinsic electrical properties of cable bacteria reveal an Arrhenius temperature dependence.

Filamentous cable bacteria exhibit long-range electron transport over centimetre-scale distances, which takes place in a parallel fibre structure with high electrical conductivity. Still, the underlying electron transport mechanism remains undisclosed. Here we determine the intrinsic electrical properties of the conductive fibres in cable bacteria from a material science perspective. Impedance spectroscopy provides an equivalent electrical circuit model, which demonstrates that dry cable bacteria filaments function as resistive biological wires. Temperature-dependent electrical characterization reveals that the conductivity can be described with an Arrhenius-type relation over a broad temperature range (- 195 °C to + 50 °C), demonstrating that charge transport is thermally activated with a low activation energy of 40-50 meV. Furthermore, when cable bacterium filaments are utilized as the channel in a field-effect transistor, they show n-type transport suggesting that electrons are the charge carriers. Electron mobility values are ~ 0.1 cm2/Vs at room temperature and display a similar Arrhenius temperature dependence as conductivity. Overall, our results demonstrate that the intrinsic electrical properties of the conductive fibres in cable bacteria are comparable to synthetic organic semiconductor materials, and so they offer promising perspectives for both fundamental studies of biological electron transport as well as applications in microbial electrochemical technologies and bioelectronics.

Using Large-Scale NO2 Data from Citizen Science for Air-Quality Compliance and Policy Support.

Citizen science projects that monitor air quality have recently drastically expanded in scale. Projects involving thousands of citizens generate spatially dense data sets using low-cost passive samplers for nitrogen dioxide (NO2), which complement data from the sparse reference network operated by environmental agencies. However, there is a critical bottleneck in using these citizen-derived data sets for air-quality policy. The monitoring effort typically lasts only a few weeks, while long-term air-quality guidelines are based on annual-averaged concentrations that are not affected by seasonal fluctuations in air quality. Here, we describe a statistical model approach to reliably transform passive sampler NO2 data from multiweek averages to annual-averaged values. The predictive model is trained with data from reference stations that are limited in number but provide full temporal coverage and is subsequently applied to the one-off data set recorded by the spatially extensive network of passive samplers. We verify the assumptions underlying the model procedure and demonstrate that model uncertainty complies with the EU-quality objectives for air-quality monitoring. Our approach allows a considerable cost optimization of passive sampler campaigns and removes a critical bottleneck for citizen-derived data to be used for compliance checking and air-quality policy use.

Quantification of Cable Bacteria in Marine Sediments via qPCR.

Cable bacteria (Deltaproteobacteria, Desulfobulbaceae) are long filamentous sulfur-oxidizing bacteria that generate long-distance electric currents running through the bacterial filaments. This way, they couple the oxidation of sulfide in deeper sediment layers to the reduction of oxygen or nitrate near the sediment-water interface. Cable bacteria are found in a wide range of aquatic sediments, but an accurate procedure to assess their abundance is lacking. We developed a qPCR approach that quantifies cable bacteria in relation to other bacteria within the family Desulfobulbaceae. Primer sets targeting cable bacteria, Desulfobulbaceae and the total bacterial community were applied in qPCR with DNA extracted from marine sediment incubations. Amplicon sequencing of the 16S rRNA gene V4 region confirmed that cable bacteria were accurately enumerated by qPCR, and suggested novel diversity of cable bacteria. The conjoint quantification of current densities and cell densities revealed that individual filaments carry a mean current of ∼110 pA and have a cell specific oxygen consumption rate of 69 fmol O2 cell-1 day-1. Overall, the qPCR method enables a better quantitative assessment of cable bacteria abundance, providing new metabolic insights at filament and cell level, and improving our understanding of the microbial ecology of electrogenic sediments.

A Cross-System Comparison of Dark Carbon Fixation in Coastal Sediments.

Dark carbon fixation (DCF) by chemoautotrophic microorganisms can sustain food webs in the seafloor by local production of organic matter independent of photosynthesis. The process has received considerable attention in deep sea systems, such as hydrothermal vents, but the regulation, depth distribution, and global importance of coastal sedimentary DCF have not been systematically investigated. Here we surveyed eight coastal sediments by means of stable isotope probing (13C-DIC) combined with bacterial biomarkers (phospholipid-derived fatty acids) and compiled additional rates from literature into a global database. DCF rates in coastal sediments range from 0.07 to 36.30 mmol C m-2 day-1, and there is a linear relation between DCF and water depth. The CO2 fixation ratio (DCF/CO2 respired) also shows a trend with water depth, decreasing from 0.09 in nearshore environments to 0.04 in continental shelf sediments. Five types of depth distributions of chemoautotrophic activity are identified based on the mode of pore water transport (advective, bioturbated, and diffusive) and the dominant pathway of microbial sulfur oxidation. Extrapolated to the global coastal ocean, we estimate a DCF rate of 0.04 to 0.06 Pg C year-1, which is less than previous estimates based on indirect measurements (0.15 Pg C year-1), but remains substantially higher than the global DCF rate at deep sea hydrothermal vents (0.001-0.002 Pg C year-1).