Maher WA, Duncan E, Dilly G, Foster S, Krikowa F, Lombi E, Scheckel K, Girguis P. Arsenic concentrations and species in three hydrothermal vent worms, Ridgeia piscesae, Paralvinella sulficola and Paralvinella palmiformis. Deep Sea Research Part I: Oceanographic Research Papers. 2016;116 :41 - 48. Publisher's VersionAbstract
Abstract Hydrothermal vents are surficial expressions of subsurface geological and hydrological processes. Fluids emitting from active vents are chemically distinct from bottom seawater, and are enriched in dissolved metals and metalloids, including arsenic. Vent organisms accumulate arsenic but the arsenic speciation in these non-photosynthetic organisms is largely unknown. Here, arsenic concentrations and chemical species were measured in three deep sea hydrothermal vent worms (Ridgeia piscesae, Paralvinella sulfincola and Paralvinella palmiformis ) from the Juan de Fuca Ridge in the Northwest pacific. R. piscesae has similar arsenic concentrations (3.8–35 µg g−1) to shallow water polychaetes while P. sulfincola and P. palmiformis have significantly higher arsenic concentrations (420–1417 and 125–321 µg g−1 respectively). R. piscesae contains appreciable quantities of inorganic arsenic (36±14%), monomethyl arsenic (2±2%), dimethyl arsenic (34±21%), an unknown methyl arsenical (7±16%), OSO3-arsenosugar (5±9%), \TETRA\ (4±5%), ThioPO4/ThioDMAE (1±2%) and an unknown thio-arsenical (12±14%). These results suggests that host and symbionts are either involved in the methylation of arsenic, or are bathed in fluids enriched in methylated arsenic as a result of free-living microbial activity. The host carrying out methylation, however, cannot be ruled out. In contrast, 96–97% of the arsenic in P. sulfincola and P. palmiformis is inorganic arsenic, likely the result of arsenic precipitation within and upon the mucus they ingest while feeding. While all worms have oxo- and thio arsenosugars (2–30%), Paralvinella also have small amounts of arsenobetaine (<0.001–0.21%). The presence of arsenosugars, arsenobetaine and other minor arsenic species in the absence of photosynthesising algae/bacteria indicates that they may be formed by vent animals in the absence of sunlight, but at this time their formation cannot be explained.
Girguis P. Microbial ecology: Here, there and everywhere. Nature Microbiology. 2016;1 :16123 - . Publisher's Version
Meier DV, Bach W, Girguis PR, Gruber-Vodicka HR, Reeves EP, Richter M, Vidoudez C, Amann R, Meyerdierks A. Heterotrophic Proteobacteria in the vicinity of diffuse hydrothermal venting. Environmental Microbiology. 2016 :n/a–n/a. Publisher's VersionAbstract
Deep-sea hydrothermal vents are highly dynamic habitats characterized by steep temperature and chemical gradients. The oxidation of reduced compounds dissolved in the venting fluids fuels primary production providing the basis for extensive life. Until recently studies of microbial vent communities have focused primarily on chemolithoautotrophic organisms. In our study, we targeted the change of microbial community compositions along mixing gradients, focusing on distribution and capabilities of heterotrophic microorganisms. Samples were retrieved from different venting areas within the Menez Gwen hydrothermal field, taken along mixing gradients, including diffuse fluid discharge points, their immediate surroundings and the buoyant parts of hydrothermal plumes. High throughput 16S rRNA gene amplicon sequencing, fluorescence in situ hybridization, and targeted metagenome analysis were combined with geochemical analyses. Close to diffuse venting orifices dominated by chemolithoautotrophic Epsilonproteobacteria, in areas where environmental conditions still supported chemolithoautotrophic processes, we detected microbial communities enriched for versatile heterotrophic Alpha- and Gammaproteobacteria. The potential for alkane degradation could be shown for several genera and yet uncultured clades. We propose that hotspots of chemolithoautotrophic life support a ‘belt’ of heterotrophic bacteria significantly different from the dominating oligotrophic microbiota of the deep sea.
Schrader PS, Reimers CE, Girguis P, Delaney J, Doolan C, Wolf M, Green D. Independent Benthic Microbial Fuel Cells Powering Sensors and Acoustic Communications with the MARS Underwater Observatory. Journal of Atmospheric and Oceanic Technology. 2016;33 (3) :607-617. Publisher's VersionAbstract
AbstractMost oceanographic instruments on the seafloor have no connections with the surface and therefore have to run on batteries and store data until recovery. To demonstrate a developing technology, sensors and acoustic modems were powered with energy harvested from the seafloor, and data were relayed acoustically in near–real time to the Monterey Accelerated Research System (MARS) observatory in Monterey Bay, California, and to surface research vessels. MARS is a cabled observatory in deep water ( 890 m) at the edge of Monterey Canyon. An acoustic modem was attached to the MARS node and configured to send out commands to, and relay data received from, remote modems. Two benthic microbial fuel cells (BMFCs) positioned approximately 0.5 km away from MARS supplied power to the remote modems and sensors. At their peak performance, these BMFCs produced continuous power densities of  35 mW m−2 (footprint area). The modems utilized in this study contained an integrated power management platform (PMP) designed to manage and store the electrical energy generated by each BMFC and to record BMFC performance parameters and sensor data on an hourly basis. Temperature and either oxygen or conductivity sensors were chosen because of their common use and environmental relevance. Acoustically transmitted data records show that the BMFCs renewed energy stores and that the oceanographic sensors measured dissolved oxygen, temperature, and conductivity reliably throughout the operational life of each BMFC system ( 6 months). These systems remained in place for more than 12 months.
Padilla CC, Bristow LA, Sarode N, Garcia-Robledo E, Gomez Ramirez E, Benson CR, Bourbonnais A, Altabet MA, Girguis PR, Thamdrup B, et al. NC10 bacteria in marine oxygen minimum zones. ISME J. 2016;10 (8) :2067 - 2071. Publisher's VersionAbstract
Bacteria of the NC10 phylum link anaerobic methane oxidation to nitrite denitrification through a unique O2-producing intra-aerobic methanotrophy pathway. A niche for NC10 in the pelagic ocean has not been confirmed. We show that NC10 bacteria are present and transcriptionally active in oceanic oxygen minimum zones (OMZs) off northern Mexico and Costa Rica. NC10 16S rRNA genes were detected at all sites, peaking in abundance in the anoxic zone with elevated nitrite and methane concentrations. Phylogenetic analysis of particulate methane monooxygenase genes further confirmed the presence of NC10. rRNA and mRNA transcripts assignable to NC10 peaked within the OMZ and included genes of the putative nitrite-dependent intra-aerobic pathway, with high representation of transcripts containing the unique motif structure of the nitric oxide (NO) reductase of NC10 bacteria, hypothesized to participate in O2-producing NO dismutation. These findings confirm pelagic OMZs as a niche for NC10, suggesting a role for this group in OMZ nitrogen, methane and oxygen cycling.
Seston SL, Beinart RA, Sarode N, Shockey AC, Ranjan P, Ganesh S, Girguis PR, Stewart FJ. Metatranscriptional Response of Chemoautotrophic Ifremeria nautilei Endosymbionts to Differing Sulfur Regimes. Frontiers in Microbiology. 2016;7 :1074. Publisher's VersionAbstract
Endosymbioses between animals and chemoautotrophic bacteria are ubiquitous at hydrothermal vents. These environments are distinguished by high physico-chemical variability, yet we know little about how these symbioses respond to environmental fluctuations. We therefore examined how the γ-proteobacterial symbionts of the vent snail Ifremeria nautilei respond to changes in sulfur geochemistry. Via shipboard high-pressure incubations, we subjected snails to 105 μM hydrogen sulfide (LS), 350 μM hydrogen sulfide (HS), 300 μM thiosulfate (TS) and seawater without any added inorganic electron donor (ND). While transcript levels of sulfur oxidation genes were largely consistent across treatments, HS and TS treatments stimulated genes for denitrification, nitrogen assimilation, and CO(2) fixation, coincident with previously reported enhanced rates of inorganic carbon incorporation and sulfur oxidation in these treatments. Transcripts for genes mediating oxidative damage were enriched in the ND and LS treatments, potentially due to a reduction in O(2) scavenging when electron donors were scarce. Oxidative TCA cycle gene transcripts were also more abundant in ND and LS treatments, suggesting that I. nautilei symbionts may be mixotrophic when inorganic electron donors are limiting. These data reveal the extent to which I. nautilei symbionts respond to changes in sulfur concentration and species, and, interpreted alongside coupled biochemical metabolic rates, identify gene targets whose expression patterns may be predictive of holobiont physiology in environmental samples.
Picard A, Gartman A, Girguis PR. What Do We Really Know about the Role of Microorganisms in Iron Sulfide Mineral Formation?. Frontiers in Earth Science. 2016;4 :68. Publisher's VersionAbstract
Iron sulfide mineralization in low-temperature systems is a result of biotic and abiotic processes, though the delineation between these two modes of formation is not always straightforward. Here we review the role of microorganisms in the precipitation of extracellular iron sulfide minerals. We summarize the evidence that links sulfur-metabolizing microorganisms and sulfide minerals in nature and we present a critical overview of laboratory-based studies of the nucleation and growth of iron sulfide minerals in microbial cultures. We discuss whether biologically derived minerals are distinguishable from abiotic minerals, possessing attributes that are uniquely diagnostic of biomineralization. These inquiries have revealed the need for additional thorough, mechanistic and high-resolution studies to understand microbially mediated formation of a variety of sulfide minerals across a range of natural environments.
Meyer JL, Jaekel U, Tully BJ, Glazer BT, Wheat GC, Lin H-T, Hsieh C-C, Cowen JP, Hulme SM, Girguis PR, et al. A distinct and active bacterial community in cold oxygenated fluids circulating beneath the western flank of the Mid-Atlantic ridge. Scientific Reports. 2016;6 :22541. Publisher's VersionAbstract
The rock-hosted, oceanic crustal aquifer is one of the largest ecosystems on Earth, yet little is known about its indigenous microorganisms. Here we provide the first phylogenetic and functional description of an active microbial community residing in the cold oxic crustal aquifer. Using subseafloor observatories, we recovered crustal fluids and found that the geochemical composition is similar to bottom seawater, as are cell abundances. However, based on relative abundances and functional potential of key bacterial groups, the crustal fluid microbial community is heterogeneous and markedly distinct from seawater. Potential rates of autotrophy and heterotrophy in the crust exceeded those of seawater, especially at elevated temperatures (25 °C) and deeper in the crust. Together, these results reveal an active, distinct, and diverse bacterial community engaged in both heterotrophy and autotrophy in the oxygenated crustal aquifer, providing key insight into the role of microbial communities in the ubiquitous cold dark subseafloor biosphere.
Ge Z, Girguis PR, Buie CR. Nanoporous microscale microbial incubators. Lab Chip. 2016;16 :480-488. Publisher's VersionAbstract
Reconstruction of phylogenetic trees based on 16S rRNA gene sequencing reveals abundant microbial diversity that has not been cultured in the laboratory. Many attribute this so-called 'great plate count anomaly' to traditional microbial cultivation techniques, which largely facilitate the growth of a single species. Yet, it is widely recognized that bacteria in nature exist in complex communities. One technique to increase the pool of cultivated bacterial species is to co-culture multiple species in a simulated natural environment. Here, we present nanoporous microscale microbial incubators (NMMI) that enable high-throughput screening and real-time observation of multi-species co-culture. The key innovation in NMMI is that they facilitate inter-species communication while maintaining physical isolation between species, which is ideal for genomic analysis. Co-culture of a quorum sensing pair demonstrates that the NMMI can be used to culture multiple species in chemical communication while monitoring the growth dynamics of individual species.
Bradley AS, Leavitt WD, Schmidt M, Knoll AH, Girguis PR, Johnston DT. Patterns of sulfur isotope fractionation during microbial sulfate reduction. Geobiology. 2016;14 (1) :91–101. Publisher's VersionAbstract
Studies of microbial sulfate reduction have suggested that the magnitude of sulfur isotope fractionation varies with sulfate concentration. Small apparent sulfur isotope fractionations preserved in Archean rocks have been interpreted as suggesting Archean sulfate concentrations of <200 μm, while larger fractionations thereafter have been interpreted to require higher concentrations. In this work, we demonstrate that fractionation imposed by sulfate reduction can be a function of concentration over a millimolar range, but that nature of this relationship depends on the organism studied. Two sulfate-reducing bacteria grown in continuous culture with sulfate concentrations ranging from 0.1 to 6 mm showed markedly different relationships between sulfate concentration and isotope fractionation. Desulfovibrio vulgaris str. Hildenborough showed a large and relatively constant isotope fractionation (34εSO4-H2S ≅ 25‰), while fractionation by Desulfovibrio alaskensis G20 strongly correlated with sulfate concentration over the same range. Both data sets can be modeled as Michaelis–Menten (MM)-type relationships but with very different MM constants, suggesting that the fractionations imposed by these organisms are highly dependent on strain-specific factors. These data reveal complexity in the sulfate concentration–fractionation relationship. Fractionation during MSR relates to sulfate concentration but also to strain-specific physiological parameters such as the affinity for sulfate and electron donors. Previous studies have suggested that the sulfate concentration–fractionation relationship is best described with a MM fit. We present a simple model in which the MM fit with sulfate concentration and hyperbolic fit with growth rate emerge from simple physiological assumptions. As both environmental and biological factors influence the fractionation recorded in geological samples, understanding their relationship is critical to interpreting the sulfur isotope record. As the uptake machinery for both sulfate and electrons has been subject to selective pressure over Earth history, its evolution may complicate efforts to uniquely reconstruct ambient sulfate concentrations from a single sulfur isotopic composition.
Sanders JG, Beichman AC, Roman J, Scott JJ, Emerson D, McCarthy JJ, Girguis PR. Baleen whales host a unique gut microbiome with similarities to both carnivores and herbivores. Nature Communications. 2015;6 :8285 - . Publisher's Version
Frank KL, Rogers KL, Rogers DR, Johnston DT, Girguis PR. Key Factors Influencing Rates of Heterotrophic Sulfate Reduction in Active Seafloor Hydrothermal Massive Sulfide Deposits. Frontiers in Microbiology. 2015;6 :1449. Publisher's VersionAbstract
Hydrothermal vents are thermally and geochemically dynamic habitats, and the organisms therein are subject to steep gradients in temperature and chemistry. To date, the influence of these environmental dynamics on microbial sulfate reduction has not been well constrained. Here, via multivariate experiments, we evaluate the effects of key environmental variables (temperature, pH, H2S, SO42-, DOC) on sulfate reduction rates and metabolic energy yields in material recovered from a hydrothermal flange from the Grotto edifice in the Main Endeavor Field, Juan de Fuca Ridge. Sulfate reduction was measured in batch reactions across a range of physico-chemical conditions. Temperature and pH were the strongest stimuli, and maximum sulfate reduction rates were observed at 50 °C and pH 6, suggesting that the in situ community of sulfate-reducing organisms in Grotto flanges may be most active in a slightly acidic and moderate thermal/chemical regime. At pH 4, sulfate reduction rates increased with sulfide concentrations most likely due to the mitigation of metal toxicity. While substrate concentrations also influenced sulfate reduction rates, energy-rich conditions muted the effect of metabolic energetics on sulfate reduction rates. We posit that variability in sulfate reduction rates reflect the response of the active microbial consortia to environmental constraints on in situ microbial physiology, toxicity, and the type and extent of energy limitation. These experiments help to constrain models of the spatial contribution of heterotrophic sulfate reduction within the complex gradients inherent to seafloor hydrothermal deposits.
Gittel A, Donhauser J, Røy H, Girguis PR, Jørgensen BB, Kjeldsen KU. Ubiquitous Presence and Novel Diversity of Anaerobic Alkane Degraders in Cold Marine Sediments. Frontiers in Microbiology. 2015;6 :1414. Publisher's VersionAbstract
Alkanes are major constituents of crude oil and are released to the marine environment by natural seepage and from anthropogenic sources. Due to their chemical inertness, their removal from anoxic marine sediments is primarily controlled by the activity of anaerobic alkane-degrading microorganisms. To conduct a comprehensive cultivation-independent survey of the diversity and distribution of anaerobic alkane degraders, we designed novel PCR primers that cover all known diversity of the 1-methylalkyl succinate synthase gene (masD/assA), which catalyzes the initial activation of alkanes. We studied masD/assA gene diversity in pristine and seepage-impacted Danish coastal sediments, as well as in sediments and alkane-degrading enrichment cultures from the Middle Valley (MV) hydrothermal vent system in the Pacific Northwest. MasD/assA genes were ubiquitously present, and the primers captured the diversity of both known and previously undiscovered masD/assA gene diversity. Seepage sediments were dominated by a single masD/assA gene cluster, which is presumably indicative of a substrate-adapted community, while pristine sediments harbored a diverse range of masD/assA phylotypes including those present in seepage sediments. This rare biosphere of anaerobic alkane degraders will likely increase in abundance in the event of seepage or accidental oil spillage. Nanomolar concentrations of short-chain alkanes (SCA) were detected in pristine and seepage sediments. Interestingly, anaerobic alkane degraders closely related to strain BuS5, the only SCA degrader in pure culture, were found in mesophilic MV enrichments, but not in cold sediments from Danish waters. We propose that the new masD/assA gene lineages in these sediments represent novel phylotypes that are either fueled by naturally occurring low levels of SCA or that metabolize medium- to long-chain alkanes. Our study highlights that masD/assA genes are a relevant diagnostic marker to identify seepage and microseepage, e.g. during prospecting for oil and gas, and may act as an indicator of anthropogenic oil spills in marine sediments.
Orcutt BN, Sylvan JB, Rogers DR, Delaney J, Lee RW, Girguis PR. Carbon fixation by basalt-hosted microbial communities. Frontiers in Microbiology. 2015;6 :904. Publisher's VersionAbstract
Oceanic crust is a massive potential habitat for microbial life on Earth, yet our understanding of this ecosystem is limited due to difficulty in access. In particular, measurements of rates of microbial activity are sparse. We used stable carbon isotope incubations of crustal samples, coupled with functional gene analyses, to examine the potential for carbon fixation on oceanic crust. Both seafloor-exposed and subseafloor basalts were recovered from different mid-ocean ridge and hot spot environments (i.e., the Juan de Fuca Ridge, the Mid-Atlantic Ridge, and the Loihi Seamount) and incubated with (13)C-labeled bicarbonate. Seafloor-exposed basalts revealed incorporation of (13)C-label into organic matter over time, though the degree of incorporation was heterogeneous. The incorporation of (13)C into biomass was inconclusive in subseafloor basalts. Translating these measurements into potential rates of carbon fixation indicated that 0.1–10 nmol C g(-1)(rock) d(-1) could be fixed by seafloor-exposed rocks. When scaled to the global production of oceanic crust, this suggests carbon fixation rates of 10(9)–10(12) g C year(-1), which matches earlier predictions based on thermodynamic calculations. Functional gene analyses indicate that the Calvin cycle is likely the dominant biochemical mechanism for carbon fixation in basalt-hosted biofilms, although the reductive acetyl-CoA pathway and reverse TCA cycle likely play some role in net carbon fixation. These results provide empirical evidence for autotrophy in oceanic crust, suggesting that basalt-hosted autotrophy could be a significant contributor of organic matter in this remote and vast environment.
Lu Z, Girguis P, Liang P, Shi H, Huang G, Cai L, Zhang L. Biological capacitance studies of anodes in microbial fuel cells using electrochemical impedance spectroscopy. Bioprocess and Biosystems Engineering. 2015;38 :1325-1333. (pdf)
Jiang Y, Liang P, Zhang C, Bian Y, Yang X, Huang X, Girguis PR. Enhancing the response of microbial fuel cell based toxicity sensors to Cu (II) with the applying of flow-through electrodes and controlled anode potentials. Bioresource Technology. 2015;190 :367-372.
Bradley AS, Leavitt WD, Schmidt M, Knoll AH, Girguis PR, Johnston DT. Patterns of microbial sulfur isotope fractionation at low sulfate concentrations. Geobiology. 2015.
Beinart RA, Gartman A, Sanders JG, Luther GW, Girguis PR. The uptake and excretion of partially oxidized sulfur expands the repertoire of energy resources metabolized by hydrothermal vent symbioses. Proceedings of the Royal Society Biological Sciences. 2015;282. (pdf)
Liao L, Wankel SD, Wu M, Cavanaugh CM, Girguis PR. Characterizing the plasticity of nitrogen metabolism by the host and symbionts of the hydrothermal vent chemoautotrophic symbioses Ridgeia piscesae. Molecular Ecology. 2014;23 :1544-1557.Abstract

Chemoautotrophic symbionts of deep sea hydrothermal vent tubeworms are known to provide their hosts with all their primary nutrition. While studies have examined how chemoautotrophic symbionts provide the association with nitrogen, fewer have examined if symbiont nitrogen metabolism varies as a function of environmental conditions. Ridgeia piscesae tubeworms flourish at Northeastern Pacific vents, occupy a range of microhabitats, and exhibit a high degree of morphological plasticity [e.g. long-skinny (LS) and short-fat (SF) phenotypes] that may relate to environmental conditions. This plasticity affords an opportunity to examine whether symbiont nitrogen metabolism varies among host phenotypes. LS and SF R.piscesae were recovered from the Axial and Main Endeavour Field hydrothermal vents. Nitrate and ammonium were quantified in Ridgeia blood, and the expression of key nitrogen metabolism genes, as well as stable nitrogen isotope ratios, was quantified in host branchial plume and symbiont-containing tissues. Nitrate and ammonium were abundant in the blood of both phenotypes though environmental ammonium concentrations were, paradoxically, lowest among individuals with the highest blood ammonium. Assimilatory nitrate reductase transcripts were always below detection, though in both LS and SF R.piscesae symbionts, we observed elevated expression of dissimilatory nitrate reductase genes, as well as symbiont and host ammonium assimilation genes. Site-specific differences in expression, along with tissue stable isotope analyses, suggest that LS and SF Ridgeia symbionts are engaged in both dissimilatory nitrate reduction and ammonia assimilation to varying degrees. As such, it appears that environmental conditions -not host phenotype-primarily dictates symbiont nitrogen metabolism.

Bose A, Gardel EJ, Vidoudez C, Parra EA, Girguis PR. Electron uptake by iron-oxidizing phototrophic bacteria. Nature Communications. 2014;5 :1-7.Abstract

Oxidation-reduction reactions underlie energy generation in nearly all life forms. Although most organisms use soluble oxidants and reductants, some microbes can access solid-phase materials as electron-acceptors or -donors via extracellular electron transfer. Many studies have focused on the reduction of solid-phase oxidants. Far less is known about electron uptake via microbial extracellular electron transfer, and almost nothing is known about the associated mechanisms. Here we show that the iron-oxidizing photoautotroph Rhodopseudomonas palustris TIE-1 accepts electrons from a poised electrode, with carbon dioxide as the sole carbon source/electron acceptor. Both electron uptake and ruBisCo form I expression are stimulated by light. Electron uptake also occurs in the dark, uncoupled from photosynthesis. Notably, the pioABC operon, which encodes a protein system essential for photoautotrophic growth by ferrous iron oxidation, influences electron uptake. These data reveal a previously unknown metabolic versatility of photoferrotrophs to use extracellular electron transfer for electron uptake.