A Lab-on-Chip Analyzer for in Situ Measurement of Soluble Reactive Phosphate: Improved Phosphate Blue Assay and Application to Fluvial Monitoring.

Here, we present a new in situ microfluidic phosphate sensor that features an improved "phosphate blue" assay which includes polyvinylpyrrolidone in place of traditional surfactants-improving sensitivity and reducing temperature effects. The sensor features greater power economy and analytical performance relative to commercially available alternatives, with a mean power consumption of 1.8 W, a detection limit of 40 nM, a dynamic range of 0.14-10 µM, and an infield accuracy of 4 ± 4.5%. During field testing, the sensor was continuously deployed for 9 weeks in a chalk stream, revealing complex relations between flow rates and phosphate concentration that suggest changing dominance in phosphate sources. A distinct diel phosphorus signal was observed under low flow conditions, highlighting the ability of the sensor to decouple geochemical and biotic effects on phosphate dynamics in fluvial environments. This paper highlights the importance of high resolution in situ sensors in addressing the current gross under-sampling of aquatic environments.

The discovery of new deep-sea hydrothermal vent communities in the southern ocean and implications for biogeography.

Since the first discovery of deep-sea hydrothermal vents along the Galápagos Rift in 1977, numerous vent sites and endemic faunal assemblages have been found along mid-ocean ridges and back-arc basins at low to mid latitudes. These discoveries have suggested the existence of separate biogeographic provinces in the Atlantic and the North West Pacific, the existence of a province including the South West Pacific and Indian Ocean, and a separation of the North East Pacific, North East Pacific Rise, and South East Pacific Rise. The Southern Ocean is known to be a region of high deep-sea species diversity and centre of origin for the global deep-sea fauna. It has also been proposed as a gateway connecting hydrothermal vents in different oceans but is little explored because of extreme conditions. Since 2009 we have explored two segments of the East Scotia Ridge (ESR) in the Southern Ocean using a remotely operated vehicle. In each segment we located deep-sea hydrothermal vents hosting high-temperature black smokers up to 382.8°C and diffuse venting. The chemosynthetic ecosystems hosted by these vents are dominated by a new yeti crab (Kiwa n. sp.), stalked barnacles, limpets, peltospiroid gastropods, anemones, and a predatory sea star. Taxa abundant in vent ecosystems in other oceans, including polychaete worms (Siboglinidae), bathymodiolid mussels, and alvinocaridid shrimps, are absent from the ESR vents. These groups, except the Siboglinidae, possess planktotrophic larvae, rare in Antarctic marine invertebrates, suggesting that the environmental conditions of the Southern Ocean may act as a dispersal filter for vent taxa. Evidence from the distinctive fauna, the unique community structure, and multivariate analyses suggest that the Antarctic vent ecosystems represent a new vent biogeographic province. However, multivariate analyses of species present at the ESR and at other deep-sea hydrothermal vents globally indicate that vent biogeography is more complex than previously recognised.

Hyperbaric biofilms on engineering surfaces formed in the deep sea.

Biofouling is a major problem for long-term deployment of sensors in the marine environment. This study showed that significant biofilm formation occurred on a variety of artificial materials (glass, copper, Delrin(™) and poly-methyl methacrylate [PMMA]) deployed for 10 days at a depth of 4700 m in the Cayman Trough. Biofilm surface coverage was used as an indicator of biomass. The lowest biofilm coverage was on copper and PMMA. Molecular analyses indicated that bacteria dominated the biofilms found on copper, Delrin(™) and PMMA with 75, 55 and 73% coverage, respectively. Archea (66%) were dominant on the glass surface simulating interior sensor conditions, whereas Eukarya comprised the highest percentage of microflora (75%) on the glass simulating the exterior of sensors. Analysis of Denaturing Gradient Gel Electrophoresis profiles indicated that copper and Delrin(™) shared the same community diversity, which was not the case for glass and PMMA, or between PMMA and copper/Delrin(™). Sequence alignment matches belonged exclusively to uncultivable microorganisms, most of which were not further classified. One extracted sequence found on glass was associated with Cowellia sp., while another extracted from the PMMA surface was associated with a bacterium in the Alterominidaceae, both γ-proteobacteria. The results demonstrate the necessity of understanding biofilm formation in the deep sea and the potential need for mitigation strategies for any kind of long-term deployment of remote sensors in the marine environment.

Lab-on-chip analyser for the in situ determination of dissolved manganese in seawater.

A spectrophotometric approach for quantification of dissolved manganese (DMn) with 1-(2-pyridylazo)-2-naphthol (PAN) has been adapted for in situ application in coastal and estuarine waters. The analyser uses a submersible microfluidic lab-on-chip device, with low power (~ 1.5 W) and reagent consumption (63 µL per sample). Laboratory characterization showed an absorption coefficient of 40,838 ± 1127 L⋅mol-1⋅cm-1 and a detection limit of 27 nM, determined for a 34.6 mm long optical detection cell. Laboratory tests showed that long-term stability of the PAN reagent was achieved by addition of 4% v/v of a non-ionic surfactant (Triton-X100). To suppress iron (Fe) interferences with the PAN reagent, the Fe(III) masking agents deferoxamine mesylate (DFO-B) or disodium 4,5-dihydroxy-1,3-benzenedisulfonate (Tiron) were added and their Fe masking efficiencies were investigated. The analyser was tested during a deployment over several weeks in Kiel Fjord (Germany), with successful acquisition of 215 in situ data points. The time series was in good agreement with DMn concentrations determined from discretely collected samples analysed via inductively coupled plasma mass spectrometry (ICP-MS), exhibiting a mean accuracy of 87% over the full deployment duration (with an accuracy of > 99% for certain periods) and clear correlations to key hydrographic parameters.

Fauna of the Kemp Caldera and its upper bathyal hydrothermal vents (South Sandwich Arc, Antarctica).

Faunal assemblages at hydrothermal vents associated with island-arc volcanism are less well known than those at vents on mid-ocean ridges and back-arc spreading centres. This study characterizes chemosynthetic biotopes at active hydrothermal vents discovered at the Kemp Caldera in the South Sandwich Arc. The caldera hosts sulfur and anhydrite vent chimneys in 1375-1487 m depth, which emit sulfide-rich fluids with temperatures up to 212°C, and the microbial community of water samples in the buoyant plume rising from the vents was dominated by sulfur-oxidizing Gammaproteobacteria. A total of 12 macro- and megafaunal taxa depending on hydrothermal activity were collected in these biotopes, of which seven species were known from the East Scotia Ridge (ESR) vents and three species from vents outside the Southern Ocean. Faunal assemblages were dominated by large vesicomyid clams, actinostolid anemones, Sericosura sea spiders and lepetodrilid and cocculinid limpets, but several taxa abundant at nearby ESR hydrothermal vents were rare such as the stalked barnacle Neolepas scotiaensis. Multivariate analysis of fauna at Kemp Caldera and vents in neighbouring areas indicated that the Kemp Caldera is most similar to vent fields in the previously established Southern Ocean vent biogeographic province, showing that the species composition at island-arc hydrothermal vents can be distinct from nearby seafloor-spreading systems. δ 13C and δ 15N isotope values of megafaunal species analysed from the Kemp Caldera were similar to those of the same or related species at other vent fields, but none of the fauna sampled at Kemp Caldera had δ 13C values, indicating nutritional dependence on Epsilonproteobacteria, unlike fauna at other island-arc hydrothermal vents.

A novel, low-cost, high performance dissolved methane sensor for aqueous environments.

A new method for in-situ detection and measurement of dissolved methane in aqueous media/environments with a limit of detection of 0.2 nM (3 sigma, and t90 approxiamtely 110s) and range (1-300 nM) is presented. The detection method is based on refractive index (RI) modulation of a modified PolyDiMethylSiloxane (PDMS) layer incorporating molecules of cryptophane-A [1] which have a selective and reversible affinity for methane [2]. The refractive index is accurately determined using surface plasmon resonance (SPR) [3]. A prototype sensor has been repeatedly tested, using a dissolved gas calibration system under a range of temperature and salinity regimes. Laboratory-based results show that the technique is specific, sensitive, and reversible. The method is suitable for miniaturization and incorporation into in situ sensor technology.

Iron Biogeochemistry in the High Latitude North Atlantic Ocean.

Iron (Fe) is an essential micronutrient for marine microbial organisms, and low supply controls productivity in large parts of the world's ocean. The high latitude North Atlantic is seasonally Fe limited, but Fe distributions and source strengths are poorly constrained. Surface ocean dissolved Fe (DFe) concentrations were low in the study region (<0.1 nM) in summer 2010, with significant perturbations during spring 2010 in the Iceland Basin as a result of an eruption of the Eyjafjallajökull volcano (up to 2.5 nM DFe near Iceland) with biogeochemical consequences. Deep water concentrations in the vicinity of the Reykjanes Ridge system were influenced by pronounced sediment resuspension, with indications for additional inputs by hydrothermal vents, with subsequent lateral transport of Fe and manganese plumes of up to 250-300 km. Particulate Fe formed the dominant pool, as evidenced by 4-17 fold higher total dissolvable Fe compared with DFe concentrations, and a dynamic exchange between the fractions appeared to buffer deep water DFe. Here we show that Fe supply associated with deep winter mixing (up to 103 nmol m-2 d-1) was at least ca. 4-10 times higher than atmospheric deposition, diffusive fluxes at the base of the summer mixed layer, and horizontal surface ocean fluxes.

Development and application of a microfluidic in-situ analyzer for dissolved Fe and Mn in natural waters.

The redox sensitive trace metals iron and manganese are two important elements that help shape the biogeochemistry of aquatic systems and thus their measurement is important. Current laboratory methods are expensive, time consuming and cannot provide the spatial and temporal resolution needed to characterize these elements in natural waters. Here we describe the first autonomous analyzer capable of providing vertical profiles as well as routine in-situ determinations of dissolved Fe(II) and Mn in aquatic environments. The spectrophotometric sensor uses microfluidic methods (Lab-on-a-chip technology) and mixes reagents and samples using a novel in-cell diffusion process. Fe(II) and Mn can be measured with a frequency of up to 12 and 6 samples per hour respectively with limits of detection of 27nM for Fe(II), 2.1% precision (n=20), and 28nM for Mn, 2.4% precision (n=19). The device combines relatively low cost, low power usage, low reagent consumption, portability, and tolerance to pressures up to at least 170 bars, with high precision and accuracy. We present data from a successful demonstration of the sensor during a cruise to the Gotland and Landsort Deep Basins of the Baltic Sea.

Talc-dominated seafloor deposits reveal a new class of hydrothermal system.

The Von Damm Vent Field (VDVF) is located on the flanks of the Mid-Cayman Spreading Centre, 13 km west of the axial rift, within a gabbro and peridotite basement. Unlike any other active vent field, hydrothermal precipitates at the VDVF comprise 85-90% by volume of the magnesium silicate mineral, talc. Hydrothermal fluids vent from a 3-m high, 1-m diameter chimney and other orifices at up to 215 °C with low metal concentrations, intermediate pH (5.8) and high concentrations (667 mmol kg(-1)) of chloride relative to seawater. Here we show that the VDVF vent fluid is generated by interaction of seawater with a mafic and ultramafic basement which precipitates talc on mixing with seawater. The heat flux at the VDVF is measured at 487±101 MW, comparable to the most powerful magma-driven hydrothermal systems known, and may represent a significant mode of off-axis oceanic crustal cooling not previously recognized or accounted for in global models.

Linking regional variation of epibiotic bacterial diversity and trophic ecology in a new species of Kiwaidae (Decapoda, Anomura) from East Scotia Ridge (Antarctica) hydrothermal vents.

We analyzed the diversity of bacterial epibionts and trophic ecology of a new species of Kiwa yeti crab discovered at two hydrothermal vent fields (E2 and E9) on the East Scotia Ridge (ESR) in the Southern Ocean using a combination of 454 pyrosequencing, Sanger sequencing, and stable isotope analysis. The Kiwa epibiont communities were dominated by Epsilon- and Gammaproteobacteria. About 454 sequencing of the epibionts on 15 individual Kiwa specimen revealed large regional differences between the two hydrothermal vent fields: at E2, the bacterial community on the Kiwa ventral setae was dominated (up to 75%) by Gammaproteobacteria, whereas at E9 Epsilonproteobacteria dominated (up to 98%). Carbon stable isotope analysis of both Kiwa and the bacterial epibionts also showed distinct differences between E2 and E9 in mean and variability. Both stable isotope and sequence data suggest a dominance of different carbon fixation pathways of the epibiont communities at the two vent fields. At E2, epibionts were putatively fixing carbon via the Calvin-Benson-Bassham and reverse tricarboxylic acid cycle, while at E9 the reverse tricarboxylic acid cycle dominated. Co-varying epibiont diversity and isotope values at E2 and E9 also present further support for the hypothesis that epibionts serve as a food source for Kiwa.

Characterisation of iron binding ligands in seawater by reverse titration.

Here we demonstrate the use of reverse titration - competitive ligand exchange-adsorptive cathodic stripping voltammetry (RT-CLE-ACSV) for the analysis of iron (Fe) binding ligands in seawater. In contrast to the forward titration, which examines excess ligands in solution, RT-CLE-ACSV examines the existing Fe-ligand complexes by increasing the concentration of added (electroactive) ligand (1-nitroso-2-naphthol) and analysis of the proportion of Fe bound to the added ligand. The data manipulation allows the accurate characterisation of ligands at equal or lower concentrations than Fe in seawater, and disregards electrochemically inert dissolved Fe such as some colloidal phases. The method is thus superior to the forward titration in environments with high Fe and low ligand concentrations or high concentrations of inert Fe. We validated the technique using the siderophore ligand ferrioxamine B, and observed a stability constant [Formula: see text] of 0.74-4.37×10(21) mol(-1), in agreement with previous results. We also successfully analysed samples from coastal waters and a deep ocean hydrothermal plume. Samples from these environments could not be analysed with confidence using the forward titration, highlighting the effectiveness of the RT-CLE-ACSV technique in waters with high concentrations of inert Fe.

A high performance microfluidic analyser for phosphate measurements in marine waters using the vanadomolybdate method.

We report a high performance autonomous analytical system based on the vanadomolybdate method for the determination of soluble reactive phosphorus in seawater. The system combines a microfluidic chip manufactured from tinted poly (methyl methacrylate) (PMMA), a custom made syringe pump, embedded control electronics and on-board calibration standards. This "lab-on-a-chip" analytical system was successfully deployed and cross-compared with reference analytical methods in coastal (south west England) and open ocean waters (tropical North Atlantic). The results of the miniaturized system compared well with a reference bench-operated phosphate auto-analyser and showed no significant differences in the analytical results (student's t-test at 95% confidence level). The optical technology used, comprising of tinted PMMA and polished fluidic channels, has allowed an improvement of two orders of magnitude of the limit of detection (52 nM) compared to currently available portable systems based on this method. The system has a wide linear dynamic range 0.1-60 µM, and a good precision (13.6% at 0.4 µM, n=4). The analytical results were corrected for silicate interferences at 0.7 µM, and the measurement frequency was configurable with a sampling throughput of up to 20 samples per hour. This portable micro-analytical system has a low reagent requirement (340 µL per sample) and power consumption (756 J per sample), and has allowed accurate high resolution measurements of soluble reactive phosphorus in seawater.

Geochemical and visual indicators of hydrothermal fluid flow through a sediment-hosted volcanic ridge in the Central Bransfield Basin (Antarctica).

In the austral summer of 2011 we undertook an investigation of three volcanic highs in the Central Bransfield Basin, Antarctica, in search of hydrothermal activity and associated fauna to assess changes since previous surveys and to evaluate the extent of hydrothermalism in this basin. At Hook Ridge, a submarine volcanic edifice at the eastern end of the basin, anomalies in water column redox potential (E(h)) were detected close to the seafloor, unaccompanied by temperature or turbidity anomalies, indicating low-temperature hydrothermal discharge. Seepage was manifested as shimmering water emanating from the sediment and from mineralised structures on the seafloor; recognisable vent endemic fauna were not observed. Pore fluids extracted from Hook Ridge sediment were depleted in chloride, sulfate and magnesium by up to 8% relative to seawater, enriched in lithium, boron and calcium, and had a distinct strontium isotope composition ((87)Sr/(86)Sr = 0.708776 at core base) compared with modern seawater ((87)Sr/(86)Sr ≈ 0.70918), indicating advection of hydrothermal fluid through sediment at this site. Biogeochemical zonation of redox active species implies significant moderation of the hydrothermal fluid with in situ diagenetic processes. At Middle Sister, the central ridge of the Three Sisters complex located about 100 km southwest of Hook Ridge, small water column E(h) anomalies were detected but visual observations of the seafloor and pore fluid profiles provided no evidence of active hydrothermal circulation. At The Axe, located about 50 km southwest of Three Sisters, no water column anomalies in E(h), temperature or turbidity were detected. These observations demonstrate that the temperature anomalies observed in previous surveys are episodic features, and suggest that hydrothermal circulation in the Bransfield Strait is ephemeral in nature and therefore may not support vent biota.

Hydrothermal vent fields and chemosynthetic biota on the world's deepest seafloor spreading centre.

The Mid-Cayman spreading centre is an ultraslow-spreading ridge in the Caribbean Sea. Its extreme depth and geographic isolation from other mid-ocean ridges offer insights into the effects of pressure on hydrothermal venting, and the biogeography of vent fauna. Here we report the discovery of two hydrothermal vent fields on the Mid-Cayman spreading centre. The Von Damm Vent Field is located on the upper slopes of an oceanic core complex at a depth of 2,300 m. High-temperature venting in this off-axis setting suggests that the global incidence of vent fields may be underestimated. At a depth of 4,960 m on the Mid-Cayman spreading centre axis, the Beebe Vent Field emits copper-enriched fluids and a buoyant plume that rises 1,100 m, consistent with >400 °C venting from the world's deepest known hydrothermal system. At both sites, a new morphospecies of alvinocaridid shrimp dominates faunal assemblages, which exhibit similarities to those of Mid-Atlantic vents.

Vailulu'u Seamount, Samoa: Life and death on an active submarine volcano.

Submersible exploration of the Samoan hotspot revealed a new, 300-m-tall, volcanic cone, named Nafanua, in the summit crater of Vailulu'u seamount. Nafanua grew from the 1,000-m-deep crater floor in <4 years and could reach the sea surface within decades. Vents fill Vailulu'u crater with a thick suspension of particulates and apparently toxic fluids that mix with seawater entering from the crater breaches. Low-temperature vents form Fe oxide chimneys in many locations and up to 1-m-thick layers of hydrothermal Fe floc on Nafanua. High-temperature (81 degrees C) hydrothermal vents in the northern moat (945-m water depth) produce acidic fluids (pH 2.7) with rising droplets of (probably) liquid CO(2). The Nafanua summit vent area is inhabited by a thriving population of eels (Dysommina rugosa) that feed on midwater shrimp probably concentrated by anticyclonic currents at the volcano summit and rim. The moat and crater floor around the new volcano are littered with dead metazoans that apparently died from exposure to hydrothermal emissions. Acid-tolerant polychaetes (Polynoidae) live in this environment, apparently feeding on bacteria from decaying fish carcasses. Vailulu'u is an unpredictable and very active underwater volcano presenting a potential long-term volcanic hazard. Although eels thrive in hydrothermal vents at the summit of Nafanua, venting elsewhere in the crater causes mass mortality. Paradoxically, the same anticyclonic currents that deliver food to the eels may also concentrate a wide variety of nektonic animals in a death trap of toxic hydrothermal fluids.