Nitrate and nitrite variability at the seafloor of an oxygen minimum zone revealed by a novel microfluidic in-situ chemical sensor

Microfluidics, or lab-on-a-chip (LOC) is a promising technology that allows the development of miniaturized chemical sensors. In contrast to the surging interest in biomedical sciences, the utilization of LOC sensors in aquatic sciences is still in infancy but a wider use of such sensors could mitigate the undersampling problem of ocean biogeochemical processes. Here we describe the first underwater test of a novel LOC sensor to obtain in situ calibrated time-series (up to 40 h) of nitrate+nitrite (ΣNOx) and nitrite on the seafloor of the Mauritanian oxygen minimum zone, offshore Western Africa. Initial tests showed that the sensor successfully reproduced water column (160 m) nutrient profiles. Lander deployments at 50, 100 and 170 m depth indicated that the biogeochemical variability was high over the Mauritanian shelf: The 50 m site had the lowest ΣNOx concentration, with 15.2 to 23.4 μM (median=18.3 μM); while at the 100 site ΣNOx varied between 21.0 and 30.1 μM over 40 hours (median = 25.1μM). The 170 m site had the highest median ΣNOx level (25.8 μM) with less variability (22.8 to 27.7 μM). At the 50 m site, nitrite concentration decreased fivefold from 1 to 0.2 μM in just 30 hours accompanied by decreasing oxygen and increasing nitrate concentrations. Taken together with the time series of oxygen, temperature, pressure and current velocities, we propose that the episodic intrusion of deeper waters via cross-shelf transport leads to intrusion of nitrate-rich, but oxygen-poor waters to shallower locations, with consequences for benthic nitrogen cycling. This first validation of an LOC sensor at elevated water depths revealed that when deployed for longer periods and as a part of a sensor network, LOC technology has the potential to contribute to the understanding of the benthic biogeochemical dynamics

Development of an Automated Detection System for Nitrite in Aquatic Environments

The main objective of the project is to develop an automated nitrite sensor for use in aquatic environments, and more specifically for use in recirculating aquaculture systems (RAS), where monitoring can help sustain a controlled environment, protect against nitrite intoxication, and promote fish health. Detecting nitrite manually with semi-quantitative colorimetric test kits, although inexpensive and simple, is prone to inter-user variability and poor sensitivity. An automated nitrite sensor has potential to provide higher resolution measurements at both concentration and time scales and can serve as a research tool for the study of filtration systems essential in maintaining a healthy RAS environment. The questions driving the project are: How to build a device that can deliver satisfactory analytical merit (e.g., sensitivity, accuracy, precision), while maintaining reliable, inexpensive, and simple operation. The research involves investigation into detection methods and state of the art instrumentation available for nitrite, production trends in chemical total analysis systems, and centers around larger questions surrounding invention and innovation. The first steps towards such a device are benchtop prototyping of the detection and fluidic modules, their integration with wet chemistry, and the validation of the analytical process carried out by the system. The project approaches the objectives with a design that relies on commercially available components and consumables and is modular and adaptable for future possible configurations. To this end, the benchtop prototype was developed as an opto-fluidic system for automated colorimetric detection. With the exception of two custom-built PVC adaptors, the entire system was built with off-the-shelf parts for around $1,000. In addition to utilizing easily replaceable components, the system was tested using commercially available and pre-made reagents based on proven chemistry (Griess assay for nitrite). Preliminary results suggest the analytical process is capable of detecting sub-micromolar nitrite concentrations (limit of detection equal to 0.18 µM) at appreciable precision, sensitivity, and accuracy in comparison to commercial instruments

Realistic measurement uncertainties for marine macronutrient measurements conducted using gas segmented flow and Lab-on-Chip techniques

Highlights • Accounting for systematic bias is required for a realistic analytical uncertainty • Gas segmented flow techniques achieved a combined uncertainties of 1-4 % • Lab-on-Chip nitrate + nitrite analysers achieved a combined uncertainties < 5% Abstract Accurate and precise measurements of marine macronutrient concentrations are fundamental to our understanding of biogeochemical cycles in the ocean. Quantifying the measurement uncertainty associated with macronutrient measurements remains a challenge. Large systematic biases (up to 10 %) have been identified between datasets, restricting the ability of marine biogeochemists to distinguish between the effects of environmental processes and analytical uncertainty. In this study we combine the routine analyses of certified reference materials (CRMs) with the application of a simple statistical technique to quantify the combined (random + systematic) measurement uncertainty associated with marine macronutrient measurements using gas segmented flow techniques. We demonstrate that it is realistic to achieve combined uncertainties of ~1-4 % for nitrate + nitrite (ΣNOx), phosphate (PO43-) and silicic acid (Si(OH)4) measurements. This approach requires only the routine analyses of CRMs (i.e. it does not require inter-comparison exercises). As CRMs for marine macronutrients are now commercially available, it is advocated that this simple approach can improve the comparability of marine macronutrient datasets and therefore should be adopted as ‘best practice’. Novel autonomous Lab-on-Chip (LoC) technology is currently maturing to a point where it will soon become part of the marine chemist’s standard analytical toolkit used to determine marine macronutrient concentrations. Therefore, it is critical that a complete understanding of the measurement uncertainty of data produced by LoC analysers is achieved. In this study we analysed CRMs using 7 different LoC ΣNOx analysers to estimate a combined measurement uncertainty of < 5%. This demonstrates that with high quality manufacturing and laboratory practices, LoC analysers routinely produce high quality measurements of marine macronutrient concentrations

Lab-on-chip for in situ analysis of nutrients in the deep sea

Microfluidic reagent-based nutrient sensors offer a promising technology to address the global undersampling of ocean chemistry but have so far not been shown to operate in the deep sea (>200 m). We report a new family of miniaturized lab-on-chip (LOC) colorimetric analyzers making in situ nitrate and phosphate measurements from the surface ocean to the deep sea (>4800 m). This new technology gives users a new low-cost, high-performance tool for measuring chemistry in hyperbaric environments. Using a combination of laboratory verification and field-based tests, we demonstrate that the analyzers are capable of in situ measurements during profiling that are comparable to laboratory-based analyses. The sensors feature a novel and efficient inertial-flow mixer that increases the mixing efficiency and reduces the back pressure and flushing time compared to a previously used serpentine mixing channel. Four separate replicate units of the nitrate and phosphate sensor were calibrated in the laboratory and showed an average limit of detection of 0.03 μM for nitrate and 0.016 μM for phosphate. Three on-chip optical absorption cell lengths provide a large linear range (to >750 μM (10.5 mg/L-N) for nitrate and >15 μM (0.47 mg/L-P) for phosphate), making the instruments suitable for typical concentrations in both ocean and freshwater aquatic environments. The LOC systems automatically collected a series of deep-sea nitrate and phosphate profiles in the northeast Atlantic while attached to a conductivity temperature depth (CTD) rosette, and the LOC nitrate sensor was attached to a PROVOR profiling float to conduct automated nitrate profiles in the Mediterranean Sea

Development of In-Situ sensors for Nutrients in Marine Waters

In particular, I focussed on the development, improvement, and deployment of autonomous analysers for on-site monitoring of nutrients that have shown promise for further application in field monitoring either on shore or in the water column (in-situ).Macronutrients include nitrate, nitrite, phosphate, and silicic acid. These macronutrients are important for primary production, and silicic acid is essential for the hard tissue of siliceous phytoplankton (e.g., diatoms). The importance of macronutrients for photosynthesis means that their supply rates have major implications for the functioning of phytoplankton communities, which are considered an carbon dioxide sink in the ocean. The analysis of macronutrients is still more difficult than physical parameters such as salinity and temperature or dissolved oxygen. In addition, analysis of macronutrients in a complex matrix such as seawater can be challenging because their concentrations are typically low, in the range from nanomolar to micromolar. This thesis reports on the development of an electrochemical approach for the determination of orthophosphate in estuarine and seawater, reporting the initial investigations of our technique with a further application in an automated analyser which uses a bipotentiostat approach to enhance data quality. In addition, the improvement of on-site determination of macronutrients with a long-term application in estuarine and coastal water is reported

Analysis of relevant technical issues and deficiencies of the existing sensors and related initiatives currently set and working in marine environment. New generation technologies for cost-effective sensors

The last decade has seen significant growth in the field of sensor networks, which are currently collecting large amounts of environmental data. This data needs to be collected, processed, stored and made available for analysis and interpretation in a manner which is meaningful and accessible to end users and stakeholders with a range of requirements, including government agencies, environmental agencies, the research community, industry users and the public. The COMMONSENSE project aims to develop and provide cost-effective, multi-functional innovative sensors to perform reliable in-situ measurements in the marine environment. The sensors will be easily usable across several platforms, and will focus on key parameters including eutrophication, heavy metal contaminants, marine litter (microplastics) and underwater noise descriptors of the MSFD. The aims of Tasks 2.1 and 2.2 which comprise the work of this deliverable are: • To obtain a comprehensive understanding and an up-to-date state of the art of existing sensors. • To provide a working basis on “new generation” technologies in order to develop cost-effective sensors suitable for large-scale production. This deliverable will consist of an analysis of state-of-the-art solutions for the different sensors and data platforms related with COMMONSENSE project. An analysis of relevant technical issues and deficiencies of existing sensors and related initiatives currently set and working in marine environment will be performed. Existing solutions will be studied to determine the main limitations to be considered during novel sensor developments in further WP’s. Objectives & Rationale The objectives of deliverable 2.1 are: • To create a solid and robust basis for finding cheaper and innovative ways of gathering data. This is preparatory for the activities in other WPs: for WP4 (Transversal Sensor development and Sensor Integration), for WP(5-8) (Novel Sensors) to develop cost-effective sensors suitable for large-scale production, reducing costs of data collection (compared to commercially available sensors), increasing data access availability for WP9 (Field testing) when the deployment of new sensors will be drawn and then realized

Benthic element cycling, fluxes and transport of solutes across the benthic boundary layer in the Mauritanian oxygen minimum zone, (SFB754) - Cruise No. M107, May 30 - July 03, 2014, Fortaleza (Brazil) - Las Palmas (Spain)

Summary A detailed multi-disciplinary research program was conducted at the Mauritanian oxygen minimum zone (OMZ). Investigations were primarily performed along a depth transect at 18°20’ N. In this area upwelling of cold, nutrient-rich deep water is strongly seasonal, predominating from April until December. Major aim was to advance understanding of how OMZs are maintained and to determine feedbacks of benthic nutrient release on the currently expanding Mauritanian OMZ under such conditions. Major focus was on (i) variability of benthic nutrient release in response to hydrodynamic forcing and regional differences in geochemistry, (ii) diapycnal and advective fluxes of nutrients, trace metals, and radio-tracer between the sediments and the stratified interior ocean as well as their entrainment into the surface mixed layer and (iii) processes involved in the respective benthic and pelagic N, Fe, and P cycles. The working program in the water column comprised a total of 73 CTD casts, 38 microstructure CTD- and 17 in situ pump deployments. Moorings and Glider were deployed at 18°20’ N and 19°50’ N. Furthermore, in the northern working area ADCP-transects and casts of Underway CTDs were conducted to follow upwelling-induced frontal systems. In situ benthic fluxes of nutrients and oxygen were conducted using the Biogeochemical Observatories BIGO I and BIGO II comprising a total of 9 deployments. Further sediment samples for biogeochemical, investigations were obtained during the deployment of 22 casts of a video guided Multiple Corer (MUC). All deployments were successful and the envisaged data and samples were collected

Chemolithotrophic and chemohetero- trophic microorganisms in sediment - and rock-hosted hydrothermal systems

Deep-sea hydrothermal vent systems are highly productive ecosystems, where reduced energy sources fuel complex communities of microorganisms, invertebrates and vertebrates. Since decades the oxidation of methane, hydrogen and inorganic sulfur compounds has been extensively studied. However, the role of inorganic nitrogen and of organic compounds as energy source has been investigated only scarcely in hydrothermal fluids, in particular at the sea floor, where hydrothermal fluids exit subsurface. The aim of my thesis was to shed light on these under-investigated topics. In my first project I studied nitrification and the involved microbes that are associated with large, nitrate-respiring and sulfur-oxidizing bacteria (SOB) of the genus Beggiatoa. These SOB formed mats and covered sulfide- and ammonia-rich hydrothermal sediments in the Guaymas Basin. In these mats, nitrification rates were measured using 15N-labeled ammonium. With up to 605 μmol N l-1 mat d-1 the nitrification rates were the highest measured for a deep-sea ecosystem. Diversity and quantitative PCR of the ammonia monooxygenase subunit A gene (amoA) indicated association of ammonia-oxidizing archaea (AOA) and bacteria (AOB) with Beggiatoa mats. In line with this, single cells of AOB and potentially ammonia-oxidizing thaumarchaotes were attached to narrow Beggiatoa-like filaments. Nitrite oxidizing bacteria were also found. Nitrifying bacteria associated with Beggiatoa mats that respire nitrate to ammonium (DNRA) could display a syntrophic consortium that internally cycle nitrogen and thereby reduce loss of bioavailable nitrogen. However, it is not clear whether large SOB in general respire nitrate also to dinitrogen. Therefore, I analyzed the genetic potential of the large SOB “Candidatus Thiomargarita nelsonii”, a close relative of Beggiatoa. The comparison to four other Beggiatoaceae identified genes for both denitrification and DNRA in “Ca. T nelsonii” and three other Beggiatoaceae. This indicates that both pathways are widely distributed among large SOB and questions the hypothesis of internal N-cycling in mats of large SOB. In my third project I investigated the microbial consumption of organic compounds that are produced in hydrothermal systems. In particular I studied acetate-assimilating heterotrophic communities in the diffuse fluids (temperature range of 4-72°C) of two rock-hosted hydrothermal systems. 16S rRNA gene-based diversity analysis and fluorescence in situ hybridization (FISH) showed that either Gammaproteobacteria or Epsilonproteobacteria rapidly grew during short-term (8-12 h) incubations with 13C-acetate. Single cells of both groups incorporated 13C-acetate as shown by nanoSIMS. Marinobacter spp. and a novel group among the Nautiliales could be heterotrophs in these systems. These are potential r-strategists that quickly respond to the fluctuating availabilities of energy sources in hydrothermal fluids

Towards improved monitoring of offshore carbon storage: A real-world field experiment detecting a controlled sub-seafloor CO2 release

Carbon capture and storage (CCS) is a key technology to reduce carbon dioxide (CO2) emissions from industrial processes in a feasible, substantial, and timely manner. For geological CO2 storage to be safe, reliable, and accepted by society, robust strategies for CO2 leakage detection, quantification and management are crucial. The STEMM-CCS (Strategies for Environmental Monitoring of Marine Carbon Capture and Storage) project aimed to provide techniques and understanding to enable and inform cost-effective monitoring of CCS sites in the marine environment. A controlled CO2 release experiment was carried out in the central North Sea, designed to mimic an unintended emission of CO2 from a subsurface CO2 storage site to the seafloor. A total of 675 kg of CO2 were released into the shallow sediments (∼3 m below seafloor), at flow rates between 6 and 143 kg/d. A combination of novel techniques, adapted versions of existing techniques, and well-proven standard techniques were used to detect, characterise and quantify gaseous and dissolved CO2 in the sediments and the overlying seawater. This paper provides an overview of this ambitious field experiment. We describe the preparatory work prior to the release experiment, the experimental layout and procedures, the methods tested, and summarise the main results and the lessons learnt