Direct visualization of mucus production by the cold-water coral <i>Lophelia pertusa</i> with digital holographic microscopy

<i>Lophelia pertusa</i> is the dominant reef-building organism of cold-water coral reefs, and is known to produce significant amounts of mucus, which could involve an important metabolic cost. Mucus is involved in particle removal and feeding processes, yet the triggers and dynamics of mucus production are currently still poorly described because the existing tools to study these processes are not appropriate. Using a novel microscopic technique—digital holographic microscopy (DHM)–we studied the mucus release of <i>L</i>. <i>pertusa</i> under various experimental conditions. DHM technology permits µm-scale observations and allows the visualization of transparent mucoid substances in real time without staining. Fragments of <i>L</i>. <i>pertusa</i> were first maintained in flow-through chambers without stressors and imaged with DHM, then exposed to various stressors (suspended particles, particulate food and air exposure) and re-imaged. Under non-stressed conditions no release of mucus was observed, whilst mucus strings and sheaths were produced in response to suspended particles (activated charcoal and drill cuttings sediment) i.e. in a stressed condition. Mucus strings and so-called ‘string balls’ were also observed in response to exposure to particulate food (brine shrimp <i>Artemia salina</i>). Upon air-exposure, mucus production was clearly visible once the fragments were returned to the flow chamber. Distinct optical properties such as optical path length difference (OPD) were measured with DHM in response to the various stimuli suggesting that different mucus types are produced by <i>L</i>. <i>pertusa</i>. Mucus produced to reject particles is similar in refractive index to the surrounding seawater, suggesting that the energy content of this mucus is low. In contrast, mucus produced in response to either food particle addition or air exposure had a higher refractive index, suggesting a higher metabolic investment in the production of these mucoid substances. This paper shows for the first time the potential of DHM technology for the detection, characterization and quantification of mucus production through OPD measurements in <i>L</i>. <i>pertusa</i>

Imaging-in-flow: digital holographic microscopy as a novel tool to detect and classify nanoplanktonic organisms

Traditional taxonomic identification of planktonic organisms is based on light microscopy, which is both time-consuming and tedious. In response, novel ways of automated (machine) identification, such as flow cytometry, have been investigated over the last two decades. To improve the taxonomic resolution of particle analysis, recent developments have focused on "imaging-in-flow," i.e., the ability to acquire microscopic images of planktonic cells in a flow-through mode. Imaging-in-flow systems are traditionally based on classical brightfield microscopy and are faced with a number of issues that decrease the classification performance and accuracy (e. g., projection variance of cells, migration of cells out of the focus plane). Here, we demonstrate that a combination of digital holographic microscopy (DHM) with imaging-in-flow can improve the detection and classification of planktonic organisms. In addition to light intensity information, DHM provides quantitative phase information, which generates an additional and independent set of features that can be used in classification algorithms. Moreover, the capability of digitally refocusing greatly increases the depth of field, enables a more accurate focusing of cells, and reduces the effects of position variance. Nanoplanktonic organisms similar in shape were successfully classified from images captured with an off-axis DHM with partial coherence. Textural features based on DHM phase information proved more efficient in separating the three tested phytoplankton species compared with shape-based features or textural features based on light intensity. An overall classification score of 92.4% demonstrates the potential of holographic-based imaging-in-flow for similar looking organisms in the nanoplankton range

Long-distance electron transport occurs globally in marine sediments

Recently, long filamentous bacteria have been reported conducting electrons over centimetre distances in marine sediments. These so-called cable bacteria perform an electrogenic form of sulfur oxidation, whereby long-distance electron transport links sulfide oxidation in deeper sediment horizons to oxygen reduction in the upper millimetres of the sediment. Electrogenic sulfur oxidation exerts a strong impact on the local sediment biogeochemistry, but it is currently unknown how prevalent the process is within the seafloor. Here we provide a state-of-the-art assessment of its global distribution by combining new field observations with previous reports from the literature. This synthesis demonstrates that electrogenic sulfur oxidation, and hence microbial long-distance electron transport, is a widespread phenomenon in the present-day seafloor. The process is found in coastal sediments within different climate zones (off the Netherlands, Greenland, the USA, Australia) and thrives on a range of different coastal habitats (estuaries, salt marshes, mangroves, coastal hypoxic basins, intertidal flats). The combination of a widespread occurrence and a strong local geochemical imprint suggests that electrogenic sulfur oxidation could be an important, and hitherto overlooked, component of the marine cycle of carbon, sulfur and other elements

Mineral formation induced by cable bacteria performing long-distance electron transport in marine sediments

Cable bacteria are multicellular, filamentous microorganisms that are capable of transporting electrons over centimeter-scale distances. Although recently discovered, these bacteria appear to be widely present in the seafloor, and when active they exert a strong imprint on the local geochemistry. In particular, their electrogenic metabolism induces unusually strong pH excursions in aquatic sediments, which induces considerable mineral dissolution, and subsequent mineral reprecipitation. However, at present, it is unknown whether and how cable bacteria play an active or direct role in the mineral reprecipitation process. To this end we present an explorative study of the formation of sedimentary minerals in and near filamentous cable bacteria using a combined approach of electron microscopy and spectroscopic techniques. Our observations reveal the formation of polyphosphate granules within the cells and two different types of biomineral formation directly associated with multicellular filaments of these cable bacteria: (i) the attachment and incorporation of clay particles in a coating surrounding the bacteria and (ii) encrustation of the cell envelope by iron minerals. These findings suggest a complex interaction between cable bacteria and the surrounding sediment matrix, and a substantial imprint of the electrogenic metabolism on mineral diagenesis and sedimentary biogeochemical cycling. In particular, the encrustation process leaves many open questions for further research. For example, we hypothesize that the complete encrustation of filaments might create a diffusion barrier and negatively impact the metabolism of the cable bacteria.</p

The impact of sedimentary alkalinity release on the water column CO₂ system in the North Sea

It has been previously proposed that alkalinity release from sediments can play an important role in the carbonate dynamics on continental shelves, lowering the <i>p</i>CO₂ of seawater and hence increasing the CO₂ uptake from the atmosphere. To test this hypothesis, sedimentary alkalinity generation was quantified within cohesive and permeable sediments across the North Sea during two cruises in September 2011 (basin-wide) and June 2012 (Dutch coastal zone). Benthic fluxes of oxygen (O₂), alkalinity (<i>A</i>_T) and dissolved inorganic carbon (DIC) were determined using shipboard closed sediment incubations. Our results show that sediments can form an important source of alkalinity for the overlying water, particularly in the shallow southern North Sea, where high <i>A</i>_T and DIC fluxes were recorded in near-shore sediments of the Belgian, Dutch and German coastal zone. In contrast, fluxes of <i>A</i>_T and DIC are substantially lower in the deeper, seasonally stratified, northern part of the North Sea. Based on the data collected, we performed a model analysis to constrain the main pathways of alkalinity generation in the sediment, and to quantify how sedimentary alkalinity drives atmospheric CO₂ uptake in the southern North Sea. Overall, our results show that sedimentary alkalinity generation should be regarded as a key component in the CO₂ dynamics of shallow coastal systems

An assessment of the precision and confidence of aquatic eddy correlation measurements

The quantification of benthic fluxes with the aquatic eddy correlation (EC) technique is based on simultaneous measurement of the current velocity and a targeted bottom water parameter (e. g., O-2, temperature). High-frequency measurements (64Hz) are performed at a single point above the seafloor using an acoustic Doppler velocimeter (ADV) and a fast-responding sensor. The advantages of aquatic EC technique are that 1) it is noninvasive, 2) it integrates fluxes over a large area, and 3) it accounts for in situ hydrodynamics. The aquatic EC has gained acceptance as a powerful technique; however, an accurate assessment of the errors introduced by the spatial alignment of velocity and water constituent measurements and by their different response times is still needed. Here, this paper discusses uncertainties and biases in the data treatment based on oxygen EC flux measurements in a large-scale flume facility with well-constrained hydrodynamics. These observations are used to review data processing procedures and to recommend improved deployment methods, thus improving the precision, reliability, and confidence of EC measurements. Specifically, this study demonstrates that 1) the alignment of the time series based on maximum cross correlation improved the precision of EC flux estimations; 2) an oxygen sensor with a response time of <0.4 s facilitates accurate EC fluxes estimates in turbulence regimes corresponding to horizontal velocities <11 cm s(-1); and 3) the smallest possible distance (<1 cm) between the oxygen sensor and the ADV's sampling volume is important for accurate EC flux estimates, especially when the flow direction is perpendicular to the sensor's orientation

Natural occurrence of microbial sulphur oxidation by long-range electron transport in the seafloor

Recently, a novel mode of sulphur oxidation was described in marine sediments, in which sulphide oxidation in deeper anoxic layers was electrically coupled to oxygen reduction at the sediment surface. Subsequent experimental evidence identified that long filamentous bacteria belonging to the family Desulfobulbaceae likely mediated the electron transport across the centimetre-scale distances. Such long-range electron transfer challenges some long-held views in microbial ecology and could have profound implications for sulphur cycling in marine sediments. But, so far, this process of electrogenic sulphur oxidation has been documented only in laboratory experiments and so its imprint on the seafloor remains unknown. Here we show that the geochemical signature of electrogenic sulphur oxidation occurs in a variety of coastal sediment environments, including a salt marsh, a seasonally hypoxic basin, and a subtidal coastal mud plain. In all cases, electrogenic sulphur oxidation was detected together with an abundance of Desulfobulbaceae filaments. Complementary laboratory experiments in intertidal sands demonstrated that mechanical disturbance by bioturbating fauna destroys the electrogenic sulphur oxidation signal. A survey of published geochemical data and 16S rRNA gene sequences identified that electrogenic sulphide oxidation is likely present in a variety of marine sediments with high sulphide generation and restricted bioturbation, such as mangrove swamps, aquaculture areas, seasonally hypoxic basins, cold sulphide seeps and possibly hydrothermal vent environments. This study shows for the first time that electrogenic sulphur oxidation occurs in a wide range of marine sediments and that bioturbation may exert a dominant control on its natural distribution

Phosphorus cycling and burial in sediments of a seasonally hypoxic marine basin

Recycling of phosphorus (P) from sediments contributes to the development of bottom-water hypoxia in many coastal systems. Here, we present results of a year-long assessment of P dynamics in sediments of a seasonally hypoxic coastal marine basin (Lake Grevelingen, the Netherlands) in 2012. Sequential phosphorus extractions (SEDEX) and X-ray absorption spectroscopy (XAS) indicate that P was adsorbed to Fe-(III)-(oxyhydr)oxides when cable bacteria were active in the surface sediments in spring. With the onset of summer hypoxia, sulphide-induced dissolution of the Fe-(III)-(oxyhydr)oxides led to P release to the pore water and overlying water. The similarity in authigenic Ca-P concentrations in the sediment and suspended matter suggest that Ca-P is not formed in situ. The P burial efficiency was ≤ 32%. Hypoxia-driven sedimentary P recycling had a major impact on the water-column chemistry in the basin in 2012. Water-column monitoring data indicate up to ninefold higher surface water concentrations of phosphate in the basin in the late 1970s and a stronger hypoxia-driven seasonal P release from the sediment. The amplified release of P from the sediment in the past is attributed to the presence of a larger pool of Fe-bound P in the basin prior to the first onset of hypoxia. Given that P is not limiting, primary production in the basin has not been affected by the decadal changes in P availability and recycling over the past 40 years. The changes in P dynamics on decadal time scales wer