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Humans and pigs share a close contact relationship, similar biological traits, and one of the highest estimated number of viruses compared to other mammalian species. The contribution and directionality of viral exchange between humans and pigs remain unclear for some of these viruses, but their transmission routes are important to characterize in order to prevent outbreaks of disease in both host species. This review collects and assesses the evidence to determine the likely transmission route of 27 viruses between humans and pigs.

Sediment resuspension is a common process in dynamic coastal settings, but its implications for remineralization and carbon turnover in seagrass meadows are poorly understood. Here, we assessed eelgrass Zostera marina metabolism in the Baltic Sea (SW Finland) using benthic flume-chambers and aquatic eddy covariance to critically evaluate the drivers of benthic O-2 exchange during dynamic flow conditions. During quiescent weather conditions, the 2 methods resolved similar metabolic rates and net ecosystem autotrophy (+/- 11. However, elevated flow speeds and sediment resuspension halfway through the study induced a 5-fold increase in the O-2 uptake rates measured by eddy covariance, whereas chamber fluxes remained relatively unchanged. Following particle resettlement, instruments were redeployed and the benthic O-2 uptake resolved by both techniques was just similar to 302 uptake up to 6-fold, mainly due to the reoxidation of reduced compounds (e.g. FeSx). This process was fully captured by the eddy O-2 fluxes, but not by the chamber incubation. Consequently, the chamber and eddy net ecosystem metabolism amounted to - 17 and -824 mmol C m(-2), respectively, throughout the study period. The rapid reoxidation and long-term effects of resuspension on benthic O-2 dynamics highlight the importance of fully capturing dynamic conditions when assessing the overall carbon turnover in coastal habitats. Future studies on the biogeochemical functioning of coastal environments should aim to capture the natural frequency and duration of resuspension events. Sediment resuspension is a common process in dynamic coastal settings, but its implications for remineralization and carbon turnover in seagrass meadows are poorly understood. Here, we assessed eelgrass Zostera marina metabolism in the Baltic Sea (SW Finland) using benthic flume-chambers and aquatic eddy covariance to critically evaluate the drivers of benthic O2 exchange during dynamic flow conditions. During quiescent weather conditions, the 2 methods resolved similar metabolic rates and net ecosystem autotrophy (±11% of each other). However, elevated flow speeds and sediment resuspension halfway through the study induced a 5-fold increase in the O2 uptake rates measured by eddy covariance, whereas chamber fluxes remained relatively unchanged. Following particle resettlement, instruments were redeployed and the benthic O2 uptake resolved by both techniques was just ~30% of the values measured before resuspension. Laboratory investigations revealed sediment resuspension could potentially increase benthic O2 uptake up to 6fold, mainly due to the reoxidation of reduced compounds (e.g. FeSx). This process was fully captured by the eddy O2 fluxes, but not by the chamber incubation. Consequently, the chamber and eddy net ecosystem metabolism amounted to -17 and -824 mmol C m-2, respectively, throughout the study period. The rapid reoxidation and long-term effects of resuspension on benthic O2 dynamics highlight the importance of fully capturing dynamic conditions when assessing the overall carbon turnover in coastal habitats. Future studies on the biogeochemical functioning of coastal environments should aim to capture the natural frequency and duration of resuspension events.

Hadal trench sediments are hotspots of biogeochemical activity in the deep sea, but the biogeochemical and ecological factors that shape benthic hadal microbial communities remain unknown. Here, we sampled ten hadal sites from two trench regions with a vertical resolution of down to 1 cm. We sequenced 16S rRNA gene amplicons using universal and archaea-specific primer sets and compared the results to biogeochemical parameters. Despite bathymetric and depositional heterogeneity we found a high similarity of microbial communities within each of the two trench axes, while composition at the phylum level varied strongly with sediment depth in conjunction with the redox stratification into oxic, nitrogenous, and ferruginous zones. As a result, communities of a given sediment horizon were more similar to each other across a distance of hundreds of kilometers within each trench, than to those of adjacent horizons from the same sites separated only by centimeters. Total organic carbon content statistically only explained a small part of the variation within and between trenches, and did not explain the community differences observed between the hadal and adjacent shallower sites. Anaerobic taxa increased in abundance at the top of the ferruginous zone, seeded by organisms deposited at the sediment surface and surviving burial through the upper redox zones. While an influence of other potential factors such as geographic isolation, hydrostatic pressure, and non-steady state depositional regimes could not be discerned, redox stratification and diagenesis appear to be the main selective forces that structure community composition in hadal sediments.

Background: Deep brain stimulation (DBS) of the dorsal subthalamic nucleus (STN) is a validated neurosurgical treatment of Parkinson’s Disease (PD). To investigate the mechanism of action, including potential DBS induced neuroplasticity, we have previously used a minipig model of Parkinson’s Disease, although the basal ganglia circuitry was not elucidated in detail.Aim: To describe the cortical projections from the primary motor cortex (M1) to the basal ganglia and confirm the presence of a cortico-striatal pathway and a hyperdirect pathway to the subthalamic nucleus, respectively, which is known to exist in primates.Materials and Methods: Five female Göttingen minipigs were injected into the primary motor cortex (n = 4) and adjacent prefrontal cortex (n = 1) with the anterograde neuronal tracer, Biotinylated Dextran Amine (BDA). 4 weeks later the animals were sacrificed and the brains cryosectioned into 30 μm thick coronal sections for subsequent microscopic analysis.Results: The hyperdirect axonal connections from the primary motor cortex were seen to terminate in the dorsolateral STN, whereas the axonal projections from the prefrontal cortex terminated medially in the STN. Furthermore, striatal tracing from the motor cortex was especially prominent in the dorsolateral putamen and less so in the dorsolateral caudate nucleus. The prefrontal efferents were concentrated mainly in the caudate nucleus and to a smaller degree in the juxtacapsular dorsal putamen, but they were also found in the nucleus accumbens and ventral prefrontal cortex.Discussion: The organization of the Göttingen minipig basal ganglia circuitry is in accordance with previous descriptions in primates. The existence of a cortico-striatal and hyperdirect basal ganglia pathway in this non-primate, large animal model may accordingly permit further translational studies on STN-DBS induced neuroplasticity of major relevance for future DBS treatments.

Copepod carcasses are prevalent in marine ecosystems and might represent an important component of the sinking flux of particulate organic carbon in the ocean. The extent to which copepod carcasses contribute to the biological carbon pump is controlled by different environmental factors, including temperature. However, the effect of temperature on the longer-term kinetics of carbon mineralization of copepod carcasses is not well-studied. We conducted laboratory experiments to quantify the carbon mineralization associated with sinking carcasses of the cosmopolitan copepod Acartia tonsa through aerobic microbial respiration at 5 temperatures (20, 16, 12, 8, and 4 degrees C). Microbial respiration rates associated with the carcasses were positively correlated with temperature and characterized by an initial short lag-phase, a rapid increase to a maximum rate, and a subsequent gradual decline in the rate of degradation. On average, 50-12 d at 20 degrees C, versus >60 d at 4 degrees C. During the incubations, most carbon mineralization occurred in the ambient seawater, likely fueled by dissolved organic carbon leaking from the carcasses into the surrounding seawater. Extrapolating measured carbon turnover and sinking rates suggests that at 20 degrees C, the mineralization of sinking copepod carcasses is constrained to the surface ocean. In contrast, at 4 degrees C, sinking copepod carcasses can reach the deep ocean before they have been completely degraded. Hence, in low-temperature regions, copepod carcasses may represent an important agent for carbon export through the biological carbon pump. Copepod carcasses are prevalent in marine ecosystems and might represent an important component of the sinking flux of particulate organic carbon in the ocean. The extent to which copepod carcasses contribute to the biological carbon pump is controlled by different environmental factors, including temperature. However, the effect of temperature on the longer-term kinetics of carbon mineralization of copepod carcasses is not well-studied. We conducted laboratory experiments to quantify the carbon mineralization associated with sinking carcasses of the cosmopolitan copepod Acartia tonsa through aerobic microbial respiration at 5 temperatures (20, 16, 12, 8, and 4°C). Microbial respiration rates associated with the carcasses were positively correlated with temperature and characterized by an initial short lag-phase, a rapid increase to a maximum rate, and a subsequent gradual decline in the rate of degradation. On average, 50% of the total carbon of the carcasses was mineralized within 6-12 d at 20°C, versus >60 d at 4°C. During the incubations, most carbon mineralization occurred in the ambient seawater, likely fueled by dissolved organic carbon leaking from the carcasses into the surrounding seawater. Extrapolating measured carbon turnover and sinking rates suggests that at 20°C, the mineralization of sinking copepod carcasses is constrained to the surface ocean. In contrast, at 4°C, sinking copepod carcasses can reach the deep ocean before they have been completely degraded. Hence, in lowtemperature regions, copepod carcasses may represent an important agent for carbon export through the biological carbon pump.

The water flow through sponges is regulated by their contractile behaviour including contraction and expansion of the aquiferous system, which leads to shifting oxygen levels in the sponge interior. Still, knowledge of spatial and temporal anoxia in sponges is lacking, but important in elucidating interactions between sponge hosts and their microbiomes. We combined 2-D luminescence lifetime imaging of oxygen with simultaneous time-lapse recordings of the sponge exhalant opening (osculum) to unveil temporal as well as spatial oxygen dynamics caused by contractile behaviour in single-osculum explants of the demosponge Halichondria panicea. The present study reveals an intrinsic concentric deoxygenation pattern in explants during episodes of osculum contraction generating an oxygen gradient with increasing concentrations towards the explant periphery. Four sponge explants faced 25 episodes with substantial changes in internal oxygen and anoxia which prevailed for 4.4 h of the total 92.0 h observation period. The 2-D images revealed that the total area of the explant experiencing anoxia during periods of osculum contraction–expansion varied between 0.01 and 13.22% and was on average 7.4 ± 4.4% for all sponge explants. Furthermore, oxygen respiration, as approximated by the rate of change of oxygen concentration during deoxygenation of the explant interior, was similar throughout the oxic parts of the explant base. The resolved 2-D dynamics provide an unprecedented insight into the internal O2 distribution of sponges and complement the traditional point measurements of oxygen sensors.

The pituitary is involved in the regulation of endocrine homeostasis. Therefore, animal models of pituitary disease based on a thorough knowledge of pituitary anatomy are of great importance. Accordingly, we aimed to perform a qualitative and quantitative description of polypeptide hormone secreting cellular components of the Göttingen minipig adenohypophysis using immunohistochemistry and stereology. Estimates of the total number of cells immune-stained for adrenocorticotrophic hormone (ACTH), prolactin (PRL), and growth hormone (GH) were obtained with the optical fractionator technique using Stereo Investigator software. Moreover, 3D reconstructions of cell distribution were made. We estimated that the normal minipig adenohypophysis contains, on average, 5.6 million GH, 3.5 million PRL, and 2.4 million ACTH producing cells. The ACTH producing cells were widely distributed, while the PRL and GH producing cells were located in clusters in the central and lateral regions of the adenohypophysis. The morphology of the hormone producing cells also differs. We visualized a clear difference in the numerical density of hormone producing cells throughout the adenohypophysis. The relative proportions of the cells analyzed in our experiment are comparable to those observed in humans, primates, and rodents; however, the distribution of cells differs among species. The distribution of GH cells in the minipig is similar to that in humans, while the PRL and ACTH cell distributions differ. The volume of the pituitary is slightly smaller than that of humans. These data provide a framework for future large animal experimentation on pituitary function in health and disease.

Particle mixing and irrigation of the seabed by benthic fauna (bioturbation) have major impacts on ecosystem functions such as remineralization of organic matter and sediment-water exchange. As a tribute to Prof. Gaston Desrosiers by the Nereis Park association, eighteen laboratories carried out a collaborative experiment to acquire a global snapshot of particle reworking by the polychaete Hediste diversicolor at 16 sites surrounding the Northern Atlantic. Organisms and soft sediments were collected during May – July at different geographical locations and, using a common laboratory protocol, particulate fluorescent tracers (‘luminophores’) were used to quantify particle transport over a 10-day period. Particle mixing was quantified using the maximum penetration depth of tracers (MPD), particle diffusive coefficients (Db), and non-local transport coefficients (r). Non-local coefficients (reflecting centimeter scale transport steps) ranged from 0.4 to 15 yr−1, and were not correlated across sites with any measured biological (biomass, biovolume) or environmental parameters (temperature, grain size, organic matter). Maximum penetration depths (MPD) averaged ~10.7 cm (6.5–14.5 cm), and were similar to the global average bioturbation depth inferred from short-lived radiochemical tracers. MPD was also not correlated with measures of size (individual biomass), but increased with grain size and decreased with temperature. Biodiffusion (Db) correlated inversely with individual biomass (size) and directly with temperature over the environmental range (Q10 ~ 1.7; 5–21 °C). The transport data were comparable in magnitude to rates reported for localized H. diversicolor populations of similar size, and confirmed some but not all correlations between sediment reworking and biological and environmental variables found in previous studies. The results imply that measures of particle reworking activities of a species from a single location can be generally extrapolated to different populations at similar conditions.

It is generally anticipated that particulate organic carbon (POC) for most part is degraded by attached microorganisms during the descent of “marine snow” aggregates toward the deep sea. There is, however, increasing evidence that fresh aggregates can reach great depth and sustain relatively high biological activity in the deep sea. Using a novel high‐pressure setup, we tested the hypothesis that increasing levels of hydrostatic pressure inhibit POC degradation in aggregates rapidly sinking to the ocean interior. Respiration activity, a proxy for POC degradation, was measured directly and continuously at up to 100 MPa (corresponding to 10 km water depth) in a rotating pressure tank that keeps the aggregates in a sinking mode. Model diatom‐bacteria aggregates, cultures of the aggregate‐forming diatom Skeletonema marinoi, and seawater microbial communities devoid of diatoms showed incomplete and complete inhibition of respiration activity when exposed to pressure levels of 10–50 and 60–100 MPa, respectively. This implies reduced POC degradation and hence enhanced POC export to hadal trenches through fast‐sinking, pressure‐exposed aggregates. Notably, continuous respiration measurements at ≥50 MPa revealed curved instead of linear oxygen time series whenever S. marinoi was present, which was not captured by discrete respiration measurements. These curvatures correspond to alternating phases of high and low respiration activity likely connected to pressure effects on unidentified metabolic processes in S. marinoi.

It is well established that benthic infauna alter sediment biogeochemistry, but the importance of their behavioural patterns in biogeochemical processes has only recently been fully appreciated. Using arrays of O2 microoptodes, siphon imaging, accelerometer loggers and different incubation approaches, we investigated the importance of behavioural patterns of the soft-shell clam Mya arenaria for benthic O2 and nitrogen dynamics. The investigations resolved a new behavioural component of buried M. arenaria: pedal water ejection (PWE). During PWE, the clams contracted their valves while briefly closing the siphon to expel oxic water previously accumulated within the mantle cavity through the pedal gape. Consequently, an upward-moving oxic plume embedded most of the shell, and oxia at the shell−sediment interface was observed for 15% of the total time of investigation. The buried clams displayed 2 additional behavioural stages: resting (R) and ventilation (V) that occurred for 57 and 28% of the time, respectively. During the V and PWE stages, the siphon of the clam was elongated above the sediment surface and surrounded by a dynamic oxic halo; both behavioural stages thus markedly increase the sediment oxygenation. The resolved irrigation patterns enhanced the benthic exchange of O2 and NH4 + as well as the benthic denitrification rate, presumably via increased benthic NO3 − supply and production. Particularly, the extensive sediment irrigation induced by the PWE stage has important implications for benthic nutrient cycling as well as for sediment oxidation and biogeochemical function of coastal sediments.

Rocky benthic communities are common in Antarctic coastal habitats; yet little is known about their carbon turnover rates. Here, we performed a broad survey of shallow ( < 65 m depth) rocky ice-scoured habitats of South Bay (Doumer Island, Western Antarctic Peninsula), combining (i) biodiversity assessments from benthic imaging, and (ii) in situ benthic dissolved oxygen (O2) exchange rates quantified by the aquatic eddy covariance technique. The 18 study sites revealed a gradual transition from macroalgae and coralline-dominated communities at ice-impacted depths (15–25 m; zone I) to large suspension feeders (e.g., sponges, bivalves) at depth zone II (25–40 m) and extensive suspension feeders at the deepest study location (zone III; 40–65 m). Gross primary production (GPP) in zone I was up to 70 mmol O2 m−2 d−1 and dark ecosystem respiration (ER) ranged from 15 to 90 mmol m−2 d−1. Zone II exhibited reduced GPP (average 1.1 mmol m−2 d−1) and ER rates from 6 to 36 mmol m−2 d−1, whereas aphotic zone III exhibited ER between 1 and 6 mmol m−2 d−1. Benthic ER exceeded GPP at all study sites, with daily net ecosystem metabolism (NEM) ranging from − 22 mmol m−2 d−1 at the shallow sites to − 4 mmol m−2 d−1 at 60 m. Similar NEM dynamics have been observed for hard-substrate Arctic habitats at comparable depths. Despite relatively high GPP during summer, coastal rocky habitats appear net heterotrophic. This is likely due to active drawdown of organic material by suspension-feeding communities that are key for biogeochemical and ecological functioning of high-latitude coastal ecosystems.

Eurythenes S.I. Smith in Scudder, 1882 (Crustacea: Amphipoda) are prevalent scavengers of the benthopelagic community from bathyal to hadal depths. While a well-studied genus, molecular systematic studies have uncovered cryptic speciation and multiple undescribed lineages. Here, we apply an integrative taxonomic approach and describe the tenth species, Eurythenes atacamensis sp. nov., based on specimens from the 2018 Atacamex and RV Sonne SO261 Expeditions to the southern sector of the Peru-Chile Trench, the Atacama Trench (24–⁠21°S). Eurythenes atacamensis sp. nov. is a large species, max. observed length 83.2 mm, possesses diagnostic features, including a short gnathopod 1 palm and a chelate gnathopod 2 palm, and a distinct genetic lineage based on a 16S rRNA and COI phylogeny. This species is a dominant bait-attending fauna with an extensive bathymetric range, spanning from 4974 to 8081 m. The RV Sonne SO261 specimens were recovered along a 10-station transect from abyssal to hadal depths and further examined for demographic and bathymetric-related patterns. Ontogenetic vertical stratification was evident across the trench axis, with only juveniles present at abyssal depths (4974–6025 m). Total length-depth analysis revealed that the size of females was unrelated to depth, whereas juveniles followed a sigmoidal relationship with a step-up in size at depths >7200 m. Thus, these bathymetric trends suggest that juveniles and females employ differing ecological strategies in subduction trench environments. This study highlights that even dominant and ecologically important species are still being discovered within the abyssal and hadal environments. Continued systematic expeditions will lead to an improved understanding of the eco-evolutionary drivers of speciation in the world’s largest ecosystem.

This study presents a novel approach resulting in the first cold-water coral reef biomass maps, used to assess associated ecosystem functions, such as carbon (C) stock and turnover. We focussed on two dominant ecosystem engineers at the Mingulay Reef Complex, the coral Lophelia pertusa (rubble, live and dead framework) and the sponge Spongosorites coralliophaga. Firstly, from combining biological (high-definition video, collected specimens), environmental (extracted from multibeam bathymetry) and ecosystem function (oxygen consumption rate values) data, we calculated biomass, C stock and turnover which can feed into assessments of C budgets. Secondly, using those values, we employed random forest modelling to predictively map whole-reef live coral and sponge biomass. The whole-reef mean biomass of S. coralliophaga was estimated to be 304 T (range 168–440 T biomass), containing 10 T C (range 5–18 T C) stock. The mean skeletal mass of the coral colonies (live and dead framework) was estimated to be 3874 T (range 507–9352 T skeletal mass), containing a mean of 209 T of biomass (range 26–515 T biomass) and a mean of 465 T C (range 60–1122 T C) stock. These estimates were used to calculate the C turnover rates, using respiration data available in the literature. These calculations revealed that the epi- and microbial fauna associated with coral rubble were the largest contributor towards C turnover in the area with a mean of 163 T C year−1 (range 149–176 T C year−1). The live and dead framework of L. pertusa were estimated to overturn a mean of 32 T C year−1 (range 4–93 T C year−1) and 44 T C year−1 (range 6–139 T C year−1), respectively. Our calculations showed that the Mingulay Reef overturned three to seven (with a mean of four) times more C than a soft-sediment area at a similar depth. As proof of concept, the supply of C needed from surface water primary productivity to the reef was inferred. Since 65–124 T C year−1 is supplied by natural deposition and our study suggested that a mean of 241 T C year−1 (range 160–400 T C year−1), was turned over by the reef, a mean of 117–176 T C year−1 (range 36–335 T C year−1) of the reef would therefore be supplied by tidal downwelling and/or deep-water advection. Our results indicate that monitoring and/or managing surface primary productivity would be a key consideration for any efforts towards the conservation of cold-water coral reef ecosystems.

Mytilid mussels form abundant, species-rich reefs on rocky substrates, but the role of this key habitat in carbon (C) cycling remains poorly understood. We performed a seasonal study on a 5 m deep photicMytilus trossulusreef in the Central Baltic Sea to investigate pathways and rates of organic C flow. Reef gross primary production (GPP) and respiration (R) were estimated seasonally using underwater O2eddy covariance on hourly and daily timescales. Photogrammetry and biotic sampling were used to quantify reef rugosity and mussel coverage, and to derive mussel filtration and biodeposition. Mussels were highly abundant, reaching ~50000 ind. m-2, and the reef structure increased the seabed surface area by 44%. GPPhourlywas up to 20 mmol O2m-2h-1and GPPdailywas up to 107 mmol O2m-2d-1, comparable to a nearby seagrass canopy. Hourly eddy fluxes responded linearly to light intensity and flow velocity, with higher velocities enhancing reef O2uptake at night. ReefRdailyexceeded GPPdailyon 12 of 13 measurement days, andRannual(29 mol O2m-2yr-1) was 3-fold larger than GPPannual. The reef sustained a productive community of microbes and fauna whose activities accounted for ~50% ofRannual. Horizontal water advection promoted food supply to the reef and likely facilitated substantial lateral C export of mussel biodeposits. Our analyses suggest that a reduction in mussel reef extent due to ongoing environmental change will have major implications for the transport and transformation of C and nutrients within the coastal Baltic Sea.

BACKGROUND: Pigs and minipigs are increasingly used as non-primate large animal models for preclinical research on nervous system disorders resulting in motor dysfunction. Knowledge of the minipig pyramidal tract is therefore essential to support such models.AIM AND METHODS: This study used 5 female Göttingen minipigs aging 11-15 months. The Göttingen minipig corticospinal tract was investigated, in the same animals, with in vivo neuronal tracing and with postmortem diffusion weighted MRI tractography to provide a thorough insight in the encephalic distribution of this primary motor pathway and its decussation at the craniocervical junction.RESULTS: The two methods similarly outlined the course of the pyramidal tract from its origin in the motor cortex down through the internal capsule to the craniocervical junction, where both methods displayed an axonal crossover at the pyramid decussation. The degree of crossover was quantified with unbiased stereology, where 81-93% of the traced corticospinal fibers crossed to the contralateral spinal cord. Accordingly, in the upper cervical spinal cord the corticospinal tract is primarily distributed in the contralateral lateral funiculus and in close relation to the gray matter, wherein some direct terminations on large ventral column gray matter neurons could be identified.DISCUSSION: The combination of neuronal tracing and tractography exploited the strengths of the respective methods to gain a better understanding of the encephalic distribution and craniocervical decussation of the Göttingen minipig corticospinal tract. Moreover, a quantification of the crossing fibers was obtained from the tracing data, which was not possible with tractography. Our data indicate that the porcine corticospinal system is quite lateralized down to the investigated upper cervical levels. However, further elucidation of this point will require a full examination of the corticospinal tracing pattern into the caudal spinal cord combined with an analysis of the direct versus indirect termination pattern on the lower motor neurons.

The copepod Calanus hyperboreus, a key species of Arctic marine ecosystems, has a partially anoxic gut that is suspected to host anaerobic microbial activities. So far, however, only dead specimens have been studied in which gut processes are quickly abolished by microbial carcass degradation. Here, live specimens were exposed to different feeding regimes and ambient oxygen levels to study the controls on copepod-associated microbial nitrogen cycling in 15N-enrichment experiments. Algae-fed copepods exhibited consistently high rates of denitrification and low or variable rates of dissimilatory nitrate reduction to ammonium (or nitrite), nitrification, and nitrous oxide release. Thus, live C. hyperboreus contribute to pelagic fixed-nitrogen loss through denitrificationthat is otherwise absent from oxygenated Arctic surface waters. The feeding-related controls on copepod-associated anaerobic nitrogen cycling aresupported by significantly reduced rates in starved copepods. In addition, freshly released fecal pellets showed similar patterns of nitrogen cycling as fedcopepods. Unlike in previous studies on copepod carcasses, low ambient oxygen levels did not stimulate anaerobic nitrogen cycling in live specimens, supportingthat the C. hyperboreus gut is anoxic, irrespective of ambient oxygen levels. Fecal pellets and carcasses retained denitrification activity for 4 d of simulated sinking. We estimate that during the time of the Arctic spring bloom, pelagic denitrification associated with live specimens, fecal pellets, and carcassesof C. hyperboreus is equivalent to ~12% of the benthic fixed-nitrogen loss. This estimate increases to ~28%, provided that the abundant sibling species C.glacialis and C. finmarchicus exhibit denitrification activities in proportion to their smaller body size

The oxygen flux between benthic systems and the water above is a widely used proxy for benthic primary production and organic carbon mineralization and is one of the most measured variables in marine and freshwater research. This presentation reviews the relatively new aquatic eddy covariance technique for measuring this flux. Because the approach relies on measurements that integrate over a large area (multiple m2) without disturbing the benthic system or the natural drivers of flux, it allows non-invasive studies of whole-system benthic metabolism that are not possible with any other approach. After summarizing the basic principles of eddy covariance, the instruments used, and key steps in the data evaluation, the rapidly growing body of research utilizing aquatic eddy covariance is presented with examples of the new kinds of insights that can be attained. These examples focus on benthic surfaces where traditional flux methods are difficult or problematic to apply and include highly dynamic benthic environments such as coral reefs, Arctic sediments, dense seagrass meadows, and permeable sands. Finally, future applications of the technique, new areas for development, and avenues to disseminate data and outcomes between new and experienced users are recommended.

Sinking phycodetrital aggregates can contribute to anaerobic nitrogen turnover as they may represent oxygen-depleted microbial hot spots in otherwise oxygenated waters. However, the dynamics of anaerobic nitrogen cycling during the long descent of aggregates through oxic or hypoxic waters are unknown. Thus, model aggregates prepared from the diatom Skeletonema marinoi were allowed to age for 4 d at high and low ambient O2 levels (70 and 15% air saturation, respectively), and changes in nitrogen transformations and microbial community structure were followed. At both O2 levels, denitrification and dissimilatory NO3– reduction to NO2– (DNRN) were the most important processes of aggregate-associated anaerobic nitrogen cycling. However, at 70% air saturation, rates of anaerobic N cycling were lower and decayed towards 0 after an early rise, whereas at 15% air saturation, they remained constantly high at average production rates of 0.66 nmol N2-N aggregate (aggr.)–1 h–1 and 0.26 nmol NO2– aggr.–1 h–1. At both O2 levels, but more pronouncedly at 70% air saturation, the microbial community underwent succession as expressed by an increase in (1) relative abundance of specific bacterial taxonomic units; (2) bacterial diversity; and (3) prokaryotic abundance. Probably, a higher carbon oxidation rate at high ambient O2 level progressively selected for microbes capable of using complex carbon polymers. Taken together, the occurrence of anoxic aggregate centers may be ephemeral at high ambient O2 levels, but persistent at low ambient O2 levels, indicating that sinking phycodetrital aggregates can remain sinks for bioavailable N in the oceans for several days, especially in O2‑depleted settings

PURPOSE: To compare knee kinematics before and after anterior cruciate ligament ACL reconstruction (ACL-R) using hamstring graft (HG) and a double-stranded iliotibial tract graft attached to Gerdy's tubercle (providing an extra-articular anterolateral tenodesis) (named the modified iliotibial tract graft = MIT). METHOD: Eighteen cadaveric knees were tested in a 6 degree of freedom kinematics rig. An optical tracking system recorded kinematics of the knee from 0 to 80 degrees of flexion applying no load, internal/external rotation (IR/ER), valgus/varus rotation (VGR/VRR), simulated pivot shift (SPS), anterior translation (AT) and posterior translation loads. The knee was tested before and after resection of the ACL and the anterolateral ligament (ALL), respectively; then after HG-ACL-R and MIT-ACL-R. Grafts were fixed at 20° of flexion. Results were compared to the intact knee. RESULTS: ACL resection resulted in a significant increase in AT (p

The Aquatic Eddy Covariance (AEC) technique has emerged as an important method to quantifyin situseafloor metabolism over large areas of heterogeneous benthic communities, enabling cross-habitat comparisons of seafloor productivity. However, the lack of a corresponding sampling protocol to perform biodiversity comparisons across habitats is impeding a full assessment of marine ecosystem metabolism. Here, we study a range of coastal benthic habitats, from rocky-bed communities defined by either perennial macroalgae or blue mussel beds to soft-sediment communities comprised of either seagrass, patches of different macrophyte species or bare sand. We estimated that the maximum contribution to the AEC metabolic flux can be found for a seafloor area of approximately 80 m2with a 5 meter upstream distance of the instrument across all the habitats. We conducted a sampling approach to characterize and quantify the dominant features of biodiversity (i.e., community biomass) within the main seafloor area of maximum metabolic contribution (i.e., gross primary production and community respiration) measured by the AEC. We documented a high biomass contribution of the macroalgalFucus vesiculosus, the seagrassZostera marinaand the macroinvertebrateMytilus edulisto the net ecosystem metabolism of the habitats. We also documented a significant role of the bare sediments for primary productivity compared to vegetated canopies of the soft sediments. The AEC also provided insight into dynamic short-term drivers of productivity such as PAR availability and water flow velocity for the productivity estimate. We regard this study as an important step forward, setting a framework for upcoming research focusing on linking biodiversity metrics and AEC flux measurements across habitats.

The important role of macroalgal canopies in the oceanic carbon (C) cycle is increasingly being recognized, but direct assessments of community productivity remain scarce. We conducted a seasonal study on a sublittoral Baltic Sea canopy of the brown algaFucus vesiculosus, a prominent species in temperate and Arctic waters. We investigated community production on hourly, daily, and seasonal timescales. Aquatic eddy covariance (AEC) oxygen flux measurements integrated ~ 40 m2of the seabed surface area and documented considerable oxygen production by the canopy year‐round. High net oxygen production rates of up to 35 ±  9 mmol m-2h-1were measured under peak irradiance of ~ 1200 µmol photosynthetically active radiation (PAR) m-2s-1in summer. However, high rates > 15 mmol m-2h-1were also measured in late winter (March) under low light intensities μmol PAR m-2s-1and water temperatures of ~ 1°C. In some cases, hourly AEC fluxes documented an apparent release of oxygen by the canopy under dark conditions, which may be due to gas storage dynamics within internal air spaces ofF. vesiculosus.Daily net ecosystem metabolism (NEM) was positive (net autotrophic) in all but one of the five measurement campaigns (December). A simple regression model predicted a net autotrophic canopy for two‐thirds of the year, and annual canopyNEMamounted to 25 mol O2m-2yr-1, approximately six‐fold higher than net phytoplankton production. Canopy C export was ~ 0.3 kg C m-2yr-1, comparable to canopy standing biomass in summer. Macroalgal canopies thus represent regions of intensified C assimilation and export in coastal waters.

Aquatic macrophytes modify the sediment biogeochemistry via radial oxygen loss (ROL) from their roots. However, the variation in ROL and its implication for nutrient availability remains poorly explored. Here, we use planar O2 optodes to investigate the spatial heterogeneity of oxic niches within the rhizosphere of Vallisneria spiralis and their alteration following variable light and ambient O2 levels. The effect of ROL on NH4 + and PO4 3− distribution in the rhizosphere was evaluated by a combination of 15N isotopic techniques, 2D sampling, and electron microscopy. A single specimen of V. spiralis could maintain an oxidised sediment volume of 41–47 cm3 and 10–27 cm3 in the rhizosphere at 100% and 38% dissolved oxygen saturation in the overlying water, respectively. Whatever the environmental conditions, the ROL was, however, very heterogeneous and dependent on root age and architecture of the root system. ROL stimulated the coupling between denitrification and nitrification in the sediment both under dark (+25 μmol N-N2 m−2 hr−1) and light (+70 μmol N-N2 m−2 hr−1) conditions. This, in combination with plant uptake, contributed to intense removal of NH4 + from the pore water. Similarly, PO4 3− was highly depleted in the rhizosphere. The detection of Fe-P plaques on the roots surface indicated substantial entrapment of P as a consequence of ROL. The extensive spatio-temporal heterogeneity of oxic and anoxic conditions ensured that aerobic and anaerobic processes co-occurred in the rhizosphere and this presumably reduced potential nutrient limitation while maximising plant fitness in an otherwise hostile reduced environment.

Cable bacteria represent a newly discovered group of filamentous microorganisms, which are capable of spatially separating the oxidative and reductive half-reactions of their sulfide-oxidizing metabolisms over centimeter distances. We investigated three ways that cable bacteria might interact with the nitrogen (N) cycle: (1) by reducing nitrate through denitrification or dissimilatory nitrate reduction to ammonium (DNRA) within their cathodic cells; (2) by nitrifying ammonium within their anodic cells; and (3) by indirectly affecting denitrification and/or DNRA by changing the Fe 2+ concentration in the surrounding sediment. We performed 15 N labeling laboratory experiments to measure these three processes using cable bacteria containing sediments from the Yarra River, Australia, and from Vilhelmsborg Sø, Denmark. Our results revealed that in the targeted systems, cable bacteria themselves did not perform significant rates of denitrification, DNRA, or nitrification. However, cable bacteria exhibited an important indirect effect, whereby they increased the Fe 2+ pool through iron sulfide dissolution. This elevated availability of Fe 2+ significantly increased DNRA and in some cases decreased denitrification. Thus, cable bacteria presence may affect the relative importance of DNRA in sediments and thus the extent by which bioavailable nitrogen is lost from the system.

Permeable sediments are common across continental shelves and are critical contributors to marine biogeochemical cycling. Organic matter in permeable sediments is dominated by microalgae, which as eukaryotes have different anaerobic metabolic pathways to prokaryotes such as bacteria and archaea. Here we present analyses of flow-through reactor experiments showing that dissolved inorganic carbon is produced predominantly as a result of anaerobic eukaryotic metabolic activity. In our experiments, anaerobic production of dissolved inorganic carbon was consistently accompanied by large dissolved H2 production rates, suggesting the presence of fermentation. The production of both dissolved inorganic carbon and H2 persisted following administration of broad spectrum bactericidal antibiotics, but ceased following treatment with metronidazole. Metronidazole inhibits the ferredoxin/hydrogenase pathway of fermentative eukaryotic H2 production, suggesting that pathway as the source of H2 and dissolved inorganic carbon production. Metabolomic analysis showed large increases in lipid production at the onset of anoxia, consistent with documented pathways of anoxic dark fermentation in microalgae. Cell counts revealed a predominance of microalgae in the sediments. H2 production was observed in dark anoxic cultures of diatoms (Fragilariopsis sp.) and a chlorophyte (Pyramimonas) isolated from the study site, substantiating the hypothesis that microalgae undertake fermentation. We conclude that microalgal dark fermentation could be an important energy-conserving pathway in permeable sediments.

Background In the past few years, an increasing number of studies have reported the potential use of microRNAs (miRNA) as circulating biomarkers for diagnosis or prognosis of a wide variety of diseases. There is, however, a lack of reproducibility between studies. Due to the high miRNA content in platelets this may partly be explained by residual platelets in the plasma samples used. When collecting fresh plasma samples, it is possible to produce cell-free/platelet-poor plasma by centrifugation. In this study, we systematically investigated whether biobanked EDTA plasma samples could be processed to be suitable for miRNA analysis. Materials and methods Blood samples were collected from ten healthy volunteers and centrifuged to produce platelet-poor-plasma (PPP) and standard biobank plasma. After one week at −80 °C the biobanked EDTA plasma was re-centrifuged by different steps to remove residual platelets. Using RT-qPCR the levels of 14 miRNAs in the different plasma preparations were compared to that of PPP. Results We were able to remove residual platelets from biobanked EDTA plasma by re-centrifugation of the thawed samples. Nevertheless, for most of the investigated miRNAs, the miRNA level was significantly higher in the re-centrifuged biobanked plasma compared to PPP, even when the platelet count was reduced to 0–1×109/L. Conclusion We found, that pre-storage centrifugation conditions have a significant impact on the measured EDTA plasma level of miRNAs known to be present in platelets. Even for the miRNAs found to be less effected, we showed that a 1.5–3 fold change in plasma levels may possible be caused by or easily overseen due to sample preparation and/or storage., Highlights • Platelet count vary significantly in standard biobanked EDTA plasma. • Residual platelets can be removed by centrifugation of thawed biobanked samples. • Re-centrifugation of biobanked plasma cannot eliminate contaminating platelet miRNAs. • Pre-storage centrifugation conditions have large impact on measured miRNA levels.

Impairment of the ubiquitin proteasome system has been implicated in Parkinson’s disease. We used positron emission tomography to investigate longitudinal effects of chronic intracerebroventricular exposure to the proteasome inhibitor lactacystin on monoaminergic projections and neuroinflammation. Göttingen minipigs were implanted in the cisterna magna with a catheter connected to a subcutaneous injection port. Minipigs were imaged at baseline and after cumulative doses of 200 and 400 μg lactacystin, respectively. Main radioligands included [11C]-DTBZ (vesicular monoamine transporter type 2) and [11C]-yohimbine (α2-adrenoceptor). [11C]-DASB (serotonin transporter) and [11C]-PK11195 (activated microglia) became available later in the study and we present their results in a smaller subset of animals for information purposes only. Striatal [11C]-DTBZ binding potentials decreased significantly by 16% after 200 μg compared to baseline, but the decrease was not sustained after 400 μg (n = 6). [11C]-yohimbine volume of distribution increased by 18–25% in the pons, grey matter and the thalamus after 200 μg, which persisted at 400 μg (n = 6). In the later subset of minipigs, we observed decreased [11C]-DASB (n = 5) and increased [11C]-PK11195 (n = 3) uptake after 200 μg. These changes may mimic monoaminergic changes and compensatory responses in early Parkinson’s disease.

The Göttingen minipig is being used increasingly in biomedical research. The anatomical structure of the porcine peripheral nervous system has been extensively characterized, but no equivalent to the dermatome map, which is so valuable in human neurophysiological research, has been created. We characterized the medullar segmental skin and muscle innervations of the minipig hind body, using neurophysiological methodology. Six adult minipigs underwent unilateral laminectomy from L2 to S3, exposing the nerve roots. The skin of the hind part of the body was divided into 36 predefined fields, based on anatomical landmarks for consistent reproducibility. We recorded the evoked potential in each exposed nerve root L2-S3 for cutaneous stimulation of each skin field, mapping the sensory innervation of the entire hind body. We subsequently recorded the motor response in seven predefined muscles during sequential stimulation of the L2-S3 nerve roots. We obtained a clear sensory evoked potential in the nerve roots during stimulation of the skin fields, allowing us to map the sensory innervation of the minipig hind body. Neurophysiological data from skin stimulation and muscle recordings enabled us to map the sensory innervation of the Göttingen minipig hind body and provide information about muscular innervation. The skin fields were sensory innervated by more than one root. The muscles each had one dominant root with minor contribution from neighboring roots. This is consistent with experimental data from human studies.

Sea-ice ecosystems are among the most extensive of Earth’s habitats; yet its autotrophic and heterotrophic activities remain poorly constrained. We employed the in situ aquatic eddy-covariance (AEC) O2 flux method and laboratory incubation techniques (H14CO3−, [3H] thymidine and [3H] leucine) to assess productivity in Arctic sea-ice using different methods, in conditions ranging from land-fast ice during winter, to pack ice within the central Arctic Ocean during summer. Laboratory tracer measurements resolved rates of bacterial C demand of 0.003–0.166 mmol C m−2 day−1 and primary productivity rates of 0.008–0.125 mmol C m−2 day−1 for the different ice floes. Pack ice in the central Arctic Ocean was overall net autotrophic (0.002–0.063 mmol C m−2 day−1), whereas winter land-fast ice was net heterotrophic (− 0.155 mmol C m−2 day−1). AEC measurements resolved an uptake of O2 by the bottom-ice environment, from ~ − 2 mmol O2 m−2 day−1 under winter land-fast ice to~ − 6 mmol O2 m−2 day−1 under summer pack ice. Flux of O2-deplete meltwater and changes in water flow velocity masked potential biological-mediated activity. AEC estimates of primary productivity were only possible at one study location. Here, productivity rates of 1.3 ± 0.9 mmol O2 m−2 day−1, much larger than concurrent laboratory tracer estimates (0.03 mmol C m−2 day−1), indicate that ice algal production and its importance within the marine Arctic could be underestimated using traditional approaches. Given careful flux interpretation and with further development, the AEC technique represents a promising new tool for assessing oxygen dynamics and sea-ice productivity in ice-covered regions.

A considerable fraction of freshwater zooplankton was recently found to consist of dead specimens that sink to the lake bottom. Such carcasses host intense microbial activities that may promote oxygen depletion at the microscale. Therefore, we tested the hypothesis that sinking zooplankton carcasses are microsites of anaerobic nitrogen cycling that contribute to pelagic fixed-nitrogen loss even in the presence of ambient oxygen. Incubation experiments were performed with the ubiquitous copepods Eudiaptomus sp. and Megacyclops gigas at different ambient oxygen levels that sinking carcasses encounter during their descent in stratified lakes. 15N-stable-isotope incubations revealed intense carcass-associated anaerobic nitrogen cycling only at low ambient oxygen levels (, Freshwater copepods, such as Eudiaptomus sp., host unique microenvironments in the water column of lakes and thereby enable diverse anaerobic nitrate conversion pathways mediated by bacterial reductases.

A protocol for quantification of circulating microRNA using TaqMan assays. MicroRNAs are purified from platelet-poor plasma using Nucleospin columns During the purification step samples are spiked with cel-miR-39 as a mean of technical normalization Normalization is performed using the ΔCt-method

Anoxic mineralization of organic matter releases dissolved inorganic carbon and produces reduced mineralization products. The reoxidation of these reduced compounds is essential for biogeochemical cycling in sediments and is mainly performed by chemoautotrophic microbes, which synthesize new organic carbon by dark CO2 fixation. At present however, the biogeochemical importance of chemoautotrophy in high-latitude sediments is largely unknown. Here, we determine the seasonal variation in sedimentary chemoautotrophic production in Kobbefjord (SW Greenland). Intact sediment cores from the fjord were incubated, and dark CO2 fixation was quantified by combining bacterial phospholipid-derived fatty acid analysis with 13C stable isotope probing (PLFA-SIP). Our results reveal a distinct seasonal cycle in chemoautotrophic activity, which increases after the spring bloom and shows lowest activity in the late winter when the fjord is covered by sea ice. The depth distribution of chemoautotrophic activity also varied seasonally, likely due to seasonal variation in the bioturbation activity of sediment infauna. Although chemoautotrophy rates (0.4 ± 0.2 mmol C m−2 d−1) were in the low range for coastal sediments, they are comparable to those from intertidal sandflats and brackish tropical lagoons, and scale with the sulfide production through sulfate reduction in the fjord. Chemoautotrophic production in these fjord sediments thus appears to be mainly driven by sulfide oxidation and can re-fix 4% of the CO2 produced by mineralization.

Copepods are exposed to a high non-predatory mortality and their decomposing carcasses act as microniches with intensified microbial activity. Sinking carcasses could thereby represent anoxic microenvironment sustaining anaerobic microbial pathways in otherwise oxic water columns. Using non-invasive O-2 imaging, we document that carcasses of Calanus finmarchicus had an anoxic interior even at fully air-saturated ambient O-2 level. The extent of anoxia gradually expanded with decreasing ambient O-2 levels. Concurrent microbial sampling showed the expression of nitrite reductase genes (nirS) in all investigated carcass samples and thereby documented the potential for microbial denitrification in carcasses. The nirS gene was occasionally expressed in live copepods, but not as consistently as in carcasses. Incubations of sinking carcasses in (15)NO3-amended seawater demonstrated denitrification, of which on average 34%+/- 17% (n=28) was sustained by nitrification. However, the activity was highly variable and was strongly dependent on the ambient O-2 levels. While denitrification was present even at air-saturation (302 mol L-1), the average carcass specific activity increased several orders of magnitude to approximate to 1 nmol d(-1) at 20% air-saturation (55 mol O-2 L-1) at an ambient temperature of 7 degrees C. Sinking carcasses of C. finmarchicus therefore represent hotspots of pelagic denitrification, but the quantitative importance as a sink for bioavailable nitrogen is strongly dependent on the ambient O-2 level. The importance of carcass associated denitrification could be highly significant in O-2 depleted environments such as Oxygen Minimum Zones (OMZ).

Mass cultivation of marine copepods as live feed requires predictable and uniform production of a standard product for the end users, the hatcheries. A previously undescribed large egg type, measuring a mean diameter of 106.5 mu m, was characterized and compared to normal eggs (82.3 mu m) from a continuous culture of Acartia tonsa. The large egg type was significantly larger than the normal eggs, and only 1 large out of approximately 100 000 normal-sized eggs was found. Mean egg densities was 1.34 +/- 0.16 g/cm(3) and 1.20 +/- 0.07 g/cm(3) for normal and large eggs, respectively. The hypothesis that large eggs were dormant, as indicated from the relatively low hatching success of 60%, within 48 h, was rejected as no significant difference was found in egg volume-specific respiration rates or histological analysis of the chorion between the two egg types. The nauplii hatched from large eggs were correspondingly large, and the copepods remained large when developing into adulthood. The fact that copepods hatched from large eggs were fertile indicates no chromosomal abnormalities and suggests that this egg type represents the upper tail end of eggs in a classical normal distribution of size vs. frequency. The next generations' eggs were not as large as the ones they originated from suggesting that the rare large eggs represent a maternal effect. The large size eggs do however pose an interesting selection potential for designing large species-specific live feed products in aquaculture. (C) 2014 Elsevier B.V. All rights reserved.

Parkinson's disease (PD) is characterized by degeneration of dopaminergic neurons in the substantia nigra leading to slowness and stiffness of limb movement with rest tremor. Using ubiquitin proteasome system inhibitors, rodent models have shown nigrostriatal degeneration and motor impairment. We translated this model to the Gottingen minipig by administering lactacystin into the medial forebrain bundle (MFB). Minipigs underwent positron emission tomography (PET) imaging with (+)-alpha-[C-11]dihydrotetrabenazine ([C-11]DTBZ), a marker of vesicular monoamine transporter 2 availability, at baseline and three weeks after the unilateral administration of 100 mu g lactacystin into the MFB. Compared to their baseline values, minipigs injected with lactacystin showed on average a 36% decrease in ipsilateral striatal binding potential corresponding to impaired presynaptic dopamine terminals. Behaviourally, minipigs displayed asymmetrical motor disability with spontaneous rotations in one of the animals. Immunoreactivity for tyrosine hydroxylase (TH) and HLA-DR-positive microglia confirmed asymmetrical reduction in nigral TH-positive neurons with an inflammatory response in the lactacystin-injected minipigs. In conclusion, direct injection of lactacystin into the MFB of minipigs provides a model of PD with reduced dopamine neurotransmission, TH-positive neuron reduction, microglial activation and behavioural deficits. This large animal model could be useful in studies of symptomatic and neuroprotective therapies with translatability to human PD.

During the last 20 years pigs have become increasingly popular in large animal translational neuroscience research as an economical and ethical feasible substitute to non-human primates. The anatomy of the pig telencephalon is, however, not well known. We present, accordingly, a detailed description of the surface anatomy and cytoarchitecture of the Gottingen minipig telencephalon based on macrophotos and consecutive high-power microphotographs of 15 μm thick paraffin embedded Nissl-stained coronal sections. In 1-year-old specimens the formalin perfused brain measures approximately 55 × 47 × 36 mm (length, width, height) and weighs around 69 g. The telencephalic part of the Gottingen minipig cerebrum covers a large surface area, which can be divided into a neocortical gyrencephalic part located dorsal to the rhinal fissure, and a ventral subrhinal part dominated by olfactory, amygdaloid, septal, and hippocampal structures. This part of the telencephalon is named the subrhinal lobe, and based on cytoarchitectural and sulcal anatomy, can be discerned from the remaining dorsally located neocortical perirhinal/insular, pericallosal, frontal, parietal, temporal, and occipital lobes. The inner subcortical structure of the minipig telencephalon is dominated by a prominent ventricular system and large basal ganglia, wherein the putamen and the caudate nucleus posterior and dorsally are separated into two entities by the internal capsule, whereas both structures ventrally fuse into a large accumbens nucleus. The presented anatomical data is accompanied by surface renderings and high-power macrophotographs illustrating the telencephalic sulcal pattern, and the localization of the identified lobes and cytoarchitectonic areas. Additionally, 24 representative Nissl-stained telencephalic coronal sections are presented as supplementary material in atlas form on http://www.cense.dk/minipig_atlas/index.html and referred to as S1–S24 throughout the manuscript.

Oxygen minimum zones (OMZs) in the ocean are of key importance for pelagic fixed-nitrogen loss (N-loss) through microbial denitrification and anaerobic ammonium oxidation (anammox). Recent studies document that zooplankton is surprisingly abundant in and around OMZs and that the microbial community associated with carcasses of a large copepod species mediates denitrification. Here, we investigate the complex N-cycling associated with sinking zooplankton carcasses exposed to the steep O2 gradient in a coastal OMZ (Golfo Dulce, Costa Rica). 15N-stable-isotope enrichment experiments revealed that the carcasses of abundant copepods and ostracods provide anoxic microbial hotspots in the pelagic zone by hosting intense anaerobic N-cycle activities even in the presence of ambient O2. Carcass-associated anaerobic N-cycling was clearly dominated by dissimilatory nitrate reduction to ammonium (DNRA) at up to 30.8 nmol NH+4 individual−1 d−1, followed by denitrification (up to 10.8 nmol N2-N individual−1 d−1), anammox (up to 1.6 nmol N2-N individual−1 d−1), and N2O production (up to 1.2 nmol N2O-N individual−1 d−1). In contrast, anaerobic N-cycling mediated by free-living bacteria proceeded mainly through anammox and denitrification in the anoxic bottom water, which underpins the distinctive microbial metabolism associated with zooplankton carcasses. Pelagic N-loss is potentially enhanced by zooplankton carcasses both directly through N2 and N2O production, and indirectly through NH+4 production that may fuel free-living anammox bacteria. We estimate that in the hypoxic water layer of Golfo Dulce, carcass-associated N2 and N2O production enhance N-loss as much as 1.4-fold at a relative carcass abundance of 36%. In the anoxic bottom water, however, N-loss is likely enhanced only marginally due to high ambient rates and low zooplankton abundance. Thus, zooplankton carcasses may enhance N-loss mainly at the hypoxic boundaries of OMZs which are usually more extensive in open-ocean than in coastal settings. Notably, these contributions by zooplankton carcasses to pelagic N-loss remain undetected by conventional, incubation-based rate measurements. Danish National Research Council/[0602-02276B]/DNRC/Dinamarca OXYGEN/[267233]/OXYGEN/Dinamarca European Research Council/[669947]/ERC/Francia UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Ciencias del Mar y Limnología (CIMAR)

Eutrophication of estuaries and coastal seas is accelerating, increasing light stress on subtidal marine plants and changing their interactions with other species. To date, we have limited understanding of how such variations in environmental and biological stress modify the impact of interactions among foundational species and eventually affect ecosystem health. Here, we used metabolomics to assess the impact of light reductions on interactions between the seagrass Zostera marina, an important habitat-forming marine plant, and the abundant and commercially important blue mussel Mytilus edulis. Plant performance varied with light availability but was unaffected by the presence of mussels. Metabolomic analysis, on the other hand, revealed an interaction between light availability and presence of M. edulis on seagrass metabolism. Under high light, mussels stimulated seagrass nitrogen and energy metabolism. Conversely, in low light mussels impeded nitrogen and energy metabolism, and enhanced responses against sulfide toxicity, causing inhibited oxidative energy metabolism and tissue degradation. Metabolomic analysis thereby revealed cryptic changes to seagrass condition that could not be detected by traditional approaches. Our findings suggest that coastal eutrophication and associated reductions in light may shift seagrass-bivalve interactions from mutualistic to antagonistic, which is important for conservation management of seagrass meadows.

Diatom-bacteria aggregates are key for the vertical transport of organic carbon in the ocean. Sinking aggregates also represent pelagic microniches with intensified microbial activity, oxygen depletion in the center, and anaerobic nitrogen cycling. Since some of the aggregate-forming diatom species store nitrate intracellularly, we explored the fate of intracellular nitrate and its availability for microbial metabolism within anoxic diatom-bacteria aggregates. The ubiquitous nitrate-storing diatom Skeletonema marinoi was studied as both axenic cultures and laboratory-produced diatom-bacteria aggregates. Stable 15N isotope incubations under dark and anoxic conditions revealed that axenic S. marinoi is able to reduce intracellular nitrate to ammonium that is immediately excreted by the cells. When exposed to a light:dark cycle and oxic conditions, S. marinoi stored nitrate intracellularly in concentrations > 60 mmol L-1 both as free-living cells and associated to aggregates. Intracellular nitrate concentrations exceeded extracellular concentrations by three orders of magnitude. Intracellular nitrate was used up within 2-3 days after shifting diatom-bacteria aggregates to dark and anoxic conditions. Thirty-one percent of the diatom-derived nitrate was converted to nitrogen gas, indicating that a substantial fraction of the intracellular nitrate pool of S. marinoi becomes available to the aggregate-associated bacterial community. Only 5% of the intracellular nitrate was reduced to ammonium, while 59% was recovered as nitrite. Hence, aggregate-associated diatoms accumulate nitrate from the surrounding water and sustain complex nitrogen transformations, including loss of fixed nitrogen, in anoxic, pelagic microniches. Additionally, it may be expected that intracellular nitrate not converted before the aggregates have settled onto the seafloor could fuel benthic nitrogen transformations.

In the world's oceans, even relatively low oxygen levels inhibit anaerobic nitrogen cycling by free-living microbes. Sinking organic aggregates, however, might provide oxygen-depleted microbial hotspots in otherwise oxygenated surface waters. Here, we show that sinking diatom aggregates can host anaerobic nitrogen cycling at ambient oxygen levels well above the hypoxic threshold. Aggregates were produced from the ubiquitous diatom Skeletonema marinoi and the natural microbial community of seawater. Microsensor profiling through the center of sinking aggregates revealed internal anoxia at ambient 40% air saturation (∼100 μmol O2 L(-1)) and below. Accordingly, anaerobic nitrate turnover inside the aggregates was evident within this range of ambient oxygen levels. In incubations with (15)N-labeled nitrate, individual Skeletonema aggregates produced NO2 (-) (up to 10.7 nmol N h(-1) per aggregate), N2 (up to 7.1 nmol N h(-1)), NH4 (+) (up to 2.0 nmol N h(-1)), and N2O (up to 0.2 nmol N h(-1)). Intriguingly, nitrate stored inside the diatom cells served as an additional, internal nitrate source for dinitrogen production, which may partially uncouple anaerobic nitrate turnover by diatom aggregates from direct ambient nitrate supply. Sinking diatom aggregates can contribute directly to fixed-nitrogen loss in low-oxygen environments in the ocean and vastly expand the ocean volume in which anaerobic nitrogen turnover is possible, despite relatively high ambient oxygen levels. Depending on the extent of intracellular nitrate consumption during the sinking process, diatom aggregates may also be involved in the long-distance export of nitrate to the deep ocean.

To investigate whether pigmentation genes involved in the melanogenic pathway (melanogenesis) contributed to melanoma predisposition, we compared pigmentary genetics with quantitative skin pigmentation measurements, the number of atypical nevi, the total nevus count, and the familial atypical multiple mole and melanoma (FAMMM) syndrome. We typed 32 pigmentary SNP markers and sequenced MC1R in 246 healthy individuals and 116 individuals attending periodic control for malignant melanoma development, 50 of which were diagnosed with FAMMM. It was observed that individuals with any two grouped MC1R variants (missense, NM_002386:c. 456C > A (p.TYR152*), or NM_002386:c.83_84insA (p.Asn29Glnfs*14) had significantly (p

A new sandwich sensor, consisting of an O 2 planar optode overlain by a thin (90 μm) DGT layer is present ed. This sensor can simultaneously resolve 2-D O 2 dynamics and trace metal fluxes in benthic substrates at a high spatial resolution. The DGT layer accumulates metals on a small particle size (0.2 μm) chelating resin and records the locally induced trace metal flux during the deployment, whereas the planar optode resolves the O 2 dynamic in near real time at the same location in the sediment. Despite its ultrathin composition, the DGT layer has high carrying capacity for trace metals with no saturation problems during application to typical coastal- or contaminated sediments. Combined with laser ablation, accumulated metal fluxes could be resolved at a resolution of ~200 μm, whereas the O 2 images had a resolution of ~100 μm. A 2-D diffusion-reaction model showed that the enhanced smearing and reduced response time of the O 2 signal associated with the additional DGT layer were marginal. To test sensor performance at realistic conditions, it was applied to an artificial bur row system consisting of permeable dialysis tubing flushed with oxygenated seawater. The measurements demonstrated localized mobilization of Ni, Cu, and Pb close to the burrow wall, where O 2 was elevated. The lat ter was also confirmed for Cu and Pb in natural sediments irrigated by the polychaete Hediste diversicolor . The sandwich sensor has great potential for investigating interrelations between O 2 dynamics and metal mobiliza tion in complex benthic systems such as burrows, rhizospheres, permeable sands, and microbial mats.

microRNA (miRNA) expression profiling and classification of tissue obtained from fine-needle aspirates (FNA) could be a major improvement of the preoperative diagnosis of thyroid nodules.Before this can be clinically implemented, a robust and non-toxic method for obtaining sufficient quantity and quality of RNA from single in vivo FNA has to be established. RNAlater is a non-toxic RNA-stabilizing agent. However, due to the high density of RNAlater, pelleting of the tissue samples is difficult, and results in low recovery of RNA that is insufficient for subsequent miRNA array expression analysis. We therefore developed a simple centrifugation method for capturing tissue stored in RNAlater on a 0.45-μm filter.FNA from 24 patients with a solitary cold thyroid nodule was stored in Trizol, liquid nitrogen, or RNAlater. The tissue stored in RNAlater was either pelleted by centrifugation or captured on the 0.45-μm filters. RNA was extracted using the Trizol method. To validate results, FNA from additional 30 patients were analyzed based on the modified RNAlater protocol.Capturing FNA tissue samples on the filters increased the RNA yield 10 fold and maintained RNA purity, permitting miRNA array expression profiling and allowing comparable levels of known miRNA-clusters regardless of preservation technique. Results were confirmed in an additional 30 patients.The modified RNAlater protocol is well suited for isolating RNA from single thyroid in vivo FNA in a clinical setting. Furthermore this permits shipping of FNA samples at room temperature from peripheral centers to a centralized array core facility.

Purpose This work describes the production and application of an aerosolised formulation of chitosan nanoparticles for improved pulmonary siRNA delivery and gene silencing in mice. Methods Aerosolised chitosan/siRNA nanoparticles were pneumatically formed using a nebulising catheter and sized by laser diffraction. In vitro silencing of aerosolised and non-aerosolised formulations was evaluated in an EGFP endogenous-expressing H1299 cell line by flow cytometry. Non-invasive intratracheal insertion of the catheter was used to study nanoparticle deposition by histological detection of Cy3-labeled siRNA and gene silencing in transgenic EGFP mouse lungs using a flow cytometric method. Results Flow cytometric analysis demonstrated minimal alteration in gene silencing efficiency before (68%) and after (62%) aerosolisation in EGFP-expressing H1299 cells. Intratracheal catheter administration in mice resulted in nanoparticle deposition throughout the entire lung in both alveoli and bronchiolar regions using low amounts of siRNA. Transgenic EGFP mice dosed with the aerosolised nanoparticle formulation showed significant EGFP gene silencing (68% reduction compared to mismatch group). Conclusions This work provides a technology platform for effective pulmonary delivery and gene silencing of RNAi therapeutics with potential use in preclinical studies of respiratory disease treatment.

Parkinson's disease is characterized by a progressive loss of substantia nigra (SN) dopaminergic neurons and the formation of Lewy bodies containing accumulated alpha-synuclein (α-syn). The pathology of Parkinson's disease is associated with neuroinflammatory microglial activation, which may contribute to the ongoing neurodegeneration. This study investigates the in vivo microglial and dopaminergic response to overexpression of α-syn. We used positron emission tomography (PET) and the 18 kDa translocator protein radioligand, [11 C](R)PK11195, to image brain microglial activation and (+)-α-[11 C]dihydrotetrabenazine ([11 C]DTBZ), to measure vesicular monoamine transporter 2 (VMAT2) availability in Gottingen minipigs following injection with recombinant adeno-associated virus (rAAV) vectors expressing either mutant A53T α-syn or green fluorescent protein (GFP) into the SN (4 rAAV-α-syn, 4 rAAV-GFP, 5 non-injected control minipigs). We performed motor symptom assessment and immunohistochemical examination of tyrosine hydroxylase (TH) and transgene expression. Expression of GFP and α-syn was observed at the SN injection site and in the striatum. We observed no motor symptoms or changes in striatal [11 C]DTBZ binding potential in vivo or striatal or SN TH staining in vitro between the groups. The mean [11 C](R)PK11195 total volume of distribution was significantly higher in the basal ganglia and cortical areas of the α-syn group than the control animals. We conclude that mutant α-syn expression in the SN resulted in microglial activation in multiple sub- and cortical regions, while it did not affect TH stains or VMAT2 availability. Our data suggest that microglial activation constitutes an early response to accumulation of α-syn in the absence of dopamine neuron degeneration.

MicroRNAs (miRNAs) are small, noncoding RNA molecules that regulate cellular differentiation, proliferation, and apoptosis. MiRNAs are expressed in a developmentally regulated and tissue-specific manner. Aberrant expression may contribute to pathological processes such as cancer, and miRNA may therefore serve as biomarkers that may be useful in a clinical environment for diagnosis of various diseases. Most miRNA profiling studies have used fresh tissue samples. However, in some types of cancer, including malignant melanoma, fresh material is difficult to obtain from primary tumors, and most surgical specimens are formalin fixed and paraffin embedded (FFPE). To explore whether FFPE material would be suitable for miRNA profiling in melanocytic lesions, we compared miRNA expression patterns in FFPE versus fresh frozen samples, obtained from 15 human melanocytic nevi. Out of microarray data, we identified 84 miRNAs that were expressed in both types of samples and represented an miRNA profile of melanocytic nevi. Our results showed a high correlation in miRNA expression (Spearman r-value of 0.80) between paired FFPE and fresh frozen material. The data were further validated by quantitative RT-PCR. In conclusion, FFPE specimens of melanocytic lesions are suitable as a source for miRNA microarray profiling.

We investigated the microalgal community in sea ice and in the water column of Kobbe- fjord, West Greenland, through an entire sea ice season. Temporal variation in physical (photosynthet- ically active radiation (PAR), temperature, brine volume) and chemical (salinity, nutrient concentra- tion) properties confirmed that sea ice is a very dynamic habitat. Nevertheless, a viable sea ice algal community was present throughout the year, with a species succession from flagellate dominance (di- noflagellates and cryptophytes) in December to February, followed by Chaetoceros simplex (a centric diatom) in March and pennate diatoms in May. Sea ice algal composition, biomass and productivity were influenced by PAR and brine volume, with colonization (enhanced by large brine volume) appar- ent during sea ice melt. Sea ice chlorophyll levels were generally low (

A mass coral spawning event on the Heron Island reef flat in 2005 provided a unique opportunity to examine the response of a coral reef ecosystem to a large episodic nutrient addition. A post-major spawning phytoplankton bloom resulted in only a small drawdown of dissolved inorganic phosphorus (DIP minimum 5 0.37 mmol L21), compared with almost complete removal of dissolved inorganic nitrogen (DIN) (minimum NO { 5 0.01 mmol L21 ;N H z 5 0.11 mmol L21), suggesting that pelagic primary production is potentially N limited on the timescale of this study. DIN, DIP, dissolved organic nitrogen (DON), and dissolved organic phosphorus were used in the production of biomass, and mass balance calculations highlighted the importance of organic forms of N and P for benthic and pelagic production in tropical coral reef environments characterized by low inorganic N and P. The input of N and P via the deposition of coral spawn and associated phytodetritus resulted in large changes to N cycling in the sediments, but only small changes to P cycling, because of the buffering capacity provided by the large pool of bioavailable P. It is most likely that this large pool of bioavailable P in the sediments drives potential N limitation of benthic coral reef communities. For example, there was sufficient bioavailable P stored in the top 10 cm of the sediment column to sustain the prespawning rates of benthic production for over 200 d. Most of the change in benthic N cycling occurred via DON and N2 pathways, driven by changes in the quantity and quality of organic matter deposited and decomposed post-major spawning. The heterotrophic and autotrophic microbial communities within the coral reef sands were able to rapidly (6 to 7 d) process the large episodic load of N and P provided by coral mass spawning.

PSII-fluorescence kinetics, and 14 C-incorporation rates in monocultures of the marine phytoplankton species Prorocentrum minimum (Pavill.) J. Schiller (Dinophyceae), Prymnesium parvum f. patelliferum (J. C. Green, D. J. Hibberd et Pienaar) A. Larsen (Coccolithophyceae), and Phaeodactylum tricornutum Bohlin (Bacillariophyceae), grown at 15� C and 80 lmol photons AEm )2 AEs )1 . Photosynthesis versus irradiance curves were measured at seven temperatures (0� C‐30� C) by all three approaches. The maximum photosynthetic rate (P C max) was strongly stimulated by temperature, reached an optimum for Pro. minimum only (20� C‐25� C), and showed a similar relative temperature response for the three applied methods, with Q10 ranging from 1.7 to 3.5. The maximum light utilization coefficient (a C ) was insensitive or decreased slightly with increasing temperature. Absolute rates of O2 production were calculated from pulse-amplitude-modulated (PAM) fluorometry measurements in combination with biooptical determination of absorbed quanta in PSII. The relationship between PAM-based O2 production and measured O2 production and 14 C assimilation showed a species-specific correlation, with 1.2‐3.3 times higher absolute values of P C max and a C when calculated from PAM data for Pry. parvum and Ph. tricornutum but equivalent for Pro. minimum. The offset seemed to be temperature insensitive and could be explained by a lower quantum yield for O2 production than the theoretical maximum (due to Mehler-type reactions). Conclusively, the PAM technique can be used to study temperature responses of photosynthesis in microalgae when paying attention to the absorption properties in PSII. Key index words: 14 C assimilation; microalgae; O2