Influence of Zooplankton Grazing on Free Dissolved Enzymes in the Sea

In the Northern Adriatic Sea, extracellular enzymatic activity was measured during a Lagrangian study following a drifting buoy for 40 h. Dissolved free enzymatic activity represented 20 to 70% of total activity depending on the type of enzyme. alpha- and beta-glucosidases exhibited a significantly higher free activity than proteolytic enzymes. In subsequent laboratory experiments we investigated the effect of zooplankton on the free enzyme pool. The 4-step approach included: (1) determination of the enzymatic activities in copepods (mainly Acartia clausi); (2) enzymatic activity in fecal pellets; (3) short- and long-term grazing experiments; and (4) degradability of free glucosidase in seawater. alpha- and beta-glucosidases, leu-aminopeptidase, lipase and chitinase were examined. Experiments in which zooplankton were selectively enriched revealed a significant increase in both particle-bound (due to the increase of bacterial density) and dissolved free enzymatic activity. Incubating water enriched in free enzymes released by zooplankton with natural bacterial consortia, we found that 70% of the original alpha- and beta-glucosidase activity remained after 22 h. The presence of microorganisms did not enhance the degradation of these enzymes as compared to autoclaved controls. We found that a considerable amount of free dissolved enzymes is lost by 0.2 mu m filtration using Nuclepore filters, thereby leading to an underestimation of dissolved enzymes by similar to 30% in our experiments. Based on our results we conclude that mesozooplankton contribute to the free enzymatic activity in natural waters especially during periods of high grazing activity

Release of cell-free enzymes by marine pelagic fungal strains

Fungi are ubiquitous organisms that secrete different enzymes to cleave large molecules into smaller ones so that can then be assimilated. Recent studies suggest that fungi are also present in the oceanic water column harboring the enzymatic repertoire necessary to cleave carbohydrates and proteins. In marine prokaryotes, the cell-free fraction is an important contributor to the oceanic extracellular enzymatic activities (EEAs), but the release of cell-free enzymes by marine fungi remains unknown. Here, to study the cell-free enzymatic activities of marine fungi and the potential influence of salinity on them, five strains of marine fungi that belong to the most abundant pelagic phyla (Ascomycota and Basidiomycota), were grown under non-saline and saline conditions (0 g/L and 35 g/L, respectively). The biomass was separated from the medium by filtration (0.2 μm), and the filtrate was used to perform fluorogenic enzymatic assays with substrate analogues of carbohydrates, lipids, organic phosphorus, sulfur moieties, and proteins. Kinetic parameters such as maximum velocity (Vmax) and half-saturation constant (Km) were obtained. The species studied were able to release cell-free enzymes, and this represented up to 85.1% of the respective total EEA. However, this differed between species and enzymes, with some of the highest contributions being found in those with low total EEA, with some exceptions. This suggests that some of these contributions to the enzymatic pool might be minimal compared to those with higher total EEA. Generally, in the saline medium, the release of cell-free enzymes degrading carbohydrates was reduced compared to the non-saline medium, but those degrading lipids and sulfur moieties were increased. For the remaining substrates, there was not a clear influence of the salinity. Taken together, our results suggest that marine fungi are potential contributors to the oceanic dissolved (i.e., cell-free) enzymatic pool. Our results also suggest that, under salinity changes, a potential effect of global warming, the hydrolysis of organic matter by marine fungal cell-free enzymes might be affected and hence, their potential contribution to the oceanic biogeochemical cycles

Seasonal dynamics of epiphytic microbial communities on marine macrophyte surfaces

Surfaces of marine macrophytes are inhabited by diverse microbial communities. Most studies focusing on epiphytic communities of macrophytes did not take into account temporal changes or applied low sampling frequency approaches. The seasonal dynamics of epiphytic microbial communities was determined in a meadow of Cymodocea nodosa invaded by Caulerpa cylindracea and in a monospecific settlement of C. cylindracea at monthly intervals. For comparison the ambient prokaryotic picoplankton community was also characterized. At the OTU level, the microbial community composition differed between the ambient water and the epiphytic communities exhibiting host-specificity. Also, successional changes were observed connected to the macrophyte growth cycle. Taxonomic analysis, however, showed similar high rank taxa (phyla and classes) in the ambient water and the epiphytic communities, with the exception of Desulfobacterota, which were only found on C. cylindracea. Cyanobacteria showed seasonal changes while other high rank taxa were present throughout the year. In months of high Cyanobacteria presence the majority of cyanobacterial sequences were classified as Pleurocapsa. Phylogenetic groups present throughout the year (e.g., Saprospiraceae, Rhodobacteraceae, members without known relatives within Gammaproteobacteria, Desulfatitalea, and members without known relatives within Desulfocapsaceae) constituted most of the sequences, while less abundant taxa showed seasonal patterns connected to the macrophyte growth cycle. Taken together, epiphytic microbial communities of the seagrass C. nodosa and the macroalga C. cylindracea appear to be host-specific and contain taxa that undergo successional changes

Eukaryotic Microbes, Principally Fungi and Labyrinthulomycetes, Dominate Biomass on Bathypelagic Marine Snow

In the bathypelagic realm of the ocean, the role of marine snow as a carbon and energy source for the deep-sea biota and as a potential hotspot of microbial diversity and activity has not received adequate attention. Here, we collected bathypelagic marine snow by gentle gravity filtration of sea water onto μm filters from similar to 1000 to 3900 m to investigate the relative distribution of eukaryotic microbes. Compared with sediment traps that select for fast-sinking particles, this method collects particles unbiased by settling velocity. While prokaryotes numerically exceeded eukaryotes on marine snow, eukaryotic microbes belonging to two very distant branches of the eukaryote tree, the fungi and the labyrinthulomycetes, dominated overall biomass. Being tolerant to cold temperature and high hydrostatic pressure, these saprotrophic organisms have the potential to significantly contribute to the degradation of organic matter in the deep sea. Our results demonstrate that the community composition on bathypelagic marine snow differs greatly from that in the ambient water leading to wide ecological niche separation between the two environments

Development and Deployment of a Point-Source Digital Inline Holographic Microscope for the Study of Plankton and Particles to a Depth of 6000 m

Bochdansky, A. B., Jericho, M. H., & Herndl, G. J. (2013). Development and deployment of a point-source digital inline holographic microscope for the study of plankton and particles to a depth of 6000 m. Limnology and Oceanography: Methods, 11, 28-40. doi: 10.4319/lom.2013.11.2

Role of Macroscopic Particles in Deep-Sea Oxygen Consumption

Macroscopic particles (\u3e 500 µg), including marine snow, large migrating zooplankton, and their fast-sinking fecal pellets, represent primary vehicles of organic carbon flux from the surface to the deep sea. In contrast, freely suspended microscopic particles such as bacteria and protists do not sink, and they contribute the largest portion of metabolism in the upper ocean. In bathy- and abyssopelagic layers of the ocean (2,000-6,000 m), however, microscopic particles may not dominate oxygen consumption. In a section across the tropical Atlantic, we show that macroscopic particle peaks occurred frequently in the deep sea, whereas microscopic particles were barely detectable. In 10 of 17 deep-sea profiles (\u3e 2,000 m depth), macroscopic particle abundances were more strongly cross-correlated with oxygen deficits than microscopic particles, suggesting that biomass bound to large particles dominates overall deep-sea metabolism

Comparison between MICRO-CARD-FISH and 16S rRNA gene clone libraries to assess the active versus total bacterial community in the coastal Arctic

We collected surface- and deep-water samples (maximum depth 300m) during the springsummer transition in the coastal Arctic along a transect in the Kongsfjorden (Ny-angstrom lesund, Spitsbergen, Norway) to determine the structure of the active versus total marine bacterioplankton community using different approaches. Catalysed reporter depositionfluorescence in situ hybridization combined with microautoradiography (MICROCARDFISH) was used to determine the abundance and activity of different bacterial groups. The bacterial communities were dominated by members of Alphaproteobacteria followed by Bacteroidetes, whereas Gammaproteobacteria were present at low abundance but exhibited a high percentage of active cells taking up leucine. The clone libraries of 16S rRNA genes (16S rDNA) and 16S rRNA from two different depths were used to decipher the bacterial community structure. Independently of the type of clone libraries analysed (16S rDNA- or 16S rRNA-based), four major and four minor taxonomic groups were detected. The bacterioplankton community was mainly dominated at both the DNA and the RNA levels by Alphaproteobacteria followed by Gammaproteobacteria. The Rhodobacteriaceae were the most abundant members of the Alphaproteobacteria in both DNA and RNA clone libraries, followed by the SAR11 clade, which was only detectable at the 16S rDNA level. Moreover, there was a general agreement between the results obtained with both techniques, although some specific phylogenetic groups, such as SAR11 and Roseobacter, deviated substantially from this relation. These discrepancies are most likely linked to different physiological states among members of the bacterioplankton community. Combined, MICROCARDFISH and DNA and RNA clone libraries, however, allowed for accurately quantifying different bacterial groups and their activity as well as a detailed phylogenetic insight into the fractions of present versus metabolically active bacterial groups

Response to Interpreting the results of oceanic mesoscale enrichment experiments: Caveats and lessons from limnology and coastal ecology

Peer Reviewe

Prokaryotic respiration and production in the meso- and bathypelagic realm of the eastern and western North Atlantic basin

We measured prokaryotic production and respiration in the major water masses of the North Atlantic down to a depth of,4,000 m by following the progression of the two branches of North Atlantic Deep Water (NADW) in the oceanic conveyor belt. Prokaryotic abundance decreased exponentially with depth from 3 to 0.4 3 105 cells mL21 in the eastern basin and from 3.6 to 0.3 3 105 cells mL21 in the western basin. Prokaryotic production measured via 3H-leucine incorporation showed a similar pattern to that of prokaryotic abundance and decreased with depth from 9.2 to 1.1 mmol C m23 d21 in the eastern and from 20.6 to 1.2 mmol C m23 d21 in the western basin. Prokaryotic respiration, measured via oxygen consumption, ranged from about 300 to 60 mmol C m23 d21 from,100 m depth to the NADW. Prokaryotic growth efficiencies of,2 % in the deep waters (depth range 1,200–4,000 m) indicate that the prokaryotic carbon demand exceeds dissolved organic matter input and surface primary production by 2 orders of magnitude. Cell-specific prokaryotic production was rather constant throughout the water column, ranging from 15 to 32 3 1023 fmol C cell21 d21 in the eastern and from 35 to 58

Estimating Carbon Flux From Optically Recording Total Particle Volume at Depths Below the Primary Pycnocline

Optical instruments can rapidly determine numbers and characteristics of water column particles with high sensitivity. Here we show the usefulness of optically assessed total particle volume below the main pycnocline to estimate carbon export in two systems: the open subarctic North Atlantic and the Ross Sea, Antarctica. Both regions exhibit seasonally high phytoplankton production and efficient export (i.e., a strong biological pump). Total particle volumes in the mesopelagic (200-300 m) were significantly correlated with those in the overlying surface mixed layer (50-60 m), indicating that most particles at depth reflect export from the surface. This connectivity, however, is modulated by the physical structure of the water column and by particle type (e.g., the presence of colonies of the haptophyte Phaeocystis antarctica versus diatoms). Evidence from both regions show that a strong pycnocline can delay or may even prevent particles from settling to deeper layers, which then succumb to disintegration, and microbial and zooplankton consumption. Strong katabatic winds in the Ross Sea may deepen the mixed layer, causing a rapid transfer of particles to mesopelagic depths through the mixed-layer pump. Independent estimates of seasonally integrated export production in the Ross Sea, based on upper water column carbon mass balance, were significantly correlated (in the order of shared variance) with (1) total particle volumes from images, (2) particulate organic carbon, and (3) chlorophyll fluorescence, all recorded at a depth range of 200-300 m. Carbon export was not significantly correlated with particle abundance measured by a Coulter counter at the same depth range. Measuring total particle volume below the primary pycnocline is therefore a useful approach to estimate carbon export at least in regions characterized by seasonally high particle export