New Tracer to Estimate Community Predation Rates of Phagotrophic Protists

Predation of eukaryotic microbes on prokaryotes is one of the most important trophic interactions on Earth, representing a major mortality term and shaping morphology and composition of prokaryotic communities. Here we introduce and validate a new tracer to determine predation rates on prokaryotes. Minicells of Escherichia coli marked with a bright green fluorescent protein (GFP) vector have many operational advantages over previously used prey analogs such as fluorescently labeled bacteria. GFP-minicells are similar in size to naturally occurring bacteria from a variety of environments including the oligotrophic open ocean and the deep sea. They are relatively stable against microbial and light degradation, are easy to grow and process, and can be produced inexpensively in large numbers. No chemical alteration of the particle surface due to heat killing and staining is involved. Grazing coefficients were compared between GFP-minicells and other GFP-modified bacteria, as well as 5-(4,6-dichlorotriazinyl) aminofluorescein (DTAF)stained cells. The grazing coefficients obtained from the removal of GFP-minicells compared favorably with estimates from tracer-independent estimates of grazing in the same experiments. Experiments with GFP-minicells resulted in community grazing coefficients similar to those reported for many different marine environments and those derived using various methods

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

Comparison of four clones of the ichthyotoxic flagellate Prymnesium

Since the mid 1980s blooms of the ichthyotoxic flagellate Prymnesium parvum have resulted in recurrent fish kills in Texas lakes, rivers, and reservoirs. South Carolina experienced a bloom of P. parvum in a brackish golf course pond in summer 2001. No dead fish were reported since the pond had no resident fish. The following year at Artesian Aquafarms in N.C., all hybrid striped bass perished to blooms of P. parvum. In the present study, clonal cultures from each of these blooms were grown in laboratory studies to determine response variation with nutrient-stressed (N-limited, P-limited) and replete cells for growth, hemolytic activity, and ichthyotoxicity. A congener, P. calathiferum originally isolated from a New Zealand bloom, was used for comparison. Of the P. parvum clones, the TX clone overall grew slower (0.21-0.31 div.d-1), had lower hemolytic activity (40-7164 units), but had the highest ichthyotoxicity (1 hr to kill fish in P-limited, 3 hrs in Replete and N-limited). This clone was the most sensitive to nutrient stress and conditioning was reduced to 1 week. In contrast, overall growth and hemolytic activity were greater in the NC (0.21-0.56 div.d-1, 77-21399 units) and SC clones (0.20-0.70 div·d-1, 45-20795 units) with lower ichthyotoxicity for both (>4 hrs in N-&P-limited). Prymnesium calathiferum showed substantially lower hemolytic activity (8-779 units), but grew faster (0.30-0.73 div.d-1) than the P. parvum clones. Nitrogen-deficient cultures were similar to or more hemolytic than P-deficient cultures for P. parvum, but the P-deficient cultures were the most ichthyotoxic. Under nutrient-replete conditions, P. calathiferum was the most ichthyotoxic of the clones with fish mortality occurring in one hour as compared to three hours for the TX clone. Toxicity in P. parvum is a complex interaction of hemolytic and ichthyotoxic components

Nasal associated lymphoid tissue of the Syrian golden hamster expresses high levels of PrPC.

The key event in the pathogenesis of the transmissible spongiform encephalopathies is a template-dependent misfolding event where an infectious isoform of the prion protein (PrPSc) comes into contact with native prion protein (PrPC) and changes its conformation to PrPSc. In many extraneurally inoculated models of prion disease this PrPC misfolding event occurs in lymphoid tissues prior to neuroinvasion. The primary objective of this study was to compare levels of total PrPC in hamster lymphoid tissues involved in the early pathogenesis of prion disease. Lymphoid tissues were collected from golden Syrian hamsters and Western blot analysis was performed to quantify PrPC levels. PrPC immunohistochemistry (IHC) of paraffin embedded tissue sections was performed to identify PrPC distribution in tissues of the lymphoreticular system. Nasal associated lymphoid tissue contained the highest amount of total PrPC followed by Peyer's patches, mesenteric and submandibular lymph nodes, and spleen. The relative levels of PrPC expression in IHC processed tissue correlated strongly with the Western blot data, with high levels of PrPC corresponding with a higher percentage of PrPC positive B cell follicles. High levels of PrPC in lymphoid tissues closely associated with the nasal cavity could contribute to the relative increased efficiency of the nasal route of entry of prions, compared to other routes of infection

NALT contains significantly more PrP^C than other lymphoid tissues.

<p>A) Western blot analysis and B) normalized quantification of lymphatic tissue PrP^C abundance. SP—spleen, SLN—submandibular lymph node, MLN—mesenteric lymph node, PP—Peyer’s patch, NALT—nasal associated lymphoid tissue.</p

PrP^C migration patterns were distinct for different lymphoid tissues.

<p>A) Migration banding patterns are consistent when comparing individual animal homogenates. SP, SLN and PP homogenates from individual animals migrate similarly. B) Line graph of lane intensity analysis demonstrating peak differences in intensity of PrP^C migration. C) Western blot of lymphoid tissue (NALT, SP, and PP) and BR control. D) Line graph of lane analysis of PrP^C intensity of lymphoid tissue indicates intensity difference and migration profile of NALT. E) Comparison of PNGase treated and untreated PrP^C from lymphoid tissue (SLN, PP and NALT and BR) demonstrates variable levels of full length and truncated PrP^C in samples. BR—brain, SP—spleen, PP—Peyer’s patch, SLN—submandibular lymph node, NALT—nasal associated lymphoid tissue.</p

Immunohistochemistry (IHC) of lymphoid tissues demonstrating presence and localization of PrP^C within B cell follicles.

<p>IHC was performed on A/B) NALT, C) MLN, D) SLN, E) PP and F) SP with the anti-PrP antibody 3F4 (B-F) or an isotype control (A). The tissue sections were processed identically using the same reagents at the same time to illustrate relative differences in PrP^C expression between tissues. The scale bar represents 100 μm. NALT—nasal associated lymphoid tissue, MLN—mesenteric lymph node, SLN—submandibular lymph node, PP—Peyer’s patch, SP—spleen.</p

Mesoscale and high-frequency variability of macroscopic particles (> 100 μm) in the Ross Sea and its relevance for late-season particulate carbon export

The Ross Sea plays a major role in the transfer of organic carbon from the surface into the deep sea due to the combination of high seasonal productivity and Antarctic bottom water formation. Here we present a particle inventory of the Ross Sea based on a combined deployment of a video particle profiler (VPP) and a high-resolution digital holographic microscope (DIHM). Long-distance (100s of kilometers) and short-distance (10s of kilometers) sections showed high variability of particle distributions that co-varied with the density structure of the water column. Particle export was apparent at sites of locally weakened pycnoclines, likely an indirect effect of nutrient mixing into the surface layer and local blooms that lead to export. Particle volume abundances at 200–300m depth were highly correlated with particle volume abundances in the upper mixed layer (<60m), consistent with particles at depth primarily the result of export rather than lateral advection. Phaeocystis antarctica (Haptophyta) colonies that were initially retained in the mixed layer sank below the euphotic zone within a period of two weeks. Fine-scale analysis at a resolution<1m revealed a significantly overdispersed (i.e., highly patchy) environment in all casts. Patchiness, as determined by the Lloyd index of patchiness and the Index of Aggregation, increased in and below the pycnocline presumably due to aggregation of particles while accumulating on density gradients. In contrast, particles in the upper mixed layer and in the nepheloid layers were more randomly distributed. In 40 of the 84 VPP depth profiles, a periodicity of particle peaks ranged from 10 to 90m with a mode of 30m, which can be regarded as the “relevant scale” or “characteristic patch size” of the vertical distribution of particles. While chlorophyll fluorescence and particle mass determined by VPP were significantly correlated at higher particle abundances, the relationship changed from cast to cast, reflecting changes in the relative contribution of fresh phytoplankton to total particle mass. Particles that sank below the main pycnocline were composed of phytoplankton, marine snow with and without embedded phytoplankton, crustacean plankton, and a surprisingly high percentage of heterotrophic (and perhaps mixotrophic) protists, such as acantharians and tintinnids. •Particle profiles using a video plankton profiler and a digital inline holographic microscope•Particle abundance at the surface is inversely related to the strength of the main pycnocline.•Particle abundance at depth is significantly correlated with that at the surface.•Phytoplankton and zooplankton comprise a large amount of exported particles.•The relationship between chlorophyll and particle abundance varies greatly