INT Toxicity over Natural Bacterial Assemblages from Surface Oligotrophic Waters: Implications for the Assessment of Respiratory Activity.

Plankton community respiration (R) is a major component of the carbon flux in aquatic ecosystems. However, current methods to measure actual respiration from oxygen consumption at relevant spatial scales are not sensitive enough in oligotrophic environments where respiration rates are very low. To overcome this drawback, more sensitive indirect enzymatic approaches are commonly used as R proxies. The in vivo electron transport system (ETSvivo) assay, which measures the reduction of (2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyl tetrazolium chloride salt, INT) to INT-formazan in the presence of natural substrate levels, was recently proposed as an indirect reliable estimation of R for natural plankton communities. However, under in vivo conditions, formazan salts could be toxic to the cells. Here, we test the toxicity of 0.2 mM of final INT concentration, widely used for ETSvivo assays, on natural bacterial assemblages collected in coastal and oceanic waters off Gran Canaria (Canary Islands, subtropical North Atlantic), in eight independent experiments. After 0.5 h of incubation, a significant but variable decline in cell viability (14-49%) was observed in all samples inoculated with INT. Moreover, INT also inhibited leucine uptake in less than 90 min of incubation. In the light of these results, we argue that enzymatic respiratory rates obtained with the ETSvivo method need to be interpreted with caution to derive R in oceanic regions where bacteria largely contribute to community respiration. Moreover, the variable toxicity on bacterial assemblages observed in our experiments questions the use of a single R/ETSvivo relationship as a universal proxy for regional studies.

A new approach for incorporating 15N isotopic data into linear inverse ecosystem models with Markov Chain Monte Carlo sampling.

Oceanographic field programs often use δ15N biogeochemical measurements and in situ rate measurements to investigate nitrogen cycling and planktonic ecosystem structure. However, integrative modeling approaches capable of synthesizing these distinct measurement types are lacking. We develop a novel approach for incorporating δ15N isotopic data into existing Markov Chain Monte Carlo (MCMC) random walk methods for solving linear inverse ecosystem models. We test the ability of this approach to recover food web indices (nitrate uptake, nitrogen fixation, zooplankton trophic level, and secondary production) derived from forward models simulating the planktonic ecosystems of the California Current and Amazon River Plume. We show that the MCMC with δ15N approach typically does a better job of recovering ecosystem structure than the standard MCMC or L2 minimum norm (L2MN) approaches, and also outperforms an L2MN with δ15N approach. Furthermore, we find that the MCMC with δ15N approach is robust to the removal of input equations and hence is well suited to typical pelagic ecosystem studies for which the system is usually vastly under-constrained. Our approach is easily extendable for use with δ13C isotopic measurements or variable carbon:nitrogen stoichiometry.

Microplastics in the Solent estuarine complex, UK: An initial assessment.

Microplastics are known to be an increasing component found within both marine sediments and the water column. This study carried out an initial assessment of the levels of microplastics present within the Solent estuarine complex, focusing specifically on the water column. A plankton net trawl survey was carried out, with samples analysed using visual observation and Fourier Transform Infrared Spectroscopy (FT-IR). The study identified significant quantities of plastics, ranging in shape, with hot spots found at confluence points within the estuary. Though the FT-IR analysis was inconclusive, the nature of the samples indicates the effect of oceanographic conditions on the prevalent types of microplastics found, which in turn identifies key local sources such as wastewater treatment plants and the plastics industry as being the dominant inputs.

Local versus basin-scale limitation of marine nitrogen fixation.

Nitrogen (N) fixation by diazotrophic plankton is the primary source of this crucial nutrient to the ocean, but the factors limiting its rate and distribution are controversial. According to one view, the ecological niche of diazotrophs is primarily controlled by the ocean through internally generated N deficits that suppress the growth of their competitors. A second view posits an overriding limit from the atmosphere, which restricts diazotrophs to regions where dust deposition satisfies their high iron (Fe) requirement, thus separating N sources from sinks at a global scale. Here we use multiple geochemical signatures of N2 fixation to show that the Fe limitation of diazotrophs is strong enough to modulate the regional distribution of N2 fixation within ocean basins--particularly the Fe-poor Pacific--but not strong enough to influence its partition between basins, which is instead governed by rates of N loss. This scale-dependent limitation of N2 fixation reconciles local observations of Fe stress in diazotroph communities with an inferred spatial coupling of N sources and sinks. Within this regime of intermediate Fe control, the oceanic N reservoir would respond only weakly to enhanced dust fluxes during glacial climates, but strongly to the reduced fluxes hypothesized under anthropogenic climate warming.

Assessment of metals bound to marine plankton proteins and to dissolved proteins in seawater.

Studies based on laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) have been performed to assess metal bound to dissolved proteins and proteins from marine plankton after two-dimensional polyacrylamide gel electrophoresis (2D PAGE). Dissolved proteins were pre-concentrated from surface seawater (60 L) by tangential ultrafiltration with 10 kDa molecular weight cut-off (MWCO) membranes and further centrifugal ultrafiltration (10 kDa) before proteins isolation by methanol/chloroform/water precipitation. Proteins isolation from plankton was assessed after different trichloroacetic acid (TCA)/acetone and methanol washing stages, and further proteins extraction with a phenol solution. LA-ICP-MS analysis of the electrophoretic profiles obtained for dissolved proteins shows the presence of Cd, Cr, Cu, and Zn in five spots analyzed. These proteins exhibit quite similar molecular weights (within the 10-14 kDa range) and pIs (from 5.8 to 7.3). Cd, Cr, Cu, and Zn have also been found to be associated to proteins isolated from plankton samples. In this case, Cd has been found to be bound to proteins of quite different molecular weight (9, 13 and 22 kDa) and pIs (4.5, 5.2, 5.5, and 10). However, trace elements such as Cr, Cu and Zn appear to be mainly bound to plankton proteins of low molecular weight and variable pI.

'Halophyte filters': the potential of constructed wetlands for application in saline aquaculture.

World consumption of seafood continues to rise, but the seas and oceans are already over-exploited. Land-based (saline) aquaculture may offer a sustainable way to meet the growing demand for fish and shellfish. A major problem of aquaculture is nutrient waste, as most of the nutrients added through feed are released into the environment in dissolved form. Wetlands are nature's water purifiers. Constructed wetlands are commonly used to treat contaminated freshwater effluent. Experience with saline systems is more limited. This paper explores the potential of constructed saline wetlands for treating the nutrient-rich discharge from land-based saline aquaculture systems. The primary function of constructed wetlands is water purification, but other ancillary benefits can also be incorporated into treatment wetland designs. Marsh vegetation enhances landscape beauty and plant diversity, and wetlands may offer habitat for fauna and recreational areas. Various approaches can be taken in utilizing plants (halophytes, macro-algae, micro-algae) in the treatment of saline aquaculture effluent. Their strengths and weaknesses are reviewed here, and a conceptual framework is presented that takes into account economic and ecological benefits as well as spatial constraints. Use of the framework is demonstrated for assessing various saline aquaculture systems in the southwestern delta region of the Netherlands.

A metaproteomic assessment of winter and summer bacterioplankton from Antarctic Peninsula coastal surface waters.

A metaproteomic survey of surface coastal waters near Palmer Station on the Antarctic Peninsula, West Antarctica, was performed, revealing marked differences in the functional capacity of summer and winter communities of bacterioplankton. Proteins from Flavobacteria were more abundant in the summer metaproteome, whereas winter was characterized by proteins from ammonia-oxidizing Marine Group I Crenarchaeota. Proteins prevalent in both seasons were from SAR11 and Rhodobacterales clades of Alphaproteobacteria, as well as many lineages of Gammaproteobacteria. The metaproteome data were used to elucidate the main metabolic and energy generation pathways and transport processes occurring at the microbial level in each season. In summer, autotrophic carbon assimilation appears to be driven by oxygenic photoautotrophy, consistent with high light availability and intensity. In contrast, during the dark polar winter, the metaproteome supported the occurrence of chemolithoautotrophy via the 3-hydroxypropionate/4-hydroxybutyrate cycle and the reverse tricarboxylic acid cycle of ammonia-oxidizing archaea and nitrite-oxidizing bacteria, respectively. Proteins involved in nitrification were also detected in the metaproteome. Taurine appears to be an important source of carbon and nitrogen for heterotrophs (especially SAR11), with transporters and enzymes for taurine uptake and degradation abundant in the metaproteome. Divergent heterotrophic strategies for Alphaproteobacteria and Flavobacteria were indicated by the metaproteome data, with Alphaproteobacteria capturing (by high-affinity transport) and processing labile solutes, and Flavobacteria expressing outer membrane receptors for particle adhesion to facilitate the exploitation of non-labile substrates. TonB-dependent receptors from Gammaproteobacteria and Flavobacteria (particularly in summer) were abundant, indicating that scavenging of substrates was likely an important strategy for these clades of Southern Ocean bacteria. This study provides the first insight into differences in functional processes occurring between summer and winter microbial communities in coastal Antarctic waters, and particularly highlights the important role that 'dark' carbon fixation has in winter.

Effect of restriction endonucleases on assessment of biodiversity of cultivable polar marine planktonic bacteria by amplified ribosomal DNA restriction analysis.

To choose a suitable restriction endonuclease for quick assessment of bacterial diversity in polar environments by ARDRA, we investigated the effect of restriction enzymes on ARDRA patterns of cultivable marine planktonic bacteria isolated from polar region. Thirty-three isolates were analyzed by ARDRA using five enzymes (HinfI, HaeIII, AluI, and the mix AfaI/MspI), respectively, resulting in different groups, each group corresponding to a particular genotype. A comparison of the ARDRA patterns was carried out, and phylogenetic position of all thirty-three bacteria was obtained by 16S rDNA sequencing. Consistent with phylogenetic analysis, ARDRA pattern comparison revealed that AluI, being sensitive and reliable enough to generate species-specific patterns, was a suitable restriction enzyme used for evaluating bacterial diversity, suggesting a combination of ARDRA with AluI and 16S rDNA sequencing can provide a simple, fast and reliable means for bacterial identification and diversity assessment in polar environments.

Metabolites from algae with economical impact.

In order to survive in a highly competitive environment, freshwater or marine algae have to develop defense strategies that result in a tremendous diversity of compounds from different metabolic pathways. Recent trends in drug research from natural sources have shown that algae are promising organisms to furnish novel biochemically active compounds. The current review describes the main substances biosynthesized by algae with potential economic impact in food science, pharmaceutical industry and public health. Emphasis is given to fatty acids, steroids, carotenoids, polysaccharides, lectins, mycosporine-like amino acids, halogenated compounds, polyketides and toxins.

Fatty acids from lipids of marine organisms: molecular biodiversity, roles as biomarkers, biologically active compounds, and economical aspects.

Because of their characteristic living environments, marine organisms produce a variety of lipids. Fatty acids constitute the essential part of triglycerides and wax esters, which are the major components of fats and oils. Nevertheless, phospholipids and glycolipids have considerable importance and will be taken into account, especially the latter compounds that excite increasing interest regarding their promising biological activities. Thus, in addition to the major polyunsaturated fatty acids (PUFA) such as eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids, a great number of various fatty acids occur in marine organisms, e.g. saturated, mono- and diunsaturated, branched, halogenated, hydroxylated, methoxylated, non-methylene-interrupted. Various unprecedented chemical structures of fatty acids, and lipid-containing fatty acids, have recently been discovered, especially from the most primitive animals such as sponges and gorgonians. This review of marine lipidology deals with recent advances in the field of fatty acids since the end of the 1990s. Different approaches will be followed, mainly developing biomarkers of trophic chains in marine ecosystems and of chemotaxonomic interest, reporting new structures, especially those with biological activities or biosynthetic interest. An important part of this review will be devoted to the major PUFA, their relevance to health and nutrition, their biosynthesis, their sources (usual and promising) and market.

Flow cytometry assessment of bacterioplankton in tropical marine environments.

Flow cytometry was used to characterize bacterioplankton from two tropical environments in Brazil: the eutrophic Guanabara Bay and the oligotrophic southwest Atlantic Ocean. Bacterial abundance was evaluated by flow cytometry, and cells were stained with SYTO 13, allowing demonstration of differences in nucleic acid content. Bacterial production was also evaluated by means of 3H-leucine incorporation. Bacterial numbers were different for both sites. In Atlantic Ocean samples, we found a maximum of 5.50 x 10(5) cells ml(-1), and low nucleic acid content organisms predominated. In Guanabara Bay, bacterial numbers were one order of magnitude higher than in the ocean, and they varied from outer bay (1.01 x 10(6) cells ml(-1)) to inner bay (6.90 x 10(6) cells ml(-1)). Bacterial activity in ocean samples varied from 4.6 to 126 ng C l(-1) h(-1), while in the bay, mean values ranged from 1.95 microg C l(-1) h(-1) (outer bay) to 7.35 microg C l(-1) h(-1) (inner bay). Values found for both parameters are characteristic of different trophic situations. These results illustrate the utility of cytometric analyses of bacterioplankton populations in characterizing their large spatial and temporal scales of distribution in aquatic ecosystems.

[Annual cycle and budgets of nutrients in the Bohai Sea].

High concentrations of dissolved inorganic nitrogen and phosphate contributed much to the environmental problems in the Bohai Sea in the last decade. It is important to understand the cycle and the budget of nutrients for the environmental management. A three-dimensional ecosystem model, coupled with a physical transport model, was adopted in this study. The simulation of the year 1982 was validated by data in 1982/1983. There were depletion from spring to summer and elevation from autumn to winter for nutrients. The higher concentration of phosphate was found at the Bohai Bay and the concentration of phosphate maintained higher level in the whole year. The higher concentration of dissolved inorganic nitrogen located near the Huanghe Estuary. Production and respiration were the most important sink and source of nutrients. The remineralization of the detritus pool can compensate 30 percent of the consumption of nutrient by the production process. The inputs of phosphates and nitrogen from river were P 0.55 x 10(3) t and N 52.7 x 10(3) t.

Assessment of oceanic productivity with the triple-isotope composition of dissolved oxygen.

Plant production in the sea is a primary mechanism of global oxygen formation and carbon fixation. For this reason, and also because the ocean is a major sink for fossil fuel carbon dioxide, much attention has been given to estimating marine primary production. Here, we describe an approach for estimating production of photosynthetic oxygen, based on the isotopic composition of dissolved oxygen of seawater. This method allows the estimation of integrated oceanic productivity on a time scale of weeks.

Domoic acid-producing diatom blooms in Monterey Bay, California: 1991-1993.

During the autumn of 1991, numerous seabird fatalities in Monterey Bay, California, led to the discovery of a new domoic acid-producing diatom, Pseudonitzschia australis. Since this initial event, sizable populations of P. australis, as well as other likely toxin producers, P. pungens f. multiseries and P. pseudodelicatissima, have occurred biannually in Monterey Bay. Using the highly sensitive FMOC-HPLC method, we detected domoic acid whenever Pseudonitzschia australis was found in the plankton, even at densities as low as 4.0 x 10(3) cells/L. Based on correlations of domoic acid and P. australis abundances and the overwhelming biovolume dominance of P. australis, we conclude that P. australis has been the major domoic acid producer during the period of our study. Our study suggests that P. australis cells may always be toxic in natural populations and that toxin concentrations on a per cell basis have no statistically significant relationship to population density or to nutrient concentrations other than silicate. Cellular levels of domoic acid were positively correlated with silicate concentrations, which is at variance with reports from prior culture experiments. These conclusions must be tentative because of the limited extent of our sampling. Nevertheless, these preliminary data indicate that further investigations of environmental conditions affiliated with cell growth and toxin production in P. australis are warranted. As a practical matter, domoic acid in the pelagic environment cannot be reliably or consistently detected by monitoring domoic acid levels in intertidal mussels. Direct measurement of domoic acid using sensitive HPLC methods is probably the most cost-effective and accurate approach for an ongoing phycotoxin monitoring program.

Rapid analytical technique for the assessment of cell metabolic activity in marine microalgae.

A standard method for the assessment of cell viability has been developed for marine phytoplankton using an inexpensive stain, fluorescein diacetate (FDA), at .75 microM for 10 min. A flow cytometer was used as the fluorescence detector, providing an assessment of viability for each individual particle. Cell size and chlorophyll fluorescence per cell were assessed simultaneously, permitting an assignment of viability to specific subpopulations, thus increasing the power of the technique. A reasonable correspondence between FDA mean fluorescence intensity per cell and an independent metabolic indicator, photosynthetic capacity measured by 14C, was found. Both FDA mean fluorescence intensity and photosynthetic capacity vary as a function of cell volume. Recovery after extended periods of darkness indicate that cells that are FDA negative may not be dead, but merely quiescent or inactive.