André Vaquer

Modelling nitrogen, primary production and oxygen in a Mediterranean lagoon. Impact of oysters farming and inputs from the watershed

An ecosystem model based on nitrogen cycling and oxygen has been developed for the Thau lagoon. It takes into account the specific features of this Mediterranean lagoon, a semi-confined system with watershed inputs and oyster farming. The ecosystem model uses currents calculated by a two-dimensional hydrodynamic model and integrated into a box model. This model is compared with a year survey data and used to estimate nitrogen and oxygen fluxes between the different ecosystem compartments. The yearly simulation shows that the ecosystem behavior is driven by meteorological forcing, especially rain which causes watershed inputs. These inputs trigger microphytoplankton growth, which is responsible for new primary production. During dry periods, nitrogen is recycled into the lagoon thanks to oysters excretion, sediment release, microzooplankton excretion and mineralization. Ammonium produced in this way is consumed by a population of pico- and nanophytoplankton causing regenerated primary production. Consequently, the ecosystem remains highly productive in summer even without external inputs. Shellfish farming also plays an important role in the whole lagoon through biodeposition. Driven by biodeposition, sediment release is the major source of nitrogen in the water column and causes oxygen reduction. The oysters contribute to the recycling activity by excretion, which supports the regenerated primary production. They are also involved in oxygen consumption by respiration which can cause local hypoxia. Further improvements are proposed before this model may become a functional environmental model for a lagoon ecosystem.

NITROGENOUS NUTRITION OF ALEXANDRIUM CATENELLA (DINOPHYCEAE) IN CULTURES AND IN THAU LAGOON, SOUTHERN FRANCE1

Alexandrium catenella (Whedon et Kofoid) Balech was isolated from Thau lagoon (northern Mediterranean) and its growth and uptake characteristics measured for nitrate, ammonium, and urea. Although affinity constants did not indicate a preference for ammonium over nitrate, there was a strong inhibition of nitrate uptake by ammonium when both nitrogen (N) sources were present. Nitrogen budgets during growth in cultures revealed major imbalances between decreases in dissolved N and increases in particulate N, indicating excretion of dissolved organic N during the early part of the growth phase and uptake during the later part. A quasi‐unialgal bloom in November 2001 (4×106 cells·L−1) allowed measurements of uptake of nitrate, nitrite, ammonium, and urea; net and gross growth rate of A. catenella; and grazing rates on this organism. The affinity constants indicate that it is not a strong competitor for the N nutrients tested when these are in low concentrations (<10 μgat N·L−1), compared with other members of the phytoplankton community. Indirect evidence from cultures indicate that dissolved organic N compounds could be important in triggering those blooms. Finally, the strongly unbalanced growth observed in the field indicates that A. catenella exhibits a storage rather than a growth response to a nutrient pulse and is adapted to low frequency events such as the passage of frontal disturbances. The disappearance of A. catenella was due to grazing that balanced growth at the peak of the bloom.

ACCLIMATION OF NITRATE UPTAKE BY PHYTOPLANKTON TO HIGH SUBSTRATE LEVELS

A literature review of data on nitrate uptake by phytoplankton suggests that nitrate levels above 20 μmol N·L−1 generally stimulated uptake rates in cultured unicellular algae and natural phytoplankton communities. This phenomenon indicates that phytoplankton cells acclimate to elevated nitrate levels by increasing their uptake capacity in a range of concentrations previously considered to be saturating. Cyanobacteria and flagellates were found to present a considerable capacity for acclimation, with low (0.1–2 μmol N·L−1) half‐saturation values (Ks) at low (5–20 μmol N·L−1) substrate levels and high (1–80 μmol N·L−1) Ks values at high (30–100 μmol N·L−1) substrate levels. However, some diatom genera (Rhizosolenia, Skeletonema, Thalassiosira) also appeared to possess a low affinity nitrate uptake system (Ks between 18 and 120 μmol N·L−1), which can help resolve the paradox of their presence in enriched seas. It follows that present models of nitrate uptake can severely underestimate the effects of high nitrate concentrations on phytoplankton dynamics and development. A more adequate approach would be to consider the possibility of multiphasic uptake involving several phase transitions as nitrate concentrations increased. Because it is a nonlinear phenomenon featuring strong thresholds, this effect appears to override that of other variables, such as irradiance, temperature, and cell size. Within the present context of eutrophication and for a range of concentrations that is becoming more and more ecologically relevant, equations are tentatively presented as a first approach to estimate Ks from ambient nitrate concentrations.

Recent applications of flow cytometry in aquatic microbial ecology

Summary— Microorganisms (unicellular algae, bacteria) constitute fundamental compartments of aquatic ecosystems because of their high concentrations and activities. The evaluation and understanding of their behaviour and role raise different problems for which traditional methodologies are often inadequate, whether they refer to global or classical microscopic analyses. Flow cytometry (FCM) has been recently used to study microorganisms in aquatic environments. Although this technology is still applied on a limited scale in our field, a large number of works has been done showing that FCM seems to be a promising tool for aquatic microbial ecology. This paper summarizes, from the literature produced during the last decade and with original data obtained in our laboratory, the main questions related to the cell identification, the evaluation of cell viability, biomasses and productions and the measurements of bacterial and phytoplanktonic activities. The representativeness of sampling and observation scales is also discussed within the framework of the FCM measurements.

Unexpected Genetic Diversity among and within Populations of the Toxic Dinoflagellate Alexandrium catenella as Revealed by Nuclear Microsatellite Markers

ABSTRACT Since 1998, blooms of Alexandrium catenella associated with paralytic shellfish poisoning have been repeatedly reported for Thau Lagoon (French Mediterranean coast). Based on data obtained for rRNA gene markers, it has been suggested that the strains involved could be closely related to the Japanese temperate Asian ribotype of the temperate Asian clade. In order to gain more insight into the origin of these organisms, we carried out a genetic analysis of 61 Mediterranean and 23 Japanese strains using both ribosomal and microsatellite markers. Whereas the phylogeny based on ribosomal markers tended to confirm the previous findings, the analysis of microsatellite sequences revealed an unexpected distinction between the French and Japanese populations. This analysis also highlighted great intraspecific diversity that was not detected with the classical rRNA gene markers. The Japanese strains are divided into two differentiated A. catenella lineages: the Sea of Japan lineage and the east coast lineage, which includes populations from the Inland Sea and the Pacific Ocean. A. catenella strains isolated from Thau Lagoon belong to another lineage. These findings indicate that microsatellite markers are probably better suited to investigations of the population genetics of this species that is distributed worldwide. Finally, application of the population genetics concepts available for macroorganisms could support new paradigms for speciation and migration in phytoplankton assemblages.

Patterns in nutrient limitation and chlorophyll a along an anthropogenic eutrophication gradient in French Mediterranean coastal lagoons

A cross-ecosystem comparison of data obtained from 20 French Mediterranean lagoons with contrasting eutrophication status provided the basis for investigating the variables that best predict chlorophyll a (Chl a) concentrations and nutrient limitation of phytoplankton biomass along a strong nutrient enrichment gradient. Summer concentrations of dissolved inorganic nitrogen (DIN) and phosphorus (DIP) comprised only a small fraction of total nitrogen (TN) and total phosphorus (TP). On the basis of inorganic nutrient concentrations, the most oligotrophic lagoons appeared to be phosphorus-limited, with a tendency towards the development of nitrogen limitation as eutrophication increased, as evidenced by decreasing DIN:DIP ratios. A weak but significantly positive relationship was found between dissolved silicate (DSi) and Chl a, reflecting DSi accumulation in the water column along the trophic state gradient and implying a progressive shift away from potential Si limitation of phytoplankton growth. Observed concentrations of Chl a were far better explained by TN and TP than by DIN and DIP concentrations, suggesting that a total nutrient based approach is likely to be the most appropriate for managing eutrophication in Mediterranean lagoons and other coastal waters. These results give credence to the idea that marine and freshwater environments respond in a similar fashion to nutrient enrichment.

Response of coastal phytoplankton to ammonium and nitrate pulses: seasonal variations of nitrogen uptake and regeneration

Seasonal variation in uptake and regeneration of ammonium and nitrate in a coastal lagoon was studied using 15N incorporation in particulate matter and by measuring changes in particulate nitrogen. Uptake and regeneration rates were two orders of magnitude lower in winter than in summer. Summer uptake values were 2.8 and 2.2 μmol N.l−1.d−1 for ammonium and nitrate, respectively. Regeneration rates were 2.9 and 2.1 μmol N.l−1.d−1 for ammonium and nitrate respectively. Regeneration/uptake ratios were often below one, indicating that water column processes were not sufficient to satisfy the phytoplankton nitrogen demand. This implies a role of other sources of nitrogen, such as macrofauna (oysters and epibionts) and sediment. Phytoplankton was well adapted to the seasonal variations in resources, with mixotrophic dinoflagellates dominant in winter, and fast growing diatoms in summer. In winter and spring, ammonium was clearly preferred to nitrate as a nitrogen source, but nitrate was an important nitrogen source in summer because of high nitrification rates. Despite low nutrient levels, the high rates of nitrogen regeneration in summer as well as the simultaneous uptake of nitrate and ammonium allow high phytoplankton growth rates which in turn enable high oyster production.

Bacterial ecology of a young equatorial hydroelectric reservoir (Petit Saut, French Guiana)

This work aims at studying the microbial and physical-chemical changes occurring in an equatorial hydroelectric reservoir Petit Saut (French Guiana), over three years, from the time it first filled.Since filling in January 1994, almost the whole water column has remained anoxic, with high concentrations of reduced elements (CH4, iron sulphides, H2S) originating from degradation of the submerged primary forest. These elements deoxygenated the water drained from the dam, as they were biologically and chemically oxidized in the river. Two major physiological guilds of microorganisms occurred. They showed a characteristic stratified arrangement in the lake, below and above an oxycline. Since flooding, we observed a cyclic development of phototrophic bacteria, assumed to be sulphur-oxidizers below the oxycline. The growth of this physiological guild seems to be strongly influenced by sulphide production and by the alternation between rainy and dry seasons. At the oxycline, the population of methane-oxidizing bacteria did not vary in the same way as the green sulphur bacteria. After a lag-phase, which was probably due to inhibition by excessive illumination in the upper water column in 1994, they developed and completely stopped diffusive methane emission in February 1995. The development of both bacterial guilds is directly correlated to changes in the level of the corresponding reduced elements from the bottom and dissolved in the lake water. The decreasing production of CH4 and sulphides induces a progressive extinction of methanotrophic bacteria and phototrophic sulphur-oxidizing bacteria. Meanwhile, the water is becoming clearer and algal populations proliferate in the upper water column, constituting an autochtonous source of organic carbon. Slowly, this aquatic ecosystem is reaching a low-activity equilibrium level related to reduced element production. It is evolving from a system ruled by an anaerobic organotrophic metabolism to an ecosystem governed by phototrophy.

Influence of nitrogen enrichment on size-fractionated in vitro carboxylase activities of phytoplankton from Thau Lagoon (Coastal Mediterranean Lagoon, France)

The influence of dissolved inorganic and organic nitrogen (DIN and DON) enrichments on pools of enzymes responsible for CO2 fixation by the Calvin-Benson (Rubisco) and beta-carboxylation pathways (beta-carboxylases) were studied in a natural plankton assemblage. The plankton community from a coastal Mediterranean lagoon were incubated in situ for 24 h with initially ammonium, nitrate and DON (taurine) enrichments and compared to a control without any enrichment. An increase of small picophytoplankton and diatom biomass was observed in the enriched inorganic nitrogen treatments 7 - 10 h after the initial enrichment. Phytoplankton biomass decreased in the control and under taurine enrichment suggesting an inorganic nitrogen limitation of phytoplankton growth.

A simple and innovative method for species identification of phytoplankton cells on minute quantities of DNA

Dinoflagellates belonging to the genus Alexandrium are often involved in harmful algal blooms. Their ecological exploration is thus essential to increase our knowledge on these toxic events. Yet, population genetic studies, taxonomic identification and environmental monitoring are hampered by major constraints: the necessity to establish monoclonal cultures from environmental samples and the sensitivity of available molecular tools. The present work describes a very simple and sensitive method for extraction and amplification of DNA at the infra-single-cell level. Its on-slide format allows for easy visual control of both quality and quantity of the templates. Combined with a semi-multiplex PCR protocol designed on the 18S-28S rDNA-ITS region of Alexandrium catenella and Alexandrium tamarense, this procedure allowed the identification and discrimination of these species from both monoclonal cultures and natural samples.