Pierre Gaudin

Spatiotemporal changes in microphytobenthos assemblages in a macrotidal flat (Bourgneuf Bay,France)1

Spatiotemporal changes in microphytobenthos species composition were investigated in relation to structural variables—biomass; life‐forms; detritus ratio, pheopigment a (pheo a):chl a; and sediment characteristics—at mesoscale in a shellfish macrotidal ecosystem. To characterize algae assemblages, multivariate analyses were performed (multidimensional scaling [MDS] ordination and clustering), supported by correlations between structural variables. Microphytobenthos, dominated by 97% diatoms belonging to 89 taxa, was constituted by two main assemblages: The first one, composed of common species (occurrence >50%), mainly nanobenthic (size <30 μm) and epipsammic (sand‐fixed cells), was associated with mixed sediments. The major species contributing to this assemblage were Navicula perminuta, Achnanthes hauckiana, Fallacia tenera, A. lanceolata var. elleptica, Amphora spp., Plagiogrammopsis vanheurkii, and Plagiogramma tenuissimum (see Table 2 for taxonomic authors). Species diversity was high (N1∼ 11), often linked to high biomass values (>70 mg chl a · m−2) and low detritus ratio. Conversely, the second assemblage comprised occasional species (occurrence from 10% to 50%), mainly microbenthic (>30 μm) and epipelic (moving cells), occurring during summer at muddy sites. This assemblage, characterized by low diversity, high biomass variability, and high detritus ratio, was dominated by Scolioneis tumida or Plagiotropis vitrea and Navicula spartinetensis. Whereas hydrodynamics globally explained the gradual assemblage changes throughout the entire mudflat and the year, oyster beds and ridge and runnel features appeared to be local spatial structuring factors allowing the establishment of specific assemblages. This study suggests a significant role for epipsammon biomass, until now underestimated, in the functioning of this turbid coastal ecosystem.

Biochemical and behavioural responses of the endobenthic bivalve Scrobicularia plana to silver nanoparticles in seawater and microalgal food

Because of their bactericidal effects, Ag nanoparticles (Ag NPs) have promising industrial development but could lead to potential ecological risks. The aim of this study was to examine the uptake and effect of silver (soluble or as lactate Ag NPs of 40 nm) at low concentrations (10 μg L(-1)) in the endobenthic bivalve Scrobicularia plana exposed, for 14 days, directly (water) or via the diet (microalgae). The stability of Ag NPs in seawater was examined using dynamic light scattering. Release of soluble Ag from Ag NPs in the experimental media was quantified by using diffusive gradient in thin film. Bioaccumulation of Ag in bivalves was measured by electrothermal atomic absorption spectrometry. Behavioural and biochemical biomarkers were determined in bivalves. Aggregation of Ag NPs and the release of soluble Ag from Ag NPs were observed in the experimental media. For both forms of Ag, bioaccumulation was much more important for waterborne than for dietary exposure. The response of oxidative stress biomarkers (catalase, glutathion S-transferase, superoxide dismutase) was more important after dietary than waterborne exposure to Ag (soluble and NPs). These defences were relatively efficient since they led to a lack of response of damage biomarkers. Burrowing was not affected for bivalves exposed directly or through the diet to both Ag forms but feeding behaviour was impaired after 10 days of dietary exposure. Since no differences of responses to Ag either soluble or nanoparticulate were observed, it seems that labile Ag released from Ag NPs was mainly responsible for toxicity.

Growth form defines physiological photoprotective capacity in intertidal benthic diatoms

In intertidal marine sediments, characterized by rapidly fluctuating and often extreme light conditions, primary production is frequently dominated by diatoms. We performed a comparative analysis of photophysiological traits in 15 marine benthic diatom species belonging to the four major morphological growth forms (epipelon (EPL), motile epipsammon (EPM-M) and non-motile epipsammon (EPM-NM) and tychoplankton (TYCHO)) found in these sediments. Our analyses revealed a clear relationship between growth form and photoprotective capacity, and identified fast regulatory physiological photoprotective traits (that is, non-photochemical quenching (NPQ) and the xanthophyll cycle (XC)) as key traits defining the functional light response of these diatoms. EPM-NM and motile EPL showed the highest and lowest NPQ, respectively, with EPM-M showing intermediate values. Like EPL, TYCHO had low NPQ, irrespective of whether they were grown in benthic or planktonic conditions, reflecting an adaptation to a low light environment. Our results thus provide the first experimental evidence for the existence of a trade-off between behavioural (motility) and physiological photoprotective mechanisms (NPQ and the XC) in the four major intertidal benthic diatoms growth forms using unialgal cultures. Remarkably, although motility is restricted to the raphid pennate diatom clade, raphid pennate species, which have adopted a non-motile epipsammic or a tychoplanktonic life style, display the physiological photoprotective response typical of these growth forms. This observation underscores the importance of growth form and not phylogenetic relatedness as the prime determinant shaping the physiological photoprotective capacity of benthic diatoms.

Light is a key factor in triggering sexual reproduction in the pennate diatom Haslea ostrearia

Sexual reproduction is an obligatory phase in the life cycle of most diatoms, as cell size decreases with successive vegetative divisions and the maximal cell size is only restored by a specialized cell, the auxospore, which follows zygote formation as a result of sexual reproduction. While in pennate diatoms the induction of sexual reproduction depends primarily on cell-cell interactions, the importance of different external factors for the induction of sexual reproduction is less well known. Here, we investigated the effects of light on sexualization in the marine benthic pennate diatom Haslea ostrearia (Gaillon) R. Simonsen. Compatible clones were crossed and exposed to different combinations of light levels, qualities, and photoperiods. Light was found to be a key factor for sexualization, and to a certain extent, to control auxosporulation in H. ostrearia. The light conditions most favorable for sexual reproduction were low irradiances (<50 micromolphotons m(-2) s(-1)) and short photoperiods (6-10 h), conditions that prevail during winter, and to a lesser extent, the higher irradiances and longer photoperiods that correspond to the spring and fall, when blooms of this organism form in the natural environment. Auxospore formation was very rare in continuous light, and maximum in presence of red radiation, while it was never observed in darkness or in radiation other than red.

Biological activities of purified marennine, the blue pigment responsible for the greening of oysters

Marennine, the blue pigment produced by the diatom Haslea ostrearia , exists in two different forms, the intra- and extracellular forms. We investigated the antibacterial, antiviral, and antiproliferative properties of both of these forms. Both forms of marennine inhibited the development of marine bacteria, in particular the pathogenic organism Vibrio aesturianus , at concentrations as low as 1 μg/mL, but they did not display any effect on a wide range of pathogenic bacteria that are relevant for food safety. Both forms of the pigment produced by H. ostrearia also exhibited antiviral activity against the HSV1 herpes virus, with intra- and extracellular marennine having EC(50) values of 24.0 and 27.0 μg/mL, respectively. These values are 2 orders of magnitude higher than the value for the reference drug, Zovirax. Moreover, both forms of marennine were effective in slowing or inhibiting the proliferation of cancer cells. This study confirms the potential of marennine as a biologically active organic molecule, which could have a protective effect on bivalves, which filter seawater and fix the pigment on their gills. Moreover, marennine could be used in food engineering and chemistry as a natural blue pigment. However, despite that it is eaten and possibly assimilated by green oyster consumers, it also deserves in depth evaluation before being considered for use as a nutraceutical.

Effect of enzymatic digestion on thallus degradation and extraction of hydrosoluble compounds from Grateloupia turuturu

The extraction of metabolites of industrial interest from seaweed tissue is difficult due to the presence of anionic polysaccharides in the cell walls. We studied enzymatic digestion of Grateloupia turuturu (Rhodophyta) to facili- tate extraction of the desired metabolites from this sea- weed. Four polysaccharidases (Onozuka R-10 cellulase, agarase, k- and i-carrageenases) were used alone and in combination. We studied the efficiency of the enzy- matic process in degrading the seaweed thallus; we also investigated the effect of the enzymatic treatment on solubilization of R-phycoerythrin and reducing sugars. The best degradation conditions for Grateloupia thalli were obtained with cellulase alone (0.48 U ml -1 ), or with a combination of cellulase/k-carrageenase. However, only the combination cellulase (0.08 U ml -1 )/k-carrageen- ase (0.01 U ml-1), or the individual enzymes wagarase or i-carrageenase at low concentration (0.55 U ml -1 and 2.5=10-3 Um l-1)x improved solubilization of R-phycoery- thrin compared to the control conditions. The simulta- neous use of polysaccharidases enabled solubilization of a large quantity of reducing sugars (40 mg glu g -1 dw). Therefore, this condition appears suitable for the pro- duction of oligosaccharides. This result also confirms indirectly the presence of agar, cellulose, k- and i-car- rageenans in the cell wall of Grateloupia turuturu.

Haslea karadagensis (Bacillariophyta): a second blue diatom, recorded from the Black Sea and producing a novel blue pigment

A new species of raphid pennate diatom producing a blue pigment, Haslea karadagensis Davidovich, Gastineau & Mouget, sp. nov., was recently isolated from the Crimean coast of the Black Sea (Ukraine). This organism is very similar to the well-known Haslea ostrearia, the first described ‘blue’ diatom, which produces marennine, the pigment involved in the greening of oysters. The Ukrainian diatom, H. karadagensis, differs slightly from H. ostrearia in the structure of its frustule, and the two organisms are unable to interbreed. Two molecular markers, rbcL and the ITS1–5.8S–ITS2 sequences, showed 2% and >50% differences, respectively, between the two species. UV-visible spectrophotometry and in vivo confocal micro-Raman spectroscopy were used to compare the pigment of H. karadagensis with marennine. Both pigments showed absorption bands in the UV and red regions, but the positions of the maxima differ between the pigments. Significant differences were observed by micro-Raman spectroscopy in the 1000–1700 cm−1 wavenumber range, revealing that the pigments are different molecules. Haslea karadagensis is the first example of a new ‘blue’ diatom and produces a novel blue pigment.

An evaluation of methods for quantifying the enzymatic degradation of red seaweed Grateloupia turuturu

For better exploitation of the red seaweed Grateloupia, enzymatic digestion of the thallus may be a way to increase access to metabolites of industrial interest. With this aim, we have tried to find a method to quantify the efficiency of enzymatic digestion. Vegetative algal material was treated with polysaccharidases (Onozuka R-10 cellulase, agarase, and Ultraflo L mixture). The proportion of degraded surface area was determined by microscopic measurement of the residue surface using imaging software and compared with the analysis of carbohydrates and R-phycoerythrin released in the incubation solution. Both the reducing carbohydrate concentration and percentage of degraded surface area appeared the most reliable methods to study enzymatic efficiency. The amount of solubilized total carbohydrates, and particularly that of R-phycoerythrin, showed non-specific variations, so no conclusions could be drawn. The application of this procedure to the screening of the efficient digestion of Grateloupia material demonstrated that cell walls were only partially digested by polysaccharidase enzymes alone. The Ultraflo L mixture and Onozuka R-10 cellulase produced a greater degradation of Grateloupia tissues and a higher release of reducing carbohydrates, whereas agarase did not display any specific action. Thus, the proposed procedure based on the quantification of residue surface area seems to be an accurate method to analyze enzymatic digestion. Other tests using different concentrations and combinations of enzymes are now required.

Heterothallism in the pennate diatom Haslea ostrearia (Bacillariophyta)

Twelve clonal cultures of the marine pennate diatom Haslea ostrearia (Gaillon) Simonsen were used to study sexual reproduction and auxosporulation. After crossing, compatible gametangia paired girdle to girdle, with no visible surrounding mucilage. Two gametes were formed per gametangium; these were not attached to the thecae of the parental frustule and gametes from one gametangium fused with the gametes from the other gametangium. Sexual reproduction was isogamous, resulting in the production of two zygotes per paired gametangia. The pattern of reproduction in H. ostrearia can be classified as type IB2a according to Geitlers system. The expansion of auxospores, which was bipolar, was accompanied by the formation of a transverse perizonium. Superfluous nuclei resulting from meiosis were not visible. The upper limit of the cell size range suitable for sexualization was c. 68 µm, i.e. about half the maximum species-specific size of about 140 µm. Sexual reproduction has been shown by previous authors to occur in clonal cultures and our observations confirm that two modes of sexual reproduction, homothallic and heterothallic, coexist in H. ostrearia.

Continuous marennin production by agar-entrapped Haslea ostrearia using a tubular photobioreactor with internal illumination.

Abstract The marine diatom Haslea ostrearia was immobilized in a tubular agar gel layer introduced into a photobioreactor of original design with internal illumination for the continuous synthesis of marennin, a blue-green pigment of biotechnological interest. Marennin was produced for a long-term period (27–43 days) and the volumetric productivity was maximum (18.7 mg day−1 l−1 gel) at the highest dilution rate (0.25 day−1) and lowest agar layer thickness (3 mm). Heterogeneous cell distribution in the agar layer revealed diffusional limitation of light and nutrients. However, the 3 mm gel thickness led to a more homogeneous cell distribution during incubation and to an increase of the whole biomass in the agar gel layer.