Loïc Ten-Hage

FIRST EVIDENCE OF PALYTOXIN ANALOGUES FROM AN OSTREOPSIS MASCARENENSIS (DINOPHYCEAE) BENTHIC BLOOM IN SOUTHWESTERN INDIAN OCEAN1

Benthic dinoflagellates of the genus Ostreopsis Schmidt are common in tropical and subtropical water, and some species produce toxins potentially involved in human intoxication events. A benthic bloom of Ostreopsis mascarenensis Quod was observed near Rodrigues Island during a survey of benthic dinoflagellates in the southwestern Indian Ocean. The morphology of O. mascarenensis was studied by LM and SEM. Preliminary screening of a crude extract of an O. mascarenensis bloom revealed neurotoxicity in mice similar to that induced by palytoxin. After partition of the crude extract, the highest toxicity was retained in the butanol‐soluble fraction, which retained hemolytic activity suggestive of palytoxin analogues. Two new toxins, mascarenotoxin‐A and ‐B, were resolved from this fraction by HPLC coupled to a diode array detector. The closed mass spectrum profile and fragmentation pattern obtained by advanced nano–electrospray ionization quadrupole time‐of‐flight mass spectrometry between purified toxins and a reference palytoxin confirmed the mascarenotoxins as palytoxin analogues. These results were confirmed by tandem mass spectrometry with the identification of specific fragment ion m/z 327. An on‐line liquid chromatography protocol coupled to tandem mass spectrometry was developed for detection of these palytoxin analogues. The present study describes the first purification, chemical, and toxicological characterization of new palytoxin analogues isolated from a benthic bloom of O. mascarenensis. These results suggest that O. mascarenensis, which is largely distributed in the southwestern Indian Ocean, could be a source of palytoxin poisoning in this tropical area.

Chemical interactions in diatoms: role of polyunsaturated aldehydes and precursors

Chemicals produced by aquatic organisms, and especially micro-organisms, have received increasing attention in the last decade for their role in shaping interactions and communities. Several cases emphasize the fact that chemical signals or defence may modulate interspecific interactions. Notably, it has been shown that diatoms, unicellular algae and key primary producers in aquatic ecosystems produce a wide range of bioactive metabolites. Among these compounds, polyunsaturated short-chain aldehydes in vitro strongly impair the reproduction of various potential grazers. In the field, the relationship between aldehyde production and reproductive failure in copepods remains unclear. Recent studies have suggested that these putative defence compounds may also be involved in intercellular communication and in interactions with competitors. Potential effects of the aldehyde precursors on various organisms have also been described. This review presents an overview of various results obtained in the last decade that could help us to understand the role of polyunsaturated aldehydes and their precursors in the ecology of diatoms. It is focused on the dichotomy between freshwater and marine environments. Indeed, most of the results on anti-proliferative aldehydes concern marine planktonic diatoms, whereas they are also known to be produced by benthic and freshwater species.

Coolia areolata sp. nov. (Dinophyceae), a new sand-dwelling dinoflagellate from the southwestern Indian Ocean

Abstract A new benthic dinoflagellate, Coolia areolata Ten-Hage, Turquet, Quod & Couté sp. nov., was isolated from sandy substrata in La Possession Bay, La Réunion, and at various other sites in the western Indian Ocean. This new species is described and illustrated by light and scanning electron micrographs and drawings. The plate formula of C. areolata is P03′7″ 6c 6?s 5′″ 2″″. Cell size ranges are 30–36.5 μm long and 28–36.5 μm wide. Cells are spherical, areolate, and covered with scattered round pores. The surface of the apical plate 1′ is smooth. The apical pore is 9–10 μm long. Cells of C. areolata are distinguished from C. monotis and C.tropicalis by thecal surface morphology, thecal plate size and shape, and apical pore length.

Colony induction and growth inhibition in Desmodesmus quadrispina (Chlorococcales) by allelochemicals released from the filamentous alga Uronema confervicolum (Ulotrichales)

In biofilms, the competition between microorganisms for light, nutrients and space is extreme. Moreover, planktonic algae can be considered as competitors insofar as they decrease the available light for the benthic algae. One of the strategies employed by microorganisms to eliminate competitors is the release of inhibiting compounds, a process known as allelopathy. Here we demonstrate that a benthic/epiphytic alga, Uronema confervicolum, produces allelopathic compounds that induce oxidative stress and growth inhibition in the planktonic Desmodesmus quadrispina. Some of these compounds can also trigger the formation of colony in D. quadrispina. As colonies have higher sedimentation rates than unicells, their induction by U. confervicolum might decrease shading. This study is the first report of colony induction in the context of alga-alga interaction. Our results also suggest the implication of mitogen-activated protein (MAP) kinases in the transduction of the signal leading to the formation of reactive oxygen species in the cells. A comparison with allelochemicals from another planktonic green alga, Monoraphidium aff. dybowski, emphasizes the specificity of colony induction by U. confervicolum, in contrast with oxidative stress which is induced by several compounds. The reciprocal production of inhibiting compounds by D. quadrispina makes this interaction an interesting example of co-evolution between two microorganisms belonging to different compartments of the ecosystem.

Prorocentrum borbonicum sp. nov. (Dinophyceae), a new toxic benthic dinoflagellate from the southwestern Indian Ocean

Abstract A new toxic benthic marine dinoflagellate, Prorocentrum borbonicum Ten-Hage, Turquet, Quod, Puiseux-Dao & Couté sp. nov., isolated from the fringing coral reef of St Leu from La Réunion Island (France, southwest Indian Ocean) is described from light and scanning micrographs. The species was identified based on shape, size, surface micromorphology, ornamentation of thecal plates, and architecture of the periflagellar area and intercalary band. Prorocentrum borbonicum is a small species (18–24 μm long and 16–20 μm wide) with a broadly ovate shape. Thecal depressions are numerous (950–1020 per valve). Two sizes of valve pores were recognized: large pores, which are scattered on the valve surface, except in the central area, and smaller pores, which are localized chiefly in the centre of the valve and at the periphery, along the intercalary band. The periflagellar area is V-shaped and accommodates a flagellar pore and an auxiliary pore, the latter being the smaller of the two. Prorocentrum borbonicum is photosynthetic, containing a golden-brown chloroplast with a centrally located pyrenoid. Prorocentrum borbonicum produces toxic compounds that are lethal to mice. The symptoms induced in mice by methanolic crude extracts indicate neurotoxicity.

Temporal patterns in epilithic biofilm -relation with a putative allelopathic activity

Temporal patterns in biomass and in the production of inhibitory compounds were assessed for a diatom- dominated epilithic biofi lm of a shallow temperate river. Biomass showed a typical seasonal pattern with the de- velopment of two peaks of biomass parallel with species successions. Analysis with self-organizing maps (SOM), a non-supervised neural network, indicated that species composition of phototrophic assemblages was completely explained by environmental parameters during the low-biomass period. When epilithic biomass was high, these fac- tors were not enough to explain species composition. This suggests that as the biomass increased, intrinsic param- eters took over from extrinsic factors in the regulation of biofi lm composition. Over the studied period, 70 % of the biofi lm samples showed inhibitory activity against one or several of the four indicator phototrophic strains tested in the bioassays. The presence of inhibitory compounds was not correlated to biofi lm biomass. The occurrence of several patterns of inhibitory activity suggested the implication of different compounds or mix of compounds, certainly linked to various compositions of algal assemblages. The SOM analysis underlined putative relationships between the occurrence of certain species and the presence of an activity inhibiting one of the indicator strains. This indicates potential targets or producers of the implied compounds. To our knowledge, this study followed for the fi rst time the temporal dynamics of inhibitory activity in a biofi lm. The frequent presence of inhibitory compounds is in agreement with the hypothesis that biofi lms are auspicious systems for the development of allelopathic interac- tions, and provides new insights for further research on allelopathic interactions in benthic phototrophic systems.

A photosynthetic rotating annular bioreactor (Taylor–Couette type flow) for phototrophic biofilm cultures

In their natural environment, the structure and functioning of microbial communities from river phototrophic biofilms are driven by biotic and abiotic factors. An understanding of the mechanisms that mediate the community structure, its dynamics and the biological succession processes during phototrophic biofilm development can be gained using laboratory-scale systems operating with controlled parameters. For this purpose, we present the design and description of a new prototype of a rotating annular bioreactor (RAB) (Taylor-Couette type flow, liquid working volume of 5.04 L) specifically adapted for the cultivation and investigation of phototrophic biofilms. The innovation lies in the presence of a modular source of light inside of the system, with the biofilm colonization and development taking place on the stationary outer cylinder (onto 32 removable polyethylene plates). The biofilm cultures were investigated under controlled turbulent flowing conditions and nutrients were provided using a synthetic medium (tap water supplemented with nitrate, phosphate and silica) to favour the biofilm growth. The hydrodynamic features of the water flow were characterized using a tracer method, showing behaviour corresponding to a completely mixed reactor. Shear stress forces on the surface of plates were also quantified by computer simulations and correlated with the rotational speed of the inner cylinder. Two phototrophic biofilm development experiments were performed for periods of 6.7 and 7 weeks with different inoculation procedures and illumination intensities. For both experiments, biofilm biomasses exhibited linear growth kinetics and produced 4.2 and 2.4 mg cm(-)² of ash-free dry matter. Algal and bacterial community structures were assessed by microscopy and T-RFLP, respectively, and the two experiments were different but revealed similar temporal dynamics. Our study confirmed the performance and multipurpose nature of such an innovative photosynthetic bioreactor for phototrophic biofilm investigations.

Bysmatrum granulosum sp. nov., a new benthic dinoflagellate from the southwestern Indian Ocean

A new benthic marine dinoflagellate, Bysmatrum granulosum Ten-Hage, Turquet, Quod & Couté, sp. nov., was obtained from sediment and coral samples from sites of La Réunion Island (SW Indian Ocean). This new species is described and illustrated by light and scanning electron micrographs. Cells are 40–50 µm long and 40–46 µm wide. The epitheca is conical and smaller than the hypotheca, which is trapezoidal with convex sides. Plate tabulation is typical for the genus (P0, X, 4′, 3a, 7″, 6c, 5s, 5‴, 2″″); epithecal plates 3′ and 4″ separate the intercalary plates 2a and 3a. Thecal plates are perforated by pores and covered by both small and minute wart-like projections, linearly arranged, radiating from the apical pore. This new species differs from the three others of the genus Bysmatrum in the following characters: cell shape and size, size of the apical pore complex and features of thecal plates (plate shapes and ornamentations).

Influence of food web structure on the biochemical composition of seston, zooplankton and recently deposited sediment in experimental freshwater mesocosms

The effects of food web structure on the quantity and biochemical composition of seston, zooplankton and recently deposited sediment in experimental freshwater mesocosms were examined. Food web structure was manipulated by addition of zooplanktivorous fish. Biochemical characterisations were carried out using lipid biomarkers (sterols, fatty acids, chlorophyll-derived compounds and long-chain alkanediols). Fish addition decreased zooplankton biomass and increased seston biomass and deposited sediment through a trophic cascade. Fish presence strongly influenced the biochemical characteristics of seston and sediment. In contrast, food web structure had a minor impact on the lipid biomarker composition of zooplankton. Although the relative abundance of sterols in the different compartments did not differ strongly between treatments, sterol profiles in seston and sediment depended on food web structure. The predominance of Δ7-sterols in seston and sediment in the fish treatment indicated a major contribution of Chlorophyceae. In contrast, the distribution of sterols in seston and sediment in the fishless treatment, dominated by cholesterol, indicated a major zooplanktonic input. The distribution of fatty acids and the relative abundance of chlorophyll-derived compounds and long-chain alkanediols agreed with the predominant contribution of phytoplankton or zooplankton to seston and sediment in the two treatments. The relative abundance of bacterial biomarkers suggested that the contribution of bacteria was rather low. The high relative abundance of polyunsaturated fatty acids (PUFAs) and the absence of stanols in sediments suggested low microbial reworking of organic matter in the recently accumulated sediments. The trophic cascade, generated by the addition of fish, increased the relative abundance of PUFAs in deposited organic matter, thus enhancing sediment quality and potential degradability.

Madinithidium gen. nov. (Bacillariophyceae), a new monoraphid diatom genus from the tropical marine coastal zone

Abstract: We studied a group of monoraphid diatom species (Bacillariophyceae, Achnanthidiaceae) found in the marine coastal environment of tropical islands of the Caribbean Sea, western Atlantic Ocean, Indian Ocean, South Pacific Ocean, the Adriatic Sea and the Black Sea. Based on light and electron microscope examination, Madinithidium was formally described as a genus new to science with the generitype defined as Madinithidium undulatum. The characteristic features of the valve structure are a strongly developed sternum and raphe sternum, transapical striae formed by a single areola (macroareolae) positioned in small depressions, elevated virgae and coaxial internal central raphe endings. Furthermore, the striae of both raphe and sternum valves are closed by finely perforated hymenes. Madinithidium and Achnanthidium species are difficult to identify correctly with light microscopy since their valves are small and finely structured. Four species recently described from the western Indian Ocean and assigned to Achnanthidium sensu lato – Achnanthidium capitatum, A. flexuistriatum, A. pseudodelicatissimum and A. scalariforme – possessed morphological features permitting their placement into Madinithidium gen. nov. The above-mentioned species were formally transferred to the new genus.