Cytotoxicity of diatom-derived oxylipins in organisms belonging to different phyla.
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The cytotoxicity of several saturated and unsaturated marine diatom-derived aldehydes and an oxo-acid have been screened in vitro and in vivo against different organisms, such as bacteria, algae, fungi, echinoderms, molluscs and crustaceans. Conjugated unsaturated aldehydes like 2E,4E-decadienal, 2E,4E-octadienal, 5E,7E-9-oxo-nonadienoic acid and 2E-decenal were active against bacteria and fungi and showed weak algicidal activity. By contrast, the saturated aldehyde decanal and the non-conjugated aldehyde 4Z-decenal had either low or no significant biological activity. In assays with oyster haemocytes, 2E,4E-decadienal exhibited a dose-dependent inhibition of cytoskeleton organisation, rate of phagocytosis and oxidative burst and a dose-dependent promotion of apoptosis. A maternal diatom diet that was rich in unsaturated aldehydes induced arrest of cell division and apoptotic cell degradation in copepod embryos and larvae, respectively. This wide spectrum of physiological pathologies reflects the potent cell toxicity of diatom-derived oxylipins, in relation to their non-specific chemical reactivity towards nucleophilic biomolecules. The cytotoxic activity is conserved across six phyla, from bacteria to crustaceans. Deregulation of cell homeostasis is supposed to induce the elimination of damaged cells through apoptosis. However, efficient protection mechanisms possibly exist in unicellular organisms. Experiments with a genetically modified yeast species exhibiting elevated membrane and/or cell wall permeability suggest that this protection can be related to the inability of the oxylipin compounds to enter the cell. (+info)
The role of the C4 pathway in carbon accumulation and fixation in a marine diatom.
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The role of a C(4) pathway in photosynthetic carbon fixation by marine diatoms is presently debated. Previous labeling studies have shown the transfer of photosynthetically fixed carbon through a C(4) pathway and recent genomic data provide evidence for the existence of key enzymes involved in C(4) metabolism. Nonetheless, the importance of the C(4) pathway in photosynthesis has been questioned and this pathway is seen as redundant to the known CO(2) concentrating mechanism of diatoms. Here we show that the inhibition of phosphoenolpyruvate carboxylase (PEPCase) by 3,3-dichloro-2-dihydroxyphosphinoylmethyl-2-propenoate resulted in a more than 90% decrease in whole cell photosynthesis in Thalassiosira weissflogii cells acclimated to low CO(2) (10 microm), but had little effect on photosynthesis in the C(3) marine Chlorophyte, Chlamydomonas sp. In 3,3-dichloro-2-dihydroxyphosphinoylmethyl-2-propenoate-treated T. weissflogii cells, elevated CO(2) (150 microm) or low O(2) (80-180 microm) restored photosynthesis to the control rate linking PEPCase inhibition with CO(2) supply in this diatom. In C(4) organic carbon-inorganic carbon competition experiments, the (12)C-labeled C(4) products of PEPCase, oxaloacetic acid and its reduced form malic acid suppressed the fixation of (14)C-labeled inorganic carbon by 40% to 50%, but had no effect on O(2) evolution in photosynthesizing diatoms. Oxaloacetic acid-dependent O(2) evolution in T. weissflogii was twice as high in cells acclimated to 10 microm rather than 22 microm CO(2), indicating that the use of C(4) compounds for photosynthesis is regulated over the range of CO(2) concentrations observed in marine surface waters. Short-term (14)C uptake (silicone oil centrifugation) and CO(2) release (membrane inlet mass spectrometry) experiments that employed a protein denaturing cell extraction solution containing the PEPCKase inhibitor mercaptopicolinic acid revealed that much of the carbon taken up by diatoms during photosynthesis is stored as organic carbon before being fixed in the Calvin cycle, as expected if the C(4) pathway functions as a CO(2) concentrating mechanism. Together these results demonstrate that the C(4) pathway is important in carbon accumulation and photosynthetic carbon fixation in diatoms at low (atmospheric) CO(2). (+info)
Sensitive determination of domoic acid using high-performance liquid chromatography with electrogenerated tris(2,2'-bipyridine)ruthenium(III) chemiluminescence detection.
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A new sensitive determination method of domoic acid based on the chemiluminescent reaction of tris(2,2'-bipyridine)ruthenium(III) has been developed. The method exhibited good reproducibility. The relative standard deviation of six replicate measurements was 1.6% for 10 ng ml(-1). A calibration graph, based on a standard domoic acid solution, was linear over the range of 1 - 500 ng ml(-1) (coefficient of correlation, r2 = 0.9995) and the detection limit was 8 pg (signal-to-noise ratio = 3) without any preconcentration and derivatization steps. This new method was successfully applied to a real sample of blue mussels spiked with 2 microg g(-1) domoic acid. (+info)
Silica morphogenesis by alternative processing of silaffins in the diatom Thalassiosira pseudonana.
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For almost 200 years scientists have been fascinated by the ornate cell walls of the diatoms. These structures are made of amorphous silica, exhibiting species-specific, mostly porous patterns in the nano- to micrometer range. Recently, from the diatom Cylindrotheca fusiformis unusual phosphoproteins (termed silaffins) and long chain polyamines have been identified and implicated in biosilica formation. However, analysis of the role of silaffins in morphogenesis of species-specific silica structures has so far been hampered by the difficulty of obtaining structural data from these extremely complex proteins. In the present study, the five major silaffins from the diatom Thalassiosira pseudonana (tpSil1H, -1L, -2H, -2L, and -3) have been isolated, functionally analyzed, and structurally characterized, mak- ing use of the recently available genome data from this organism. Surprisingly, the silaffins of T. pseudonana and C. fusiformis share no sequence homology but are similar regarding amino acid composition and post-translational modifications. Silaffins tpSil1H and -2H are higher molecular mass isoforms of tpSil1L and -2L, respectively, generated in vivo by alternative processing of the same precursor polypeptides. Interestingly, only tpSil1H and -2H but not tpSil1L and -2L induce the formation of porous silica patterns in vitro, suggesting that the alternative processing event is an important step in morphogenesis of T. pseudonana biosilica. (+info)
A live bioprobe for studying diatom-surface interactions.
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Atomic force microscopy has been employed to compare the adhesion of Navicula species I diatoms to surfaces of a hydrophobic elastomer, Intersleek, and a hydrophilic mineral, mica. This was accomplished using tipless atomic force microscopy cantilevers functionalized with live diatom cells. Both surfaces were tested with the same diatom bioprobe. Force versus distance curves generated during these experiments revealed comparable cell adhesion strengths on Intersleek and mica, indicating that Navicula diatoms secrete extracellular polymeric substances with hydrophobic and hydrophilic properties. A statistical analysis of force curves was carried out and the average values of works of detachment of a diatom from Intersleek and mica surfaces were determined. (+info)
The genome of the diatom Thalassiosira pseudonana: ecology, evolution, and metabolism.
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Diatoms are unicellular algae with plastids acquired by secondary endosymbiosis. They are responsible for approximately 20% of global carbon fixation. We report the 34 million-base pair draft nuclear genome of the marine diatom Thalassiosira pseudonana and its 129 thousand-base pair plastid and 44 thousand-base pair mitochondrial genomes. Sequence and optical restriction mapping revealed 24 diploid nuclear chromosomes. We identified novel genes for silicic acid transport and formation of silica-based cell walls, high-affinity iron uptake, biosynthetic enzymes for several types of polyunsaturated fatty acids, use of a range of nitrogenous compounds, and a complete urea cycle, all attributes that allow diatoms to prosper in aquatic environments. (+info)
The role of complex lipids in the synthesis of bioactive aldehydes of the marine diatom Skeletonema costatum.
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Diatoms are unicellular plants broadly present in freshwater and marine ecosystems, where they play a primary role in sustaining the marine food chain. In the last 10 years, there has been accumulating evidence that diatoms may have deleterious effects on the hatching success of zooplankton crustaceans such as copepods, thus affecting dynamics of planktonic populations and limiting secondary production. At the molecular level, failure to hatch is ascribed to the presence of a family of inhibitory oxylipins, which we propose to collectively name polyunsaturated short-chain aldehydes (abbreviated here as PUSCAs). Here we describe the origin of PUSCAs produced by the marine diatom Skeletonema costatum via a lipoxygenase-mediated pathways involving non-esterified polyunsaturated fatty acids (PUFA). Experiments with complex lipids proved the pivotal role of chloroplast-derived glycolipids, especially monogalactosyldiacylglycerol (MGDG), in providing hexadecatrienoic acid (C16:3 omega-4), hexadecatetraenoic acid (C16:4 omega-1) and eicosapentaenoic acid (C20:5 omega-3) to the downstream process leading to 2E,4Z-octadienal (C8:2 omega-4), 2E,4Z,7-octatrienal (C8:3 omega-1) and 2E,4Z-heptadienal (C7:2 omega-3), respectively. Under physiological conditions, the hydrolytic process is associated to galactolipid hydrolyzing enzyme capable of removing fatty acids from both sn positions of glycerol. (+info)
Changes in bacterioplankton composition under different phytoplankton regimens.
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The results of empirical studies have revealed links between phytoplankton and bacterioplankton, such as the frequent correlation between chlorophyll a and bulk bacterial abundance and production. Nevertheless, little is known about possible links at the level of specific taxonomic groups. To investigate this issue, seawater microcosm experiments were performed in the northwestern Mediterranean Sea. Turbulence was used as a noninvasive means to induce phytoplankton blooms dominated by different algae. Microcosms exposed to turbulence became dominated by diatoms, while small phytoflagellates gained importance under still conditions. Denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene fragments showed that changes in phytoplankton community composition were followed by shifts in bacterioplankton community composition, both as changes in the presence or absence of distinct bacterial phylotypes and as differences in the relative abundance of ubiquitous phylotypes. Sequencing of DGGE bands showed that four Roseobacter phylotypes were present in all microcosms. The microcosms with a higher proportion of phytoflagellates were characterized by four phylotypes of the Bacteroidetes phylum: two affiliated with the family Cryomorphaceae and two with the family Flavobacteriaceae. Two other Flavobacteriaceae phylotypes were characteristic of the diatom-dominated microcosms, together with one Alphaproteobacteria phylotype (Roseobacter) and one Gammaproteobacteria phylotype (Methylophaga). Phylogenetic analyses of published Bacteroidetes 16S rRNA gene sequences confirmed that members of the Flavobacteriaceae are remarkably responsive to phytoplankton blooms, indicating these bacteria could be particularly important in the processing of organic matter during such events. Our data suggest that quantitative and qualitative differences in phytoplankton species composition may lead to pronounced differences in bacterioplankton species composition. (+info)