Haloperoxidase mediated quorum quenching by Nitzschia cf pellucida: study of the metabolization of N-acyl homoserine lactones by a benthic diatom

Diatoms are known to produce a variety of halogenated compounds, which were recently shown to have a role in allelopathic interactions between competing species. The production of these compounds is linked to haloperoxidase activity. This research, has shown that this system may also be involved in diatom-bacteria interactions via the H2O2 dependent inactivation of a type of quorum sensing (QS) molecule, i.e., N-beta-ketoacylated homoserine lactones (AHLs), by a natural haloperoxidase system from the benthic diatom Nitzschia cf pellucida. The AHL degradation pathway towards corresponding halogenated derivatives was elucidated via HPLC-MS analysis and the synthesis of a broad series of novel halogenated AHL analogues as reference compounds. Furthermore, their biological activity as quorum sensing modulators was directly compared and evaluated against a series of naturally occurring beta-keto-AHLs. It has been demonstrated that the loss of the QS activity results from the final cleavage of the halogenated N-acyl chain of the signal molecules

N-acylated homoserine lactone-derived tetramic acids as algicidal compounds

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The fine-tuning of NPQ in diatoms relies on the regulation of both xanthophyll cycle enzymes

AbstractDiatoms possess an efficient mechanism to dissipate photons as heat in conditions of excess light, which is visualized as the Non-Photochemical Quenching of chlorophyll a fluorescence (NPQ). In most diatom species, NPQ is proportional to the concentration of the xanthophyll cycle pigment diatoxanthin formed from diadinoxanthin by the diadinoxanthin de-epoxidase enzyme. The reverse reaction is performed by the diatoxanthin epoxidase. Despite the xanthophyll cycle’s central role in photoprotection, its regulation is not yet well understood. The proportionality between diatoxanthin and NPQ allowed us to calculate the activity of both xanthophyll cycle enzymes in the model diatom Phaeodactylum tricornutum from NPQ kinetics. From there, we explored the light-dependency of the activity of both enzymes. Our results demonstrate that a tight regulation of both enzymes is key to fine-tune NPQ: (i) the rate constant of diadinoxanthin de-epoxidation is low under a light-limiting regime but increases as photosynthesis saturates, probably due to the thylakoidal proton gradient ΔpH (ii) the rate constant of diatoxanthin epoxidation exhibits an optimum under low light and decreases in the dark due to an insufficiency of the co-factor NADPH as well as in higher light through an as yet unresolved inhibition mechanism, that is unlikely to be related to the ΔpH. We observed that the suppression of NPQ by an uncoupler was due to an accelerated diatoxanthin epoxidation enzyme rather than to the usually hypothesized inhibition of the diadinoxanthin de-epoxidation enzyme.</jats:p

Behavioural versus physiological photoprotection in epipelic and epipsammic benthic diatoms

Benthic diatoms are dominant primary producers in intertidal marine sediments, which are characterized by widely fluctuating and often extreme light conditions. To cope with sudden increases in light intensity, benthic diatoms display both behavioural and physiological photoprotection mechanisms. Behavioural photoprotection is restricted to raphid pennate diatoms, which possess a raphe system that enables motility and hence positioning in sediment light gradients (e.g. via vertical migration into the sediment). The main physiological photoprotection mechanism is to dissipate excess light energy as heat, measured as Non-Photochemical Quenching (NPQ) of chlorophyll fluorescence. A trade-off between vertical migration and physiological photoprotection (NPQ) in benthic diatoms has been hypothesized before, but this has never been formally tested. We exposed five epipelic diatom species (which move in between sediment particles) and four epipsammic diatom species (which live in close association with individual sand grains) to high light conditions, and characterized both NPQ and the relative magnitude of the migratory response to high light. Our results reveal the absence of a significant downward migratory response in an araphid diatom, but also in several raphid epipsammic diatoms, while all epipelic species showed a significant migratory response upon high light exposure. In all epipsammic species the upregulation of NPQ was rapid and pronounced; NPQ relaxation in low light conditions, however, occurred faster in the araphid diatom, compared with the raphid epipsammic species. In contrast, all epipelic species lacked a strong and flexible NPQ response and showed higher susceptibility to photodamage when not able to migrate. While overall our results support the vertical migration-NPQ trade-off, the lack of strong relationships between the capacity for vertical migration and NPQ within the epipsammic and epipelic groups suggests that other factors as well, such as cell size, substrate type and photoacclimation, may influence photoprotective strategies

Assessing the suitability of imaging-PAM fluorometry for monitoring growth of benthic diatoms

High-throughput monitoring of growth using Pulse Amplitude Modulated (PAM) chlorophyll fluorescence imaging (Imaging-PAM) is an important tool for experimental ecological and physiological research on benthic diatom cultures, as due to substrate adhesion non-destructive subsampling for biomass determination is difficult and/or not representative. We evaluated the reliability of Imaging-PAM for quantifying benthic diatom biomass by comparing biomass determinations based on PAM fluorescence parameters to those based on cells counts and chlorophyll a measurements for cultures of the diatoms Cylindrotheca closterium and Seminavis robusta in different growth phases (exponential vs stationary) and cell size ranges. We found that PAM fluorescence imaging is particularly suited to monitor growth during the exponential phase. The dark adapted minimal fluorescence (F-0) parameter proved to be the most reliable fluorescence-based biomass proxy, especially for establishing complete growth curves, as it was the most robust parameter to growth phase changes. A dark adaptation period prior to the PAM measurements improved the quantification of the stationary but not of the exponential cultures. Under the right conditions, dark adaptation can thus be omitted for exponentially growing cultures to enhance throughput. The use of multiwell plates for growth monitoring with the Imaging-PAM is feasible even though there can be a slight biomass underestimation near of the edges of the plate. Based on these findings, we formulate specific guidelines for using Imaging-PAM fluorometry in experiments with benthic diatom cultures. We especially recommend taking caution when working under conditions which potentially influence the condition of the photosynthetic apparatus of the algae

Auxin function in the brown alga Dictyota dichotoma

Auxin controls body plan patterning in land plants and has been proposed to play a similar role in the development of brown algae (Phaeophyta) despite their distant evolutionary relationship with land plants. The mechanism of auxin action in brown algae remains controversial because of contradicting conclusions derived from pharmacological studies on Fucus. In this study, we used Dictyota dichotoma as a model system to show that auxin plays a role during the apical-basal patterning of the embryo of brown algae. Indole-3-acetic acid (IAA) was detectable in D. dichotoma germlings and mature tissue. Although two-celled D. dichotoma zygotes normally develop a rhizoid from one pole and a thallus meristem from the other, addition of exogenous auxins to one-celled embryos affected polarization, and both poles of the spheroidal embryo developed into rhizoids instead. The effect was strongest at lower pH and when variable extrinsic informational cues were applied. 2-[4-(diethylamino)-2-hydroxybenzoyl]benzoic acid (BUM), an inhibitor of ABC-B/multi-drug resistance/P-glycoprotein (ABCB) subfamily of transporters in land plants, affected rhizoid formation by increasing rhizoid branching and inducing ectopic rhizoids. An in silico survey of auxin genes suggested a diverse range of biosynthesis genes and transport genes, such as PIN-LIKES (PILS) and ATP-binding cassete subfamily (ABCB) transporters, from land plants have homologues in D. dichotoma and Ectocarpus siliculosus. Together with reports on auxin function in basal lineages of green algae, these results suggest that auxin function predates the divergence between the green and brown lineage and the transition towards land plants

Nematodes stimulate biomass accumulation in a multispecies diatom biofilm

While the effects of abiotic parameters on microbial tidal biofilms are relatively well-documented, the effects of grazing and/or bioturbation by meiofauna are poorly understood. We investigated the impact of a natural nematode assemblage on the biomass and microbial community structure of a multispecies diatom biofilm. Nematodes stimulated diatom biomass accumulation of the biofilm and caused a shift in diatom community structure. Higher diatom biomass accumulation in the presence of nematodes could be the result of increased diatom biomass production through nutrient regeneration resulting from grazing or bioturbation, and/or through shifts in interspecific interactions between diatoms (e.g. competition) through selective grazing. Alternatively, lower biomass in the controls may be due to higher secretion of diatom production in the form of bound extracellular polymeric substances (EPS). Our observations underscore that meiobenthos, and especially nematodes, are important for the structure and production of tidal biofilms