Modification of Living Diatom, Thalassiosira weissflogii, with a Calcium Precursor through a Calcium Uptake Mechanism: A Next Generation Biomaterial for Advanced Delivery Systems.

The diatom's frustule, characterized by its rugged and porous exterior, exhibits a remarkable biomimetic morphology attributable to its highly ordered pores, extensive surface area, and unique architecture. Despite these advantages, the toxicity and nonbiodegradable nature of silica-based organisms pose a significant challenge when attempting to utilize these organisms as nanotopographically functionalized microparticles in the realm of biomedicine. In this study, we addressed this limitation by modulating the chemical composition of diatom microparticles by modulating the active silica metabolic uptake mechanism while maintaining their intricate three-dimensional architecture through calcium incorporation into living diatoms. Here, the diatom Thalassiosira weissflogii was chemically modified to replace its silica composition with a biodegradable calcium template, while simultaneously preserving the unique three-dimensional (3D) frustule structure with hierarchical patterns of pores and nanoscale architectural features, which was evident by the deposition of calcium as calcium carbonate. Calcium hydroxide is incorporated into the exoskeleton through the active mechanism of calcium uptake via a carbon-concentrating mechanism, without altering the microstructure. Our findings suggest that calcium-modified diatoms hold potential as a nature-inspired delivery system for immunotherapy through antibody-specific binding.

Reactive oxygen species mediated extracellular polymeric substances production assisting the recovery of Thalassiosira pseudonana from polystyrene micro and nanoplastics exposure.

As emerging pollutants in the aquatic environments, micro- and nano-plastics (MNPs) aroused widespread environmental concerns for their potential threats to the ecological health. Previous research has proved that microalgae growth could recover from the MNPs toxicities, in which the extracellular polymeric substances (EPS) might play the key role. In order to comprehensively investigate the recovery process of microalgae from MNPs stress and the effecting mechanisms of EPS therein, this study conducted a series of experiments by employing two sizes (0.1 and 1 µm) of polystyrene (PS) MNPs and the marine model diatom Thalassiosira pseudonana during 14 days. The results indicated: the pigments accumulations and photosynthetic recovery of T. pseudonana under MPs exposure showed in the early stage (4-5 days), while the elevation of reactive oxygen species (ROS) and EPS contents lasted longer time period (7-8 days). EPS was aggregated with MNPs particles and microalgal cells, corresponding to the increased settlement rates. More increase of soluble (SL)-EPS contents was found than bound (B)-EPS under MNPs exposure, in which the increase of the protein proportion and humic acid-like substances in SL-EPS was found, thus facilitating aggregates formation. ROS was the signaling molecule mediating the overproduction of EPS. The transcriptional results further proved the enhanced EPS biosynthesis on the molecular level. Therefore, this study elucidated the recovery pattern of microalgae from MNPs stress and linked "ROS-EPS production changes-aggregation formation" together during the growth recovery process, with important scientific and environmental significance.

Rich diversity and active spatial-temporal dynamics of Thalassiosira species revealed by time-series metabarcoding analysis.

Thalassiosira is a species-rich genus in Bacillariophyta that not only contributes positively as primary producer, but also poses negative impacts on ecosystems by causing harmful algal blooms. Although taxonomical studies have identified a large number of Thalassiosira species, however, the composition of Thalassiosira species and their geographical distribution in marine ecosystems were not well understood due primarily to the lack of resolution of morphology-based approaches used previously in ecological expeditions. In this study, we systematically analyzed the composition and spatial-temporal dynamic distributions of Thalassiosira in the model marine ecosystem Jiaozhou Bay by applying metabarcoding analysis. Through analyzing samples collected monthly from 12 sampling sites, 14 Thalassiosira species were identified, including five species that were not previously reported in Jiaozhou Bay, demonstrating the resolution and effectiveness of metabarcoding analysis in ecological research. Many Thalassiosira species showed prominent temporal preferences in Jiaozhou Bay, with some displaying spring-winter preference represented by Thalassiosira tenera, while others displaying summer-autumn preference represented by Thalassiosira lundiana and Thalassiosira minuscula, indicating that the temperature is an important driving factor in the temporal dynamics. The application of metabarcoding analysis, equipped with appropriate molecular markers with high resolution and high specificity and databases of reference molecular marker sequences for potential all Thalassiosira species, will revolutionize ecological research of Thalassiosira species in Jiaozhou Bay and other marine ecosystems.

Phosphorus forms and zinc concentrations affect the physiological ecology and sinking rate of Thalassiosira weissflogii.

The combined effects of phosphorus (P) forms and zinc (Zn) concentrations on diatom silicification remain unclear. In this study, we investigate the effects of different Zn concentrations on the growth, cellular silicon content and sinking rate of Thalassiosira weissflogii under different P forms. The results showed that under the dissolved inorganic phosphorus (DIP) treatments, the specific growth rate of T. weissflogii in Zn limitation culture was significantly lower than that in Zn-replete culture. However, T. weissflogii cellular silicon content and sinking rate increased. Moreover, the reduced specific growth rate (7 %, p < 0.05), enhanced ALP activity (63 %, p < 0.05), and sinking rate (20 %, p < 0.05) for Zn-deplete T. weissflogii implied that the bioavailability of dissolved organic phosphorus (DOP) was depressed under Zn deplete medium. This study demonstrates that the physiological ecology and sinking rate of the diatom T. weissflogii were affected by both individual and combined changes in P forms and Zn concentrations.

Cytochrome c oxidase subunit I gene in Thalassiosira nordenskioeldii strains inhabiting in cold and warm sea waters.

From the biota beneath the sea ice in Lake Saroma, which is adjacent to Sea of Okhotsk, a diatom culture of Saroma 16 was isolated. Strutted processes and a labiate process in Saroma 16 were characteristic of those in Thalassiosira nordenskioeldii. Similarity search analysis showed that the 826-bp rbcL-3P region sequence of this strain was 100% identical to multiple sequences registered as T. nordenskioeldii in a public database. The 4305-bp PCR-amplified mitochondrial cytochrome c oxidase subunit I (COI) gene (COI)-5P region of Saroma 16 included a 1060-bp open reading frame (ORF), which was interrupted by 934-bp and 2311-bp introns that included frame-shifted ORFs encoding reverse-transcriptase (RTase)-like proteins. Previous reports showed that a strain of the same species, CNS00052, originating from the East China Sea included no introns in the COI, whereas North Atlantic Ocean strains of the same species, such as CCMP992, CCMP993, and CCMP997, included a 2.3-kb intron in the same position as Saroma 16.

Marine diatom algae cultivation in simulated dairy wastewater and biomass valorization.

Liquid byproducts and organic wastes generated from dairy processing units contribute as the largest source of industrial food wastewater. Though bacteria-mediated treatment strategies are largely implemented, a more effective and innovative management system is needed of the hour. Thus, the current study involves the cultivation of centric diatoms, Chaetoceros gracilis, and Thalassiosira weissflogii in simulated dairy wastewater (SDWW) formulated using varying amounts of milk powder with artificial seawater f/2 media (ASW). The results revealed that cell density and biomass productivity were highest in the 2.5% SDWW treatment cultures of both the strains, the maximum being in C. gracilis (7.5 × 106 cells mL - 1; 21.1 mg L-1 day-1). Conversely, the total carotenoid, chrysolaminarin, and phenol content were negatively impacted by SDWW. However, a considerable enhancement in the total lipid content was reported in the 2.5% SDWW culture of both species. Furthermore, the fatty acid profiling revealed that though the total polyunsaturated fatty acid (PUFA) content was highest in the control setups, the total mono polyunsaturated fatty acid (MUFA) content was higher in the 5% SDWW setups (30.66% in C. gracilis and 33.21% in T. weissflogii). In addition to it, in the cultures utilizing energy from external carbon sources provided by SDWW, the biodiesel produced was also enhanced owing to the heightened cetane number. Thus, the current study evidently highlights the organic carbon acquisition potential of marine diatoms with the scope of providing sustainable biorefinery.

Warming modulates the photosynthetic performance of Thalassiosira pseudonana in response to UV radiation.

Diatoms form a major component of phytoplankton. These eukaryotic organisms are responsible for approximately 40% of primary productivity in the oceans and contribute significantly to the food web. Here, the influences of ultraviolet radiation (UVR) and ocean warming on diatom photosynthesis were investigated in Thalassiosira pseudonana. The organism was grown at two temperatures, namely, 18°C, the present surface water temperature in summer, and 24°C, an estimate of surface temperature in the year 2,100, under conditions of high photosynthetically active radiation (P, 400-700 nm) alone or in combination with UVR (P + UVR, 295-700 nm). It was found that the maximum photochemical yield of PSII (Fv/Fm) in T. pseudonana was significantly decreased by the radiation exposure with UVR at low temperature, while the rise of temperature alleviated the inhibition induced by UVR. The analysis of PSII subunits turnover showed that high temperature alone or worked synergistically with UVR provoking fast removal of PsbA protein (KPsbA), and also could maintain high PsbD pool in T. pseudonana cells. With the facilitation of PSII repair process, less non-photochemical quenching (NPQ) occurred at high temperature when cells were exposed to P or P + UVR. In addition, irrespective of radiation treatments, high temperature stimulated the induction of SOD activity, which partly contributed to the higher PSII repair rate constant (Krec) as compared to KPsbA. Our findings suggest that the rise in temperature could benefit the photosynthetic performance of T. pseudonana via modulation of its PSII repair cycle and protective capacity, affecting its abundance in phytoplankton in the future warming ocean.

Induction of photosynthesis under anoxic condition in Thalassiosira pseudonana and Euglena gracilis: interactions between fermentation and photosynthesis.

Introduction: In their natural environment, microalgae can be transiently exposed to hypoxic or anoxic environments. Whereas fermentative pathways and their interactions with photosynthesis are relatively well characterized in the green alga model Chlamydomonas reinhardtii, little information is available in other groups of photosynthetic micro-eukaryotes. In C. reinhardtii cyclic electron flow (CEF) around photosystem (PS) I, and light-dependent oxygen-sensitive hydrogenase activity both contribute to restoring photosynthetic linear electron flow (LEF) in anoxic conditions. Methods: Here we analyzed photosynthetic electron transfer after incubation in dark anoxic conditions (up to 24 h) in two secondary microalgae: the marine diatom Thalassiosira pseudonana and the excavate Euglena gracilis. Results: Both species showed sustained abilities to prevent over-reduction of photosynthetic electron carriers and to restore LEF. A high and transient CEF around PSI was also observed specifically in anoxic conditions at light onset in both species. In contrast, at variance with C. reinhardtii, no sustained hydrogenase activity was detected in anoxic conditions in both species. Discussion: Altogether our results suggest that another fermentative pathway might contribute, along with CEF around PSI, to restore photosynthetic activity in anoxic conditions in E. gracilis and T. pseudonana. We discuss the possible implication of the dissimilatory nitrate reduction to ammonium (DNRA) in T. pseudonana and the wax ester fermentation in E. gracilis.

Common environmental stress responses in a model marine diatom.

Marine planktonic diatoms are among the most important contributors to phytoplankton blooms and marine net primary production. Their ecological success has been attributed to their ability to rapidly respond to changing environmental conditions. Here, we report common molecular mechanisms used by the model marine diatom Thalassiosira pseudonana to respond to 10 diverse environmental stressors using RNA-Seq analysis. We identify a specific subset of 1076 genes that are differentially expressed in response to stressors that induce an imbalance between energy or resource supply and metabolic capacity, which we termed the diatom environmental stress response (d-ESR). The d-ESR is primarily composed of genes that maintain proteome homeostasis and primary metabolism. Photosynthesis is strongly regulated in response to environmental stressors but chloroplast-encoded genes were predominantly upregulated while the nuclear-encoded genes were mostly downregulated in response to low light and high temperature. In aggregate, these results provide insight into the molecular mechanisms used by diatoms to respond to a range of environmental perturbations and the unique role of the chloroplast in managing environmental stress in diatoms. This study facilitates our understanding of the molecular mechanisms underpinning the ecological success of diatoms in the ocean.

Ribosome Profiling in the Model Diatom Thalassiosira pseudonana.

Diatoms are an important group of eukaryotic microalgae, which play key roles in marine biochemical cycling and possess significant biotechnological potential. Despite the importance of diatoms, their regulatory mechanisms of protein synthesis at the translational level remain largely unexplored. Here, we describe the detailed development of a ribosome profiling protocol to study translation in the model diatom Thalassiosira pseudonana, which can easily be adopted for other diatom species. To isolate and sequence ribosome-protected mRNA, total RNA was digested, and the ribosome-protected fragments were obtained by a combination of sucrose-cushion ultracentrifugation and polyacrylamide gel electrophoresis for size selection. To minimize rRNA contamination, a subtractive hybridization step using biotinylated oligos was employed. Subsequently, fragments were converted into sequencing libraries, enabling the global quantification and analysis of changes in protein synthesis in diatoms. The development of this novel ribosome profiling protocol represents a major expansion of the molecular toolbox available for diatoms and therefore has the potential to advance our understanding of the translational regulation in this important group of phytoplankton. © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Ribosome profiling in Thalassiosira pseudonana Alternate Protocol: Ribosome profiling protocol for diatoms using sucrose gradient fractionation.

Decreasing O2 availability reduces cellular protein contents in a marine diatom.

Anthropogenic activities and climate change are exacerbating marine deoxygenation. Apart from aerobic organisms, reduced O2 also affects photoautotrophic organisms in the ocean. This is because without available O2, these O2 producers cannot maintain their mitochondrial respiration, especially under dim-light or dark conditions, which may disrupt the metabolism of macromolecules including proteins. We used growth rate, particle organic nitrogen and protein analyses, proteomics, and transcriptomics to determine cellular nitrogen metabolism of the diatom Thalassiosira pseudonana grown under three O2 levels in a range of light intensities at nutrient-rich status. The ratio of protein nitrogen to total nitrogen under ambient O2 level among different light intensities was about 0.54-0.83. At the lowest light intensity, decreased O2 had a stimulatory effect on protein content. When light intensity increased to moderate and high or inhibitory levels, decreased O2 reduced the protein content, with maximum values of 56 % at low O2 and 60 % at hypoxia, respectively. In addition, cells growing under low O2 or hypoxic status exhibited a decreased rate of nitrogen assimilation associated with decreased protein content, which was associated with downregulated expression of genes related to nitrate transform and protein synthesis and upregulated expression of genes related to protein degradation. Our results suggest that decreased O2 reduces the protein content of phytoplankton cells, which might degrade grazer nutrition and thus affect marine food chains under the scenario of increasingly deoxygenated/hypoxic waters in future.

Distribution, toxicity and bioaccumulation of trace metals in environmental matrices of an estuary in a protected area.

Mangroves are productive ecosystems that are highly threatened by anthropogenic activities. We investigated the environmental quality of the Serinhaém river estuary located in a legally protected area. Through chemical analysis of sediments and tissues of Cardisoma guanhumi, in addition to bioassays with elutriate involving Nitokra sp. and Thalassiosira pseudonana, we determined the contamination status and risk factors related to trace metals in the estuary. For the sediment, the concentrations of Cr and Ni were above the limit established by CONAMA n° 454/2012 in the "City" site, and Cr above the TEL in all sampling sites. Ecotoxicological tests showed high toxicity in samples from "City" and "Tributary". The elements Cr, Mn, Ni and Zn were also higher in crabs from these sites. Cr levels exceeded the Brazilian limit for food consumption. The bioaccumulation factor was not significant. However, the overall analysis proved that this estuary is increasingly impacted by anthropogenic pressure.

Phytoplankton composition from Araçá Bay and São Sebastião Channel, São Paulo, Brazil

: Despite its small area, Araçá Bay (AB) holds cultural, historical, and economic value and displays great benthic biodiversity. Thus, it is crucial to monitor its environmental health, including knowing the main groups of phytoplankton and their temporal variability. The shallow waters of Araçá Bay are continuously modified by the complex hydrography of the adjacent São Sebastião channel (SSC), challenging standard experimental designs for phytoplankton collection. Here we report changes in phytoplankton composition at intervals of five to six weeks from September 2013 to August 2014 in both Araçá Bay and SSC. Samples were collected twice daily for three consecutive days to increase taxonomic resolution. Our goal was to provide an inventory of species occurrences to aid future public policies and environmental management of the area. Analyses revealed high species richness and 166 different phytoplankton taxa. Diatoms and dinoflagellates were always numerically dominant, but taxa occurrence changed markedly. Diatoms of the genera Pseudo-nitzschia were abundant during spring and summer concurrently to signatures of South Atlantic Central Water in the SSC, while Thalassiosira occurred when waters displayed relatively lower salinity. The inventory demonstrated several potentially harmful species of microalgae and cyanobacteria, strongly suggesting investments in monitoring programs in this area that currently experience an increase in population.

Improving the genome and proteome annotations of the marine model diatom Thalassiosira pseudonana using a proteogenomics strategy.

Diatoms are unicellular eukaryotic phytoplankton that account for approximately 20% of global carbon fixation and 40% of marine primary productivity; thus, they are essential for global carbon biogeochemical cycling and climate. The availability of ten diatom genome sequences has facilitated evolutionary, biological and ecological research over the past decade; however, a complimentary map of the diatom proteome with direct measurements of proteins and peptides is still lacking. Here, we present a proteome map of the model marine diatom Thalassiosira pseudonana using high-resolution mass spectrometry combined with a proteogenomic strategy. In-depth proteomic profiling of three different growth phases and three nutrient-deficient samples identified 9526 proteins, accounting for ~ 81% of the predicted protein-coding genes. Proteogenomic analysis identified 1235 novel genes, 975 revised genes, 104 splice variants and 234 single amino acid variants. Furthermore, our quantitative proteomic analysis experimentally demonstrated that a considerable number of novel genes were differentially translated under different nutrient conditions. These findings substantially improve the genome annotation of T. pseudonana and provide insights into new biological functions of diatoms. This relatively comprehensive diatom proteome catalog will complement available diatom genome and transcriptome data to advance biological and ecological research of marine diatoms. Supplementary Information: The online version contains supplementary material available at 10.1007/s42995-022-00161-y.

Localization and characterization θ carbonic anhydrases in Thalassiosira pseudonana.

Carbonic anhydrase (CA) is a crucial component for the operation of CO2-concentrating mechanisms (CCMs) in the majority of aquatic photoautotrophs that maintain the global primary production. In the genome of the centric marine diatom, Thalassiosira pseudonana, there are four putative gene sequences that encode θ-type CA, which was a type of CA recently identified in marine diatoms and green algae. In the present study, specific subcellular locations of four θCAs, TpθCA1, TpθCA2, TpθCA3, and TpθCA4 were determined by expressing GFP-fused proteins of these TpθCAs in T. pseudonana. As a result, C-terminal GFP fusion proteins of TpθCA1, TpθCA2, and TpθCA3 were all localized in the chloroplast; TpθCA2 was at the central chloroplast area, and the other two TpθCAs were throughout the chloroplast. Immunogold-labeling transmission electron microscopy was further performed for the transformants expressing TpθCA1:GFP and TpθCA2:GFP with anti-GFP-monoclonal antibody. TpθCA1:GFP was localized in the free stroma area, including the peripheral pyrenoid area. TpθCA2:GFP was clearly located as a lined distribution at the central part of the pyrenoid structure, which was most likely the pyrenoid-penetrating thylakoid. Considering the presence of the sequence encoding the N-terminal thylakoid-targeting domain in the TpθCA2 gene, this localization was likely the lumen of the pyrenoid-penetrating thylakoid. On the other hand, TpθCA4:GFP was localized in the cytoplasm. Transcript analysis of these TpθCAs revealed that TpθCA2 and TpθCA3 were upregulated in atmospheric CO2 (0.04% CO2, LC) levels, while TpθCA1 and TpθCA4 were highly induced under 1% CO2 (HC) condition. The genome-editing knockout (KO) of TpθCA1, by CRISPR/Cas9 nickase, gave a silent phenotype in T. pseudonana under LC-HC conditions, which was in sharp agreement with the case of the previously reported TpθCA3 KO. In sharp contrast, TpθCA2 KO is so far unsuccessful, suggesting a housekeeping role of TpθCA2. The silent phenotype of KO strains of stromal CAs suggests that TpαCA1, TpθCA1, and TpθCA3 may have functional redundancy, but different transcript regulations in response to CO2 of these stromal CAs suggest in part their independent roles.

Phytoplankton competition and resilience under fluctuating temperature.

Environmental variability is an inherent feature of natural systems which complicates predictions of species interactions. Primarily, the complexity in predicting the response of organisms to environmental fluctuations is in part because species' responses to abiotic factors are non-linear, even in stable conditions. Temperature exerts a major control over phytoplankton growth and physiology, yet the influence of thermal fluctuations on growth and competition dynamics is largely unknown. To investigate the limits of coexistence in variable environments, stable mixed cultures with constant species abundance ratios of the marine diatoms, Phaeodactylum tricornutum and Thalassiosira pseudonana, were exposed to different temperature fluctuation regimes (n = 17) under high and low nitrogen (N) conditions. Here we demonstrate that phytoplankton exhibit substantial resilience to temperature variability. The time required to observe a shift in the species abundance ratio decreased with increasing fluctuations, but coexistence of the two model species under high N conditions was disrupted only when amplitudes of temperature fluctuation were high (±8.2°C). N limitation caused the thermal amplitude for disruption of species coexistence to become lower (±5.9°C). Furthermore, once stable conditions were reinstated, the two species differed in their ability to recover from temperature fluctuations. Our findings suggest that despite the expectation of unequal effect of fluctuations on different competitors, cycles in environmental conditions may reduce the rate of species replacement when amplitudes remain below a certain threshold. Beyond these thresholds, competitive exclusion could, however, be accelerated, suggesting that aquatic heatwaves and N availability status are likely to lead to abrupt and unpredictable restructuring of phytoplankton community composition.

Efficient gene replacement by CRISPR/Cas-mediated homologous recombination in the model diatom Thalassiosira pseudonana.

CRISPR/Cas enables targeted genome editing in many different plant and algal species including the model diatom Thalassiosira pseudonana. However, efficient gene targeting by homologous recombination (HR) to date is only reported for photosynthetic organisms in their haploid life-cycle phase. Here, a CRISPR/Cas construct, assembled using Golden Gate cloning, enabled highly efficient HR in a diploid photosynthetic organism. Homologous recombination was induced in T. pseudonana using sequence-specific CRISPR/Cas, paired with a dsDNA donor matrix, generating substitution of the silacidin, nitrate reductase and urease genes by a resistance cassette (FCP:NAT). Up to c. 85% of NAT-resistant T. pseudonana colonies screened positive for HR by nested PCR. Precise integration of FCP:NAT at each locus was confirmed using an inverse PCR approach. The knockout of the nitrate reductase and urease genes impacted growth on nitrate and urea, respectively, while the knockout of the silacidin gene in T. pseudonana caused a significant increase in cell size, confirming the role of this gene for cell-size regulation in centric diatoms. Highly efficient gene targeting by HR makes T. pseudonana as genetically tractable as Nannochloropsis and Physcomitrella, hence rapidly advancing functional diatom biology, bionanotechnology and biotechnological applications targeted on harnessing the metabolic potential of diatoms.

Interlinking diatom frustule diversity from the abyss of the central Arabian Sea to surface processes: physical forcing and oxygen minimum zone.

This study analyzed the diversity and abundance of diatom frustules including the ancillary parameters using the core top sediments from five locations (21, 19, 15, 13, and 11°N) along the central Arabian Sea (64°E), an area profoundly influenced by atmospheric forcing (monsoons) and oxygen minimum zone (OMZ) with high spatial variability. Significantly higher organic carbon (0.97 ± 0.05%) and diatom frustules (5.92 ± 0.57 × 104 valves g-1) were noticed in the north (21, 19, 15°N) where natural nutrient enrichment via open-ocean upwelling, winter convection, and lateral advection support large diatom-dominated phytoplankton blooms and intense OMZ. Conversely, the south (13, 11°N) depicted significantly lower organic carbon (0.74 ± 0.08%) as well as frustules (4.02 ± 0.87 × 104 valves g-1) as this area mostly remains nutrient-poor dominated by small-medium-sized phytoplankton. The north was dominated by large-sized diatoms like Coscinodiscus that could escape grazing and sink consequently due to higher ballasting. Furthermore, the presence of the intense OMZ in the north might reduce grazing pressure (low zooplankton stock) and mineralization speed facilitating higher phytodetritus transport. Relatively smaller chain-forming centric (Thalassiosira) and pennate diatoms (Pseudo-nitzschia, Fragilaria, Nitzschia, etc.) were found throughout the transect with higher abundance in the south. The euphotic diatom diversity from the existing literature was compared with the frustule diversity from the sediments suggesting not all diatoms make their way to the abyss. Such distinct spatial north-south variability in diatom frustule size as well as abundance could be attributed to cell size, grazing, and water column mineralization rates related to OMZ.

Cytotoxic Potential of the Marine Diatom Thalassiosira rotula: Insights into Bioactivity of 24-Methylene Cholesterol.

Marine microalgae are receiving great interest as sustainable sources of bioactive metabolites for health, nutrition and personal care. In the present study, a bioassay-guided screening allowed identifying an enriched fraction from SPE separation of the methanolic extract of the marine diatom Thalassiosira rotula with a chemically heterogeneous composition of cytotoxic molecules, including PUFAs, the terpene phytol, the carotenoid fucoxanthin and the phytosterol 24-methylene cholesterol (24-MChol). In particular, this latter was the object of deep investigation aimed to gain insight into the mechanisms of action activated in two tumour cell models recognised as resistant to chemical treatments, the breast MCF7 and the lung A549 cell lines. The results of our studies revealed that 24-MChol, in line with the most studied ß-sitosterol (ß-SIT), showed cytotoxic activity in a 3-30 µM range of concentration involving the induction of apoptosis and cell cycle arrest, although differences emerged between the two sterols and the two cancer systems when specific targets were investigated (caspase-3, caspase-9, FAS and TRAIL).

Elevated CO2 modulates the physiological responses of Thalassiosira pseudonana to ultraviolet radiation.

Diatoms account for a large proportion of marine primary productivity, they tend to be the predominant species in the phytoplankton communities in the surface ocean with frequent and large light fluctuations. To understand the impacts of increased CO2 on diatoms' capacity in exploitation of variable solar radiation, we cultured a model diatom Thalassiosira pseudonana with 400 or 1000ppmv CO2 and exposed it to high photosynthetically active radiation (PAR) alone or PAR plus ultraviolet radiation (UVR) to examine its physiological performances. The results showed that the maximum photochemical efficiency (Fv/fm) was significantly reduced by high PAR and PAR + UVR in T. pseudonana, UVR-induced inhibition on PSII activity was exacerbated by high CO2. PSII activity drops coincide approximately with PsbA content in the cells exposed to high PAR or PAR + UVR, which was pronounced at high CO2. The removal of PsbD in T. pseudonana cells declined under high CO2 during UVR exposure, limiting the repair capacity of PSII. In addition, high CO2 reversed the induction of energy-dependent form of NPQ by UVR to the increase of Y(No), indicating the severe damage of the photoprotective reactions. Our findings suggest that the adverse impacts of UVR on PSII function of T. pseudonana were aggravated by the elevated CO2 through modulating its capacity in repair and protection, which thereby would influence its abundance and competitiveness in phytoplankton communities.