Unveiling the ecological resilience and industrial potential of Skeletonema marinoi through mixotrophic cultivation in Nordic winter condition.

Mixotrophy, the concurrent use of inorganic and organic carbon in the presence of light for microalgal growth, holds ecological and industrial significance. However, it is poorly explored in diatoms, especially in ecologically relevant species like Skeletonema marinoi. This study strategically employed mixotrophic metabolism to optimize the growth of a strain of Skeletonema marinoi (Sm142), which was found potentially important for biomass production on the west coast of Sweden in winter conditions. The aim of this study was to discern the most effective organic carbon sources by closely monitoring microalgal growth through the assessment of optical density, chlorophyll a fluorescence, and biomass concentration. The impact of various carbon sources on the physiology of Sm142 was investigated using photosynthetic and respiratory parameters. The findings revealed that glycerol exhibited the highest potential for enhancing the biomass concentration of Sm142 in a multi-cultivator under the specified experimental conditions, thanks to the increase in respiration activity. Furthermore, the stimulatory effect of glycerol was confirmed at a larger scale using environmental photobioreactors simulating the winter conditions on the west coast of Sweden; it was found comparable to the stimulation by CO2-enriched air versus normal air. These results were the first evidence of the ability of Skeletonema marinoi to perform mixotrophic metabolism during the winter and could explain the ecological success of this diatom on the Swedish west coast. These findings also highlight the importance of both organic and inorganic carbon sources for enhancing biomass productivity in harsh winter conditions.

Characterization of trehalolipid biosurfactant produced by the novel marine strain Rhodococcus sp. SP1d and its potential for environmental applications.

BACKGROUND: Biosurfactants are surface-active compounds with environmental and industrial applications. These molecules show higher biocompatibility, stability and efficiency compared to synthetic surfactants. On the other hand, biosurfactants are not cost-competitive to their chemical counterparts. Cost effective technology such as the use of low-cost substrates is a promising approach aimed at reducing the production cost. This study aimed to evaluate the biosurfactant production and activity by the novel strain Rhodococcus sp. SP1d by using different growth substrates. Therefore, to exploit the biosurfactant synthesized by SP1d for environmental applications, the effect of this compound on the bacteria biofilm formation was evaluated. Eventually, for a possible bioremediation application, the biosurfactant properties and its chemical characteristics were investigated using diesel as source of carbon. RESULTS: Rhodococcus sp. SP1d evidence the highest similarity to Rhodococcus globerulus DSM 43954T and the ability to biosynthesize surfactants using a wide range of substrates such as exhausted vegetable oil, mineral oil, butter, n-hexadecane, and diesel. The maximum production of crude biosurfactant after 10 days of incubation was reached on n-hexadecane and diesel with a final yield of 2.38 ± 0.51 and 1.86 ± 0.31 g L- 1 respectively. Biosurfactants produced by SP1d enhanced the biofilm production of P. protegens MP12. Moreover, the results showed the ability of SP1d to produce biosurfactants on diesel even when grown at 10 and 18 °C. The biosurfactant activity was maintained over a wide range of NaCl concentration, pH, and temperature. A concentration of 1000 mg L- 1 of the crude biosurfactant showed an emulsification activity of 55% towards both xylene and olive oil and a reduction of 25.0 mN m- 1 of surface tension of water. Eventually, nuclear magnetic resonance spectroscopy indicated that the biosurfactant is formed by trehalolipids. CONCLUSIONS: The use of low-cost substrates such as exhausted oils and waste butter reduce both the costs of biosurfactant synthesis and the environmental pollution due to the inappropriate disposal of these residues. High production yields, stability and emulsification properties using diesel and n-hexadecane as substrates, make the biosurfactant produced by SP1d a sustainable biocompound for bioremediation purpose. Eventually, the purified biosurfactant improved the biofilm formation of the fungal antagonistic strain P. protegens MP12, and thus seem to be exploitable to increase the adherence and colonization of plant surfaces by this antagonistic strain and possibly enhance antifungal activity.

Two-phase microalgae cultivation for RAS water remediation and high-value biomass production.

The overall goal of this study was to provide solutions to innovative microalgae-based technology for wastewater remediation in a cold-water recirculating marine aquaculture system (RAS). This is based on the novel concept of integrated aquaculture systems in which fish nutrient-rich rearing water will be used for microalgae cultivation. The produced biomass can be used as fish feed, while the cleaned water can be reused, to create a highly eco-sustainable circular economy. Here, we tested three microalgae species Nannochloropis granulata (Ng), Phaeodactylum tricornutum (Pt), and Chlorella sp (Csp) for their ability to remove nitrogen and phosphate from the RAS wastewater and simultaneously produce high-value biomass, i.e., containing amino acids (AA), carotenoids, and polyunsaturated fatty acids (PUFAs). A high yield and value of biomass were achieved for all species in a two-phase cultivation strategy: i) a first phase using a medium optimized for best growth (f/2 14x, control); ii) a second "stress" phase using the RAS wastewater to enhance the production of high-value metabolites. Ng and Pt performed best in terms of biomass yield (i.e., 5-6 g of dry weight, DW.L-1) and efficient cleaning of the RAS wastewater from nitrite, nitrate, and phosphate (i.e., 100% removal). Csp produced about 3 g L-1 of DW and reduced efficiently only nitrate, and phosphate (i.e., about 76% and 100% removal, respectively). The biomass of all strains was rich in protein (30-40 % of DW) containing all the essential AA except Methionine. The biomass of all three species was also rich in PUFAs. Finally, all tested species are excellent sources of antioxidant carotenoids, including fucoxanthin (Pt), lutein (Ng and Csp) and ß-carotene (Csp). All tested species in our novel two-phase cultivation strategy thus showed great potential to treat marine RAS wastewater and provide sustainable alternatives to animal and plant proteins with extra added values.

Mixotrophy in a Local Strain of Nannochloropsis granulata for Renewable High-Value Biomass Production on the West Coast of Sweden

A local strain of Nannochloropsis granulata (Ng) has been reported as the most productive microalgal strain in terms of both biomass yield and lipid content when cultivated in photobioreactors that simulate the light and temperature conditions during the summer on the west coast of Sweden. To further increase the biomass and the biotechnological potential of this strain in these conditions, mixotrophic growth (i.e., the simultaneous use of photosynthesis and respiration) with glycerol as an external carbon source was investigated in this study and compared with phototrophic growth that made use of air enriched with 1-2% CO2. The addition of either glycerol or CO2-enriched air stimulated the growth of Ng and theproduction of high-value long-chain polyunsaturated fatty acids (EPA) as well as the carotenoid canthaxanthin. Bioassays in human prostate cell lines indicated the highest antitumoral activity for Ng extracts and fractions from mixotrophic conditions. Metabolomics detected betaine lipids specifically in the bioactive fractions, suggesting their involvement in the observed antitumoral effect. Genes related to autophagy were found to be upregulated by the most bioactive fraction, suggesting a possible therapeutic target against prostate cancer progression. Taken together, our results suggest that the local Ng strain can be cultivated mixotrophically in summer conditions on the west coast of Sweden for the production of high-value biomass containing antiproliferative compounds, carotenoids, and EPA.

Mixotrophy in diatoms: Molecular mechanism and industrial potential.

Diatoms are microalgae well known for their high variability and high primary productivity, being responsible for about 20% of the annual global carbon fixation. Moreover, they are interesting as potential feedstocks for the production of biofuels and high-value lipids and carotenoids. Diatoms exhibit trophic flexibility and, under certain conditions, they can grow mixotrophically by combing photosynthesis and respiration. So far, only a few species of diatoms have been tested for their mixotrophic metabolism; in some cases, they produced more biomass and with higher lipid content when grown under this condition. Phaeodactylum tricornutum is the most studied diatom species for its mixotrophic metabolism due to available genome sequence and molecular tools. However, studies in additional species are needed to better understand the conservation of this process in diatoms and its potential in industrial applications. Here, we describe the photosynthetic and respiratory pathways involved in mixotrophy and provide an overview of the trophic variability in diatoms. This review also highlights promising areas of industrial applications for diatoms when cultivated under mixotrophy.

Boosting Biomass Quantity and Quality by Improved Mixotrophic Culture of the Diatom Phaeodactylum tricornutum.

Diatoms are photoautotrophic unicellular algae and are among the most abundant, adaptable, and diverse marine phytoplankton. They are extremely interesting not only for their ecological role but also as potential feedstocks for sustainable biofuels and high-value commodities such as omega fatty acids, because of their capacity to accumulate lipids. However, the cultivation of microalgae on an industrial scale requires higher cell densities and lipid accumulation than those found in nature to make the process economically viable. One of the known ways to induce lipid accumulation in Phaeodactylum tricornutum is nitrogen deprivation, which comes at the expense of growth inhibition and lower cell density. Thus, alternative ways need to be explored to enhance the lipid production as well as biomass density to make them sustainable at industrial scale. In this study, we have used experimental and metabolic modeling approaches to optimize the media composition, in terms of elemental composition, organic and inorganic carbon sources, and light intensity, that boost both biomass quality and quantity of P. tricornutum. Eventually, the optimized conditions were scaled-up to 2 L photobioreactors, where a better system control (temperature, pH, light, aeration/mixing) allowed a further improvement of the biomass capacity of P. tricornutum to 12 g/L.

A systems-wide understanding of photosynthetic acclimation in algae and higher plants.

The ability of phototrophs to colonise different environments relies on robust protection against oxidative stress, a critical requirement for the successful evolutionary transition from water to land. Photosynthetic organisms have developed numerous strategies to adapt their photosynthetic apparatus to changing light conditions in order to optimise their photosynthetic yield, which is crucial for life on Earth to exist. Photosynthetic acclimation is an excellent example of the complexity of biological systems, where highly diverse processes, ranging from electron excitation over protein protonation to enzymatic processes coupling ion gradients with biosynthetic activity, interact on drastically different timescales from picoseconds to hours. Efficient functioning of the photosynthetic apparatus and its protection is paramount for efficient downstream processes, including metabolism and growth. Modern experimental techniques can be successfully integrated with theoretical and mathematical models to promote our understanding of underlying mechanisms and principles. This review aims to provide a retrospective analysis of multidisciplinary photosynthetic acclimation research carried out by members of the Marie Curie Initial Training Project, AccliPhot, placing the results in a wider context. The review also highlights the applicability of photosynthetic organisms for industry, particularly with regards to the cultivation of microalgae. It intends to demonstrate how theoretical concepts can successfully complement experimental studies broadening our knowledge of common principles in acclimation processes in photosynthetic organisms, as well as in the field of applied microalgal biotechnology.

Investigating mixotrophic metabolism in the model diatom Phaeodactylum tricornutum.

Diatoms are prominent marine microalgae, interesting not only from an ecological point of view, but also for their possible use in biotechnology applications. They can be cultivated in phototrophic conditions, using sunlight as the sole energy source. Some diatoms, however, can also grow in a mixotrophic mode, wherein both light and external reduced carbon contribute to biomass accumulation. In this study, we investigated the consequences of mixotrophy on the growth and metabolism of the pennate diatom Phaeodactylum tricornutum, using glycerol as the source of reduced carbon. Transcriptomics, metabolomics, metabolic modelling and physiological data combine to indicate that glycerol affects the central-carbon, carbon-storage and lipid metabolism of the diatom. In particular, provision of glycerol mimics typical responses of nitrogen limitation on lipid metabolism at the level of triacylglycerol accumulation and fatty acid composition. The presence of glycerol, despite provoking features reminiscent of nutrient limitation, neither diminishes photosynthetic activity nor cell growth, revealing essential aspects of the metabolic flexibility of these microalgae and suggesting possible biotechnological applications of mixotrophy.This article is part of the themed issue 'The peculiar carbon metabolism in diatoms'.

The Water to Water Cycles in Microalgae.

In oxygenic photosynthesis, light produces ATP plus NADPH via linear electron transfer, i.e. the in-series activity of the two photosystems: PSI and PSII. This process, however, is thought not to be sufficient to provide enough ATP per NADPH for carbon assimilation in the Calvin-Benson-Bassham cycle. Thus, it is assumed that additional ATP can be generated by alternative electron pathways. These circuits produce an electrochemical proton gradient without NADPH synthesis, and, although they often represent a small proportion of the linear electron flow, they could have a huge importance in optimizing CO2 assimilation. In Viridiplantae, there is a consensus that alternative electron flow comprises cyclic electron flow around PSI and the water to water cycles. The latter processes include photosynthetic O2 reduction via the Mehler reaction at PSI, the plastoquinone terminal oxidase downstream of PSII, photorespiration (the oxygenase activity of Rubisco) and the export of reducing equivalents towards the mitochondrial oxidases, through the malate shuttle. In this review, we summarize current knowledge about the role of the water to water cycles in photosynthesis, with a special focus on their occurrence and physiological roles in microalgae.

Energetic coupling between plastids and mitochondria drives CO2 assimilation in diatoms.

Diatoms are one of the most ecologically successful classes of photosynthetic marine eukaryotes in the contemporary oceans. Over the past 30 million years, they have helped to moderate Earth's climate by absorbing carbon dioxide from the atmosphere, sequestering it via the biological carbon pump and ultimately burying organic carbon in the lithosphere. The proportion of planetary primary production by diatoms in the modern oceans is roughly equivalent to that of terrestrial rainforests. In photosynthesis, the efficient conversion of carbon dioxide into organic matter requires a tight control of the ATP/NADPH ratio which, in other photosynthetic organisms, relies principally on a range of plastid-localized ATP generating processes. Here we show that diatoms regulate ATP/NADPH through extensive energetic exchanges between plastids and mitochondria. This interaction comprises the re-routing of reducing power generated in the plastid towards mitochondria and the import of mitochondrial ATP into the plastid, and is mandatory for optimized carbon fixation and growth. We propose that the process may have contributed to the ecological success of diatoms in the ocean.

Membrane glycerolipid remodeling triggered by nitrogen and phosphorus starvation in Phaeodactylum tricornutum.

Diatoms constitute a major phylum of phytoplankton biodiversity in ocean water and freshwater ecosystems. They are known to respond to some chemical variations of the environment by the accumulation of triacylglycerol, but the relative changes occurring in membrane glycerolipids have not yet been studied. Our goal was first to define a reference for the glycerolipidome of the marine model diatom Phaeodactylum tricornutum, a necessary prerequisite to characterize and dissect the lipid metabolic routes that are orchestrated and regulated to build up each subcellular membrane compartment. By combining multiple analytical techniques, we determined the glycerolipid profile of P. tricornutum grown with various levels of nitrogen or phosphorus supplies. In different P. tricornutum accessions collected worldwide, a deprivation of either nutrient triggered an accumulation of triacylglycerol, but with different time scales and magnitudes. We investigated in depth the effect of nutrient starvation on the Pt1 strain (Culture Collection of Algae and Protozoa no. 1055/3). Nitrogen deprivation was the more severe stress, triggering thylakoid senescence and growth arrest. By contrast, phosphorus deprivation induced a stepwise adaptive response. The time scale of the glycerolipidome changes and the comparison with large-scale transcriptome studies were consistent with an exhaustion of unknown primary phosphorus-storage molecules (possibly polyphosphate) and a transcriptional control of some genes coding for specific lipid synthesis enzymes. We propose that phospholipids are secondary phosphorus-storage molecules broken down upon phosphorus deprivation, while nonphosphorus lipids are synthesized consistently with a phosphatidylglycerol-to-sulfolipid and a phosphatidycholine-to-betaine lipid replacement followed by a late accumulation of triacylglycerol.

Ions channels/transporters and chloroplast regulation.

Ions play fundamental roles in all living cells and their gradients are often essential to fuel transports, to regulate enzyme activities and to transduce energy within and between cells. Their homeostasis is therefore an essential component of the cell metabolism. Ions must be imported from the extracellular matrix to their final subcellular compartments. Among them, the chloroplast is a particularly interesting example because there, ions not only modulate enzyme activities, but also mediate ATP synthesis and actively participate in the building of the photosynthetic structures by promoting membrane-membrane interaction. In this review, we first provide a comprehensive view of the different machineries involved in ion trafficking and homeostasis in the chloroplast, and then discuss peculiar functions exerted by ions in the frame of photochemical conversion of absorbed light energy.