Untargeted Metabolomics Unveil Changes in Autotrophic and Mixotrophic Galdieria sulphuraria Exposed to High-Light Intensity.

The thermoacidophilic red alga Galdieria sulphuraria has been optimizing a photosynthetic system for low-light conditions over billions of years, thriving in hot and acidic endolithic habitats. The growth of G. sulphuraria in the laboratory is very much dependent on light and substrate supply. Here, higher cell densities in G. sulphuraria under high-light conditions were obtained, although reductions in photosynthetic pigments were observed, which indicated this alga might be able to relieve the effects caused by photoinhibition. We further describe an extensive untargeted metabolomics study to reveal metabolic changes in autotrophic and mixotrophic G. sulphuraria grown under high and low light intensities. The up-modulation of bilayer lipids, that help generate better-ordered lipid domains (e.g., ergosterol) and keep optimal membrane thickness and fluidity, were observed under high-light exposure. Moreover, high-light conditions induced changes in amino acids, amines, and amide metabolism. Compared with the autotrophic algae, higher accumulations of osmoprotectant sugars and sugar alcohols were recorded in the mixotrophic G. sulphuraria. This response can be interpreted as a measure to cope with stress due to the high concentration of organic carbon sources. Our results indicate how G. sulphuraria can modulate its metabolome to maintain energetic balance and minimize harmful effects under changing environments.

Glycolytic Shunts Replenish the Calvin-Benson-Bassham Cycle as Anaplerotic Reactions in Cyanobacteria.

The recent discovery of the Entner-Doudoroff (ED) pathway as a third glycolytic route beside Embden-Meyerhof-Parnas (EMP) and oxidative pentose phosphate (OPP) pathway in oxygenic photoautotrophs requires a revision of their central carbohydrate metabolism. In this study, unexpectedly, we observed that deletion of the ED pathway alone, and even more pronounced in combination with other glycolytic routes, diminished photoautotrophic growth in continuous light in the cyanobacterium Synechocystis sp. PCC 6803. Furthermore, we found that the ED pathway is required for optimal glycogen catabolism in parallel to an operating Calvin-Benson-Bassham (CBB) cycle. It is counter-intuitive that glycolytic routes, which are a reverse to the CBB cycle and do not provide any additional biosynthetic intermediates, are important under photoautotrophic conditions. However, observations on the ability to reactivate an arrested CBB cycle revealed that they form glycolytic shunts that tap the cellular carbohydrate reservoir to replenish the cycle. Taken together, our results suggest that the classical view of the CBB cycle as an autocatalytic, completely autonomous cycle that exclusively relies on its own enzymes and CO2 fixation to regenerate ribulose-1,5-bisphosphate for Rubisco is an oversimplification. We propose that in common with other known autocatalytic cycles, the CBB cycle likewise relies on anaplerotic reactions to compensate for the depletion of intermediates, particularly in transition states and under fluctuating light conditions that are common in nature.

Physiological and Ecological Aspects of Chlorella sorokiniana (Trebouxiophyceae) Under Photoautotrophic and Mixotrophic Conditions.

Mixotrophy is a metabolic strategy in which an organism is autotrophic and heterotrophic simultaneously. Considering that the aquatic environment provides several organic sources of carbon, it is probably common for microalgae to perform mixotrophy and not only photoautotrophy, but little is known about microalgae mixotrophy. The present work aimed at investigating the growth, photosynthetic activity, morphology, and biochemical composition of the microalga Chlorella sorokiniana in mixotrophic and photo-mixotrophic conditions, comparing it with photoautotrophy. The results showed pH changes after glucose addition, reaching pH 11.62 in mixotrophic and 10.47 in sequential photo-mixotrophic cultures, which limited the microalgal growth. Highest biomass was obtained in the mixotrophic culture in comparison with the sequential photo-mixotrophic one. Rapid light saturation curves showed that α (photosynthetic efficiency, 1.69) and relative electron transport rate (rETR; 565.61) were higher in the mixotrophic cultures, whereas the highest Ik (irradiance saturation, 386.68) was obtained in the photoautotrophic ones. In the sequential photo-mixotrophic cultures, photosynthetic activity varied during glucose consumption, decreasing the maximum quantum yield Fv/Fm after glucose addition, indicating change in metabolism, from photoautotrophy to mixotrophy by the microalga. The results showed that the mixotrophic cultures had higher production of chlorophyll a (6.26 mg mL-1), cell density (6.62 × 107 cell mL-1), and lipids (0.06 pg µm-3). Sequential photo-mixotrophic cultures showed the highest biovolume (360.5 µm3 cell-1) and total carbohydrates (0.026 pg µm-3). The protein concentration was 3.2 and 2.4 times higher in photoautotrophy and photo-mixotrophic growth, respectively, than in mixotrophy, but lipids were three times higher under mixotrophy. The biochemical changes we observed indicate that the microalga's plasticity in face of new environmental characteristics, such as the presence of organic carbon, can change the flow of energy through natural ecosystems.

Novel quantitative insights into carbon sources for synthesis of poly hydroxybutyrate in Synechocystis PCC 6803.

Many freshwater cyanobacteria accumulate polyhydroxybutyrate (PHB) under nitrogen or phosphorus deprivation. While prior literature has shed lights on transcriptomic and metabolomic changes in the model cyanobacterium Synechocystis PCC 6803 cells, the quantitative contributions of the newly fixed carbon following nitrogen deprivation or the externally added acetate to PHB synthesis are not clear. Similarly, it is not clear how photomixotrophy affects precursor contributions. In this study, we show that (i) the pre-growth mode (photoautotrophic or photomixotrophic), while significantly impacting glycogen levels, does not have any significant effect on PHB levels, (ii) the carbon fixed following nitrogen deprivation contributes 26% of C for PHB synthesis in photoautotrophically pre-grown cells and its contribution to the PHB synthesis goes down with the addition of acetate at the resuspension phase or with photomixotrophic pre-growth, (iii) the acetate added at the start of nitrogen deprivation, doubles the intracellular PHB levels and contributes 44-48% to PHB synthesis and this value is not greatly affected by how the cells were pre-grown. Indirectly, the labeling studies also show that the intracellular C recycling is the most important source of precursors for PHB synthesis, contributing about 74-87% of the C for PHB synthesis in the absence of acetate. The addition of acetate significantly reduces its contribution. In photoautotrophic pre-growth followed by acetate addition under nitrogen starvation, the contribution of intracellular C reduces to about 34%. Thus, our study provides several novel quantitative insights on how prior nutritional status affects the precursor contributions for PHB synthesis.

Chlamydomonas reinhardtii responding to high light: a role for 2-propenal (acrolein).

High light causes photosystem II to generate singlet oxygen (1 O2 ), a reactive oxygen species (ROS) that can react with membrane lipids, releasing reactive electrophile species (RES), such as acrolein. To investigate how RES may contribute to light stress responses, Chlamydomonas reinhardtii was high light-treated in photoautotrophic and mixotrophic conditions and also in an oxygen-enriched atmosphere to elevate ROS production. The responses were compared to exogenous acrolein. Non-photochemical quenching (NPQ) was higher in photoautotrophic cells, as a consequence of a more de-epoxidized state of the xanthophyll cycle pool and more LHCSR3 protein, showing that photosynthesis was under more pressure than in mixotrophic cells. Photoautotrophic cells had lowered α-tocopherol and ß-carotene contents and a higher level of protein carbonylation, indicators of elevated 1 O2 production. Levels of glutathione, glutathione peroxidase (GPX5) and glutathione-S-transferase (GST1), important antioxidants against RES, were also increased in photoautotrophic cells. In parallel to the wild-type, the LHCSR3-deficient npq4 mutant was high light-treated, which in photoautotrophic conditions exhibited particular sensitivity under elevated oxygen, the treatment that induced the highest RES levels, including acrolein. The npq4 mutant had more GPX5 and GST1 alongside higher levels of carbonylated protein and a more oxidized glutathione redox state. In wild-type cells glutathione contents doubled after 4 h treatment, either with high light under elevated oxygen or with a non-critical dose (600 ppm) of acrolein. Exogenous acrolein also increased GST1 levels, but not GPX5. Overall, RES-associated oxidative damage and glutathione metabolism are prominently associated with light stress and potentially in signaling responses of C. reinhardtii.

Dispensability of a sulfolipid for photoautotrophic cell growth and photosynthesis in a marine cyanobacterium, Synechococcus sp. PCC 7002.

Sulfoquinovosyl diacylglycerol, which mainly comprises thylakoid membranes in oxygenic photosynthetic organisms, plays species-dependent roles in freshwater microbes. In this study, a sulfoquinovosyl-diacylglycerol deficient mutant was generated in a cyanobacterium, Synechococcus sp. PCC 7002, for the first time among marine microbes to gain more insight into its physiological significance. The mutation had little deleterious impact on photoautotrophic cell growth, and functional and structural properties of the photosystem II complex. These findings were similar to previous observations for a freshwater cyanobacterium, Synechococcus elongatus PCC 7942, but were distinct from those for another freshwater cyanobacterium, Synechocystis sp. PCC 6803, and a green alga, Chlamydomonas reinhardtii, both of which require sulfoquinovosyl diacylglycerol for cell growth and/or photosystem II. Therefore, the functionality of PSII to dispense with sulfoquinovosyl diacylglycerol in Synechococcus sp. PCC 7002, similar to that in Synechococcus elongatus PCC 7942, seemed to have been excluded from the evolution of the PSII complex from cyanobacteria to green algal chloroplasts. Meanwhile, sulfoquinovosyl diacylglycerol was found to contribute to photoheterotrophic growth of Synechococcus sp. PCC 7002, which revealed a novel species-dependent strategy for utilizing SQDG in physiological processes.

A new bioenergetic and thermodynamic approach to batch photoautotrophic growth of Arthrospira (Spirulina) platensis in different photobioreactors and under different light conditions.

Photobioreactor configuration, mode of operation and light intensity are known to strongly impact on cyanobacteria growth. To shed light on these issues, kinetic, bioenergetic and thermodynamic parameters of batch Arthrospira platensis cultures were estimated along the time at photosynthetic photon flux density (PPFD) of 70µmolm(-2)s(-1) in different photobioreactors with different surface/volume ratio (S/V), namely open pond (0.25cm(-1)), shaken flask (0.48cm(-1)), horizontal photobioreactor (HoP) (1.94cm(-1)) and helicoidal photobioreactor (HeP) (3.88cm(-1)). Maximum biomass concentration and productivity remarkably increased with S/V up to 1.94cm(-1). HoP was shown to be the best-performing system throughout the whole runs, while HeP behaved better only at the start. Runs carried out in HoP increasing PPFD from 40 to 100µmolm(-2)s(-1) revealed a progressive enhancement of bioenergetics and thermodynamics likely because of favorable light distribution. HoP appeared to be a promising configuration to perform high-yield indoor cyanobacterial cultures.

The Type II NADPH Dehydrogenase Facilitates Cyclic Electron Flow, Energy-Dependent Quenching, and Chlororespiratory Metabolism during Acclimation of Chlamydomonas reinhardtii to Nitrogen Deprivation.

When photosynthetic organisms are deprived of nitrogen (N), the capacity to grow and assimilate carbon becomes limited, causing a decrease in the productive use of absorbed light energy and likely a rise in the cellular reduction state. Although there is a scarcity of N in many terrestrial and aquatic environments, a mechanistic understanding of how photosynthesis adjusts to low-N conditions and the enzymes/activities integral to these adjustments have not been described. In this work, we use biochemical and biophysical analyses of photoautotrophically grown wild-type and mutant strains of Chlamydomonas reinhardtii to determine the integration of electron transport pathways critical for maintaining active photosynthetic complexes even after exposure of cells to N deprivation for 3 d. Key to acclimation is the type II NADPH dehydrogenase, NDA2, which drives cyclic electron flow (CEF), chlororespiration, and the generation of an H(+) gradient across the thylakoid membranes. N deprivation elicited a doubling of the rate of NDA2-dependent CEF, with little contribution from PGR5/PGRL1-dependent CEF The H(+) gradient generated by CEF is essential to sustain nonphotochemical quenching, while an increase in the level of reduced plastoquinone would promote a state transition; both are necessary to down-regulate photosystem II activity. Moreover, stimulation of NDA2-dependent chlororespiration affords additional relief from the elevated reduction state associated with N deprivation through plastid terminal oxidase-dependent water synthesis. Overall, rerouting electrons through the NDA2 catalytic hub in response to photoautotrophic N deprivation sustains cell viability while promoting the dissipation of excess excitation energy through quenching and chlororespiratory processes.

Microdroplet photobioreactor for the photoautotrophic culture of microalgal cells.

Microalgae, unicellular photoautotrophic microorganisms, have attracted great attention for the production of biofuel and high-value products, but the commercial use of microalgae has been limited by low photosynthetic productivity. To overcome this limitation, it is required to develop an efficient platform for the rapid evaluation of photoautotrophic growth performance and productivity of microalgal strains. Here we describe a droplet-based photobioreactor for high-throughput analysis of the photoautotrophic growth of microalgal cells. By integrating micropillar arrays and adjusting the height of the microchamber, we could accurately monitor the growth kinetics of microalgae in an immobilized microdroplet and improve the transfer rate of CO2 into the microdroplet photobioreactor with an increased contact area between the microdroplet and PDMS surface. The improvement of CO2 transfer into the microdroplet was also confirmed by improved microalgal cell growth and a decrease in pH measured using colorimetric and fluorescence-based assays. The photoautotrophic growth kinetics of Chlorella vulgaris were measured under different CO2 concentrations (ambient, 1%, 2.5%, 5% and 7.5%) and light intensity (35, 55, 100, 150, and 200 µmol photons per m(2) per s) conditions, which are key factors for photoautotrophic growth. Chlorella vulgaris in a microdroplet showed better cell growth performance compared to a flask culture due to the reduced shading effects and improved mass transfer. Finally, we could evaluate the photoautotrophic growth performance of four microalgal strains (Chlorella vulgaris, Chlorella protothecoides, Chlorella sorokiniana and Neochloris oleoabundans) for 120 hours. These results demonstrate that our microdroplet system can be used as an efficient photobioreactor for the rapid evaluation of the photoautotrophic growth of microalgal strains under various conditions.

An efficient screening method for the isolation of heterotrophic bacteria influencing growth of diatoms under photoautotrophic conditions.

Interactions between photoautotrophic diatoms and heterotrophic bacteria are important for the biogeochemical C-cycle in the oceans. Additionally, biofilms formed by diatoms and bacteria are the initiating step of biofouling processes, which causes high costs in shipping. Despite this ecological and economical importance, the knowledge about biochemical and molecular mechanisms underlying these interkingdom interactions is relatively small. For analyzing these mechanisms, laboratory model systems are required. In this study, an efficient screening method for isolating bacteria influencing photoautotrophic diatom growth was established. First, diatom cultures of Phaeodactylum tricornutum and Thalassiosira pseudonana were made axenic by applying ß-lactam antibiotics. Second, a non-invasive method for measuring growth of multiple parallel diatom cultures by chlorophyll fluorescence was established. This method allowed semi-quantitative chlorophyll determination of cultures with up to 3 µg (chlorophyll) ml(-1). Axenic diatom cultures were then used for enriching bacteria and led to the isolation of 24 strains influencing growth of both diatom strains in various ways. For example, Rheinheimera sp. strain Tn16 inhibited growth of T. pseudonana, while it stimulated growth and cell aggregation of P. tricornutum. Thus, this screening method is appropriate for isolating heterotrophic bacteria showing different interactions with different diatom species ranging from synergistic to antagonistic. In consecutive applications, this method will be useful to screen for bacterial mutants with altered phenotypes regarding the influence on diatom growth.

Fine-Tuning of Photoautotrophic Protein Production by Combining Promoters and Neutral Sites in the Cyanobacterium Synechocystis sp. Strain PCC 6803.

Cyanobacteria are photosynthetic cell factories that use solar energy to convert CO2 into useful products. Despite this attractive feature, the development of tools for engineering cyanobacterial chassis has lagged behind that for heterotrophs such as Escherichia coli or Saccharomyces cerevisiae. Heterologous genes in cyanobacteria are often integrated at presumptively "neutral" chromosomal sites, with unknown effects. We used transcriptome sequencing (RNA-seq) data for the model cyanobacterium Synechocystis sp. strain PCC 6803 to identify neutral sites from which no transcripts are expressed. We characterized the two largest such sites on the chromosome, a site on an endogenous plasmid, and a shuttle vector by integrating an enhanced yellow fluorescent protein (EYFP) expression cassette expressed from either the Pcpc560 or the Ptrc1O promoter into each locus. Expression from the endogenous plasmid was as much as 14-fold higher than that from the chromosome, with intermediate expression from the shuttle vector. The expression characteristics of each locus correlated predictably with the promoters used. These findings provide novel, characterized tools for synthetic biology and metabolic engineering in cyanobacteria.

Enhanced autotrophic astaxanthin production from Haematococcus pluvialis under high temperature via heat stress-driven Haber-Weiss reaction.

High temperatures (30-36 °C) inhibited astaxanthin accumulation in Haematococcus pluvialis under photoautotrophic conditions. The depression of carotenogenesis was primarily attributed to excess intracellular less reactive oxygen species (LROS; O2 (-) and H2O2) levels generated under high temperature conditions. Here, we show that the heat stress-driven inefficient astaxanthin production was improved by accelerating the iron-catalyzed Haber-Weiss reaction to convert LROS into more reactive oxygen species (MROS; O2 and OH·), thereby facilitating lipid peroxidation. As a result, during 18 days of photoautotrophic induction, the astaxanthin concentration of cells cultured in high temperatures in the presence of iron (450 µM) was dramatically increased by 75 % (30 °C) and 133 % (36 °C) compared to that of cells exposed to heat stress alone. The heat stress-driven Haber-Weiss reaction will be useful for economically producing astaxanthin by reducing energy cost and enhancing photoautotrophic astaxanthin production, particularly outdoors utilizing natural solar radiation including heat and light for photo-induction of H. pluvialis.

Screening and characterization of oleaginous Chlorella strains and exploration of photoautotrophic Chlorella protothecoides for oil production.

The growth and oil production of nine Chlorella strains were comparatively assessed and Chlorellaprotothecoides CS-41 demonstrated the greatest lipid production potential. The effects of different nitrogen forms and concentrations, phosphorus concentrations and light intensities on growth and oil production were studied in laboratory columns. C. protothecoides CS-41 accumulated lipids up to 55% of dry weight, with triacylglycerol and oleic acid being 71% of total lipids and 59% of total fatty acids, respectively. High biomass and lipid productivities were achieved in outdoor panel PBRs, up to 1.25 and 0.59 g L(-1) day(-1), or 44. 1 and 16.1 g m(-2) day(-1), respectively. A two-stage cultivation strategy was proposed to enhance the algal biomass and lipid production. This is the first comprehensive investigation of both indoor and outdoor photoautotrophic C. protothecoides cultures for oil production, and C. protothecoides CS-41 represents a promising biofuel feedstock worthy of further exploration.

Downregulation of a putative plastid PDC E1α subunit impairs photosynthetic activity and triacylglycerol accumulation in nitrogen-starved photoautotrophic Chlamydomonas reinhardtii.

The chloroplast pyruvate dehydrogenase complex (cpPDC) catalyses the oxidative decarboxylation of pyruvate forming acetyl-CoA, an immediate primer for the initial reactions of de novo fatty acid (FA) synthesis. Little is known about the source of acetyl-CoA in the chloroplasts of photosynthetic microalgae, which are capable of producing high amounts of the storage lipid triacylglycerol (TAG) under conditions of nutrient stresses. We generated Chlamydomonas reinhardtii CC-1618 mutants with decreased expression of the PDC2_E1α gene, encoding the putative chloroplast pyruvate dehydrogenase subunit E1α, using artificial microRNA. A comparative study on the effects of PDC2_E1α silencing on FAs and TAG production in C. reinhardtii, grown photoautotrophically and mixotrophically, with and without a nitrogen source in the nutrient medium, was carried out. Reduced expression of PDC2 _E1α led to a severely hampered photoautotrophic growth phenotype with drastic impairment in TAG accumulation under nitrogen deprivation. In the presence of acetate, downregulation of PDC2_E1α exerted little to no effect on TAG production and photosynthetic activity. In contrast, under photoautotrophic conditions, especially in the absence of a nitrogen source, a dramatic decline in photosynthetic oxygen evolution and photosystem II quantum yield against a background of the apparent over-reduction of the photosynthetic electron chain was recorded. Our results suggest an essential role of cpPDC in the supply of carbon precursors for de novo FA synthesis in microalgae under conditions of photoautotrophy. A shortage of this supply is detrimental to the nitrogen-starvation-induced synthesis of storage TAG, an important carbon and energy sink in stressed Chlamydomonas cells, thereby impairing the acclimation ability of the microalga.

Novel flat-plate photobioreactors for microalgae cultivation with special mixers to promote mixing along the light gradient.

Novel flat-plate photobioreactors (PBRs) with special mixers (type-a, type-b, and type-c) were designed based on increased mixing degree along the light gradient. The hydrodynamic and light regime characteristic of the novel PBRs were investigated through computational fluid dynamics. Compared with the control reactor without mixer, the novel reactors can effectively increase liquid velocity along the light gradient, the frequency of light/dark (L/D) cycles, and the algal growth rates of Chlorella pyrenoidosa. The maximum biomass concentrations in type-a, type-b, and type-c reactors were 42.9% (1.3 g L(-1)), 31.9% (1.2 g L(-1)), and 20.9% (1.1 g L(-1)) higher than that in the control reactor (0.91 g L(-1)), respectively, at an aeration rate of 1.0 vvm. Correlation analysis of algal growth rate with the characteristics of mixing and light regime shows the key factors affecting algal photoautotrophic growth are liquid velocity along the light gradient and L/D cycles rather than the macro-mixing degree.

Environmental pH affects photoautotrophic growth of Synechocystis sp. PCC 6803 strains carrying mutations in the lumenal proteins of PSII.

Synechocystis sp. strain PCC 6803 grows photoautotrophically across a broad pH range, but wild-type cultures reach a higher density at elevated pH; however, photoheterotrophic growth is similar at high and neutral pH. A number of PSII mutants each lacking at least one lumenal extrinsic protein, and carrying a second PSII lumenal mutation, are able to grow photoautotrophically in BG-11 medium at pH 10.0, but not pH 7.5. We investigated the basis of this pH effect and observed no pH-specific change in variable fluorescence yield from PSII centers of the wild type or the pH-dependent ΔPsbO:ΔPsbU and ΔPsbV:ΔCyanoQ strains; however, 77 K fluorescence emission spectra indicated increased coupling of the phycobilisome (PBS) antenna at pH 10.0 in all mutants. DNA microarray data showed a cell-wide response to transfer from pH 10.0 to pH 7.5, including decreased mRNA levels of a number of oxidative stress-responsive transcripts. We hypothesize that this transcriptional response led to increased tolerance against reactive oxygen species and in particular singlet oxygen. This response enabled photoautotrophic growth of the PSII mutants at pH 10.0. This hypothesis was supported by increased resistance of all strains to rose bengal at pH 10.0 compared with pH 7.5.

Lipid production by Chlorella vulgaris after a shift from nutrient-rich to nitrogen starvation conditions.

A two-stage process, composed of growth under nutrient-rich conditions followed by cultivation under nitrogen starvation and controlled conditions of phosphate, light intensity, aeration, and carbon sources was applied for lipid production by the green alga Chlorella vulgaris. Using conditions without addition of nitrogen, 2mg/L PO(4)-P, light intensity of 100µmol/m(2)/s and 0.25vvm of air, about 43% of dry cell weight accumulated as lipids after 12h, which equates to a lipid productivity of 77.8mg/L/d. In a medium containing 5mg/L NO(3)-N and 2mg/L PO(4)-P, and at a light intensity of 100µmol/m(2)/s and 0.25vvm of 2% CO(2), about 53% of dry cell weight consisted of lipids after 24h, representing a lipid productivity of 77.1mg/L/d. The low amount of nutrients, moderate aeration and light intensity were helpful for increasing lipid productivity.

Retrograde signaling and photoprotection in a gun4 mutant of Chlamydomonas reinhardtii.

GUN4 is a regulatory subunit of Mg-chelatase involved in the control of tetrapyrrole synthesis in plants and cyanobacteria. Here, we report the first characterization of a gun4 insertion mutant of the unicellular green alga Chlamydomonas reinhardtii. The mutant contains 50% of chlorophyll as compared to wild-type and accumulates ProtoIX. In contrast to the increase in LHC transcription, the accumulation of most LHC proteins is drastically diminished, implying posttranscriptional down-regulation in the absence of transcriptional coordination. We found that 803 genes change their expression level in gun4 as compared to wild-type, by RNA-Seq, and this wide-ranging effect on transcription is apparent under physiological conditions. Besides LHCs, we identified transcripts encoding enzymes of the tetrapyrrole pathway and factors involved in signal transduction, transcription, and chromatin remodeling. Moreover, we observe perturbations in electron transport with a strongly decreased PSI-to-PSII ratio. This is accompanied by an enhanced activity of the plastid terminal oxidase (PTOX) that could have a physiological role in decreasing photosystem II excitation pressure.

Mixotrophic cultivation of Chlorella vulgaris using industrial dairy waste as organic carbon source.

Growth parameters and biochemical composition of the green microalga Chlorella vulgaris cultivated under different mixotrophic conditions were determined and compared to those obtained from a photoautotrophic control culture. Mixotrophic microalgae showed higher specific growth rate, final biomass concentration and productivities of lipids, starch and proteins than microalgae cultivated under photoautotrophic conditions. Moreover, supplementation of the inorganic culture medium with hydrolyzed cheese whey powder solution led to a significant improvement in microalgal biomass production and carbohydrate utilization when compared with the culture enriched with a mixture of pure glucose and galactose, due to the presence of growth promoting nutrients in cheese whey. Mixotrophic cultivation of C. vulgaris using the main dairy industry by-product could be considered a feasible alternative to reduce the costs of microalgal biomass production, since it does not require the addition of expensive carbohydrates to the culture medium.

Enhancement of microalgal biomass and lipid productivities by a model of photoautotrophic culture with heterotrophic cells as seed.

For overcoming the long period of seed cultured photoautotrophically and inadequate cell supply for the inoculation of microalgae photoautotrophic cultivation, a model for the photoautotrophic culture of three Chlorella species with heterotrophic cells as seed was investigated. The model can not only take advantages of rapid cell growth in heterotrophic process for preparation of cells as seed but also increase the biomass and lipid productivities of the microalgae cultivated photoautotrophically. The results showed that biomass productivities of Chlorella pyrenoidosa, Chlorella ellipsoidea and Chlorella vulgaris cultured by heterotrophy were 20.9, 26.9 and 25.2 times higher than those by photoautotrophy in seed culturing period. In the subsequent photoautotrophic culture, the biomass and lipid productivities of C. pyrenoidosa, C. ellipsoidea and C. vulgaris with heterotrophic seed were 1.91, 1.51, 1.48 and 1.66, 1.37, 1.42 times higher than those with photoautotrophic seed. Furthermore, the culture model was also carried out successfully outdoor.