Red Microalga Galdieria Sulphuraria Research Articles

Mixotrophic and heterotrophic growth of Galdieria sulphuraria using buttermilk as a carbon source

The growth of the polyextremophile red microalga Galdieria sulphuraria ACUF 064 was evaluated under mixotrophy and heterotrophy in a 13-L lab-scale stirred photobioreactor, using buttermilk as a carbon source. Beforehand, G. sulphuraria ACUF 064 growth on glucose, galactose and lactose was evaluated. No significant differences were observed in terms of growth when lactose, glucose or galactose was used as a carbon source. Overall, the biomass yield on carbon was 70% higher in mixotrophy than heterotrophy for galactose (2.0 gx gC−1) and lactose (2.0 gx gC−1), while it was similar for glucose (1.5 gx gC−1). At the same time, the yield of biomass on nitrogen for cultures grown in lactose was the lowest in comparison to all the other tested substrates. This was also supported by a higher C-phycocyanin content, with 5.9% wC-PC/wx as the highest value in mixotrophy. A preliminary experiment in flasks under mixotrophic conditions with different buttermilk dilutions revealed that a dilution ratio of 40% v/v of buttermilk (corresponding to 2.0 gC L−1) was optimal for biomass production. Finally, G. sulphuraria ACUF 064 was grown in the optimal buttermilk dilution ratio in a 13 L photobioreactor. The highest biomass productivity was also obtained in mixotrophy (0.55 gx L−1 d−1), corresponding to a carbon removal of 61%. Overall, lactose-containing substrates such as buttermilk hold promise as a substrate for the growth of G. sulphuraria while revalorizing an industrial effluent.

Cell adaptation of the extremophilic red microalga Galdieria sulphuraria to the availability of carbon sources.

Global energy demand and fossil fuels impact on climate can be partially managed by an increase in the use of biofuels for transports and industries. Biodiesel production is generally preceded by a transesterification process of the green biomass triacylglycerols that generates large amounts of glycerol as a by-product. In this study, the extremophilic red microalga Galdieria sulphuraria 074W was cultivated in heterotrophy. The microalgal growth parameters and biomass composition were compared when grown on an equivalent molar concentration of carbon of either glucose or glycerol as unique carbon source. The maximal biomass reached in these two conditions was not significantly different (∼2.5 g.L–1). Fatty acid profile, protein and storage carbohydrate contents were also statistically similar, irrespectively of the metabolized carbon source. We also observed that the pigment content of G. sulphuraria cells decreased during heterotrophic growth compared to photoautotrophic cultivated cells, and that this diminution was more important in the presence of glucose than glycerol: cells were yellowish in the presence of glucose and green in the presence of glycerol. The pigmentation was restored when glucose was totally consumed in the medium, suggesting that the presence of glucose repressed pigment synthesis. Based on this observation, a transcriptome analysis was performed in order to better understand the mechanisms involved in the loss of color mediated by darkness and by glucose in G. sulphuraria. Three conditions were analyzed: heterotrophy with glycerol or glucose and phototrophy. This allowed us to understand the transcriptional response of cells to light and dark environments both at the nuclear and chloroplast levels, and to show that transcription of gene families, acquired by horizontal gene transfer, such as sugar, amino acid, or acetate transporters, were involved in the response to the availability of different (in)organic sources.

Growth under Different Trophic Regimes and Synchronization of the Red Microalga Galdieria sulphuraria.

The extremophilic unicellular red microalga Galdieria sulphuraria (Cyanidiophyceae) is able to grow autotrophically, or mixo- and heterotrophically with 1% glycerol as a carbon source. The alga divides by multiple fission into more than two cells within one cell cycle. The optimal conditions of light, temperature and pH (500 µmol photons m−2 s−1, 40 °C, and pH 3; respectively) for the strain Galdieria sulphuraria (Galdieri) Merola 002 were determined as a basis for synchronization experiments. For synchronization, the specific light/dark cycle, 16/8 h was identified as the precondition for investigating the cell cycle. The alga was successfully synchronized and the cell cycle was evaluated. G. sulphuraria attained two commitment points with midpoints at 10 and 13 h of the cell cycle, leading to two nuclear divisions, followed subsequently by division into four daughter cells. The daughter cells stayed in the mother cell wall until the beginning of the next light phase, when they were released. Accumulation of glycogen throughout the cell cycle was also described. The findings presented here bring a new contribution to our general understanding of the cell cycle in cyanidialean red algae, and specifically of the biotechnologically important species G. sulphuraria.

Comparison of Galdieria growth and photosynthetic activity in different culture systems

In the last years, the acidothermophilic red microalga Galdieria sulphuraria has been increasingly studied for industrial applications such as wastewater treatment, recovery of rare earth elements, production of phycobilins. However, even now it is not possible an industrial cultivation of this organism because biotechnological research on G. sulphuraria and allied species is relatively recent and fragmented. Having in mind a possible scale-up for commercial applications, we have compared the growth and photosynthetic performance of G. sulphuraria in four suspended systems (Inclined bubble column, Decanter Laboratory Flask, Tubular Bioreactor, Ultra-flat plate bioreactor) and one immobilized system (Twin Layer Sytem). The results showed that G. sulphuraria had the highest growth, productivity and photosynthetic performance, when grown on the immobilized system, which also offers some economics advantages.

Comparative study on light attenuation models of Galdieria sulphuraria for efficient production of phycocyanin

The thermo-acidophilic red microalga Galdieria sulphuraria is valuable especially for the intracellular thermostable phycocyanin with broad application prospects. Heterotrophy or photoinduction had been studied to enhance the production of phycocyanin at small-scale indoors. The aim of this study was to scale up the photoinduction process. The Cornet model could well describe and predict the light attenuation in G. sulphuraria suspension compared with the Lambert-Beer and hyperbolic models. Volume-averaged light intensity was the key parameter for photoinduction by testing respectively in 1-L column bioreactor, 3-L and 15-L flat plate photobioreactors indoors and outdoors, and supposed to be 600 μmol photons m−2 s−1. Accordingly, amplifying photoinduction in 1.35-m2 raceway pond outdoors was successfully carried out with the final cell dry weight of 2.37 g L−1 and phycocyanin up to 10.32%, which was comparable with that from Spirulina/Arthrospira and 3.82 times higher than that of heterotrophic cells of G. sulphuraria.

Biomass and phycocyanin content of heterotrophic Galdieria sulphuraria 074G under maltodextrin and granular starches\u2013feeding conditions

A major disadvantage of microalgal cultivation is limited biomass yields due to the autotrophic lifestyle of most microalgal species. Heterotrophic growth on a suitable carbon source and oxygen can overcome such limitations. The red microalga Galdieria sulphuraria strain 074G grows heterotrophically on glucose and a number of other carbon sources while constitutively producing photopigments, including the blue-colored phycocyanin, a natural food colorant. Galdieria sulphuraria strain 074G grew well on maltodextrins as well as on granular starch in combination with the enzyme cocktail Stargen002. The maltodextrin cultures produced 2 mg phycocyanin per gram substrate, being slightly more than on glucose. The phycocyanin extracted from maltodextrin-grown cultures was thermostable up to 55 °C. Maltodextrins can be a cheap alternative to glucose syrups for the production of phycocyanin as natural food colorant.

The glycogen of Galdieria sulphuraria as alternative to starch for the production of slowly digestible and resistant glucose polymers

Highly branched glucose polymers produced from starch are applied in various products, such as peritoneal dialysis solutions and sports drinks. Due to its insoluble, granular nature, the use of native starch as substrate requires an energy consuming pre-treatment to achieve solubilization at the expense of process costs. Glycogen, like starch, is also a natural glucose polymer that shows more favorable features, since it is readily soluble in cold water and more accessible by enzymes. The extremophilic red microalga Galdieria sulphuraria accumulates large amounts of a small, highly branched glycogen that could represent a good alternative to starch as substrate for the production of highly branched glucose polymers. In the present work, we analyzed the structure-properties relationship of this glycogen in its native form and after treatment with amyloglucosidase and compared it to highly branched polymers produced from potato starch. Glycogen showed lower susceptibility to digestive enzymes and significantly decreased viscosity in solution compared to polymers derived from starch, properties conferred by its shorter side chains and higher branch density. The action of amyloglucosidase on native glycogen was somewhat limited due to the high branch density but resulted in the production of a hyperbranched polymer that was virtually resistant to digestive enzymes.

Floridoside production by the red microalga Galdieria sulphuraria under different conditions of growth and osmotic stress.

Floridoside is a compatible solute synthesized by red algae that has attracted considerable attention due to its promising antifouling and therapeutic properties. However, research on industrial applications of floridoside is hampered by limited compound availability and the development of a production process yielding high amounts of this glycoside has not been explored yet. In the present work, floridoside accumulation by the red microalgae Galdieria sulphuraria under different conditions was investigated in order to optimize the production of this glycoside in this microalgae. G. sulphuraria shows consider advantages over other red algae as potential industrial producer of floridoside due to its unicellular nature, its ability to grow heterotrophically in complete darkness and its acidophilic lifestyle. The main compatible solute accumulated by G. sulphuraria under salt stress was purified, identified as floridoside by 1H-NMR and used as standard for quantification. Our results showed that applying the osmotic stress after the cells had grown first in medium with no salt resulted in higher floridoside yields compared to those obtained in cells growing under osmotic stress from the beginning. Among several parameters tested, the use of glycerol as carbon source for cell growth showed the most significant impact on floridoside accumulation, which reached a maximum of 56.8 mg/g dry biomass.

Characterization of the highly branched glycogen from the thermoacidophilic red microalga Galdieria sulphuraria and comparison with other glycogens

The thermoacidophilic red microalga Galdieria sulphuraria synthesizes glycogen when growing under heterotrophic conditions. Structural characterization revealed that G. sulphuraria glycogen is the most highly branched glycogen described to date, with 18% of α-(1→6) linkages. Moreover, it differs from other glycogens because it is composed of short chains only and has a substantially smaller molecular weight and particle size. The physiological role of this highly branched glycogen in G. sulphuraria is discussed.

Lipidomic analysis of the extremophilic red alga Galdieria sulphuraria in response to changes in pH

Analysis of polar lipids from the red micro-alga Galdieria sulphuraria by means of the HILIC-LC/high resolution ESI-tandem MS determined 14 classes of lipids including identification of tens of molecular species of lipids and their regioisomers. The extremophilic alga G. sulphuraria uses several mechanisms for adaptation to low pH and high temperature, the major change being a replacement of cellular phospholipids by betaine lipids. We also found a change in the ratio of regioisomers of the polar lipids, i.e. phospho-, glyco- and betaine lipids, in dependence on the culture medium pH. We hypothesize that biosynthesis of polar lipids via the prokaryotic and/or eukaryotic pathway is dependent on ambient pH.

Microalgae as human food: chemical and nutritional characteristics of the thermo-acidophilic microalga Galdieria sulphuraria

The use of microalgae as a food source is still poorly developed because of the technical difficulties related to their cultivation and the limited knowledge about their chemical composition and nutritional value. The unicellular red microalga Galdieria sulphuraria has a very high daily productivity and its cultivation under acidic conditions avoided any bacterial contamination. G. sulphuraria can be cultured under autotrophic and heterotrophic conditions: in this study a screening of 43 strains showed that in the latter case a duplication of biomass production was obtained. The proximate composition (protein, carbohydrates, fiber and lipids), the micronutrient content (carotenoids, phycobiliproteins, chlorophylls and vitamins) together with the antioxidant activity of the biomass produced by a selected strain of G. sulphuraria under both cultivation conditions were determined. Results showed that the material is rich in proteins (26-32%) and polysaccharides (63-69%) and poor in lipids. Under heterotrophic cultivation conditions, the lipid moiety mainly contained monounsaturated fatty acids. Among micronutrients, some B group vitamins are present, beta-carotene is the main carotenoid and phycobiliproteins are present under both cultivating conditions. G. sulphuraria proteins are strictly associated with polysaccharide components and therefore not digestible. In the second part of the work, an extraction protocol using Viscozyme L, a commercial enzymatic preparation containing a mixture of polysaccharidases, was developed which made G. sulphuraria proteins a good substrate for human gastrointestinal enzymes. All in all, the data suggested that G. sulphuraria biomass has a potential use as food ingredients both for protein-rich or insoluble dietary fibre-rich applications. The low concentration of lipids and the absence of green color make this microalgae source particularly useful for the addition to many food preparations.

Far-red light-regulated efficient energy transfer from phycobilisomes to photosystem I in the red microalga Galdieria sulphuraria and photosystems-related heterogeneity of phycobilisome population

Phycobilisomes (PBS) are the major photosynthetic antenna complexes in cyanobacteria and red algae. In the red microalga Galdieria sulphuraria, action spectra measured separately for photosynthetic activities of photosystem I (PSI) and photosystem II (PSII) demonstrate that PBS fraction attributed to PSI is more sensitive to stress conditions and upon nitrogen starvation disappears from the cell earlier than the fraction of PBS coupled to PSII. Preillumination of the cells by actinic far-red light primarily absorbed by PSI caused an increase in the amplitude of the PBS low-temperature fluorescence emission that was accompanied by the decrease in PBS region of the PSI 77 K fluorescence excitation spectrum. Under the same conditions, fluorescence excitation spectrum of PSII remained unchanged. The amplitude of P700 photooxidation in PBS-absorbed light at physiological temperature was found to match the fluorescence changes observed at 77 K. The far-red light adaptations were reversible within 2–5 min. It is suggested that the short-term fluorescence alterations observed in far-red light are triggered by the redox state of P700 and correspond to the temporal detachment of the PBS antenna from the core complexes of PSI. Furthermore, the absence of any change in the 77 K fluorescence excitation cross-section of PSII suggests that light energy transfer from PBS to PSI in G. sulphuraria is direct and does not occur through PSII. Finally, a novel photoprotective role of PBS in red algae is discussed.

Functional Characterization of the Plastidic Phosphate Translocator Gene Family from the Thermo-Acidophilic Red Alga Galdieria sulphuraria Reveals Specific Adaptations of Primary Carbon Partitioning in Green Plants and Red Algae

In chloroplasts of green plants and algae, CO(2) is assimilated into triose-phosphates (TPs); a large part of these TPs is exported to the cytosol by a TP/phosphate translocator (TPT), whereas some is stored in the plastid as starch. Plastidial phosphate translocators have evolved from transport proteins of the host endomembrane system shortly after the origin of chloroplasts by endosymbiosis. The red microalga Galdieria sulphuraria shares three conserved putative orthologous transport proteins with the distantly related seed plants and green algae. However, red algae, in contrast to green plants, store starch in their cytosol, not inside plastids. Hence, due to the lack of a plastidic starch pool, a larger share of recently assimilated CO(2) needs to be exported to the cytosol. We thus hypothesized that red algal transporters have distinct substrate specificity in comparison to their green orthologs. This hypothesis was tested by expression of the red algal genes in yeast (Saccharomyces cerevisiae) and assessment of their substrate specificities and kinetic constants. Indeed, two of the three red algal phosphate translocator candidate orthologs have clearly distinct substrate specificities when compared to their green homologs. GsTPT (for G. sulphuraria TPT) displays very narrow substrate specificity and high affinity; in contrast to green plant TPTs, 3-phosphoglyceric acid is poorly transported and thus not able to serve as a TP/3-phosphoglyceric acid redox shuttle in vivo. Apparently, the specific features of red algal primary carbon metabolism promoted the evolution of a highly efficient export system with high affinities for its substrates. The low-affinity TPT of plants maintains TP levels sufficient for starch biosynthesis inside of chloroplasts, whereas the red algal TPT is optimized for efficient export of TP from the chloroplast.

The genome of the thermoacidophilic red microalga Galdieria sulphuraria encodes a small family of secreted class III peroxidases that might be involved in cell wall modification

We report the identification of a small family of secreted class III plant peroxidases (Prx) from the genome of the unicellular thermoacidophilic red alga Galdieria sulphuraria (Cyanidiaceae). Apart from two class I ascorbate peroxidases and one cytochrome c peroxidase, the red algal genome encodes four class III plant peroxidases, thus complementing the short list of algal cell wall peroxidases (Passardi et al. in Genomics 89:567-579, 2007). We have characterized the family gene structure, analyzed the extracellular space and cell wall fraction of G. sulphuraria for the presence of peroxidase activity and used shotgun proteomics to identify candidate extracellular peroxidases. For a detailed enzymatic characterization, we have purified a secreted peroxidase (GsPrx04) from the cell-free medium using hydrophobic interaction chromatography. The enzyme proved heat and acid-stable and exhibited an apparent molecular mass of 40 kDa. Comparative genomics between endolithically growing G. sulphuraria and a close relative, the obligatory aquatic, cell wall-less Cyanidioschyzon merolae, revealed that class III peroxidases only occur in the terrestrial microalga, thus supporting the key function of these enzymes in the process of land colonization.

EST-analysis of the thermo-acidophilic red microalga Galdieriasulphuraria reveals potential for lipid A biosynthesis and unveils the pathway of carbon export from rhodoplasts

When we think of extremophiles, organisms adapted to extreme environments, prokaryotes come to mind first. However, the unicellular red micro-alga Galdieria sulphuraria (Cyanidiales) is a eukaryote that can represent up to 90% of the biomass in extreme habitats such as hot sulfur springs with pH values of 0-4 and temperatures of up to 56 degrees C. This red alga thrives autotrophically as well as heterotrophically on more than 50 different carbon sources, including a number of rare sugars and sugar alcohols. This biochemical versatility suggests a large repertoire of metabolic enzymes, rivaled by few organisms and a potentially rich source of thermo-stable enzymes for biotechnology. The temperatures under which this organism carries out photosynthesis are at the high end of the range for this process, making G. sulphuraria a valuable model for physical studies on the photosynthetic apparatus. In addition, the gene sequences of this living fossil reveal much about the evolution of modern eukaryotes. Finally, the alga tolerates high concentrations of toxic metal ions such as cadmium, mercury, aluminum, and nickel, suggesting potential application in bioremediation. To begin to explore the unique biology of G. sulphuraria , 5270 expressed sequence tags from two different cDNA libraries have been sequenced and annotated. Particular emphasis has been placed on the reconstruction of metabolic pathways present in this organism. For example, we provide evidence for (i) a complete pathway for lipid A biosynthesis; (ii) export of triose-phosphates from rhodoplasts; (iii) and absence of eukaryotic hexokinases. Sequence data and additional information are available at http://genomics.msu.edu/galdieria.