Unicellular Microalgae Research Articles

Promoting the Growth of Haematococcus lacustris under High Light Intensity through the Combination of Light/Dark Cycle and Light Color

The unicellular microalgae Haematococcus lacustris is an astaxanthin-rich organism that is widely used for commercial cultivation, but its main limitation is its relatively low biomass yield. It is widely accepted that the use of appropriate high light intensity could promote algal growth; however, H. lacustris is very sensitive to high-intensity light, and its growth can be readily arrested by inappropriate illumination. To exploit the growth-promoting benefit of higher light intensities while avoiding growth arrestment, we examined the growth of H. lacustris under high light intensities using various light profiles, including different light colors and light/dark cycles. The results show that light color treatments could not alleviate cellular stress under high light intensities; however, it was interesting to find that red light was favored the most by cells out of all the colors. In terms of the light/dark cycle, the 2/2 h light/dark cycle treatment was shown to lead to the highest specific growth rate, which was 46% higher than that achieved in the control treatment (18 μmol/m2·s light intensity, white light). Therefore, in further experiments, the 2/2 h light/dark cycle with red-light treatment was examined. The results show that this combination enabled a significantly higher specific growth rate, which was 66.5% higher than that achieved in the control treatment (18 μmol/m2·s light intensity, white light). These findings offer new strategies for the optimization of illumination for the growth of H. lacustris and lay the foundations for more reasonable lighting utilization for the cultivation of commercially valuable algal species.

Assessment of Eicosapentaenoic Acid (EPA) Production from Filamentous Microalga Tribonema aequale: From Laboratory to Pilot-Scale Study.

It has long been explored to use EPA-rich unicellular microalgae as a fish oil alternative for production of the high-value omega-3 fatty acid eicosapentaenoic acid (EPA, 20:5, n-3). However, none of the efforts have ever reached commercial success. This study reported a filamentous yellow-green microalga Tribonema aequale that possesses the ability to grow rapidly and synthesize significant amounts of EPA. A series of studies were conducted in a glass column photobioreactor under laboratory culture conditions and in pilot-scale open raceway ponds outdoors. The emphasis was placed on the specific nutrient requirements and the key operational parameters in raceway ponds such as culture depth and mixing regimes. When optimized, T. aequale cells contained 2.9% of EPA (w/w) and reached a very high biomass concentration of 9.8 g L−1 in the glass column photobioreactor. The cellular EPA content was increased further to 3.5% and the areal biomass and EPA productivities of 16.2 g m−2 d−1 and 542.5 mg m−2 d−1, respectively, were obtained from the outdoor pilot-scale open raceway ponds, which were the record high figures reported thus far from microalgae-based EPA production. It was also observed that T. aequale was highly resistant to microbial contamination and easy for harvesting and dewatering, which provide two additional competitive advantages of this filamentous microalga over the unicellular counterparts for potential commercial production of EPA and other derived co-products.

Current advances in recovery and biorefinery of fucoxanthin from Phaeodactylum tricornutum

The growing human population has put significant pressure on the market for food sources and nutritional supplements, necessitating careful consideration of high costs, supply, and long-term sustainability. The same is true for the biorefinery of carotenoids where the high extraction costs significantly raise the end-product costs. Many brown seaweeds and unicellular microalgae contain fucoxanthin, an orange-colored pigment found in their chloroplasts. Because of its several medicinal benefits (anti-cancer, anti-diabetic, anti-oxidant, etc), it is an important carotenoid. As a result, a lot of research is being done on fucoxanthin production and extraction from a variety of macroalgal and microalgal sources. When compared to other microalgae, Phaeodactylum tricornutum, a model diatom, is one of the most abundant producers of fucoxanthin. The focus of this paper is on extracting fucoxanthin from Phaeodactylum tricornutum, with Chaetoceros calcitrans and Isochrysis galbana receiving only a passing mention. The cultivation, harvesting, and drying processes of the aforesaid diatoms are briefly discussed in the upstream processing of fucoxanthin. The final section of this paper discusses various traditional (e.g., maceration extraction, Soxhlet extraction, and steam distillation) and new (e.g., Sc-CO2, ultrasound, microwave, pressured liquid, enzyme, and electric field-based extractions) extraction procedures for fucoxanthin. Finally, the limitations of present extraction strategies are discussed, as well as the potential for cost-effective technologies such as liquid biphasic flotation (LBF) to be used for fucoxanthin recovery.

Eco-technological method for carbon dioxide biosorption and molecular mechanism of the RuBisCO enzyme from unicellular microalga Chlorella vulgaris RDS03: a synergistic approach

Eco-technological method for carbon dioxide biosorption and molecular mechanism of the RuBisCO enzyme from unicellular microalga Chlorella vulgaris RDS03: a synergistic approach

Innovative method to grow the probiotic Lactobacillus reuteri in the omega3-rich microalga Isochrysis galbana

Microalgae are natural sources of valuable bioactive compounds, such as polyunsaturated fatty acids (PUFAs), that show antioxidant, anti-inflammatory, anticancer and antimicrobial activities. The marine microalga Isochrysis galbana (I. galbana) is extremely rich in ω3 PUFAs, mainly eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Probiotics are currently suggested as adjuvant therapy in the management of diseases associated with gut dysbiosis. The Lactobacillus reuteri (L. reuteri), one of the most widely used probiotics, has been shown to produce multiple beneficial effects on host health. The present study aimed to present an innovative method for growing the probiotic L. reuteri in the raw seaweed extracts from I. galbana as an alternative to the conventional medium, under conditions of oxygen deprivation (anaerobiosis). As a result, the microalga I. galbana was shown for the first time to be an excellent culture medium for growing L. reuteri. Furthermore, the gas-chromatography mass-spectrometry analysis showed that the microalga-derived ω3 PUFAs were still available after the fermentation by L. reuteri. Accordingly, the fermented compound (FC), obtained from the growth of L. reuteri in I. galbana in anaerobiosis, was able to significantly reduce the adhesiveness and invasiveness of the harmful adherent-invasive Escherichia coli to intestinal epithelial cells, due to a cooperative effect between L. reuteri and microalgae-released ω3 PUFAs. These findings open new perspectives in the use of unicellular microalgae as growth medium for probiotics and in the production of biofunctional compounds.

A Novel Bifunctional Wax Ester Synthase Involved in Early Triacylglycerol Accumulation in Unicellular Green Microalga Haematococcus pluvialis Under High Light Stress.

The bulk of neutral lipids, including astaxanthin esters and triacylglycerols (TAGs), are accumulated in the green microalga Haematococcus pluvialis under high light (HL) stress. In this study, a novel bifunctional wax ester synthase (WS) gene was cloned from H. pluvialis upon HL stress. The overexpression of HpWS restored the biosynthesis of wax esters and TAGs in neutral lipid-deficient yeast mutant Saccharomyces cerevisiae H1246 fed with C18 alcohol and C18:1/C18:3 fatty acids, respectively. Under HL stress, HpWS was substantially upregulated at the transcript level, prior to that of the type I diacylglycerol:acyl-CoA acyltransferase encoding gene (HpDGAT1). HpDGAT1 is the major TAG synthase in H. pluvialis. In addition, the application of xanthohumol (a DGAT1/2 inhibitor) in the H. pluvialis cells did not completely eliminate the TAG biosynthesis under HL stress at 24 h. These results indicated that HpWS may contribute to the accumulation of TAGs in H. pluvialis at the early stage under HL stress. In addition, the overexpression of HpWS in Chlamydomonas reinhardtii bkt5, which is engineered to produce free astaxanthin, enhanced the production of TAGs and astaxanthin. Our findings broaden the understanding of TAG biosynthesis in microalgae and provide a new molecular target for genetic manipulation in biotechnological applications.

In Situ 3D Bioprinting Living Photosynthetic Scaffolds for Autotrophic Wound Healing.

Three-dimensional (3D) bioprinting has been extensively explored for tissue repair and regeneration, while the insufficient nutrient and oxygen availability in the printed constructs, as well as the lack of adaptive dimensions and shapes, compromises the overall therapeutic efficacy and limits their further application. Herein, inspired by the natural symbiotic relationship between salamanders and algae, we present novel living photosynthetic scaffolds by using an in situ microfluidic-assisted 3D bioprinting strategy for adapting irregular-shaped wounds and promoting their healing. As the oxygenic photosynthesis unicellular microalga (Chlorella pyrenoidosa) was incorporated during 3D printing, the generated scaffolds could produce sustainable oxygen under light illumination, which facilitated the cell proliferation, migration, and differentiation even in hypoxic conditions. Thus, when the living microalgae-laden scaffolds were directly printed into diabetic wounds, they could significantly accelerate the chronic wound closure by alleviating local hypoxia, increasing angiogenesis, and promoting extracellular matrix (ECM) synthesis. These results indicate that the in situ bioprinting of living photosynthetic microalgae offers an effective autotrophic biosystem for promoting wound healing, suggesting a promising therapeutic strategy for diverse tissue engineering applications.

Biosorption of Heavy Metals From Tannery Effluents by Using Green Unicellular Microalga, Tetradesmus obliquus RDRL01

In present study, ecofriendly biological treatment of tannery effluent was performed by using unicellular microalga Tetradesmus obliquus strain RDRL01, isolated from Sipcot, Vellore, Tamil Nadu (India). The microalga reduced heavy metals content viz., chromium, cobalt, nickel, cadmium, lead, zinc and copper by 99.1, 98.2, 97.4, 94.3, 98.3, 96.8 and 96.3%, respectively, after 15 days treatment of tannery effluent. Treated effluent had 3.51 g biomass L-1 and treated effluent biomass contained 330.5 mg carbohydrate g-1 and yielded 13.5 mL bioethanol g-1. The functional groups identified by Fourier transform infrared analysis were carboxylic acids, alkyl, hydroxyl, and carbonyl. Purified bioethanol was confirmed by HPLC, and chromato-gram peak showed the retention time of 5.35 min. Microalgae T. obliquus RDRL01 demonstrated good biosorption efficiency and removed various toxic heavy metal pollutants from tannery effluent.

Critical assessment of the filamentous green microalga Oedocladium carolinianum for astaxanthin and oil production

The filamentous microalga Oedocladium carolinianum that was isolated from soil was found to be capable of producing large amounts of astaxanthin under stress culture conditions. When the culture conditions were optimized, the maximum specific growth rate of 0.37 d−1 and the maximum biomass concentration of 10.12 g L−1 were obtained after 18 days of cultivation in 1-liter glass columns (inner diameter: 5 cm) under laboratory conditions. When subjected to culture conditions of nitrogen starvation and 2 g L−1 NaCl-induced salinity stress, however, the cells produced up to 3.91% w/w astaxanthin and a high astaxanthin productivity of 24.2 mg L−1 d−1 was obtained. Analysis by HPLC-MS revealed that the majority of astaxanthin was in fatty acid esterified forms with a typical molecular ratio of free, monoester and diester astaxanthin of 1:18:81. The biosynthesis of astaxanthin coincided with that of fatty acids, and the total fatty acid content reached 40% w/w or more. The technical feasibility of mass culture of O. carolinianum was tested in a 7000-L inclined thin-layer photobioreactor and a 10,000-L tubular photobioreactor in a greenhouse. This demonstrated that O. carolinianum grew rapidly in a nitrogen replete BG-11 culture medium and the maximum biomass concentrations obtained in the inclined thin-layer photobioreactor and tubular photobioreactor were 3.74 g L−1 and 3.07 g L−1 respectively, resulting in maximum biomass productivities of 276 mg L−1 d−1 and 198 mg L−1 d−1, respectively. Although small populations of a few zooplankton species occurred in the two types of photobioreactors, none grazed on O. carolinianum and they grazed on invading unicellular microalgae instead. It was therefore concluded that O. carolinianum is a promising microalga for sustainable co-production of astaxanthin and fatty acids.

Uptake of Cu2+ by unicellular microalga Chlorella vulgaris from synthetic wastewaters is attenuated by polystyrene microspheres

Aquatic and terrestrial ecosystems are receiving micro- and macro-plastic pollutants alarmingly from various anthropogenic activities. The complications caused by microplastics are largely unexplored and need substantial studies. In the current study, we investigated the repressive effects of negatively and positively charged polystyrene microspheres of two variable sizes (0.05 and 0.5 μm) on functioning of unicellular green microalgae. For the purpose, a pollution-resistant microalgal species was isolated and identified by 18 S rRNA gene sequencing as Chlorella vulgaris. The functioning of the pure-cultured microalgal cells was then assessed in terms of their better metal (Cu2+) uptake potential with and without the provision of PS microspheres. The algal cells up took Cu2+ significantly (90% at 75 mg/L) after 15 days of aerobic incubation. However, positively charged polystyrene microspheres remarkably affected the uptake of Cu2+ and it was comparatively reduced to almost 50%, while negatively charged microspheres couldn't influence the Cu2+ uptake potential of C. vulgaris. In addition, size of the microspheres insignificantly affected the metal uptake potential of the microalgae. Unveiled facts of this investigation will be helpful for designing economical and efficient remedial systems based on the in-situ implication of microalgae.

Exploring the concurrent use of microalgae Coelastrella sp. for electricity generation and dairy wastewater treatment

This paper illustrates the potential of unicellular microalgae in electricity production in concurrence with wastewater treatment. Our study focused on the isolation of microalgal strains from various aquatic sources. Cyclic voltammetry was performed to detect the electrogenic activity and out of 18 algal isolates, six algal strains were screened. Coelastrella sp. from the paddy field revealed the well-defined redox peak and its analyzed EPS production was 13.79 mg/mL which paves way to hydrated biofilm matrix assisting in better electrogenicity. Its potential to remove nutrients in wastewater was assessed under lab-scale race way pond. Significant decrease in N, P, BOD, COD, and TOC was observed. The removal efficiency of NH4-N, NO3-N, P, BOD, COD and TOC were 90.38, 90.24, 66.75, 67.15, 69.44, and 83.51 respectively. To the best of our knowledge, EPS production, electrogenic activity and their utility in waste water treatment are reported for the first time in Coelastrella sp.

Dynamic modelling of microalgae growth under micro-aeration conditions

Microalgae cultivation is a critical prerequisite of biomass and biofuels production, which is conductive to the reduction of fossil energy consumptions, but there are the hydrodynamic challenges. This study proposed a dynamic modelling of microalgae growth using experimental and computational fluid dynamics (CFD) methods. Photosynthesis performances of unicellular microalgae in photobioreactors (PBRs) were investigated. In a regions-dividing airlift PBR-B, flow field and cell-cycle progression of Chlorella vulgaris were studied. It demonstrated adverse effects of the Karman Vortex Street on microalgae exponential growth. High-levels biomass productivity in flow stagnate region emerges. In a PBR-O, cell density and fluorescence intensity of Chlorella vulgaris were elevated by 2.62 % and 46.69 % in the 9th day of cultivation. Therefore, Chlorella vulgaris growth profiles were predicted using this mathematical modelling successfully. It benefits for the retrofits of commercial shear-sensitivity microalgae culture that require aeration in bio-energy industry.

Effect of phytochemical vanillic acid on the growth and lipid accumulation of freshwater microalga Euglena gracilis.

A feasible approach against the low yield of microalgae biomass involves the use of a stimulator for microalgal growth. In this research, vanillic acid present in the hydrolysate of agricultural waste, was applied to the cultivation of unicellular microalga Euglena gracilis. At the optimal dosage of 800mgL-1 vanillic acid, biomass yield at treatment increased 2.08-fold. Correspondingly, the content of chlorophyll a and carotenoids was 3.48 and 2.69 fold than of the control ground, respectively. Increased in cell aspect ratio demonstrated that the alga was more active after vanillic acid treatment. Furthermore, relative lipid and carbohydrate content were analyzed using Fourier transform infrared spectroscopy, the result showed that vanillic acid increased the lipid contentin algal cells without sacrificing biomass, which would be a promising way for future biofuel production.

Проблеми використання мікробіологічних систем у процесі ремісії вуглекислого газу із атмосферного повітря

The initiatives of the Kyoto Protocol, the Paris Convention on Climate Change 2015, the X Prize Prize Fund for Revolutionary Innovation and the announcement by Grand Elon Musk of the invention of the best carbon capture technology with a total budget of $ 100 million suggest that air absorption from carbon. In our opinion, the use of biological photosynthetic crops and technological solutions based on them, which will ensure the absorption of carbon dioxide, is a great prospect, as this process has already changed the Earth's atmosphere in the distant past. In addition, the use of plants will not only absorb carbon dioxide from the atmosphere, but also obtain additional raw materials rich in hydrocarbons and lipids, as well as generate oxygen. This technological solution to the problem of the presence of greenhouse gases establishes the feasibility of using the process of growing unicellular microalgae in photoreactor systems. Energy and technical and economic indicators of functioning of carbon dioxide utilization systems on the basis of standard approaches are determined. The need to create a system of automatic analysis of the state of culture based on the use of artificial intelligence and systems of automatic pattern recognition has been identified. The necessity of integration of the heat accumulator into the structure of the photoreactor for smoothing of daily temperature differences (which in turn minimized energy consumption) and systems of additional lighting and mixing is proved. A number of own designs of photoreactors and further ways of their modernization on use of polymeric films for isolation of working space of photoreactors for the purpose of avoidance of infection of culture are offered.

Enhancement of Pigments Production by Nannochloropsis oculata Cells in Response to Bicarbonate Supply

In this study, the effects of bicarbonate addition on growth and pigment contents of the unicellular microalga Nannochloropsis oculata, were evaluated. N. oculata represents an interesting source of biomolecules widely used for food supplements and nutraceuticals. The bicarbonate was supplemented to microalgae cultures at concentrations of 0, 6, 18, 30, 42 and 60 mM. The cultures supplemented with salt at highest concentrations (42 and 60 mM) showed a significant increase in algal growth, demonstrated by the optical density spread. The intracellular content of pigments such as chlorophyll a and total carotenoids reached the highest values in cells from cultures supplied with bicarbonate. In fact, concentrations of bicarbonate from 30 to 60 mM strongly improved, for a short period of only 72 h, the cellular levels of chlorophylls and carotenoids. These are interesting pigments with commercial applications. The utilization of bicarbonate could represent an interesting sustainable opportunity to improve microalgae cultivation for cellular growth and pigment contents.

Underwater Light Manipulation by the Benthic Diatom Ctenophora pulchella: From PAR Efficient Collection to UVR Screening.

Several species of diatoms, unicellular microalgae which constitute the main component of phytoplankton, are characterized by an impressive photosynthetic efficiency while presenting a noticeable tolerance versus exposure to detrimental UV radiation (UVR). In particular, the growth rate of the araphid diatom Ctenophora pulchella is not significantly affected by harsh treatments with UVR, even in absence of detectable, specific UV-absorbing pigments and even if it is not able to avoid high UV exposure by motility. In this work we applied a multi-disciplinary approach involving numerical computation, photonics, and biological parameters in order to investigate the possible role of the frustule, micro- and nano-patterned silica shell which encloses the cell, in the ability of C. pulchella to efficiently collect photosynthetic active radiation (PAR) and to simultaneously screen the protoplasm from UVR. The characterization of the photonic properties of the frustule has been accompanied by in vivo experiments conducted in water in order to investigate its function as optical coupler between light and plastids.

Regeneration of the Eyespot and Flagellum in Euglena gracilis during Cell Division.

Cell division of unicellular microalgae is a fascinating process of proliferation, at which whole organelles are regenerated and distributed to two new lives. We performed dynamic live cell imaging of Euglena gracilis using optical microscopy to elucidate the mechanisms involved in the regulation of the eyespot and flagellum during cell division and distribution of the organelles into the two daughter cells. Single cells of the wild type (WT) and colorless SM-ZK cells were confined in a microfluidic device, and the appearance of the eyespot (stigma) and emergent flagellum was tracked in sequential video-recorded images obtained by automatic cell tracking and focusing. We examined 12 SM-ZK and 10 WT cells and deduced that the eyespot diminished in size and disappeared at an early stage of cell division and remained undetected for 26–97 min (62 min on average, 22 min in deviation). Subsequently, two small eyespots appeared and were distributed into the two daughter cells. Additionally, the emergent flagellum gradually shortened to zero-length, and two flagella emerged from the anterior ends of the daughter cells. Our observation revealed that the eyespot and flagellum of E. gracilis are degraded once in the cell division, and the carotenoids in the eyespot are also decomposed. Subsequently, the two eyespots/flagella are regenerated for distribution into daughter cells. As a logical conclusion, the two daughter cells generated from a single cell division possess the equivalent organelles and each E. gracilis cell has eternal or non-finite life span. The two newly regenerated eyespot and flagellum grow at different rates and mature at different timings in the two daughter cells, resulting in diverse cell characteristics in E. gracilis.

Approaches and involved principles to control pH/pCO2 stability in algal cultures

Experimental cultures of both microalgae and macroalgae are commonly carried out by phycologists or environmental biologists to look into morphological, physiological, and molecular responses to aquatic environmental changes. However, the species of inorganic carbon in algae cultures is often altered by algal photosynthetic CO2 removal and/or bicarbonate utilization. The pH changes associated with altered carbonate chemistry in cultures impact physiological processes in microalgae and macroalgae even at their exponential growth phases, since extra energy is required to sustain intracellular acid–base homeostasis. Usually, pH increases during light period due to inorganic carbon uptake and utilization for photosynthesis and decreases during dark period because of respiratory CO2 release. Therefore, to obtain relevant data aimed for physiological and/or molecular responses of algae to changed levels of environmental factors, stability of pH/pCO2 in the cultures should be considered and controlled to rule out impacts of carbonate chemistry and pH changes. In this work, principles involved in changing pH processes in algal cultures are mechanistically analyzed and several approaches to control pH and pCO2 are introduced. In order to sustain stability of pH/pCO2, the principles underline the following key points: (1) maintaining the rate of photosynthetic C removal less than or equal to the rate of CO2 dissolution into the cultures which are aerated; or (2) sustaining dilute cultures with very low cell density without aeration, so that photosynthetic C removal is small enough not to cause significant pH/pCO2 changes; or (3) stabilizing the changes in micro-environments surrounding the cells or thallus. To maintain pH drift < 1% in growing typical unicellular microalgae, the recommended cell concentration ranges from 50 × 103 to 200 × 103 mL−1 with aeration (air replacement rate of ca 500–1000 mL L−1 min−1) in semi-continuous cultures of < 1 L, and it ranges from 100 to 5000 cells mL−1 for diatoms and from 100 to 100 × 103 cells mL−1 for coccolithophores in dilute cultures without aeration, respectively. For macroalgae, maintaining the thalli in flowing through- system or in semi-continuous cultures (continuously control algal biomass density) is recommended.

Unicellular microalgae vs. filamentous algae for wastewater treatment and nutrient recovery

Microalgae-based wastewater treatment is a promising platform since it enables nutrient recovery. The use of filamentous algae could improve this technology by reducing harvesting cost and enhancing biomass management. The potential of 12 unicellular and 12 filamentous algae strains was evaluated for the treatment and nutrient recovery of a raw wastewater from a slaughterhouse (RWS). Chlorococcum sp. FAUBA-42 (unicellular) and Stigeoclonium sp. FAUBA-10 (filamentous) were selected due to their biomass production using RWS as the sole nutrient source. Comprehensive analysis of batch and semicontinuous regime was carried out for both strains. Semicontinuous cultures showed three times higher biomass productivity than batch regime. The highest biomass productivity (0.45 g DW L−1 d−1) and photosynthetic quantum yield (1.6 g DW mol photons−1) was reached by the filamentous strain Stigeoclonium FAUBA-10 under 2.5 d HRT. Both strains showed nutrient removal above 92% under 5 d and 2.5 d HRT. Biomass of Stigeoclonium FAUBA-10 can be easily harvested by filtration through mesh screens (<341 μm) which is an operational benefit.

Structural evidence for extracellular silica formation by diatoms

The silica cell wall of diatoms, a widespread group of unicellular microalgae, is an exquisite example for the ability of organisms to finely sculpt minerals under strict biological control. The prevailing paradigm for diatom silicification is that this is invariably an intracellular process, occurring inside specialized silica deposition vesicles that are responsible for silica precipitation and morphogenesis. Here, we study the formation of long silicified extensions that characterize many diatom species. We use cryo-electron tomography to image silica formation in situ, in 3D, and at a nanometer-scale resolution. Remarkably, our data suggest that, contradictory to the ruling paradigm, these intricate structures form outside the cytoplasm. In addition, the formation of these silica extensions is halted at low silicon concentrations that still support the formation of other cell wall elements, further alluding to a different silicification mechanism. The identification of this unconventional strategy expands the suite of mechanisms that diatoms use for silicification.