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All sourcesNASA NTRSDOE OSTI
DOE OSTI2026

Microalgae Commodities Production with a Direct Air Capture Process (Final Technical Report)

This report documents the work performed under Award Number DE-EE0009276, "Microalgae Commodities Production with a Direct Air Capture Process," to advance the technology for large-scale, low-cost microalgae biomass production in the United States. The project's overarching goal was to decouple microalgae production from enriched CO 2 point sources, such power plant flue gases, which otherwise limit the national resource potential for algal biomass.

09 BIOMASS FUELS↗
DOE OSTI2024

Simple and Effective Squash-PCR for Rapid Genotyping of Industrial Microalgae

Microalgae are recognized for their versatility in providing renewable energy, biopharmaceuticals, and nutraceuticals, attributed to their sustainable, renewable, and cost-effective nature. Genetic engineering has proven highly effective in enhancing microalgae production. PCR-based genotyping is the primary method for screening genetically transformed microalgae cells. Recently, we developed a novel PCR method, namely Squash-PCR, and employed it for the molecular analysis of industrially important fungi and yeasts. In this study, we successfully implemented the Squash-PCR technique in 12 industrially significant algae species. This approach offers a quick and reliable means of obtaining DNA templates directly from squashed algal cells, eliminating the need for time-consuming and labor-intensive cultivation and genomic DNA extraction steps. Our results demonstrate the effectiveness of Squash-PCR in detecting and characterizing target genes of interest in 12 different algae species. Overall, this study establishes the Squash-PCR method as a valuable tool for molecular studies in algae, enabling researchers to rapidly screen and manipulate genetic traits in diverse algal species.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Optimized production of a bioactive human recombinant protein from the microalgae Chlamydomonas reinhardtii grown at high density in a fed-batch bioreactor

Microalgae have been identified as an alternative platform to produce high-quality biomass and subsequent bioproducts, such as foods, feeds, nutritional supplements, recombinant proteins, and biofuels. Traditional biotechnological hosts for therapeutic proteins, such as the bacteria Escherichia coli and mammalian CHO cells, have long been established as the dominate platforms, but recent advances have shown that microalgae can potentially serve as an alternative platform. In the present study, we examine the potential of the microalga Chlamydomonas reinhardtii to produce a complex human recombinant protein in a high-density heterotrophic culture. The recombinant human protein, ICAM-1, was targeted for secretion to the extracellular media of the culture from cells grown in a bioreactor using a fed-batch strategy to achieve high cell density. Ultimately, this resulted in a maximum biomass titer of 40 g/L and a recombinant protein titer of 50 mg/L. The algal-produced ICAM-1 protein showed comparable bioactivity to mammalian cell culture produced ICAM-1, as measured using binding assays for its native ligand LFA-1. This work shows that C. reinhardtii is a viable option to produce complex recombinant proteins, with native biological activity, at high concentrations using a fed batch heterotrophic growth strategy.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

Impacts of feeding three strains of microalgae alone or in combination on growth performance, protein metabolism, and meat quality of broiler chickens

Variations in nutrient compositions, especially amino acid (AA) profiles, among microalgal species may enable a superior feeding outcome from a combined than singular supplementation in poultry diets. Therefore, a feeding trial was conducted to compare the effects of three strains of microalgal biomass supplemented alone or in combination to replace 5 % (starter) and 10 % (grower) soybean meal (on weight-to-weight basis) on growth performance, protein metabolism, and meat quality of broiler chickens. Day-old Cornish Cross male chicks (total = 180) were divided into 5 groups (6 cages/treatment, 6 birds/cage) and fed a corn-soybean meal basal diet (BD), BD + H117 (Chlorella sp., H117), BD + C985 (Tetraselmis sp., C985), BD + Nannochloropsis oceanica (NO), and BD + H117 + C985 + NO (Combination). Feeding any of the microalgae diets did not alter growth performance nor meat quality including texture, pH, color, and water holding capacity of breast and thigh meats. However, the breast weight percentages were decreased (P < 0.05) by feeding the C985, NO, and Combination diets. Compared with the BD, the 4 microalgal diets led to higher (P < 0.05) plasma uric acid and protein concentrations at weeks 3 and (or) 6. The mRNA levels of MAFbx, MURF1, FOXO1, and calpastatin in the breast and thigh muscles were altered by the microalgal diets but not those of genes associated with other quality traits. In conclusion, replacing 5 % or 10 % soybean meal with three sources of microalgae in broiler diets decreased breast weights percentage but not absolute weight. Furthermore, feeding chickens with the combination of three microalgae did not restore the breast loss and induced different expressions of genes related to muscle hypertrophy or atrophy.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Transportome remodeling of a symbiotic microalga inside a planktonic host

Abstract Metabolic exchange is one of the foundations of symbiotic associations between organisms and is a driving force in evolution. In the ocean, photosymbiosis between heterotrophic hosts and microalgae is powered by photosynthesis and relies on the transfer of organic carbon to the host (e.g. sugars). Yet, the identity of transferred carbohydrates as well as the molecular mechanisms that drive this exchange remain largely unknown, especially in unicellular photosymbioses that are widespread in the open ocean. Combining genomics, single-holobiont transcriptomics, and environmental metatranscriptomics, we revealed the transportome of the marine microalga Phaeocystis in symbiosis within acantharia, with a focus on sugar transporters. At the genomic level, the sugar transportome of Phaeocystis is comparable to non-symbiotic haptophytes. By contrast, we found significant remodeling of the expression of the transportome in symbiotic microalgae compared to the free-living stage. More particularly, 36% of sugar transporter genes were differentially expressed. Several of them, such as GLUTs, TPTs, and aquaporins, with glucose, triose-phosphate sugars, and glycerol as potential substrates, were upregulated at the holobiont and community level. We also showed that algal sugar transporter genes exhibit distinct temporal expression patterns during the day. This reprogramed transportome indicates that symbiosis has a major impact on sugar fluxes within and outside the algal cell, and highlights the complexity and the dynamics of metabolic exchanges between partners. This study improves our understanding of the molecular players of the metabolic connectivity underlying the ecological success of planktonic photosymbiosis and paves the way for more studies on transporters across photosymbiotic models.

Environmental Sciences & Ecology↗
DOE OSTI2024

Examining Microalgae Growth with Respect to Cell Count and Salinity

In recent years, microalgae have been identified as potential source of biofuel due to their ability to grow abundantly in little time. As photosynthetic organisms, microalgae only need water, light, micronutrients, and carbon dioxide to yield algal biomass. Several applications for this biomass are being studied, such as the extraction of biofuel (Zhang et al. 2022) as well as wastewater treatment (Rude et al. 2022). Two specific microalgae species have been identified as uniquely prolific: Picochlorum celeri, a saltwater species, and Chlorella sorokiniana 1116, a freshwater species. Both species showcased survival in less-than-ideal conditions and showcased hardiness in their respective waters (Krishnan et al. 2021), making them popular specimen to be studied for a variety of applications. This project involved two components, the first component focusing on the development of a growth curve for P. celeri and the second studying varying salt concentration in both species.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Charting the state of GEMs in microalgae: progress, challenges, and innovations

Genome-scale metabolic models (GEMs) provide a systems-level framework for understanding and engineering microalgal metabolism. This review explores the evolution of GEMs in microalgae, highlighting advances in light modeling, automation, and multi-omics integration. Special emphasis is placed on Chlamydomonas reinhardtii as a model species. Limitations of current models, particularly for microalgae, are discussed, alongside promising developments in dynamic modeling and machine learning. Together, these innovations chart a path toward more predictive, adaptable GEMs that can accelerate biotechnological applications of microalgae in sustainable production systems.

Plant Sciences↗
DOE OSTI2024

2023 Billion-Ton Report: An Assessment of U.S. Renewable Carbon Resources - Microalgae

Microalgae is a unique biomass resource that does not need to compete for land and water with other biomass feedstocks because it can be cultivated on low-quality unencumbered land using noncompetitive water types including saline and wastewater. It is included as a complementary resource alongside other biomass feedstocks reported in this study, albeit at higher biomass production costs reflective of more capital-intensive farming operations than typical for terrestrial biomass. Higher biomass costs can be offset by the potential to produce value-added coproducts unique to compositional constituents of microalgae. Relative to the 2016 Billion-Ton Report, this chapter reflects the latest analysis from the 2022 Algae Harmonization Update, which uses the latest parameterized and high-performing saline algal strain, second-generation carbon capture of point-source waste CO 2 , and high-pressure pipeline transport resolved to specific point-source types, saline water sourcing up to 40,000 mg/L total dissolved solids for source and makeup water salinity, blowdown water treatment and recycle, and brine disposal handling. National-scale algal biomass availability potential was calculated at 152 million tons/yr ash-free dry weight (AFDW) (191 million tons/yr dry weight) at an average biomass

09 BIOMASS FUELS↗
DOE OSTI2023

Do betaine lipids replace phosphatidylcholine as fatty acid editing hubs in microalgae?

Acyl editing refers to a deacylation and reacylation cycle on a lipid, which allows for fatty acid desaturation and modification prior to being removed and incorporated into other pools. Acyl editing is an important determinant of glycerolipid synthesis and has been well-characterized in land plants, thus this review begins with an overview of acyl editing in plants. Much less is known about acyl editing in algae, including the extent to which acyl editing impacts lipid synthesis and on which lipid substrate(s) it occurs. This review compares what is known about acyl editing on its major hub phosphatidylcholine (PC) in land plants with the evidence for acyl editing of betaine lipids such as diacylglyceryltrimethylhomoserine (DGTS), the structural analog that replaces PC in several species of microalgae. In land plants, PC is also known to be a major source of fatty acids and diacylglycerol (DAG) for synthesis of the neutral lipid triacylglycerol (TAG). We review the evidence that DGTS contributes substantially to TAG accumulation in algae as a source of fatty acids, but not as a precursor to DAG. We conclude with evidence of acyl editing on other membrane lipid substrates in plants and algae apart from PC or DGTS, and discuss future analyses to elucidate the role of DGTS and other betaine lipids in acyl editing in microalgae.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

An ensemble data assimilation modeling system for operational outdoor microalgae growth forecasting

Microalgae have received increasing attention as a potential feedstock for biofuel or biobased products. Forecasting the microalgae growth is beneficial for managers in planning pond operations and harvesting decisions. This study proposed a biomass forecasting system comprised of the Huesemann Algae Biomass Growth Model (BGM), the Modular Aquatic Simulation System in Two Dimensions (MASS2), ensemble data assimilation (DA), and numerical weather prediction Global Ensemble Forecast System (GEFS) ensemble meteorological forecasts. The novelty of this study is to seek the use of ensemble DA to improve both BGM and MASS2 model initial conditions with the assimilation of biomass and water temperature measurements and consequently improve short-term biomass forecasting skills. This study introduces the theory behind the proposed integrated biomass forecasting system, with an application undertaken in pseudo-real-time in three outdoor ponds cultured with Chlorella sorokiniana in Delhi, California, United States. Results from all three case studies demonstrate that the biomass forecasting system improved the short-term (i.e., 7-day) biomass forecasting skills by about 60% on average, comparing to forecasts without using the ensemble DA method. Given the satisfactory performances achieved in this study, it is probable that the integrated BGM-MASS2-DA forecasting system can be used operationally to inform managers in making pond operation and harvesting planning decisions.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

An attached microalgae platform for recycling phosphorus through biologically mediated fertilizer formation and biomass cultivation

Nutrient management is a global challenge for protecting water bodies from eutrophication and for retaining and sustainably recycling phosphorus within the biosphere. This challenge is especially important for water resource recovery facilities (WRRFs) in jurisdictions that limit nutrient loads in plant effluent. A microalgae-based biofilm platform has been designed, constructed, and tested to remove phosphorus and nitrogen from anaerobic digester (AD) effluent filtrate through cultivating biomass and inducing the precipitation of a mineral called struvite (NH₄MgPO₄ · 6H₂O) using a rotating algae biofilm reactor (RABR). RABRs function by rotating a growth substratum through nutrient rich water and then into the atmosphere, immersing microalgae in both sunlight and water. Photosynthesis is utilized in RABR operation to enhance struvite formation by increasing the pH value within the biofilm through the uptake of carbon dioxide from solution. Measurements of pH trended higher with depth through the biofilm when exposed to light confirming the function of photosynthesis in increasing pH and, as a consequence, struvite formation. In addition, reducing RPM allows more time for water to evaporate through exposure of the biofilm to the atmosphere and provides a management option to exceed struvite solubility product. Struvite precipitation was predicted based on chemical analysis and MINTEQ modeling of ADE, was confirmed using Scanning Electron Microscopy and Energy Dispersive X-ray Spectroscopy, and quantified by determining ash content. Increase in pH within the biofilm was confirmed as photosynthetic photon flux density increased. While struvite precipitation and removal from wastewater is conventionally accomplished through physicochemical methods, this research is the first report of struvite enhanced formation through algae biofilm-based photosynthesis processes.

09 BIOMASS FUELS↗
DOE OSTI2025

Ultrastructure of the Endoplasmic Reticulum in Eukaryotic Microalgae

ABSTRACT The endoplasmic reticulum (ER) is a large and highly dynamic component of the eukaryotic endomembrane system. In eukaryotic microalgae, it plays six distinct roles: (1) It envelopes the chromatin to form thenucleus. (2) It forms cisternae in the cytoplasm, some of which scaffold the synthesis of proteins destined for incorporation into membranes or for secretion. (3) It associates withGolgicisternae to scaffold the synthesis of glycosylated proteins. (4) It associates with theplasma membraneto mediate the synthesis and secretion of hydrophobic molecules. (5) It mediates the synthesis of cytoplasmiclipid bodies. (6) In lineages harboring complex plastids of red algal ancestry, it forms thechloroplast ER, which envelops the primary chloroplast envelope. In this review, these systems are illustrated using the quick‐freeze deep‐etch electron microscopy (QFDEEM) technique, which lifts up the topological configurations adopted by this gossamer system. A key finding is that in all the complex microalgae examined except dinoflagellates, the inner nuclear envelope membrane associates directly with the plastid‐contiguous membrane of the chloroplast ER at foci designated as chloroplast‐nuclear junctions. These junctions may play a role in regulating the maintenance and physiology of the complex organelles.

Microbiology↗
DOE OSTI2022

Genetic compensation of triacylglycerol biosynthesis in the green microalga Chlamydomonas reinhardtii

Genetic compensation has been proposed to explain phenotypic differences between gene knockouts and knockdowns in several metazoan and plant model systems. With the rapid development of reverse genetic tools such as CRISPR/Cas9 and RNAi in microalgae, it is increasingly important to assess whether genetic compensation affects the phenotype of engineered algal mutants. While exploring triacylglycerol (TAG) biosynthesis pathways in the model alga Chlamydomonas reinhardtii, it was discovered that knockout of certain genes catalyzing rate-limiting steps of TAG biosynthesis, type-2 diacylglycerol acyltransferase genes (DGTTs), triggered genetic compensation under abiotic stress conditions. Genetic compensation of a DGTT1 null mutation by a related PDAT gene was observed regardless of the strain background or mutagenesis approach, for example, CRISPR/Cas 9 or insertional mutagenesis. However, no compensation was found in the PDAT knockout mutant. The effect of PDAT knockout was evaluated in a Δvtc1 mutant, in which PDAT was upregulated under stress, resulting in a 90% increase in TAG content. Knockout of PDAT in the Δvtc1 background induced a 12.8-fold upregulation of DGTT1 and a 272.3% increase in TAG content in Δvtc1/pdat1 cells, while remaining viable. These data suggest that genetic compensation contributes to the genetic robustness of microalgal TAG biosynthetic pathways, maintaining lipid and redox homeostasis in the knockout mutants under abiotic stress. This work demonstrates examples of genetic compensation in microalgae, implies the physiological relevance of genetic compensation in TAG biosynthesis under stress, and provides guidance for future genetic engineering and mutant characterization efforts.

09 BIOMASS FUELS↗
DOE OSTI2024

Utilization of microalgae for agricultural runoff remediation and sustainable biofuel production through an integrated biorefinery approach

Generally wastewater such agricultural runoff is considered a nuisance; however, it could be harnessed as a potential source of nutrients like nitrates and phosphates in integrated biorefinery context. In the current study, microalgae Chlorella sp. S5 was used for bioremediation of agricultural runoff and the leftover algal biomass was used as a potential source for production of biofuels in an integrated biorefinery context. The microalgae Chlorella sp. S5 was cultivated on Blue Green (BG 11) medium and a comprehensive optimization of different parameters including phosphates, nitrates, and pH was carried out to acquire maximum algal biomass enriched with high lipids content. Dry biomass was quantified using the solvent extraction technique, while the identification of nitrates and phosphates in agricultural runoff was carried out using commercial kits. The algal extracted lipids (oils) were employed in enzymatic trans-esterification for biodiesel production using whole-cell biomass of Bacillus subtilis Q4 MZ841642. The resultant fatty acid methyl esters (FAMEs) were analyzed using Fourier transform infrared (FTIR) spectroscopy and gas chromatography coupled with mass spectrometry (GC–MS). Subsequently, both the intact algal biomass and its lipid-depleted algal biomass were used for biogas production within a batch anaerobic digestion setup. Interestingly, Chlorella sp. S5 demonstrated a substantial reduction of 95% in nitrate and 91% in phosphate from agricultural runoff. The biodiesel derived from algal biomass exhibited a noteworthy total FAME content of 98.2%, meeting the quality standards set by American Society for Testing and Materials (ASTM) and European union (EU) standards. Furthermore, the biomethane yields obtained from whole biomass and lipid-depleted biomass were 330.34 NmL/g VS added and 364.34 NmL/g VS added , respectively. In conclusion, the findings underscore the potent utility of Chlorella sp. S5 as a multi-faceted resource, proficiently employed in a sequential cascade for treating agricultural runoff, producing biodiesel, and generating biogas within the integrated biorefinery concept.

09 BIOMASS FUELS↗
DOE OSTI2022

Convergent evolution and horizontal gene transfer in Arctic Ocean microalgae

Microbial communities in the world ocean are affected strongly by oceanic circulation, creating characteristic marine biomes. The high connectivity of most of the ocean makes it difficult to disentangle selective retention of colonizing genotypes (with traits suited to biome specific conditions) from evolutionary selection, which would act on founder genotypes over time. The Arctic Ocean is exceptional with limited exchange with other oceans and ice covered since the last ice age. To test whether Arctic microalgal lineages evolved apart from algae in the global ocean, we sequenced four lineages of microalgae isolated from Arctic waters and sea ice. Here we show convergent evolution and highlight geographically limited HGT as an ecological adaptive force in the form of PFAM complements and horizontal acquisition of key adaptive genes. Notably, ice-binding proteins were acquired and horizontally transferred among Arctic strains. A comparison with Tara Oceans metagenomes and metatranscriptomes confirmed mostly Arctic distributions of these IBPs. The phylogeny of Arctic-specific genes indicated that these events were independent of bacterial-sourced HGTs in Antarctic Southern Ocean microalgae.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Native Microalgae-Bacteria Consortia: A Sustainable Approach for Effective Urban Wastewater Bioremediation and Disinfection

Urban wastewater is a significant by-product of human activities. Conventional urban wastewater treatment plants have limitations in their treatment, mainly concerning the low removal efficiency of conventional and emerging contaminants. Discharged wastewater also contains harmful microorganisms, posing risks to public health, especially by spreading antibiotic-resistant bacteria and genes. Therefore, this study assesses the potential of a native microalgae-bacteria system (MBS) for urban wastewater bioremediation and disinfection, targeting NH4+-N and PO43−-P removal, coliform reduction, and antibiotic resistance gene mitigation. The MBS showed promising results, including a high specific growth rate (0.651 ± 0.155 d−1) and a significant average removal rate of NH4+-N and PO43−-P (9.05 ± 1.24 mg L−1 d−1 and 0.79 ± 0.06 mg L−1 d−1, respectively). Microalgae-induced pH increase rapidly reduces coliforms (r > 0.9), including Escherichia coli, within 3 to 6 days. Notably, the prevalence of intI1 and the antibiotic resistance genes sul1 and blaTEM are significantly diminished, presenting the MBS as a sustainable approach for tertiary wastewater treatment to combat eutrophication and reduce waterborne disease risks and antibiotic resistance spread.

Sousa, Joana F.↗
NASA NTRSDate not supplied

Space Algae: Understanding the Genomic Impacts on Microalgae After Growth in the International Space Station

Plants and microbes can be used for biological support of crewed space missions. The radiation and microgravity environment of spaceflight is expected to increase genetic mutation of all organisms. It is essential to understand how spaceflight impacts mutation rates in photosynthetic organisms to enable appropriate countermeasures and ensure productivity during long duration and deep space missions. The Space Algae flight experiments to the International Space Station (ISS) are studying the genomic stability of microalgae that could potentially be used in biological life support systems. Space Algae-1 grew ultraviolet light mutagenized Chlamydomonas reinhardtii in the VEGGIE plant growth chamber for approximately 40 mitotic generations over one month on the ISS. Whole genome sequencing from pooled cell samples every 10 generations revealed that spaceflight cultures had an ~50% increase in DNA polymorphisms relative to ground controls. These mutations had a novel base substitution signature and suggested a risk that microalgae may be unstable for long-term production in space. Space Algae-2 is focusing on the edible cyanobacterium Arthrospira platensis, commonly known as Spirulina. This experiment seeks to grow serial cultures to allow the organism to evolve in long-term spaceflight. Biological responses of the cells to spaceflight will be assessed with multi-omics analyses to determine mutation load, gene/protein expression, metabolic/nutritional composition, and cell morphology.

Algae↗
NASA NTRSDate not supplied

Space Algae: Understanding the Genomic Impacts on Microalgae After Growth in the International Space Station

Plants and microbes can be used for biological support of crewed space missions. The radiation and microgravity environment of spaceflight is expected to increase genetic mutation of all organisms. It is essential to understand how spaceflight impacts mutation rates in photosynthetic organisms to enable appropriate countermeasures and ensure productivity during long duration and deep space missions. The Space Algae flight experiments to the International Space Station (ISS) are studying the genomic stability of microalgae that could potentially be used in biological life support systems. Space Algae-1 grew ultraviolet light mutagenized Chlamydomonas reinhardtii in the VEGGIE plant growth chamber for approximately 40 mitotic generations over one month on the ISS. Whole genome sequencing from pooled cell samples every 10 generations revealed that spaceflight cultures had an ~50% increase in DNA polymorphisms relative to ground controls. These mutations had a novel base substitution signature and suggested a risk that microalgae may be unstable for long-term production in space. Space Algae-2 is focusing on the edible cyanobacterium Arthrospira platensis, commonly known as Spirulina. This experiment seeks to grow serial cultures to allow the organism to evolve in long-term spaceflight. Biological responses of the cells to spaceflight will be assessed with multi-omics analyses to determine mutation load, gene/protein expression, metabolic/nutritional composition, and cell morphology.

Algae↗