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Two novel Patescibacteria: Phycocordibacter aenigmaticus gen. nov. sp. nov. and Minusculum obligatum gen. nov. sp. nov., both associated with microalgae optimized for carbon dioxide sequestration from flue gas

The functional roles of bacterial symbionts associated with microalgae remain understudied despite the importance of microalgae in biotechnology and environmental microbiology. 16S rRNA gene sequencing was conducted to analyze bacterial communities associated with two microalgae optimized for growth with flue gas containing 5%–10% CO 2 . Two dominant bacteria with no taxonomic classification beyond the class level (Paceibacteria) were discovered repeatedly in the most productive algal cultures. Long-read metagenomic sequencing was conducted to yield high-quality metagenomes, from which two novel species were discovered under the Seqcode (seqco.de/r:ywe1blo2), Phycocordibacter aenigmaticus gen. nov. sp. nov. and Minusculum obligatum gen. nov. sp. nov. The genus Phycocordibacter gen. nov. was proposed as the nomenclatural type of the family Phycocordibacteraceae fam. nov. and the order Phycocordibacterales ord. nov. Both bacteria possessed features typical of Patescibacteria such as reduced genomes (<800 kbp), lack of complete glycolysis and tricarboxylic acid (TCA) cycle pathways, and inability to synthesize amino acids. Instead, they rely on the reductive pentose phosphate pathway (Calvin cycle) for essential biosynthesis and redox balance. P. aenigmaticus may also rely on elemental sulfur oxidation (sdo), partial nitrite reduction (nirK), and sulfur-related amino acid metabolism (SAMe → SAH). Both bacteria were found in high relative abundance in cultures of Tetradesmus obliquus HTB1 (freshwater) and Nannochloropsis oceanica IMET1 (marine), suggesting a tight association with microalgae in various environments. The absence of full metabolic pathways for energy production suggests extreme metabolic limitations and obligate symbiosis, most likely with other bacteria associated with the microalgae.

54 ENVIRONMENTAL SCIENCES↗

Induction of D-xylose uptake and expression of NAD(P)H-linked xylose reductase and NADP + -linked xylitol dehydrogenase in the oleaginous microalga Chlorella sorokiniana

The heterotrophic and mixotrophic culture of oleaginous microalgae is a promising process to produce biofuel feedstock due to the advantage of fast growth. Various organic carbons have been explored for this application. However, despite being one of the most abundant and economical sugar resources in nature, D-xylose has never been demonstrated as a carbon source for wild-type microalgae. The purpose of the present work was to identify the feasibility of D-xylose utilization by the oleaginous microalga Chlorella sorokiniana. The sugar uptake kinetic analysis was performed with 14 C-labeled sugars and the data showed that the D-glucose induced algal cells (the alga was heterotrophically grown on D-glucose and then harvested as D-glucose induced cells) exhibited a remarkably increased D-xylose uptake rate. The maximum D-xylose transport rate was 3.8 nmol min -1 mg -1 dry cell weight (DCW) with K m value of 6.8 mM. D-xylose uptake was suppressed in the presence of D-glucose, D-galactose and D-fructose but not L-arabinose and D-ribose. The uptake of D-xylose activated the related metabolic pathway, and the activities of a NAD(P)H-linked xylose reductase (XR) and a unique NADP + -linked xylitol dehydrogenase (XDH) were detected in C. sorokiniana. Compared with the culture in the dark, the consumption of D-xylose increased 2 fold under light but decreased to the same level with addition of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), indicating that extra chemical energy from the light-dependent reaction contributed the catabolism of D-xylose for C. sorokiniana. An inducible D-xylose transportation system and a related metabolic pathway were discovered for microalga for the first time. The transportation of D-xylose across the cell membrane of C. sorokiniana could be realized by an inducible hexose symporter. The uptake of D-xylose subsequently activated the expression of key catalytic enzymes that enabled D-xylose entering central metabolism. Results of this research are useful to better understand the D-xylose metabolic pathway in the microalga C. sorokiniana and provide a target for genetic engineering to improve D-xylose utilization for microalgal lipid production.

09 BIOMASS FUELS↗

Defatted Biomass of the Microalga, Desmodesmus sp., Can Replace Fishmeal in the Feeds for Atlantic salmon

Microalgal biomass is a potential feed ingredient that can replace fishmeal and ensure sustainability standards in aquaculture. To understand the efficacy of the defatted biomass from the marine microalga, Desmodesmus sp. a 70-day feeding study was performed with Atlantic salmon (Salmo salar) smolts. Three groups of fish (av. wt. 167 g) were offered either a control feed (without the microalga) or the microalga-containing (10/20%) feeds. At the end of the feeding period, the growth indices (condition factor, specific growth rate) and survival of the microalga-fed fish were not significantly different from the respective values of the control fish, but the feed conversion ratios were inferior. The proximate composition of the whole body of salmon from the three groups did not vary significantly. Compared to the control fish, the 10% alga-fed fish had lower lipid content in their filet. The protein and lipid digestibility in the three feeds did not differ significantly, but the digestibility of energy in the 10% alga-feed was significantly lower than that of the control feed. Furthermore, comparison of the distal intestinal proteome of Atlantic salmon revealed that the expressions of Alpha-2-HS-glycoprotein-like (Ahsg), Myosin-11 isoform X1 (My11) and Dihydrolipoyl dehydrogenase, mitochondrial-like (Dld) were altered by the microalgal feeding. Examination of the physiological status of the fish based on the serum antioxidant capacities did not reveal any alga-feed-related differences. Moreover, the expression of the selected immune and inflammatory marker genes and the micromorphological observations did not indicate any aberration in the intestinal health of the microalga-fed fish. It is possible to include 20% of defatted Desmodesmus sp. in the feeds of Atlantic salmon.

59 BASIC BIOLOGICAL SCIENCES↗

Isolation of phosphorus-hyperaccumulating microalgae from revolving algal biofilm (RAB) wastewater treatment systems

Excess phosphorus (P) in wastewater effluent poses a serious threat to aquatic ecosystems and can spur harmful algal blooms. Revolving algal biofilm (RAB) systems are an emerging technology to recover P from wastewater before discharge into aquatic ecosystems. In RAB systems, a community of microalgae take up and store wastewater P as polyphosphate as they grow in a partially submerged revolving biofilm, which may then be harvested and dried for use as fertilizer in lieu of mined phosphate rock. In this work, we isolated and characterized a total of 101 microalgae strains from active RAB systems across the US Midwest, including 82 green algae, 9 diatoms, and 10 cyanobacteria. Strains were identified by microscopy and 16S/18S ribosomal DNA sequencing, cryopreserved, and screened for elevated P content (as polyphosphate). Seven isolated strains possessed at least 50% more polyphosphate by cell dry weight than a microalgae consortium from a RAB system, with the top strain accumulating nearly threefold more polyphosphate. These top P-hyperaccumulating strains include the green alga Chlamydomonas pulvinata TCF-48 g and the diatoms Eolimna minima TCF-3d and Craticula molestiformis TCF-8d, possessing 11.4, 12.7, and 14.0% polyphosphate by cell dry weight, respectively. As a preliminary test of strain application for recovering P, Chlamydomonas pulvinata TCF-48 g was reinoculated into a bench-scale RAB system containing Bold basal medium. The strain successfully recolonized the system and recovered twofold more P from the medium than a microalgae consortium from a RAB system treating municipal wastewater. These isolated P-hyperaccumulating microalgae may have broad applications in resource recovery from various waste streams, including improving P removal from wastewater.

54 ENVIRONMENTAL SCIENCES↗

Considerations for the Design of a Gas Transport System for Co-location of Microalgae Cultivation with CO 2 Sources

Co-locating microalgae cultivation facilities with sources of waste CO 2 may present opportunities for cost savings that could benefit the algal biomass industry and industries or utilities that have incentives to manage carbon emissions. However, the cost savings have not been quantified. We compare the cost of utilizing CO 2 from flue gas transported to 405-ha, base-case, open-pond microalgae facilities to the commercial purchase price of CO 2 . Sources of CO 2 include coal- and natural gas-fired power plants and ethanol, ammonia, and cement production plants in the United States. The transport of CO 2 -containing gases to the microalgae cultivation facility requires infrastructure and electricity. This engineering analysis explores the parameters that affect the infrastructure and transport distance over which emitted flue gases can be cost-effectively transported to microalgae cultivation facilities, compared to the cost of commercially purchased CO 2 (i.e., the break-even distance). Parameters that are varied include size of facility pond area, productivity, and daily duration of waste CO 2 emissions that are transported to algae. Under our assumptions and under all cases in this study, cost savings can be achieved by co-locating microalgae facilities near waste CO 2 sources. The break-even distance, which is estimated here to range from under 2 to 80 km, depends primarily on the concentration of CO 2 in flue gas, algae productivity, facility size, and pipeline system design considerations. Greater break-even distances are simulated if electricity cost is minimized, rather than if capital cost is minimized.

09 BIOMASS FUELS↗

Is Genetic Engineering a Route to Enhance Microalgae-Mediated Bioremediation of Heavy Metal-Containing Effluents?

Contamination of the biosphere by heavy metals has been rising, due to accelerated anthropogenic activities, and is nowadays, a matter of serious global concern. Removal of such inorganic pollutants from aquatic environments via biological processes has earned great popularity, for its cost-effectiveness and high efficiency, compared to conventional physicochemical methods. Among candidate organisms, microalgae offer several competitive advantages; phycoremediation has even been claimed as the next generation of wastewater treatment technologies. Furthermore, integration of microalgae-mediated wastewater treatment and bioenergy production adds favorably to the economic feasibility of the former process—with energy security coming along with environmental sustainability. However, poor biomass productivity under abiotic stress conditions has hindered the large-scale deployment of microalgae. Recent advances encompassing molecular tools for genome editing, together with the advent of multiomics technologies and computational approaches, have permitted the design of tailor-made microalgal cell factories, which encompass multiple beneficial traits, while circumventing those associated with the bioaccumulation of unfavorable chemicals. Previous studies unfolded several routes through which genetic engineering-mediated improvements appear feasible (encompassing sequestration/uptake capacity and specificity for heavy metals); they can be categorized as metal transportation, chelation, or biotransformation, with regulation of metal- and oxidative stress response, as well as cell surface engineering playing a crucial role therein. This review covers the state-of-the-art metal stress mitigation mechanisms prevalent in microalgae, and discusses putative and tested metabolic engineering approaches, aimed at further improvement of those biological processes. Finally, current research gaps and future prospects arising from use of transgenic microalgae for heavy metal phycoremediation are reviewed.

Ranjbar, Saeed (ORCID:0000000256518605)↗

Microalgae Commodities from Coal-Fired Power Plant Flue Gas CO 2 (Draft Final Report, October 1, 2015 to June 30, 2018)

The Orlando Utilities Commission (OUC) operates an approximately 900-MWe coal fired-power plant at Stanton Energy Center (SEC), which was the site for this project to develop an integrated process for microalgae utilization of flue gas CO 2 for the production of biofuels and animal feeds. Microalgae have both potentially high yields (tons biomass/acre-year) and can be used for biofuels or high value animal feeds. The high biomass yields of microalgae cultures allows efficient use of land and water resources. They can also use wastewaters, recycle fertilizers, and have lower overall greenhouse gas emissions than conventional biofuels or animal feeds. However, many technical issues and technology gaps related to large-scale algae biomass production still must be resolved before such a process can be scaled-up. In this project, techno-economic analyses (TEAs) and life cycle assessments (LCAs) were carried out, projecting the costs and impacts of large-scale algae biomass cultivation for production of biofuels and animal feeds using OUC-SEC flue gas CO 2 . In addition, 3.5-m 2 (~ 40-ft 2 ) experimental raceway ponds were used for the cultivation of microalgae at OUC-SEC and also at the University of Florida. A large-scale algae farm was designed for a 1,300-acre site (~500-ha) located near the OUC-SEC plant, consisting of 400 hectares (1,000 acres) of paddle wheel mixed raceway ponds (each 4 ha), receiving flue gas piped approximately 3 km (2 miles) from the SEC plant. In the first year of this project (Budget Period 1, BP1), the TEA and LCA studies focused on the production of biogas produced from algae biomass and its use to replace coal at the power plant. However, due to the resulting very high cost of CO 2 mitigation, the process was modified to produce compressed natural gas (‘renewable natural gas’, RNG) for transportation fuel from the biogas, in conjunction with wastewater treatment by the algae cultures. During Budget Period 2 (BP2), the TEA and LCA studies focused on the production of animal feeds by the algae farm, using fresh water and agricultural fertilizers. In this case, CO 2 mitigation costs would be entirely covered if the algal biomass could be sold at between a two to three-fold premium above soybean feed, based on a higher nutritional value for the algal biomass. In support of the above analyses, microalgae cultivation experiments were carried out for over two years during this project in small 3.5-m 2 ponds at both SEC and the University of Florida. The most important results were the demonstration at SEC of the cultivation, on flue gas CO 2 , of filamentous algae biomass, which can be harvested at much lower cost than processes required for the colloidal algae species, as currently used by almost all algal mass culture biofuels and animal feeds projects. The yield of biogas production from the algal biomass was investigated by the University of Florida, and the value of the algal biomass as animal feed was projected based on biochemical analysis of the SEC ad UF biomasses. The conclusion from TEA/LCA carried out for the biogas case in BP1 was that a 400-ha algae system producing biomethane for co-firing at OUC-SEC could generate 27,500 MWh/yr of renewable power, offsetting 28,400 Mg CO 2eq /yr, which amounted to less than 1% of the CO 2 emitted by the SEC plant. (Mg = metric ton = 1.10 short tons). The overall cost was projected at $820/Mg of CO 2eq reduction, making this process unfeasible under any possible scenario. More favorable economics were projected when the biogas was converted to higher value transportation fuel, RNG, which also would benefit from renewable fuel (RINs) and possibly other credits. However, only by combining RNG production with municipal wastewater treatment in the algal ponds would CO 2 utilization result in no net cost or even be economically beneficial (i.e., generate income to the power plant). However, in that case the amount of CO 2 utilization would be even lower than that in the biogas case, because a large fraction of the carbon required would be supplied from the wastewater and CO 2 recovered during biogas upgrading to RNG. In brief, use of power plant CO 2 emissions for algal biofuels production was not favorable, even with optimistic assumptions about biomass productivity and biogas yields. For the animal feed case, developed during BP2, algal biomass would replace soybean feeds. The same scale 400-ha algal production plant was projected to produce 26,280 Mg per year of animal feed with a utilization of 45,300 Mg flue gas CO 2 taken up by the algal biomass. However, the process would result in a projected reduction of 112,500 Mg CO 2eq emissions per year, or roughly 1.9% of OUC-SEC emissions, mainly due to the large CO 2eq emissions from conventional soybean production. A major factor in this analysis were greenhouse gas emissions from land use changes. The cost of CO 2 utilization was projected at $116/Mg if the biomass sold for the same price as soybean meal, but if sold as a premium feed (e.g., based on high omega-3 fatty acid and carotenoid content), the power plant could actually charge for the CO 2 utilized. These economic projections are sensitive to assumptions underlying the TEA and LCA models, including biomass composition and the financial models used to project CAPEX and OPEX. Further development of an algal animal feed production using coal-fired power plant flue gas CO 2 is recommended based on the TEA and LCA, as well as experimental results. The required research and development is detailed in a technology gap analysis (Section 11), which focuses on the demonstrations of the cultivation of higher value filamentous algal biomass at OUC-SEC.

01 COAL, LIGNITE, AND PEAT↗

Comparative Study of Flash and Acid Hydrolysis of Microalgae (Scenedesmus sp.) for the Recovery of Biochemicals and Production of Porous Biocarbon Nanosheets

An integrated biorefinery concept is a novel and economical process intensification methodology for efficient utilization of biomass components. In this research, microalgae (Scenedesmus sp.) slurry with biomass concentration of 8.5 wt.% was parallelly fractionated using two techniques: 'flash hydrolysis (FH)' and 'acid hydrolysis (AH)'. FH was performed at 240 degrees C with a residence time of 10 +/- 2 s in a continuous flow reactor, whereas AH was performed at 155 degrees C and reaction time of 15 min in a batch reactor. About 63% of microalgal biomass was solubilized in liquid hydrolysate through both FH and AH. However, AH had an advantage over FH in recovering microalgae proteins and carbohydrates. FAME recovery through solvent extraction from FH and AH derived wet solids (insoluble microalgae) was 40 and 63%, respectively. Finally, the FH- and AH-derived post extraction solid residue was thermally activated using K2CO3 to produce highly microporous biocarbon nanosheets with BET surface areas of 712 and 1289 m2 g-1, respectively. Overall, an integrated process was developed using two potential hydrolysis techniques to maximize utilization of microalgae components.

BASIC BIOLOGICAL SCIENCES,BIOMASS FUELS↗

Establishment of stable synthetic mutualism without co-evolution between microalgae and bacteria demonstrated by mutual transfer of metabolites (NanoSIMS isotopic imaging) and persistent physical association (Fluorescent in situ hybridization)

The demonstration of a mutualistic interaction requires evidence of benefits for both partners as well as stability of the association over multiple generations. A synthetic mutualism between the freshwater microalga Chlorella sorokiniana and the soil-derived plant growth-promoting bacterium (PGPB) Azospirillum brasilense was created when both microorganisms were co-immobilized in alginate beads. Using stable isotope enrichment experiments followed by high-resolution secondary ion mass spectrometry (SIMS) imaging of single cells, we demonstrated transfer of carbon and nitrogen compounds between the two partners. Further, using fluorescent in situ hybridization (FISH), mechanical disruption and scanning electron microscopy, we demonstrated the stability of their physical association for a period of 10 days after the aggregated cells were released from the beads. The bacteria significantly enhanced the growth of the microalgae while the microalgae supported growth of the bacteria in a medium where it could not otherwise grow. In conclusion, we propose that this microalga-bacterium association is a true synthetic mutualism independent of co-evolution. (155 words).

59 BASIC BIOLOGICAL SCIENCES↗

Recent advancements in the genetic engineering of microalgae

The development of more sustainable food, feed, and bio-products is critical to mitigating the environmental stresses facing our world today. Algae, which includes seaweeds, eukaryotic microalgae, and cyanobacteria, are a promising platform to achieving this, as they have low energy and space requirements, are safe for human and animal consumption, and can be manipulated to produce a diversity of valuable bioproducts. This review focuses on microalgae, both eukaryotic and cyanobacteria. In the past, addressing the major challenges of bringing microalgal production systems to an economically viable scale only had a relatively small genetic toolset to work with, in comparison to other microbial systems such as bacteria and yeast. Expanding the molecular tools available for genetic engineering of microalgae will lead to higher product yields, and accelerate the development of new microalgal bioproducts for commercial applications, thereby supporting the shift towards more environmentally friendly products. In this review, we highlight significant advances from recent years on the design of microalgal expression vectors, discovery of genetic regulatory elements (promoters and transcription factors), optimization of transformation methods, and development of new strain improvement techniques, all aimed at advancing microalgae to become a more efficient biomanufacturing platform. We then discuss how these tools have been applied to improving recombinant protein production, and to enhance metabolic pathway engineering.

09 BIOMASS FUELS↗

A systematic multicriteria-based approach to support product portfolio selection in microalgae biorefineries

Here this work proposes and applies a sequential approach of objective methods to aid the decision-making process for the deployment of microalgae biorefineries. The strategy combines Multicriteria Decision Analysis (MCDA) and weight assignment methods to simultaneously consider technical, economic, and environmental criteria to (1) outrank the best bioproduct options from different biomass fractions present in microalgae biomass at different ratios (namely carbohydrates, lipids, and protein) and (2) define the most suitable biorefining pathways associated with specific pairings of microalgae strains and cultivation conditions. The first part of the assessment identified succinic acid, acrylic acid, and citric acid as the top-ranked bioproducts from carbohydrates, polyurethane from lipids, and thermoplastic extrusion co-feed from protein. The second step of the analysis determined that, when production of a hydrocarbon fuel is desired, the compositional profile of a strain is paramount in defining the biorefining setup that should be pursued. In summary, microalgae lipids should be sent to the production of hydrocarbon fuels if the ratio between neutral lipids and fermentable carbohydrates is higher than roughly 1, with carbohydrates and protein being converted to the higher-value products noted above. Finally, this result was corroborated through process simulations, which indicated superior economic and environmental metrics when strains are paired with suitable conversion pathways identified through MCDA based on their compositional profiles. The outcomes of this work provide clear, objective, guidelines for establishing the best biorefining approach for a large suite of biochemical compositions as a screening method prior to employing detailed process simulations alongside rigorous techno-economic and life-cycle assessments.

09 BIOMASS FUELS↗

A disposable bulk-acoustic-wave microalga trapping device for real-time water monitoring

Recently, there has been an increase in the use of algal physiology as a convenient and useful indicator for monitoring of water quality in aquatic ecosystems. However, current methods for monitoring algal physiology are high-cost, power intensive, and have limited sensitivity for practical samples in which algal concentration is low. In order to alleviate these problems, we developed a simple-to-fabricate acoustophoretic particle trapping device that can effectively enrich various types of microalgae such as chlorella and diatoms for real-time monitoring of water quality. This microalgae trap is fabricated by embedding a piezoelectric resonator in a single channel. In comparison to previously reported acoustophoretic particle traps, this device has a simple structure and does not require sheath flows, which makes the device low cost and simple to fabricate and operate. Using Chlorella kessleri and the marine diatom Thalassiosira pseudonana as model algae, we report the microalgae trap has demonstrated notable particle trapping efficiencies between 82 % to 74 % for Chlorella with a through-flow of 0.5 μl/min to 2.7 μl/min and 85 % to 79 % for diatoms with a 0.5 μl/min to 4 μl/min through-flow. The trap has also shown to simultaneously trap Chlorella and diatoms at different heights of the microchannel. This device has high promise for trapping, separating and manipulating microalgae.

47 OTHER INSTRUMENTATION↗

Development of a high-productivity, halophilic, thermotolerant microalga Picochlorum renovo

Abstract Microalgae are promising biocatalysts for applications in sustainable fuel, food, and chemical production. Here, we describe culture collection screening, down-selection, and development of a high-productivity, halophilic, thermotolerant microalga, Picochlorum renovo . This microalga displays a rapid growth rate and high diel biomass productivity (34 g m −2 day −1 ), with a composition well-suited for downstream processing. P. renovo exhibits broad salinity tolerance (growth at 107.5 g L −1 salinity) and thermotolerance (growth up to 40 °C), beneficial traits for outdoor cultivation. We report complete genome sequencing and analysis, and genetic tool development suitable for expression of transgenes inserted into the nuclear or chloroplast genomes. We further evaluate mechanisms of halotolerance via comparative transcriptomics, identifying novel genes differentially regulated in response to high salinity cultivation. These findings will enable basic science inquiries into control mechanisms governing Picochlorum biology and lay the foundation for development of a microalga with industrially relevant traits as a model photobiology platform.

09 BIOMASS FUELS↗

Life-cycle analysis of microalgae-based polyurethane foams

Polyurethane plastics are essential in many consumer and commercial products such as insulation, furniture, automotive interiors, and clothing. Pathways for producing polyurethane from microalgae offer an opportunity to reduce greenhouse gas emissions and other environmental impacts and can incorporate processes that avoid the use of toxic isocyanates typically used in conventional polyurethane production processes. In this study, the greenhouse gas emissions, fossil energy, and water consumption of biobased polyurethane and biobased non-isocyanate polyurethane were evaluated via life-cycle analysis using the R&D Greenhouse Gases, Regulated Emissions, and Energy Use in Technologies model. Microalgae-based polyurethane foam was found to achieve greenhouse gas emission reductions of up to 79% compared with conventional polyurethane foam production. The greenhouse gas reductions for the non-isocyanate microalgae polyurethane pathway are slightly lower at 58% compared with conventional polyurethane foam. However, it offers additional benefits by reducing toxicity potential compared to the isocyanate polyurethane pathway. The analysis also included a biorefinery-level analysis to evaluate the impact of incorporating polyurethane production into fuel-processing microalgae biorefineries. The sensitivity analyses conducted in this study reveal that improved algae cultivation strategies can lead to decreases of up to 127% and 80% in GHG emissions from the baseline process of Bio-PU and Bio-NIPU, respectively. Likewise, implementation of renewable electricity can result in up to 128% and 74% lower GHG emissions compared to the baseline production of Bio-PU and Bio-NIPU, respectively. Finally, the analysis evaluated different coproduct handling methods including displacement and allocation (based on mass, energy, and market-value). The results suggest that it is important to consider both the displacement and allocation methods as these led to significant differences in the environmental impacts.

36 MATERIALS SCIENCE↗

Modulation of xanthophyll cycle impacts biomass productivity in the marine microalga Nannochloropsis

Life on earth depends on photosynthetic primary producers that exploit sunlight to fix CO 2 into biomass. Approximately half of global primary production is associated with microalgae living in aquatic environments. Microalgae also represent a promising source of biomass to complement crop cultivation, and they could contribute to the development of a more sustainable bioeconomy. Photosynthetic organisms evolved multiple mechanisms involved in the regulation of photosynthesis to respond to highly variable environmental conditions. While essential to avoid photodamage, regulation of photosynthesis results in dissipation of absorbed light energy, generating a complex trade-off between protection from stress and light-use efficiency. This work investigates the impact of the xanthophyll cycle, the light-induced reversible conversion of violaxanthin into zeaxanthin, on the protection from excess light and on biomass productivity in the marine microalgae of the genus Nannochloropsis. Zeaxanthin is shown to have an essential role in protection from excess light, contributing to the induction of nonphotochemical quenching and scavenging of reactive oxygen species. On the contrary, the overexpression of zeaxanthin epoxidase enables a faster reconversion of zeaxanthin to violaxanthin that is shown to be advantageous for biomass productivity in dense cultures in photobioreactors. These results demonstrate that zeaxanthin accumulation is critical to respond to strong illumination, but it may lead to unnecessary energy losses in light-limiting conditions and accelerating its reconversion to violaxanthin provides an advantage for biomass productivity in microalgae.

59 BASIC BIOLOGICAL SCIENCES↗

Development and Validation of a Process Model and Open-Source Process Simulator for Microalgae-Based Tertiary Phosphorus Recovery

Microalgae-based tertiary wastewater treatment has the potential to meet stringent effluent phosphorus limits, with the added benefit of producing a marketable feedstock. However, the lack of validated mechanistic models and their implementation in process simulators have limited the adoption of this technology. In this study, an updated lumped pathway metabolic model (Phototrophic-Mixotrophic Process Model, PM 2 ), including both photoautotrophic and heterotrophic metabolisms of microalgae, was developed to predict effluent phosphorus concentration and biomass yield in response to dynamic influent and varying environmental conditions. The model was implemented in QSDsan – an open-source, Python-based design and simulation platform – for robust simulation under uncertainty. A global sensitivity analysis was performed to prioritize model parameters for calibration. The model was then calibrated and validated using batch experimental data and 45 days of continuous online monitoring data from a full-scale (568 m 3 ·d -1 ) microalgae-based tertiary wastewater treatment plant (EcoRecover process). In particular, along with dynamic influent composition, temperature and light intensity data with diel variation were provided as model inputs to reflect the microalgal behavior under day-night cycling. Overall, the QSDsan-based microalgae process simulator was able to predict effluent phosphorus within 0.02–0.04 mg-P·L -1 , while also capturing the general trends of state variables according to nutrient availability.

Lumped pathway metabolic model↗

Volatile Metabolites Emission by In Vivo Microalgae—An Overlooked Opportunity?

Fragrances and malodors are ubiquitous in the environment, arising from natural and artificial processes, by the generation of volatile organic compounds (VOCs). Although VOCs constitute only a fraction of the metabolites produced by an organism, the detection of VOCs has a broad range of civilian, industrial, military, medical, and national security applications. The VOC metabolic profile of an organism has been referred to as its ‘volatilome’ (or ‘volatome’) and the study of volatilome/volatome is characterized as ‘volatilomics’, a relatively new category in the ‘omics’ arena. There is considerable literature on VOCs extracted destructively from microalgae for applications such as food, natural products chemistry, and biofuels. VOC emissions from living (in vivo) microalgae too are being increasingly appreciated as potential real-time indicators of the organism’s state of health (SoH) along with their contributions to the environment and ecology. This review summarizes VOC emissions from in vivo microalgae; tools and techniques for the collection, storage, transport, detection, and pattern analysis of VOC emissions; linking certain VOCs to biosynthetic/metabolic pathways; and the role of VOCs in microalgae growth, infochemical activities, predator-prey interactions, and general SoH.

Biochemistry & Molecular Biology↗

Multi-Wavelength Based Optical Density Sensor for Autonomous Monitoring of Microalgae

A multi-wavelength based optical density sensor unit was designed, developed, and evaluated to monitor microalgae growth in real time. The system consisted of five main components including (1) laser diode modules as light sources (2) photodiodes as detectors, (3) driver circuit, (4) flow cell and (5) sensor housing temperature controller. The sensor unit was designed to be integrated into any microalgae culture system for both real time and non-real time optical density measurements and algae growth monitoring applications. It was shown that the sensor unit was capable of monitoring the dynamics and physiological changes of the microalgae culture in real-time. Algae biomass concentration was accurately estimated with optical density measurements at 650, 685 and 780 nm wavelengths used by the sensor unit. The sensor unit was able to monitor cell concentration as high as 1.05 g L -1 (1.51x10 8 cells mL -1 ) during the culture growth without any sample preparation for the measurements. Since high cell concentrations do not need to be diluted using the sensor unit, the system has the potential to be used in industrial microalgae cultivation systems for real time monitoring and control applications that can lead to improved resource use efficiency.

09 BIOMASS FUELS↗