Bicistronic expression of nuclear transgenes in Chlamydomonas reinhardtii.

In eukaryotic organisms, proteins are typically translated from monocistronic messenger RNAs containing a single coding sequence (CDS). However, recent long transcript sequencing identified 87 nuclear polycistronic mRNAs in Chlamydomonas reinhardtii natively carrying multiple co-expressed CDSs. In this study, we investigated the dynamics of 22 short intergenic sequences derived from these native polycistronic loci by their application in genetic constructs for synthetic transgene expression. A promising candidate sequence was identified based on the quantification of transformation efficiency and expression strength of a fluorescence reporter protein. Subsequently, the expression of independent proteins from one mRNA was verified by cDNA amplification and protein molecular mass characterization. We demonstrated engineered bicistronic expression in vivo to drive successful co-expression of several terpene synthases with the selection marker aphVIII. Bicistronic transgene design resulted in significantly increased (E)-α-bisabolene production of 7.95 mg L-1 from a single open reading frame, 18.1× fold higher than previous reports. Use of this strategy simplifies screening procedures for identification of high-level expressing transformants, does not require the application of additional fluorescence reporters, and reduces the nucleotide footprint compared to classical monocistronic expression cassettes. Although clear advantages for bicistronic transgene expression were observed, this strategy was found to be limited to the aphVIII marker, and further studies are necessary to gain insights into the underlying mechanism that uniquely permits this co-expression from the algal nuclear genome.

Lipid turnover through lipophagy in the newly identified extremophilic green microalga Chlamydomonas urium.

Autophagy is a central degradative pathway highly conserved among eukaryotes, including microalgae, which remains unexplored in extremophilic organisms. In this study, we described and characterized autophagy in the newly identified extremophilic green microalga Chlamydomonas urium, which was isolated from an acidic environment. The nuclear genome of C. urium was sequenced, assembled and annotated in order to identify autophagy-related genes. Transmission electron microscopy, immunoblotting, metabolomic and photosynthetic analyses were performed to investigate autophagy in this extremophilic microalga. The analysis of the C. urium genome revealed the conservation of core autophagy-related genes. We investigated the role of autophagy in C. urium by blocking autophagic flux with the vacuolar ATPase inhibitor concanamycin A. Our results indicated that inhibition of autophagic flux in this microalga resulted in a pronounced accumulation of triacylglycerols and lipid droplets (LDs). Metabolomic and photosynthetic analyses indicated that C. urium cells with impaired vacuolar function maintained an active metabolism. Such effects were not observed in the neutrophilic microalga Chlamydomonas reinhardtii. Inhibition of autophagic flux in C. urium uncovered an active recycling of LDs through lipophagy, a selective autophagy pathway for lipid turnover. This study provided the metabolic basis by which extremophilic algae are able to catabolize lipids in the vacuole.

Features of the microalga Raphidocelis subcapitata: physiology and applications.

The microalga Raphidocelis subcapitata was isolated from the Nitelva River (Norway) and subsequently deposited in the collection of the Norwegian Institute of Water Research as "Selenastrum capricornutum Printz". This freshwater microalga, also known as Pseudokirchneriella subcapitata, acquired much of its notoriety due to its high sensitivity to different chemical species, which makes it recommended by different international organizations for the assessment of ecotoxicity. However, outside this scope, R. subcapitata continues to be little explored. This review aims to shed light on a microalga that, despite its popularity, continues to be an "illustrious" unknown in many ways. Therefore, R. subcapitata taxonomy, phylogeny, shape, size/biovolume, cell ultra-structure, and reproduction are reviewed. The nutritional and cultural conditions, chronological aging, and maintenance and preservation of the alga are summarized and critically discussed. Applications of R. subcapitata, such as its use in aquatic toxicology (ecotoxicity assessment and elucidation of adverse toxic outcome pathways) are presented. Furthermore, the latest advances in the use of this alga in biotechnology, namely in the bioremediation of effluents and the production of value-added biomolecules and biofuels, are highlighted. To end, a perspective regarding the future exploitation of R. subcapitata potentialities, in a modern concept of biorefinery, is outlined. KEY POINTS: • An overview of alga phylogeny and physiology is critically reviewed. • Advances in alga nutrition, cultural conditions, and chronological aging are presented. • Its use in aquatic toxicology and biotechnology is highlighted.

Effects of high dietary inclusion of Arthrospira platensis, either extruded or supplemented with a super-dosing multi-enzyme mixture, on broiler growth performance and major meat quality parameters.

BACKGROUND: This investigation assessed the effects of high dietary inclusion of Spirulina (Arthrospira platensis) on broiler chicken growth performance, meat quality and nutritional attributes. For this, 120 male broiler chicks were housed in 40 battery brooders (three birds per brooder). Initially, for 14 days, a standard corn and soybean meal diet was administered. Subsequently, from days 14 to 35, chicks were assigned to one of the four dietary treatments (n = 10 per treatment): (1) control diet (CTR); (2) diet with 15% Spirulina (SP); (3) diet with 15% extruded Spirulina (SPE); and (4) diet with 15% Spirulina plus a super-dosing enzymes supplement (0.20% pancreatin extract and 0.01% lysozyme) (SPM). RESULTS: Throughout the experimental period, both SP and SPM diets resulted in decreased final body weight and body weight gain compared to control (p < 0.001), with the SPE diet showing comparable results to CTR. The SPE diet prompted an increase in average daily feed intake (p = 0.026). However, all microalga treatments increased the feed conversion ratio compared to CTR. Dietary inclusion of Spirulina notably increased intestinal content viscosity (p < 0.010), which was mitigated by the SPM diet. Spirulina supplementation led to lower pH levels in breast meat 24 h post-mortem and heightened the b* colour value in both breast and thigh meats (p < 0.010). Furthermore, Spirulina contributed to an increased accumulation of total carotenoids, n-3 polyunsaturated fatty acids (PUFA), and saturated fatty acids (SFA), while diminishing n-6 PUFA, thus altering the n-6/n-3 and PUFA/SFA ratios favourably (p < 0.001). However, it also reduced zinc concentration in breast meat (p < 0.001). CONCLUSIONS: The findings indicate that high Spirulina levels in broiler diets impair growth due to increased intestinal viscosity, and that extrusion pre-treatment mitigates this effect. Despite reducing digesta viscosity, a super-dosing enzyme mix did not improve growth. Data also indicates that Spirulina enriches meat with antioxidants and n-3 PUFA but reduces α-tocopherol and increases saturated fats. Reduced zinc content in meat suggests the need for Spirulina biofortification to maintain its nutritional value.

Enhancing microalgae biomass production: Exploring improved scraping frequency in a hybrid cultivation system.

Recently, hybrid systems, such as those incorporating high-rate algal ponds (HRAPs) and biofilm reactors (BRs), have shown promise in treating domestic wastewater while cultivating microalgae. In this context, the objective of the present study was to determine an improved scraping frequency to maximize microalgae biomass productivity in a mix of industrial (fruit-based juice production) and domestic wastewater. The mix was set to balance the carbon/nitrogen ratio. The scraping strategy involved maintaining 1 cm wide stripes to retain an inoculum in the reactor. Three scraping frequencies (2, 4, and 6 days) were evaluated. The findings indicate that a scraping frequency of each 2 days provided the highest biomass productivity (18.75 g total volatile solids m-2 d-1). The species' behavior varied with frequency: Chlorella vulgaris was abundant at 6-day intervals, whereas Tetradesmus obliquus favored shorter intervals. Biomass from more frequent scraping demonstrated a higher lipid content (15.45%). Extrapolymeric substance production was also highest at the 2-day frequency. Concerning wastewater treatment, the system removed 93% of dissolved organic carbon and ∼100% of ammoniacal nitrogen. Combining industrial and domestic wastewater sources to balance the carbon/nitrogen ratio enhanced treatment efficiency and biomass yield. This study highlights the potential of adjusting scraping frequencies in hybrid systems for improved wastewater treatment and microalgae production.

Biofuel from wastewater-grown microalgae: A biorefinery approach using hydrothermal liquefaction and catalyst upgrading.

Third-generation biofuels from microalgae are becoming necessary for sustainable energy. In this context, this study explores the hydrothermal liquefaction (HTL) of microalgae biomass grown in wastewater, consisting of 30% Chlorella vulgaris, 69% Tetradesmus obliquus, and 1% cyanobacteria Limnothrix planctonica, and the subsequent upgrading of the produced bio-oil. The novelty of the work lies in integrating microalgae cultivation in wastewater with HTL in a biorefinery approach, enhanced using a catalyst to upgrade the bio-oil. Different temperatures (300, 325, and 350 °C) and reaction times (15, 30, and 45 min) were tested. The bio-oil upgrading occurred with a Cobalt-Molybdenum (CoMo) catalyst for 1 h at 375 °C. Post-HTL, although the hydrogen-to-carbon (H/C) ratio decreased from 1.70 to 1.38-1.60, the oxygen-to-carbon (O/C) ratio also decreased from 0.39 to 0.079-0.104, and the higher heating value increased from 20.6 to 36.4-38.3 MJ kg-1. Palmitic acid was the main component in all bio-oil samples. The highest bio-oil yield was at 300 °C for 30 min (23.4%). Upgrading increased long-chain hydrocarbons like heptadecane (5%), indicating biofuel potential, though nitrogenous compounds such as hexadecanenitrile suggest a need for further hydrodenitrogenation. Aqueous phase, solid residues, and gas from HTL can be used for applications such as biomass cultivation, bio-hydrogen, valuable chemicals, and materials like carbon composites and cement additives, promoting a circular economy. The study underscores the potential of microalgae-derived bio-oil as sustainable biofuel, although further refinement is needed to meet current fuel standards.

Differences in Exudates Between Strains of Chlorella sorokiniana Affect the Interaction with the Microalga Growth-Promoting Bacteria Azospirillum brasilense : Differences in Exudates Between Strains of Chlorella sorokiniana Affect the Interaction with the Microalga Growth-Promoting Bacteria Azospirillum brasilense.

The microalga Chlorella sorokiniana and the microalgae growth-promoting bacteria (MGPB) Azospirillum brasilense have a mutualistic interaction that can begin within the first hours of co-incubation; however, the metabolites participating in this initial interaction are not yet identified. Nuclear magnetic resonance (NMR) was used in the present study to characterize the metabolites exuded by two strains of C. sorokiniana (UTEX 2714 and UTEX 2805) and A. brasilense Cd when grown together in an oligotrophic medium. Lactate and myo-inositol were identified as carbon metabolites exuded by the two strains of C. sorokiniana; however, only the UTEX 2714 strain exuded glycerol as the main carbon compound. In turn, A. brasilense exuded uracil when grown on the exudates of either microalga, and both microalga strains were able to utilize uracil as a nitrogen source. Interestingly, although the total carbohydrate content was higher in exudates from C. sorokiniana UTEX 2805 than from C. sorokiniana UTEX 2714, the growth of A. brasilense was greater in the exudates from the UTEX 2714 strain. These results highlight the fact that in the exuded carbon compounds differ between strains of the same species of microalgae and suggest that the type, rather than the quantity, of carbon source is more important for sustaining the growth of the partner bacteria.

Metabonomics analysis of microalga Scenedesmus obliquus under ciprofloxacin stress.

The wide use of antibiotics in aquaculture has triggered global ecological security issue. Microalgal bioremediation is a promising strategy for antibiotics elimination due to carbon recovery, detoxification and various ecological advantages. However, a lack of understanding with respect to the corresponding regulation mechanism towards antibiotic stress may limit its practical applicability. The microalga Scenedesmus obliquus was shown to be capable of effectively eliminating ciprofloxacin (CIP), which is a common antibiotic used in aquaculture. However, the corresponding transcriptional alterations require further investigation and verification at the metabolomic level. Thus, this study uncovered the metabolomic profiles and detailed toxic and defense mechanisms towards CIP in S. obliquus using untargeted metabolomics. The enhanced oligosaccharide/polyol/lipid transport, up-regulation of carbohydrate and arachidonic acid metabolic pathways and increased energy production via EMP metabolism were observed as defense mechanisms of microalgal cells to xenobiotic CIP. The toxic metabolic responses included: (1) down-regulation of parts of mineral and organic transporters; (2) electrons competition between antibiotic and NAD during intracellular CIP degradation; and (3) suppressed expression of the hem gene in chlorophyll biosynthesis. This study describes the metabolic profile of microalgae during CIP elimination and reveals the key pathways from the perspective of metabolism, thereby providing information on the precise regulation of antibiotic bioremediation via microalgae.

Photosynthetic performance of Chlamydopodium (Chlorophyta) cultures grown in outdoor bioreactors.

The microalga Chlamydopodium fusiforme MACC-430 was cultured in two types of outdoor pilot cultivation units-a thin-layer cascade (TLC) and a raceway pond (RWP) placed in a greenhouse. This case study aimed to test their potential suitability for cultivation scale-up to produce biomass for agriculture purposes (e.g., as biofertilizer or biostimulant). The culture response to the alteration of environmental conditions was evaluated in "exemplary" situations of good and bad weather conditions using several photosynthesis measuring techniques, namely oxygen production, and chlorophyll (Chl) fluorescence. Validation of their suitability for online monitoring in large-scale plants has been one of the objectives of the trials. Both techniques were found fast and robust reliable to monitor microalgae activity in large-scale cultivation units. In both bioreactors, Chlamydopodium cultures grew well in the semi-continuous regime using daily dilution (0.20-0.25 day-1). The biomass productivity calculated per volume was significantly (about 5 times) higher in the RWPs compared to the TLCs. The measured photosynthesis variables showed that the build-up of dissolved oxygen concentration in the TLC was higher, up to 125-150% of saturation (%sat) as compared to the RWP (102-104%sat). As only ambient CO2 was available, its shortage was indicated by a pH increase due to photosynthetic activity in the thin-layer bioreactor at higher irradiance intensities. In this setup, the RWP was considered more suitable for scale-up due to higher areal productivity, lower construction and maintenance costs, the smaller land area required to maintain large culture volumes, as well as lower carbon depletion and dissolved oxygen build-up. KEY POINTS: • Chlamydopodium was grown in both raceways and thin-layer cascades in pilot-scale. • Various photosynthesis techniques were validated for growth monitoring. • In general, raceway ponds were evaluated as more suitable for cultivation scale-up.

The effect of Dunaliella tertiolecta supplementation on diet-induced obesity in UCP1-deficient mice.

We previously demonstrated that dietary supplementation with Dunaliella tertiolecta (DT) increases uncoupling protein 1 (UCP1) expression in brown adipose tissue (BAT) and improves diet-induced obesity (DIO) in C57BL/6 J mice at thermoneutrality (30 °C). Here, we investigated whether DT improves DIO in a thermoneutral UCP1-deficient (KO) animal. KO mice were fed a high-fat diet supplemented with DT for 12 weeks. Compared to control group without DT, body weight was significantly reduced in DT group with no difference in food intake. Dunaliella tertiolecta-supplemented mice exhibited lower adiposity and well-maintained multilocular morphology in BAT, in which a significant increase in gene expression of PR domain containing 16 was detected in DT group compared to control group. Moreover, increase in UCP2 level and/or decrease in ribosomal protein S6 phosphorylation were detected in adipose tissues of DT group relative to control group. These results suggest that DT supplementation improves DIO by stimulating UCP1-independent energy dissipation at thermoneutrality.

Therapeutic Potentials of Microalgae and Their Bioactive Compounds on Diabetes Mellitus.

Diabetes mellitus is a metabolic disorder characterized by hyperglycemia due to impaired insulin secretion, insulin resistance, or both. Oxidative stress and chronic low-grade inflammation play crucial roles in the pathophysiology of diabetes mellitus. There has been a growing interest in applying natural products to improve metabolic derangements without the side effects of anti-diabetic drugs. Microalgae biomass or extract and their bioactive compounds have been applied as nutraceuticals or additives in food products and health supplements. Several studies have demonstrated the therapeutic effects of microalgae and their bioactive compounds in improving insulin sensitivity attributed to their antioxidant, anti-inflammatory, and pancreatic ß-cell protective properties. However, a review summarizing the progression in this topic is lacking despite the increasing number of studies reporting their anti-diabetic potential. In this review, we gathered the findings from in vitro, in vivo, and human studies to discuss the effects of microalgae and their bioactive compounds on diabetes mellitus and the mechanisms involved. Additionally, we discuss the limitations and future perspectives of developing microalgae-based compounds as a health supplement for diabetes mellitus. In conclusion, microalgae-based supplementation has the potential to improve diabetes mellitus and be applied in more clinical studies in the future.

The Antiviral Potential of Algal Lectins.

Algae have emerged as fascinating subjects of study due to their vast potential as sources of valuable metabolites with diverse biotechnological applications, including their use as fertilizers, feed, food, and even pharmaceutical precursors. Among the numerous compounds found in algae, lectins have garnered special attention for their unique structures and carbohydrate specificities, distinguishing them from lectins derived from other sources. Here, a comprehensive overview of the latest scientific and technological advancements in the realm of algal lectins with a particular focus on their antiviral properties is provided. These lectins have displayed remarkable effectiveness against a wide range of viruses, thereby holding great promise for various antiviral applications. It is worth noting that several alga species have already been successfully commercialized for their antiviral potential. However, the discovery of a diverse array of lectins with potent antiviral capabilities suggests that the field holds immense untapped potential for further expansion. In conclusion, algae stand as a valuable and versatile resource, and their lectins offer an exciting avenue for developing novel antiviral agents, which may lead to the development of cutting-edge antiviral therapies.

Extraction of Valuable Biomolecules from the Microalga Haematococcus pluvialis Assisted by Electrotechnologies.

The freshwater microalga Haematococcus pluvialis is well known as the cell factory for natural astaxanthin, which composes up to 4-7% of its total dry weight. The bioaccumulation of astaxanthin in H. pluvialis cysts seems to be a very complex process that depends on different stress conditions during its cultivation. The red cysts of H. pluvialis develop thick and rigid cell walls under stress growing conditions. Thus, the biomolecule extraction requires general cell disruption technologies to reach a high recovery rate. This short review provides an analysis of the different steps in H. pluvialis's up and downstream processing including cultivation and harvesting of biomass, cell disruption, extraction and purification techniques. Useful information on the structure of H. pluvialis's cells, biomolecular composition and properties and the bioactivity of astaxanthin is collected. Special emphasis is given to the recent progress in application of different electrotechnologies during the growth stages and for assistance of the recovery of different biomolecules from H. pluvialis.

Advanced pathway engineering for phototrophic putrescine production.

The polyamine putrescine (1,4-diaminobutane) contributes to cellular fitness in most organisms, where it is derived from the amino acids ornithine or arginine. In the chemical industry, putrescine serves as a versatile building block for polyamide synthesis. The green microalga Chlamydomonas reinhardtii accumulates relatively high putrescine amounts, which, together with recent advances in genetic engineering, enables the generation of a powerful green cell factory to promote sustainable biotechnology for base chemical production. Here, we report a systematic investigation of the native putrescine metabolism in C. reinhardtii, leading to the first CO2 -based bio-production of putrescine, by employing modern synthetic biology and metabolic engineering strategies. A CRISPR/Cas9-based knockout of key enzymes of the polyamine biosynthesis pathway identified ornithine decarboxylase 1 (ODC1) as a gatekeeper for putrescine accumulation and demonstrated that the arginine decarboxylase (ADC) route is likely inactive and that amine oxidase 2 (AMX2) is mainly responsible for putrescine degradation in C. reinhardtii. A 4.5-fold increase in cellular putrescine levels was achieved by engineered overexpression of potent candidate ornithine decarboxylases (ODCs). We identified unexpected substrate promiscuity in two bacterial ODCs, which exhibited co-production of cadaverine and 4-aminobutanol. Final pathway engineering included overexpression of recombinant arginases for improved substrate availability as well as functional knockout of putrescine degradation, which resulted in a 10-fold increase in cellular putrescine titres and yielded 200 mg/L in phototrophic high cell density cultivations after 10 days.

Intermediates formed during natural attenuation of C9 aromatics under simulated marine conditions: Identification, transformation pathway, and toxicity to microalgae.

C9 aromatics - benzene hydrocarbon containing nine carbon atoms among - leakage accident has caused serious damage to the marine ecology near Quangang District, Fujian Province, China. The ecological restoration of the accident sea area is basically realized through natural attenuation. To determine whether the natural attenuation of C9 aromatics in the marine environment will generate highly toxic intermediates, and thus cause more serious harm to marine ecology, the intermediates of C9 aromatics (n-propylbenzene, isopropylbenzene, 2-ethyltoluene, 3-ethyltoluene, 4-ethyltoluene, 1,2,3-trimethylbenzene, 1,2,4-trimethylbenzene, 1,3,5-trimethylbenzene, and indene) in the process of natural attenuation were studied under the marine conditions simulated by a microcosm. The acute toxic effects of 12 intermediates with longer residual time on Phaeodactylum tricornutum were also ascertained. Twenty natural attenuation intermediates of C9 aromatics were identified. These products primarily include the derivatives of phenols, aromatic alcohols, aromatic aldehydes, aromatic ketones, and aromatic acids, as well as an aromatic lactone compound. No intermediates of 1,3,5-trimethylbenzene and indene during the attenuation process were determined. The indirect photooxidation initiated by hydroxyl radical might play an essential role in the formation of intermediates of C9 aromatic. Based on the 96-h EC50 values for P. tricornutum, the toxicity of the 12 intermediates, in descending order, was: 4-ethylphenol, 2-methylacetophenone, 2,3-dimethylbenzyl alcohol, 4-methylacetophenone, 3-methylacetophenone, 1-phenyl-1-propanol, 1-(2-methylphenyl) ethanol, 2-phenyl-2-propanol, 3,4-dimethylbenzoic acid, 2,4-dimethylbenzoic acid, 2,5-dimethylbenzoic acid, then 4-tolylacetic acid. The 96-h EC50 values of the intermediates of C9 aromatics to P. tricornutum ranged from 8.4 to 199.1 mg/L, which were lower than that of their corresponding parent compound. The findings provided essential fundamental insights for the assessment of marine environmental risk of C9 aromatics leakage accidents, and subsequent emergency disposal countermeasures.

Characterization and Optimization of Culture Conditions for Aurantiochytrium sp. SC145 Isolated from Sand Cay (Son Ca) Island, Vietnam, and Antioxidative and Neuroprotective Activities of Its Polyunsaturated Fatty Acid Mixture.

Aurantiochytrium is a heterotrophic marine microalga that has potential industrial applications. The main objectives of this study were to isolate an Aurantiochytrium strain from Sand Cay (Son Ca) Island, Vietnam, optimize its culture conditions, determine its nutritional composition, extract polyunsaturated fatty acids (PUFAs) in the free (FFA) and the alkyl ester (FAAE) forms, and evaluate the antioxidation and neuroprotection properties of the PUFAs. Aurantiochytrium sp. SC145 can be grown stably under laboratory conditions. Its culture conditions were optimized for a dry cell weight (DCW) of 31.18 g/L, with total lipids comprising 25.29%, proteins 7.93%, carbohydrates 15.21%, and carotenoid at 143.67 µg/L of DCW. The FAAEs and FFAs extracted from Aurantiochytrium sp. SC145 were rich in omega 3-6-9 fatty acids (40.73% and 44.00% of total fatty acids, respectively). No acute or subchronic oral toxicity was determined in mice fed with the PUFAs in FFA or FAAE forms at different doses over 90 days. Furthermore, the PUFAs in the FFA or FAAE forms and their main constituents of EPA, DHA, and ALA showed antioxidant and AChE inhibitory properties and neuroprotective activities against damage caused by H2O2- and amyloid-ß protein fragment 25-35 (Aß25-35)-induced C6 cells. These data suggest that PUFAs extracted from Aurantiochytrium sp. SC145 may be a potential therapeutic target for the treatment of neurodegenerative disorders.

Supercritical CO2 Extract from Microalga Tetradesmus obliquus: The Effect of High-Pressure Pre-Treatment.

High-pressure pre-treatment followed by supercritical carbon dioxide (ScCO2) extraction (300 bar, 40 °C) was applied for the attainment of the lipophilic fraction of microalga Tetradesmus obliquus. The chemical profile of supercritical extracts of T. obliquus was analyzed by ultra-high-performance liquid chromatography-high-resolution mass spectrometry with electrospray ionization (UHPLC-ESI-HRMS). Moreover, the impact of ScCO2 on the microbiological and metal profile of the biomass was monitored. The application of the pre-treatment increased the extraction yield approximately three-fold compared to the control. In the obtained extracts (control and pre-treated extracts), the identified components belonged to triacylglyceroles, fatty acid derivatives, diacylglycerophosphocholines and diacylglycerophosphoserines, pigments, terpenes, and steroids. Triacylglycerols (65%) were the most dominant group of compounds in the control extract. The pre-treatment decreased the percentage of triacylglycerols to 2%, while the abundance of fatty acid derivatives was significantly increased (82%). In addition, the pre-treatment led to an increase in the percentages of carotenoids, terpenoids, and steroids. Furthermore, it was determined that ScCO2 extraction reduced the number of microorganisms in the biomass. Considering its microbiological and metal profiles, the biomass after ScCO2 can potentially be used as a safe and important source of organic compounds.

DASH cryptochrome 1, a UV-A receptor, balances the photosynthetic machinery of Chlamydomonas reinhardtii.

Drosophila, Arabidopsis, Synechocystis, Homo (DASH) cryptochromes belong to the cryptochrome/photolyase family and can act as DNA repair enzymes. In bacteria and fungi, they also can play regulatory roles, but in plants their biological functions remain elusive. Here, we characterize CRY-DASH1 from the green alga Chlamydomonas reinhardtii. We perform biochemical and in vitro photochemical analysis. For functional characterization, a knock-out mutant of cry-dash1 is used. CRY-DASH1 protein is localized in the chloroplast and accumulates at midday. Although the photoautotrophic growth of the mutant is significantly reduced compared to the wild-type (WT), the mutant has increased levels of photosynthetic pigments and a higher maximum photochemical efficiency of photosystem II (PS II). Hyper-stacking of thylakoid membranes occurs together with an increase in proteins of the PS II reaction center, D1 and its antenna CP43, but not of their transcripts. CRY-DASH1 binds fully reduced flavin adenine dinucleotide and the antenna 5,10-methenyltetrahydrofolate, leading to an absorption peak in the UV-A range. Supplementation of white light with UV-A increases photoautotrophic growth of the WT but not of the cry-dash1 mutant. These results suggest a balancing function of CRY-DASH1 in the photosynthetic machinery and point to its role as a photoreceptor for the UV-A range separated from the absorption of photosynthetic pigments.

The ATG4 protease integrates redox and stress signals to regulate autophagy.

Autophagy is a highly conserved degradative pathway that ensures cellular homeostasis through the removal of damaged or useless intracellular components including proteins, membranes, or even entire organelles. A main hallmark of autophagy is the biogenesis of autophagosomes, double-membrane vesicles that engulf and transport to the vacuole the material to be degraded and recycled. The formation of autophagosomes responds to integrated signals produced as a consequence of metabolic reactions or different types of stress and is mediated by the coordinated action of core autophagy-related (ATG) proteins. ATG4 is a key Cys-protease with a dual function in both ATG8 lipidation and free ATG8 recycling whose balance is crucial for proper biogenesis of the autophagosome. ATG4 is conserved in the green lineage, and its regulation by different post-translational modifications has been reported in the model systems Chlamydomonas reinhardtii and Arabidopsis. In this review, we discuss the major role of ATG4 in the integration of stress and redox signals that regulate autophagy in algae and plants.

Attenuation pathways of erythromycin and biochemical responses related to algal growth and lipid synthesis in a microalga-effluent system.

Microalgal cultivation in municipal wastewater treatment plants (WWTPs) can realize the coupling of wastewater treatment and microalgae energy utilization, however, the residual antibiotics in effluents from WWTPs affect the growth of microalgae. In this study, green alga (Scenedesmus obliquus) cells were inoculated into the effluents to ascertain the attenuation pathways of erythromycin (ERY) and the biochemical responses of microalga in a microalga-effluent system. Results showed that hydrolysis, photolysis, and biodegradation (including bioadsorption) cause the attenuation of ERY in a microalga-effluent system, and the biodegradation (including bioadsorption) has the greatest removal rate (reaching a maximum of 57.87%), followed by hydrolysis (reaching a maximum of 34.13%), and photolysis (less than 5%) after five days. The photosynthetic pigment contents in cells of microalga decreased the most (by 35.66% for chlorophyll a), and the production of ROS was stimulated (by 33.75%) after five-day exposure to ERY at an initial concentration of 100 µg/L. Meanwhile, the activity of ribulose-1,5-biphosphate carboxylase (RuBPCase) decreased by 55.65%, and the activity of acetyl-CoA carboxylase (ACCase) increased by 55.65%. The ROS level, photosynthetic pigment content, and RuBPCase activity were extremely significantly correlated with each other (P < 0.01), indicating that exposure to ERY changed those biochemical responses related to the rate of photosynthesis of microalga, inhibiting the growth thereof. On the other hand, exposure to ERY increased lipid production by microalga through the induced ACCase activity.