Search

The utilization of residual sludge by microalgae represents an environmentally sustainable method for resource recovery. In this study, Tetradesmus obliquus was cultured in hydrolysate derived from toxic sludge. Under symbiotic conditions with bacteria, Tetradesmus obliquus demonstrated enhanced toxin degradation capability and biomass accumulation, which exhibited a 1.39-fold increase in algal cell density, a 1.50-fold increase in Rubisco activity, and a total protein content of 341.83 ± 6.99 mg/L on the 30th day of cultivation. Metabolic utilization of substances in the hydrolysate by microalgae led to a toxicity removal rate of up to 60.43% by day 10. Phenylalanine showed the most significant increase among essential amino acids, and transcriptomic profiling identified genes (gene_16399, gene_16602) involved in phenylalanine enrichment. Macrotranscriptomics showed that bacteria upregulated the TCS system and tryptophan metabolism, supplying microalgae with more CO 2 and IAA, which enhanced amino acid enrichment. This study established a non-toxic and biomass-accumulating bacterial-algal co-cultivation system., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Utilizing microalgae to capture flue gas pollutants is an effective strategy for mitigating greenhouse gas emissions. However, existing carbon-fixing microalgae exhibit poor tolerance towards acidic flue gas. In this study, the Desmodesmus sp. SZ-1, which can thrive in acidic environments and efficiently sequester CO 2 , was isolated. Desmodesmus sp. SZ-1 exhibited strong acid tolerance ability, with an average carbon fixation rate of 497.6 mg/L/d under 10 % CO 2 and pH 3.5. Physiological analysis revealed that SZ-1 responded to high CO 2 by increasing chlorophyll levels while coping with acidic stress by activating antioxidant enzymes. Genome analysis revealed a large number of carbon fixation and acid adaptation genes, involved in membrane lipid biosynthesis, H + pumps, molecular chaperones, peroxidase system, amino acid synthesis, and carbonic anhydrase. This study provides a novel algal resource for mitigating acid gas emissions and a comprehensive genetic database for genetically modifying microalgae., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background: Canthaxanthin is a ketocarotenoid with high antioxidant activity, and it is primarily produced by microalgae, among which Nannochloropsis oceanica, a marine alga widely used for aquaculture. In the last decade, N. oceanica has become a model organism for oleaginous microalgae to develop sustainable processes to produce biomolecules of interest by exploiting its photosynthetic activity and carbon assimilation properties. N. oceanica can accumulate lipids up to 70% of total dry weight and contains the omega-3 fatty acid eicosapentaenoic acid (EPA) required for both food and feed applications. The genome sequence, other omics data, and synthetic biology tools are available for this species, including an engineered strain called LP-tdTomato, which allows homologous recombination to insert the heterologous genes in a highly transcribed locus in the nucleolus region. Here, N. oceanica was engineered to induce high ketocarotenoid and canthaxanthin production., Results: We used N. oceanica LP-tdTomato strain as a background to express the key enzyme for ketocarotenoid production, a β-carotene ketolase (CrBKT) from Chlamydomonas reinhardtii. Through the LP-tdTomato strain, the transgene insertion by homologous recombination in a highly transcribed genomic locus can be screened by negative fluorescence. The overexpression of CrBKT in bkt transformants increased the content of carotenoids and ketocarotenoids per cell, respectively, 1.5 and 10-fold, inducing an orange/red color in the bkt cell cultures. Background (LP) and bkt lines productivity were compared at different light intensities from 150 to 1200 µmol m -2 s -1 : at lower irradiances, the growth kinetics of bkt lines were slower compared to LP, while higher productivity was measured for bkt lines at 1200 µmol m -2 s -1 . Despite these results, the highest canthaxanthin and ketocarotenoids productivity were obtained upon cultivation at 150 µmol m -2 s -1 ., Conclusions: Through targeted gene redesign and heterologous transformation, ketocarotenoids and canthaxanthin content were significantly increased, achieving 0.3% and 0.2% dry weight. Canthaxanthin could be produced using CO 2 as the only carbon source at 1.5 mg/L titer. These bkt-engineered lines hold potential for industrial applications in fish or poultry feed sectors, where canthaxanthin and ketocarotenoids are required as pigmentation agents., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Not applicable. Competing interests: Authors declare a competing financial interest: a patent application about the use of the BKT gene has been submitted by the University of Verona and granted at national level (Patent application n. 102021000027824 and PCT/IB2022/060381 “modified β-carotene ketolase (BKT), corresponding nucleic acid and microalgae strain comprising the same) having M.B. and S.C. among inventors. M.B. is the CEO and shareholder of asteasier srl, a company which could commercially exploit the results reported in this paper., (© 2024. The Author(s).)

Protein post-translational modifications (PTMs) regulate protein functions but remain poorly characterized in Nannochloropsis . This study examined three PTMs: lysine acetylation (Kac), lysine β-hydroxybutyrylation (Kbhb), and phosphorylation. Using LC-MS/MS, we identified 4571 Kac sites, 7812 Kbhb sites, and 6237 phosphorylation sites across 2455, 3109, and 2786 proteins, respectively. Subcellular localization analysis revealed significant overlaps between Kac and Kbhb proteins, primarily in the chloroplast, cytosol, and nucleus, while phosphorylated proteins were predominantly located in the nucleus and chloroplast. Motif analysis highlighted specific amino acid enrichments around modification sites, with several motifs conserved. Additionally, 529 proteins harbored all three PTMs, underscoring the potential regulatory interplay. Kac, Kbhb, and phosphorylated proteins were particularly abundant in glycolysis, the TCA cycle, carbon fixation, and lipid metabolism pathways, influencing energy production and lipid accumulation. Based on previous transcriptome data under nutrient-limited conditions, these frequently modified key enzymes appear to be vital components in the response to abiotic stress. The presence of histone modifications related to Kac and Kbhb might also point to the epigenetic regulation in gene expression and stress adaptation. This comprehensive PTM landscape in N. oceanica provides a foundation of valuable insights into future metabolic engineering and biotechnological applications.

In the present study, nine Enterobacteriaceae species present in wastewater were isolated and identified, and loop-mediated isothermal amplification (LAMP) was developed for the detection of Enterobacteriaceae by designing primers based on the mcr-1, KPC, OXA-23, and VIM genes, which are recognized markers of antimicrobial resistance (AMR) transmission during microalgal bioremediation treatment. The developed assays successfully detected four strains positive for mcr-1 gene-asociated resistance ( Acinetobacter baylyi , Klebsiella pneumoniae , Morganella morganii , and Serratia liquefaciens ), three strains for KPC gene-associated resistance ( Acinetobacter sp., Escherichia coli 15499, and Morganella morganii ), seven strains for OXA-23 gene-associated resistance ( Acinetobacter baylyi , Enterobacter hormaechi , Enterobacter cloacae , Escherichia coli 15922, Escherichia coli 51446, Morganella morganii , and Serratia liquefaciens ), and three strains for resistance to the VIM gene-associated resistance ( Acinetobacter baylyi , Acinetobacter sp., and Enterobacter hormaechi ) from a single colony. A reduction in microbiological load of 93.6% was achieved at 15 colony-forming units (CFU) mL -1 , utilizing EMB agar and LAMP values of 0.142 ± 0.011 for the mcr-1 gene, 0.212 ± 0.02 for the KPC gene, 0.233 ± 0.006 for the OXA-23 gene, and 0.219 ± 0.035 for the VIM gene. Furthermore, bioremediation efficiency values of 71.6% and 75% for total nitrogen and phosphorus, respectively, were observed at 72 h of treatment in open pond microalgal remediation systems (MRS). This study demonstrated that the LAMP technique is faster and more sensitive than traditional detection methods, such as CFU, for Enterobacteriaceae. Consequently, this method may be considered for the detection of microbiological quality indicators within the water treatment industry., Competing Interests: The authors declare that they have no competing interests., (© 2024 Cameron et al.)

Microalgae-bacteria consortia (MBC) system has been shown to enhance the efficiency of microalgae in wastewater treatment, yet its effectiveness in treating levofloxacin (LEV) wastewater remains unexplored. This study compared the treatment of LEV wastewater using pure Chlorella pyrenoidosa (PA) and its MBC constructed with activated sludge bacteria. The results showed that MBC improved the removal efficiency of LEV from 3.50-5.41 % to 33.62-57.20 % by enhancing the growth metabolism of microalgae. The MBC increased microalgae biomass and extracellular polymeric substance (EPS) secretion, yet reduced photosynthetic pigment content compared to the PA. At the phylum level, Proteobacteria and Actinobacteriota are the major bacteria in MBC. Furthermore, the transcriptome reveals that the growth-promoting effects of MBC are associated with the up-regulation of genes encoding the glycolysis, the citrate cycle (TCA cycle), and the pentose phosphate pathway. Enhanced carbon fixation, coupled with down-regulation of photosynthetic electron transfer processes, suggests an energy allocation mechanism within MBC. The up-regulation of porphyrin and arachidonic acid metabolism, along with the expression of genes encoding LEV-degrading enzymes, provides evidence of MBC's superior tolerance to and degradation of LEV. Overall, these findings lead to a better understanding of the underlying mechanisms through which MBC outperforms PA in treating LEV wastewater., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Microalgae-based glycolate production through the photorespiratory pathway is considered an environmentally friendly approach. However, the potential for glycolate production is limited by photoautotrophic cultivation with low cell density and existing strains. In this study, a targeted knockout approach was used to disrupt the key photorespiration enzyme, Chlamydomonas reinhardtii hydroxypyruvate reductase 1 (CrHPR1), leading to a significant increase in glycolate production of 280.1 mg/L/OD 750 . The highest potency yield reached 2.1 g/L under optimized mixotrophic conditions, demonstrating the possibility of synchronizing cell growth with glycolate biosynthesis in microalgae. Furthermore, the hypothesis that the cell wall-deficient mutant facilitates glycolate excretion was proposed and validated by comparing the glycolate accumulation trends of various Chlamydomonas reinhardtii strains. This study will facilitate the development of microalgae-based biotechnology and shed lights on the continuous advancement of green biomanufacturing for industrial application., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Monoclonal antibodies (mAbs) have become indispensable tools in various fields, from research to therapeutics, diagnostics, and industries. However, their production, primarily in mammalian cell culture systems, is cost-intensive and resource-demanding. Microalgae, diverse photosynthetic microorganisms, are gaining attention as a favorable option for manufacturing mAbs and various other recombinant proteins. This review explores the potential of microalgae as a robust expression system for biomanufacturing high-value proteins. It also highlights the diversity of microalgae species suitable for recombinant protein. Nuclear and chloroplast genomes of some microalgae have been engineered to express mAbs and other valuable proteins. Codon optimization, vector construction, and other genetic engineering techniques have significantly improved recombinant protein expression in microalgae. These accomplishments demonstrate the potential of microalgae for biopharmaceutical manufacturing. Microalgal biotechnology holds promise for revolutionizing the production of mAbs and other therapeutic proteins, offering a sustainable and cost-effective solution to address critical healthcare needs., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Given that crop yields are strongly limited by nitrogen, engineering crop plants with self-nitrogen-fertilization capacity holds great promise for sustainable agriculture. Recently, a nitrogen-fixing organelle has been characterized in the unicellular marine microalgae Braarudosphaera bigelowii. Engineering a nitrogen-fixing organelle into the non-nitrogen-fixing crops could benefit both environmental sustainability and global food security., Competing Interests: Declaration of interests The authors declare no conflicts of interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Since microalgae face various environmental stresses for the high production of biofuels, multiple studies have been performed to determine if microalgae are resistant to these various stresses. In this study, the viability of cells under various abiotic stresses was investigated by introducing a putative germin-like protein (GLP) from Neopyropia yezoensis , which was known to be related in the resistance to abiotic stresses. The expression of GLP in Chlamydomonas reinhardtii allowed cells to grow better in various abiotic stress environments. In nitrogen starvation conditions, recombinant cells accumulated the lipid droplet 1.46-fold more than wild-type cells and responded more rapidly to form palmelloid forms. Under high-temperature, hydrogen peroxide conditions and saline stress, the survival rate was increased 3.5 times, 2.19 times, and 3.19 times in recombinant C. reinhardtii with GLP, respectively. The expression level of genes related to pathways in response to various stresses increased 2-fold more under those conditions. This result will be useful for the development of microalgae that can grow better and produce more biofuels under different stress conditions.

The phenolic antioxidant 2,6-Di-tert-butyl-p-cresol (BHT) has been detected in various environments and is considered a potential threat to aquatic organisms. Algal-bacterial interactions are crucial for maintaining ecosystem balance and elemental cycling, but their response to BHT remains to be investigated. This study analyzed the physiological and biochemical responses of the microalga Isochrysis galbana and the changes of associated bacterial communities under different concentrations of BHT stress. Results showed that the biomass of I. galbana exhibited a decreasing trend with increasing BHT concentrations up to 40 mg/L. The reduction in chlorophyll, carotenoid, and soluble protein content of microalgal cells was also observed under BHT stress. The production of malondialdehyde and the activities of superoxide dismutase, peroxidase, and catalase were further determined. Scanning electron microscopy analysis revealed that BHT caused surface rupture of the algal cells and loss of intracellular nutrients. Proteomic analysis demonstrated the upregulation of photosynthesis and citric acid cycle pathways as a response to BHT stress. Additionally, BHT significantly increased the relative abundance of specific bacteria in the phycosphere, including Marivita, Halomonas, Marinobacter, and Alteromonas. Further experiments confirmed that these bacteria had the ability to utilize BHT as the sole carbon resource for growth, and genes related to the degradation of phenolic compounds were detected through pangenome analysis., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

DpAP2 is a transcription factor regulating carotenoid biosynthesis pathway. It was speculated that MeJA significantly decreased expression of DpAP2 gene, then the decreasing DpAP2 expression significantly inhibited expression of some key enzyme genes such as PSY, PDS and GGPS in carotenoid biosynthesis pathway. In contrast, it was speculated that GA3 significantly increased expression of DpAP2 gene, then the increasing DpAP2 expression significantly increased expression of some key enzyme genes such as PDS and GGPS in carotenoid biosynthesis pathway. To increase the content of carotenoid, we evaluated the effect of DpAP2 overexpression on carotenoid accumulation in D. parva. Transgenic D. parva showed a higher carotenoid content (3.18 mg/g DW) compared with control group (2.13 mg/g DW) at 9 d. The dosage effects of exogenous hormones MeJA and GA3 were found in D. parva cells treated with different concentrations of MeJA (10, 20, 50, 100 μM) and GA3 (10, 20, 50, 100 μM). The high concentrations of MeJA (10-100 μM) inhibited the accumulation of carotenoid, and the relative expression of DpAP2, PSY, PDS and GGPS decreased significantly. On the contrary, the relative expression of DpAP2, PDS and GGPS increased significantly when D. parva was treated with 10, 20, 50 and 100 μM GA3, which promoted the biosynthesis of carotenoid. Therefore, we inferred that there was a hierarchical regulation from hormone, transcription factor, key enzyme gene to carotenoid accumulation in carotenoid biosynthesis. Carotenoid biosynthesis was enhanced by DpAP2 overexpression (1.4930 fold of control) and exogenous substances such as GA3 (1.5889 fold of control), which laid a foundation for massive accumulation of carotenoids in microalgae. In the future, further studies were required to demonstrate the complex regulatory network., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Ruan et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Microalgae are significantly influenced by light quality and quantity, whether in their natural habitats or under laboratory and industrial culture conditions. The present study examines the adaptive responses of the marine microalga Tetraselmis chuii to different light regimes, using a cost-effective filtering method and a multi-omics approach. Microalgal growth rates were negatively affected by all filtered light regimes. After six days of cultivation, growth rate for cultures exposed to blue and green filtered light was 67%, while for red filter was 83%, compared to control cultures. Transcriptomic analysis revealed that the usage of green filters resulted in upregulation of transcripts involved in ribosome biogenesis or coding for elongation factors, exemplified by a 2.3-fold increase of TEF3. On the other hand, a 2.7-fold downregulation was observed in photosynthesis-related petJ. Exposure to blue filtered light led to the upregulation of transcripts associated with pyruvate metabolism, while photosynthesis was negatively impacted. In contrast, application of red filter induced minor transcriptomic alterations. Regarding metabolomic analysis, sugars, amino acids, and organic acids exhibited significant changes under different light regimes. For instance, under blue filtered light sucrose accumulated over 6-fold, while aspartic acid content decreased by 4.3-fold. Lipidomics analysis showed significant accumulation of heptadecanoic and linoleic acids under green and red light filters. Together, our findings indicate that filter light can be used for targeted metabolic manipulation., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Chernozem soil is a valuable resource and contains a great diversity of microorganisms that play a global role in the process of soil formation, the species diversity of which has changed over the last five years under the influence of different agrotechnologies. For the first time, under the conditions of the Central Chernozem region, grain and fallow crop rotation, studies using the DNA-metabarcoding method were carried out to study the taxonomic structure of bacteria, fungi, cyanobacteria, and microalgae communities in the arable horizon of typical medium loamy chernozem under winter wheat cultivation. A comparative analysis of the composition of the genotypes showed significant differences in the presented level of mineral nutrition of the soil NPK (60) and NPK (100) compared with the control variant. After processing the 16S and 18S rRNA datasets, a similar trend of decreasing numbers of pro- and eukaryotic species was found from 6296 (control without MF) to 5310 with NPK (60) and to 4643 with NPK (100), respectively. The Chao1 index indicated that the expected diversity within the prokaryotic group was higher in the control without MF at 211, but decreased to 182 and 193 with NPK (60) and NPK (100) fertilizers, respectively. Analysis of the eukaryotic group revealed a 2.6- and 2.9-fold decrease in diversity by class and genus, respectively, depending on the nutritional levels in agrotechnologies, owing to the use of MF. In the prokaryotic community, Alphaproteobacteria microorganisms predominated at an amount of 14.20-14.46%, with Cyanophyceae accounting for 5.2-9.9%. The diversity of eukaryotes was smaller than the number of classes of prokaryotes; the main dominant were Zygnematophyceae 19.5-41%, Chlorophyceae occupied 10.4-15.8%. On the other hand, the doses of fertilizers used contributed to the emergence of dominant species adapted to high doses of mineral nutrients for plants., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Main Conclusion: PPI analysis deepens our knowledge in critical processes like carbon fixation and nutrient sensing. Moreover, signaling networks, including pathways like MAPK/ERK and TOR, provide valuable information in how microalgae respond to environmental changes and stress. Additionally, species-species interaction networks for microalgae provide a comprehensive understanding of how different species interact within their environments. This review examines recent advancements in the study of biological networks within microalgae, with a focus on the intricate interactions that define these organisms. It emphasizes how network biology, an interdisciplinary field, offers valuable insights into microalgae functions through various methodologies. Crucial approaches, such as protein-protein interaction (PPI) mapping utilizing yeast two-hybrid screening and mass spectrometry, are essential for comprehending cellular processes and optimizing functions, such as photosynthesis and fatty acid biosynthesis. The application of advanced computational methods and information mining has significantly improved PPI analysis, revealing networks involved in critical processes like carbon fixation and nutrient sensing. The review also encompasses transcriptional networks, which play a role in gene regulation and stress responses, as well as metabolic networks represented by genome-scale metabolic models (GEMs), which aid in strain optimization and the prediction of metabolic outcomes. Furthermore, signaling networks, including pathways like MAPK/ERK and TOR, are crucial for understanding how microalgae respond to environmental changes and stress. Additionally, species-species interaction networks for microalgae provide a comprehensive understanding of how different species interact within their environments. The integration of these network biology approaches has deepened our understanding of microalgal interactions, paving the way for more efficient cultivation and new industrial applications., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The application of the fermentative bacteria and microalgae coupling system in the wastewater treatment has been studied, but there remains few knowledge regarding the organic and inorganic carbon metabolism within this system. In this study, the carbon metabolism of microalgae and fermentative bacteria was elucidated by 13 C stable isotope labeling and functional gene prediction, respectively. The 13 C glucose and 13 C NaHCO 3 were used as stable isotope tracers to clarify the organic and inorganic carbon metabolism of microalgae, indicating that approximately 71.5 % of the Acetyl-CoA in microalgae was synthesized from organic carbon sources, while 26.8 % was synthesized through the utilization of inorganic carbon sources. Inorganic carbon sources can enhance the activity of photosynthetic system and facilitate the Calvin cycle. Considering the adequate organic carbon sources and insufficient inorganic carbon sources in the fermentative bacteria and microalgae coupling system, NaHCO 3 was added to improve carbon utilization of microalgae. The maximum microalgal lipid yield reached 1130.37 mg/L with 1000 mg/L NaHCO 3 supplementation. Functional gene prediction was used to analysis the effect of various carbon composition on the bacterial carbon metabolism. Notably, the additional inorganic carbon sources increased the abundance of bacterial functional genes associated with the fermentation and acetic acids synthesis, which was advantageous for VFAs production and further promoted microalgae growth. This study can gain a deeper understanding of microbial metabolic mechanisms during the operation of fermentative bacteria and microalgae system, and improve its sustained operational stability., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Lincomycin (LIN) in pharmaceutical wastewater would enter municipal wastewater treatment plants and decrease their performance, leading to residual LIN enter the natural environment and pose serious eco-risk. In this study, a bacterium-microalgae consortium co-immobilized filter (BMCCF) was established and used to remove LIN in artificial pharmaceutical wastewater treatment plants effluents (PWWTPE). LIN removal mechanisms and degradation products' eco-toxicity was studied, and the abundance change of class 1 integrase gene (intI1) and antibiotic resistance genes (ARGs) was monitored. As a result, 98.54% of 82 mg L -1 LIN was removed within 7 days, LIN removal was mainly attributed to bio-degradation by the Bacillus subtilis strain, and LIN degradation products were less toxic than their substrate. Therefore, the BMCCF established in this study provides a promising alternative for the bio-treatment of pharmaceutical wastewater containing high concentration of LIN., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Microalgae, with the ability to harness solar energy to fix CO 2 and convert it into organic compounds, have emerged as promising green cell factories. With the rapid development of cutting-edge biotechnologies, the research and application of photosynthetic microalgae have been expanding, leading to comprehensive and in-depth engineering of microalgae. The synthetic biology and genome editing technologies have enabled the applications of microalgae in medicine, agriculture, food, energy, and the environment. However, the survival and spreading of engineered microalgae in the natural environment pose potential safety risks to ecosystems and human health. To curb the risks caused by the spreading of engineered microalgae in the environment, biosafety policies should be formulated for engineered microalgae and the prevention and control technologies should be developed. Toward this goal, researchers have developed biocontainment systems, including positive strategies such as the design of toxic protein-based kill switches and passive strategies such as knocking out essential genes to construct the strains with nutritional deficiencies, thereby spatially containing engineered microalgae. This article summarizes the application of cutting-edge biotechnologies in the engineering of microalgae, the biosafety risks and management regulations associated with the escape of engineered microalgae, and the progress in novel biocontainment technologies established for engineered microalgae. Finally, this article gives insights into the future development direction of microalgae biocontainment.

Euglena gracilis is a unique microalga that lacks a cell wall and is able to grow under different trophic culture conditions. In this study, cell growth, biomass production, and changes in the ultrastructure of E. gracilis cells cultivated photoautotrophically, mixotrophically, and under sequential-heterotrophy-photoinduction (SHP) were assessed. Mixotrophy induced the highest cell growth and biomass productivity (6.27 ± 0.59 mg/L/d) in E. gracilis, while the highest content of fatty acids, 2.69 ± 0.04% of dry cell weight (DCW) and amino acids, 38.16 ± 0.08% of DCW was obtained under SHP condition. E. gracilis also accumulated significantly higher saturated fatty acids and lower unsaturated fatty acids when cultivated under SHP condition. Transcriptomic analysis showed that the expression of photosynthetic genes (PsbA, PsbC, F-type ATPase alpha and beta) was lower, carbohydrate and protein synthetic genes (glnA, alg14 and fba) were expressed higher in SHP-culture cells when compared to other groups. Different trophic conditions also induced changes in the cell ultrastructure, where paramylon and starch granules were more abundant in SHP-cultured cells. The findings generated in this study illustrated that aerobic SHP cultivation of E. gracilis possesses great potential in human and animal feed applications., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Oxidative stress is a universal interpretation for the toxicity mechanism of nanoplastics to microalgae. However, there is a lack of deeper insight into the regulation mechanism in microalgae response to oxidative stress, thus affecting the prevention and control for nanoplastics hazard. The integrated analysis of transcriptomics and metabolomics was employed to investigate the mechanism for the oxidative stress response of Chlorella pyrenoidosa to nanoplastics and subsequently lock the according core pathways and driver genes induced. Results indicated that the linoleic acid metabolism, glycine (Gly)-serine (Ser)-threonine (Thr) metabolism, and arginine and proline metabolism pathways of C. pyrenoidosa were collectively involved in oxidative stress. The analysis of linoleic acid metabolism suggested that nanoplastics prompted algal cells to secrete more allelochemicals, thereby leading to destroy the immune system of cells. Gly-Ser-Thr metabolism and arginine and proline metabolism pathways were core pathways involved in algal regulation of cell membrane function and antioxidant system. Key genes, such as LOX2.3, SHM1, TRPA1, and proC1, are drivers of regulating the oxidative stress of algae cells. This investigation lays the foundation for future applications of gene editing technology to limit the hazards of nanoplastics on aquatic organism., Competing Interests: Declaration of Competing interest This paper is our unpublished work and it has not been submitted to any other journal for reviews. Thus, it has not been previously published, in whole or in part, and it is not under consideration by any other journal. Additionally, I confirm that both authors are aware of, and accept responsibility for, the manuscript. There is no conflict of interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

L-Lactate is a commodity chemical used in various fields. Microorganisms have produced L-lactate via lactic fermentation using saccharides derived from crops as carbon sources. Recently, L-lactate production using microalgae, whose carbon source is carbon dioxide, has been spotlighted because the prices of the crops have increased. A red alga Cyanidioschyzon merolae produce L-lactate via lactic fermentation under dark anaerobic conditions. The L-lactate titer of C. merolae is higher than those of other microalgae but lower than those of heterotrophic bacteria. Therefore, an increase in the L-lactate titer is required in C. merolae. L-Lactate dehydrogenase (L-LDH) catalyzes the reduction of pyruvate to L-lactate during lactic fermentation. C. merolae possesses five isozymes of L-LDH. The results of previous transcriptome analysis suggested that L-LDHs are the key enzymes in the lactic fermentation of C. merolae. However, their biochemical characteristics, such as catalytic efficiency and tolerance for metabolites, have not been revealed. We compared the amino acid sequences of C. merolae L-LDHs (CmLDHs) and characterized one of the isozymes, CmLDH1. BLAST analysis revealed that the sequence similarities of CmLDH1 and the other isozymes were above 99%. The catalytic efficiency of CmLDH1 under its optimum conditions was higher than those of L-LDHs of other organisms. ATP decreased the affinity and turnover number of CmLDH1 for NADH. These findings contribute to understanding the characteristics of L-LDHs of microalgae and the regulatory mechanisms of lactic fermentation in C. merolae., (© 2024. The Author(s).)

Marine microbes that have for eons been adapted to stable salinity regimes are confronted with sudden decreases in salinity in the Arctic Ocean. The episodic freshening is increasing due to climate change with melting multi-year sea-ice and glaciers, greater inflows from rivers, and increased precipitation. To investigate algal responses to lowered salinity, we analyzed the responses and acclimatation over 24 h in a non-model Arctic marine alga (pelagophyte CCMP2097) following transfer to realistic lower salinities. Using RNA-seq transcriptomics, here we show rapid differentially expressed genes related to stress oxidative responses, proteins involved in the photosystem and circadian clock, and those affecting lipids and inorganic ions. After 24 h the pelagophyte adjusted to the lower salinity seen in the overexpression of genes associated with freezing resistance, cold adaptation, and salt tolerance. Overall, a suite of ancient widespread pathways is recruited enabling the species to adjust to the stress of rapid salinity change., (© 2024. The Author(s).)

Dunaliella salina is an innovative expression system due to its distinct advantages such as high salt tolerance, low susceptibility to contamination, and the absence of the cell wall. While nuclear transformation has been extensively studied, research on D. salina chloroplast transformation remains in the preliminary stages. In this study, we established an efficient chloroplast expression system for D. salina using Golden Gate assembly. We developed a D. salina toolkit comprising essential components such as chloroplast-specific promoters, terminators, homologous fragments, and various vectors. We confirmed its functionality by expressing the EGFP protein. Moreover, we detailed the methodology of the entire construction process. This expression system enables the specific targeting of foreign genes through simple homologous recombination, resulting in stable expression in chloroplasts. The toolkit achieved a relatively high transformation efficiency within a shorter experimental cycle. Consequently, the construction and utilization of this toolkit have the potential to enhance the efficiency of transgenic engineering in D. salina and advance the development of microalgal biofactories., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

In the current study, systems biology approach was applied to get a deep insight regarding the regulatory mechanisms of Chromochloris zofingiensis under overall stress conditions. Meta-analysis was performed using p-values combination of differentially expressed genes. To identify the informative models related to stress conditions, two distinct weighted gene co-expression networks were constructed and preservation analyses were performed using medianRankand Zsummary algorithms. Moreover, functional enrichment analysis of non-preserved modules was performed to shed light on the biological performance of underlying genes in the non-preserved modules. In the next step, the gene regulatory networks between top hub genes of non-preserved modules and transcription factors were inferred using ensemble of trees algorithm. Results showed that the power of beta = 7 was the best soft-thresholding value to ensure a scale-free network, leading to the determination of 12 co-expression modules with an average size of 128 genes. Preservation analysis showed that the connectivity pattern of the six modules including the blue, black, yellow, pink, greenyellow, and turquoise changed during stress condition which defined as non-preserved modules. Examples of enriched pathways in non-preserved modules were Oxidative phosphorylation", "Vitamin B6 metabolism", and "Arachidonic acid metabolism". Constructed regulatory network between identified TFs and top hub genes of non-preserved module such as Cz06g10250, Cz03g12130 showed that some specific TFs such as C3H and SQUAMOSA promoter binding protein (SBP) specifically regulates the specific hubs. The current findings add substantially to our understanding of the stress responsive underlying mechanism of C. zofingiensis for future studies and metabolite production programs., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Bahman Panahi. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The catastrophic loss of aquatic life in the Central European Oder River in 2022, caused by a toxic bloom of the haptophyte microalga Prymnesium parvum (in a wide sense, s.l.), underscores the need to improve our understanding of the genomic basis of the toxin. Previous morphological, phylogenetic, and genomic studies have revealed cryptic diversity within P. parvum s.l. and uncovered three clade-specific (types A, B, and C) prymnesin toxins. Here, we used state-of-the-art long-read sequencing and assembled the first haplotype-resolved diploid genome of a P. parvum type B from the strain responsible for the Oder disaster. Comparative analyses with type A genomes uncovered a genome-size expansion driven by repetitive elements in type B. We also found conserved synteny but divergent evolution in several polyketide synthase (PKS) genes, which are known to underlie toxin production in combination with environmental cues. We identified an approximately 20-kbp deletion in the largest PKS gene of type B that we link to differences in the chemical structure of types A and B prymnesins. Flow cytometry and electron microscopy analyses confirmed diploidy in the Oder River strain and revealed differences to closely related strains in both ploidy and morphology. Our results provide unprecedented resolution of strain diversity in P. parvum s.l. and a better understanding of the genomic basis of toxin variability in haptophytes. The reference-quality genome will enable us to better understand changes in microbial diversity in the face of increasing environmental pressures and provides a basis for strain-level monitoring of invasive Prymnesium in the future., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: The steady world population growth and the current climate emergency crisis demand the development of sustainable methods to increase crop performance and resilience to the abiotic and biotic stresses produced by global warming. Microalgal extracts are being established as sustainable sources to produce compounds that improve agricultural yield, concurrently contributing during their production process to atmospheric CO 2 abatement through the photosynthetic activity of microalgae., Results: In the present study, we characterize the transcriptomic response in the model plant Arabidopsis thaliana and the plant of horticultural interest Solanum lycopersicum to the foliar application of a microalgae-based commercial preparation LRM™ (AlgaEnergy, Madrid, Spain). The foliar spray of LRM™ has a substantial effect over both transcriptomes potentially mediated by various compounds within LRM™, including its phytohormone content, activating systemic acquired resistance, possibly mediated by salicylic acid biosynthetic processes, and drought/heat acclimatization, induced by stomatal control and wax accumulation during cuticle development. Specifically, the agronomic improvements observed in treated S. lycopersicum (tomato) plants include an increase in the number of fruits, an acceleration in flowering time and the provision of higher drought resistance. The effect of LRM™ foliar spray in juvenile and adult plants was similar, producing a fast response detectable 2 h from its application that was also maintained 24 h later., Conclusion: The present study improves our knowledge on the transcriptomic effect of a novel microalgal extract on crops and provides the first step towards a full understanding of the yield and resistance improvement of crops. © 2024 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry., (© 2024 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.)

Ubiquitous distribution of pharmaceutical contaminants in environment has caused unexpected adverse effects on ecological organisms; however, how microorganisms recover from their toxicities remains largely unknown. In this study, we comprehensively investigated the effect of a representative pollutant, doxylamine (DOX) on a freshwater microalgal species, Chlorella sp. by analyzing the growth patterns, biochemical changes (total chlorophyll, carotenoid, carbohydrate, protein, and antioxidant enzymes), and transcriptomics. We found toxicity of DOX on Chlorella sp. was mainly caused by disrupting synthesis of ribosomes in nucleolus, and r/t RNA binding and processing. Intriguingly, additional bicarbonate enhanced the toxicity of DOX with decreasing the half-maximum effective concentrations from 15.34 mg L -1 to 4.63 mg L -1 , which can be caused by inhibiting fatty acid oxidation and amino acid metabolism. Microalgal cells can recover from this stress via upregulating antioxidant enzymatic activities to neutralize oxidative stresses, and photosynthetic pathways and nitrogen metabolism to supply more energies and cellular signaling molecules. This study extended our understanding on how microalgae can recover from chemical toxicity, and also emphasized the effect of environmental factors on the toxicity of these contaminants on aquatic microorganisms., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The enzyme phospholipase A 2 (PLA 2 ) plays a crucial role in acyl remodeling of phospholipids via the Lands' cycle, and consequently alters fatty acid compositions in triacylglycerol (TAG). In this study, a full-length cDNA sequence coding Myrmecia incisa phospholipase A 2 (MiPLA 2 ) was cloned using the technique of rapid amplification of cDNA ends. Comparison of the 1082-bp cDNA with its corresponding cloned DNA sequence revealed that MiPLA 2 contained 3 introns. Mature MiPLA 2 (mMiPLA 2 ) had a conserved Ca 2+ -binding loop and a catalytic site motif that has been recognized in plant secretory PLA 2 (sPLA 2 ) proteins. Correspondingly, phylogenetic analysis illustrated that MiPLA 2 was clustered within GroupXIA of plant sPLA 2 proteins. To ascertain the function of MiPLA 2 , the cDNA coding for mMiPLA 2 was subcloned into the vector pET-32a to facilitate the production of recombinant mMiPLA 2 in Escherichia coli. Recombinant mMiPLA 2 was purified and used for the in vitro enzyme reaction. Thin-layer chromatography profiles of the catalytic products generated by recombinant mMiPLA 2 indicated a specificity for cleaving sn-2 acyl chains from phospholipids, thereby functionally characterizing MiPLA 2 . Although recombinant mMiPLA 2 displayed a strong preference for phosphatidylethanolamine, it preferentially hydrolyzes arachidonic acid (ArA) at the sn-2 position of phosphatidylcholine. Results from the fused expression of p1300-sp-EGFP-mMiPLA 2 illustrated that MiPLA 2 was localized in the intercellular space of onion epidermis. Furthermore, the positive correlation between MiPLA 2 transcription and free ArA levels were established. Consequently, the role of mMiPLA 2 in the biosynthesis of ArA-rich TAG was elucidated. This study helps to understand how M. incisa preferentially uses ArA to synthesize TAG., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Masson SAS.)

Omega-3 polyunsaturated fatty acids (PUFAs), such as eicosapentaenoic acid (EPA; C20:5n-3) and docosahexaenoic acid (DHA; C22:6n-3) are widely used as additives in fish feed in the aquaculture sector. To date, the supply of omega-3 PUFAs have heavily depended upon fish oil production. As the need for omega-3 PUFAs supply for the growing population increases, a more sustainable approach is required to keep up with the demand. The oleaginous diatom Fistulifera solaris is known to synthesize EPA with the highest level among autotrophically cultured microalgae, however, this species does not accumulate significant amounts of DHA, which, in some cases, is required in aquaculture rather than EPA. This is likely due to the lack of expression of essential enzymes namely Δ5 elongase (Δ5ELO) and Δ4 desaturase. In this study, we identified endogenous Δ5ELO genes in F. solaris and introduced recombinant expression cassettes harboring Δ5ELO into F. solaris through bacterial conjugation. As a result, it managed to induce the synthesis of docosapentaenoic acid (DPA; C22:5n-3), a direct precursor of DHA. This study paves the way for expanding our understanding of the omega-3 PUFAs pathway using endogenous genes in the oleaginous diatom., (Copyright © 2024 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.)

The lag phase is key in resuming bacterial growth, but it remains underexplored particularly in environmental bacteria. Here we use transcriptomics and 13 C-labelled metabolomics to show that the lag phase of the model marine bacterium Phaeobacter inhibens is shortened by methylated compounds produced by the microalgal partner, Emiliania huxleyi. Methylated compounds are abundantly produced and released by microalgae, and we show that their methyl groups can be collected by bacteria and assimilated through the methionine cycle. Our findings underscore the significance of methyl groups as a limiting factor during the lag phase and highlight the adjustability of this growth phase. In addition, we show that methylated compounds, typical of photosynthetic organisms, prompt diverse reductions in lag times in bacteria associated with algae and plants, potentially favouring early growth in some bacteria. These findings suggest ways to accelerate bacterial growth and underscore the significance of studying bacteria within an environmental context., (© 2024. The Author(s).)

The production of biologics in mammalian cells is hindered by some limitations including high production costs, prompting the exploration of other alternative expression systems that are cheaper and sustainable like microalgae. Successful productions of biologics such as monoclonal antibodies have already been demonstrated in the diatom Phaeodactylum tricornutum ; however, limited production yields still remain compared to mammalian cells. Therefore, efforts are needed to make this microalga more competitive as a cell biofactory. Among the seventeen reported accessions of P. tricornutum , ten have been mainly studied so far. Among them, some have already been used to produce high-value-added molecules such as biologics. The use of "omics" is increasingly being described as useful for the improvement of both upstream and downstream steps in bioprocesses using mammalian cells. Therefore, in this context, we performed an RNA-Seq analysis of the ten most used P. tricornutum accessions (Pt1 to Pt10) and deciphered the differential gene expression in pathways that could affect bioproduction of biologics in P. tricornutum . Our results highlighted the benefits of certain accessions such as Pt9 or Pt4 for the production of biologics. Indeed, these accessions seem to be more advantageous. Moreover, these results contribute to a better understanding of the molecular and cellular biology of P. tricornutum .

Carotenoids, with their diverse biological activities and potential pharmaceutical applications, have garnered significant attention as essential nutraceuticals. Microalgae, as natural producers of these bioactive compounds, offer a promising avenue for sustainable and cost-effective carotenoid production. Despite the ability to cultivate microalgae for its high-value carotenoids with health benefits, only astaxanthin and β-carotene are produced on a commercial scale by Haematococcus pluvialis and Dunaliella salina, respectively. This review explores recent advancements in genetic engineering and cultivation strategies to enhance the production of lutein by microalgae. Techniques such as random mutagenesis, genetic engineering, including CRISPR technology and multi-omics approaches, are discussed in detail for their impact on improving lutein production. Innovative cultivation strategies are compared, highlighting their advantages and challenges. The paper concludes by identifying future research directions, challenges, and proposing strategies for the continued advancement of cost-effective and genetically engineered microalgal carotenoids for pharmaceutical applications.

Epitranscriptomics is considered as a new regulatory step in eukaryotes for developmental processes and stress responses. The aim of this study was, for the first time, to identify RNA methyltransferase (writers) and demethylase (erasers) in four investigated species, i.e., the dinoflagellates Alexandrium tamutum and Amphidinium carterae , the diatom Cylindrotheca closterium , and the green alga Tetraselmis suecica . As query sequences for the enzymatic classes of interest, we selected those ones that were previously detected in marine plants, evaluating their expression upon nutrient starvation stress exposure. The hypothesis was that upon stress exposure, the activation/deactivation of specific writers and erasers may occur. In microalgae, we found almost all plant writers and erasers (ALKBH9B, ALKBH10B, MTB, and FIP37), except for three writers (MTA, VIRILIZER, and HAKAI). A sequence similarity search by scanning the corresponding genomes confirmed their presence. Thus, we concluded that the three writer sequences were lacking from the studied transcriptomes probably because they were not expressed in those experimental conditions, rather than a real lack of these genes from their genomes. This study showed that some of them were expressed only in specific culturing conditions. We also investigated their expression in other culturing conditions (i.e., nitrogen depletion, phosphate depletion, and Zinc addition at two different concentrations) in A. carterae , giving new insights into their possible roles in regulating gene expression upon stress.

Microalgal biomass for biofuel production, integration into functional food, and feed supplementation has generated substantial interest worldwide due to its high growth rate, non-competitiveness for agronomic land, ease of cultivation in containments, and presence of several bioactive molecules. In this study, genetic engineering tools were employed to develop transgenic lines of freshwater microalga Chlorella vulgaris with a higher starch content, by up-regulating ADP-glucose pyrophosphorylase (AGPase), which is a rate-limiting enzyme in starch biosynthesis. Expression of the Escherichia coli glgC (AGPase homolog) gene in C. vulgaris led to an increase in total carbohydrate content up to 45.1% (dry cell weight, DCW) in the transgenic line as compared to 34.2% (DCW) in the untransformed control. The starch content improved up to 16% (DCW) in the transgenic alga compared to 10% (DCW) in the control. However, the content of total lipid, carotenoid, and chlorophyll decreased differentially in the transgenic lines. The carbohydrate-rich biomass from the transgenic algal line was used to produce bioethanol via yeast fermentation, which resulted in a higher ethanol yield of 82.82 mg/L as compared to 54.41 mg/L from the untransformed control. The in vitro digestibility of the transgenic algal starch revealed a resistant starch content of up to 7% of total starch. Faster growth of four probiotic bacterial species along with a lowering of the pH of the growth medium indicated transgenic alga to exert a positive prebiotic effect. Taken together, the study documents the utilization of genetically engineered C. vulgaris with enriched carbohydrates as bioethanol feedstock and functional food ingredients., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

The CCT (CO, COL and TOC1) gene family has been elucidated to be involved in the functional differentiation of the products in various plant species, but their specific mechanisms are poorly understood. In the present investigation, we conducted a genome-wide identification and phylogenetic analysis of CCT genes from microalgae to legumes. A total of 700 non-redundant members of the CCT gene family from 30 species were identified through a homology search. Phylogenetic clustering with Arabidopsis and domain conservation analysis categorized the CCT genes into three families. Multiple sequence alignment showed that the CCT domain contains important amino acid residues, and each CCT protein contains 24 conserved motifs, as demonstrated by the motif analysis. Whole-genome/segment duplication, as well as tandem duplication, are considered to be the driving forces in the evolutionary trajectory of plant species. This comprehensive investigation into the proliferation of the CCT gene family unveils the evolutionary dynamics whereby WGD/segment duplication is the predominant mechanism contributing to the expansion of the CCT genes. Meanwhile, the examination of the gene expression patterns revealed that the expression patterns of CCT genes vary in different tissues and at different developmental stages of plants, with high expression in leaves, which is consistent with the molecular regulation of flowering in photosynthesis by CCT. Based on the protein-protein interaction analysis of CCT genes in model plants, we propose that the CCT gene family synergistically regulates plant development and flowering with light-signaling factors (PHYs and PIFs) and MYB family transcription factors. Understanding the CCT gene family's molecular evolution enables targeted gene manipulation for enhanced plant traits, including optimized flowering and stress resistance., Competing Interests: The authors declare no conflicts of interest.

Euglena gracilis (E. gracilis), pivotal in the study of photosynthesis, endosymbiosis, and chloroplast development, is also an industrial microalga for paramylon production. Despite its importance, E. gracilis genome exploration faces challenges due to its intricate nature. In this study, we achieved a chromosome-level de novo assembly (2.37 Gb) using Illumina, PacBio, Bionano, and Hi-C data. The assembly exhibited a contig N50 of 619 Kb and scaffold N50 of 1.12 Mb, indicating superior continuity. Approximately 99.83% of the genome was anchored to 46 chromosomes, revealing structural insights. Repetitive elements constituted 58.84% of the sequences. Functional annotations were assigned to 39,362 proteins, enhancing interpretative power. BUSCO analysis confirmed assembly completeness at 80.39%. This first high-quality E. gracilis genome offers insights for genetics and genomics studies, overcoming previous limitations. The impact extends to academic and industrial research, providing a foundational resource., (© 2024. The Author(s).)

Increasing antivirals in surface water caused by their excessive consumption pose serious threats to aquatic organisms. Our recent research found that the input of antiviral drug arbidol to algal bloom water can induce acute toxicity to the growth and metabolism of Microcystis aeruginosa, resulting in growth inhibition, as well as decrease in chlorophyll and ATP contents. However, the toxic mechanisms involved remained obscure, which were further investigated through transcriptomic analysis in this study. The results indicated that 885-1248 genes in algae were differentially expressed after exposure to 0.01-10.0 mg/L of arbidol, with the majority being down-regulated. Analysis of commonly down-regulated genes found that the cellular response to oxidative stress and damaged DNA bonding were affected, implying that the stress defense system and DNA repair function of algae might be damaged. The down-regulation of genes in porphyrin metabolism, photosynthesis, carbon fixation, glycolysis, tricarboxylic acid cycle, and oxidative phosphorylation might inhibit chlorophyll synthesis, photosynthesis, and ATP supply, thereby hindering the growth and metabolism of algae. Moreover, the down-regulation of genes related to nucleotide metabolism and DNA replication might influence the reproduction of algae. These findings provided effective strategies to elucidate toxic mechanisms of contaminants on algae in algal bloom water., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The phycosphere is an essential ecological niche for the proliferation of antibiotic resistance genes (ARGs). However, how ARGs' potential hosts change and the driving mechanism of metabolites under antibiotic stress in the phycosphere have seldom been researched. We investigated the response of Chlorella pyrenoidosa and the structure and abundance of free-living (FL) and particle-attached (PA) bacteria, ARGs, and metabolites under sulfadiazine by using real-time quantitative PCR, 16 S rRNA high-throughput. The linkage of key bacterial communities, ARGs, and metabolites through correlations was established. Through analysis of physiological indicators, Chlorella pyrenoidosa displayed a pattern of "low-dose promotion and high-dose inhibition" under antibiotic stress. ARGs were enriched in the PA treatment groups by 117 %. At the phylum level, Proteobacteria, Bacteroidetes, and Actinobacteria as potential hosts for ARGs. At the genus level, potential hosts included Sphingopyxis, SM1A02, Aquimonas, Vitellibacter, and Proteiniphilum. Middle and high antibiotic concentrations induced the secretion of metabolites closely related to potential hosts by algae, such as phytosphingosine, Lysophosphatidylcholine, and α-Linolenic acid. Therefore, changes in bacterial communities indirectly influenced the distribution of ARGs through alterations in metabolic products. These findings offer essential details about the mechanisms behind the spread and proliferation of ARGs in the phycosphere., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Microalgae are considered to be more useful and effective to use in biomass production than other photosynthesis organisms. However, microalgae need to be altered to acquire more desirable traits for the relevant purpose. Although neutron radiation is known to induce DNA mutations, there have been few studies on its application to microalgae, and the optimal relationship between irradiation intensity and mutation occurrence has not been established. In this study, using the unicellular red alga Cyanidioschyzon merolae as a model, we analyzed the relationship between the absorbed dose of two types of neutrons, high-energy (above 1 MeV) and thermal (around 25 meV) neutrons, and mutation occurrence while monitoring mutations in URA5.3 gene encoding UMP synthase. As a result, the highest mutational occurrence was observed when the cells were irradiated with 20 Gy of high-energy neutrons and 13 Gy of thermal neutrons. Using these optimal neutron irradiation conditions, we next attempted to improve the lipid accumulation of Euglena gracilis, which is a candidate strain for biofuel feedstock production. As a result, we obtained several strains with a maximum 1.3-fold increase in lipid accumulation compared with the wild-type. These results indicate that microalgae breeding by neutron irradiation is effective., (© 2024. The Author(s).)

Marine microbes are important in biogeochemical cycling, but the nature and magnitude of their contributions are influenced by their associated viruses. In the presence of a lytic virus, cells that have evolved resistance to infection have an obvious fitness advantage over relatives that remain susceptible. However, susceptible cells remain extant in the wild, implying that the evolution of a fitness advantage in one dimension (virus resistance) must be accompanied by a fitness cost in another dimension. Identifying costs of resistance is challenging because fitness is context-dependent. We examined the context dependence of fitness costs in isolates of the picophytoplankton genus Micromonas and their co-occurring dsDNA viruses using experimental evolution. After generating 88 resistant lineages from two ancestral Micromonas strains, each challenged with one of four distinct viral strains, we found resistance led to a 46% decrease in mean growth rate under high irradiance and a 19% decrease under low. After a year in culture, the experimentally selected lines remained resistant, but fitness costs had attenuated. Our results suggest that the cost of resistance in Micromonas is dependent on environmental conditions and the duration of population adaptation, illustrating the dynamic nature of fitness costs of viral resistance among marine protists., (© 2024 John Wiley & Sons Ltd.)

Over the past decade, the focus on omega (ω)-3 fatty acids from microalgae has intensified due to their diverse health benefits. Bioprocess optimization has notably increased ω-3 fatty acid yields, yet understanding of the genetic architecture and metabolic pathways of high-yielding strains remains limited. Leveraging genomics, transcriptomics, proteomics, and metabolomics tools can provide vital system-level insights into native ω-3 fatty acid-producing microalgae, further boosting production. In this review, we explore 'omics' studies uncovering alternative pathways for ω-3 fatty acid synthesis and genome-wide regulation in response to cultivation parameters. We also emphasize potential targets to fine-tune in order to enhance yield. Despite progress, an integrated omics platform is essential to overcome current bottlenecks in optimizing the process for ω-3 fatty acid production from microalgae, advancing this crucial field., Competing Interests: Declaration of interests The authors declare no competing interest., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Light management strategy is crucial for improving microalgal production in terms of higher biomass and economically valuable bioactive molecules. However, green light has received less attention in developing light managements for algae and higher plant due to its low absorption rate by chlorophyll. In this study, the effects of green light supplementation, in the combination with red and blue light were investigated in Haematococcus pluvialis. 10% and 20% of green light supplementations were applied in 3:2 ratios of red and blue LED light combinations as an expense of red-light. Growth rates, chlorophyll concentration, and dry weight were measured to assess the growth kinetics of H. pluvialis along with the relative transcript accumulations of four mRNAs: Rubisco, PTOX 2 , PsaB, and PsbS. Growth rates, chlorophyll concentrations and dry weight were found significantly higher in presence of 10% green light supplementation compared to red and blue light combinations. The relative transcript accumulations of Rubisco and PsbS genes showed significant upregulation at the end of the experiments (with the fold change of 42.91 ± 12.08 and 98.57 ± 27.38, respectively, relative to the beginning of the experiments) compared to combinations of red and blue light (fold change of 19.09 ± 3.0 and 47.77 ± 14.21, respectively, relative to beginning of the experiments). PsaB and PTOX 2 transcripts did not show significant accumulation differences between treatments. It seems that green light has a dose dependent additive effect on the growth rate of H. pluvialis. The upregulation of Rubisco and PsbS may indicate green light dependent carbon assimilation and light-harvesting response in H. pluvialis., (© 2024 American Institute of Chemical Engineers.)

Main Conclusion: Nitrogen stress altered important lipid parameters and related genes in Chlorella pyrenoidosa via ROS and Ca 2+ signaling. The mutual interference between ROS and Ca 2+ signaling was also uncovered. The changed mechanisms of lipid parameters (especially lipid classes and unsaturation of fatty acids) in microalgae are not completely well known under nitrogen stress. Therefore, Chlorella pyrenoidosa was exposed to 0, 0.5, 1 and 1.5 g L -1 NaNO 3 for 4 days. Then, the physiological and biochemical changes were measured. It was shown that the total lipid contents, neutral lipid ratios as well as their related genes (accD and DGAT) increased obviously while the polar lipid ratios, degrees of unsaturation as well as their related genes (PGP and desC) decreased significantly in nitrogen stress groups. The obvious correlations supported that gene expressions should be the necessary pathways to regulate the lipid changes in C. pyrenoidosa under nitrogen stress. The changes in ROS and Ca 2+ signaling as well as their significant correlations with corresponding genes and lipid parameters were analyzed. The results suggested that ROS and Ca 2+ may regulate these gene expressions and lipid changes in C. pyrenoidosa under nitrogen stress conditions. This was verified by the subordinate tests with an ROS inhibitor and calcium reagents. It also uncovered the clues of mutual interference between ROS and Ca 2+ signaling. To summarize, this study revealed the signaling pathways of important lipid changes in microalgae under N stress., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Antibiotic residues in mariculture wastewater seriously affect the aquatic environment. Antibiotic Resistance Genes (ARGs) produced under antibiotic stress flow through the environment and eventually enter the human body, seriously affecting human health. Microalgal-bacterial symbiotic system (MBSS) can remove antibiotics from mariculture and reduce the flow of ARGs into the environment. This review encapsulates the present scenario of mariculture wastewater, the removal mechanism of MBSS for antibiotics, and the biomolecular information under metagenomic assay. When confronted with antibiotics, there was a notable augmentation in the extracellular polymeric substances (EPS) content within MBSS, along with a concurrent elevation in the proportion of protein (PN) constituents within the EPS, which limits the entry of antibiotics into the cellular interior. Quorum sensing stimulates the microorganisms to produce biological responses (DNA synthesis - for adhesion) through signaling. Oxidative stress promotes gene expression (coupling, conjugation) to enhance horizontal gene transfer (HGT) in MBSS. The microbial community under metagenomic detection is dominated by aerobic bacteria in the bacterial-microalgal system. Compared to aerobic bacteria, anaerobic bacteria had the significant advantage of decreasing the distribution of ARGs. Overall, MBSS exhibits remarkable efficacy in mitigating the challenges posed by antibiotics and resistant genes from mariculture wastewater., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Nannochloropsis gaditana , a microalga known for its photosynthetic efficiency, serves as a cell factory, producing valuable biomolecules such as proteins, lipids, and pigments. These components make it an ideal candidate for biofuel production and pharmaceutical applications. In this study, we genetically engineered N. gaditana to overexpress the enzyme fructose-1,6-bisphosphatase (cyFBPase) using the Hsp promoter, aiming to enhance sugar metabolism and biomass accumulation. The modified algal strain, termed NgFBP, exhibited a 1.34-fold increase in cyFBPase activity under photoautotrophic conditions. This modification led to a doubling of biomass production and an increase in eicosapentaenoic acid (EPA) content in fatty acids to 20.78-23.08%. Additionally, the genetic alteration activated the pathways related to glycine, protoporphyrin, thioglucosides, pantothenic acid, CoA, and glycerophospholipids. This shift in carbon allocation towards chloroplast development significantly enhanced photosynthesis and growth. The outcomes of this study not only improve our understanding of photosynthesis and carbon allocation in N. gaditana but also suggest new biotechnological methods to optimize biomass yield and compound production in microalgae.

Overloading of nutrients such as nitrogen causes eutrophication of freshwater bodies. The spread of antibiotic resistance genes (ARGs) poses a threat to ecosystems. However, studies on the enrichment and spread of ARGs from increased nitrogen loading in algal-bacterial symbiotic systems are limited. In this study, the transfer of extracellular kanamycin resistance (KR) genes from large (RP4) small (pEASY-T1) plasmids into the intracellular and extracellular DNA (iDNA, eDNA) of the inter-algal environment of Chlorella pyrenoidosa was investigated, along with the community structure of free-living (FL) and particle-attached (PA) bacteria under different nitrogen source concentrations (0-2.5 g/L KNO 3 ). The results showed that KR gene abundance in the eDNA adsorbed on solid particles (D-eDNA) increased initially and then decreased with increasing nitrogen concentration, while the opposite was true for the rest of the free eDNA (E-eDNA). Medium nitrogen concentrations promoted the transfer of extracellular KR genes into the iDNA attached to algal microorganisms (A-iDNA), eDNA attached to algae (B-eDNA), and the iDNA of free microorganisms (C-iDNA); high nitrogen contributed to the transfer of KR genes into C-iDNA. The highest percentage of KR genes was found in B-eDNA with RP4 plasmid treatment (66.2%) and in C-iDNA with pEASY-T1 plasmid treatment (86.88%). In addition, dissolved oxygen (DO) significantly affected the bacterial PA and FL community compositions. Nephelometric turbidity units (NTU) reflected the abundance of ARGs in algae. Proteobacteria, Cyanobacteria, Bacteroidota, and Actinobacteriota were the main potential hosts of ARGs. These findings provide new insights into the distribution and dispersal of ARGs in the phytoplankton inter-algal environment., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The development of wastewater treatment processes capable of reducing and fixing carbon is currently a hot topic in the wastewater treatment field. Microalgae possess a natural carbon-fixing advantage, and microalgae that can symbiotically coexist with indigenous bacteria in actual wastewater attract more significant attention. Ultraviolet (UV) mutagenesis and dissolved organic carbon (DOC) acclimation were applied to strengthen the carbon-fixing performance of microalgae in this study. The mechanisms associated with microalgal water purification ability, gene regulation at the molecular level and photosynthetic potential under different trophic modes resulting from carbon fixation and transformation were disclosed. The superior performance of Chlorella sp. MHQ2 was eventually screened out among a large number of mutants generated from 3 wild-type Chlorella strains. Results indicated that the dry cell weight of the optimal species Chlorella sp. HQ mutant MHQ2 was 1.91 times that of the wild strain in the pure algal system, more carbon from municipal wastewater (MW) were transferred to the microalgae and re-entered into the biological cycle through resource utilization. In addition, COD, NH 3 -N and TP removal efficiencies of MW by Chlorella sp. MHQ2 were found to increase to 95.8% (1.1-times), 96.4% (1.4-times), and 92.9% (1.2-times), respectively, under the extra DOC supply and the assistance of indigenous bacteria in the MW. In the transcriptome analysis of the logarithmic phase, the glycolytic pathway was inhibited, and the pentose phosphate pathway was mainly carried out for microalgal life activities, further promoting efficient energy utilization. Upon analysis of carbon capture capacity and photosynthetic potential in trophic mode, the addition of NaHCO 3 increased the photosynthetic rate of Chlorella sp. MHQ2 in mixotrophy whereas it was attenuated in autotrophy. This study could provide a new perspective for the study of resource utilization and microalgae carbon- fixing mechanisms in the actual wastewater treatment process., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The pregnane X receptor (PXR) is a nuclear hormone receptor that plays a pivotal role in regulating gene expression in response to various ligands, particularly xenobiotics. In this context, the aim of this study was to shed light on the ligand affinity and functions of four NR1J1 paralogs identified in the marine mussel Mytilus galloprovincialis , employing a dual-luciferase reporter assay. To achieve this, the activation patterns of these paralogs in response to various toxins, including freshwater cyanotoxins (Anatoxin-a, Cylindrospermopsin, and Microcystin-LR, -RR, and -YR) and marine algal toxins (Nodularin, Saxitoxin, and Tetrodotoxin), alongside natural compounds (Saint John's Wort, Ursolic Acid, and 8-Methoxypsoralene) and microalgal extracts ( Tetraselmis , Isochrysis, LEGE 95046, and LEGE 91351 extracts), were studied. The investigation revealed nuanced differences in paralog response patterns, highlighting the remarkable sensitivity of MgaNR1J1γ and MgaNR1J1δ paralogs to several toxins. In conclusion, this study sheds light on the intricate mechanisms of xenobiotic metabolism and detoxification, particularly focusing on the role of marine mussel NR1J1 in responding to a diverse array of compounds. Furthermore, comparative analysis with human PXR revealed potential species-specific adaptations in detoxification mechanisms, suggesting evolutionary implications. These findings deepen our understanding of PXR-mediated metabolism mechanisms, offering insights into environmental monitoring and evolutionary biology research.

Carbon capture and biochemical storage are some of the primary drivers of photosynthetic yield and productivity. To elucidate the mechanisms governing carbon allocation, we designed a photosynthetic light response test system for genetic and metabolic carbon assimilation tracking, using microalgae as simplified plant models. The systems biology mapping of high light-responsive photophysiology and carbon utilization dynamics between two variants of the same Picochlorum celeri species, TG1 and TG2 elucidated metabolic bottlenecks and transport rates of intermediates using instationary 13 C-fluxomics. Simultaneous global gene expression dynamics showed 73% of the annotated genes responding within one hour, elucidating a singular, diel-responsive transcription factor, closely related to the CCA1/LHY clock genes in plants, with significantly altered expression in TG2. Transgenic P. celeri TG1 cells expressing the TG2 CCA1/LHY gene, showed 15% increase in growth rates and 25% increase in storage carbohydrate content, supporting a coordinating regulatory function for a single transcription factor., (© 2024. The Author(s).)

Microalgae are regarded as sustainable and potent chassis for biotechnology. Their capacity for efficient photosynthesis fuels dynamic growth independent from organic carbon sources and converts atmospheric CO 2 directly into various valuable hydrocarbon-based metabolites. However, approaches to gene expression and metabolic regulation have been inferior to those in more established heterotrophs (e.g., prokaryotes or yeast) since the genetic tools and insights in expression regulation have been distinctly less advanced. In recent years, however, these tools and their efficiency have dramatically improved. Various examples have demonstrated new trends in microalgal biotechnology and the potential of microalgae for the transition towards a sustainable bioeconomy., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)