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• Microalgae accumulate high levels of oil under stress, but the underlying biosynthetic pathways are not fully understood. We sought to identify key regulators of lipid metabolism under stress conditions. • We found that the Chlamydomonas reinhardtii gene encoding the MYB-type transcription factor MYB1 is highly induced under stress conditions. Two myb1 mutants accumulated less total fatty acids and storage lipids than their parental strain upon nitrogen (N) depletion. • Transcriptome analysis revealed that genes involved in lipid metabolism are highly enriched in the wild-type but not in the myb1-1 mutant after 4 h of N depletion. Among these genes were several involved in the transport of fatty acids from the chloroplast to the endoplasmic reticulum (ER): acyl-ACP thioesterase (FAT1), Fatty Acid EXporters (FAX1, FAX2), and long-chain acyl-CoA synthetase1 (LACS1). Furthermore, overexpression of FAT1 in the chloroplast increased lipid production. These results suggest that, upon N depletion, MYB1 promotes lipid accumulation by facilitating fatty acid transport from the chloroplast to the ER. • This study identifies MYB1 as an important positive regulator of lipid accumulation in C. reinhardtii upon N depletion, adding another player to the established regulators of this process, including NITROGEN RESPONSE REGULATOR 1 (NRR1) and TRIACYLGLYCEROL ACCUMULATION REGULATOR 1 (TAR1).

Sub-Saharan Africa is facing food security challenges due, in part, to decades of soil nitrogen (N) depletion. Applying N fertilizer could increase crop yields and replenish soil N pools. From 2010 to 2015, field experiments conducted in Embu and Kiboko, Kenya and Harare, Zimbabwe investigated yield and N uptake response of six maize ( Zea mays L.) hybrids to four N fertilizer rates (0 to 160 kg N ha −1 ) in continuous maize production systems. The N recovery efficiency (NRE), cumulative N balance, and soil N content in the upper 0.9 m of soil following the final harvest were determined at each N rate. Plant and soil responses to N fertilizer applications did not differ amongst hybrids. Across locations and N rates, NRE ranged from 0.4 to 1.8 kg kg −1 . Higher NRE values in Kiboko and Harare occurred at lower post-harvest soil inorganic N levels. The excessively high NRE value of 1.8 kg kg −1 at 40 kg N ha −1 in Harare suggested that maize hybrids deplete soil inorganic N most at low N rates. Still, negative cumulative N balances indicated that inorganic soil N depletion occurred at all N rates in Embu and Harare (up to − 193 and − 167 kg N ha −1 , respectively) and at the 40 kg N ha −1 rate in Kiboko (− 72 kg N ha −1 ). Overall, maize N uptake exceeded fertilizer N applied and so, while yields increased, soil N pools were not replenished, especially at low total soil N levels (< 10,000 kg N ha −1 in top 0.9 m).

The great challenge of reducing soil nutrient depletion and assuring agricultural system productivity in low-income countries caused by limited synthetic fertilizer use necessitates local and cost-effective nutrient sources. We estimated the changes of the nitrogen budget of agricultural systems in the East African Community from 1961 to 2018 to address the challenges of insufficient nitrogen inputs and serious soil nitrogen depletion in agricultural systems of the East African Community region. Results showed that total nitrogen input increased from 12.5 kg N ha -1 yr -1 in the 1960s to 21.8 kg N ha -1 yr -1 in the 2000s and 27 kg N ha -1 yr -1 in the 2010s. Total nitrogen crop uptake increased from 12.8 kg N ha -1 yr -1 in the 1960s to 18.2 kg N ha -1 yr -1 in the 2000s and 21.8 kg N ha -1 yr -1 in the 2010s. Soil nitrogen stock increased from -2.0 kg N ha -1 yr -1 in the 1960s to -0.5 kg N ha -1 yr -1 in the 2000s and 0.3 kg N ha -1 yr -1 in the 2010s. Our results allow us to substantiate for the first time that soil nitrogen depletion decreases with increasing input of nitrogen in agricultural systems of the East African Community region. This suggests that increases in nitrogen inputs through biological nitrogen fixation and animal manure are the critical nitrogen management practices to curb soil nitrogen depletion and sustain agricultural production systems in the East African Community region in order to meet food demand for a growing population.

Docosahexaenoic acid (DHA) is a high-value omega-3 polyunsaturated fatty acid (PUFA) with critical roles in human health and nutrition, and Aurantiochytrium species have emerged as promising microbial cell factories for sustainable and scalable DHA biosynthesis, achieving yields of up to 43% of total fatty acids. To elucidate the regulatory mechanisms underlying lipid overproduction in this organism, we performed an integrated proteomic and transcriptomic analysis of the high-yield mutant Aurantiochytrium sp. YHPM1 under a two-stage nitrogen depletion strategy, at 48 and 108 h, the strain exhibited a biphasic metabolic response. Initially, β-oxidation and amino-acid catabolism were suppressed. This was followed by enhanced fatty-acid (FA) and triacylglycerol (TAG) biosynthesis. Moreover, key lipid biosynthetic enzymes, including DGAT, DLAT, and GPAT were markedly upregulated at both transcriptomic and proteomic levels, while β-oxidation enzymes, such as ACADM and FOX2 were suppressed. Notably, 8% of transcripts exhibited discordant transcript–protein expression, most prominently at 48 h, suggesting extensive post-transcriptional regulation. Overall, these findings demonstrate a phased metabolic reprogramming strategy in YHPM1 and advance our understanding of post-transcriptional mechanisms governing DHA accumulation, thereby facilitating the industrial-scale production of DHA using Aurantiochytrium.

Background This batch study deals with the relation between lipid as well as triglyceride contents in Nannochloropsis salina and variation in culture conditions such as nitrogen concentration and temperature. Results The tested parameters caused reduction in growth expressed as cell count, optical density and dry weight, as well strongly involved in lipids and triglycerides accumulation and significantly affected the lipid productivity. At the beginning of the work, the concentration of nitrogen in the medium was reduced to three quarter, half and quarter of the original f2 medium while the temperature kept constant. After that, the optimal nitrogen concentration (quarter of the original media) giving high lipid yield was tested with different temperature degrees from 15 to 35°C with five degree intervals. Although the growth was insignificantly influenced, a considerable increase in lipid and triglyceride (56.1 and 15.1% of dry weight respectively) was observed when the concentration of nitrogen in the medium was reduced to the quarter. Moreover, 59.3% lipid and 17.1% triglyceride on the basis of dry weight were obtained by the combination of 25% nitrogen concentration and 30°C. Simple regressions recommended that the interaction effect of nitrogen limitation and temperature on lipid and triglyceride accumulation was not as fundamental as for nitrogen limitation stress. Conclusion The degree of nitrogen availability in the combination of temperature effect has been identified as the critical determinant for the maximal production of lipid in N. salina . Nevertheless, major advances in this field can be considered by studying more stresses techniques and genetic strategies.

Beta-glucans are polysaccharides that can be used for different applications, for example as an immunomodulator in food or feed or for managing high cholesterol levels. Certain microalgae species use beta-glucans as energy storage, accumulating them during nutrient depletion. In this study, we examined and compared beta-glucan production during nitrogen depletion in three different algae species, Phaeodactylum tricornutum, Monodopsis subterranea and Cylindrotheca fusiformis, grown in artificially illuminated flat panel airlift reactors, in order to determine the most promising microalgae species for beta-glucan production. Co-products such as fatty acids (especially eicosapentaenoic acid) and the carotenoid fucoxanthin (not produced by M. subterranea) were also considered. Biomass analysis showed that P. tricornutum cultures reached a maximal beta-glucan content of 317 ± 9 mg gDW−1, M. subterranea cultures reached 188 ± 6 mg gDW−1 and C. fusiformis cultures reached 129 ± 13 mg gDW−1. Furthermore, beta-glucan production was faster in P. tricornutum cultures. However, the maximum volumetric beta-glucan concentration reached was higher in M. subterranea cultures compared to P. tricornutum cultures as M. subterranea cultures produced more biomass during nitrogen depletion. In terms of possible co-products, P. tricornutum produced fucoxanthin and EPA, whereas M. subterranea did not produce fucoxanthin. However, M. subterranea exhibited a higher EPA content, which remained above 45 mg g−1 even after several days of nitrogen depletion. Overall, our results suggest that P. tricornutum and M. subterranea are both suitable species for beta-glucan production in flat panel airlift reactors.

Microalgae have gained attention due to their higher reproduction rate and lipid productivity. In particular, various stress conditions lead to an overproduction of lipids in microalgae cells. The study investigated the influence of additional CO2 introduced with air into the reactor during biomass growth of Chlorella sp. Additionally, increased phosphorus concentration in the medium under stress cultivation (low nitrogen concentration) was examined. The partial pressure of CO2 and its increased availability to Chlorella sp. in the cultivation medium increased biomass growth (1.4 times) and chlorophyll content (2.5 times) in microalgae cells. A high phosphorus fertilizer significantly increased lipid production under stress conditions with CO2 supply to 85.2 mg/g (2.6 times) and without CO2 to 73.8 mg/g (2.2 times). A high concentration of phosphorus in the culture medium stimulated the synthesis of C16:0 (about 38–45%) and C18:1 CIS9 (about 24–30%). The results confirm that the fertilizers can be used as a culture medium to induce stress and stimulate lipid production. Adjusting the composition of the fertilizers and controlling the additional CO2 supply could prove beneficial to increase the content of the desired fatty acids.

Cell walls of microalgae consist of a polysaccharide and glycoprotein matrix providing the cells with a formidable defense against its environment. We characterized enzymes that can digest the cell wall and weaken this defense for the purpose of protoplasting or lipid extraction. A growth inhibition screen demonstrated that chitinase, lysozyme, pectinase, sulfatase, β-glucuronidase, and laminarinase had the broadest effect across the various Chlorella strains tested and also inhibited Nannochloropsis and Nannochloris strains. Chlorella is typically most sensitive to chitinases and lysozymes, both enzymes that degrade polymers containing N-acetylglucosamine. Using a fluorescent DNA stain, we developed rapid methodology to quantify changes in permeability in response to enzyme digestion and found that treatment with lysozyme in conjunction with other enzymes has a drastic effect on cell permeability. Transmission electron microscopy of enzymatically treated Chlorella vulgaris indicates that lysozyme degrades the outer surface of the cell wall and removes hair-like fibers protruding from the surface, which differs from the activity of chitinase. This action on the outer surface of the cell causes visible protuberances on the cell surface and presumably leads to the increased settling rate when cells are treated with lysozyme. We demonstrate radical ultrastructural changes to the cell wall in response to treatment with various enzyme combinations which, in some cases, causes a greater than twofold increase in the thickness of the cell wall. The enzymes characterized in this study should prove useful in the engineering and extraction of oils from microalgae.

Rising oil prices and concerns over climate change have resulted in more emphasis on research into renewable biofuels from microalgae. Unlike plants, green microalgae have higher biomass productivity, will not compete with food and agriculture, and do not require fertile land for cultivation. However, microalgae biofuels currently suffer from high capital and operating costs due to low yields and costly extraction methods. Microalgae grown under optimal conditions produce large amounts of biomass but with low neutral lipid content, while microalgae grown in nutrient starvation accumulate high levels of neutral lipids but are slow growing. Producing lipids while maintaining high growth rates is vital for biofuel production because high biomass productivity increases yield per harvest volume while high lipid content decreases the cost of extraction per unit product. Therefore, there is a need for metabolic engineering of microalgae to constitutively produce high amounts of lipids without sacrificing growth. Substrate availability is a rate-limiting step in balancing growth and fatty acid (FA) production because both biomass and FA synthesis pathways compete for the same substrates, namely acetyl-CoA and NADPH. In this review, we discuss the efforts made for improving biofuel production in plants and microorganisms, the challenges faced in achieving lipid productivity, and the important role of precursor supply for FA synthesis. The main focus is placed on the enzymes which catalyzed the reactions supplying acetyl-CoA and NADPH.

BackgroundMicroalgal starch is regarded as a promising alternative to crop-based starch for biorefinery such as the production of biofuels and bio-based chemicals. The single or separate use of inorganic carbon source, e.g., CO2 and NaHCO3, caused aberrant pH, which restricts the biomass and starch production. The present study applied an in situ CO2–NaHCO3 system to regulate photosynthetic biomass and starch production along with starch quality in a marine green microalga Tetraselmis subcordiformis under nitrogen-depletion (−N) and nitrogen-limitation (±N) conditions.ResultsThe CO2 (2%)–NaHCO3 (1 g L−1) system stabilized the pH at 7.7 in the −N cultivation, under which the optimal biomass and starch accumulation were achieved. The biomass and starch productivity under −N were improved by 2.1-fold and 1.7-fold, respectively, with 1 g L−1 NaHCO3 addition compared with the one without NaHCO3 addition. NaHCO3 addition alleviated the high-dCO2 inhibition caused by the single CO2 aeration, and provided sufficient effective carbon source HCO3− for the maintenance of adequate photosynthetic efficiency and increase in photoprotection to facilitate the biomass and starch production. The amylose content was also increased by 44% under this CO2–bicarbonate system compared to the single use of CO2. The highest starch productivity of 0.73 g L−1 day−1 under −N cultivation and highest starch concentration of 4.14 g L−1 under ±N cultivation were both achieved with the addition of 1 g L−1 NaHCO3. These levels were comparable to or exceeded the current achievements reported in studies. The addition of 5 g L−1 NaHCO3 under ±N cultivation led to a production of high-amylose starch (59.3% of total starch), which could be used as a source of functional food.ConclusionsThe in situ CO2–NaHCO3 system significantly improved the biomass and starch production in T. subcordiformis. It could also regulate the starch quality with varied relative amylose content under different cultivation modes for diverse downstream applications that could promote the economic feasibility of microalgal starch-based biofuel production. Adoption of this system in T. subcordiformis would facilitate the CO2 mitigation couple with its starch-based biorefinery.