Isochrysis Zhangjiangensis Research Articles

Effect of heavy metals in aquaculture water on the growth of microalgae and their migration mechanism in algae-shellfish system

This research the bait microalgae Chlorella sp. and Isochrysis zhangjiangensis were fed to Ruditapes philippinarum (clams) using two types of cultures, monoculture and co-culture, simulating the conditions, and presence, of common heavy metals Cu2+, Pb2+ and Cd2+ in cultured waters. The effects of microalgae growth, antioxidant activity, and changes in nutrient TN and TP were investigated. The metal contents of the culture solution, algal cells, and four tissues of Ruditapes philippinarum were analyzed. The results showed that the highest nutrient removal efficiency was achieved under the co-culture of three heavy metals, with the removal rates of TN and TP ranging from 47.26% to 51.36% and 78.28% to 87.72%, respectively. The higher tolerance of Chlorella sp. to heavy metals compared to Isochrysis zhangjiangensis was probably due to the higher glutathione and super oxide dimutase activities exhibited by Chlorella sp. cells, which increased by 28% and 40.64%, respectively. Isochrysis zhanjiangensis, Chlorella sp. and their co-culture group had the highest removal rate of Pb2+, which were 20.15%, 26.56% and 29.19%, respectively, followed by Cd2+ and Cu2+. The enrichment of the four tissues from the Ruditapes philippinarum organism for heavy metals, was in the order of visceral mass > gill > coat membrane > muscle. The biotransfer coefficients of the three metals in the microalgae-Ruditapes philippinarum food chain biomagnification ranged from 0.24 to 0.79, which indicated that there was no significant biomagnification of the metals in the transfer process of the food chain. The results of this study reveal the mechanism of heavy metal transport in the algae-shellfish system, which would provide a theoretical basis for improving the ecological environment, and can improve the quality of cultured shellfish products.

Enhancing the Thermotolerance of Isochrysis zhangjiangensis Through Co-culturing With Algoriphagusmarincola.

Isochrysis zhangjiangensis is an important microalgal species used as bait in aquaculture. However, its optimal cultivation temperature is around 25°C, limiting its use in summer when temperature is higher. To overcome this limitation, we aimed to develop a consortia of I. zhangjiangensis and bacteria that are more resistant to heat stress. Here, six thermotolerance-promoting bacterial strains were isolated from the culture of a heat-tolerant mutant strain of I. zhangjiangensis (IM), and identified as Algoriphagus marincola, Nocardioides sp., Pseudidiomarina sp., Labrenzia alba, Nitratireductor sp., and Staphylococcus haemolyticus. Further, co-culturing I. zhangjiangensis with A. marincola under high temperature conditions increased cell density, chlorophyll a, PSII maximum photochemical efficiency (Fv/Fm), and soluble protein content of microalgae. The presence of A. marincola positively influenced the activities of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and total antioxidant capacity (T-AOC) in I. zhangjiangensis cells, while concurrently reducing the levels of reactive oxygen species (ROS). Additionally, gene expression studies confirmed that co-culturing with A. marincola upregulated the expression of antioxidant-related genes (sod and pod) and stress tolerance genes (heat shock protein genes). Our findings indicate that A. marincola effectively helps I. zhangjiangensis withstand high temperature stress, leading to improved yield of microalgae during high temperature conditions. The thermotolerance-promoting bacteria can be exploited as potential inoculants for enhancing the productivity and sustainability of bait microalgae in aquaculture.

High-Purity Fucoxanthin Can Be Efficiently Prepared from Isochrysis zhangjiangensis by Ethanol-Based Green Method Coupled with Octadecylsilyl (ODS) Column Chromatography.

The exploitation of new economically valuable microalgae as a sustainable source of minor high-value products can effectively promote the full utilization of microalgae. The efficient preparation of minor products from microalgae remains the challenge, owing to the coexistence of various components with a similar polarity in the microalgae biomass. In this study, a novel approach based on the sustainable-oriented strategy for fucoxanthin (FX) production was proposed, which consisted of four steps, including the culture of microalga, ethanol extraction, ODS column chromatography, and ethanol precipitation. The high-purity FX (around 95%) was efficiently obtained in a total recovery efficiency of 84.28 ± 2.56%. This study reveals that I. zhangjiangensis is a potentially promising feedstock for FX production and firstly provides a potentially eco-friendly method for the scale-up preparation of FX from the microalga I. zhangjiangensis.

Nutrient Deprivation Coupled with High Light Exposure for Bioactive Chrysolaminarin Production in the Marine Microalga Isochrysis zhangjiangensis.

Chrysolaminarin, a kind of water-soluble bioactive β-glucan produced by certain microalgae, is a potential candidate for food/pharmaceutical applications. This study identified a marine microalga Isochrysis zhangjiangensis, in which chrysolaminarin production was investigated via nutrient (nitrogen, phosphorus, or sulfur) deprivations (-N, -P, or -S conditions) along with an increase in light intensity. A characterization of the antioxidant activities of the chrysolaminarin produced under each condition was also conducted. The results showed that nutrient deprivation caused a significant increase in chrysolaminarin accumulation, though this was accompanied by diminished biomass production and photosynthetic activity. -S was the best strategy to induce chrysolaminarin accumulation. An increase in light intensity from 80 (LL) to 150 (HL) µE·m−2·s−1 further enhanced chrysolaminarin production. Compared with -N, -S caused more suitable stress and reduced carbon allocation toward neutral lipid production, which enabled a higher chrysolaminarin accumulation capacity. The highest chrysolaminarin content and concentration reached 41.7% of dry weight (%DW) and 632.2 mg/L, respectively, under HL-S, with a corresponding productivity of 155.1 mg/L/day achieved, which exceeds most of the photoautotrophic microalgae previously reported. The chrysolaminarin produced under HL-N (Iz-N) had a relatively competitive hydroxyl radical scavenging activity at low concentrations, while the chrysolaminarin produced under HL-S (Iz-S) exhibited an overall better activity, comparable to the commercial yeast β-glucan, demonstrating I. zhangjiangensis as a promising bioactive chrysolaminarin producer from CO2.

Integrated metabolic tools reveal carbon alternative in Isochrysis zhangjiangensis for fucoxanthin improvement

This study explored the regulation of photosystem and central carbon metabolism in cell growth and fucoxanthin accumulation of Isochrysis zhangjiangensis via transcriptome analysis, targeted metabolite measurements, and flux balance analysis. High light promoted biomass accumulation but dramatically decreased fucoxanthin productivity. It suppressed the active photosystem and reduced chlorophyll content, but improved metabolic flux of Calvin–Benson–Bassham and tricarboxylic acid cycle for massive biomass accumulation. The CO2 fixation was largely dependent on mitochondrial energy illustrated by the integrated metabolic tools. At a molecular level, glyceraldehyde–3–phosphate, acetyl–CoA, and pyruvate contents increased at exponential phase under high light, which tended to participate into fatty acid biosynthesis by the up-regulated ACCase. However, high light inhibited most genes involved in fucoxanthin biosynthesis and induced diadinoxanthin cycle to diatoxanthin form. Therefore, constant light at 100 μmol m−2 s−1 balancing biomass concentration and fucoxanthin content provided the highest fucoxanthin productivity at 3.06 mg L-1 d-1.

Analysis of Relationship Between Bacterial Community and High Temperature Tolerance of Isochrysis zhangjiangensis

Analysis of Relationship Between Bacterial Community and High Temperature Tolerance of Isochrysis zhangjiangensis

Cultivation of aquaculture feed Isochrysis zhangjiangensis in low-cost wave driven floating photobioreactor without aeration device

This study aimed to use floating photobioreactor (PBR) to produce microalgae biomass for aquaculture applications, and this was tested with cultivation of Isochrysis zhangjiangensis. The highest cell density of 16.1 ± 0.61 × 106 cell L–1 was obtained in an outdoor culture with a depth of 5.0 cm in 1.0 m2 floating PBR, but deeper culture resulted in higher biomass productivity. Large-scale cultivation at size of 10 m2 (1000 L) produced the highest cell density of 17.8 × 106 cell L–1 and highest biomass productivity of 0.115 g L–1 d–1, which was at the same level as that for flat-panel PBR (100 L). This developed technique provides an innovative approach to produce microalgae on site for use as fresh aquaculture feed, as well as fresh cells for use as seed inoculums for large-area aquaculture water bodies. This approach provides not only a low-cost microalgae production system but also better integration between microalgae production and aquaculture.

The heat-tolerance evaluation of an Isochrysis zhangjiangensis mutant generated by atmospheric and room temperature plasmas

Isochrysis zhangjiangensis is widely used in the marine aquaculture as larval feed, especially for filter feeding cultures, as well as a good candidate for biofuels. However, the optimal cultivation temperature for I. zhangjiangensis is below 30 °C and this stain is seriously affected by high temperature, which causes the limited application during the summer. I. zhangjiangensis IM130005 is a strain generated by atmospheric and room temperature plasmas with relative higher growth rate and lipid production than the wide strain (WT), with the ability to tolerate several hours’ high temperature during the outdoor cultivation. Here, a detailed comparison was performed by continuous monitoring growth, chlorophyll fluorescence and fatty acid profile between IM13005 and WT under a mimic temperature shock to the summer outdoor cultivation. Based on a nearly 20% increase of total fatty acid in IM13005, which was majorly contributed by saturated or monounsaturated FAs in form of neutral lipids, within 5 h under the heat shock, the fatty acids and lipids synthesis variation were postulated as the physiological reason for the high temperature tolerance.

Effect of different phosphorus concentrations on biodiesel production from Isochrysis zhangjiangensis under nitrogen sufficiency or deprivation condition.

The effects of two nitrogen (N) concentrations combining with three phosphorus (P) concentrations on Isochrysis zhangjiangensis growth and formation of fatty acid (FA) were investigated in this study. Biomass concentration, mass fraction, and productivity of FA in I. zhangjiangensis were low in N-deprived media. Under both N and P sufficiency conditions, the intake of P and N was 40 times and 4.7 times of the normal algal growth condition, respectively, indicating I. zhangjiangensis had the potential for removing P and N from high concentrated N and P salinity wastewater. This study also showed that P deficiency in N sufficient medium increased the FA content, however, the difference between P limitation and P deprivation was not significant (P > 0.05). In N sufficient and P limitation medium, FA productivity was the highest, with a composition suitable for biofuel, so, this condition was the optimal condition for biodiesel production from I. zhangjiangensis.

The Light Regime Effect on Triacylglycerol Accumulation of Isochrysis zhangjiangensis

Stress state of microalgal cells is caused under unfavorable conditions such as disordered light regime and depleted nitrogen. The stress state can impair photosynthetic efficiency, inhibit cell growth and result in the accumulation of triacylglycerol (TAG) from protective mechanisms. Continuous light or nitrogen starvation was applied on microalgae and performed effectively on inducing TAG production. To evaluate the light regime effect on inducing TAG production, the effect of different light regimes on nitrogen-starved Isochrysis zhangjiangensis was investigated in this work. The continuous light and nitrogen starvation elevated TAG content of biomass by 73% and 193%, respectively. Furthermore, the TAG accumulation of I. zhangjiangensis cell under nitrogen starvation decreased under aggravated stress from continuous illumination. Our results demonstrated that culturing the cells with 14L: 10D light regime under nitrogen starvation is the optimal mode to achieve maximal accumulation of TAG. A recovery in light regime was necessary for I. zhangjiangensis cultivation.

Storage carbon metabolism of Isochrysis zhangjiangensis under different light intensities and its application for co-production of fucoxanthin and stearidonic acid

This study explored the co-production of fucoxanthin and stearidonic acid from Isochrysis zhangjiangensis by investigating its carbon metabolism under different light intensities. Results showed high light inhibited the synthesis of fucoxanthin and stearidonic acid, while promoted cell growth and enhanced cellular lipid content compared with low light, achieving 2.4 g/L and 28.55%, respectively. Low light accelerated the accumulation of fucoxanthin and stearidonic acid, which obtained 23.29 mg/g and 17.16% (of total fatty acid). In combination with the molecular analysis, low light redirected carbon skeletons into glyceraldehyde-3-phosphate and diverted into carotenoid especially fucoxanthin. While, high light redistributed the skeletons to Malonyl CoA, citrate and α-Ketoglutarate and then oriented into lipid metabolism. The highest fucoxanthin and stearidonic acid productivity was 2.94 mg L−1 d−1 and 4.33 mg L−1 d−1, respectively, which revealed I. zhanjiangensis is a potential strain for the co-production of fucoxanthin and stearidonic acid.

New insights into the CO2-steady and pH-steady cultivations of two microalgae based on continuous online parameter monitoring

CO2-steady and pH-steady cultivations are two common strategies used in the cultivation of microalgae for the control of carbon supply. However, the differences between the two strategies have seldom been discussed due to the lack of an appropriate evaluation platform. Here, a precise comparison of CO2-steady and pH-steady modes was carried out using Nannochloropsis oceanica IMTE1 and Isochrysis zhangjiangensis on the Algal Station system, a comprehensive process-monitoring platform specifically designed for microalgae R&D. The physicochemical variables, optical density, dry weight and chlorophyll fluorescence parameters, as well as the environmental parameters, pH, temperature, light intensity and CO2 concentration, were detected online at 20-minute intervals. The I. zhangjiangensis biomass concentration increased by 17% under the pH-steady mode compared to the CO2-steady mode, whereas there was no difference in the N. oceanica biomass concentration. By using Algal Station, a notable difference was also found in the periodical change of Fv/Fm between these two strains. This is the first systematic study of Fv/Fm variation during light and dark cycles in microalgae. The results obtained by Algal Station are important for basic physiological studies and dynamic process optimizations for more efficient microalgae cultivation.

Enhancement of the Production of Bioactive Microalgal Metabolites by Ultraviolet Radiation (UVA 365 nm).

Two marine microalgae, Nitzschia closterium and Isochrysis zhangjiangensis, treated for 9 days with photosynthetically active radiation (PAR), were subjected to 3 days of ultraviolet light band A at 365 nm (UVA365) followed by a 3 day post-UVA PAR treatment. Compared with the control that only had PAR treatment, UVA365 treatment significantly ( p < 0.05) increased the amounts of total xanthophylls in N. closterium and I. zhangjiangensis by up to 3.53- and 1.23-fold, respectively, after 3 days of UVA365 irradiation. In particular, UVA365 treatment markedly ( p < 0.05) increased two major bioactive xanthophyll pigments, fucoxanthin in N. closterium and diadinoxanthin in I. zhangjiangensis, by up to 1.97- and 1.52-fold, respectively. UVA365 treatment significantly ( p < 0.05) increased the accumulation of another bioactive microalgal metabolite, total mycosporine-like amino acids, in N. closterium and I. zhangjiangensis by up to 1.40- and 1.30-fold, respectively. UVA365 irradiation has potential as an efficient approach for enhancing the production of valuable microalgal metabolites.

Regulatory mechanisms of oxidative species and phytohormones in marine microalgae Isochrysis zhangjiangensis under nitrogen deficiency

Oxidative species and phytohormones are important factors in plants that can regulate stress responses. In this study, the content variations of four oxidative species, O2−, H2O2, ClO−, and NO, and the phytohormones, salicylic acid (SA), jasmonic acid (JA), and folic acid (FA), were investigated in the microalgae Isochrysis zhangjiangensis under nitrogen deficiency. The O2− and H2O2 contents were found to be positively correlated with the photosynthetic activity of the microalgae, while the opposite relationship was observed for NO and ClO−. In higher plants, there is a balance model for NO and ROS interactions, which revealed that the cooperation of NO and H2O2 regulates the pathogen-induced hypersensitive response-associated cell death. Because the NO and H2O2 levels in both nitrogen-replete and nitrogen-depleted I. zhangjiangensis were not simultaneously high, we proposed that as a unicellular lower plant, the microalgae I. zhangjiangensis follows this model and regulates the different types of intracellular oxidative species, especially when they were cultured from nitrogen sufficient condition to corresponding nitrogen deficient condition, to avoid cell death. Together with the corresponding changes in the SA, JA and FA contents in the nitrogen-stressed microalgae, we proposed that the balance model of oxidative species interactions, as well as the correlations among nitrogen assimilation, photosynthesis and the biosyntheses of SA, JA, and FA, may originally exist in algae, and plants have followed this model and maintained these correlations all along to protect themselves from stressful conditions during their evolutions from alga to higher plants.

Determination of internal controls for quantitative gene expression of Isochrysis zhangjiangensis at nitrogen stress condition

Isochrysis zhangjiangensis is a potential marine microalga for biodiesel production, which accumulates lipid under nitrogen limitation conditions, but the mechanism on molecular level is veiled. Quantitative real-time polymerase chain reaction (qPCR) provides the possibility to investigate the gene expression levels, and a valid reference for data normalization is an essential prerequisite for firing up the analysis. In this study, five housekeeping genes, actin (ACT), α-tubulin (TUA), s-tubulin (TUB), ubiquitin (UBI), 18S rRNA (18S) and one target gene, diacylglycerol acyltransferase (DGAT), were used for determining the reference. By analyzing the stabilities based on calculation of the stability index and on operating the two types of software, geNorm and bestkeeper, it showed that the reference genes widely used in higher plant and microalgae, such as UBI, TUA and 18S, were not the most stable ones in nitrogen-stressed I. zhangjiangensis, and thus are not suitable for exploring the mRNA expression levels under these experimental conditions. Our results show that ACT together with TUB is the most feasible internal control for investigating gene expression under nitrogen-stressed conditions. Our findings will contribute not only to future qPCR studies of I. zhangjiangensis, but also to verification of comparative transcriptomics studies of the microalgae under similar conditions.

Free amino acids and small molecular acids profiling of marine microalga Isochrysis zhangjiangensis under nitrogen deficiency

Isochrysis zhangjiangensis is a marine microalga with potential usage in biofuel production. The alga is known to exhibit enhanced lipid and polysaccharide storage under nitrogen deficiency conditions, though the regulatory mechanism has not been elucidated.In this study, to monitor the metabolic transition in response to nitrogen starvation, the intracellular metabolite fluctuation within 32h was profiled by gas chromatography–mass spectrometry and liquid chromatography–mass spectrometry scanned in selected ion monitoring mode for the first time. These techniques identified and quantified the levels of 14 SMAs, 2 carbohydrates involved in the TCA cycle and glycolysis, and 28 free amino acids (AAs). The pulsed increase of pyruvate, which is the precursor of acetyl-CoA and fatty acids (FAs), indicated a potential to produce more FAs. Surprisingly, although overall AAs showed a decreasing trend under the experimental conditions, Ala and Phe showed increased levels initially. It has been postulated that a carbon flux switch between lipids and polysaccharides exists in I. zhangjiangensis. To be feasible for biofuel production, enhancement of carbon flux to lipids is expected. Facilitating the intracellular lipid storage, together with nitrogen deficiency, a β-1,3-glucanase inhibitor (Micafungin) was introduced to block the formation of chrysolaminarin, the major polysaccharide in I. zhangjiangensis. The treatment resulted in 10% reduction of polysaccharides and 50% transient increase of total lipids. AAs and other metabolites involved in the TCA cycle and glycolysis, were also perturbed by the treatment with Micafungin, among which gamma-aminobutyric acid (GABA) was profoundly affected although the effects of Micafungin were only transient. This study indicates that β-1,3-glucanase could be a good candidate for further genetic manipulation and that the possibility of regulating GABA for high oil yield warrants detailed study.

Coordinated regulation of nitrogen supply mode and initial cell density for energy storage compounds production with economized nitrogen utilization in a marine microalga Isochrysis zhangjiangensis

Lipids and carbohydrates are main energy storage compounds (ESC) of microalgae under stressed conditions and they are potential feedstock for biofuel production. Yet, the sustainable and commercially successful production of ESC in microalgae needs to consider nitrogen utilization efficiency. Here the impact of different initial cell densities (ICDs) on ESC accumulation in Isochrysis zhangjiangensis under two nitrogen supply modes (an initially equal concentration of nitrogen per-cell in the medium (N1) and an equal total concentration of nitrogen in the culture system (N2)) were investigated. The results demonstrated that the highest ESC yield (1.36gL−1) at N1, which included a maximal nitrogen supply in the cultivation system, and the highest ESC content (66.5%) and ESC productivity per mass of nitrogen (3.28gg−1 (N) day−1) at N2, were all obtained under a high ICD of 8.0×106cellsmL−1. Therefore I. zhangjiangensis qualifies for ESC-enriched biomass production with economized nitrogen utilization.

Nutritional stress effects under different nitrogen sources on the genes in microalga Isochrysis zhangjiangensis and the assistance of Alteromonas macleodii in releasing the stress of amino acid deficiency

The expressions of nine nitrogen assimilation-associated genes, NRT2, NAR1, NIA2, NIR, GLN2, GLSF, GSN1, GDH, and AAT2, in the microalga Isochrysis zhangjiangensis were investigated to unveil the effects of limitations of various nitrogen sources (NaNO3 , NH4 Cl, NaNO2 , and an amino acid mixture) on the microalgae. The results demonstrated that the NRT2, NAR1, GLN2, GSN1, and AAT2 genes were highly expressed in lipid-rich microalgae under inorganic nitrogen-deficient conditions and they decreased after nitrogen resupply. Significant increases in the expressions of NAR1, GLN2, and GLSF were found in nitrate-depleted microalgae, whereas significant increases in the expressions of NRT2, NAR1, GLN2, and GSN1 were found in nitrite-depleted microalgae. Significant increases in the expressions of only NRT2 and GSN1 were found in ammonium-depleted microalgae (P<0.05). Except for the NRT2, other genes were expressed at lower levels under amino acid-deficient conditions compared with amino acid-sufficient controls. The expression of the NIA2 gene decreased in nitrogen-depleted microalgae regardless of the initial nitrogen source. However, the results of fatty acid analyses showed that the features of fatty acid profiles followed a similar mode, in which the percentage compositions of C16:0 and C18:1Δ(9) increased in nitrogen-depleted cells and that of C16:1Δ(9) , C18:3Δ(9,12,15) , C18:4Δ(6,9,12,15) , and C18:5Δ(3,6,9,12,15) decreased, regardless of the type of nitrogen source applied. It was also found that the epiphytic bacterium Alteromonas macleodii played a particularly important role in releasing microalgae from the stress of amino acid deficiency. These findings also provide a foundation for regulating microalgal lipid production through manipulation of the nitrogen assimilation-associated genes.

Coordinated response of photosynthesis, carbon assimilation, and triacylglycerol accumulation to nitrogen starvation in the marine microalgae Isochrysis zhangjiangensis (Haptophyta)

The photosynthetic performance, carbon assimilation, and triacylglycerol accumulation of Isochrysis zhangjiangensis under nitrogen-deplete conditions were studied to understand the intrinsic correlations between them. The nitrogen-deplete period was divided into two stages based on the photosynthetic parameters. During the first stage, carbon assimilation was not reduced compared with that under favorable conditions. The marked increase in triacylglycerols and the variation in the fatty acid profile suggested that triacylglycerols were mainly derived from de novo synthesized acyl groups. In the second stage, the triacylglycerol content continued increasing while the carbohydrate content decreased from 44.0% to 26.3%. These results indicated that the intracellular conversion of carbohydrates to triacylglycerols occurred. Thus, we propose that sustainable carbon assimilation and incremental triacylglycerol production can be achieved by supplying appropriate amounts of nitrogen in medium to protect the photosynthetic process from severe damage using the photosynthetic parameters as indicators.

Identification of carbohydrates as the major carbon sink of the marine microalga Isochrysis zhangjiangensis (Haptophyta) and optimization of its productivity by nitrogen manipulation

Microalgae represent a potential feedstock for biofuel production. During cultivation under nitrogen-depleted conditions, carbohydrates, rather than neutral lipids, were the major carbon sink of the marine microalga Isochrysis zhangjiangensis (Haptophyta). Carbohydrates reached maximum levels of 21.2pgcell−1 on day 5, which was an increase of more than 7-fold from day 1, while neutral lipids simultaneously increased 1.9-fold from 4.0 to 7.6pgcell−1 during the ten-day nitrogen-depleted cultivation. The carbohydrate productivity of I. zhangjiangensis was improved by optimization of the nitrate supply mode. The maximum carbohydrate concentration was 0.95gL−1 under batch cultivation, with an initial nitrogen concentration of 31.0mgL−1, which was 2.4-fold greater than that achieved under nitrogen-depleted conditions. High performance liquid chromatography (HPLC) analysis showed that the accumulated carbohydrate in I. zhangjiangensis was composed of glucose. These results show that I. zhangjiangensis represents an ideal carbohydrate-enriched bioresource for biofuel production.