Erythromycin mediates co-flocculation between cyanobacterium Synechocystis sp. PCC 6803 and filamentous fungi in liquid cultivation without organic compounds.

Photoautotrophic cyanobacteria assimilate the greenhouse gas carbon dioxide as their sole carbon source for producing useful bioproducts. However, harvesting the cells from their liquid media is a major bottleneck in the process. Thus, an easy-to-harvest method, such as auto-flocculation, is desirable. Here, we found that cyanobacterium Synechocystis sp. PCC 6803 co-flocculated with a natural fungal contamination in the presence of the antibiotic erythromycin (EM) but not without EM. The fungi in the co-flocculated biomass were isolated and found to consist of five species with the filamentous Purpureocillium lilacinum and Aspergillus protuberus making up 71% of the overall fungal population. The optimal co-cultivation for flocculation was an initial 5 mg (fresh weight) of fungi, an initial cell density of Synechocystis of 0.2 OD730, 10 µM EM, and 14 days of cultivation in 100 mL of BG11 medium with no organic compound. This yielded 248 ± 28 mg/L of the Synechocystis-fungi flocculated biomass from 560 ± 35 mg/L of total biomass, a 44 ± 2% biomass flocculation efficiency. Furthermore, the EM treated Synechocystis cells in the Synechocystis-fungi flocculate had a normal cell color and morphology, while those in the axenic suspension exhibited strong chlorosis. Thus, the occurrence of the Synechocystis-fungi flocculation was mediated by EM, and the co-flocculation with the fungi protected Synechocystis against the development of chlorosis. Transcriptomic analysis suggested that the EM-mediated co-flocculation was a result of down-regulation of the minor pilin genes and up-regulation of several genes including the chaperone gene for pilin regulation, the S-layer protein genes, the exopolysaccharide-polymerization gene, and the genes for signaling proteins involved in cell attachment and abiotic-stress responses. The CuSO4 stress can also mediate Synechocystis-fungi flocculation but at a lower flocculation efficiency than that caused by EM. The EM treatment may be applied in the co-culture between other cyanobacteria and fungi to mediate cell bio-flocculation.

Detoxification of water hyacinth hydrolysate mediated by exopolysaccharide-based hydrogel enhances hydrogen and methane production.

In this study, the exopolysaccharide from cyanobacteria was used for detoxification of acid hydrolysate of water hyacinth biomass. Exopolysaccharide-hydrogel showed phenolics and furans removal of 86 % and 97 %, respectively, with sugar recovery of 98.3 %. The fermentation of detoxified acid hydrolysate was integrated with that of pretreated biomass subjected to enzymatic saccharification derived from commercial cellulose (ESF) or from microbe (MSF). The maximum hydrogen production of 69.2 mL/g-VS was obtained in MSF, which is 1.2- and 1.6-fold higher than ESF and undetoxified acid hydrolysate, respectively. Additionally, the methane production of 12.6 mL/g-VS by mixed methanogenic consortia was obtained using the spent liquor containing volatile fatty acids. This enhanced hydrogen and methane production in subsequent microbial processes is mainly attributed to the selective removal of inhibitors in combination with an integrated carbohydrate utilization.

Microalgal-bacterial immobilized co-culture as living biofilters for nutrient recovery from synthetic wastewater and their potential as biofertilizers.

This study investigates nutrient recovery from synthetic municipal wastewater using co-immobilized cultures of Chlorella vulgaris TISTR 8580 (CV) and plant growth-promoting bacteria, Bacillus subtilis TISTR 1415 (BS) as living biofilters for a subsequent biofertilizer activity. The optimal condition for nutrient recovery was at the 1:1 ratio of CV/BS using mixed guar gum/carrageenan (GG/CG) binders. After 7-day wastewater treatment, the living biofilters removed 86.7 ± 0.5% of ammonium and 99.3 ± 0.3% of phosphates and were tested subsequently as biofertilizers for 20 days to grow selected plants. The highest optimal biomass and chlorophyll a content was 2 ± 0.3 g (CV/BS 3:1) and 12.4 ± 0.7 µg/g (CV/BS 1:1) from cucumber respectively, however, the close-to-neutral pH (8.0 ± 0.3) was observed from sunflower using CV/BS 1:1 living biofilters. Conclusively, the designed living biofilters exhibit the potential to recover nutrients from wastewater and be used as biofertilizers for circular agriculture.

Physicochemical and photocatalytic properties of biogenic ZnO and its chitosan nanocomposites for UV-protection and antibacterial activity on coated textiles.

The textiles for medical use and the purification of textile factory effluents have become the most crucial part of the human healthcare sector. In this study bioactive compounds produced by four distinct plant extracts were used for the synthesis of zinc oxide nanoparticles. The four different ZnO nanoparticles were comprehensively characterized by different analytical techniques. XRD analysis revealed the crystalline nature and phase purity of the ZnO nanoparticles. FTIR spectra provided information on the function of plant extracts in the stabilization or capping process. The size distribution and morphological diversity of the nanoparticles were further clarified by SEM and TEM images. The photocatalytic degradation activity of the four ZnO nanoparticles on two different dyes showed that ZnO nanoparticles prepared from A. indica were most effective for the degradation of 98 % and 91 % of Rhodamine B and Alizarin red dye respectively. The selected ZnO nanoparticles from A. indica were used to prepare ZnO-chitosan nanocomposites before coating on cotton fabrics. The hydrophobicity, UV protection factor, and antibacterial activity of ZnO-chitosan nanocomposites, when coated on cotton fabrics, were also examined. The overall results demonstrated the ZnO and ZnO-chitosan nanocomposite prepared in the present study as a promising material for environmental remediation application.

Augmentin loaded functionalized halloysite nanotubes: A sustainable emerging nanocarriers for biomedical applications.

Clay minerals such as Halloysite nanotubes (HNTs), abundantly available green nanomaterial, exhibit a significant advantage in biomedical applications such as drug delivery, antibacterial and antimicrobials, tissue engineering or regeneration, etc. Because of the mesoporous structure and high absorbability, HNTs exhibit great potential as a nanocarrier in drug delivery applications. The sulfuric acid treatment enhances the surface area of the HNTs and thereby improves their drug-loading capacity by enlarging their lumen space/inner diameter. In the present investigation, based on the literature that supports the efficacy of drug loading after acid treatment, a dual treatment was performed to functionalize the HNTs surface. First, the HNTs were etched and functionalized using sulfuric acid. The acid-functionalized HNTs underwent another treatment using (3-aminopropyl) triethoxysilane (APTES) to better interact the drug molecules with the HNTs surfaces for efficient drug loading. Augmentin, a potential drug molecule of the penicillin group, was used for HNTs loading, and their antibacterial properties, cytotoxicity, and cumulative drug release (%) were evaluated. Different characterization techniques, such as X-ray diffractometer (XRD) and Fourier Transform Infra-Red (FT-IR), confirm the loading of Augmentin to the APTES@Acid HNTs. TEM images confirm the effective loading of the drug molecule with the HNTs. The drug encapsulation efficiency shows 40.89%, as confirmed by the Thermogravimetric Analysis (TGA). Also, the Augmentin-loaded APTES@Acid HNTs exhibited good antibacterial properties against E. coli and S. aureus and low cytotoxicity, as confirmed by the MTT assay. The drug release studies confirmed the sustainable release of Augmentin from the APTES@Acid HNTs. Hence, the treated HNTs can be considered as a potential nanocarrier for effectively delivering Augmentin and promoting enhanced therapeutic benefits.

Novel smart fiber/metal/chitosan composite as a filter for self-detoxifying photocatalytic wastewater remediation and biomedical applications.

Wastewater treatment remains the most significant delinquent issue world-wide. Generally, wastewater treatment involves filtration followed by acidified de-emulsification through photocatalytic reduction. The aim of the present study is to reduce the use of nanoparticles in wastewater treatment and also to find an appropriate alternative to replace cotton fiber filters used in water treatment plant. The cotton fiber filters are highly prone to bacterial film development leading to bactericidal degradation of the fibers. We developed a ZnO-chitosan nanocomposite coated fiber for wastewater treatment to enhance its photocatalytic activity under acidic condition. The fiber showed high degree of photocatalytic degradation activity, reducing rhodamine B dye, chemical oxygen demand and chromium levels in the synthetic wastewater to 37, 79 and 51% respectively under highly acidic condition. Additionally, ZnO-chitosan nanocomposite did not cause mortality on Danio rerio embryo after 72 h incubation. The ZnO-chitosan nanocomposite coated fiber showed strong antibacterial activity against Escherichia coli and Staphylococcus aureus with a reduction of 96% and 99% respectively. This study demonstrated the potential of a novel smart fiber in wastewater treatment and biomedical applications.

Extracellular self-photosensitizer combined with metal oxide-based nano bio-hybrid system encapsulated by alginate improves hydrogen production in the presence of oxygen.

The photocatalytic nano-biohybrid systems have great potential for the conversion of solar energy to fermentative hydrogen production. Herein, a whole-cell nano-biohybrid system consisting of biosynthesized cadmium sulfide, Enterobacter aerogenes cells, and metal oxide nanoparticles was constructed. The system was encapsulated with sodium alginate and used for light-driven biohydrogen production under anaerobic and in the presence of oxygen conditions. After 48 h incubation in the presence of oxygen, the E. aerogenes cells with the encapsulated hybrid system yielded 2.7 mmol H2/mmol glucose, a 13.5-fold higher than that of the E. aerogenes cells without encapsulation. The encapsulated hybrid system could produce hydrogen for up to 96 h and could produce hydrogen even under natural sunlight conditions. These results revealed that efficient hydrogen production is possible in the presence of oxygen. Overall, the present study demonstrated the potential of using proper nano-biohybrid system with encapsulation for the production of hydrogen under ambient air condition.

Determination of Poly(3-hydroxybutyrate) Content in Cyanobacterium Synechocystis sp. PCC 6803 Using Acid Hydrolysis Followed by High-performance Liquid Chromatography.

Various photoautotrophic cyanobacteria accumulate intracellular poly(3-hydroxybutyrate) (PHB) granules. This protocol can be used for determining the PHB contents of the cells as % PHB weight per dry cell weight using acid hydrolysis followed by high-performance liquid chromatography (HPLC). This HPLC analysis is rapid, with a running time of approximately 5 min per sample. The technique can accurately determine PHB concentrations in the range of 2-1,000 µg/mL PHB. However, this technique is not applicable for determining the contents of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) in cyanobacteria.

Characterization and Homology Modeling of Catalytically Active Recombinant PhaCAp Protein from Arthrospira platensis.

Polyhydroxybutyrate (PHB) is a biocompatible and biodegradable polymer that has the potential to replace fossil-derived polymers. The enzymes involved in the biosynthesis of PHB are ß-ketothiolase (PhaA), acetoacetyl-CoA reductase (PhaB), and PHA synthase (PhaC). PhaC in Arthrospira platensis is the key enzyme for PHB production. In this study, the recombinant E. cloni®10G cells harboring A. platensis phaC (rPhaCAp) was constructed. The overexpressed and purified rPhaCAp with a predicted molecular mass of 69 kDa exhibited Vmax, Km, and kcat values of 24.5 ± 2 µmol/min/mg, 31.3 ± 2 µM and 412.7 ± 2 1/s, respectively. The catalytically active rPhaCAp was a homodimer. The three-dimensional structural model for the asymmetric PhaCAp homodimer was constructed based on Chromobacterium sp. USM2 PhaC (PhaCCs). The obtained model of PhaCAp revealed that the overall fold of one monomer was in the closed, catalytically inactive conformation whereas the other monomer was in the catalytically active, open conformation. In the active conformation, the catalytic triad residues (Cys151-Asp310-His339) were involved in the binding of substrate 3HB-CoA and the CAP domain of PhaCAp involved in the dimerization.

Scenedesmus sp. strain SD07 cultivation in municipal wastewater for pollutant removal and production of lipid and exopolysaccharides.

In this study, an efficient microalgal strain SD07 was isolated from pond wastewater and identified as Scenedesmus sp. using the 18S rRNA gene sequence analysis. The strain SD07 was grown in a variety of concentrations (25-100%) of municipal wastewater. Scenedesmus sp. strain SD07 grown in 75% diluted wastewater produced a higher amount of biomass (1.93 ± 0.10 g L-1), and removal of chemical oxygen demand (COD), ammonium (NH4+), total nitrogen (TN) and total phosphate (TP) by 91.36%, 88.41%, 93.26% and 96.32%, respectively from wastewater. The harvested strain SD07 biomass has protein, carbohydrate and lipid contents of 35%, 20.4% and 33%, respectively. Fatty acid profiles revealed that the strain SD07 lipids mainly consist of palmitic acid (40.5%), palmitoleic acid (19%), linoleic acid (17%) and oleic acid (13.2%). Furthermore, strain SD07 cultured in 75% diluted wastewater produced 378 mg L-1 of exopolysaccharides (EPS). The EPS was utilized as a biostimulant in the cultivation of Solanum lycopersicum under salinity stress. In summary, these findings suggest that this Scenedesmus sp. strain SD07 can be employed for wastewater treatment as well as the production of valuable biomass, high-quality algal oil and EPS.

Chemical Triggering Cyanobacterial Glycogen Accumulation: Methyl Viologen Treatment Increases Synechocystis sp. PCC 6803 Glycogen Storage by Enhancing Levels of Gene Transcript and Substrates in Glycogen Synthesis.

Two-stage cultivation is effective for glycogen production by cyanobacteria. Cells were first grown under adequate nitrate supply (BG11) to increase biomass and subsequently transferred to nitrogen deprivation (-N) to stimulate glycogen accumulation. However, the two-stage method is time-consuming and requires extensive energy. Thus, one-stage cultivation that enables both cell growth and glycogen accumulation is advantageous. Such one-stage method could be achieved using a chemical triggering glycogen storage. However, there is a limited study on such chemicals. Here, nine compounds previously reported to affect cyanobacterial cellular functions were examined in Synechocystis sp. PCC 6803. 2-Phenylethanol, phenoxyethanol, 3-(3,4-dichlorophenyl)-1,1-dimethylurea and methyl viologen can stimulate glycogen accumulation. The oxidative stress agent, methyl viologen significantly increased glycogen levels up to 57% and 69% [w/w dry weight (DW)] under BG11 and -N cultivation, respectively. One-stage cultivation where methyl viologen was directly added to the pre-grown culture enhanced glycogen storage to 53% (w/w DW), compared to the 10% (w/w DW) glycogen level of the control cells without methyl viologen. Methyl viologen treatment reduced the contents of total proteins (including phycobiliproteins) but caused increased transcript levels of glycogen synthetic genes and elevated levels of metabolite substrates for glycogen synthesis. Metabolomic results suggested that upon methyl viologen treatment, proteins degraded to amino acids, some of which could be used as a carbon source for glycogen synthesis. Results of oxygen evolution and metabolomic analysis suggested that photosynthesis and carbon fixation were not completely inhibited upon methyl viologen treatment, and these two processes may partially generate upstream metabolites required for glycogen synthesis.

Enhanced polyglucan contents in divergent cyanobacteria under nutrient-deprived photoautotrophy: transcriptional and metabolic changes in response to increased glycogen accumulation in nitrogen-deprived Synechocystis sp. PCC 6803.

Cyanobacteria accumulate polyglucan as main carbohydrate storage. Here, the cellular polyglucan content was determined in 27 cyanobacterial strains from 25 genera. The polyglucan contents were significantly enhanced in 20 and 23 strains under nitrogen (-N) and phosphate (-P) deprivation, respectively. High polyglucan accumulation was not associated with particular evolutionary groups but was strain specific. The highest polyglucan accumulations of 46.2% and 52.5% (w/w dry weight; DW) were obtained under -N in Synechocystis sp. PCC 6803 (hereafter Synechocystis) and Chroococcus limneticus, respectively. In Synechocystis, 80-97% (w/w) of the polyglucan was glycogen. Transcriptome and metabolome analyses during glycogen accumulation under -N were determined in Synechocystis. The genes responsible for the supply of the substrates for glycogen synthesis: glycerate-1,3-phosphate and fructose-1,6-phosphate, were significantly up-regulated. The genes encoding the enzymes converting succinate to malate in TCA cycle, were significantly down-regulated. The genes encoding the regulator proteins which inhibits metabolism at lower part of glycolysis pathway, were also significantly up-regulated. The transcript levels of PII protein and the level of 2-oxoglutarate, which form a complex that inhibits lower part of glycolysis pathway, were significantly increased. Thus, the increased Synechocystis glycogen accumulation under -N was likely to be mediated by the increased supply of glycogen synthesis substrates and metabolic inhibitions at lower part of glycolysis pathway and TCA cycle.

Advancement on mixed microalgal-bacterial cultivation systems for nitrogen and phosphorus recoveries from wastewater to promote sustainable bioeconomy.

Biological wastewater treatment is a promising and environmentally friendly method that utilises living microorganisms to remediate water and enable recovery or conversion of contaminants into valuable products. For many decades, microalgae and cyanobacteria, photosynthetic living microorganisms, have been explored extensively for wastewater bioremediation. They can be used for recovering valuable nutrients such as nitrogen and phosphorous from secondary effluents and capable of transforming those nutrients into marketable products such as biofuels, biofertilisers, nutraceutical, and pigments for promoting a Bio-Circular Green economy. In recent years, there has been a shift towards mixing compatible microalgae with bacteria, which is inspired by their natural symbiotic relationships to increase nitrogen and phosphorus recoveries. With this enhanced bioremediation, recovery of polluted wastes can be intensified and higher biomass quality (with high nutrient density) can be achieved. This review focuses on the state-of-the-art of mixed microalgal-bacterial cultivating systems. A comprehensive comparison of existing studies that used Chlorella species as microalgae in various mixed microalgal-bacterial cultivating systems (suspension, biofilm, and immobilisation) for nitrogen and phosphorus recoveries from wastewater is conducted. Key technical challenges such as balancing microalgae and bacteria species, pH regulation, light distribution, biomass harvesting, and biomass conversion are also discussed. From the data comparisons among different cultivation systems, it has been suggested that immobilisation appears to require less amount of operational light compared to the suspended and biofilm-based systems for similar nitrogen and phosphorus removal efficiencies.

Simazine Enhances Dark Fermentative H2 Production by Unicellular Halotolerant Cyanobacterium Aphanothece halophytica.

The halotolerant cyanobacterium Aphanothece halophytica is a potential H2 producer that induces H2 evolution under nitrogen deprivation. H2 is mainly produced via the catabolism of stored glycogen under dark anaerobic condition. H2 evolution is catalyzed by O2-sensitive bidirectional hydrogenase. The aim of this study was to improve H2 production by A. halophytica using various kinds of inhibitors. Among all types of inhibitors, simazine efficiently promoted the highest H2 production under dark conditions. High simazine concentration and long-term incubation resulted in a decrease in cell and chlorophyll concentrations. The optimal simazine concentration for H2 production by A. halophytica was 25 µM. Simazine inhibited photosynthetic O2 evolution but promoted dark respiration, resulting in a decrease in O2 level. Hence, the bidirectional hydrogenase activity and H2 production was increased. A. halophytica showed the highest H2 production rate at 58.88 ± 0.22 µmol H2 g-1 dry weight h-1 and H2 accumulation at 356.21 ± 6.04 µmol H2 g-1 dry weight after treatment with 25 µM simazine under dark anaerobic condition for 2 and 24 h, respectively. This study demonstrates the potential of simazine for the enhancement of dark fermentative H2 production by A. halophytica.

Chemicals Affecting Cyanobacterial Poly(3-hydroxybutyrate) Accumulation: 2-Phenylethanol Treatment Combined with Nitrogen Deprivation Synergistically Enhanced Poly(3-hydroxybutyrate) Storage in Synechocystis sp. PCC6803 and Anabaena sp. TISTR8076.

Various photoautotrophic cyanobacteria increase the accumulation of bioplastic poly(3-hydroxybutyrate) (PHB) under nitrogen deprivation (-N) for energy storage. Several metabolic engineering enhanced cyanobacterial PHB accumulation, but these strategies are not applicable in non-gene-transformable strains. Alternatively, stimulating PHB levels by chemical exposure is desirable because it might be applied to various cyanobacterial strains. However, the study of such chemicals is still limited. Here, 19 compounds previously reported to affect bacterial cellular processes were evaluated for their effect on PHB accumulation in Synechocystis sp. PCC6803, where 3-(3,4-dichlorophenyl)-1,1-dimethylurea, methyl viologen, arsenite, phenoxyethanol and 2-phenylethanol were found to increase PHB accumulation. When cultivated with optimal nitrate supply, Synechocystis contained less than 0.5% [w/w dry weight (DW)] PHB, while cultivation under -N conditions increased the PHB content to 7% (w/w DW). Interestingly, the -N cultivation combined with 2-phenylethanol exposure reduced the Synechocystis protein content by 27% (w/w DW) but significantly increased PHB levels up to 33% (w/w DW), the highest ever reported photoautotrophic cyanobacterial PHB accumulation in a wild-type strain. Results from transcriptomic and metabolomic analysis suggested that under 2-phenylethanol treatment, Synechocystis proteins were degraded to amino acids, which might be subsequently utilized as the source of carbon and energy for PHB biosynthesis. 2-Phenylethanol treatment also increased the levels of metabolites required for Synechocystis PHB synthesis (acetyl-CoA, acetoacetyl-CoA, 3-hydroxybutyryl-CoA and NADPH). Additionally, under -N, the exposure to phenoxyethanol and 2-phenylethanol increased the PHB levels of Anabaena sp. from 0.4% to 4.1% and 6.6% (w/w DW), respectively. The chemicals identified in this study might be applicable for enhancing PHB accumulation in other cyanobacteria.

Metabolic transformation of cyanobacteria for biofuel production.

World-wide, an emerging demand is moving towards the biofuels to replace the fossil fuels. In alternative biofuel production strategies, cyanobacteria have unique characteristic of accumulating glycogen, lipid, and fuel molecules through natural mechanisms. Moreover, the cyanobacteria can be easily engineered to synthesis a plenty of fuel molecules from CO2. To obtain the fuel molecule from cyanobacteria, various techniques were invented in which the metabolic engineering is found to be a prerequisite to develop an economically feasible process. The expression of indigenous or heterologous pathways plays an important role in developing successful production process. In addition, the engineering of photosynthetic apparatus, destruction of competitive pathways and improvement of tolerance were also proven to improve the product specific synthesis. Although various metabolic engineering approaches have been developed, there are certain obstacles when it comes to implementation for the production. In this review, the important biosynthetic pathways for biofuels, alteration of other genes to improve the actual pathway and possibilities of developing cyanobacterial fuel production have been elaborated.

Microalgal Biorefinery Concepts' Developments for Biofuel and Bioproducts: Current Perspective and Bottlenecks.

Microalgae have received much interest as a biofuel feedstock. However, the economic feasibility of biofuel production from microalgae does not satisfy capital investors. Apart from the biofuels, it is necessary to produce high-value co-products from microalgae fraction to satisfy the economic aspects of microalgae biorefinery. In addition, microalgae-based wastewater treatment is considered as an alternative for the conventional wastewater treatment in terms of energy consumption, which is suitable for microalgae biorefinery approaches. The energy consumption of a microalgae wastewater treatment system (0.2 kW/h/m3) was reduced 10 times when compared to the conventional wastewater treatment system (to 2 kW/h/m3). Microalgae are rich in various biomolecules such as carbohydrates, proteins, lipids, pigments, vitamins, and antioxidants; all these valuable products can be utilized by nutritional, pharmaceutical, and cosmetic industries. There are several bottlenecks associated with microalgae biorefinery. Hence, it is essential to promote the sustainability of microalgal biorefinery with innovative ideas to produce biofuel with high-value products. This review attempted to bring out the trends and promising solutions to realize microalgal production of multiple products at an industrial scale. New perspectives and current challenges are discussed for the development of algal biorefinery concepts.

Overexpression of fatty acid synthesis genes in Synechocystis sp. PCC 6803 with disrupted glycogen synthesis increases lipid production with further enhancement under copper induced oxidative stress.

In the present study, fatty acid synthesis genes such as alpha and beta subunits of acetyl CoA carboxylase (accA and accD) were overexpressed in the glgC (Glucose-1-phosphate adenylyltransferase) knockout Synechocystis sp. PCC 6803. The biomass and lipid contents were evaluated in both the wild type and the engineered strains after copper treatment. The maximum lipid production of 0.981 g/L with the productivity of 81.75 mg/L/d was obtained from the copper treated ΔglgC + A-OX strain, which showed a 3.3-fold increase compared to the untreated wild type with satisfactory biodiesel properties. After copper treatment the knockout strain improved the unsaturated fatty acids level contributing to the increase of the saturated and mono-unsaturated ratio with improvement of the fuel quality. Copper induced oxidative stress also improved the photosynthetic pigments in engineered strains leading to increased tolerance against oxidative stress in the engineered strains. The copper treatment increased the antioxidant enzyme activities in the engineered strains especially in ΔglgC + A-OX strain. The carbon flux to lipid synthesis was enhanced by the engineered strains particularly with the knockout-overexpression strains. The Synechocystis sp. PCC 6803 engineered with ΔglgC + A-OX showed high potential for fuel production after the copper treatment.