Comparison of Metabarcoding and Microscopy Methodologies to Analyze Diatom Communities in Five Estuaries Along the Southern Coast of the Korean Peninsula.

The study of microalgal communities is critical for understanding aquatic ecosystems. These communities primarily comprise diatoms (Heterokontophyta), with two methods commonly used to study them: Microscopy and metabarcoding. However, these two methods often deliver different results; thus, their suitability for analyzing diatom communities is frequently debated and evaluated. This study used these two methods to analyze the diatom communities in identical water samples and compare the results. The taxonomy of the species constituting the diatom communities was confirmed, and both methods showed that species belonging to the orders Bacillariales and Naviculales (class Bacillariophyceae) are the most diverse. In the lower taxonomic levels (family, genus, and species), microscopy tended to show a bias toward detecting diatom species (Nitzschia frustulum, Nitzschia inconspicua, Nitzschia intermedia, Navicula gregaria, Navicula perminuta, Navicula recens, Navicula sp.) belonging to the Bacillariaceae and Naviculaceae families. The results of the two methods differed in identifying diatom species in the communities and analyzing their structural characteristics. These results are consistent with the fact that diatoms belonging to the genera Nitzschia and Navicula are abundant in the communities; furthermore, only the Illumina MiSeq data showed the abundance of the Melosira and Entomoneis genera. The results obtained from microscopy were superior to those of Illumina MiSeq regarding species-level identification. Based on the results obtained via microscopy and Illumina MiSeq, it was revealed that neither method is perfect and that each has clear strengths and weaknesses. Therefore, to analyze diatom communities effectively and accurately, these two methods should be combined.

Enhanced glutathione content improves lateral root development and grain yield in rice plants.

KEY MESSAGE: Enhanced glutathione content improves lateral root development by positively regulating the transcripts of root development genes responsive to glutathione treatment, thereby increasing the overall productivity of rice plants. Glutathione is primarily known as a cellular antioxidant molecule, but its role in lateral root development in rice plants has not been elucidated. Here, we have investigated its role in lateral root development of rice Oryza sativa L. Exogenous glutathione (GSH) promoted both the number and length of lateral roots in rice, and the GSH biosynthesis inhibitor buthionine sulfoximine (BSO) significantly reduced these parameters, compared to untreated plants. The inhibition by BSO was reversed with exogenous GSH. Transcript profiling by RNA-seq revealed that expression of the transcription factor genes DREB and ERF and the hormone-related genes AOS, LOX, JAZ, and SAUR were significantly downregulated in the BSO-treated plants and, in contrast, upregulated in plants treated with GSH and with GSH and BSO together. We generated OsGS-overexpressing transgenic plants in which the transgene is controlled by the abiotic-stress-inducible OsRab21 promoter to study the effect of endogenously increased GSH levels. In cold stress, transgenic rice plants enhanced stress tolerance and lateral root development by maintaining redox homeostasis and improving upregulating the expression of transcription factors and hormone-related genes involved in lateral root development. We observed improved root growth of OsGS-overexpressing plants in paddy fields compared to the wild-type controls. These traits may have alleviated transplanting stress during early growth in the field and accounted for the increased productivity. These results provide information and perspectives on the role of GSH in gene expression, lateral root development, and grain yield in rice.

Enhanced biomass and oxidative stress tolerance of Synechococcus elongatus PCC 7942 overexpressing the DHAR gene from Brassica juncea.

OBJECTIVES: To improve the oxidative stress tolerance, biomass yield, and ascorbate/dehydroascorbate (AsA/DHA) ratio of Synechococcus elongatus PCC 7942 in the presence of H2O2, by heterologous expression of the dehydroascorbate reductase (DHAR) gene from Brassica juncea (BrDHAR). RESULTS: Under H2O2 stress, overexpression of BrDHAR in the transgenic strain (BrD) of S. elongatus greatly increased the AsA/DHA ratio. As part of the AsA recycling system, the oxidative stress response induced by reactive oxygen species was enhanced, and intracellular H2O2 level decreased. In addition, under H2O2 stress conditions, the BrD strain displayed increased growth rate and biomass, as well as higher chlorophyll content and deeper pigmentation than did wild-type and control strains. CONCLUSION: By maintaining the AsA pool and redox homeostasis, the heterologous expression of BrDHAR increased S. elongatus tolerance to H2O2 stress, improving the biomass yield under these conditions. The results suggest that the BrD strain of S. elongatus, with its ability to attenuate the deleterious effects of ROS caused by environmental stressors, could be a promising platform for the generation of biofuels and other valuable bioproducts.

Crystal structures of aldehyde deformylating oxygenase from Limnothrix sp. KNUA012 and Oscillatoria sp. KNUA011.

The cyanobacterial aldehyde deformylating oxygenase (cADO) is a key enzyme that catalyzes the unusual deformylation of aliphatic aldehydes for alkane biosynthesis and can be applied to the production of biofuel in vitro and in vivo. In this study, we determined crystal structures of two ADOs from Limnothrix sp. KNUA012 (LiADO) and Oscillatoria sp. KNUA011 (OsADO). The structures of LiADO and OsADO resembled those of typical cADOs, consisting of eight α-helices found in ferritin-like di-iron proteins. However, structural comparisons revealed that while the LiADO active site was vacant of iron and substrates, the OsADO active site was fully occupied, containing both a coordinated metal ion and substrate. Previous reports indicated that helix 5 is capable of adopting two distinct conformations depending upon the existence of bound iron. We observed that helix 5 of OsADO with an iron bound in the active site presented as a long helix, whereas helix 5 of LiADO, which lacked iron in the active site, presented two conformations (one long and two short helices), indicating that an equilibrium exists between the two states in solution. Furthermore, acquisition of a structure having a fully occupied active site is unique in the absence of higher iron concentrations as compared with other cADO structures, wherein low affinity for iron complicates the acquisition of crystal structures with bound iron. An in-depth analysis of the ADO apo-enzyme, the enzyme with substrate bound, and the enzyme with both iron and substrate bound provided novel insight into substrate-binding modes in the absence of a coordinated metal ion and suggested a separate two-step binding mechanism for substrate and iron co-factors. Moreover, our results provided a comprehensive structural basis for conformational changes induced by binding of the substrate and co-factor.

Homologous expression of cytosolic dehydroascorbate reductase increases grain yield and biomass under paddy field conditions in transgenic rice (Oryza sativa L. japonica).

Dehydroascorbate reductase (DHAR, EC 1.8.5.1) maintains redox pools of ascorbate (AsA) by recycling oxidized AsA to reduced AsA. To investigate whether DHAR affects rice yield under normal environmental conditions, cDNA-encoding DHAR (OsDHAR1) was isolated from rice and used to develop OsDHAR1-overexpressing transgenic rice plants, under the regulation of a maize ubiquitin promoter. Incorporation and expression of the transgene in transgenic rice plants was confirmed by genomic polymerase chain reaction (PCR), semi-quantitative reverse transcription PCR (RT-PCR), western blot, and enzyme activity. The expression levels were at least twofold higher in transgenic (TG) rice plants than in control wild-type (WT) rice plants. In addition, OsDHAR1-overexpression in seven-independent homologous transgenic plants, as compared to WT plants, increased photosynthetic capacity and antioxidant enzyme activities under paddy field conditions, which led to an improved AsA pool and redox homeostasis. Furthermore, OsDHAR1 overexpression significantly improved grain yield and biomass due to the increase of culm and root weights and to enhance panicle and spikelet numbers in the same seven independent TG rice plants during the farming season (2010 and 2011) in South Korea. The OsDHAR protein contained the redox-active site (Cys20), as well as the conserved GSH-binding region, GSH-binding motif, glutathione-S-transferase (GST) N-terminal domain, C-terminal domain interface, and GST C-terminal domain. Therefore, our results indicate that OsDHAR1 overexpression, capable of functioning in AsA recycling, and protein folding increases environmental adaptation to paddy field conditions by the improving AsA pool and redox homeostasis, which enhances rice grain yield and biomass.

Expression of salt-induced 2-Cys peroxiredoxin from Oryza sativa increases stress tolerance and fermentation capacity in genetically engineered yeast Saccharomyces cerevisiae.

Peroxiredoxins (Prxs), also termed thioredoxin peroxidases (TPXs), are a family of thiol-specific antioxidant enzymes that are critically involved in cell defense and protect cells from oxidative damage. In this study, a putative chloroplastic 2-Cys thioredoxin peroxidase (OsTPX) was identified by proteome analysis from leaf tissue samples of rice (Oryza sativa) seedlings exposed to 0.1 M NaCl for 3 days. To investigate the relationship between the OsTPX gene and the stress response, OsTPX was cloned into the yeast expression vector p426GPD under the control of the glyceraldehyde-3-phosphate dehydrogenase (GPD1) promoter, and the construct was transformed into Saccharomyces cerevisiae cells. OsTPX expression was confirmed by semi-quantitative reverse transcription-polymerase chain reaction and western blot analyses. OsTPX contained two highly conserved cysteine residues (Cys114 and Cys236) and an active site region (FTFVCPT), and it is structurally very similar to human 2-Cys Prx. Heterologous OsTPX expression increased the ability of the transgenic yeast cells to adapt and recover from reactive oxygen species (ROS)-induced oxidative stresses, such as a reduction of cellular hydroperoxide levels in the presence of hydrogen peroxide and menadione, by improving redox homeostasis. OsTPX expression also conferred enhanced tolerance to tert-butylhydroperoxide, heat shock, and high ethanol concentrations. Furthermore, high OsTPX expression improved the fermentation capacity of the yeast during glucose-based batch fermentation at a high temperature (40 °C) and at the general cultivation temperature (30 °C). The alcohol yield in OsTPX-expressing transgenic yeast increased by approximately 29 % (0.14 g g(-1)) and 21 % (0.12 g g(-1)) during fermentation at 40 and 30 °C, respectively, compared to the wild-type yeast. Accordingly, OsTPX-expressing transgenic yeast showed prolonged cell survival during the environmental stresses produced during fermentation. These results suggest that heterologous OsTPX expression increases acquired tolerance to ROS-induced oxidative stress by improving cellular redox homeostasis and improves fermentation capacity due to improved cell survival during fermentation, especially at a high temperature.

Expression of dehydrin gene from Arctic Cerastium arcticum increases abiotic stress tolerance and enhances the fermentation capacity of a genetically engineered Saccharomyces cerevisiae laboratory strain.

We investigated Arctic plants to determine if they have a specific mechanism enabling them to adapt to extreme environments because they are subject to such conditions throughout their life cycles. Among the cell defense systems of the Arctic mouse-ear chickweed Cerastium arcticum, we identified a stress-responsive dehydrin gene CaDHN that belongs to the SK5 subclass and contains conserved regions with one S segment at the N-terminus and five K segments from the N-terminus to the C-terminus. To investigate the molecular properties of CaDHN, the yeast Saccharomyces was transformed with CaDHN. CaDHN-expressing transgenic yeast (TG) cells recovered more rapidly from challenge with exogenous stimuli, including oxidants (hydrogen peroxide, menadione, and tert-butyl hydroperoxide), high salinity, freezing and thawing, and metal (Zn(2+)), than wild-type (WT) cells. TG cells were sensitive to copper, cobalt, and sodium dodecyl sulfate. In addition, the cell survival of TG cells was higher than that of WT cells when cells at the mid-log and stationary stages were exposed to increased ethanol concentrations. There was a significant difference in cultures that have an ethanol content >16 %. During glucose-based batch fermentation at generally used (30 °C) and low (18 °C) temperatures, TG cells produced a higher alcohol concentration through improved cell survival. Specifically, the final alcohol concentrations were 13.3 and 13.2 % in TG cells during fermentation at 30 and 18 °C, respectively, whereas they were 10.2 and 9.4 %, respectively, in WT cells under the same fermentation conditions. An in vitro assay revealed that purified CaDHN acted as a reactive oxygen species scavenger by neutralizing H2O2 and a chaperone by preventing high temperature-mediated catalase inactivation. Taken together, our results show that CaDHN expression in transgenic yeast confers tolerance to various abiotic stresses by improving redox homeostasis and enhances fermentation capacity, especially at low temperatures (18 °C).

The Prokaryotic Microalga Limnothrix redekei KNUA012 to Improve Aldehyde Decarbonylase Expression for Use as a Biological Resource.

The prokaryotic microalga Limnothrix redekei KNUA012 isolated from a freshwater bloom sample from Lake Hapcheon, Hapcheon-gun, South Korea, was investigated for its potential as a biofuel feedstock. Microalgae produce straight-chain alkanes/alkenes from acyl carrier protein-linked fatty acyls via aldehyde decarbonylase (AD; EC 1.2.1.3), which can convert aldehyde intermediates into various biofuel precursors, such as alkanes and free fatty acids. In L. redekei KNUA012, long-chain ADs can convert fatty aldehyde intermediates into alkanes. After heterologous AD expression in Escherichia coli (pET28-AD), we identified an AD in L. redekei KNUA012 that can synthesize various alkanes, such as pentadecane (C15H32), 8-heptadecene (C17H34), and heptadecane (C17H36). These alkanes can be directly used as fuels without transesterification. Biodiesel constituents including dodecanoic acid (C13H26O2), tetradecanoic acid (C15H30O2), 9-hexa decenoic acid (C17H32O2), palmitoleic acid (C17H32O2), hexadecanoic acid (C17H34O2), 9-octadecenoic acid (C19H36O2), and octadecanoic acid (C19H38O2) are produced by L. redekei KNUA012 as the major fatty acids. Our findings suggest that Korean domestic L. redekei KNUA012 is a promising resource for microalgae-based biofuels and biofuel feedstock.

Glutathione reductase from Brassica rapa affects tolerance and the redox state but not fermentation ability in response to oxidative stress in genetically modified Saccharomyces cerevisiae.

To determine whether the exogenous expression of glutathione reductase (GR) from Brassica rapa subsp. pekinensis (BrGR) can reduce the deleterious effects of unfavorable conditions, we constructed a transgenic Saccharomyces cerevisiae strain bearing the GR gene cloned into the yeast expression vector, pVTU260. BrGR expression was confirmed by semi reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, immunoblotting analysis and an enzyme assay. Ectopic BrGR-expression improved cellular glutathione (GSH) homeostasis after higher GSH accumulation in the transgenic yeast than in the wild-type yeast under H(2)O(2)-induced oxidative stress. The BrGR-expressing yeast strain induced the activation of metabolic enzymes (Hxt, G6PDH, GAPDH and Ald), antioxidant systems (Gpx, Trx2, Trx3, Trr1, Tsa1 and porin) and molecular chaperones (Hsp104, Hsp90, Hsp70, Hsp42, Hsp26, Grp, Sti1 and Zpr1), which led to lower oxidative protein damage after a reduction in the level of cellular ROS in the BrGR-expressing yeast strain exposed to H(2)O(2) than in the wild-type yeast strain. BrGR-expression increased the ability to adapt and recover from H(2)O(2)-induced oxidative stress and various stressors, including heat shock, menadione, tert-butyl hydroperoxide, heavy metals, sodium dodecyl sulfate, ethanol and NaCl, but did not affect fermentation capacity. These results suggest that ectopic BrGR expression confers acquired tolerance by improving proteostasis and redox homeostasis through co-activation of various cell rescue proteins against ROS-induced oxidative stress in yeast cells.

The microbial communities (bacteria, algae, zooplankton, and fungi) improved biofloc technology including the nitrogen-related material cycle in Litopenaeus vannamei farms.

Microbes are essential in biofloc technology for controlling nitrogen levels in water. The composition and function of microorganisms with biofloc systems were reported; however, data on microorganisms other than bacteria, such as algae (which are essential in the nitrogen cycle) and zooplankton (which are bacterial and algal predators), remain limited. The microbial communities (including bacteria, algae, zooplankton, and fungi) were investigated in shrimp farms using biofloc technology. Using Illumina MiSeq sequencing, the V4 region of 18S rRNA and the V3-V4 region of 16S rRNA were utilized for the analysis of the eukaryotic and prokaryotic microbial communities. As a result, it was found that the biofloc in the shrimp farm consisted of 48.73%-73.04% eukaryotic organisms and 26.96%-51.27% prokaryotic organisms. In these shrimp farms, prokaryotic microbial communities had higher specie richness and diversity than eukaryotic microbial communities. However, the eukaryotic microbial communities were more abundant than their prokaryotic counterparts, while algae and zooplankton dominated them. It was discovered that the structures of the microbial communities in the shrimp farms seemed to depend on the effects of predation by zooplankton and other related organisms. The results provided the nitrogen cycle in biofloc systems by the algal and bacterial groups in microbial communities.

Over-Expression of Dehydroascorbate Reductase Improves Salt Tolerance, Environmental Adaptability and Productivity in Oryza sativa.

Abiotic stress induces reactive oxygen species (ROS) generation in plants, and high ROS levels can cause partial or severe oxidative damage to cellular components that regulate the redox status. Here, we developed salt-tolerant transgenic rice plants that overexpressed the dehydroascorbate reductase gene (OsDHAR1) under the control of a stress-inducible sweet potato promoter (SWPA2). OsDHAR1-expressing transgenic plants exhibited improved environmental adaptability compared to wild-type plants, owing to enhanced ascorbate levels, redox homeostasis, photosynthetic ability, and membrane stability through cross-activation of ascorbate-glutathione cycle enzymes under paddy-field conditions, which enhanced various agronomic traits, including root development, panicle number, spikelet number per panicle, and total grain yield. dhar2-knockdown plants were susceptible to salt stress, and owing to poor seed maturation, exhibited reduced biomass (root growth) and grain yield under paddy field conditions. Microarray revealed that transgenic plants highly expressed genes associated with cell growth, plant growth, leaf senescence, root development, ROS and heavy metal detoxification systems, lipid metabolism, isoflavone and ascorbate recycling, and photosynthesis. We identified the genetic source of functional genomics-based molecular breeding in crop plants and provided new insights into the physiological processes underlying environmental adaptability, which will enable improvement of stress tolerance and crop species productivity in response to climate change.

Enhancement of Biomass Production in Colony-Forming Green Algae, Botryosphaerella sudetica, Under Mixotrophic Cultivation.

In this study, we characterized the potential of colony-forming green algae, Botryosphaerella sudetica KNUA107, isolated from Ulleung Island, South Korea, as a bioresource and analyzed the effects of mixotrophic cultivation on its bioresource production efficiency. Internal transcribed spacer (ITS) (ITS1, 5.8S, and ITS2), ribulose bisphosphate carboxylase large subunit (rbcL), and elongation factor Tu (tufa) regions were used for molecular identification and phylogenetic analysis. B. sudetica KNUA107 had a strong relationship with the green algae of Botryococcus and Botryosphaerella genera, which are colony-forming species, and was also associated with members of the Neochloris genus. To improve biomass productivity, we tested mixotrophic cultivation conditions using several organic carbon sources. Glucose supplementation stimulated B. sudetica KNUA107 growth and reduced the time needed to reach the stationary phase. In addition, the colony size was 1.5-2.0 times larger with glucose than in photoautotrophic cultures, and settleability improved in proportion to colony size. The total lipid content and biomass productivity were also higher in cultures supplemented with glucose. Among the lipid components, saturated fatty acids and monounsaturated fatty acids had the highest proportion. Our study suggests that B. sudetica KNUA107, which has enhanced efficiency in biomass production and lipid components under mixotrophic cultivation, has high potential as a bioresource.

Effect of Different Cultivation Modes (Photoautotrophic, Mixotrophic, and Heterotrophic) on the Growth of Chlorella sp. and Biocompositions.

In the past, biomass production using microalgae culture was dependent on inorganic carbon sources as microalgae are photosynthetic organisms. However, microalgae utilize both organic and inorganic carbon sources, such as glucose. Glucose is an excellent source of organic carbon that enhances biomass yield and the content of useful substances in microalgae. In this study, photoautotrophic, mixotrophic, and heterotrophic cultivation conditions were applied to three well-known strains of Chlorella (KNUA104, KNUA114, and KNUA122) to assess biomass productivity, and compositional changes (lipid, protein, and pigment) were evaluated in BG11 media under photoautotrophic, mixotrophic, and heterotrophic conditions utilizing different initial concentrations of glucose (5, 10, 15, 20, and 25 g L-1). Compared to the photoautotrophic condition (biomass yield: KNUA104, 0.35 ± 0.04 g/L/d; KNUA114, 0.40 ± 0.08 g/L/d; KNUA122, 0.38 ± 0.05 g/L/d) glucose was absent, and the biomass yield improved in the mixotrophic (glucose: 20 g L-1; biomass yield: KNUA104, 2.99 ± 0.10 g/L/d; KNUA114, 5.18 ± 0.81 g/L/d; KNUA122, 5.07 ± 0.22 g/L/d) and heterotrophic conditions (glucose: 20 g L-1; biomass yield: KNUA104, 1.72 ± 0.26 g/L/d; KNUA114, 4.26 ± 0.27 g/L/d; KNUA122, 4.32 ± 0.32 g/L/d). All strains under mixotrophic and heterotrophic conditions were optimally cultured when 15-20 g L-1 initial glucose was provided. Although bioresourse productivity improved under both mixotrophic and heterotrophic conditions where mixotrophic conditions were found to be optimal as the yields of lipid and pigment were also enhanced. Protein content was less affected by the presence of light or the concentration of glucose. Under mixotrophic conditions, the highest lipid content (glucose: 15 g L-1; lipid content: 68.80 ± 0.54%) was obtained with Chlorella vulgaris KNUA104, and enhanced pigment productivity of Chlorella sorokiniana KNUA114 and KNUA122 (additional pigment yield obtained with 15 g L-1 glucose: KNUA 114, 0.33 ± 0.01 g L-1; KNUA122, 0.21 ± 0.01 g L-1). Also, saturated fatty acid (SFA) content was enhanced in all strains (SFA: KNUA104, 29.76 ± 1.31%; KNUA114, 37.01 ± 0.98%; KNUA122, 33.37 ± 0.17%) under mixotrophic conditions. These results suggest that mixotrophic cultivation of Chlorella vulgaris and Chlorella sorokiniana could improve biomass yield and the raw material quality of biomass.

Environmental Factors Associated with the Eukaryotic Microbial Community and Microalgal Groups in the Mountain Marshes of South Korea.

The diversity indices of eukaryotic microalgal groups in the Jeonglyeongchi, Waegok, and Wangdeungjae marshes of Mount Jiri, Korea, were measured using Illumina MiSeq and culture-based analyses. Waegok marsh had the highest species richness, with a Chao1 value of 828.00, and the highest levels of species diversity, with Shannon and Simpson index values of 6.36 and 0.94, respectively, while Wangdeungjae marsh had the lowest values at 2.97 and 0.75, respectively. The predominant species in all communities were Phagocata sibirica (Jeonglyeongchi, 68.64%), Aedes albopictus (Waegok, 34.77%), Chaetonotus cf. (Waegok, 24.43%), Eimeria sp. (Wangdeungjae, 26.17%), and Eumonhystera cf. (Wangdeungjae, 22.27%). Relative abundances of the microalgal groups Bacillariophyta (diatoms) and Chlorophyta (green algae) in each marsh were respectively: Jeonglyeongchi 1.38% and 0.49%, Waegok 7.0% and 0.3%, and Wangdeungjae 10.41% and 4.72%. Illumina MiSeq analyses revealed 34 types of diatoms and 13 types of green algae. Only one diatom (Nitzschia dissipata) and five green algae (Neochloris sp., Chlamydomonas sp., Chlorococcum sp., Chlorella vulgaris, Scenedesmus sp.) were identified by a culture-based analysis. Thus, Illumina MiSeq analysis can be considered an efficient tool for analyzing microbial communities. Overall, our results described the environmental factors associated with geographically isolated mountain marshes and their respective microbial and microalgal communities.

Expression of yeast cyclophilin A (Cpr1) provides improved stress tolerance in Escherichia coli.

Cyclophilins contain the conserved activity of cis-trans peptidyl-prolyl isomerase that is implicated in protein folding and function as molecular chaperones. The yeast cyclophilin A gene (cpr1) was subcloned to the prokaryotic expression vector pKM260. It was found that the expression of Cpr1 drastically increased the cell viability of E. coli BL21 in the presence of abiotic stress conditions, such as cadmium, copper, hydrogen peroxide, heat, and SDS. Thus, this study illustrates the importance of Cpr1 as a molecular chaperone that improved cellular stress responses when E. coli cells were exposed to adverse conditions, and it also shows the possibility of increasing the stability of E. coli strains utilized for the production of recombinant proteins.

Characterization of Chlorella sorokiniana and Chlorella vulgaris fatty acid components under a wide range of light intensity and growth temperature for their use as biological resources.

This study aims to characterize the potential of three strains of microalgal species (Chlorella sorokiniana KNUA114 and KNUA122; C. vulgaris KNUA104) for use as feedstock, based on their fatty acid compositions. Each strain was molecularly identified using four marker genes (ITS, SSU, rbcL, and tufA) and phylogenetically characterized. C. sorokiniana and C. vulgaris collected from Ulleung Island, South Korea, were homologous with other known species groups. Samples' fatty acid components were measured using GC/MS analysis in growth temperatures of 10 °C, 25 °C, and 35 °C. The growth rate of C. sorokiniana strains was higher than that of C. vulgaris under high-temperature conditions, confirming the potential industrial applicability of the former as feedstock material. Additionally, saturated fatty acid contents and productivities increased as biological resources of the C. sorokiniana strains were higher than those of C. vulgaris under high light intensity and temperature conditions. These results suggest that the fatty acid components of C. sorokiniana strains may potentially be used as biological resources (e.g., feedstock).

Effects of Co-culture on Improved Productivity and Bioresource for Microalgal Biomass Using the Floc-Forming Bacteria Melaminivora Jejuensis.

Bacterial and algal floc formation was induced by inoculating three species of wastewater-derived bacteria (Melaminivora jejuensis, Comamonas flocculans, and Escherichia coli) into algal cultures (Chlorella sorokiniana). Bacterial and algal flocs formed in algal cultures inoculated with M. jejuensis and C. flocculans, and these flocs showed higher sedimentation rates than pure algal culture. The floc formed by M. jejuensis (4988.46 ± 2589.81 µm) was 10-fold larger than the floc formed by C. flocculans (488.60 ± 226.22 µm), with a three-fold higher sedimentation rate (M. jejuensis, 91.08 ± 2.32% and C. flocculans, 32.55 ± 6.33%). Biomass and lipid productivity were improved with M. jejuensis inoculation [biomass, 102.25 ± 0.35 mg/(L·day) and 57.80 ± 0.20 mg/(L·day)] compared with the productivity obtained under pure algal culture conditions [biomass, 78.00 ± 3.89 mg/(L·day) and lipids, 42.26 ± 2.11 mg/(L·day)]. Furthermore, the fatty acid composition of the biomass produced under pure algal culture conditions was mainly composed of C16:0 (43.67%) and C18:2 (45.99%), whereas the fatty acid composition of the biomass produced by M. jejuensis was mainly C16:0 (31.80%), C16:1 (24.45%), C18:1 (20.23%), and C18:2 (16.11%). These results suggest the possibility of developing an efficient method for harvesting microalgae using M. jejuensis and provide information on how to improve biomass productivity using floc-forming bacteria.

Expression of Heterologous OsDHAR Gene Improves Glutathione (GSH)-Dependent Antioxidant System and Maintenance of Cellular Redox Status in Synechococcus elongatus PCC 7942.

An excess of reactive oxygen species (ROS) can cause severe oxidative damage to cellular components in photosynthetic cells. Antioxidant systems, such as the glutathione (GSH) pools, regulate redox status in cells to guard against such damage. Dehydroascorbate reductase (DHAR, EC 1.8.5.1) catalyzes the glutathione-dependent reduction of oxidized ascorbate (dehydroascorbate) and contains a redox active site and glutathione binding-site. The DHAR gene is important in biological and abiotic stress responses involving reduction of the oxidative damage caused by ROS. In this study, transgenic Synechococcus elongatus PCC 7942 (TA) was constructed by cloning the Oryza sativa L. japonica DHAR (OsDHAR) gene controlled by an isopropyl ß-D-1-thiogalactopyranoside (IPTG)-inducible promoter (Ptrc) into the cyanobacterium to study the functional activities of OsDHAR under oxidative stress caused by hydrogen peroxide exposure. OsDHAR expression increased the growth of S. elongatus PCC 7942 under oxidative stress by reducing the levels of hydroperoxides and malondialdehyde (MDA) and mitigating the loss of chlorophyll. DHAR and glutathione S-transferase activity were higher than in the wild-type S. elongatus PCC 7942 (WT). Additionally, overexpression of OsDHAR in S. elongatus PCC 7942 greatly increased the glutathione (GSH)/glutathione disulfide (GSSG) ratio in the presence or absence of hydrogen peroxide. These results strongly suggest that DHAR attenuates deleterious oxidative effects via the glutathione (GSH)-dependent antioxidant system in cyanobacterial cells. The expression of heterologous OsDHAR in S. elongatus PCC 7942 protected cells from oxidative damage through a GSH-dependent antioxidant system via GSH-dependent reactions at the redox active site and GSH binding site residues during oxidative stress.

Illumina MiSeq Analysis and Comparison of Freshwater Microalgal Communities on Ulleungdo and Dokdo Islands.

Ulleungdo and Dokdo are volcanic islands with an oceanic climate located off the eastern coast of South Korea. In the present study, we used barcoded Illumina MiSeq to analyze eukaryotic microalgal genera collected from Seonginbong, the highest peak on Ulleungdo, and from groundwater sites on Dongdo and Seodo Islands, which are part of Dokdo. Species richness was significantly greater in the Seonginbong samples than in the Dongdo and Seodo samples, with 834 operational taxonomic units (OTUs) identified from Seonginbong compared with 203 OTUs and 182 OTUs from Dongdo and Seodo, respectively. Taxonomic composition analysis was also used to identify the dominant microalgal phyla at each of the three sites, with Chlorophyta (green algae) the most abundant phyla on Seonginbong and Dongdo, and Bacillariophyta (diatoms) the most abundant on Seodo. These findings suggest that differences in the abundances of Chlorophyta and Bacillariophyta species in the Seonginbong, Dongdo, and Seodo samples are due to variations in species richness and freshwater resources at each sampling location. To the best of our knowledge, this is the first report to detail freshwater microalgal communities on Ulleungdo and Dokdo. As such, the number of species identified in the Seonginbong, Dongdo, and Seodo samples might be an indicator of the ecological differences among these sites and varying characteristics of their microbial communities. Information regarding the microalgal communities also provides a basis for understanding the ecological interactions between microalgae species and other eukaryotic microorganisms.Ulleungdo and Dokdo are volcanic islands with an oceanic climate located off the eastern coast of South Korea. In the present study, we used barcoded Illumina MiSeq to analyze eukaryotic microalgal genera collected from Seonginbong, the highest peak on Ulleungdo, and from groundwater sites on Dongdo and Seodo Islands, which are part of Dokdo. Species richness was significantly greater in the Seonginbong samples than in the Dongdo and Seodo samples, with 834 operational taxonomic units (OTUs) identified from Seonginbong compared with 203 OTUs and 182 OTUs from Dongdo and Seodo, respectively. Taxonomic composition analysis was also used to identify the dominant microalgal phyla at each of the three sites, with Chlorophyta (green algae) the most abundant phyla on Seonginbong and Dongdo, and Bacillariophyta (diatoms) the most abundant on Seodo. These findings suggest that differences in the abundances of Chlorophyta and Bacillariophyta species in the Seonginbong, Dongdo, and Seodo samples are due to variations in species richness and freshwater resources at each sampling location. To the best of our knowledge, this is the first report to detail freshwater microalgal communities on Ulleungdo and Dokdo. As such, the number of species identified in the Seonginbong, Dongdo, and Seodo samples might be an indicator of the ecological differences among these sites and varying characteristics of their microbial communities. Information regarding the microalgal communities also provides a basis for understanding the ecological interactions between microalgae species and other eukaryotic microorganisms.

ASR Enhances Environmental Stress Tolerance and Improves Grain Yield by Modulating Stomatal Closure in Rice.

Abscisic acid-, stress-, and ripening-induced (ASR) genes are involved in responding to abiotic stresses, but their precise roles in enhancing grain yield under stress conditions remain to be determined. We cloned a rice (Oryza sativa) ASR gene, OsASR1, and characterized its function in rice plants. OsASR1 expression was induced by abscisic acid (ABA), salt, and drought treatments. Transgenic rice plants overexpressing OsASR1 displayed improved water regulation under salt and drought stresses, which was associated with osmolyte accumulation, improved modulation of stomatal closure, and reduced transpiration rates. OsASR1-overexpressing plants were hypersensitive to exogenous ABA and accumulated higher endogenous ABA levels under salt and drought stresses, indicating that OsASR1 is a positive regulator of the ABA signaling pathway. The growth of OsASR1-overexpressing plants was superior to that of wild-type (WT) plants under paddy field conditions when irrigation was withheld, likely due to improved modulation of stomatal closure via modified ABA signaling. The transgenic plants had higher grain yields than WT plants for four consecutive generations. We conclude that OsASR1 has a crucial role in ABA-mediated regulation of stomatal closure to conserve water under salt- and drought-stress conditions, and OsASR1 overexpression can enhance salinity and drought tolerance, resulting in improved crop yields.