De Novo Transcriptomic Analysis of an Oleaginous Microalga: Pathway Description and Gene Discovery for Production of Next-Generation Biofuels

Background: Eustigmatos cf. polyphem is a yellow-green unicellular soil microalga belonging to the eustimatophyte with high biomass and considerable production of triacylglycerols (TAGs) for biofuels, which is thus referred to as an oleaginous microalga. The paucity of microalgae genome sequences, however, limits development of gene-based biofuel feedstock optimization studies. Here we describe the sequencing and de novo transcriptome assembly for a non-model microalgae species, E. cf. polyphem, and identify pathways and genes of importance related to biofuel production. Results: We performed the de novo assembly of E. cf. polyphem transcriptome using Illumina paired-end sequencing technology. In a single run, we produced 29,199,432 sequencing reads corresponding to 2.33 Gb total nucleotides. These reads were assembled into 75,632 unigenes with a mean size of 503 bp and an N50 of 663 bp, ranging from 100 bp to.3,000 bp. Assembled unigenes were subjected to BLAST similarity searches and annotated with Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) orthology identifiers. These analyses identified the majority of carbohydrate, fatty acids, TAG and carotenoids biosynthesis and catabolism pathways in E. cf. polyphem. Conclusions: Our data provides the construction of metabolic pathways involved in the biosynthesis and catabolism of carbohydrate, fatty acids, TAG and carotenoids in E. cf. polyphem and provides a foundation for the molecular genetics and functional genomics required to direct metabolic engineering efforts that seek to enhance the quantity and character o

Effects of nutrients and light intensity on the growth and biochemical composition of a marine microalga Odontella aurita

Algal biotechnology has advanced greatly in the past three decades. Many microalgae are now cultivated to produce bioactive substances. Odontella aurita is a marine diatom industrially cultured in outdoor open ponds and used for human nutrition. For the first time, we have systematically investigated the effects of culture conditions in cylindrical glass columns and flat-plate photobioreactors, including nutrients (nitrogen, phosphorus, silicon, and sulfur), light intensity and light path, on O. aurita cell growth and biochemical composition (protein, carbohydrate, β-1,3-glucan, lipids, and ash). The optimal medium for photoautotrophic cultivation of O. aurita contained 17.65 mmol/L nitrogen, 1.09 mmol/L phosphorus, 0.42 mmol/L silicon, and 24.51 mmol/L sulfur, yielding a maximum biomass production of 6.1–6.8 g/L and 6.7–7.8 g/L under low and high light, respectively. Scale-up experiments were conducted with flat-plate photobioreactors using different light-paths, indicating that a short light path was more suitable for biomass production of O. aurita. Analyses of biochemical composition showed that protein content decreased while carbohydrate (mainly composed of β-1,3-glucan) increased remarkably to about 50% of dry weight during the entire culture period. The highest lipid content (19.7% of dry weight) was obtained under 0.11 mmol/L silicon and high light conditions at harvest time. Fatty acid Profiles revealed that 80% were C14, C16, and C20, while arachidonic acid and eicosapentaenoic acid (EPA) accounted for 1.6%–5.6% and 9%–20% of total fatty acids, respectively. High biomass production and characteristic biochemical composition Profiles make O. aurita a promising microalga for the production of bioactive components, such as EPA and β-1,3-glucan

Serum Cystatin C within 24 hours after admission: a potential predictor for acute kidney injury in Chinese patients with community acquired pneumonia

AbstractBackground Acute kidney injury (AKI) is common in patients with community-acquired pneumonia (CAP), and is associated with poor prognosis. Therefore, in this study, we evaluated whether AKI in Chinese patients with CAP could be well predicted by serum Cystatin C within 24 h after admission.Methods Univariate and multivariate logistic regression analyses were used to investigate independent factors of AKI in patients with CAP.Results Totally, 2716 patients with CAP were included in this study. 766 (28%) patients developed AKI. After multivariate logistic regression analysis, serum Cystatin C (odds ratio [OR] 4.27, 95% confidence interval [CI] 3.36–5.44; p < 0.001) was an independent factor for AKI in patients with CAP. Serum Cystatin C had an area under the receiver operating characteristic curve (AUC) of 0.81 for predicting AKI, with an optimal cutoff value of 1.37 mg/L, computing 68% sensitivity, 80% specificity. Furthermore, serum Cystatin C within 24 h after admission still had a good and stable prediction efficiency for AKI in various subgroups (age, gender, hypertension, diabetes, coronary artery disease, cardiac insufficiency, cerebrovascular disease, atrial fibrillation, chronic obstructive pulmonary disease, chronic kidney disease, and tumor, albumin, anemia, platelet count, white blood cell count, and uric acid, confusion, uremia, respiratory rate, blood pressure, and age 65 years or older [CURB-65] score, acute respiratory failure, intensive care unit admission, and mechanical ventilation) of patients with CAP (AUCs: 0.69–0.84).Conclusion Serum Cystatin C within 24 h after admission appears to be a good biomarker for predicting AKI in Chinese patients with CAP

COG annotations of putative proteins.

<p>All putative proteins were aligned to the COG database and can be classified functionally into at least 25 molecular families. A, RNA processing and modification; B, Chromatin structure and dynamics; C, Energy production and conversion; D, Cell cycle control, cell division, chromosome partitioning; E, Amino acid transport and metabolism; F, Nucleotide transport and metabolism; G, Carbohydrate transport and metabolism; H, Coenzyme transport and metabolism; I, Lipid transport and metabolism; J, Translation, ribosomal structure and biogenesis; K, Transcription; L, Replication, recombination and repair; M, Cell wall/membrane/envelope biogenesis; N, Cell motility; O, Posttranslational modification, protein turnover, chaperones; P, Inorganic ion transport and metabolism; Q, Secondary metabolites biosynthesis, transport and catabolism; R, General function prediction only; S, Function unknown; T, Signal transduction mechanisms; U, Intracellular trafficking, secretion, and vesicular transport; V, Defense mechanisms; W, Extracellular structures; Y, Nuclear structure; Z, Cytoskeleton.</p

Fatty acid biosynthesis pathway reconstructed based on the <i>de novo</i> assembly and annotation of <i>E.</i> cf. <i>polyphem</i> transcriptome.

<p>Identified enzymes are shown in boxes and include: ACCase, acetyl-CoA carboxylase (EC: 6.4.1.2); MAT, malonyl-CoA ACP transacylase (EC: 2.3.1.39); KAS, 3-ketoacyl ACP synthase (KAS I, EC: 2.3.1.41; KASII, EC: 2.3.1.179; KAS III, EC: 2.3.1.180); KAR, 3-ketoacyl ACP reductase (EC: 1.1.1.100); HD, 3-hydroxy acyl-CoA dehydratase (EC: 4.2.1.-); EAR, enoyl-ACP reductase (NADH) (EC: 1.3.1.9); AAD, Δ9 Acyl-ACP desaturase (EC: 1.14.19.2); OAT, oleoyl-ACP thioesterase (EC: 3.1.2.14); Δ12D, Δ12(ω6)-desaturase (EC: 1.4.19.6); Δ15D, Δ15(ω3)-desaturase (EC: 1.4.19.-); Δ5D, Δ5- desaturase(EC: 1.14.99.-), Δ6D, Δ6- desaturase(EC: 1.14.99.-) and Δ6E, Δ6-elongase (EC: 6.21.3.-). The fatty acid biosynthesis pathway in <i>E.</i> cf. <i>polyphem</i> produces saturated, PA, palmitic acid (16:0) and SA, stearic acid (18:0), and unsaturated fatty acids OA, oleic acid (18:1ω9); LA, linoleic acid (18:2ω6); ALA, α-linolenic acid (18:3ω3); SDA, stearidonic acid (18:4ω3); ETA, eicosatetraenoic acid (20:4ω3) and EPA, eicosapentaenoic acid (20:5ω3).</p

Summary for the <i>E.</i> cf. <i>polyphem</i> transcriptome.

<p>Summary for the <i>E.</i> cf. <i>polyphem</i> transcriptome.</p

Chrysolaminarin biosynthesis and degradation pathway reconstructed based on the <i>de novo</i> assembly and annotation of <i>E.</i> cf. <i>polyphem</i> transcriptome.

<p>Identified enzymes are shown in boxes and include: UGPase, UDP glucose pyrophosphorylase (EC: 2.7.7.9); UDPG, chrysolaminarin synthase (EC: 2.4.1.34); exo-Glu, exo-1,3-β-glucanase (EC: 3.2.1.58); endo-Glu, endo-1,3-β-glucanase (EC: 3.2.1.39) and BGL, β-glucosidases (EC: 3.2.1.21). G-1-P, glucose-1-phosphate; PPi, pyrophosphate.</p

Glycolysis pathway reconstructed based on the <i>de novo</i> assembly and annotation of <i>E.</i> cf. <i>polyphem</i> transcriptome.

<p>Identified enzymes are shown in boxes and include: HK, hexokinase (EC:2.7.1.1); GCK, glucokinase (EC: 2.7.1.2); G6PI, glucose-6-phosphate isomerase (EC: 5.3.1.9); PFK, phosphofructokinase-6 (EC: 2.7.1.11); FBA, fructose-bisphosphate aldolase (EC:4.1.2.13); TPI, triose-phosphate isomerase (EC: 5.3.1.1); GAPDH, glyceraldehyde-3-phosphate dehydrogenase (EC: 1.2.1.9, 1.2.1.12); GPDH, glycerol-3-phosphate dehydrogenase (EC:1.1.1.8); PGK, phosphoglycerate kinase (EC: 2.7.2.3); PGAM, phosphoglycerate mutase (EC: 5.4.2.1); ENO, enolase (EC: 4.2.1.11); PK, pyruvate kinase (EC: 2.7.1.40); PDC, pyruvate decarboxylase (EC: 4.1.1.1); ADH, alcohol dehydrogenase (EC: 1.1.1.1); PDHC, the pyruvate dehydrogenase complex consisting of PDHB, pyruvate dehydrogenase (acetyl-transferring) (EC: 1.2.4.1), DLAT, dihydrolipoamide acetyltransferase (EC: 2.3.1.12), DLD, dihydrolipoyl dehydrogenase (EC: 1.8.1.4). G-6-P, glucose-6-phosphate; F-6-P, fructose 6-phosphate; FBP, fructose-1,6-bisphosphate; GA3P, glyceraldehyde-3-phosphate; DHAP, dihydroxyacetone phosphate; G-3-P, glycerol-3-phosphate; 1,3BPG, 1, 3-bisphosphoglycerate; 3PG, 3-phosphoglycerate; 2PG, 2-phosphoglycerate; PEP, phosphoenolpyruvate.</p

Annotation of non-redundant consensus sequences.

<p>All 14,982 CDS sequences generated by ESTscan were annotated though Swissprot, Nr, GO, KEGG, and COG databases.</p