I-Son Ng

Phototrophic cultivation of a thermo-tolerant Desmodesmus sp. for lutein production: Effects of nitrate concentration, light intensity and fed-batch operation

Four indigenous thermo-tolerant Desmodesmus sp. strains were examined for their ability to produce lutein. Among them, Desmodesmus sp. F51 was the best strain for this purpose. The medium composition, nitrate concentration and light intensity were manipulated to improve the phototrophic growth and lutein production of Desmodesmus sp. F51. It was found that a nitrogen-sufficient condition was required for lutein accumulation, while a high light intensity enhanced cell growth but caused a decrease in the lutein content. The best cell growth and lutein production occurred when the light intensity and initial nitrate concentration were 600 μmol/m(2)/s and 8.8 mM, respectively. The fed-batch cultivation strategy was shown to further improve lutein production. The highest lutein productivity (3.56±0.10 mg/L/d) and content (5.05±0.20 mg/g) were obtained when pulse-feeding of 2.2 mM nitrate was employed. This study demonstrated the potential of using Desmodesmus sp. F51 as a lutein producer in practical applications.

Understanding interactive characteristics of bioelectricity generation and reductive decolorization using Proteus hauseri.

This first-attempt study quantitatively explored interactive characteristics of bioelectricity generation and dye decolorization in air-cathode single-chamber microbial fuel cells (MFCs) using indigenous Proteus hauseri ZMd44. After approx. 15 cycles (30 days) acclimatization in dye-bearing cultures, P. hauseri could express its stable capability of simultaneous bioelectricity generation and color removal (SBP&CR) in MFCs. Evidently, appropriate acclimation strategy for formation of the electrochemically active anodic biofilm played a crucial role to enhance the performance of SBP&CR in MFCs. Gradually increased supplementations of C.I. reactive blue 160 resulted in progressively decreased decay rate of bioelectricity generation. That is, a dye decolorized in a faster rate would result in a lower capability for bioelectricity generation and vice versa. In addition, a reduced dye with less toxicity potency (e.g., 2-aminophenol) might work as a redox mediator of electron transport to anodic biofilm for bioelectricity generation in MFCs.

Dynamic synergistic effect on Trichoderma reesei cellulases by novel β-glucosidases from Taiwanese fungi.

Dynamic synergistic effects in cellulosic bioconversion have been revealed between Trichoderma reesei cellulases and β-glucosidases (BGLs) from six Taiwanese fungi. A high level of synergy (8.9-fold) was observed with the addition of Chaetomella raphigera BGL to T. reesei cellulases. In addition, the C. raphigera BGL possessed the highest activity (V(max)/K(m)=46.6 U/mg mM) and lowest glucose inhibition (Ki=4.6mM) with the substrate 4-nitrophenyl β-d-glucopyranoside. For the natural cellobiose substrate, however, the previously isolated Aspergillus niger BGL Novo-188 had the highest V(max)/K(m) (0.72 U/mg mM) and lowest Ki (59.5mM). The demonstrated dynamic synergistic effects between some BGLs and the T. reesei cellulase system suggest that BGLs not only prevent the inhibition by cellobiose, but also enhance activities of endo- and exo-cellulases in cellulosic bioconversion. Comparisons of kinetic parameters and synergism analyses between BGLs and T. reesei cellulases can be used for further optimization of the cellulosic bioconversion process.

Impact of carbon and nitrogen feeding strategy on high production of biomass and docosahexaenoic acid (DHA) by Schizochytrium sp. LU310

A new isolated Schizochytrium sp. LU310 from the mangrove forest of Wenzhou, China, was found as a high producing microalga of docosahexaenoic acid (DHA). In this study, the significant improvements for DHA fermentation by the batch mode in the baffled flasks (i.e. higher oxygen supply) were achieved. By applied the nitrogen-feeding strategy in 1000 mL baffled flasks, the biomass, DHA concentration and DHA productivity were increased by 110.4%, 117.9% and 110.4%, respectively. Moreover, DHA concentration of 21.06 g/L was obtained by feeding 15 g/L of glucose intermittently, which was an increase of 41.25% over that of the batch mode. Finally, an innovative strategy was carried out by intermittent feeding carbon and simultaneously feeding nitrogen. The maximum DHA concentration and DHA productivity in the fed-batch cultivation reached to 24.74 g/L and 241.5 mg/L/h, respectively.

Enhanced integration of large DNA into E. coli chromosome by CRISPR/Cas9.

Metabolic engineering often necessitates chromosomal integration of multiple genes but integration of large genes into Escherichia coli remains difficult. CRISPR/Cas9 is an RNA‐guided system which enables site‐specific induction of double strand break (DSB) and programmable genome editing. Here, we hypothesized that CRISPR/Cas9‐triggered DSB could enhance homologous recombination and augment integration of large DNA into E. coli chromosome. We demonstrated that CRISPR/Cas9 system was able to trigger DSB in >98% of cells, leading to subsequent cell death, and identified that mutagenic SOS response played roles in the cell survival. By optimizing experimental conditions and combining the λ‐Red proteins and linear dsDNA, CRISPR/Cas9‐induced DSB enabled homologous recombination of the donor DNA and replacement of lacZ gene in the MG1655 strain at efficiencies up to 99%, and allowed high fidelity, scarless integration of 2.4, 3.9, 5.4, and 7.0 kb DNA at efficiencies approaching 91%, 92%, 71%, and 61%, respectively. The CRISPR/Cas9‐assisted gene integration also functioned in different E. coli strains including BL21 (DE3) and W albeit at different efficiencies. Taken together, our methodology facilitated precise integration of dsDNA as large as 7 kb into E. coli with efficiencies exceeding 60%, thus significantly ameliorating the editing efficiency and overcoming the size limit of integration using the commonly adopted recombineering approach. Biotechnol. Bioeng. 2017;114: 172–183.

Deciphering simultaneous bioelectricity generation and dye decolorization using Proteus hauseri

This first-attempt study disclosed how and why electron-shuttling mediators were capable to stimulate bioelectricity-generating capabilities of dye-bearing microbial fuel cells (MFCs) using Proteus hauseri. Due to significant biotoxicity of 4-aminophenol (4AP) and the absence of electron-mediating potential of 3AP, only 2AP among all isomers could work as an exogenous mediator to stimulate bioelectricity generation of P. hauseri. Dye toxicity to cells on anodic biofilm in MFCs apparently affected the performance of simultaneous bioelectricity production and color removal (SBP&CR) in MFCs. Plus, dose-response analysis upon toxicity potency of reactive blue 160 revealed that cells on anodic biofilm in MFCs had a higher tolerance to reactive blue 160 than suspended cells. Apparently, augmentation of electron mediator(s) with low toxicity was a feasible means to facilitate bioelectricity-generating capability of SBP&CR.

Deciphering mediating characteristics of decolorized intermediates for reductive decolorization and bioelectricity generation.

As decolorized intermediates could play a role of electron-shuttling mediator to enhance the performance of dye decolorization and bioelectricity generation, this study selected model compounds with auxochromes (e.g., benzene-1,2-diol, 1,2-diaminobenzene) to explore how chemical structure(s) affected color removal and power producing capabilities in microbial fuel cells (MFCs). According to cyclic voltammetry, respiratory testing and MFC data, promising electron-shuttling capabilities of aforementioned compounds were revealed using Proteus hauseri ZMd44, Aeromonas sp. C78, Acinetobacter johnsonii NIUx72 bearing MFCs. These findings clearly indicated that chemical structure(s) of decolorized mediators directly affected characteristics of simultaneous reductive decolorization and bioelectricity generation in MFCs, suggesting feasible operation strategy of MFCs for industrial applications.

Flotation: A promising microalgae harvesting and dewatering technology for biofuels production

Microalgal biomass as renewable energy source is believed to be of great potential for reliable and sustainable biofuels production. However, microalgal biomass production is pinned by harvesting and dewatering stage thus hindering the developing and growing microalgae biotechnology industries. Flotation technology applied in mineral industry could be potentially applied in microalgae harvesting and dewatering, however substantial knowledge on different flotation units is essential. This paper presents an overview on different flotation units as promising cost‐effective technologies for microalgae harvesting thus bestowing for further research in development and commercialization of microalgae based biofuels. Dispersed air flotation was found to be less energy consuming. Moreover, Jameson cell flotation and dispersed ozone flotation are believed to be energy efficient microalgae flotation approaches. Microalgae harvesting and dewatering by flotation is still at embryonic stage, therefore extended studies with the focus on life cycle assessment, sustainability of the flotation unit, optimization of the operating parameters using different algal species is imperative. Though there are a number of challenges in microalgae harvesting and dewatering, with well designed and developed cultivation, harvesting/dewatering, extraction and conversion technologies, progressively, microalgae technology will be of great potential for biological carbon sequestration, biofuels and biochemicals production.

Synergistic effect of Trichoderma reesei cellulases on agricultural tea waste for adsorption of heavy metal Cr(VI)

This is the first attempt to study the synergistic effect between Trichoderma reesei cellulases and the abundant agricultural tea waste in absorption of heavy metal Cr(VI) as well as its kinetic model development. The properties of tea waste were first analyzed by near infrared spectroscopy (NIR), particle size distribution (PSD), X-ray photoelectron spectroscopy (XPS) and scanning electron microscopy (SEM) examination with EDX for comparison between its original (UN-TW) and cellulase-hydrolyzed (TRCEL-TW) conditions. Then, an advanced kinetic model in the form of -d[Cr(VI)]/dt = A[H+](n)e(-Ea/RT) [Cr(VI)](m)(0), which can successfully predict the time-dependent Cr(VI) concentration of various pHs, initial Cr(VI) concentrations and temperatures was developed. The demonstrated synergistic effects of T. reesei cellulases on tea waste suggested that cellulosic material provides more accessibility area for absorption of heavy metal. This study also provides an alternative approach to remove toxic Cr(VI) from aqueous solutions and extend the utilization of agricultural tea waste.

Exploring new strains of dye-decolorizing bacteria

This study unveiled a new strategy to explore new indigenous strains with excellent decolorization capabilities from freshwaters and seawaters. Two new bacterial decolorizers DX2b and SH7b, which have the capability to decolorize textile dyes, were isolated from Cross-Strait Taiwan and China. According to PCR-augmented 16S rRNA gene analyses for strain identification, >99% of nucleotide sequences in isolated strains were identical to type strains Rahnella aquatilis, Acinetobacter guillouiae, Microvirgula aerodenitrificans, and Pseudomonas sp. Time-series inspection upon azoreductase activity assay and generation of decolorized intermediates all confirmed in parallel with reductive decolorization of new decolorizers DX2b and SH7b. The result also showed that bacterial decolorization of these new strains was mainly catalyzed via the enzymatic expression of azoreductase and riboflavin reductase, and biosorption seemed not to play a crucial role color removal (approximately <10%).