Methyl jasmonate treatment affects the regulation of the 2-C-methyl-D-erythritol 4-phosphate pathway and early steps of the triterpenoid biosynthesis in Chlamydomonas reinhardtii

© 2019 Elsevier B.V. Terpenoids are a large and diverse class of naturally occurring metabolites serving many industrial applications and natural roles. Economically important terpenoids are often produced in low abundance from their natural sources, making their industrial-scale production challenging or uneconomical, therefore engineered microorganisms are frequently used as heterologous production platforms. Photosynthetic microorganisms, such as the green alga Chlamydomonas reinhardtii, represent promising systems to produce terpenoids in a cost-effective and sustainable manner, but knowledge about the regulation of their terpenoid metabolism remains limited. Here we report on the investigation of the phytohormone methyl jasmonate (MeJA) as elicitor of algal terpenoid synthesis. We treated C. reinhardtii cells in mid-exponential growth phase with three different concentrations of MeJA (0.05, 0.5 and 1 mM). The highest concentration of MeJA affected the photosynthetic activity of the cells, arrested the growth and up-regulated key genes of the 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway, leading to a significant increase in intermediates of this pathway, squalene and (S)-2,3-epoxysqualene, while the abundance of cycloartenol, and two main sterols (ergosterol and 7-dehydroporiferasterol) decreased. These data suggest the redirection of the carbon flux towards the synthesis of yet uncharacterised triterpenoid secondary metabolites upon MeJA treatment. Our results offer important new insights into the regulation of the triterpenoid metabolism in C. reinhardtii and raise important questions on hormonal signalling in microalgae. Phytohormone treatment is tested for the first time in algae, where it holds great potential for identifying key transcriptional regulators of the MEP pathway as targets for future metabolic engineering studies for improve production of high-value triterpenoids

Genome sequencing as a new window into the microbial community of membrane bioreactors – A critical review

© 2019 Elsevier B.V. Recent developed sequencing techniques have resulted in a new and unprecedented way to study biological wastewater treatment, in which most organisms are uncultivable. This review provides (i) an insight on state-of-the-art sequencing techniques and their limitations; (ii) a critical assessment of the microbial community in biological reactor and biofouling layer in a membrane bioreactor (MBR). The data from high-throughput sequencing has been used to infer microbial growth conditions and metabolisms of microorganisms present in MBRs at the time of sampling. These data shed new insight to two fundamental questions about a microbial community in the MBR process namely the microbial composition (who are they?) and the functions of each specific microbial assemblage (what are their function?). The results to date also highlight the complexity of the microbial community growing on MBRs. Environmental conditions are dynamic and diverse, and can influence the diversity and structural dynamics of any given microbial community for wastewater treatment. The benefits of understanding the structure of microbial communities on three major aspects of the MBR process (i.e. nutrient removal, biofouling control, and micropollutant removal) were symmetrically delineated. This review also indicates that the deployment of microbial community analysis for a practical engineering context, in terms of process design and system optimization, can be further realized

Methyl Jasmonate and Methyl-β-Cyclodextrin Individually Boost Triterpenoid Biosynthesis in <i>Chlamydomonas Reinhardtii</i> UVM4.

The commercialisation of valuable plant triterpenoids faces major challenges, including low abundance in natural hosts and costly downstream purification procedures. Endeavours to produce these compounds at industrial scale using microbial systems are gaining attention. Here, we report on a strategy to enrich the biomass of the biotechnologically-relevant Chlamydomonas reinhardtii strain UVM4 with valuable triterpenes, such as squalene and (S)-2,3-epoxysqualene. C. reinhardtii UVM4 was subjected to the elicitor compounds methyl jasmonate (MeJA) and methyl-β-cyclodextrine (MβCD) to increase triterpene yields. MeJA treatment triggered oxidative stress, arrested growth, and altered the photosynthetic activity of the cells, while increasing squalene, (S)-2,3-epoxysqualene, and cycloartenol contents. Applying MβCD to cultures of C. reinhardtii lead to the sequestration of the two main sterols (ergosterol and 7-dehydroporiferasterol) into the growth medium and the intracellular accumulation of the intermediate cycloartenol, without compromising cell growth. When MβCD was applied in combination with MeJA, it counteracted the negative effects of MeJA on cell growth and physiology, but no synergistic effect on triterpene yield was observed. Together, our findings provide strategies for the triterpene enrichment of microalgal biomass and medium

Validation of a cationic polyacrylamide flocculant for the harvesting fresh and seawater microalgal biomass

© 2019 Elsevier B.V. A simple, efficient, and fast settling flocculation technique to harvest microalgal biomass was demonstrated using a proprietary cationic polyacrylamide flocculant for a freshwater (Chlorella vulgaris) and a marine (Phaeodactylum tricornutum) microalgal culture at their mid-stationary growth phase. The optimal flocculant doses were 18.9 and 13.7 mg/g of dry algal biomass for C. vulgaris and P. tricornutum, respectively (equivalent to 7 g per m3 of algal culture for both species). The obtained optimal dose was well corroborated with changes in cell surface charge, and culture solution optical density and turbidity. At the optimal dose, charge neutralization of 64 and 86% was observed for C. vulgaris and P. tricornutum algal cells, respectively. Algae recovery was independent of the culture solution pH in the range of pH 6 to 9. Algal biomass recovery was achieved of 100 and 90% for C vulgaris and P. tricornutum respectively, and over 98% medium recovery was achievable by simple decanting

Molecular physiology reveals ammonium uptake and related gene expression in the seagrass Zostera muelleri

© 2016 Elsevier Ltd Seagrasses are important marine foundation species, which are presently threatened by coastal development and global change worldwide. The molecular mechanisms that drive seagrass responses to anthropogenic stresses, including elevated levels of nutrients such as ammonium, remains poorly understood. Despite the evidence that seagrasses can assimilate ammonium by using glutamine synthetase (GS)/glutamate synthase (glutamine-oxoglutarate amidotransferase or GOGAT) cycle, the regulation of this fundamental metabolic pathway has never been studied at the gene expression level in seagrasses so far. Here, we combine (i) reverse transcription quantitative real-time PCR (RT-qPCR) to measure expression of key genes involved in the GS/GOGAT cycle, and (ii) stable isotope labelling and mass spectrometry to investigate 15N-ammonium assimilation in the widespread Australian species Zostera muelleri subsp. capricorni (Z. muelleri). We demonstrate that exposure to a pulse of ammonium in seawater can induce changes in GS gene expression of Z. muelleri, and further correlate these changes in gene expression with 15N-ammonium uptake rate in above- and below-ground tissue

Maintenance of Geobacter-dominated biofilms in microbial fuel cells treating synthetic wastewater

© 2015 Elsevier B.V. Geobacter-dominated biofilms can be selected under stringent conditions that limit the growth of competing bacteria. However, in many practical applications, such stringent conditions cannot be maintained and the efficacy and stability of these artificial biofilms may be challenged. In this work, biofilms were selected on low-potential anodes (-0.36V vs Ag/AgCl, i.e. -0.08V vs SHE) in minimal acetate or ethanol media. Selection conditions were then relaxed by transferring the biofilms to synthetic wastewater supplemented with soil as a source of competing bacteria. We tracked community succession and functional changes in these biofilms. The Geobacter-dominated biofilms showed stability in their community composition and electrochemical properties, with Geobacter sp. being still electrically active after six weeks in synthetic wastewater with power densities of 100±19mW·m-2 (against 74±14mW·m-2 at week 0) for all treatments. After six weeks, the ethanol-selected biofilms, despite their high taxon richness and their efficiency at removing the chemical oxygen demand (0.8g·L-1 removed against the initial 1.3g·L-1 injected), were the least stable in terms of community structure. These findings have important implications for environmental microbial fuel cells based on Geobacter-dominated biofilms and suggest that they could be stable in challenging environments

Comment on microbial community composition Is unaffected by anode potential

While bioelectrochemical systems (BESs) such as microbial fuel cells (MFCs) or microbial electrolysis cells (MECs) remain promising technologies, their widespread use for the sustainable production of energy from wastewaters is yet to be realized. Establishment of anode-respiring bacteria (ARB) at the anode surface is considered an important determinant of BES performance. Because ARB use the anode for respiration, it should be possible to use anode potential as a mechanism to select various ARB communities with different electron transfer properties

Photosynthetic biocathode enhances the power output of a sediment-type microbial fuel cell

Conventional microbial fuel cells (MFCs) consist of biological anodes and abiotic cathodes separated by a proton-exchange membrane. The abiotic cathode usually catalyses the reduction of oxygen to produce water by means of expensive catalysts such as platinum. The cathodic reaction is often limiting in MFCs and researchers are now focusing on efficient, low-cost catalysts to improve oxygen reduction at the cathode. This paper describes a photosynthetic biocathode in a sediment-type MFC constructed without a proton-exchange membrane. The carbon and stainless steel cathode did not contain any catalyst, but was covered in a biofilm composed of a complex community including microalgae and cyanobacteria. Although electroactive species were detected in the cathode biofilm, no biocatalysis of oxygen reduction was observed. Enhancement of the current output was mostly due to the production of pure oxygen near the cathode surface by the photosynthetic biofilm. Photosynthesis could produce dissolved oxygen levels approximately four times higher than oxygen levels obtained by aeration. The MFC was able to generate a maximum power density of 11 mW/m2 (projected anode area) over six months without feeding. © 2014 The Royal Society of New Zealand