Metabolites from Phototrophic Prokaryotes and Algae Volume 2

A special issue of Metabolites (ISSN 2218-1989). This special issue belongs to the section "Plant Metabolism".

Deadline for manuscript submissions: closed (30 June 2019) | Viewed by 42883

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Special Issue Editors


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Guest Editor
Department of Biosciences, College of Science, Swansea University, Swansea, UK
Interests: microalgae; cyanobacteria; marine microbial ecology; algal natural products; ‘omics’
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Guest Editor
Department of Biosciences, College of Science, Swansea University, Swansea SA2 8PP, UK
Interests: metabolomics and lipidomics; co-culture techniques; microbial consortia; microalgal biotechnology (biorefinery); nutrient remediation and resource recovery

Special Issue Information

Dear Colleagues,

Algae (here including phototrophic prokaryotes) are a polyphyletic collection of aquatic organisms having an enormous diversity in terms of form and function. Ubiquitous in fresh and marine environments, their contribution to global primary production approximates that of terrestrial organisms and their role in regulating carbon and nitrogen cycles is essential to maintaining life on our planet.

In addition to the important ecological role that algae play in global carbon and nitrogen cycles, these organisms are increasingly emerging as being important in biotechnology. Their ability to fix carbon through photosynthesis, their high productivities compared to plants and the production of some unique groups of metabolites makes them an attractive proposition to fulfilling the drive towards a sustainable and low carbon bio-based circular economy. 

However, our understanding of metabolism and metabolite production in algae currently lags behind that in plants. From an ecological perspective better understanding of metabolic pathways and metabolite production (both intracellular and extracellular) will lead to improved understanding on the role of algae in cycling elements within aquatic environments and to improved assessments of aquatic primary production. Additionally, from a biotechnological perspective, a better understanding will lead to improved yield and the ability to manipulate algal growth for bio-production purposes. There is also potential to apply our understanding on manipulation of metabolite pathways and production to plant and other non-plant systems.

From both an ecological and biotechnological perspective, we need improved understanding on acclimation and adaptation strategies to both abiotic (e.g., nutrients, light, temperature and salinity) and biotic factors (e.g., microbial consortia interactions, predator-prey interactions and bacterial/viral infection). In particular, we need improved understanding on shifts in allocation between primary and secondary metabolism and metabolites and on carbon and nitrogen allocation.  

For this Special Issue we welcome research papers and reviews based around metabolomics to improve knowledge on the metabolome and metabolism in algae with a focus on carbon and nitrogen resource allocation. We welcome papers from an ecological or from a biotechnological and/or waste water remediation perspective through from single cell analysis to complex microbial consortia systems.  Analytical approach focused papers, including extraction, technical measurements and bioinformatics, are also welcome.  

Dr. Carole Llewellyn
Dr. Rahul Vijay Kapoore
Guest Editors

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Keywords

  • Stress adaptation
  • Nutrient metabolism
  • Biotechnology
  • Ecology
  • Physiology
  • Metabolomics

Published Papers (10 papers)

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Research

13 pages, 2427 KiB  
Article
Far-Red Light Acclimation for Improved Mass Cultivation of Cyanobacteria
by Alla Silkina, Bethan Kultschar and Carole A. Llewellyn
Metabolites 2019, 9(8), 170; https://doi.org/10.3390/metabo9080170 - 19 Aug 2019
Cited by 10 | Viewed by 4038
Abstract
Improving mass cultivation of cyanobacteria is a goal for industrial biotechnology. In this study, the mass cultivation of the thermophilic cyanobacterium Chlorogloeopsis fritschii was assessed for biomass production under light-emitting diode white light (LEDWL), far-red light (FRL), and combined white light and far-red [...] Read more.
Improving mass cultivation of cyanobacteria is a goal for industrial biotechnology. In this study, the mass cultivation of the thermophilic cyanobacterium Chlorogloeopsis fritschii was assessed for biomass production under light-emitting diode white light (LEDWL), far-red light (FRL), and combined white light and far-red light (WLFRL) adaptation. The induction of chl f was confirmed at 24 h after the transfer of culture from LEDWL to FRL. Using combined light (WLFRL), chl f, a, and d, maintained the same level of concentration in comparison to FRL conditions. However, phycocyanin and xanthophylls (echinone, caloxanthin, myxoxanthin, nostoxanthin) concentration increased 2.7–4.7 times compared to LEDWL conditions. The productivity of culture was double under WLFRL compared with LEDWL conditions. No significant changes in lipid, protein, and carbohydrate concentrations were found in the two different light conditions. The results are important for informing on optimum biomass cultivation of this species for biomass production and bioactive product development. Full article
(This article belongs to the Special Issue Metabolites from Phototrophic Prokaryotes and Algae Volume 2)
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24 pages, 1601 KiB  
Article
Euglena Central Metabolic Pathways and Their Subcellular Locations
by Sahutchai Inwongwan, Nicholas J. Kruger, R. George Ratcliffe and Ellis C. O’Neill
Metabolites 2019, 9(6), 115; https://doi.org/10.3390/metabo9060115 - 14 Jun 2019
Cited by 15 | Viewed by 5595
Abstract
Euglenids are a group of algae of great interest for biotechnology, with a large and complex metabolic capability. To study the metabolic network, it is necessary to know where the component enzymes are in the cell, but despite a long history of research [...] Read more.
Euglenids are a group of algae of great interest for biotechnology, with a large and complex metabolic capability. To study the metabolic network, it is necessary to know where the component enzymes are in the cell, but despite a long history of research into Euglena, the subcellular locations of many major pathways are only poorly defined. Euglena is phylogenetically distant from other commonly studied algae, they have secondary plastids bounded by three membranes, and they can survive after destruction of their plastids. These unusual features make it difficult to assume that the subcellular organization of the metabolic network will be equivalent to that of other photosynthetic organisms. We analysed bioinformatic, biochemical, and proteomic information from a variety of sources to assess the subcellular location of the enzymes of the central metabolic pathways, and we use these assignments to propose a model of the metabolic network of Euglena. Other than photosynthesis, all major pathways present in the chloroplast are also present elsewhere in the cell. Our model demonstrates how Euglena can synthesise all the metabolites required for growth from simple carbon inputs, and can survive in the absence of chloroplasts. Full article
(This article belongs to the Special Issue Metabolites from Phototrophic Prokaryotes and Algae Volume 2)
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22 pages, 3053 KiB  
Article
Improved Algal Toxicity Test System for Robust Omics-Driven Mode-of-Action Discovery in Chlamydomonas reinhardtii
by Stefan Schade, Emma Butler, Steve Gutsell, Geoff Hodges, John K. Colbourne and Mark R. Viant
Metabolites 2019, 9(5), 94; https://doi.org/10.3390/metabo9050094 - 10 May 2019
Cited by 5 | Viewed by 3886
Abstract
Algae are key components of aquatic food chains. Consequently, they are internationally recognised test species for the environmental safety assessment of chemicals. However, existing algal toxicity test guidelines are not yet optimized to discover molecular modes of action, which require highly-replicated and carefully [...] Read more.
Algae are key components of aquatic food chains. Consequently, they are internationally recognised test species for the environmental safety assessment of chemicals. However, existing algal toxicity test guidelines are not yet optimized to discover molecular modes of action, which require highly-replicated and carefully controlled experiments. Here, we set out to develop a robust, miniaturised and scalable Chlamydomonas reinhardtii toxicity testing approach tailored to meet these demands. We primarily investigated the benefits of synchronised cultures for molecular studies, and of exposure designs that restrict chemical volatilisation yet yield sufficient algal biomass for omics analyses. Flow cytometry and direct-infusion mass spectrometry metabolomics revealed significant and time-resolved changes in sample composition of synchronised cultures. Synchronised cultures in sealed glass vials achieved adequate growth rates at previously unachievably-high inoculation cell densities, with minimal pH drift and negligible chemical loss over 24-h exposures. Algal exposures to a volatile test compound (chlorobenzene) yielded relatively high reproducibility of metabolic phenotypes over experimental repeats. This experimental test system extends existing toxicity testing formats to allow highly-replicated, omics-driven, mode-of-action discovery. Full article
(This article belongs to the Special Issue Metabolites from Phototrophic Prokaryotes and Algae Volume 2)
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17 pages, 3001 KiB  
Article
Effects of Copper and pH on the Growth and Physiology of Desmodesmus sp. AARLG074
by Nattaphorn Buayam, Matthew P. Davey, Alison G. Smith and Chayakorn Pumas
Metabolites 2019, 9(5), 84; https://doi.org/10.3390/metabo9050084 - 30 Apr 2019
Cited by 12 | Viewed by 3876
Abstract
Copper (Cu) is a heavy metal that is widely used in industry and as such wastewater from mining or industrial operations can contain high levels of Cu. Some aquatic algal species can tolerate and bioaccumulate Cu and so could play a key role [...] Read more.
Copper (Cu) is a heavy metal that is widely used in industry and as such wastewater from mining or industrial operations can contain high levels of Cu. Some aquatic algal species can tolerate and bioaccumulate Cu and so could play a key role in bioremediating and recovering Cu from polluted waterways. One such species is the green alga Desmodesmus sp. AARLG074. The aim of this study was to determine how Desmodesmus is able to tolerate large alterations in its external Cu and pH environment. Specifically, we set out to measure the variations in the Cu removal efficiency, growth, ultrastructure, and cellular metabolite content in the algal cells that are associated with Cu exposure and acidity. The results showed that Desmodesmus could remove up to 80% of the copper presented in Jaworski’s medium after 30 min exposure. There was a decrease in the ability of Cu removal at pH 4 compared to pH 6 indicating both pH and Cu concentration affected the efficiency of Cu removal. Furthermore, Cu had an adverse effect on algal growth and caused ultrastructural changes. Metabolite fingerprinting (FT-IR and GC-MS) revealed that the polysaccharide and amino acid content were the main metabolites affected under acid and Cu exposure. Fructose, lactose and sorbose contents significantly decreased under both acidic and Cu conditions, whilst glycerol and melezitose contents significantly increased at pH 4. The pathway analysis showed that pH had the highest impact score on alanine, aspartate and glutamate metabolism whereas Cu had the highest impact on arginine and proline metabolism. Notably both Cu and pH had impact on glutathione and galactose metabolism. Full article
(This article belongs to the Special Issue Metabolites from Phototrophic Prokaryotes and Algae Volume 2)
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15 pages, 3011 KiB  
Article
Intracellular and Extracellular Metabolites from the Cyanobacterium Chlorogloeopsis fritschii, PCC 6912, During 48 Hours of UV-B Exposure
by Bethan Kultschar, Ed Dudley, Steve Wilson and Carole A. Llewellyn
Metabolites 2019, 9(4), 74; https://doi.org/10.3390/metabo9040074 - 16 Apr 2019
Cited by 17 | Viewed by 4685
Abstract
Cyanobacteria have many defence strategies to overcome harmful ultraviolet (UV) stress including the production of secondary metabolites. Metabolomics can be used to investigate this altered metabolism via targeted and untargeted techniques. In this study we assessed the changes in the intra- and extracellular [...] Read more.
Cyanobacteria have many defence strategies to overcome harmful ultraviolet (UV) stress including the production of secondary metabolites. Metabolomics can be used to investigate this altered metabolism via targeted and untargeted techniques. In this study we assessed the changes in the intra- and extracellular low molecular weight metabolite levels of Chlorogloeopsis fritschii (C. fritschii) during 48 h of photosynthetically active radiation (PAR) supplemented with UV-B (15 µmol m−2 s−1 of PAR plus 3 µmol m−2 s−1 of UV-B) and intracellular levels during 48 h of PAR only (15 µmol m−2 s−1) with sampling points at 0, 2, 6, 12, 24 and 48 h. Gas chromatography–mass spectrometry (GC–MS) was used as a metabolite profiling tool to investigate the global changes in metabolite levels. The UV-B time series experiment showed an overall significant reduction in intracellular metabolites involved with carbon and nitrogen metabolism such as the amino acids tyrosine and phenylalanine which have a role in secondary metabolite production. Significant accumulation of proline was observed with a potential role in stress mitigation as seen in other photosynthetic organisms. 12 commonly identified metabolites were measured in both UV-B exposed (PAR + UV-B) and PAR only experiments with differences in significance observed. Extracellular metabolites (PAR + UV-B) showed accumulation of sugars as seen in other cyanobacterial species as a stress response to UV-B. In conclusion, a snapshot of the metabolome of C. fritschii was measured. Little work has been undertaken on C. fritschii, a novel candidate for use in industrial biotechnology, with, to our knowledge, no previous literature on combined intra- and extracellular analysis during a UV-B treatment time-series. This study is important to build on experimental data already available for cyanobacteria and other photosynthetic organisms exposed to UV-B. Full article
(This article belongs to the Special Issue Metabolites from Phototrophic Prokaryotes and Algae Volume 2)
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11 pages, 1585 KiB  
Communication
The Bacterial Phytoene Desaturase-Encoding Gene (CRTI) is an Efficient Selectable Marker for the Genetic Transformation of Eukaryotic Microalgae
by Ana Molina-Márquez, Marta Vila, Javier Vigara, Ana Borrero and Rosa León
Metabolites 2019, 9(3), 49; https://doi.org/10.3390/metabo9030049 - 12 Mar 2019
Cited by 10 | Viewed by 4013
Abstract
Genetic manipulation shows great promise to further boost the productivity of microalgae-based compounds. However, selection of microalgal transformants depends mainly on the use of antibiotics, which have raised concerns about their potential impacts on human health and the environment. We propose the use [...] Read more.
Genetic manipulation shows great promise to further boost the productivity of microalgae-based compounds. However, selection of microalgal transformants depends mainly on the use of antibiotics, which have raised concerns about their potential impacts on human health and the environment. We propose the use of a synthetic phytoene desaturase-encoding gene (CRTIop) as a selectable marker and the bleaching herbicide norflurazon as a selective agent for the genetic transformation of microalgae. Bacterial phytoene desaturase (CRTI), which, unlike plant and algae phytoene desaturase (PDS), is not sensitive to norflurazon, catalyzes the conversion of the colorless carotenoid phytoene into lycopene. Although the expression of CRTI has been described to increase the carotenoid content in plant cells, its use as a selectable marker has never been testedin algae or in plants. In this study, a version of the CRTI gene adapted to the codon usage of Chlamydomonas has been synthesized, and its suitability to be used as selectable marker has been shown. The microalgae were transformed by the glass bead agitation method and selected in the presence of norflurazon. Average transformation efficiencies of 550 colonies µg−1 DNA were obtained. All the transformants tested had incorporated the CRTIop gene in their genomes and were able to synthesize colored carotenoids. Full article
(This article belongs to the Special Issue Metabolites from Phototrophic Prokaryotes and Algae Volume 2)
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19 pages, 1378 KiB  
Article
Modulation of Polar Lipid Profiles in Chlorella sp. in Response to Nutrient Limitation
by Daniel A. White, Paul A. Rooks, Susan Kimmance, Karen Tait, Mark Jones, Glen A. Tarran, Charlotte Cook and Carole A. Llewellyn
Metabolites 2019, 9(3), 39; https://doi.org/10.3390/metabo9030039 - 28 Feb 2019
Cited by 17 | Viewed by 4125
Abstract
We evaluate the effects of nutrient limitation on cellular composition of polar lipid classes/species in Chlorella sp. using modern polar lipidomic profiling methods (liquid chromatography–tandem mass spectrometry; LC-MS/MS). Total polar lipid concentration was highest in nutrient-replete (HN) cultures with a significant reduction in [...] Read more.
We evaluate the effects of nutrient limitation on cellular composition of polar lipid classes/species in Chlorella sp. using modern polar lipidomic profiling methods (liquid chromatography–tandem mass spectrometry; LC-MS/MS). Total polar lipid concentration was highest in nutrient-replete (HN) cultures with a significant reduction in monogalactosyldiacylglycerol (MGDG), phosphatidylglycerol (PG), phosphatidylcholine (PC), and phosphatidylethanolamine (PE) class concentrations for nutrient-deplete (LN) cultures. Moreover, reductions in the abundance of MGDG relative to total polar lipids versus an increase in the relative abundance of digalactosyldiacylglycerol (DGDG) were recorded in LN cultures. In HN cultures, polar lipid species composition remained relatively constant throughout culture with high degrees of unsaturation associated with acyl moieties. Conversely, in LN cultures lipid species composition shifted towards greater saturation of acyl moieties. Multivariate analyses revealed that changes in the abundance of a number of species contributed to the dissimilarity between LN and HN cultures but with dominant effects from certain species, e.g., reduction in MGDG 34:7 (18:3/16:4). Results demonstrate that Chlorella sp. significantly alters its polar lipidome in response to nutrient limitation, and this is discussed in terms of physiological significance and polar lipids production for applied microalgal production systems. Full article
(This article belongs to the Special Issue Metabolites from Phototrophic Prokaryotes and Algae Volume 2)
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17 pages, 4454 KiB  
Article
An Improved Genome-Scale Metabolic Model of Arthrospira platensis C1 (iAK888) and Its Application in Glycogen Overproduction
by Amornpan Klanchui, Sudarat Dulsawat, Kullapat Chaloemngam, Supapon Cheevadhanarak, Peerada Prommeenate and Asawin Meechai
Metabolites 2018, 8(4), 84; https://doi.org/10.3390/metabo8040084 - 26 Nov 2018
Cited by 14 | Viewed by 3685
Abstract
Glycogen-enriched biomass of Arthrospira platensis has increasingly gained attention as a source for bioethanol production. To study the metabolic capabilities of glycogen production in A. platensis C1, a genome-scale metabolic model (GEM) could be a useful tool for predicting cellular behavior and suggesting [...] Read more.
Glycogen-enriched biomass of Arthrospira platensis has increasingly gained attention as a source for bioethanol production. To study the metabolic capabilities of glycogen production in A. platensis C1, a genome-scale metabolic model (GEM) could be a useful tool for predicting cellular behavior and suggesting strategies for glycogen overproduction. New experimentally validated GEM of A. platensis C1 namely iAK888, which has improved metabolic coverage and functionality was employed in this research. The iAK888 is a fully functional compartmentalized GEM consisting of 888 genes, 1,096 reactions, and 994 metabolites. This model was demonstrated to reasonably predict growth and glycogen fluxes under different growth conditions. In addition, iAK888 was further employed to predict the effect of deficiencies of NO3, PO43−, or SO42− on the growth and glycogen production in A. platensis C1. The simulation results showed that these nutrient limitations led to a decrease in growth flux and an increase in glycogen flux. The experiment of A. platensis C1 confirmed the enhancement of glycogen fluxes after the cells being transferred from normal Zarrouk’s medium to either NO3, PO43−, or SO42−-free Zarrouk’s media. Therefore, iAK888 could be served as a predictive model for glycogen overproduction and a valuable multidisciplinary tool for further studies of this important academic and industrial organism. Full article
(This article belongs to the Special Issue Metabolites from Phototrophic Prokaryotes and Algae Volume 2)
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16 pages, 1296 KiB  
Article
Quenching for Microalgal Metabolomics: A Case Study on the Unicellular Eukaryotic Green Alga Chlamydomonas reinhardtii
by Rahul Vijay Kapoore and Seetharaman Vaidyanathan
Metabolites 2018, 8(4), 72; https://doi.org/10.3390/metabo8040072 - 31 Oct 2018
Cited by 4 | Viewed by 3874
Abstract
Capturing a valid snapshot of the metabolome requires rapid quenching of enzyme activities. This is a crucial step in order to halt the constant flux of metabolism and high turnover rate of metabolites. Quenching with cold aqueous methanol is treated as a gold [...] Read more.
Capturing a valid snapshot of the metabolome requires rapid quenching of enzyme activities. This is a crucial step in order to halt the constant flux of metabolism and high turnover rate of metabolites. Quenching with cold aqueous methanol is treated as a gold standard so far, however, reliability of metabolomics data obtained is in question due to potential problems connected to leakage of intracellular metabolites. Therefore, we investigated the influence of various parameters such as quenching solvents, methanol concentration, inclusion of buffer additives, quenching time and solvent to sample ratio on intracellular metabolite leakage from Chlamydomonas reinhardtii. We measured the recovery of twelve metabolite classes using gas chromatography mass spectrometry (GC-MS) in all possible fractions and established mass balance to trace the fate of metabolites during quenching treatments. Our data demonstrate significant loss of intracellular metabolites with the use of the conventional 60% methanol, and that an increase in methanol concentration or quenching time also resulted in higher leakage. Inclusion of various buffer additives showed 70 mM HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) to be suitable. In summary, we recommend quenching with 60% aqueous methanol supplemented with 70 mM HEPES (−40 °C) at 1:1 sample to quenching solvent ratio, as it resulted in higher recoveries for intracellular metabolites with subsequent reduction in the metabolite leakage for all metabolite classes. Full article
(This article belongs to the Special Issue Metabolites from Phototrophic Prokaryotes and Algae Volume 2)
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13 pages, 1524 KiB  
Article
The Effect of High-Intensity Ultraviolet Light to Elicit Microalgal Cell Lysis and Enhance Lipid Extraction
by Thomas Sydney, Jo-Ann Marshall-Thompson, Rahul Vijay Kapoore, Seetharaman Vaidyanathan, Jagroop Pandhal and J. Patrick A. Fairclough
Metabolites 2018, 8(4), 65; https://doi.org/10.3390/metabo8040065 - 15 Oct 2018
Cited by 21 | Viewed by 4294
Abstract
Currently, the energy required to produce biofuel from algae is 1.38 times the energy available from the fuel. Current methods do not deliver scalable, commercially viable cell wall disruption, which creates a bottleneck on downstream processing. This is primarily due to the methods [...] Read more.
Currently, the energy required to produce biofuel from algae is 1.38 times the energy available from the fuel. Current methods do not deliver scalable, commercially viable cell wall disruption, which creates a bottleneck on downstream processing. This is primarily due to the methods depositing energy within the water as opposed to within the algae. This study investigates ultraviolet B (UVB) as a disruption method for the green algae Chlamydomonas reinhardtii, Dunaliella salina and Micractinium inermum to enhance solvent lipid extraction. After 232 seconds of UVB exposure at 1.5 W/cm2, cultures of C. reinhardtii (culture density 0.7 mg/mL) showed 90% disruption, measured using cell counting, correlating to an energy consumption of 5.6 MJ/L algae. Small-scale laboratory tests on C. reinhardtii showed bead beating achieving 45.3 mg/L fatty acid methyl esters (FAME) and UV irradiation achieving 79.9 mg/L (lipids solvent extracted and converted to FAME for measurement). The alga M. inermum required a larger dosage of UVB due to its thicker cell wall, achieving a FAME yield of 226 mg/L, compared with 208 mg/L for bead beating. This indicates that UV disruption had a higher efficiency when used for solvent lipid extraction. This study serves as a proof of concept for UV irradiation as a method for algal cell disruption. Full article
(This article belongs to the Special Issue Metabolites from Phototrophic Prokaryotes and Algae Volume 2)
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