Molecular mechanisms for photosynthetic carbon partitioning into storage neutral lipids in Nannochloropsis oceanica under nitrogen-depletion conditions

Polysaccharides are a major carbon/energy-reservoir in microalgae, yet their relationship with another form of carbon/energy storage, triacylglycerol (TAG), is poorly understood. Here employing oleaginous microalga Nannochloropsis oceanica as a model, we probed the crosstalk between carbohydrate metabolism and TAG accumulation by tracking the temporal dynamics of lipidomes, monosaccharides and polysaccharides and transcripts of selected genes over 14 days under nitrogen-depleted (N-) and nitrogen-replete (N+) conditions. Glucose, galactose and mannitol were the main monosaccharides in IMET1, and laminarin may be the storage polysaccharide that competes for carbon precursors with TAG. Transcriptional expression analysis revealed that the beta-1,3-glucan degradation and pyruvate dehydrogenases pathways were the main regulatory components involved in driving carbon flow to TAG synthesis. Furthermore, temporal changes of lipidomes and transcripts of glycerolipid metabolism genes were indicative of possible conversion of membrane lipids to TAG, especially under an early stage of nitrogen deprivation conditions. A carbon partitioning model for Nannochloropsis oceanica was proposed, in which beta-1,3-glucan metabolism, acetyl-CoA synthesis and membrane lipid turnover/degradation, in addition to de novo fatty acid synthesis, all contributed to TAG synthesis. (C) 2014 Elsevier B.V. All rights reserved

Carotenoid Distribution in Living Cells of Haematococcus pluvialis (Chlorophyceae)

Haematococcus pluvialis is a freshwater unicellular green microalga belonging to the class Chlorophyceae and is of commercial interest for its ability to accumulate massive amounts of the red ketocarotenoid astaxanthin (3,3′-dihydroxy-β,β-carotene-4,4′-dione). Using confocal Raman microscopy and multivariate analysis, we demonstrate the ability to spectrally resolve resonance–enhanced Raman signatures associated with astaxanthin and β-carotene along with chlorophyll fluorescence. By mathematically isolating these spectral signatures, in turn, it is possible to locate these species independent of each other in living cells of H. pluvialis in various stages of the life cycle. Chlorophyll emission was found only in the chloroplast whereas astaxanthin was identified within globular and punctate regions of the cytoplasmic space. Moreover, we found evidence for β-carotene to be co-located with both the chloroplast and astaxanthin in the cytosol. These observations imply that β-carotene is a precursor for astaxanthin and the synthesis of astaxanthin occurs outside the chloroplast. Our work demonstrates the broad utility of confocal Raman microscopy to resolve spectral signatures of highly similar chromophores in living cells

A Method to Track Targets in Three-Dimensional Space Using an Imaging Sonar

This paper introduces a methodology applying an imaging sonar for three-dimensional (3D) target tracking underwater. The key process in this work involves obtaining the target’s position in space using two images of the same scene, acquired by an adaptive resolution imaging sonar (ARIS) at different positions. A data association algorithm was designed to connect the same target in image sequences. The goal of this work was to track multiple targets in 3D space. The ARIS provides sequences of bi-dimensional images from the backscattered energy according to the range and azimuth. The challenge involved determining the missing elevation information for the observed object within the sonar detection range. By computing the geometrical transformation between the acquisition planar images and the cubical space, using only the sonar information that included the posture and moving speed of the ARIS, the target’s elevation information was obtained. To evaluate the performance of the proposed method, an indoor experiment was conducted using the ARIS. On the basis of the experimental results, we confirmed that the proposed method effectively obtained the target’s position in 3D space. A moving target simulation was also conducted, and the results showed that this method was effective for moving targets. Finally, a field experiment was performed to obtain the vertical distribution and track the 3D trajectories of fish

Assessment of Eicosapentaenoic Acid (EPA) Production from Filamentous Microalga Tribonema aequale: From Laboratory to Pilot-Scale Study

It has long been explored to use EPA-rich unicellular microalgae as a fish oil alternative for production of the high-value omega-3 fatty acid eicosapentaenoic acid (EPA, 20:5, n-3). However, none of the efforts have ever reached commercial success. This study reported a filamentous yellow-green microalga Tribonema aequale that possesses the ability to grow rapidly and synthesize significant amounts of EPA. A series of studies were conducted in a glass column photobioreactor under laboratory culture conditions and in pilot-scale open raceway ponds outdoors. The emphasis was placed on the specific nutrient requirements and the key operational parameters in raceway ponds such as culture depth and mixing regimes. When optimized, T. aequale cells contained 2.9% of EPA (w/w) and reached a very high biomass concentration of 9.8 g L−1 in the glass column photobioreactor. The cellular EPA content was increased further to 3.5% and the areal biomass and EPA productivities of 16.2 g m−2 d−1 and 542.5 mg m−2 d−1, respectively, were obtained from the outdoor pilot-scale open raceway ponds, which were the record high figures reported thus far from microalgae-based EPA production. It was also observed that T. aequale was highly resistant to microbial contamination and easy for harvesting and dewatering, which provide two additional competitive advantages of this filamentous microalga over the unicellular counterparts for potential commercial production of EPA and other derived co-products

Regulation of Light Spectra on Cell Division of the Unicellular Green Alga Haematococcus pluvialis: Insights from Physiological and Lipidomic Analysis

Commercial scale production of natural astaxanthin is currently conducted through cultivation of the green alga Haematococcus pluvialis. This study comprehensively investigated the impact of seven different light spectra on the growth, morphology and photosynthesis of H. pluvialis vegetative cells. Further, the lipidomes of vegetative H. pluvialis grown under various light spectra were qualitatively and quantitatively analyzed using liquid chromatography/mass spectrometry (LC/MS). The results showed the existence of blue light—alone or with red light—promoted cell division, while pure red light or white light enabled increased cell sizes, cellular pigment, starch and lipid contents, and biomass production. Although the photosynthetic performance of H. pluvialis measured as chlorophyll a fluorescence was not significantly affected by light spectra, the lipid profiles, particularly chloroplast membrane lipids, showed remarkable changes with light spectra. The contents of most lipid species in the blue/red light 1/2 group, which showed the fastest cell division, remained at a moderate level compared with those under other light spectra, indicating the fastest dividing cells were featured by a fine-tuned lipid profile. From biotechnical perspective, this comprehensive study can provide insights into the development of appropriate light regimes to promote the cell density or biomass of H. pluvialis mass culture

Two filamentous microalgae as feed ingredients improved flesh quality and enhanced antioxidant capacity and immunity of the gibel carp (Carassius auratus gibelio)

A 40-day experiment was conducted to investigate the effects of two filamentous microalgae as feed ingredients on growth performance, tissue fatty acid profiles, pigmentation and immunity of gibel carp (204.83 +/- 2.17 g, mean +/- SD). Three diets (control, Oedocladium sp. and Tribonema sp.) were formulated. In the control diet, no microalgal meal was added and into the Oedocladium and Tribonema diets were added 40 g/kg Oedocladium sp. meal and 50 g/kg Tribonema sp. meal, respectively. Compared to the control, the addition of Oedocladium sp. and Tribonema sp. had significant effects on tissue fatty acid profiles, antioxidant capacity and immunity without compromising growth, body composition and pigmentation. The addition of Tribonema sp. and Oedocladium sp. significantly increased the EPA, DHA profiles and the ratio of n-3/n-6 polyunsaturated fatty acids, and simultaneously decreased the n-6 polyunsaturated fatty acids profile in fish muscles. Moreover, addition of Tribonema sp. to the diet significantly increased muscle palmitoleic profile and EPA + DHA contents. Furthermore, the addition of either microalga significantly increased the total superoxide dismutase activity level and the complement 3 and immunoglobulin M contents in the plasma. These results demonstrate that Oedocladium sp. and Tribonema sp. can be used as feed ingredients to improve flesh quality and increase the immunity of fish.</p

Effect of initial biomass density on growth and astaxanthin production of Haematococcus pluvialis in an outdoor photobioreactor

Initial biomass density (IBD) is an important factor that affects the viability and productivity of microalgae particularly when sunlight is used for photosynthesis. In this paper, the effect of IBD on photosynthesis, growth, and astaxanthin production of the green microalga Haematococcus pluvialis during the astaxanthin induction stage was studied in a glass column photobioreactor during different seasons. Of seven IBDs, i.e., 0.1, 0.5, 0.8, 1.5, 2.7, 3.5, and 5.0 g L-1 tested, 0.8 g L-1 IBD was optimal and resulted in the highest astaxanthin productivity of 17.1 mg L-1 day(-1). Severe photoinhibition of photosynthesis occurred at low IBD (e.g., 0.1 g L-1) cultures, especially in the winter, and severe light limitation to individual cells in high IBD cultures (> 2.7 g L-1) were responsible for reduced astaxanthin production. This was the first report quantitatively assessing IBD as the key limiting factor for astaxanthin production in H. pluvialis outdoor cultivation

A Lipid Bodies-Associated Galactosyl Hydrolase Is Involved in Triacylglycerol Biosynthesis and Galactolipid Turnover in the Unicellular Green Alga Chlamydomonas reinhardtii

Monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) are the main constituent lipids of thylakoid and chloroplast envelop membranes. Many microalgae can accumulate large amounts of triacylglycerols (TAGs) under adverse environmental conditions, which is accompanied by degradation of the photosynthetic membrane lipids. However, the process mediating the conversion from galactolipids to TAG remains largely unknown. In this study, we performed genetic and biochemical analyses of galactosyl hydrolases (CrGH) identified in the proteome of lipid bodies of the green microalga Chlamydomonas reinhardtii. The recombinant CrGH was confirmed to possess galactosyl hydrolase activity by using o-nitrophenyl-β-D-galactoside as the substrate, and the Michaelis constant (Km) and Kcat of CrGH were 13.98 μM and 3.62 s−1, respectively. Comparative lipidomic analyses showed that the content of MGDG and DGDG increased by 14.42% and 24.88%, respectively, in the CrGH-deficient mutant as compared with that of the wild type cc4533 grown under high light stress conditions, and meanwhile, the TAG content decreased by 32.20%. Up-regulation of CrGH at both a gene expression and protein level was observed under high light stress (HL) conditions. In addition, CrGH was detected in multiple subcellular localizations, including the chloroplast envelope, mitochondria, and endoplasmic reticulum membranes. This study uncovered a new paradigm mediated by the multi-localized CrGH for the conversion of the photosynthetic membranes to TAGs

High-throughput fluorescence-activated cell sorting for cell wall-deficient microalgal mutants screening

Bioavailability and extraction efficiency of the natural antioxidant astaxanthin from the unicellular green alga Haematococcus pluvialis have been limited by the rigid algal cell walls. A high-throughput fluorescence-activated cell sorting (FACS) pipeline was developed in this study with the aim to screen cell wall-deficient mutants. The fluorescein-conjugated lectin-Ricinus communis Agglutinin I (RCA(120)) was used as a marker for binding the sugar moieties of the algal cell walls, which enabled discriminating the algal strains that differed in the cell walls. By utilizing the established FACS pipeline, a cell wall-deficient mutant (i.e. mutant-264) with the significantly reduced RCA(120)-fluorescein intensity was screened from the mutant library. The total content of the cell wall-associated sugars was reduced by 43.9% in the mutant-264 as compared to that of the wild type, with significant decrease in mannose, mannitol and galactose. The dry weight, astaxanthin content and extraction efficiency were improved by 21.1%, 13.1% and 16.5%, respectively, in mutant-264 as compared to that of the wild type. This study not only demonstrated the feasibility of utilization of the FACS-based high-throughput pipeline for cell wall mutant screening, but also delivered a novel promising H. pluvialis strain with multiple improved traits for industrial applications