Larisa Semenova

Accumulation of Astaxanthin by a New Haematococcus pluvialis Strain BM1 from the White Sea Coastal Rocks (Russia)

We report on a novel arctic strain BM1 of a carotenogenic chlorophyte from a coastal habitat with harsh environmental conditions (wide variations in solar irradiance, temperature, salinity and nutrient availability) identified as Haematococcus pluvialis Flotow. Increased (25‰) salinity exerted no adverse effect on the growth of the green BM1 cells. Under stressful conditions (high light, nitrogen and phosphorus deprivation), green vegetative cells of H. pluvialis BM1 grown in BG11 medium formed non-motile palmelloid cells and, eventually, hematocysts capable of a massive accumulation of the keto-carotenoid astaxanthin with a high nutraceutical and therapeutic potential. Routinely, astaxanthin was accumulated at the level of 4% of the cell dry weight (DW), reaching, under prolonged stress, 5.5% DW. Astaxanthin was predominantly accumulated in the form of mono- and diesters of fatty acids from C16 and C18 families. The palmelloids and hematocysts were characterized by the formation of red-colored cytoplasmic lipid droplets, increasingly large in size and number. The lipid droplets tended to merge and occupied almost the entire volume of the cell at the advanced stages of stress-induced carotenogenesis. The potential application of the new strain for the production of astaxanthin is discussed in comparison with the H. pluvialis strains currently employed in microalgal biotechnology.

Stress-induced changes in pigment and fatty acid content in the microalga Desmodesmus sp. Isolated from a White Sea hydroid

Effects of light intensity, nitrogen availability, and inoculum density on growth and the content of esterified fatty acids (FA), chlorophylls, and carotenoids in Desmodesmus sp. 3Dp86E-1 chlorophyte alga isolated from the White Sea hydroid Dynamena pumila L. were investigated. The growth of algae in the complete BG-11 medium was not limited by irradiances up to 480 μE/(m2 s) PAR but depended on the inoculum density. Under nitrogen starvation conditions, high-intensity light retarded growth of the microalga; this effect was less pronounced in the cultures initiated at high inoculum densities. The highest FA percentage in biomass (30% at the 3rd day of cultivation) was detected in nitrogen-starving cultures grown under high light conditions; however, the highest volumetric FA content (0.25 g/L) was attained on a complete medium at 480 μE/(m2 s). An increase in the content of oleic acid (18:1) on the background of a decrease in linolenic acid (18:3) was characteristic of the microalga under stress conditions. The microalga was found to be non-carotenogenic. Nitrogen starvation brought about a dramatic decrease in chlorophyll content on the background of relatively constant carotenoid content. On nitrogen-deplete medium, the high light did not trigger the adaptive response of the pigment apparatus. The changes in absorption spectra of Desmodesmus sp. 3Dp86E-1 cell suspensions reflected the increase in relative contribution of carotenoids to light absorption by the microalgal cells; these changes were tightly related with FA accumulation. The mechanisms of acclimation of Desmodesmus sp. 3Dp86E-1 to high light and nitrogen starvation are discussed in view of possible biotechnological applications of this alga.

Physiological plasticity of symbiotic Desmodesmus (Chlorophyceae) isolated from taxonomically distant white sea invertibrates

The physiological heterogeneity of closely related symbiotic microalgae from taxonomically distant animal hosts was studied for the first time. Three strains of unicellular algae from the genus Desmodesmus (Chlorophyceae) isolated from White Sea benthic invertebrates were used as the object in this study. The effects of nitrogen starvation and high light intensity on the growth, changes in chlorophyll and total carotenoid contents and fatty acid content and composition of the microalgal cell lipids were followed. Nitrogen starvation declined the biomass accumulation rate as well as chlorophyll and carotenoid contents on the background of the enhanced fatty acid accumulation in all strains studied. The ultrastructural study revealed the reduction of photosynthetic apparatus and an increase in the proportion of cell volume occupied by oil bodies and starch grains as well as an increase in the cell wall thickness. A decline in the effective per cell irradiance in the cultures of higher cell density, as a rule, slowed down the changes in pigment and fatty acids composition characteristic of nitrogen starvation. The rates of biomass accumulation and cell biochemical composition under the nitrogen-starvation conditions were strain-specific. In most cases, the decline in chlorophylls proceeded at a higher rate in comparison with that of carotenoid decline. In two of the three strains studied, both these process occurred synchronously with the decline in the unsaturation of the cell lipid fatty acids. The possibilities of biotechnological application of the symbiotic microalgae are discussed with the peculiarities of their stress physiology in mind.

Effects of glucose during photoheterotrophic growth of the cyanobacterium Calothrix sp. PCC 7601 capable for chromatic adaptation

Photoheterotrophic growth of a filamentous cyanobacterium Calothrix sp. PCC 7601, which is capable for complementary chromatic adaptation, in the presence of glucose was accompanied by changes in the content of phycobiliproteins. Glucose, a source of energy and a metabolism regulator, differently affected the level of major phycobilisome pigments, phycocyanin (PC) and phycoerythrin (PE) in the cells. When red light enhanced PC synthesis, glucose enhanced it additionally. When green light suppressed PC synthesis, glucose did not affect it. Under both light regimes, glucose inhibited PE synthesis. Thus, glucose oppositely affected the content of two major phycobiliproteins. Glucose not only affected the ratio between phycobiliproteins but also decreased the content of carotenoids, inhibited activity of photosystem II, and affected cell sizes. A stereochemical analog of glucose, 2-deoxy-D-glucose, induced effects similar to those of glucose. A comparison with the effects of red and green light demonstrated that glucose acted on Calothrix similarly to red light and oppositely to green light.

A new subarctic strain of Tetradesmus obliquus. Part I: Identification and fatty acid profiling

We isolated a new subarctic strain Tetradesmus obliquus IPPAS S-2023 (Scenedesmaceae, Chlorophyceae) from rock baths in the White Sea. To verify its taxonomic assignment, internal transcribed spacer 2 (ITS2) of the strain was sequenced and its secondary structure was compared with predicted ITS2 secondary structures of Scenedesmaceae. The analysis of the ITS2 made it possible to assign the new strain IPPAS S-2023 to the species T. obliquus. The ultrastructural studies and energy-dispersive X-ray spectroscopy (EDX) analysis revealed a marked accumulation of vacuolar inclusions enriched in phosphorus and nitrogen (N) as well as cytoplasmiс oil bodies. Most of predicted properties of biodiesel derived from the fatty acids profile of the strain grown in the N-free medium complied with the requirements of European and American standards. The results suggest that the new subarctic strain T. obliquus IPPAS S-2023 is a promising candidate for nutrients biosequestration and for biodiesel production. In a companion paper, we assess its biomass production capability and suitability and demonstrated suitability of IPPAS S-2023 as a reference strain for studies on elevated CO2 stress effects selection of carbon dioxide-tolerant microalgae by comparison with a CO2-tolerant strain IPPAS S-2014.

A new subarctic strain of Tetradesmus obliquus. Part II: comparative studies of CO2-stress tolerance

A huge interest in CO2-tolerant microalgae is fueled by development of CO2-biomitigation methods based on intensive cultivation of microalgae. Still, the mechanisms of CO2-tolerance are scarcely investigated. Previously, we described a symbiotic Desmodesmus sp. IPPAS S-2014 from a White Sea hydroid tolerant to extremely high (20–100%) CO2 levels. In the present work, we compared its ultrastructural and physiological responses to those of a novel free-living White Sea strain of Tetradesmus obliquus IPPAS S-2023 characterized in the companion paper. The strain S-2023 is closely related to Desmodesmus sp. IPPAS S-2014 but lacks its tolerance to extremely high CO2 (it is unable to survive at 100% CO2 and exhibits a reduced-growth phenotype when sparged with 20% CO2: air mixture). We compared the responses of the cell organization and photosynthetic activity to 20% CO2 in the tolerant and the intolerant White Sea chlorophytes using chlorophyll fluorescence measurements and ultrastructural analysis (transmission electron microscopy). The features peculiar to the CO2-intolerant chlorophyte include (i) inability to maintain pH homeostasis, (ii) a steady decline in the photosynthetic activity of the cells, (iii) a reduction of the photosynthetic membranes, and (iv) delayed accumulation of starch (starch grains) and its subsequent conversion to reserve lipids (oil bodies). Nitrogen starvation enhances the effects of high-CO2 stress in the CO2-intolerant microalga. The results of this work are discussed in the context of selection of tolerant algal strains for CO2 biomitigation applications.

Stress-induced changes in the ultrastructure of the photosynthetic apparatus of green microalgae

In photosynthetic organisms including unicellular algae, acclimation to and damage by environmental stresses are readily apparent at the level of the photosynthetic apparatus. Phenotypic manifestations of the stress responses include rapid and dramatic reduction of photosynthetic activity and pigment content aimed at mitigating the risk of photooxidative damage. Although the physiological and molecular mechanisms of these events are well known, the ultrastructural picture of the stress responses is often elusive and frequently controversial. We analyzed an extensive set of transmission electron microscopy images of the microalgal cells obtained across species of Chlorophyta and in a wide range of growth conditions. The results of the analysis allowed us to pinpoint distinct ultrastructural changes typical of normal functioning and emergency reduction of the chloroplast membrane system under high light exposure and/or mineral nutrient starvation. We demonstrate the patterns of the stress-related ultrastructural changes including peculiar thylakoid rearrangements and autophagy-like processes and provide an outlook on their significance for implementation of the stress responses.

ASSESSMENT OF A NEW CHLORELLA VULGARIS (CHLOROPHYTA) IPPAS C-2015 STRAIN FOR APPLICATION IN POULTRY WASTEWATER BIOREMEDIATION

The potential of the use of a new microalga strain Chlorella vulgaris IPPAS C-2015 (Chlorophyta, Trebouxiophyceae) for poultry wastewater treatment has been studied. The artificial wastewater (AWW) from chicken litter that mimicked real poultry wastewater was prepared, and the efficiency of the bioremoval of the inorganic anion from it and destruction of the organic contaminant in it during the new strain of C. vulgaris semi-continuous cultivation was assessed. After three days of C. vulgaris culturing, the initial nitrate and orthophosphate levels in AWW decreased by more than 90% and more than 48%, respectively, and organic compounds were degraded by 80% on average (judging by chemical oxygen consumption). During the cultivation of the microalgae in AWW, the bacteria associated with the C. vulgaris pre-culture gradually replaced the bacteria characteristic of the AWW. The microalga biomass grown in AWW contained a great amount of polyunsaturated long-chain fatty acids from the С 18 family. The capacities of the new C. vulgaris strain of being used in the combined poultry wastewater treatment and utilization of the resulting biomass are discussed together with the potential advantages of the microalgae-based over conventional biological wastewater treatment technologies.