Responses of Heterotrophic Cultures of Chlorella vulgaris Beyerinck to Darkness and Light. II. Action Spectrum for and Mechanism of the Light Requirement for Heterotrophic Growth

Chlorella vulgaris Beyerinck (Emerson's strain), fails to grow in the dark even when sugars are provided. This phenomenon was clearly demonstrated in the alga, C. vulgaris, for which the growth rate in darkness on a glucose medium remained constant for 2 days and then declined to approach zero. Pigment concentrations also declined in darkness. Changes in flow rate of 1% CO₂-in-air from zero to 7 ml per minute caused a progressive increase in the dark growth rate over a 5-day period, but did not maintain growth in the dark. Rates above 7 ml per minute produced no changes in growth rates.

White light intensities below the compensation point of the alga maintained heterotrophic growth. The saturation value for this response was 0.8 μw/cm². White light also initiated growth in nongrowing cultures transferred from darkness to light.

The action spectrum for heterotrophic growth indicated a porphyrin as the active pigment. Light in the 425 mμ region was 4 times as effective as white light in stimulating heterotrophic growth. A secondary peak of growth stimulation occurred in the 575 mμ region.

The respiration of glucose by the alga was stimulated by low intensities of white light. This response was not immediate, but was clearly present after the third day of incubation.

Malonate and cyanide were inhibitory to growth of C. vulgaris on inorganic medium or glucose medium under 300 ft-c of white light. These data suggested that succinic dehydrogenase and cytochrome oxidase systems were present.

Substances inhibitory to growth were excreted into the medium under dark-growth conditions, and 2 of these substances were indentified as formic and acetic acids.

The evidence suggested that respiration of glucose cannot proceed for an extended period of time in darkness. The reason for this is postulated to be the lack of a cytochrome or a cytochrome precursor.

     

Responses of Two Coastal Algae (Skeletonema costatum and Chlorella vulgaris) to Changes in Light and Iron Levels1

Iron (Fe) is essential for phytoplankton growth and photosynthesis, and is proposed to be an important factor regulating algal blooms under replete major nutrients in coastal environments. Here, Skeletonema costatum, a typical red-tide diatom species, and Chlorella vulgaris, a widely distributed Chlorella, were chosen to examine carbon fixation and Fe uptake by coastal algae under dark and light conditions with different Fe levels. The cellular carbon fixation and intracellular Fe uptake were measured via ¹⁴C and ⁵⁵Fe tracer assay, respectively. Cell growth, cell size, and chlorophyll-α concentration were measured to investigate the algal physiological variation in different treatments. Our results showed that cellular Fe uptake proceeds under dark and the uptake rates were comparable to or even higher than those in the light for both algal species. Fe requirements per unit carbon fixation were also higher in the dark resulting in higher Fe: C ratios. During the experimental period, high Fe addition significantly enhanced cellular carbon fixation and Fe uptake. Compared to C. vulgaris, S. costatum was the common dominant bloom species because of its lower Fe demand but higher Fe uptake rate. This study provides some of the first measurements of Fe quotas in coastal phytoplankton cells, and implies that light and Fe concentrations may influence the phytoplankton community succession when blooms occur in coastal ecosystems.     

Morphometric and Stereologic Analysis of Chlorella vulgaris Under Heterotrophic Growth Conditions

A computer-supported determination of stereological parameterswas used to study the possible ultrastructural changes of Chlorellavulgaris UAM 101 under photolithotrophic, mixotrophic and photoheterotrophicconditions of growth. Data recording was carried out througha semi-automatic digitizing image analysis system instead ofthe current method of superimposition of an array of short lines. Glucose promoted drastic physiological changes [Martínezand Orús, 1991. Plant Physiology (in press)], which stronglyaffected the size of the cells and volume densities of storagematerials. However, volumetric ratios of the mitochondrion orchloroplast active fraction were not affected by the presenceof glucose, probably indicating that these ratios are characteristicof each species. Cell wall ultrastructure was also analysed and the presenceof sporopollenin demonstrated, in contrast to the thin and sporopollenin-lackingcell wall generally described for Chlorella vulgaris species. Chlorella vulgaris, photoautotrophic, mixotrophic, photoheterotrophic, image analysis, stereology     

The Accumulation of Carbohydrate in Chlorella vulgaris under Heterotrophic Conditions

The accumulation of carbohydrate in two strains of Chlorellavulgaris under autotrophic and heterotrophic conditions hasbeen examined. Cells of the Emerson strain, which cannot dividein the dark on a glucose medium, accumulate much more carbohydrateunder heterotrophic conditions than they do when growing autotrophically.Much of the extra carbohydrate is in the form of polysaccharide,particularly starch. The extent of this accumulation is notrelated to the size of the cells. In the Brannon no. 1strain,where there is no light requirement for cell division, thereis also a marked accumulation of carbohydrate, chiefly of somepolysaccharide other than starch. The relation between the carbohydrateaccumulation and the contrasting behaviour of the two strainsunder fully heterotrophic conditions is discussed.     

Alkalinity-induced aggregation in Chlorella vulgaris I. Changes in cell volume and cell-wall structure

In Chlorella vulgaris cell aggregation, the clustering of singlecells into groups is induced by an alkaline pH (9.5). The processof alkalinity-induced aggregation may be divided into two stages:the first stage (0–24 hr after exposure to the alkalinepH) is characterized by enhanced precipitation of cells fromthe medium, as well as by a seven fold increase in cell volume.The second stage (24–120 hr) is associated with a furtherincrease in the extent of cell precipitation in the culture,which seems to result from the aggregation of clusters of enlargedcells. Electron micrographs reveal the existence, at this phase,of a number of autospores in the cells within a modified multi-layeredmother cell wall. The pectin content of cells at this stageis twice that of control cells grown at pH 6.3. In addition,the relative content of the different pectin fractions is modifiedas a result of the exposure to alkalinity. It is suggested thatthe aggregates result from the repeated failure of the cellsto detach from their original mother cell walls, thus formingclusters which represent several generations of cells. ¹Present address: Division of Food Storage, ARO Beit Dagan,P.O.B. 6, Israel. (Received September 3, 1979; )     

植物激素在小球藻异养培养中的作用

本文研究了几种植物激素对小球藻异养培养的影响。结果表明,IAA、IBA及6-BA三种植物激素均不同程度地促进了小球藻的异养生长,培养≤36h时,IAA或IBA以20mg／L的促进小球藻异养生长的效应最大,100mg／L IAA或IBA则抑制了藻的生长;>36h时,100mg／L IAA或IBA表现出促进小球藻生长的效应,并最终获最大净A540增长量;6-BA以0.1 mg／L的促进作用最大。IBA与6-BA组合同样表现出促进小球藻异养生长的效应,但并非IBA和6-BA简单的加合效应,5 mg／LIBA与6-BA组合的效应维持6-BA单因子的作用趋势,20mg／LIBA与lmg/L6-BA组合的效应大于与0.1mg／L6-BA组合的,100mg／LIBA与0.1mg／L6-BA组合的效应在≤36h时大于与1mg／L6-BA组合的,>36h时则相反。另外,高浓度IBA(≥20mg／L)与6-BA组合抑制了前中期异养藻对葡萄糖的吸收,但加速了中后期葡萄的吸收。再者,IBA与6-BA组合加速了异养小球藻对No-3的吸收。     

Growth, Pigment Synthesis, and Ultrastructural Responses of Phaseolus vulgaris L. cv. Blue Lake to Intermittent and Flashing Light

Growing bean plants (Phaseolus vulgaris L. cv. Blue Lake) on cycles of 1 minute light-1 minute dark or 5 minutes light-5 minutes dark, providing an integrated 12 hours light-12 hours dark per day for each set of plants, led to production after 21 days of new leaves low or lacking in chloroplast pigments. Subsequently, dry weight increase was sharply cut. Leaf area was affected by the light regimes after the second week of growth. By the fourth week, plants on the 1 minute light-1 minute dark cycle showed about one-half the leaf area of the controls. Shoot growth was favored over root growth to the greatest degree on the 1 minute light-1 minute dark regimes. Chlorophyll a/b ratios were close to 3.0 in all of the intermittent light regimes, but the total amounts of chlorophyll in milligrams per primary leaf were higher from day 9 to day 23 for the 12 hour light-12 hours dark controls than for other plants.

Although they produced chlorophyll, the plants receiving 1 or 2 milliseconds per second of light continued to lose weight at the same rate as the dark controls; thus, it is assumed there was no net photosynthesis. Plants receiving flashing light allocated significantly more food reserves from the seed to roots than did dark controls. Total chlorophyll formation was significantly accelerated by 2 milliseconds per second light. With 1 millisecond per second light, it took 5 days longer to achieve the same level of chlorophyll. After the 18th day, there was a steady decline in chlorophyll, b degrading more rapidly than a.

It is thought that several light-driven reactions are involved in the observed pigment synthesis, photosynthesis, food allocation, and growth of bean. Some of these reactions may be cyclic and others linear. Collectively, they must reach a harmonic point for normal metabolism and development to occur. Because time courses for each of these reactions are different, the intermittent and flashing light technique offers the possibility of individually studying some of the key light-driven reactions.

     

谷氨酸对异养培养小球藻生长的影响

在不添加其他氮源的小球藻异养培养基中,谷氨酸可促进小球藻的生长,但基本不增加叶绿素的合成;在以铵盐为氮源时,谷氨酸可明显促进小球藻对铵盐的利用,促进小球藻生物量增加和叶绿素合成;在以硝酸盐为氮源时,谷氨酸可增加小球藻的生物量,对叶绿素含量无明显影响。     

植物激素在小球藻异养培养中的作用

本文研究了几种植物激素对小球藻异养培养的影响。结果表明 ,IAA、IBA及 6_BA三种植物激素均不同程度地促进了小球藻的异养生长 ,培养≤ 36h时 ,IAA或IBA以 2 0mg/L的促进小球藻异养生长的效应最大 ,1 0 0mg/LIAA或IBA则抑制了藻的生长 ;>36h时 ,1 0 0mg/LIAA或IBA表现出促进小球藻生长的效应 ,并最终获最大净A540 增长量 ;6_BA以 0 1mg/L的促进作用最大。IBA与 6_BA组合同样表现出促进小球藻异养生长的效应 ,但并非IBA和 6_BA简单的加合效应 ,5mg/LIBA与 6_BA组合的效应维持 6_BA单因子的作用趋势 ,2 0mg/LIBA与 1mg/L 6_BA组合的效应大于与 0 .1mg/L 6_BA组合的 ,1 0 0mg/LIBA与 0 .1mg/L 6_BA组合的效应在≤ 36h时大于与 1mg/L 6_BA组合的 ,>36h时则相反。另外 ,高浓度IBA(≥ 2 0mg/L)与 6_BA组合抑制了前中期异养藻对葡萄糖的吸收 ,但加速了中后期葡萄的吸收。再者 ,IBA与 6_BA组合加速了异养小球藻对NO_3 的吸收。     

Changes in the Contents of Inactive Complexes of Photosystem II in Chlorella Cells Incubated in the Light and Darkness

A prolonged (20–24 h) dark incubation of Chlorella pyrenoidosa algae at 37–38° did not diminish the relative yield of the variable chlorophyll fluorescence (F _v/F _m) and enhanced the relative contribution of the slow phase (sF _v) to the kinetics of F _v increase. Iodoacetamide, a nonmetabolized glucose analog, 2-deoxyglucose (2-DG), an inhibitor of protein synthesis, cycloheximide, and a decrease in the temperature of dark incubation to 18–20° prevented this sF _v increase. Both the illumination of dark-incubated cells and the addition of 2-DG in darkness restored the initial level of sF _v. In the light-grown chlorella cells, the relative contribution of sF _v reversibly declined with lowering light intensity and increased when ₂ was excluded from the bubbling mixture. The authors presume that the slow phase in the kinetics of F _v increase is related to the functioning of the fraction of the photosystem II complexes with a destabilized primary quinone acceptor of electron, and the content of these complexes in the cell depends on the plastoquinone redox state.     

Stomatal Responses of Tradescantia albiflora to Changing Air Humidity in Light and in Darkness

Tradescantia albiflora has green variegated and white leaves.Its stomatal apparatus consists of the guard cells and two pairsof subsidiary cells. Investigations were carried out by observingthe stomata microscopically by means of a video system in situin a CO₂ exchange chamber and by simultaneously measuring thegas exchange of the leaves. In response to air humidity changes,stomatal movements in T. albiflora begin, owing to turgor changes,in the polar and lateral subsidiary cells. The stomatal responseof green leaves to changes of air humidity showed typical transientand oscillatory phases prior to steady-state reactions. In darkness,stomata closed when air humidity decreased; however, they didnot reopen when air humidity was raised again. Stomata of illuminatedwhite leaves responded like those of green leaves in darkness.With increasing soil water stress stomata responded to changingair humidity with reductions of the transient phases and a decreasingtendency to reopen when air humidity became high again. CO₂deficiency of the air caused the stomata to open in the dark,and interacted with the air humidity effect in such a way thatstomata of green leaves responded to air humidity changes indarkness in a similar way as they did in light. Key words: Stomata, humidity response, green and white leaf areas, CO₂ deficient air     

硒胁迫对小球藻光合色素含量和生长的影响

50 mg/L的硒胁迫下,小球藻中β-胡萝卜素、类叶黄素、叶绿素(叶绿素a、叶绿素b)的含量先明显增加,之后逐渐下降;而在800 mg/L硒胁迫下,各种光合色素含量均明显下降;小球藻活体细胞在693 nm处出现叶绿素a的吸收峰,其吸收值在硒胁迫后明显减弱;在室温荧光发射光谱中,700 nm处的发射峰随着硒浓度的增加而显著下降;荧光激发光谱表明:硒胁迫使小球藻的能量传递受阻,传递效率降低;藻体中水溶性蛋白含量随着硒胁迫的增强而下降.等离子体原子发射光谱(ICP-AES)研究结果表明:随着硒浓度增加,藻体中Mg2 、Ca2 、K 和Na 含量呈明显降低趋势,而培养液中这些离子的浓度则不断增加.     

Acute Behavioral Responses to Light and Darkness in Nocturnal Mus musculus and Diurnal Arvicanthis niloticus

The term masking refers to immediate responses to stimuli that override the influence of the circadian timekeeping system on behavior and physiology. Masking by light and darkness plays an important role in shaping an organism's daily pattern of activity. Nocturnal animals generally become more active in response to darkness (positive masking) and less active in response to light (negative masking), and diurnal animals generally have opposite patterns of response. These responses can vary as a function of light intensity as well as time of day. Few studies have directly compared masking in diurnal and nocturnal species, and none have compared rhythms in masking behavior of diurnal and nocturnal species. Here, we assessed masking in nocturnal mice (Mus musculus) and diurnal grass rats (Arvicanthis niloticus). In the first experiment, animals were housed in a 12:12 light-dark (LD) cycle, with dark or light pulses presented at 6 Zeitgeber times (ZTs; with ZT0 = lights on). Light pulses during the dark phase produced negative masking in nocturnal mice but only at ZT14, whereas light pulses resulted in positive masking in diurnal grass rats across the dark phase. In both species, dark pulses had no effect on behavior. In the 2nd experiment, animals were kept in constant darkness or constant light and were presented with light or dark pulses, respectively, at 6 circadian times (CTs). CT0 corresponded to ZT0 of the preceding LD cycle. Rhythms in masking responses to light differed between species; responses were evident at all CTs in grass rats but only at CT14 in mice. Responses to darkness were observed only in mice, in which there was a significant increase in activity at CT 22. In the 3rd experiment, animals were kept on a 3.5:3.5-h LD cycle. Surprisingly, masking was evident only in grass rats. In mice, levels of activity during the light and dark phases of the 7-h cycle did not differ, even though the same animals had responded to discrete photic stimuli in the first 2 experiments. The results of the 3 experiments are discussed in terms of their methodological implications and for the insight they offer into the mechanisms and evolution of diurnality.     

Increase of Copper Toxicity to Growth of Chlorella vulgaris with Increase of Light Intensity

Abstract In the absence of inhibitory concentrations of copper, the photoautotrophic growth of Chlorella vulgaris INETI58C at 27°C exhibited a higher specific growth rate and reached a higher maximal concentration of biomass, under irradiance of 150 W m⁻², compared with 100 W m⁻². However, when the mineral growth medium was supplemented with CuSO₄ (range 40–80 μM), algal growth was significantly affected at the higher light intensity. In the presence of Cu²⁺, the increase in dry biomass was uncoupled from the increase in cell concentration since more than 16 autospores gathered together, inside the enlarged mother cell, suggesting that copper arrested the normal bursting of the mother cell wall. At the higher irradiance, growth medium supplementation with 80 μM of CuSO₄ led to bleaching of photosynthetic pigments. No growth was observed, while, under the lower irradiance, growth was only slightly inhibited. Results clearly showed that copper toxicity to growth of C. vulgaris was strongly influenced by light intensity. Higher light intensity elicits lethal or sublethal Cu²⁺ damage at concentrations lower than the threshold level for damage at lower light intensities. Cu²⁺ may elicit lethal or sublethal light damage at irradiances lower than the threshold level for unpolluted aquatic systems. Received: 17 January 1997; Accepted 15 April 1997     

The effects of pH and temperature on orthophosphate removal by immobilized Chlorella vulgaris

Summary When Chlorella vulgaris was immobilized in calcium alginate beads, it removed more than 90% phosphate (10mg P/L) added to artificial wastewater at pH 3 to 9 and from 10 to 30°C. Free cells, however, only removed 40–60% of added phosphate at low pH (3–5) and at 10°C. Immobilized C. vulgaris is shown to have great potentialities for removing phosphate from low pH wastewater and at low temperature.     

The Heterotrophic Nutrition of Chlorella vulgaris (Brannon No. 1 Strain): With two Figures In the Text

A range of sugars, sugar alcohols, sugar phosphates, organicacids, and monohydric alcohols have been tested as carbon sourcesfor growth and as respiratory substrates using Chlorella vulgaris,Brannon I, grown in darkness. Much higher rates of growth and respiration were obtained withd-glucose than with any other substance tested. Ethanol (at0·005 M.) sustained both growth and respiration at c.50 per cent, of the level with glucose (0·028 M. or higher).Evidence was obtained that the organism can become ‘adapted’to utilize d-galactose and sucrose as effective carbon sources.Sustained growth was not obtained with any of the other substancestested. The glucose monophosphates, methanol and certain organic acids(oxalacetate, -ketoglutarate, cis-aconitate, and pyruvate) clearlystimulated oxygen uptake but to a less extent than ethanol.The other substances tested were either inhibitory to respirationor inactive or of very low activity as substrates. The growth in darkness and in liquid culture of Chlorella whensupplied with d-glucose was insensitive to pH over the range4·5 to 7·0 and was markedly enhanced by a highlevel of aeration. Gains in cellular dry weight ranging from45 to 90 per cent, of the weight of d-glucose disappearing fromthe culture medium were recorded in growth experiments; measurementsof CO₂ evolution in the Warburg indicated retention of up totwo-thirds of the glucose-C in cell material.