PHOTOINHIBITION OF MECHANICALLY STIMULABLE BIOLUMINESCENCE IN THE AUTOTROPHIC DINOFLAGELLATE CERATIUM FUSUS (PYRROPHYTA)1

Ceratium fusus (Ehrenb.) Dujardin was exposed to light of different wavelengths and photon flux densities (PFDs) to examine their effects on mechanically stimulable bioluminescence (MSL). Photoinhibition of MSL was proportional to the logarithm of PFD. Exposure to I μmol photons·m^?2s^?1 of broadband blue light (ca. 400–500 nm) produced near-complete photoinhibition (≥90% reduction in MSL) with a threshold at ca. 0.01 μmol photons·m^?2·s^?1. The threshold of photoinhibition was ca. an order of magnitude greater for both broadband green (ca. 500–580 nm) and red light (ca. 660–700 nm). Exposure to narrow spectral bands (ca. 10 nm half bandwidth) from 400 and 700 nm at a PFD of 0.1 μmol photons·m^?2·s^?1 produced a maximal response of photoinhibition in the blue wavelengths (peak ca. 490 nm). A photoinhibition response (≥ 10%) in the green (ca. 500–540 nm) and red wavelengths (ca. 680 nm) occurred only at higher PFDs (1 and 10 μmol photons·m^?2·s^?1). The spectral response is similar to that reported for Gonyaulax polyedra Stein and Pyrocystis lunula Schütt and unlike that of Alexandrium tamarense (Lebour) Balech et Tangen. The dinoflagellate's own bioluminescence is two orders of magnitude too low to result in self-photoinhibition. The quantitative relationships developed in the laboratory predict photoinhibition of bioluminescence in populations of C. fusus in the North Atlantic Ocean.     

OCCURRENCE AND ABUNDANCE OF GREEN-FLUORESCING DINOFLAGELLATES IN SURFACE WATERS OF THE NORTHWEST ATLANTIC AND NORTHEAST PACIFIC OCEANS1

We have cultured green fluorescing heterotrophic dinoflagellates whose continuous green fluorescence is due to an unidentified compound, probably a flavin, that excites with blue (～460 nm) light and emits green (～535 nm) light. No evidence of bioluminescence was found, but we note that compounds with similar fluorescence characteristics have been associated with bioluminescence in other taxa. These cells, all naked gymnodinoids, are widespread and abundant in the Northwest Atlantic and Northeast Pacific Oceans (10³–10⁵ L^?1). They comprise 4–100% of the total heterotrophic dinoflagellate component which, in turn, is usually equivalent magnitude to the phototrophic naked dinoflagellate component of the phytoplankton community.     

Bioluminescence of Marine Dinoflagellates : I. An underwater photometer for day and night measurements

Portable light-baffled underwater photometers have been designed for the measurement of dinoflagellate bioluminescence by day and night. Maximal light emission is obtained by mechanical stimulation in a defined volume. The pump which stimulates the dinoflagellates also constantly replenishes the sample volume so that continuous measurements are possible. Evidence for both diurnal variation and vertical migration is presented. Using luminous bacteria for calibration a single dinoflagellate has been found to emit of the order of 10¹⁰ light quanta per flash. The technique suggests that large scale mapping of bioluminescence is feasible.     

Diversity of the Luciferin Binding Protein Gene in Bioluminescent Dinoflagellates – Insights from a New Gene in Noctiluca scintillans and Sequences from Gonyaulacoid Genera

Dinoflagellate bioluminescence systems operate with or without a luciferin binding protein, representing two distinct modes of light production. However, the distribution, diversity, and evolution of the luciferin binding protein gene within bioluminescent dinoflagellates are not well known. We used PCR to detect and partially sequence this gene from the heterotrophic dinoflagellate Noctiluca scintillans and a group of ecologically important gonyaulacoid species. We report an additional luciferin binding protein gene in N. scintillans which is not attached to luciferase, further to its typical combined bioluminescence gene. This supports the hypothesis that a profound re‐organization of the bioluminescence system has taken place in this organism. We also show that the luciferin binding protein gene is present in the genera Ceratocorys, Gonyaulax, and Protoceratium, and is prevalent in bioluminescent species of Alexandrium. Therefore, this gene is an integral component of the standard molecular bioluminescence machinery in dinoflagellates. Nucleotide sequences showed high within‐strain variation among gene copies, revealing a highly diverse gene family comprising multiple gene types in some organisms. Phylogenetic analyses showed that, in some species, the evolution of the luciferin binding protein gene was different from the organism's general phylogenies, highlighting the complex evolutionary history of dinoflagellate bioluminescence systems.     

Action spectrum for a low intensity, rapid photoinhibition of mechanically stimulable bioluminescence in the marine dinoflagellates Gonyaulax catenella, G. acatenella, and G. tamarensis

The mechanically stimulable bioluminescence of members of the Gonyaulax catenella group can be maximally photoinhibited by exposure to as few as 10¹³ quanta/cm², a factor 10⁴ times smaller than that required for comparable photoinhibition in Gonyaulax polyedra and all other photosynthetic bioluminescent dinoflagellates investigated. Following an irradiation pulse there is an initial time lag of one minute, followed by a rapid decrease in mechanical stimulability to approximately 1% of the dark unirradiated control with a firstorder rate constant as high as 0.01 sec^?1. Action spectra for all three species imply a pigment with a single absorption band having a maximum at 562 nm and a half band width of 105 nm within the spectral range 325 nm to 775 nm. Photoinhibition appears to decrease either the sensitivity of the shear receptor mechanism or the efficiency of signal transmission in the dinoflagellates, since chemically stimulable bioluminescence is unaffected by these exposures.     

Behavioral responses of the coastal copepod Acartia hudsonica (Pinhey) to simulated dinoflagellate bioluminescence

Light pulses were used to mimic dinoflagellate bioluminescence and test its effects on the swimming behavior of Acartia hudsonica (Pinhey). The horizontal swimming patterns of the copepod were tracked and described using a video-computer system. Single flashes of light of 60 ms duration, with a wavelength of peak emission of 475 nm and an intensity of 2 μE · m^?2 · s^?1 caused a “startle” response consisting of a short burst of high speed swimming. A series of these flashes repeated every 5 s resulted in higher average swimming speed, more swimming speed bursts, and straighter paths. These behavioral changes are similar to those previously found for A. hudsonica in the presence of bioluminescent dinoflagellates. The effects of altering the intensity, duration, and color of the simulated dinoflagellate flash were also tested. Our results support the hypothesis that dinoflagellate bioluminescence is a highly evolved adaptation for repelling nocturnal grazers.     

Effects of light and temperature on the growth of Takayama helix (Dinophyceae): mixotrophy as a survival strategy against photoinhibition

Takayama helix is a mixotrophic dinoflagellate that can feed on diverse algal prey. We explored the effects of light intensity and water temperature, two important physical factors, on its autotrophic and mixotrophic growth rates when fed on Alexandrium minutum CCMP1888. Both the autotrophic and mixotrophic growth rates and ingestion rates of T. helix on A. minutum were significantly affected by photon flux density. Positive growth rates of T. helix at 6–58 μmol photons · m^?2 · s^?1 were observed in both the autotrophic (maximum rate = 0.2 · d^?1) and mixotrophic modes (0.4 · d^?1). Of course, it did not grow both autotrophically and mixotrophically in complete darkness. At ≥247 μmol photons · m^?2 · s^?1, the autotrophic growth rates were negative (i.e., photoinhibition), but mixotrophy turned these negative rates to positive. Both autotrophic and mixotrophic growth and ingestion rates were significantly affected by water temperature. Under both autotrophic and mixotrophic conditions, it grew at 15–28°C, but not at ≤10 or 30°C. Therefore, both light intensity and temperature are critical factors affecting the survival and growth of T. helix.     

Nyctohemeral variations of marine bioluminescence in the Mediterranean and the northeastern Atlantic]

Bioluminescence measurements have been made using a bathyphotometer allowing the determination of stimulated light intensities down to 2,000 m depth, in the Mediterranean Sea on the Almeria-Oran front, during the winter 1997-1998, and in the northeastern Atlantic, on the Armorican continental shelf, during summers 1999 and 2000. Bioluminescence is weaker in the Mediterranean than in the Atlantic. In the epipelagic waters, day/night variations appear clearly, stimulated bioluminescence is higher at night than during the day. These diel variations can be explained by vertical migration of bioluminescent organisms and by photoinhibition of dinoflagellate bioluminescence. Fluorescence measurements made at the same time give information about potential bioluminescent sources, autotrophic and heterotrophic.     

OLIGONUCLEOTIDE PRIMERS FOR THE DETECTION OF BIOLUMINESCENT DINOFLAGELLATES REVEAL NOVEL LUCIFERASE SEQUENCES AND INFORMATION ON THE MOLECULAR EVOLUTION OF THIS GENE1

Bioluminescence is reported in members of 18 dinoflagellate genera. Species of dinoflagellates are known to have different bioluminescent signatures, making it difficult to assess the presence of particular species in the water column using optical tools, particularly when bioluminescent populations are in nonbloom conditions. A “universal” oligonucleotide primer set, along with species and genus‐specific primers specific to the luciferase gene were developed for the detection of bioluminescent dinoflagellates. These primers amplified luciferase sequences from bioluminescent dinoflagellate cultures and from environmental samples containing bioluminescent dinoflagellate populations. Novel luciferase sequences were obtained for strains of Alexandrium cf. catenella (Whedon et Kof.) Balech and Alexandrium fundyense Balech, and also from a strain of Gonyaulax spinifera (Clap. et Whitting) Diesing, which produces bioluminescence undetectable to the naked eye. The phylogeny of partial luciferase sequences revealed five significant clades of the dinoflagellate luciferase gene, suggesting divergence among some species and providing clues on their molecular evolution. We propose that the primers developed in this study will allow further detection of low‐light‐emitting bioluminescent dinoflagellate species and will have applications as robust indicators of dinoflagellate bioluminescence in natural water samples.     

THE ROLE OF CALCIUM IN FLOW‐STIMULATED BIOLUMINESCENCE OF THE RED TIDE DINOFLAGELLATE LINGULODINIUM POLYEDRUM

Many marine planktonic dinoflagellates emit flashes of light in response to either laminar or turbulent flows as well as direct mechanical stimulation. The production of a flash of light is known to be mediated by a proton‐mediated action potential across the vacuolar membrane; the mechanotransduction process initiating this action potential is unknown. Here we report on an investigation into the role of Ca⁺² in the mechanotransduction process regulating bioluminescence in the red tide dinoflagellate Lingulodinium polyedrum. Calcium ionophores and low concentrations of the membrane‐disrupting agent digitonin stimulated bioluminescence only when calcium was present in the media or added with the agent, indicating that the flash‐triggering vacuolar action potential is specifically stimulated by a calcium influx. A variety of known calcium channel blockers or antagonists inhibited mechanically stimulated bioluminescence but did not affect cellular bioluminescent capacity. In many cases the inhibitory affect occurred after only a brief exposure. In addition, gadolinium (Gd⁺³), a blocker of many stretch‐activated ion channels, caused potent inhibition of mechanically stimulated bioluminescence. The order of potency of the transition metals tested was La⁺³ > Gd⁺³ > Co⁺² > Mn⁺² > Ni⁺², similar to their potency as blockers of known calcium channels. Experiments with a quantified shear flow demonstrated that flow‐stimulated bioluminescence depended on the level of extracellular calcium. Future work will elucidate the signaling pathway involving calcium‐mediated flow‐stimulated mechanotransduction. Our goal is to use bioluminescence as a proxy for the initial cellular mechanotransduction events triggered by fluid flow.     

CHLOROPLAST PIGMENT PATTERNS IN DINOFLAGELLATES1

The chlorophylls and carotenoids of 22 species of dinoflagellates were analysed by thin layer chromatography, using 2-dimensional sucrose plates, and 1-dimensional polyethylene plates for chlorophylls c₁ and c₂. Peridinin was the major carotenoid in 19 of the species, while fucoxanthin was the major carotenoid in 3. In the peridinin-containing species, 5 carotenoid fractions, constituting more than 95% of the total carotenoids, were always present. These were peridinin (± neo-peridinin), averaging 64% of the total carotenoid, diadinoxanthin, dinoxanthin, β-carotene and a polar, unidentified pink xanthophyll. Six other carotenoid fractions occurred in minor or trace quantities among the species, but were not identified. Two of these had, a wide distribution; the other 4 were restricted to one or 2 species. The chlorophyll content of the dinoflagellate cultures ranged from 1–141 μg chlorophyll a + c/10⁶ cells, a pattern which was broadly correlated with cell size. In the peridinin-containing species the ratio of chlorophyll a to c on a molar basis was approximately 2 (range 1.60–4.39); in the fucoxanthin-containing species this ratio was approximately 4 (range 2.65–5.73). Both chlorophylls c₁ and c₂ occurred in the fucoxanthin-containing dinoflagellates, and only chlorophyll c₂ (one exception) occurred in the peridinin-containing dinoflagellates. These patterns of chlorophyll c and major carotenoid correspond to patterns previously observed in the Pyrrhophyta and the Chrysophyta, suggesting different phylogenetic origins for the “dinoflagellate” chloroplasts.     

PLANKTONIC SARCODINES: MICROHABITAT FOR OCEANIC DINOFLAGELLATES1

Two morphologically distinct species of free-swimming dinoflagellates belonging to the genus Gyrodinium utilize the spine and rhizopodial environments of planktonic foraminifera and colonial radiolaria as microhabitats. Up to 84% of the sarcodines examined in a given population were associated with these dinoflagellates at densities up to 20,000 cells per sarcodine in some radiolarian colonies. Both dinoflagellate species possess chloroplasts, indicating they are capable of autotrophy. ¹⁴C-labelling experiments with the radiolarian-associated dinoflagellate demonstrate that it can take up inorganic carbon under both light and dark conditions. Ultrastructural evidence suggests the foraminiferal dinoflagellate may be capable of phagotrophy. Hence, these algae should be considered mixotrophs. An unusual cytoplasmic extension used for attachment and possibly feeding occurs in the foraminiferal-associated Gyrodinium and is documented with electron microscopy. Ultrastructural examination suggests this organelle may be hydrostatically controlled and may be an extension of the sac pusule.     

The mechanical triggering of bioluminescence in marine dinoflagellates: chemical basis

In both photosynthetic (Pyrodinium bahamense, Gonyaulax polyedra, Pyrocystis Iunula, P. noctiluca, P. fusiformis) and nonphotosynthetic (Noctiluca miliaris) bioluminescent dinoflagellates chemical stimulation can by-pass mechanical stimulation. The effective ions are Ca⁺⁺, K⁺, NH₄⁺ and H⁺. Other chemicals found effective are those implicated in Ca⁺⁺ transport or binding. There are interspecies differences in degrees of mechanical and chemical stimulability. Photoinhibition of mechanical stimulability is the result of two effects, the first a reduction in total bioluminescence potential and the second a decrease in mechanical stimulability resulting experimentally in a decreased rate of light emission. This latter effect can be reversed with Ca⁺⁺ ions. Chemicals which bind Ca⁺⁺ or displace Ca⁺⁺ can mimic the effects of photoinhibition. The chemical inhibition of mechanical stimulability is also reversed by Ca⁺⁺ ions. A scheme is proposed which is consistent for all species examined.     

Bottom-up and top-down controls of the microbial food web in the Southern Ocean: experiments with manipulated microcosms

Summary We have studied bottom-up and top-down control of the Southern Ocean microbial food web by microcosm experiments. Water from the Weddell Sea and Weddell Scotia Confluence were used for the experiments. Microcosms were manipulated by nutrients and light, and by size-selective screening. Incubation at the higher light level doubled phytoplankton growth rates from 0.12 to 0.24 day^–1 in the Weddell experiment and from 0.15 to 0.30 day^–1 in the Confluence experiment. Nutrient enrichment had no significant effect on growth rates in either of the experiments, indicating that phytoplankton growth was not nutrient-limited. In the microcosms where dinoflagellate growth rate was different, high dinoflagellate numbers were reflected as depressed nanoflagellate growth as well as depressed growth of phytoplankton, suggesting that dinoflagellates controlled both heterotrophic nanoflagellates and autotrophic nanoplankton. Only during short periods, when dinoflagellate numbers were low, could exponential growth of nanoflagellates be demonstrated. Bacterioplankton growth rates were, on average, 0.26 day^–1 in the Weddell experiment and 0.22 day^–1 in the Confluence experiment. Bacteria were controlled by heterotrophic nanoflagellates. Potential growth rates up to 0.75 day^–1 were measured from batch cultures without predators. With the microcosm experiments, we could demonstrate a strong top-down control by dinoflagellates on phytoplankton and on heterotrophic nanoflagellates, and a control by heterotrophic nanoflagellates on bacteria. We could also demonstrate weak nutrient limitation on autotrophs and substrate limitation on heterotrophic bacteria. In the two study areas, biomass production and carbon flow were mediated mainly by organisms that passed through a 20 m net and had growth rates in the order of 0.20 to 0.30 day^–1.Data presented here were collected during the European Polarstern Study (EPOS) sponsored by the European Science Foundation     

Immobilized Growth of the Peridinin-Producing Marine Dinoflagellate Symbiodinium in a Simple Biofilm Photobioreactor

Products from phototrophic dinoflagellates such as toxins or pigments are potentially important for applications in the biomedical sciences, especially in drug development. However, the technical cultivation of these organisms is often problematic due to their sensitivity to hydrodynamic (shear) stress that is a characteristic of suspension-based closed photobioreactors (PBRs). It is thus often thought that most species of dinoflagellates are non-cultivable at a technical scale. Recent advances in the development of biofilm PBRs that rely on immobilization of microalgae may hold potential to circumvent this major technical problem in dinoflagellate cultivation. In the present study, the dinoflagellate Symbiodinium voratum was grown immobilized on a Twin-Layer PBR for isolation of the carotenoid peridinin, an anti-cancerogenic compound. Biomass productivities ranged from 1.0 to 11.0 g m^?2 day^?1 dry matter per vertical growth surface and a maximal biomass yield of 114.5 g m^?2, depending on light intensity, supplementary CO₂, and type of substrate (paper or polycarbonate membrane) used. Compared to a suspension culture, the performance of the Twin-Layer PBRs exhibited significantly higher growth rates and maximal biomass yield. In the Twin-Layer PBR a maximal peridinin productivity of 24 mg m^?2 day^?1 was determined at a light intensity of 74 μmol m^?2 s^?1, although the highest peridinin content per dry weight (1.7 % w/w) was attained at lower light intensities. The results demonstrate that a biofilm-based PBR that minimizes hydrodynamic shear forces is applicable to technical-scale cultivation of dinoflagellates and may foster biotechnological applications of these abundant marine protists.     

SPONTANEOUS AND STIMULATED BIOLUMINESCENCE OF THE DINOFLAGELLATE CERATOCORYS HORRZDA (PERIDINIALES)1

This is the first report of spontaneous bioluminescence in the autotrophic dinoflagellate Ceratocorys horrida von Stein. Bioluminescence was measured, using an automated data acquisition system, in a strain of cultured cells isolated from the Sargasso Sea. Ceratocorys horrida is only the second dinoflagellate species to exhibit rhythmicity in the rate of spontaneous flashing, flash quantum flux (intensity), and level of spontaneous glowing. The rate of spontaneous flashing was maximal during hours 2–4 of the dark phase [i.e. circadian time (CT)16–18 for a 14:10 h LD cycle (LD14:10)], with approximately 2% of the population flashing-min^?1, a rate approximately one order of magnitude greater than that of the dinoflagellate Gonyaulax polyedra. Flash quantum flux was also maximal during this period. Spontaneous flashes were 134 ms in duration with a maximum flux (intensity) of 3.1×10⁹ quanta-s^?1. Light emission presumably originated from blue fluorescent microsources distributed in the cell periphery and not from the spines. Values of both spontaneous flash rate and maximum flux were independent of cell concentration. Isolated cells also produced spontaneous flashes. Spontaneous glowing was dim except for a peak of 6.4× 10⁴quanta-s^?1 cell^?1, which occurred at CT22.9 for LD14:10 and at CT22.8 for LD12:12. The total integrated emission of spontaneous flashing and glowing during the dark phase was 4×10⁹ quantacell^?1, equivalent to the total stimulable luminescence. The rhythms for C. horrida flash and glow behavior were similar to those of Gonyaulax polyedra, although flash rate and quantum flux were greater. Spontaneous bioluminescence in C. horrida may be a circadian rhythm because it persisted for at least three cycles in constant dark conditions. This is also the first detailed study of the stimulated bioluminescence of C. horrida, which also displayed a diurnal rhythm. Cultures exhibited >200 times more mechanically stimulated bioluminescence during the dark phase than during the light phase. Mechanical stimulation during the dark phase resulted in 6.7 flashes. cell^?1; flashes were brighter and longer in duration than spontaneous flashes. Cruise-collected cells exhibited variability in quantum flux with few differences in flash kinetics. The role of dinoflagellate spontaneous bioluminescence in the dynamics of near-surface oceanic communities is unknown, but it may be an important source of natural in situ bioluminescence.     

Contrasting effects of high‐intensity photosynthetically active radiation on two bloom‐forming dinoflagellates

While light limitation can inhibit bloom formation in dinoflagellates, the potential for high‐intensity photosynthetically active radiation (PAR) to inhibit blooms by causing stress or damage has not been well‐studied. We measured the effects of high‐intensity PAR on the bloom‐forming dinoflagellates Alexandrium fundyense and Heterocapsa rotundata. Various physiological parameters (photosynthetic efficiency F_v/F_m, cell permeability, dimethylsulfoniopropionate [DMSP], cell volume, and chlorophyll‐a content) were measured before and after exposure to high‐intensity natural sunlight in short‐term light stress experiments. In addition, photosynthesis‐irradiance (P‐E) responses were compared for cells grown at different light levels to assess the capacity for photophysiological acclimation in each species. Experiments revealed distinct species‐specific responses to high PAR. While high light decreased F_v/F_m in both species, A. fundyense showed little additional evidence of light stress in short‐term experiments, although increased membrane permeability and intracellular DMSP indicated a response to handling. P‐E responses further indicated a high light‐adapted species with Chl‐a inversely proportional to growth irradiance and no evidence of photoinhibition; reduced maximum per‐cell photosynthesis rates suggest a trade‐off between photoprotection and C fixation in high light‐acclimated cells. Heterocapsa rotundata cells, in contrast, swelled in response to high light and sometimes lysed in short‐term experiments, releasing DMSP. P‐E responses confirmed a low light‐adapted species with high photosynthetic efficiencies associated with trade‐offs in the form of substantial photoinhibition and a lack of plasticity in Chl‐a content. These contrasting responses illustrate that high light constrains dinoflagellate community composition through species‐specific stress effects, with consequences for bloom formation and ecological interactions within the plankton.     

RESPONSE OF PROROCENTRUM MARIAE-LEBOURIAE (DINOPHYCEAE) TO LIGHT OF DIFFERENT SPECTRAL QUALITIES AND IRRADIANCES: GROWTH AND PIGMENTATION1

Growth and pigment concentrations of the, estuarine dinoflagellate, Prorocentrum mariae-lebouriae (Parke and Ballantine) comb. nov., were measured in cultures grown in white, blue, green and red radiation at three different irradiances. White irradiances (400–800 nm) were 13.4, 4.0 and 1.8 W · m^?2 with photon flux densities of 58.7 ± 3.5, 17.4 ± 0.6 and 7.8 ± 0.3 μM quanta · m^?2· s^?1, respectively. All other spectral qualities had the same photon flux densities. Concentrations of chlorophyll a and chlorophyll c were inversely related to irradiance. A decrease of 7- to 8-fold in photon flux density resulted in a 2-fold increase in chlorophyll a and c and a 1.6- to 2.4-fold increase in both peridinin and total carotenoid concentrations. Cells grown in green light contained 22 to 32% more peridinin per cell and exhibited 10 to 16% higher peridinin to chlorophyll a ratios than cells grown in white light. Growth decreased as a function of irradiance in white, green and red light grown cells but was the same at all blue light irradiances. Maximum growth rates occurred at 8 μM quanta · m^?2· s^?1 in blue light, while in red and white light maximum growth rates occurred at considerably higher photon flux densities (24 to 32 μM quanta · m^?2· s^?1). The fastest growth rates occurred in blue and red radiation. White radiation producing maximum growth was only as effective as red and blue light when the photon flux density in either the red or blue portion of the white light spectrum was equivalent to that of a red or of blue light treatment which produced maximum growth rates. These differences in growth and pigmentation indicate that P. mariae-lebouriae responds to the spectral quality under which it is grown.     

Carotenoid-to-chlorophyll ratio as a proxy for assay of total fatty acids and arachidonic acid content in the green microalga Parietochloris incisa

The relationships between pigment (carotenoid and chlorophyll) content with accumulation of total fatty acids (TFA) and arachidonic acid (AA) were studied in the green microalga Parietochloris incisa (Trebouxiophyceae, Chlorophyta) grown under different PFDs (35, 200, and 400 μmol photons m⁻² s⁻¹) and nitrogen availabilities. The growth of P. incisa under higher light and nitrogen deficiency was accompanied by accumulation of FA, an increase in carotenoid and a decline in chlorophyll content. It was found that the carotenoid-to-chlorophyll ratio (but not the individual pigment content) correlates closely with the volumetric content of both TFA and AA. Analysis of scattering-compensated absorption spectra of P. incisa suspensions revealed their tight relationship in the blue-green range of the spectrum with the carotenoid-to-chlorophyll ratio, TFA, and AA content. These findings allowed the development of algorithms for the non-destructive assay of TFA and AA in cell suspensions in the ranges of 0.09–3.04 and 0.04–1.7 μg mL⁻¹, with accuracy of 0.06 and 0.01 μg mL⁻¹, respectively, via analytically measured carotenoid-to-chlorophyll ratio and using the ratio of absorption coefficients at 510 and 678 nm, with accuracy of 0.07 and 0.02 μg mL⁻¹, respectively. The feasibility of obtaining essential spectral information concerning the physiological condition of P. incisa using a standard spectrophotometer is also shown.     

Variability in the bioluminescence response of the dinoflagellate Pyrocystis lunula

Light emission in dinoflagellates is induced by water motions. But although it is known that mechanical stimulations of these organisms trigger the bioluminescent response, the exact mechanism that involves some cell membrane excitations by fluid motions is not yet fully understood and is still controversial. We show in this experimental study that the accelerated shear flow, created by abrupt rotations of one or both co-axial cylinders of a Couette shearing chamber excites the light emission from cultured dinoflagellates Pyrocystis lunula. Following our first results published earlier that state that pure laminar shear does not excite the main bioluminescent response in dinoflagellates, our present experiments show that both shear and acceleration in the flow are needed to trigger the bioluminescent response. Besides, the probability to stimulate this bioluminescent response under acceleration and shear is deduced from the response curves. This response follows a Gaussian distribution that traduces a heterogeneity in individual cell thresholds for the stimulation of bioluminescence in a dinoflagellate population. All these results will have a repercussion in the possible applications of dinoflagellate bioluminescence in flow visualizations and measurements. Moreover, this study opens a new way in studying mechanically-induced stimulus thresholds at the cell level.