Light response curve methodology and possible implications in the application of chlorophyll fluorescence to benthic diatoms

Chlorophyll a fluorescence has been increasingly applied to benthic microalgae, especially diatoms, for measurements of electron transport rate (ETR) and construction of rapid light response curves (RLCs) for the determination of photophysiological parameters [mainly the maximum relative ETR (rETR_max), the light saturation coefficient (E _k) and the maximum light use coefficient (α)]. Various problems with the estimation of ETR from the microphytobenthos have been identified, especially in situ. This study further examined the effects of light history of the cells and light dose accumulation during RLCs on the fluorescence measurements of ETR using the benthic diatom Navicula phyllepta. RLCs failed to saturate when using incremental increases in irradiance, however, curves with decreasing irradiance did saturate. Patterns indicating photoacclimation in response to light histories were observed, with higher rETR_max and E _k, and lower α, at high light compared to low light. However, these differences could be negated by increasing the RLC irradiance duration from 30 to 60 s. It is suggested that problems arose as a result of rapid fluorescence variations due to ubiquinone (Q_A) oxidation and non-photochemical chlorophyll fluorescence quenching (NPQ) which depended upon the light history of the cells and the RLCs accumulated light dose. Also, RLCs with irradiance duration of 10 s were shown to have a high level of error possibly specific to the fluorimeter programming. It is suggested that RLCs, using a Diving-PAM fluorimeter on benthic diatoms, should be run using decreasing irradiance steps of 30 s duration.     

Effects of light exposure on the retention of kleptoplastic photosynthetic activity in the sacoglossan mollusc <Emphasis Type="Italic">Elysia viridis</Emphasis>

The effects of light exposure on the photosynthetic activity of kleptoplasts were studied in the sacoglossan mollusc Elysia viridis. The photosynthetic activity of ingested chloroplasts was assessed in vivo by non-destructively measuring photophysiological parameters using pulse amplitude modulation (PAM) fluorometry. Animals kept under starvation were exposed to two contrasting light conditions, 30 μmol photons m⁻² s⁻¹ (low light, LL), and 140 μmol photons m⁻² s⁻¹ (high light, HL), and changes in photosynthetic activity were monitored by measuring the maximum quantum yield of photosystem II (PSII), F _v/F _m, the minimum fluorescence, F _o, related to chlorophyll a content, and by measuring rapid light-response curves (RLC) of relative electron transport rate (rETR). RLCs were characterised by the initial slope of the curve, α_RLC, related to efficiency of light capture, and the maximum rETR level, rETR_m,RLC, determined by the carbon-fixation metabolism. Starvation induced the decrease of all photophysiological parameters. However, the retention of photosynthetic activity (number of days for F _v/F _m > 0), as well as the rate and the patterns of its decrease over time, varied markedly with light exposure. Under HL conditions, a rapid, exponential decrease was observed for F _v/F _m, α_RLC and rETR_m,RLC, F _o not showing any consistent trend of variation, and retention times ranged between 6 and 15 days. These results suggested that the retention of chloroplast functionality is limited by photoinactivation of PSII reaction center protein D1. In contrast, under LL conditions, a slower decrease in all parameters was found, with retention times varying from 15 to 57 days. F _v/F _m, α_RLC and rETR_m,RLC exhibited a bi-phasic pattern composed by a long phase of slow decrease in values followed by a rapid decline, whilst F _o decayed exponentially. These results were interpreted as resulting from lower rates of D1 photoinactivation under low light and from the gradual decrease in carbon provided by photosynthesis due to reduction of functional photosynthetic units.     

Chlorophyll fluorescence as a proxy for microphytobenthic biomass: alternatives to the current methodology

Pulse amplitude modulated (PAM) fluorescence has been used as a proxy of microphytobenthic biomass after a dark adaptation period of 15 min to stabilise the minimum fluorescence yield (F _o ¹⁵). This methodology was investigated for in situ migratory and ex situ engineered non-migratory biofilms, comparing dark adaptation to low (5% ambient) and far-red light treatments over different emersion periods. Far-red and low light reduced potential errors resulting from light history effects, by reversal of non-photochemical quenching after 5 min of treatment, compared to over 10 min required by conventional dark adaptation. An in situ decline of minimum fluorescence yield over 15 min was observed during the dark adaptation for migratory biofilms, but was not observed in the non-migratory biofilms indicating that the major cause of decline was downward vertical migration of cells into the sediment. This pattern occurred in far-red light after 10 min, but not for the low light treatment, indicating that low light maintained the biomass at the surface of the sediment. It is therefore concluded that low light treatment is a better option than conventional dark adaptation for the measurement of minimum fluorescence as a proxy of microphytobenthic biomass.     

Time-course of photoadaptation in the photosynthesis-irradiance relationship of a dinoflagellate exhibiting photosynthetic periodicity

Cultures of the marine dinoflagellate Glenodinium sp. were light-shifted and rates of photoadaptation determined by monitoring changes in cell volume, growth rate, pigmentation, parameters of the photosynthesisirradiance (P-I) curves and respiration. To approximate physiological conditions of field populations, cells were cultured on an alternating light-dark cycle of 12hL:12hD, which introduced a daily periodicity of photosynthesis. One result of the present study was to demonstrate how specific parameters of the P-I relationship influenced by periodicity of the light: dark cycle are distinguished from photosynthetic parameters influenced by changes in light level. Under steady-state conditions, rates of both light-saturated (P_max) and light-limited photosynthesis changed in unison over the day; these changes were not related to pigmentation, and displayed their maxima midday. This close relationship between P_max and the slope (a) of the cellular P-I curves in steadystate conditions was quickly adjusted when growth illumination was altered. Rates of light-limited photosynthesis were increased under low light conditions and the periodicity of cellular photosynthesis was maintained. The short-term responses of the P-I relationship to changing light level was different, depending on (1) whether the light shift was from high to low light or vice versa, and (2) whether the high light levels were sufficient to promote maximal photosynthesis rates. Major increases in the photosynthetic carotenoid peridinin, associated with a single type of light-harvesting chromo protein in the chloroplast, was observed immediately upon shifting high light cultures to low light conditions. Following pigment synthesis, significant increases in rates of light-limited photosynthesis were observed in about one-tenth the generation time, while cellular photosynthetic potential was unaffected. it is suggested that general results were consistent with suggested that general results were consistent with earlier reports that the major photoadaptive strategy of Glenodinium sp. is to alter photosynthetic unit (PSU) size. Photoadaptive response times to high light were light-dependent, but appeared to be shower than photoadaptive responses to low light. If light intensities were bright enough to maximize growth rates, photosynthetic response times were on the order of a generation period and pigmentation fell quickly as cells divided at a faster rate. If light-intensities were not sufficient to maximize growth rates, then pigment content did not decline, while rates of light-limited photosynthesis declined quickly. In all cases, photoadaptation was followed best by monitoring fast changes in half saturation constants for photosynthesis, rather than fluctuating changes in pigmentation. Results compared well with time-course phenomena reported for other groups of phytoplankton. Overall, results suggest phytoplankton can bring about photo-induced changes in photosynthesis very quickly and thus accommodate widely fluctuating light regimes over short periods of time.     

Short-term variability in the photosynthetic activity of microphytobenthos as detected by measuring rapid light curves using variable fluorescence

Short-term variability in the photosynthetic activity of microphytobenthos assemblages was studied by measuring chlorophyll fluorescence rapid light curves (RLC), using pulse amplitude modulated (PAM) fluorometry. Measurements carried out on undisturbed samples under dark–light cycles revealed large diel oscillations in both the initial slope of the RLC () and in the maximum relative electron transport rate (ETR_m). Short-term variations in RLC parameters were also observed, closely following changes in incident photon irradiance (E). Increases in irradiance were followed by decreases in and increases in ETR_m, resulting in significant correlations between the light-saturation parameter E_k and E. These results were interpreted as resulting from the onset of reversible energy-dissipating, non-photochemical quenching mechanisms and of compensatory high light-induced activation of carbon metabolism activity. Short-term RLC variability was shown to result mainly from physiological causes and to be detectable only by using short (10–20 s) light steps during RLC construction. Dark-adapted samples kept under constant conditions exhibited apparently endogenous rhythms in RLC parameters and in the maximum quantum yield, F_v/F_m, coincident with vertical migratory movements occurring during subjective photoperiods. These fluctuations appeared to result from the interaction between migratory rhythms and the physiological responses, and from the endogenous activation of processes affecting both the efficiency of energy transfer from light-harvesting antennae to the photosystem II (PSII) reaction centres or from non-radiative pathways (F_v/F_m, ) and the reactions downstream of PSII (ETR_m).Communicated by M. Kühl, Helsingør     

Effects of altered photic regimes on diel patterns of species-specific photosynthesis

Diel patterns of photosynthesis were measured for two polar diatoms (Coscinodiscus sp. and Porosira pseudodenticulata) collected in September 1985 from McMurdo Sound, Antarctica, and four temperate dinoflagellates (Gonyaulax hyalina, Gymnodinium splendens, Dinophysis caudata, and Glenodinium sp.) collected in July 1985 and January 1986 from the Southern California Bight, California, USA. For phytoplankton incubated under three combinations of photoperiod and irradiance, distinct diel patterns of light-saturated (P _max) and light-limited (P _L ) photosynthesis were found for (i) different species isolated from the same environment, and (ii) polar diatoms and temperate dinoflagellates. The time of day when the maximum rate of P _max occurred was influenced by both irradiance and daylength for the polar diatoms but not by daylength for three out of four temperate dinoflagellates. The range of values of the ratio of maximum to minimum rates of photosynthesis (P _max:P _min) was similar for polar diatoms and temperate dinoflagellates. The results of this study suggest that changes in irradiance or photoperiod could influence species-specific patterns of photosynthesis in nature. As a consequence, in light-limited environments differential reproductive success could result from these diel patterns, and ultimately be reflected in temporal and spatial differences in community structure.     

Photosynthetic characteristics of marine phytoplankton from contrasting physical environments

Two suites of phytoplankton samples have been collected in consecutive years at various times over a day from selected depths within vertically mixed and stratified water columns in the western Irish Sea, in order to provide a range of possible light histories within the populations collected. Values for the maximum rate of ¹⁴C retention (P _max) and the initial slope of the ¹⁴C retention: light intensity curve () were obtained. Supra-thermocline samples from the stratified water exhibited higher P _max values than corresponding subthermocline samples. Higher values of were also generally associated with samples from the supra-thermocline zone of the stratified region. Differences in the depth distribution of P _max and in the mixed water were small, except in the presence of a shallow thermocline. In one suite of samples from the stratified water, a diurnal increase in the P _max values of the supra-thermocline samples was observed. P _max values obtained from the samples from the mixed water were interpreted in relation to the distribution obtained from the samples from the stratified zone. Data from both the contrasting sites visited for one sample suite demonstrated a two-phase relationship between the chlorophyll a concentration and both P _max and . The rates of ¹⁴C retention of the first suite of samples were estimated by two techniques. The average differences in the retention were greater in samples from the sub-as opposed to suprathermocline zone. No trends were apparent in the smaples from the mixed waters.     

Relationship between photosynthesis and non-photochemical quenching of chlorophyll fluorescence in two red algae with different carotenoid compositions

Algae are continuously exposed to short-term fluctuations in irradiance. We investigate how two red algae species regulate photosynthetic efficiency to cope with such changes and identify some strategies that differ from higher plants. Two red algae, Gracilaria domingensis and Kappaphycus alvarezii, with antheraxanthin and lutein as major xanthophylls, respectively, reacted to the onset of low light (below E _k) with a substantial decrease of NPQ. This is different from higher plants, but similar to previous observations in, e.g. cyanobacteria where it indicates an increase in the effective absorbance cross-section of Photosystem II (PSII) by state transition. Kinetic studies in continuous light revealed a high susceptibility of PSII to light stress ((1-q _P)/NPQ) in K. alvarezii immediately after the sudden onset of high light, followed by a decrease. This was caused by a slower onset of NPQ in K. alvarezii, followed by acclimation. In G. domingensis, susceptibility of PSII to light stress was stable with time, but absolute values of (1-q _P)/NPQ were higher than in K. alvarezii. These observations suggest that K. alvarezii may be better adapted to high light levels, but is less well prepared for large sudden changes in irradiation. In K. alvarezii, photosynthesis continued to increase with increasing irradiation when NPQ was saturated. As (1-q _P) and NPQ were still balanced in this situation, most likely, processes other than photosynthetic oxygen release are responsible for the increasing net O₂ production observed.     

Different patterns of carotenoid composition and photosynthesis acclimation in two tropical red algae

To be able to survive, marine macroalgae in shallow coastal waters need mechanisms for short-term acclimation to fast changes in their environment. Of major importance are mechanisms that regulate the efficiency of photosynthesis by protecting PS II from photo-oxidative damage. Carotenoids, xanthophyll cycles and non-photochemical quenching (NPQ) are central constituents of such protection mechanisms. Red algae as a group do not have a universal carotenoid composition. We screened ten red algal species and selected two species, originating from similar ecological conditions but with different carotenoid compositions, for use in irradiance-acclimation experiments. We selected the tropical intertidal species Gracilaria domingensis and Kappaphycus alvarezii with antheraxanthin and lutein as major xanthophylls, respectively. Simultaneous in vivo fluorescence and O₂ evolution experiments were performed at different irradiance levels, which allowed a direct comparison of overall photosynthetic performance with NPQ. Interconversions of xanthophylls (violaxanthin, zeaxanthin, β-cryptoxanthin and one unidentified carotenoid) did occur in G. domingensis, but not in response to sudden exposure to light. Thus, NPQ was not correlated with any xanthophyll cycle during short-term acclimation to light. G. domingensis had five times higher weight-specific photosynthetic rates than K. alvarezii, which can be explained by the thicker thallus of K. alvarezii. Chlorophyll-specific gross photosynthetic rates were higher in K. alvarezii, but net rates were the same for both species. G. domingensis showed an immediate strong onset of NPQ upon exposure to irradiance, followed by downregulation to the NPQ level required. In K. alvarezii NPQ increased slowly until the required NPQ level was reached. At high irradiance G. domingensis downregulated photosynthesis while K. alvarezii continued to produce O₂ even at 2,000 μmol photons m⁻² s⁻¹ without NPQ increase. The strategy of K. alvarezii may provide short-term gains but with the risk of oxidative damage. The fast onset of NPQ in G. domingensis even at subsaturating irradiance as well as downregulation of photosynthesis when NPQ is saturated might provide this species with a competitive advantage under conditions of changing irradiance in the field.     

Light-shade adaptation by the oceanic dinoflagellates Pyrocystis noctiluca and P. fusiformis

Species-specific rates of photosynthetic carbon uptake (P), chlorophyll a content and P versus irradiance (P-I), have been measured for cells of Pyrocystis noctiluca and P. fusiformis isolated from natural populations collected in the euphotic zone within and below the surface mixed layer in the Sargasso Sea. These same measurements and the assay for ribulose bis-phosphate carboxylase (RuBP-Case), have been made for cultures of P. noctiluca in a 12 h L: 12 h D photoperiod at 9 different constant or at changing light intensities. In nature chl a cell^-1 was constant throughout the euphotic zone. The photosynthetic capacity (P_max), of cells captured below the surface mixed layer was lower by a factor of 10 compared with cells collected from the surface mixed layer. The P_max for P. noctiluca collected and incubated within the surface mixed layer was the same as for cell cultures grown under high light, non nutrient-limiting conditions, suggesting that photosynthesis in the natural system was not nutrient limited. In laboratory cultures under constant low light intensities, chl a cell^-1 increased by a factor of 5 while both P_max and RuBPCase activity decreased by a factor of ca 4 compared with high light intensities. In changing light intensities both P_max and RuBPCase activities were decreased by factors of 4 during low light intervals while chl a cell^-1 approached a constant intermediate value. The change in chl a cell^-1 in response to prolonged exposure to constant low light intensities was first order with a rate constant of 0.33 d^-1. For all irradiance conditions in culture, the P-I dependence could be described by the simple Michaelis-Menten formula. The ratio of P_max to K_I, (the light intensity where P=P_max/2) was a constant with a Coefficient of Variation of 12%: The constancy of this ratio, the parallel changes in RuBPCase activity with P_max and the constant chl a cell^-1 in the Sargasso Sea imply that for P. noctiluca and presumably P. fusiformis in nature, a dark enzymatic step rather than changes in photosynthetic pigment concentrations may regulate the photosynthetic capacity in the changing photic environment.Contribution no. 1141 from McCollum-Pratt Institute and Department of Biology, The Johns Hopkins University. Supported by DOE contract no. EY 76S20 3278, NSF no. OCE 76-02571 and ONR no. N300014-81-C-0062     

Growth,photosynthesis and carbon metabolism in the temperate marine diatom Skeletonema costatum adapted to low temperature and low photon-flux density

The temperate diatom Skeletonema costatum (Grev.) Cleve was grown in low temperature and/or low light conditions. The cultures were acclimatized for at least three months before experiments were begun. Our data indicate that the initial slope of the photosynthesis vs irradiance curve () is controlled predominantly by light history and the light-saturated photosynthesis (P _max) by temperature. The number of divisions per day decreased with decreasing light intensity, but was identical for cultures grown at 3° or 18°C. The metabolic pathways of inorganic carbon fixation were not fundamentally affected by low temperature or low light intensity, but both these factors increased labelling of C₃ compounds, synthesized by the Calvin-Benson cycle, and decreased that of phosphoenolpyruvate (PEP) and other metabolites. This indicates an enhancement of ribulose-1,5-bisphosphate (RuBP) carboxylase activity, which is the first step in the C₃ pathway (3-phosphoglycerate and sugar phosphate synthesis); this may optimize cell functions. At low temperatures, a seven-fold increase in RuBP carboxylase activity per cell was observed. S. costatum is able to adapt to low irradiance by increasing and decreasing I _k (the ratio of P _max:, light intensity at onset of light saturation), and to low temperature by increasing its cellular chlorophyll a and RuBP carboxylase content. However, in the latter case, adaptation is not optimal. This study revealed two main features: (1) there is evidence that RuBP carboxylase has a key function in adjustment to high rates of photosynthesis at suboptimal temperatures or irradiances; (2) adaptive mechanisms are dynamic processes and the role of the time scale in physiological adaptation should be considered.     

The theoretical basis for estimating phytoplankton production and specific growth rate from chlorophyll, light and temperature data

Phytoplankton maximum specific growth rate, μ_max, and maximum photosynthetic quantum yield, Φ_max, can be related mathematically via the photosynthetic light curve, P(I). A model is presented in which maximum quantum yield defines the initial slope of the light curve and is assumed to be a known constant, while maximum specific growth rate defines the light-saturated region of the curve and is assumed to be a known function of temperature. The effect of introducing μ_max(T) into P(I, Φ_max) is to replace the unknown, temperature-dependent light saturation parameter with a term involving the ratio μ_max/Φ_max. The advantage of writing P(I) in terms of both μ_max and Φ_max is that those parameters are particularly well documented in the literature. Consequently, estimates of nutrient-unlimited phytoplankton growth and production rates can be based solely on the constants μ_max, Φ_max and k_c (light absorption per unit of chlorophyll) and the free variables light, temperature and chlorophyll concentration. Rate estimates appear to be accurate to within a factor of two for an extremely wide range of conditions.One particularly significant result of introducing μ_max into P(I, Φ_max) is that the carbon : chlorophyll ration, θ, appears explicitly. It is possible to derive an expression for optimum θ based on the assumption that adaptive changes in carbon/chlorophyll occur so as to maximize the specific growth rate for given conditions of light and temperature. Laboratory and field data are compiled from the literature to test the formulae presented here.     

Light absorption,photosynthesis and growth of Nannochloris atomus in nutrient-saturated cultures

Nannochloris atomus was maintained in exponential growth at photon flux densities (PFD) from 400 to 700 nm, ranging from 10 to 200 mol m^-2 s^-1. Growth was lightsaturated at PFDs in excess of 100 mol m^-2 s^-1, with a mean light-saturated growth rate at 23 °C of 1.5×10^-5s^-1 (1.2 d^-1). The light-limited growth rates extrapolated to a compensation PFD for growth that was not significantly different from zero, although no changes in cell numbers were observed in a single culture incubated at a PFD of 1.0 mol m^-2s^-1. Dark-respiration rates were independent of PFD, averaging 1.7×10^-6 mol O₂ mol^-1 C s^-1 (0.14 mol O₂ mol^-1 C d^-1). The maximum photon (quantum) efficiency of photosynthesis was also independent of PFD, with a mean value of 0.12 mol O₂ mol^-1 photon. The chlorophyll a-specific light absorption cross-section ranged from 3 to 6×10^-3 m² mg^-1 chl a and was lowest at low PFDs due to intracellular self-shading of pigments associated with high cell-chlorophyll a contents. The C:chl a ratio increased from 10 to 40 mg C mg^-1 chl a between PFDs of 14 and 200 mol m^-2 s^-1. These new observations for N. atomus are compared with our previous observations for the diatom Phaeodactylum tricornutum in terms of an energy budget for microalgal growth.     

Primary production as influenced by diel periodicity of phytoplankton photosynthesis

Diel periodicity in parameters of photosynthesis-irradiance (P-I) curves was incorporated into calculations of integral daily phytoplankton production for the Santa Barbara Channel off southern California (USA). Model equations of the relationship between photosynthesis and light were used in combination with observed slope () and asymptote (P _max) values presented in the preceding paper. Primary production was always 19 to 39% less than comparable estimates obtained with the assumption of constant maximum daily and P _max values. Regardless of which P-I formulation was used of 6 tested, observed production (using a temporal series of simulated in situ incubations) ranged from 13% less to 25% more than estimates from constant midday and P _max values. The amplitude and timing of diel oscillations differed somewhat among 3 field stations. Maximum to minimum ratios ranged from approximately 3 to 5 for , and 4 to 6 for P _max. The differences in amplitude and timing of oscillations in P-I curves both contributed to errors in calculating phytoplankton production. Thus, photosynthetic periodicity in the upwelling area of the Santa Barbara Channel influences phytoplankton production. There were oscillations in both and P _max, and the time-dependence of these parameters should be considered to improve the accuracy of predictive models of primary productivity.     

Effects of salinity and possible interactions with temperature and pH on growth and photosynthesis of <Emphasis Type="Italic">Halophila johnsonii</Emphasis> Eiseman

The effects of salinity, temperature, and pH variations on growth, survival, and photosynthetic rates of the seagrass Halophila johnsonii Eiseman were examined. Growth and survival responses to salinity were characterized by aquarium experiments in which plants were exposed to seven different salinity treatments (0, 10, 20, 30, 40, 50, and 60 psu) during 15 days. Photosynthetic behavior was assessed for short-term salinity exposures (1 or 20 h) by incubation experiments in biological oxygen demand (BOD) bottles and by measuring photosynthesis versus irradiance (PI) responses in an oxygen electrode chamber. In the bottle experiments the possible effects of interactions between salinity and temperature (15, 25, and 35°C) or pH (5, 6, 7, and 8.2) were also examined. Growth and survival of H. johnsonii were significantly affected by salinity, with maximum rates obtained at 30 psu. Salinity also altered the parameters of the PI curves. Light-saturated photosynthesis (P _max) and the photosynthetic efficiency at subsaturating light (α) increased significantly up to an optimum of 40 psu, decreasing again at the highest salinities. Dark respiration rates and compensating irradiance (I _c) showed minimum values at 40 and 50 psu, while light-saturation point (I _k) was maximum at 30–50 psu. An interaction between salinity and temperature was not found although an increase of temperature alone produced an increase in α, P _max, respiration rates, and I _k. An interaction between salinity and pH was only found in the P _max response: P _max increased with pH=5 at 30 psu. In addition, reducing the pH increased α significantly. In the BOD bottles experiment a significant reduction in the dark respiration with decreasing pH was observed, but the opposite trend was observed in the photosynthetic rate. These results suggest that the endemic seagrass H. johnsonii could be negatively affected by hypo- or hypersalinity conditions, although salinity changes did not seem to alter the tolerance of this species to other environmental factors, such as temperature or pH.     

In vivo and in vitro differences in chloroplast functionality in the two north Atlantic sacoglossans (Gastropoda,Opisthobranchia) <Emphasis Type="Italic">Placida dendritica</Emphasis> and <Emphasis Type="Italic">Elysia viridis</Emphasis>

The photosynthetic functionality in chloroplasts in the two sacoglossan molluscs Placida dendritica and Elysia viridis from the Trondheim fjord in Norway was studied. P. dendritica and E. viridis with no functional chloroplasts in their digestive system were introduced to the green macroalgae Codium fragile. Our results showed that P. dendritica was not able to retain functional (photosynthetic) chloroplasts. Transmission electron microscopy (TEM) showed that chloroplasts were directly digested when phagocytosed into the digestive cells. Four stages of chloroplast degradation were observed. A corresponding operational quantum yield of chl a fluorescence (Φ_PSII ~ 0) indicated autofluorescence, and the presence of highly degraded chl a supported these observations. In contrast, E. viridis was able to retain functional chloroplasts. For this species it took only 1 week for the chloroplasts inside the digestive cells to acquire the same Φ_PSII and light utilisation coefficient (α) as C. fragile kept under the same light conditions. Data for 8 days showed a 2–6-fold increase in the maximum photosynthetic rate (P _max) and light saturation index (E _k) relative to C. fragile. This increase in available light was probably caused by a reduced package effect in the digestive gland of E. viridis relative to C. fragile, resulting in a partial photoacclimation response by reducing the turnover time of electrons (τ). Isolated pigments from C. fragile compared to E. viridis showed the same levels of photosynthetic pigments (chl a and b, neoxanthin, violaxanthin, siphonaxanthin, siphonein and β,ε-carotene) relative to μg chl a (w:w), indicating that the chloroplasts in E. viridis did not synthesise any new pigments. After 73 days of starvation, it was estimated that chloroplasts in E. viridis were able to stay photosynthetic 5–9 months relative to the size of the slugs, corresponding to an RFC of level 8 (a retention ability to retain functional chloroplasts (RFC) for more than 3 months). The reduction in Φ_PSII, P _max and α as a function of time was caused by a reduction in chloroplast health and number (chloroplast thylakoid membranes and PSII are degraded). These observations therefore conclude that chloroplasts from C. fragile cannot divide or synthesise new pigments when retained by E. viridis, but are able to partially photoacclimate by decreasing τ as a response to more light. This study also points to the importance of siphonaxanthin and siphonein as chemotaxonomic markers for the identification of algal sources of functional chloroplasts.     

Diel oscillations of the photosynthesis-irradiance (P-I) relationship in natural assemblages of phytoplankton

Diel oscillations in the photosynthesis-irradiance (P-I) relationship are described for marine phytoplankton assemblages at 6 stations in an upwelling area off the southern California coast (USA) between May and August 1980. The initial slope () and asymptote (P _max) of P-I curves changed significantly over the day; both parameters were in phase and had similar changes in amplitude. The diel oscillations in photosynthesis appeared unrelated to changes in chlorophyll a concentrations. Amplitudes of daily variations in photosynthesis ranged from approximately 3 to 9, as measured by the maximum to minimum ratio for photosynthetic capacity (P _max). Diatom-rich samples collected during an upwelling event and those dominated by dinoflagellates both had midday to early afternoon maxima in and P _max. Samples from other locations had peak photosynthetic activity later in the afternoon. The relationship between and P _max was consistent in all phytoplankton samples analyzed, with a surprisingly high correlation considering the spatial and temporal scales encompassed in this study. These results indicate that the photosynthesis-irradiance (P-I) relationship is time-dependent and, moreover, that changes in and P _max are closely coupled for a variety of natural phytoplankton assemblages.     

Response of zooxanthellae in symbiosis with the Mediterranean corals Cladocora caespitosa and Oculina patagonica to elevated temperatures

Scleractinian symbiotic corals living in the Ligurian Sea (NW Mediterranean Sea) have experienced warm summers during the last decade, with temperatures rapidly increasing, within a few days, to 3–4°C above the mean value of 24°C. The effect of elevated temperatures on the photosynthetic efficiency of zooxanthellae in symbiosis with temperate corals has not been well investigated. In this study, the corals, Cladocora caespitosa and Oculina patagonica were collected in the Ligurian Sea (44°N, 9°E), maintained during 2 weeks at the mean summer temperature of 24°C and then exposed during 48 h to temperatures of 24 (control), 27, 29 and 32°C. Chlorophyll (chl) fluorescence parameters [F _v/F _m, electron transport rate (ETR), non-photochemical quenching (NPQ)] were measured using pulse amplitude modulated (PAM) fluorimetry before, during the thermal increase, and after 1 and 7 days of recovery (corals maintained at 24°C). Zooxanthellae showed a broad tolerance to temperature increase, since their density remained unchanged and there was no significant reduction in their maximum quantum yield (F _v/F _m) or ETR up to 29°C. This temperature corresponded to a 5°C increase compared to the mean summer temperature (24°C) in the Ligurian Sea. At 32°C, there was a significant decrease in chl contents for both corals. This decrease was due to a reduction in the chl/zooxanthellae content. For C. caespitosa, there was also a decrease in ETR_max, not associated with a change in F _v/F _m or in the non-photochemical quenching (NPQ); for O. patagonica, both ETR_max and F _v/F _m significantly decreased, and NPQ_max showed a significant increase. Damages to the photosystem II appeared to be reversible in both corals, since F _v/F _m values returned to normal after 1 day at 24°C. Zooxanthellae in symbiosis with the Mediterranean corals investigated can therefore be considered as resistant to short-term increases in temperature, even well above the maximum temperatures experienced by these corals in summer.     

Chlorophyll fluorescence measures of seagrasses <Emphasis Type="Italic">Halophila ovalis</Emphasis> and <Emphasis Type="Italic">Zostera capricorni</Emphasis> reveal differences in response to experimental shading

In coastal waters and estuaries, seagrass meadows are often subject to light deprivation over short time scales (days to weeks) in response to increased turbidity from anthropogenic disturbances. Seagrasses may exhibit negative physiological responses to light deprivation and suffer stress, or tolerate such stresses through photo-adaptation of physiological processes allowing more efficient use of low light. Pulse Amplitude Modulated (PAM) fluorometery has been used to rapidly assess changes in photosynthetic responses along in situ gradients in light. In this study, however, light is experimentally manipulated in the field to examine the photosynthesis of Halophila ovalis and Zostera capricorni. We aimed to evaluate the tolerance of these seagrasses to short-term light reductions. The seagrasses were subject to four light treatments, 0, 5, 60, and 90% shading, for a period of 14 days. In both species, as shading increased the photosynthetic variables significantly (P < 0.05) decreased by up to 40% for maximum electron transport rates (ETR_max) and 70% for saturating irradiances (E_k). Photosynthetic efficiencies (α) and effective quantum yields (ΔF/Fm′) increased significantly (P < 0.05), in both species, for 90% shaded plants compared with 0% shaded plants. H. ovalis was more sensitive to 90% shading than Z. capricorni, showing greater reductions in ETR_max, indicative of a reduced photosynthetic capacity. An increase in E_k, Fm′ and ΔF/Fm′ for H. ovalis and Z. capricorni under 90% shading suggested an increase in photochemical efficiency and a more efficient use of low-photon flux, consistent with photo-acclimation to shading. Similar responses were found along a depth gradient from 0 to10 m, where depth related changes in ETR_max and E_k in H. ovalis implied a strong difference of irradiance history between depths of 0 and 5–10 m. The results suggest that H. ovalis is more vulnerable to light deprivation than Z. capricorni and that H. ovalis, at depths of 5–10 m, would be more vulnerable to light deprivation than intertidal populations. Both species showed a strong degree of photo-adaptation to light manipulation that may enable them to tolerate and adapt to short-term reductions in light. These consistent responses to changes in light suggest that photosynthetic variables can be used to rapidly assess the status of seagrasses when subjected to sudden and prolonged periods of reduced light.     

Effects of nitrate on the diurnal vertical migration,carbon to nitrogen ratio,and the photosynthetic capacity of the dinoflagellate Gymnodinium splendens

A non-thecate dinoflagellate, Gymnodinium splendens, was studied in a 12 d laboratory experiment in 2.0x0.25 m containers in which light, temperature, and nutrients could be manipulated. Under a 12 h light: 12 h dark cycle, the dinoflagellates exhibited diurnal vertical migrations, swimming downward before the dark period began and upward before the end of the dark period. This vertical migration probably involved geotaxis and a diel rhythm, as well as light-mediated behavior. The vertical distribution of nitrate affected the behavior and physiology of the dinoflagellate. When nitrate was present throughout the container, the organisms resembled those in exponential batch culture both in C:N ratios and photosynthetic capacity (P_max); moreover, they migrated to the surface during the day. In contrast, when nitrate was depleted, C:N ratios increased, P_max decreased, and the organisms formed a subsurface layer at a depth corresponding to the light level at which photosynthesis saturated. When nitrate was present only at the bottom of the tank, C:N ratios of the population decreased until similar to those of nutrient-saturated cells and P_max increased; however, the dinoflagellates behaved the same as nutrient-depleted cells, forming a subsurface layer during the light period. Field measurements revealed a migratory subsurface chlorophyll maximum layer dominated by G. splendens. It was just above the nitracline during the day, and in the nitracline during the night, which concurs with our laboratory observations.