Solar Radiation-Induced Mortality of Marine Pico-Phytoplankton in the Oligotrophic Ocean[Dagger]

Aug 11, 04:00 AM

Current Headlines: By Agusti, Susana Llabres, Moira

ABSTRACT We examined the response of pico-phytoplankton communities sampled at the equatorial, tropical and temperate Central Atlantic Ocean to subsurface underwater solar radiation in order to test the generality of the reported cell mortality for these populations when exposed to high ultra violet radiation (UVR) and photosynthetically active radiation. The natural communities of pico-phytoplankton populations tested experienced high cell mortality when exposed to high solar radiation, despite inhabiting tropical waters. Synechococcus and eukaryotes were more resistant to solar radiation than Prochlorococcus. The decay rates of all pico- phytoplankton groups examined tended to be much higher when exposed to total solar radiation than when UVB-R was filtered out. We also show that even short exposures of 30 min to high solar radiation were able to induce cell mortality in Prochlorococcus. The variability in the decay rates of living Prochlorococcus cells were strongly related to the condition of the original population. However, Synechococcus decay rates were higher in populations from the tropical area, with eukaryotes sensitivity increasing with increasing the trophic degree. The data reported in this study and in the literature revealed contrasting capacities of Prochlorococcus, Synechococcus and eukaryotes to survive under high solar radiation. Although the mechanisms involved are as yet unclear, their elucidation may help explain niche partitioning among these organisms in the ocean.

INTRODUCTION

Pico-phytoplankton, comprising the smallest photosynthetic organisms characterized by cell diameters < 2 [mu]m, dominates the biomass and primary production in the oligotrophic ocean (1), where their small size represents an advantage to grow at the characteristically extremely low nutrient concentrations (2,3). The oligotrophic ocean comprises 70% of the ocean surface, largely distributed along tropical and subtropical regions. In addition to the high incident ultraviolet radiation (UVR) at these latitudes, oligotrophic waters are highly transparent, allowing a deep penetration of visible and ultraviolet radiation (4). Pico- phytoplankton is thus likely to be exposed to high UVR.

The small size of pico-phytoplankton has been argued to be a disadvantage under high solar radiation exposure, as pico-sized organisms should have difficulties to accommodate within their tiny cells the significant quantities of sunscreen substances required to avoid radiation damage (5). Indeed, pico-phytoplankton populations have been recently reported to be highly sensitivity to solar radiation (6), with the exposure to natural levels of UVR and photosynthetically active radiation (PAR) inducing considerable cell mortality in the subtropical Atlantic Ocean. The cyanobacteria Prochlorococcus, the smallest (about 0.5 [mu]m diameter) and most abundant photosynthetic organism in the tropical ocean, appeared to be most sensitive to UVR (6,7), showing high cell mortality rates when exposed to high natural levels of underwater UVR (6). Pico- eukaryotes showed lower, although significant cell death, and cells of the cyanobacteria Synechococcus, appeared to be most resistant to solar radiation (6). Whereas pico-phytoplankton cell death increases with increasing UVR and PAR doses, there is a considerable and unexplained variability in the cell death supported by different populations at comparable UVR and PAR doses (6).

In this study, we examine the response of natural populations of the cyanobacteria Prochlorococcus and Synechococcus, and pico- eukaryotes, the organisms forming pico-phytoplanktonic communities in the Central Atlantic Ocean, to solar radiation to test the generality of the reported cell mortality in response to exposure to ambient UVR and PAR levels. We do so through the meta-analysis of a series of experiments run with natural communities of pico- phytoplankton from the equatorial, tropical and temperate Central Atlantic Ocean exposed to natural increased levels of underwater solar radiation, and compare the results with those reported previously for communities in the Northern East Atlantic (6) and Mediterranean Sea. For the most sensitive species, we test whether cell mortality can be induced after short (minutes) exposures to high natural UVR. We examine whether the differences in pico- phytoplankton mortality can be explained by differences in ambient environmental conditions and the extent of stress the communities experienced in situ. We also discuss the consequences of UVR- induced mortality of pico-phytoplankton for the functioning of the oligotrophic ocean.

MATERIALS AND METHODS

Experiments with natural communities of plankton were performed during the cruises LATITUDE-3 and LATITUDE-4, on board the Spanish R/ V BIO Hesperides. LATITUDE-3 started at Las Palmas de Gran Canaria on November 1, 1999 and ended in Montevideo on November 20, 1999. The cruise LATITUDE-4 started in Rio de Janeiro on March 16, 2000, and ended on April 4, 2000, at the Canary archipelago. A total of nine stations distributed along the South and North Atlantic Subtropical Gyre, the Interequatorial Zone and the South and North temperate Atlantic were sampled for the experiments (Table 1). Station sampling started at 9:00 A.M. (local time) with CTD casts down to 500 m, along the cruises. Seawater samples were collected in Niskin bottles attached to a Rossette-CTD system from 5 m depth and used in experiments performed in Stations #5, #7, #9, #12 and #14 (LATITUDE-3) and Stations #8 and #15 (LATITUDE-4) and from 30 m depth for experiments performed in Stations #6 and #10 (LATITUDE- 4).

The depth of the upper mixed layer (UML), an index of the stability of the water column, was calculated from the CTD profiles as the shallowest depth at which s^sub t^ (water density) differs from surface values by more than 0.05 kg m^sup -3^ (8).

Underwater PAR was measured at each station during LATITUDE-4 from the surface to 200 m depth, by using a 4pi underwater quantum sensor (LI-192SA) attached to a SATLANTIC-OCP 100 automatic profiler, as described in Agusti (9). Light conditions in the experiments were tested by measuring underwater levels of PAR in the incubators at noon by using a 4pi underwater quantum sensor (LI- 192SA). The levels of irradiance to which the communities were exposed during the experiments were equivalent to that reaching 0.5- 1.5 m depth at the stations, as indicated by the water profiles. Incident solar radiation was measured and automatically recorded every 10 min during the two cruises using a Radiation Sensor 2770 (Aandera Instruments) installed in a meteorological station placed over the bridge deck. The sensor is sensitive in the range 300-2500 nm, but is covered by a glass (borosilicate) dome, filtering out most incident UVB radiation. Doses of incident solar radiation, in J cm^sup -2^, received by planktonic communities during the different experiments were calculated by integrating the radiation values for the duration of each experiment.

The experiments consisted of incubating seawater samples in duplicate quartz, glass and black bottles (100 mL) submersed underwater (0.5 m) in open-air tank incubators of 1000 L placed on deck in an area free of shadows, receiving solar radiation, and with sea surface recirculating water to maintain the in situ temperature. Water samples were incubated shortly after sampling (i.e. half an hour) with experiments starting at 10:00 to 10:30 A.M. (local time). The total duration of the experiments was 7-8 h and duplicated samples to determine abundance of total and living cells were taken from quartz and glass treatments every 2-4 h, depending on the experiments. The experiments run with communities from Stations #10 and #15 were also sampled after the first 30 min of exposure. Experiment from Station #15 lasted 3 h. Dark controls were sampled at the beginning and at the end of the experiment. Quartz bottles allowed all the radiation (UVR + PAR) to pass through, while glass bottles were borosilicate that filtered most UVB-R with 50% transmission at 313 nm.

The variability in the abundance of Prochlorococcus, Synechococcus and pico-photosynthetic eukaryotic cells during the experiments was determined on board in replicated fresh subsamples from each replicated treatment bottle by using a FACSCALIBUR (Becton- Dickinson) flow cytometer, fitted with a 488 nm laser and a photomultiplier for forward-scattered light detection. An aliquot of a calibrated solution of 1-[mu]m-diameter high-green fluorescent beads (Polysciences, Inc.) added to the samples was used as an internal standard for the quantification of cell concentration. Beads concentration in the standard solution was calculated by filtering replicated aliquots onto black nuclepore filters and counting the beads under an epifluorescence microscope. The red, green and orange fluorescence emissions and the forward and side scattering of the cells and beads were used to detect different cell populations and to differentiate them from the fluorescent beads.

Table 1. Cruise, position and general characteristics of the stations sampled for the experiments reported.

The abundance of living pico-cyanobacteria cells was analyzed throughout the experiments at the beginning and in all the sampling intervals using a cell membrane permeability test (cell digestion assay, 10) as modified for phytoplankton cells by Agusti and Sanchez (11). The cell digestion assay is based on the brief exposure of the cells to the enzymes trypsin and DNAse I, which enter the cytoplasm of cells with damaged plasma membranes (i.e. necrotic or advanced apoptotic cells) resulting in the entire digestion of the cells while having little or no effect on the viability, morphology or function of live cells (11). The digestion of dead cells is based on the fragmentation and hydrolysis of DNA by DNAse I, and peptide hydrolysis by trypsin, which penetrates the damaged cells. The digested dead cells are undetectable by optical observation and lose any fluorescence signals (both autofluorescence and fluorescence from staining), and are therefore effectively removed from the population (11). Stock solutions of DNAse I (400 [mu]g mL^sup 1^ in Hank's Balanced Salt Solution [HBSS]) and trypsin (1% in HBSS) were prepared in HBSS medium (Sigma Co.) and kept frozen at - 65[degrees]C until use. Viability tests by the cell digestion assay were made on fresh samples by adding 200 [mu]L of DNAse I stock solution to replicated 1 mL of seawater samples followed by incubation for 15 min at 36[degrees]C. Then, 200 [mu]L of trypsin solution was added, followed by incubation for an additional 30 min at 35[degrees]C (11). At the end of the incubation, the samples were placed on ice to stop enzyme reaction and the abundance of pico- phytoplanktonic cells remaining after the assay was quantified by using a flow cytometer, as described above.

The cells remaining after the enzymatic treatment, i.e. those having intact membranes, were considered to represent living or viable cells, whereas dead ones, with compromised membranes, were digested by the enzymatic cocktail and were undetectable by the flow cytometer. The fraction of living pico-cyanobacteria cells in the samples was calculated by dividing the concentration of living cells after the enzyme treatment by the cell concentration in the untreated sample, which represent the total (dead plus living) cell concentration.

Published data of pico-phytoplankton cell death induced by UVR and PAR were collected and used in the analysis. Only data of decay rates of living Prochlorococcus, Synechococcus and Eukaryotic cells corresponding to communities exposed to 100% subsurface total solar radiation (incubated in quartz bottles) were used. Data from the Mediterranean Sea (M. Llabres, S. Agusti, P. Alonso-Laita, G. H. Hendl, unpublished) were used in the analysis.

RESULTS

The pico-phytoplankton communities examined were sampled from waters covering a large geographical area, including communities sampled in the temperate, tropical and intertropical areas of the Atlantic Ocean (Table 1). All the waters sampled were warm, with temperatures ranging from 19.7[degrees]C to 28.55[degrees]C (Table 1). The chlorophyll a (Chl a) concentration was low throughout, as expected from these largely oligotrophic areas of the ocean, and the depth of the mixed layer (UML) varied considerably across the study area (Table 1). Pico-phytoplankton populations of the cyanobacteria Prochlorococcus and Synechococcus and eukaryotic aryotic pico- phytoplankton cells were always present in the communities sampled. Doses of incident solar radiation received during the experiments varied from 1615 to 2615 J cm^sup -2^ (Table 1).

Pico-phytoplankton communities exposed to subsurface natural solar radiation in the experiments generally experienced enced a decline in the total abundance and that of living cells of Prochlorococcus, Synechococcus and eukaryotic pico-phytoplankton toplankton (Fig. 1), indicative of cell death induction. The communities growing in dark controls, however, did not show a significant decrease in the abundance of the pico-phytoplankton plankton populations, showing values of living cells close to those quantified at the beginning of the experiment. Prochlorococcus rococcus living cells abundance in the experiment of Station #10 decreased from 1.45 x 10^sup 5^ +- 5300 cells mL^sup -1^ at the initial time to 6.1 x 10^sup 3^ +- cells mL^sup -1^ in the quartz bottles, although 1.58 x 10^sup 5^ +- 4100 living cells mL^sup - 1^, similar to initial values (P = 0.86), were quantified in the dark controls at the end of the experiment indicating that the decline in the population was induced by solar radiation. In the light treatments, the decline was most pronounced, indicating greater cell mortality, in the populations of Prochlorococcus (Fig. 1), which showed decay rates of living cells varying from 0 to 0.39 h^sup -1^, greater than those observed for Synechococcus and for eukaryotic pico-phytoplankton (P < 0.05), which varied from 0 to 0.14 h^sup -1^ and from 0 to 0.16 h^sup -1^, respectively (Fig. 1), showing Synechococcus and eukaryotes similar averaged decay rates (P = 0.96). The decline of living cells were lower when UVB-R was filtered out (Fig. 1). The decay rates observed when UVB-R was filtered out were also higher for Prochlorococcus populations, than for the other two pico-phytoplankton groups.

The experiments from Stations #6 and #10, which were performed with communities from 30 m depth instead of those from surface waters (5 m) as performed in all other stations, showed a much steeper decline in the total abundance of Prochlorococcus and that of living cells than that observed in the experiments using surface water (Fig. 1). Prochlorococcus showed the highest decay rates of 0.42 and 0.65 h^sup -1^ in the experiments with populations sampled from 30 m depth, more than those observed for populations from surface water (Fig. 1) (P < 0.05). Synechococcus and eukaryotes showed decay rates similar to those observed in experiments with surface water populations (P = 0.88).

On average, Prochlorococcus was the most sensitive group to total solar radiation, as indicated by the high average decay rate of 0.178 +- 0.055 h^sup -1^ (mean +- SE) which represented an average half-life of 3.8 h when exposed to subsurface total solar radiation, significantly higher than those observed for Synechococcus and Eukaryotes (P < 0.05). Eukaryotes and Synechococcus showed averaged decay rates an order of magnitude lower (0.077 +- 0.038 and 0.075 +- 0.023 h-1, mean +- SE, for eukaryotes and Synechococcus, respectively), representing longer and similar (P = 0.89) average half-lives of 8.9 and 9.18 h, respectively. The induction of Prochlorococcus cell death was reduced when filtering out UVB-R, resulting in a more extended average half-life of 8.64 h (P < 0.05). Similar results were obtained for eukaryotes and Synechococcus, for which average half-lives after filtering out UVB-R increased to 14.6 and 22.8 h, respectively, although differences were not significant (P > 0.05). Indeed, the decay rates of all pico-phytoplankton groups examined tended to be much higher when exposed to total solar radiation than when UVB-R was filtered out, as reflected in the consistent departures from the 1:1 line in the relationship between decay rates observed under both treatments (Fig. 2).

Sampling at shorter time intervals was performed in two of the experiments (Fig. 3) where samples were taken after 30 min of exposition to total solar radiation. A decay in the abundance of living cells and in the abundance of the total population was detected after 30 min in Prochlorococcus populations, parallel to an increase in the proportion of dead cells in the experiment conducted at Station #10 (Fig. 3). In the experiment with communities from Station #15, the total abundance of Prochlorococcus cells and that of living cells declined, but the proportion of dead cells decreased after 30 min of exposition to total solar radiation (Fig. 3), indicating that dead cells had already lysed within 30 min from the onset of exposure.

A comparison across experiments also showed that the pico- phytoplankton cell mortality rates induced by total solar radiation were high among communities (Figs. 1 and 3). The variability in the decay rates of living cells did not show a clear latitudinal trend for Prochlorococcus and eukaryotes. However, the lower decay rates for Synechococcus cells were observed at the experiments performed with populations from the Stations #14 and #15 located in the temperate area of the South and North Atlantic, respectively (Fig. 4), and the highest decay rates of Synechococcus cells were observed at the experiments performed with populations sampled from the equatorial and the intertropical regions (Fig. 4).

There was no significant relationship between the mortality rates induced by solar radiation in the three pico-phytoplanktonic groups and the depth of the UML or the light extinction coefficient at the different stations (R^sup 2^ < 0.15, P > 0.20). For the eukaryotic pico-phytoplankton we observed, however, a strong negative relationship between the decay rates of living cells induced by total solar radiation and the Chl a concentration in the waters sampled (R^sup 2^ = 0.73. P < 0.001), whereas there was no such relationship for the other two pico-phytoplankton groups (P > 0.05). For Prochlorococcus, however, the variability in the decay rates of living cells was strongly related to the condition of the original population from subsurface (5 m) waters, as represented by the percentage of living cells found in the population at the onset of the experiments (R^sup 2^ = 0.77, P < 0.001).

Figure 1. The decline in the abundance of living cells of natural populations of the cyanobacteria Prochlorococcus and Synechococcus and picoeukaryotes with time from the exposure to subsurface underwater solar radiation. The different lines represent the fitted linear regressions obtained for each experiment. (a) Experiments performed with surface (5 m) communities exposed to total solar radiation (UVR + PAR). (b) Experiments performed with communities sampled at 30 m depth and exposed to total solar radiation (UVR + PAR). (c) Experiments performed with surface (5 m) communities exposed to solar radiation filtered to exclude UVB-R. (d) Experiments performed with communities sampled at 30 m depth and exposed to solar radiation filtered to exclude UVB-R. Figure 2. The relationship between cell decay rates of the cyanobacteria Prochlorococcus and Synechococcus and pico-eukaryotes exposed to total solar radiation and to solar radiation filtered to exclude UVB- R. The solid line shows the 1:1 line expected if decay rates were similar in the presence and absence of UVB-R.

We extended the analysis of the variability in pico- phytoplankton cell death by total solar radiation by including comparable results reported in the literature (Table 2). The decay rates of Prochlorococcus living cells induced by total solar radiation collected from the literature varied from 0.05 to 0.4 h^sup -1^ (Table 2), similar to the range observed in this study (Table 2). For Synechococcus and eukaryotes, however, literature data represented lower cell mortality rates when exposed to high irradiance than that reported in this study, with maximum decay rates of living cells of 0.065 h^sup -1^ reported for both Synechococcus and eukaryotes in the literature, well below the maximum decay rates induced by exposure to total solar radiation in the Atlantic populations examined in this study (Table 2). Literature data, however, extended the geographical origin of the populations and the range of variability of the water properties at the stations sampled (Table 2).

By including the data from the literature in the analysis, we confirmed the independence of the decay rates of living cells of the three pico-phytoplankton groups of the depth of the UML and the extinction coefficient. Also, the relationship between the decay rates of eukaryotes and the Chl a concentration in our dataset disappeared when data from other areas were included in the analysis (R^sup 2^ = 0.002, P > 0.05). Yet, inclusion of the literature data confirmed the strong and positive relationship between the decay rates of living cells and the condition of the population, as indicated by the percentage of living Prochlorococcus cells at the onset of the experiments (Fig. 5, R^sup 2^ = 0.62, P < 0.001).

DISCUSSION

Recent reports showed experimentally that natural levels of underwater UVR and PAR can induce considerable cell death in natural pico-phytoplankton communities (6) helping to explain the considerable phytoplankton cell death and lysis reported recently for the oligotrophic ocean (12,13). A high proportion of dying cells within pico-phytoplankton communities has also been found to be prevalent in oligotrophic waters (9,11). Nutrient deficiency, virus infection and low light have been identified as the factors causing phytoplankton cell death in the ocean (12-16). Recent reports, however, identified the penetration of high doses of UVR and PAR as an important stressor for pico-phytoplankton, explaining the vertical distribution of dead pico-phytoplankton cells in the Central Atlantic (9). The findings that pico-phytoplankton populations are highly sensitive to UVR (6) support the hypothesis that solar radiation could be an important factor determining the variability of phytoplankton cell death in the ocean.

The results obtained in the present study further show that the high sensitivity of pico-phytoplankton to ambient levels of solar radiation described previously (6) appear to be a general phenomenon. Natural populations of Prochlorococcus, Synechococcus and eukaryotes from several locations from the Atlantic Ocean declined when exposed to full solar radiation. The decay rates induced by solar radiation during the experiments were similar to those reported for North East Atlantic pico-phytoplankton exposed to high levels of solar radiation (100% and 50% light treatments of Llabres and Agusti [6]) and to the rates reported for populations from the Mediterranean Sea (Llabres et al., submitted). The lethal effect of solar radiation resulted in half-lives of 3 h for Prochlorococcus, similar to those reported previously for other natural populations of this taxa (6,8). Synechococcus and eukaryotes were more resistant to solar radiation than Prochlorococcus, as indicated by the lower decay rates and half-lives obtained. Half- lives of Prochlorococcus increased considerably to values around 8 h when exposed to the treatment where UVB-R was filtered out, indicating that UVB-R is a major factor in causing cell death in these organisms (6,7); similarly, Synechococcus and eukaryotes cell death was considerably reduced when UVB-R was screened as observed (6,7).

Figure 3. (a) Decay with time in the abundance of the total population (continuous line) and living Prochlorococcus cells (dotted line) observed after 30 min and 2-3 h of exposure to total solar radiation in the experiments from Stations #10 and #15. (b) and (c) Changes in the percentage of Prochlorococcus dead cells at the sampling intervals shown in (a).

Figure 4. The latitudinal distribution of decay rates (of living cells) of Atlantic Synechococcus populations when exposed to subsurface total solar radiation. The latitude values corresponded to the position where populations were sampled and exposed to subsurface underwater solar radiation. Solid circles represent data from experiments performed during this study, squares represent literature data from similar experiments performed with Atlantic Synechococcus populations (6). The line represents the tendency by fitting a polynomial model.

We showed that even short-time exposures of 30 min to 100% UVR and PAR were able to induce cell death in Prochlorococcus. the most sensitive taxa. The results of short time exposures also suggested that high UVR and PAR could also trigger the lysis of the cells that were already dead within the populations examined. The demonstration of considerable damage to Prochlorococcus cells upon short-term, 30 min, exposure to high UVR and PAR does not only highlight the sensitivity of these organisms to high solar radiation but also implies that extreme care should be taken to avoid exposure of samples containing this organism to direct sunlight during manipulation in cruises and experimental research.

Dying cells were identified by using a membrane permeability test (cell digestion assay, 11), as dead cells are characterized by increased membrane permeability (10,17). UVR has been described to induce damage in cell membranes (18,19), suggesting that the exposure of dead Prochlorococcus cells with compromised membranes to UVR has triggered the lysis of the dying cells by implementing damage of the already injured cell membranes. However, this suggested phenomena should require a more specific study to be analyzed throughout.

We found considerable differences in sensitivity to ambient levels of UVR among the three taxa and among experiments, ranging from no cell death to fast cell death rates upon exposure to high ambient UVR levels. It is remarkable, however, that the decay rates of Prochlorococcus, Synechococcus and eukaryotes induced by solar radiation varied independently, despite the three populations had been exposed to similar doses during the experiments. This indicated contrasting capacities of these taxa to overcome the cell damage induced by UVR and PAR (20). which vary strongly among the three pico-phytoplankton groups. Synechococcus has been identified to be the most resistant to UVR and PAR of the three pico-phytoplankton groups (6,7). This assessment was supported by our experiments, where Synechococcus showed the smallest decay rates and the longer half-lives, with the differences with the other taxa being particularly large when UVR was filtered out, indicating Synechococcus cells to be highly resistant to high PAR levels. A variety of specific mechanisms of Synechococcus to overcome UVR damage have been reported (21-23), which have not been detected for the other two groups. The cell decay rates of Synechococcus induced by UVR and PAR were found to vary with the geographical origin of the population, with the highest cell death rates induced in populations from the equatorial zone. This may signal the existence of differences between Synechococcus strains in their capacity for photoprotection and repairing damages induced by UVR, a possibility that should be tested in future studies.

Table 2. Reported decay rates (mu, h^sup -1^) of living cells in natural communities of pico-phytoplankton exposed to increased levels of underwater total solar radiation (UVR + PAR) from a variety of experiments reported in the literature.

Figure 5. The relationship between the decay rates of Prochlorococcus living cells when exposed to natural subsurface levels of total solar radiation and the percentage of living cells in the sampled populations at the onset of the experiments. Solid circles represent data from this study and squares represent data from the literature (M. Llabres. S. Agusti, P. Alonso-Laita, G.H. Hendl, unpublished).

The variability in the decay rates of eukaryotic cells induced by exposure to high natural levels of UVR and PAR between experiments was strongly related, as indicated by regression analysis, to the Chl a concentration in the original sample, with lower pico- eukaryotic cell death induced by exposure to solar radiation in the populations from waters with sparse phytoplankton communities. Pico- eukaryotes are very small organisms whose taxonomic identification by traditional methods is cumbersome, but which are represented in a broad variety of taxonomic groups of microalgae (24). Despite the absence of taxonomic information on the species included in each community tested here, the results suggest that the eukaryotic communities sampled at the most eutrophic waters have developed more efficient reparation or photoprotective systems against UVR exposition than the communities from the less oligotrophic waters sampled did. However, the relationship between pico-eukaryote cell death rates induced by exposure to solar radiation and Chl a concentration did not fit when the literature data were included. This lack of fit was, however, attributable to the inclusion of experimental results from the community growing in the area influenced by the NW African upwelling (experiment from Station #66, 6), which represents a far more eutrophic environment than that in the other experiments. Pico-eukaryotes in these productive environments may differ from those in oligotrophic waters and the capacity to resist solar radiation and repair-associated damages may also be affected by resource availability (20). Moreover, bacterial populations differ in their capacity to repair DNA damage and different systems have been described among phytoplanktonic groups (20) with photoreactivation as a DNA repair mechanism in some species of microalgae (24,25), other groups presenting nucleotide excision repair pathway or both systems (25,26). Contrary to those observed for the other two pico-phytoplankton groups, differences in Prochlorococcus cell death induced by solar radiation among experiments were explained by the variability in the percentage of living cells found in the source populations. Populations including a low proportion of living cells, indicative that these populations were subject to high stress in situ, were less resistant to UVR than Prochlorococcus populations composed of a high proportion of healthy, living cells. This result is in agreement with previous findings indicating that different DNA repair efficiencies could strongly depend on the initial level of damage (25,26).

Prochlorococcus showed the highest sensitivity to solar radiation among the pico-phytoplankters, and their high cell mortality induced by solar radiation has been attributed by Llabres and Agusti (6) to its very small size (0.3 [mu]m cell radius) that is below the theoretical threshold size needed to accommodate sunscreen substances in its tiny cells (5). Prochlorococcus, the smallest among oxygenic photosynthetic organisms, is characterized by the smallest genome known for any one organism (27,28), which should limit its capacity to synthesize reparatory and photoprotective systems, which would compound with the absence of important genes for DNA repair (22), rendering Prochlorococcus highly vulnerable to damage. The highest cell death induced in Prochlorococcus was observed, however, in the experiments performed with populations sampled from deeper layers (30 m depth) and exposed to subsurface solar radiation, indicating that the surface populations tested in most of the experiments were more resistant than those growing deeper. This suggests that, despite the strong limitations of Prochlorococcus to overcome cell injuries induced by UVR and high PAR described, surface populations exposed to high solar radiation have developed higher resistance to UVR and PAR than those in deeper layers, indicative of some adaptive capacity to high solar radiation in Prochlorococcus that should be examined further.

Solar ultraviolet radiation (UVR; 280-400 nm) is increasingly recognized to exert a major influence on biological and chemical processes in the aquatic environment, including the growth and production of phytoplankton (29-32). There is increasing evidence that pico-phytoplankton, the primary producers dominating the extensive oligotrophic areas of the ocean, experience high cell death due to exposure to natural underwater levels of solar radiation corresponding to those in the upper meters of the water column. The small size of these organisms is likely a primary factor limiting their capacity to develop adequate levels of photoprotection and repair systems, and, in consequence, rendering these organisms highly vulnerable to high ambient irradiance levels, which causes pico-phytoplanktonic cell death. The meta-analysis of a broad set of experimental data reported here and the literature revealed contrasting capacities of Prochlorococcus, Synechococcus and eukaryotes to survive high solar radiation. The mechanisms involved, genetic, physiological and cellular, are as yet unclear, but their elucidation may help explain niche partitioning among these organisms in the ocean.

Acknowledgements-This work was supported by the project LATITUDE (MAR98-1676E) and the project RODA (CTM2004-06842-CO3-02) funded by the Spanish Ministry of Education and Science to S.A. M.L. was supported by the EC project THRESHOLDS. We thank the technical UTM personnel and the BIO Hesperidos crew for their professional assistance and help during the cruise. We are grateful to R. Martinez, S. Loureiro, B. Casas and A. Orfila for sampling assistance and to C. M. Duarte for valuable comments on the manuscript.

[dagger] This invited paper is part of the Symposium-in-Print: UV Effects in Aquatic and Terrestrial Environments.

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Susana Agusti* and Moira Llabres

Instituto Mediterraneo de Estudios Avanzados (IMEDEA), CSIC- UIB, Esporles, Balearic Islands, Spain

Received 30 August 2006; accepted 12 March 2007; DOI: 10.1111/ J.1751-1097.2007.00144.x

* Corresponding author email: sagusti@uib.es (Susana Agusti)

(c) 2007 The Authors. Journal Compilation. The American Society of Photobiology 0031-8655/07

Copyright American Society for Photobiology Jul/Aug 2007

(c) 2007 Photochemistry and Photobiology. Provided by ProQuest Information and Learning. All rights Reserved.