Photosynthetic metabolism in microphytobenthos : modulation by temperature and dissolved inorganic carbon

Tese de doutoramento, Biologia (Ecofisiologia), Universidade de Lisboa, Faculdade de Ciências, 2017Main primary producers of estuarine systems include benthic microalgae, predominantly diatoms, and cyanobacteria that colonize subtidal and intertidal zones, commonly referred to as microphytobenthos (MPB). MPB communities account for about 50% of the primary productivity of estuarine systems (e.g. MacIntyre et al., 1996; Barranguet et al., 1998; Underwood and Kromkamp, 1999; Serôdio and Catarino, 2000). This is highly relevant considering that estuarine systems are among the most productive ecosystems in the biosphere (Constanza et al., 1997). Despite the extreme variability of environmental parameters (irradiance, temperature and salinity) that characterizes the estuarine habitat, these communities are able to display high photosynthetic rates. It is common to distinguish two types of MPB communities: epipelic communities (typical of muddy sediments) composed of motile species and epipsammic communities (typical of sandy sediments) composed largely by non-motile species attached to sand grains. Contrary to epipsammic, epipelic diatoms exhibit circadian and circatidal migratory rhythms (Palmer and Round, 1967; Palmer, 1973; Admiraal et al., 1982; Serôdio and Catarino, 2000). Being MPB critical for the function of estuarine and coastal ecosystems, understanding the impact of climate change on its structure and productivity is mandatory. In particular, the increase of temperature and inorganic carbon availability can markedly affect photosynthesis, diversity and productivity of estuarine MPB communities. However, studies of the impact of climate changes on MPB are still scarce, and in particular the effects of the interaction between increased temperature and inorganic carbon availability are, to a large extent, still unknown. Thus, in this project we studied the effects of temperature and dissolved inorganic carbon (DIC) availability in two MPB communities of the Tagus estuary, in central Portugal, applying new methods of remote sensing, namely laser induced fluorescence (LIF) and modulated imaging fluorescence (Imaging-PAM). The LIF methodology (chapter 2) was used to trace migratory rhythms of benthic epipelic microalgae typical of mud sediments. Indeed, it was confirmed that, during the diurnal low tide these microalgae migrate to the surface of sediment to drive photosynthesis and then migrate in depth before the arrival of the tide. Furthermore, epipelic diatoms show photophobic migration when exposed to high light levels, a process known as behavioral photoprotection. This technique proved to be adequate to the study of MPB communities, both for the determination of surface biomass and the study of migratory rhythms. The possibility of using LIF for middle-range remote sensing under natural light conditions opens good perspectives for its use in environmental monitoring of estuarine systems. In the last decades, conventional pulse amplitude modulated fluorescence (PAM) has been increasingly applied to estuarine MPB communities (Serôdio et al., 1997; Kromkamp et al., 1998; Paterson et al., 1998; Serôdio et al., 2001; Honeywill et al., 2002; Perkins et al., 2002; Serôdio, 2003; Stephens et al., 2003; Forster and Kromkamp, 2004; Murphy et al., 2004; Serôdio, 2004; Forster and Jesus, 2005; Murphy et al., 2005). Lower resolution imaging systems allow the mapping of fluorescence parameters over large areas, making it a unique technique to study the spatial heterogeneity of the photosynthetic activity across an autotrophic surface (Scholes and Rolfe, 1996; Hill et al., 2004. This can be particularly relevant in the study of MPB biofilms, characterized by a “patchy,” heterogeneous distribution and can provide useful information about the spatial distribution of biomass and productivity in sediments. Yet, important hardware differences exist between conventional and imaging systems and therefore a comparative study of the results obtained with these technologies in MPB was required (chapter 3). In fact, we observed significant differences between the results obtained with conventional and imaging systems, especially in what refers to the analysis of rapid fluorescence curves as a function of irradiance (RLCs, Rapid Light Curves). Lower values of α (initial slope of the rETR vs. E curve), ETRmax (maximum relative ETR), Ek (light saturation parameter) and Fv/Fm (maximum quantum efficiency of photosystem II of dark-adapted samples) were obtained using the Imaging- PAM system. The discrepancy between instruments was dependent on sample type, being more pronounced in mud than in sandy MPB sediments. The differences may be largely explained by the interplay between the different depth-integration of the fluorescence signal, dependent on the thickness of the photosynthetic layer, and the different attenuation coefficients of the light sources used by the two types of instrument. It is therefore important to take into account the specificity of each fluorescence system when interpreting and comparing chlorophyll fluorescence data of MPB communities, taking full advantage of the application of imaging fluorescence to the MPB communities, in particular in what refers to its unique ability to study the spatial heterogeneity of their photosynthetic activity. The in-depth understanding of the primary production processes of MPB is a prerequisite to assess the impacts of climate change on estuarine systems. Among the global change variables, increased temperature and atmospheric CO2 (and concomitantly dissolved inorganic carbon, DIC) are expected to have the most significant impact over the primary productivity in estuaries. Therefore, we studied their effect, as well as their interaction, on the photosynthesis of MPB biofilms. The short-term effect of increasing temperature in the photosynthetic activity of two communities of MPB (Alcochete and Trancão) was studied (chapter 4). The Trancão MPB community had higher photosynthetic electron transport capacity (higher ETRmax) being photoacclimated at higher irradiance (higher EK) and had lower energy conversion efficiency at limiting irradiance (lower α) .The differences in the species composition and size class may explain the results obtained for the two sites, since smaller cells are generally more metabolically active. However, for the both MPB communities photosynthetic capacity increased with temperature up to 35 °C and decreased with extreme temperature of 42 °C. Photosynthetic efficiencies at low irradiance (α) were not affected by the temperature in the 15-35 °C range, and decreased at 42 °C. These MPB estuarine communities were able to increase the photosynthetic capacity under exposure to short-term high temperatures sediments, similar to happens during the summer midday in the diurnal low tides. We also studied the effect of dissolved inorganic carbon availability (DIC) in photosynthesis, on the two intertidal MPB communities, taking as reference the model diatom species Phaeodactylum tricornutum, using fluorescence imaging (on intact MPB communities) and polarographic oxygen measurement (in cell suspensions) (chapter 5). The parameters of DIC acquisition kinetics (Vmax and Km (DIC)) estimated for MPB communities were higher ((Km (DIC) = 0.31 mM; Vmax = 7.78 nmol min-1 μg (Chl a)-1), than those obtained for Phaeodactylum tricornutum (Km (DIC) = 0.23 mM; Vmax = 4.64 nmol min-1 μg (Chl a)-1), showing that these communities have species with low affinity for DIC and high photosynthetic capacity. The net photosynthesis of MPB suspensions reached saturation at DIC concentrations of 1-1.5 mM, 17 lower than those found in the sediment top 5 mm interstitial water, suggesting that the photosynthesis of these communities is not limited by DIC availability. However, when NaHCO3 was added to intact MPB samples, an increase in the maximum photosynthetic electron transport rate (ETRmax) was observed. These results suggest that the local depletion of DIC in the photic layer of the sediment, where the microalgae accumulate during diurnal low tide, limits the photosynthetic activity of MPB, providing the first experimental evidence of DIC limitation of the productivity of intertidal MPB communities. The interactive effects of temperature and DIC in biomass, species composition and photosynthetic performance of MPB was studied using a microcosm ELSS ('experimental life support system') with tide regime (chapter 6). The longer term effect of increased temperature (24 ° C) had a negative effect on MPB biomass and photosynthetic performance under both simulated DIC conditions (pH = 7.4 and pH = 8.0). Furthermore, at elevated temperatures there was a change in the relative abundance of the major species of diatoms and an increase of cyanobacteria. The long term effect of increasing DIC ( pH = 7.4) under low temperature (18 °C) had a positive effect on biomass MPB, possibly due to decreased local depletion of dissolved inorganic carbon. No significant effects were found on the relative abundance of the major groups of microalgae. Our results suggest that the interactive effects of increased temperature and DIC availability of estuarine MPB communities can have a negative impact on the structure, diversity and productivity of these communities, eventually affecting the entire estuarine ecosystems. Therefore, a deepen understanding of the potential impacts of climate change in these complex ecosystems is required