Kristian Spilling

Decadal-scale changes of dinoflagellates and diatoms in the anomalous baltic sea spring bloom.

The algal spring bloom in the Baltic Sea represents an anomaly from the winter-spring bloom patterns worldwide in terms of frequent and recurring dominance of dinoflagellates over diatoms. Analysis of approximately 3500 spring bloom samples from the Baltic Sea monitoring programs revealed (i) that within the major basins the proportion of dinoflagellates varied from 0.1 (Kattegat) to >0.8 (central Baltic Proper), and (ii) substantial shifts (e.g. from 0.2 to 0.6 in the Gulf of Finland) in the dinoflagellate proportion over four decades. During a recent decade (1995-2004) the proportion of dinoflagellates increased relative to diatoms mostly in the northernmost basins (Gulf of Bothnia, from 0.1 to 0.4) and in the Gulf of Finland, (0.4 to 0.6) which are typically ice-covered areas. We hypothesize that in coastal areas a specific sequence of seasonal events, involving wintertime mixing and resuspension of benthic cysts, followed by proliferation in stratified thin layers under melting ice, favors successful seeding and accumulation of dense dinoflagellate populations over diatoms. This head-start of dinoflagellates by the onset of the spring bloom is decisive for successful competition with the faster growing diatoms. Massive cyst formation and spreading of cyst beds fuel the expanding and ever larger dinoflagellate blooms in the relatively shallow coastal waters. Shifts in the dominant spring bloom algal groups can have significant effects on major elemental fluxes and functioning of the Baltic Sea ecosystem, but also in the vast shelves and estuaries at high latitudes, where ice-associated cold-water dinoflagellates successfully compete with diatoms.

Inducing autoflocculation in the diatom Phaeodactylum tricornutum through CO2 regulation

The effect of pH on flocculation was studied using the diatom Phaeodactylum tricornutum and the green algae Scenedesmus cf. obliquus as surrogate species. There was a distinct, species-specific threshold of pH where flocculation started. P. tricornutum started to flocculate at pHxa010.5 and S. cf. obliquus at pHxa011.3. Above this threshold, settling rates up to 360xa0cmxa0h−1 were observed for P. tricornutum and the concentrating factor was up to 60-fold. The combined effect of pH, turbulence, and cell density on flocculation of P. tricornutum was additionally studied in a factorial 53-design experiment. pH was the most important factor affecting flocculation, but at the pH threshold (pHxa010.5), the concentrating factor was increased by increasing cell density and turbulence. Algae increases the pH during photosynthesis, and the P. tricornutum and S. cf. obliquus cultures increased the pH to a maximum of 10.8 and 9.5, respectively, after discontinuing the CO2 supply. For P. tricornutum, this was above the flocculation threshold, and rapid settling of this species due to increased pH was observed in a matter of hours after the CO2 supply was turned off. This could be used as a simple, low-cost, initial dewatering step for this species.

Functional group richness: implications of biodiversity for light use and lipid yield in microalgae.

Currently, very few studies address the relationship between diversity and biomass/lipid production in primary producer communities for biofuel production. Basic studies on the growth of microalgal communities, however, provide evidence of a positive relationship between diversity and biomass production. Recent studies have also shown that positive diversity–productivity relationships are related to an increase in the efficiency of light use by diverse microalgal communities. Here, we hypothesize that there is a relationship between diversity, light use, and microalgal lipid production in phytoplankton communities. Microalgae from all major freshwater algal groups were cultivated in treatments that differed in species richness and functional group richness. Polycultures with high functional group richness showed more efficient light use and higher algal lipid content with increasing species richness. There was a clear correlation between light use and lipid production in functionally diverse communities. Hence, a powerful and cost‐effective way to improve biofuel production might be accomplished by incorporating diversity related, resource‐use‐dynamics into algal biomass production.

Interaction Effects of Light, Temperature and Nutrient Limitations (N, P and Si) on Growth, Stoichiometry and Photosynthetic Parameters of the Cold-Water Diatom Chaetoceros wighamii

Light (20-450 μmol photons m-2 s-1), temperature (3-11°C) and inorganic nutrient composition (nutrient replete and N, P and Si limitation) were manipulated to study their combined influence on growth, stoichiometry (C:N:P:Chl a) and primary production of the cold water diatom Chaetoceros wighamii. During exponential growth, the maximum growth rate (~0.8 d-1) was observed at high temperture and light; at 3°C the growth rate was ~30% lower under similar light conditions. The interaction effect of light and temperature were clearly visible from growth and cellular stoichiometry. The average C:N:P molar ratio was 80:13:1 during exponential growth, but the range, due to different light acclimation, was widest at the lowest temperature, reaching very low C:P (~50) and N:P ratios (~8) at low light and temperature. The C:Chl a ratio had also a wider range at the lowest temperature during exponential growth, ranging 16-48 (weight ratio) at 3°C compared with 17-33 at 11°C. During exponential growth, there was no clear trend in the Chl a normalized, initial slope (α*) of the photosynthesis-irradiance (PE) curve, but the maximum photosynthetic production (Pm) was highest for cultures acclimated to the highest light and temperature. During the stationary growth phase, the stoichiometric relationship depended on the limiting nutrient, but with generally increasing C:N:P ratio. The average photosynthetic quotient (PQ) during exponential growth was 1.26 but decreased to <1 under nutrient and light limitation, probably due to photorespiration. The results clearly demonstrate that there are interaction effects between light, temperature and nutrient limitation, and the data suggests greater variability of key parameters at low temperature. Understanding these dynamics will be important for improving models of aquatic primary production and biogeochemical cycles in a warming climate.

MICROSENSOR MEASUREMENTS OF THE EXTERNAL AND INTERNAL MICROENVIRONMENT OF FUCUS VESICULOSUS (PHAEOPHYCEAE) 1

We investigated the O2, pH, and irradiance microenvironment in and around the tissue of the brown alga Fucus vesiculosus L. using microsensors. Microsensors are ideal tools for gaining new insights into what limits and controls macroalgal activity and growth at very fine spatial (<100u2003μm) and temporal (seconds) scales. This first microsensor investigation of a fucoid macroalga revealed differences in the microenvironment and metabolic activities at the level of different cell layers and thallus structures. F. vesiculosus responded quickly to rapid shifts in irradiance resulting in a highly dynamic microenvironment around and within its thallus. In combination with detailed morphological studies and molecular approaches, microsensors offer a promising toolbox to quantitatively describe structural and functional adaptations of macroalgae to environmental conditions, such as flow and light climate, as well as their physiological responses to environmental stressors.

False-positive diatom test: A real challenge? A post-mortem study using standardized protocols

The main criticism of the validity of the diatom test for the diagnosis of drowning is based on the potential ante- and post-mortem penetration of diatoms and the finding of diatoms in bodies of non-drowned human beings. However, qualitative and quantitative studies on diatoms in organs of the non-drowned have yielded both conflicting and contradictory results. In the present study, we have analysed under standardised methods the diatom content in several organs of 14 non-drowned human bodies. Overall, only 9 diatoms (6 entire, 3 fragmented) were disclosed in 6 of the 14 non-drowned bodies. Each of these 6 cadavers had only a single positive organ. Six diatoms were found in the bone marrow, 2 in the lung, and one in the pleural liquid. No diatoms were recovered from the brain, liver, kidney, or blood samples of any of these 14 bodies. Moreover, in five additional cadavers, whose lungs were injected, prior autopsy, with a 3.5L solution containing a bi-cellulate diatom culture (Thalassiosira baltica, Thalassiosira levanderi) via tracheostomy, a few diatoms appeared in the pleural cavity and in the blood from the left heart chamber, but none in any other internal organs investigated. The results of the presented study demonstrate that the issue of the false-positive diatom test should not be a logical impediment to the performance of the diatom method. However, strict and standardized protocols aimed at avoiding contamination during sample preparation must be used, appropriate separation values set and taxonomic analysis of all diatoms performed.

Lipid content in 19 brackish and marine microalgae: influence of growth phase, salinity and temperature

Algal lipids provide essential fatty acids for higher trophic levels in the marine food web, and understanding the fatty acid composition in phytoplankton is critical for evaluating its value as a diet. Nineteen microalgal species, mainly originating from the Baltic Sea, covering major algal classes were grown in different growth conditions. Samples were taken during both the exponential and stationary growth phases and analysed regarding their fatty acid methyl esters and free fatty acids. Our results show that across all screened species, total fatty acids increased significantly from exponential to stationary growth phase. Furthermore, it was observed that warm-water species contained more lipids and differed in their lipid profile as compared with the cold-water species. Brackish water species also showed a slightly higher lipid content than the marine species, but their lipid profile was not significantly different. Plotting changes in lipids against changes in cell nitrogen revealed a significant dependency between decrease in cell nitrogen and increase in lipids across all tested species.

Nile Red staining of phytoplankton neutral lipids: species-specific fluorescence kinetics in various solvents

Nile Red (NR) staining potentially offers a simple method for monitoring lipid accumulation in microalgal cultivation. However, variable staining efficiencies and methods have been reported. The effect of dimethyl sulfoxide (DMSO), ethylene glycol (EG) and glycerol on NR penetration with four different phytoplankton species representing different taxonomical groups was studied. Treatment with the solvents enhanced the NR fluorescence of the diatom Phaeodactylum tricornutum during kinetic fluorescence measurements, but high concentrations of solvents were needed. None of the solvents improved NR staining of the green alga Chlorella pyrenoidosa and Scenedesmus obliquus, which are known to be difficult to stain due to their thick and rigid cell walls. The naked Isochrysis sp. cells stained best without solvents. The results confirm that NR staining protocol needs to be optimized for each species.

The effect of high pH on structural lipids in diatoms

We tested the hypothesis that increased pH reduces the amount of structural lipids. To do this, we used three different diatoms (Phaeodactylum tricornutum CCAP strain, P. tricornutum TV strain and Amphiprora sp). We tested the effect of rapid increase from pHu20097.5 to 10 by adding NaOH. The total lipid content was reduced by 13, 36 and 47xa0% in the P. tricornutum CCAP strain, TV strain and Amphiprora sp., respectively, 1xa0h after increasing the pH. The P. tricornutum CCAP strain was used for further testing the effect of pH on the lipid content during active growth. This strain was cultivated at pHu20097.5 and 10, and the pH was regulated by the CO2 inflow. The growth rate was similar (0.3xa0day−1) in both pH treatments, but the lipid content in the pHu200910 treatment was on average 28xa0% lower than in the pHu20097.5 treatment. Our data support the hypothesis that structural lipids are reduced when pH increases to high levels. The results suggest that regulating the pH during algae cultivation could be used to refine the lipid composition in the harvested algal biomass.

Contrasting seasonality in optical-biogeochemical properties of the Baltic Sea

Optical-biogeochemical relationships of particulate and dissolved organic matter are presented in support of remote sensing of the Baltic Sea pelagic. This system exhibits strong seasonality in phytoplankton community composition and wide gradients of chromophoric dissolved organic matter (CDOM), properties which are poorly handled by existing remote sensing algorithms. Absorption and scattering properties of particulate matter reflected the seasonality in biological (phytoplankton succession) and physical (thermal stratification) processes. Inherent optical properties showed much wider variability when normalized to the chlorophyll-a concentration compared to normalization to either total suspended matter dry weight or particulate organic carbon. The particle population had the largest optical variability in summer and was dominated by organic matter in both seasons. The geographic variability of CDOM and relationships with dissolved organic carbon (DOC) are also presented. CDOM dominated light absorption at blue wavelengths, contributing 81% (median) of the absorption by all water constituents at 400 nm and 63% at 442 nm. Consequentially, 90% of water-leaving radiance at 412 nm originated from a layer (z90) no deeper than approximately 1.0 m. With water increasingly attenuating light at longer wavelengths, a green peak in light penetration and reflectance is always present in these waters, with z90 up to 3.0–3.5 m depth, whereas z90 only exceeds 5 m at biomass < 5 mg Chla m-3. High absorption combined with a weakly scattering particle population (despite median phytoplankton biomass of 14.1 and 4.3 mg Chla m-3 in spring and summer samples, respectively), characterize this sea as a dark water body for which dedicated or exceptionally robust remote sensing techniques are required. Seasonal and regional optical-biogeochemical models, data distributions, and an extensive set of simulated remote-sensing reflectance spectra for testing of remote sensing algorithms are provided as supplementary data.