Sigrid Haande

Diversity of Microcystis aeruginosa isolates (Chroococcales, Cyanobacteria) from East-African water bodies

With exception of South Africa, very little is known about the presence and abundance of toxic cyanobacteria and cyanobacterial blooms on the African continent. The close proximity between society and nature, and the use of the sparse water resources as drinking water in large parts of Africa, lead to the recognition that more knowledge on toxic cyanobacterial blooms is of major importance. The bloom forming cyanobacterium Microcystis aeruginosa is known to produce cyclic heptatoxins (microcystins) which can be toxic to humans. In this study the morphological, genetic, and chemical characters of 24 strains of M. aeruginosa from several water bodies in Kenya and Uganda, some of them used as drinking water sources, were examined. The M. aeruginosa strains possessed different levels of diversity depending on characterisation method. Four morphotypes were identified based on the traditional morphological approach, 10 genotypes by DNA sequence comparison of the PC-IGS and ITS1 rDNA regions, and 10 chemotypes based on MALDI-TOF-MS oligopeptide analysis. Only 4 of the 24 isolated strains from East Africa were found to produce microcystins, while oligopeptides belonging to the aeruginosin and cyanopeptolin class were detected in most strains.

Morphological and Phylogenetic Analysis of Anabaenopsis abijatae and Anabaenopsis elenkinii (Nostocales, Cyanobacteria) from Tropical Inland Water Bodies

Anabaenopsis spp. are heterocytous cyanobacteria commonly found in tropical, subtropical, and temperate water bodies. So far, the knowledge about the phylogeny of this genus is poor. Therefore, we have isolated 15 Anabaenopsis spp. strains from Kenyan and Mexican alkaline and saline water bodies and from a Ugandan freshwater body and studied the morphology and phylogeny in a polyphasic approach. Morphologically, the investigated strains could be discriminated in two groups. One group was containing six Anabaenopsis abijatae and A. cf. abijatae strains with up to more than 500 vegetative cells in one filament, mostly single intercalary heterocyte formation, and the ability to branch out. The other group comprised nine strains of Anabaenopsis elenkinii with short filaments with up to 38 vegetative cells, intercalary heterocytes in pairs, and no ability to branch out. The morphological differences were reflected in the two distinct clusters, which were found in the phylogenetic trees of 16S rDNA and PC-IGS. While the high 16S rDNA similarity values >97.5% found between all investigated A. abijatae and A. elenkinii strains support the assignment of these two species to one single genus, the morphological differences and the low similarity values (<87.3) found in PC-IGS sequences between the two clusters indicate two separate genera. A close morphological and phylogenetic relationship was found for A. abijatae and Anabaenopsis (Cyanospira) rippkae.

LC-MS Analysis with Thiol Derivatization to Differentiate [Dhb7]- from [Mdha7]-Microcystins: Analysis of Cyanobacterial Blooms, Planktothrix Cultures and European Crayfish from Lake Steinsfjorden, Norway

Kinetic studies showed that [Asp(3), Dhb(7)]MC-RR reacted with mercaptoethanol hundreds of times more slowly than MC-RR and a range of other [Mdha(7)]-containing microcystin congeners. The difference in reaction rate was sufficiently large that derivatization of microcystin-containing samples with mercaptoethanol, followed by LC-MS analysis, clearly discriminated between microcystins containing the isobaric [Dhb(7)]- and [Mdha(7)]-groups. Application of this approach, using LC-MS with both-ion trap and triple-quadrupole mass spectrometers, to water samples and Planktothrix cultures from Lake Steinsfjorden, Norway, demonstrated the presence of [Asp(3), Dhb(7)]MC-RR (5), [Asp(3)]MC-RY (14), and [Asp(3)]MC-LY (16), as well as analogues tentatively identified as [Asp(3)]MC-RR (4), [Asp(3), DMAdda(5), Dhb(7)]MC-LR (6), [Asp(3), Dhb(7)]MC-HtyR (8), [Asp(3)]MC-HtyR (9), [Asp(3), Dhb(7)]MC-LR (10), [Asp(3)]MC-LR (11), [Asp(3), Dhb(7)]MC-RY (15), and [Asp(3), Dhb(7)]MC-LY (17), together with low levels of several other analogues. This is the first use of this thiol-based LC-MS approach to identify Dhb-containing microcystins, and allowed identification of LC-MS peaks in a mixture of [Mdha(7)]- and [Dhb(7)]-congeners of [Asp(3)]MC-RR (4, 5), -RY (14, 15), and -LY (16, 17) in the samples from L. Steinsfjorden. This is also the first report of MC-RY-congeners outside of Africa, or in Planktothrix spp. Analysis of European crayfish (Astacus astacus) taken from L. Steinsfjorden revealed the presence of only trace levels of microcystins in the edible parts.

The Red Queen Race between Parasitic Chytrids and Their Host, Planktothrix : A Test Using a Time Series Reconstructed from Sediment DNA

Parasitic chytrid fungi (phylum Chytridiomycota) are known to infect specific phytoplankton, including the filamentous cyanobacterium Planktothrix. Subspecies, or chemotypes of Planktothrix can be identified by the presence of characteristic oligopeptides. Some of these oligopeptides can be associated with important health concerns due to their potential for toxin production. However, the relationship between chytrid parasite and Planktothrix host is not clearly understood and more research is needed. To test the parasite - host relationship over time, we used a sediment core extracted from a Norwegian lake known to contain both multiple Planktothrix chemotype hosts and their parasitic chytrid. Sediment DNA of chytrids and Planktothrix was amplified and a 35-year coexistence was found. It is important to understand how these two antagonistic species can coexistence in a lake. Reconstruction of the time series showed that between 1979–1990 at least 2 strains of Planktothrix were present and parasitic pressure exerted by chytrids was low. After this period one chemotype became dominant and yet showed continued low susceptibility to chytrid parasitism. Either environmental conditions or intrinsic characteristics of Planktothrix could have been responsible for this continued dominance. One possible explanation could be found in the shift of Planktothrix to the metalimnion, an environment that typically consists of low light and decreased temperatures. Planktothrix are capable of growth under these conditions while the chytrid parasites are constrained. Another potential explanation could be due to the differences between cellular oligopeptide variations found between Planktothrix chemotypes. These oligopeptides can function as defense systems against chytrids. Our findings suggest that chytrid driven diversity was not maintained over time, but that the combination of environmental constraints and multiple oligopeptide production to combat chytrids could have allowed one Planktothrix chemotype to have dominance despite chytrid presence.

Spectrophotometric Analysis of Pigments: A Critical Assessment of a High-Throughput Method for Analysis of Algal Pigment Mixtures by Spectral Deconvolution.

The Gauss-peak spectra (GPS) method represents individual pigment spectra as weighted sums of Gaussian functions, and uses these to model absorbance spectra of phytoplankton pigment mixtures. We here present several improvements for this type of methodology, including adaptation to plate reader technology and efficient model fitting by open source software. We use a one-step modeling of both pigment absorption and background attenuation with non-negative least squares, following a one-time instrument-specific calibration. The fitted background is shown to be higher than a solvent blank, with features reflecting contributions from both scatter and non-pigment absorption. We assessed pigment aliasing due to absorption spectra similarity by Monte Carlo simulation, and used this information to select a robust set of identifiable pigments that are also expected to be common in natural samples. To test the method’s performance, we analyzed absorbance spectra of pigment extracts from sediment cores, 75 natural lake samples, and four phytoplankton cultures, and compared the estimated pigment concentrations with concentrations obtained using high performance liquid chromatography (HPLC). The deviance between observed and fitted spectra was generally very low, indicating that measured spectra could successfully be reconstructed as weighted sums of pigment and background components. Concentrations of total chlorophylls and total carotenoids could accurately be estimated for both sediment and lake samples, but individual pigment concentrations (especially carotenoids) proved difficult to resolve due to similarity between their absorbance spectra. In general, our modified-GPS method provides an improvement of the GPS method that is a fast, inexpensive, and high-throughput alternative for screening of pigment composition in samples of phytoplankton material.

Environmental conditions determine the course and outcome of phytoplankton chytridiomycosis

Chytrid fungi are highly potent parasites of phytoplankton. They are thought to force phytoplankton organisms into an evolutionary arms race with high population diversity as the outcome. The underlying selection regime is known as Red Queen dynamics. However, our study suggests a more complex picture for chytrid parasitism in the cyanobacterium Planktothrix. Laboratory experiments identified a “cold thermal refuge”, inside which Planktothrix can grow without chytrid infection. A field study in two Norwegian lakes underlined the ecological significance of this finding. The study utilized sediment DNA as a biological archive in combination with existing monitoring data. In one lake, temperature and light conditions forced Planktothrix outside the thermal refuge for most of the growing season. This probably resulted in Red Queen dynamics as suggested by a high parasitic pressure exerted by chytrids, an increase in Planktothrix genotype diversity over time, and a correlation between Planktothrix genotype diversity and duration of bloom events. In the second lake, a colder climate allowed Planktothrix to largely stay inside the thermal refuge. The parasitic pressure exerted by chytrids and Planktothrix genotype diversity remained low, indicating that Planktothrix successfully evaded the Red Queen dynamics. Episodic Planktothrix blooms were observed during spring and autumn circulation, in the metalimnion or under the ice. Interestingly, both lakes were dominated by the same or related Planktothrix genotypes. Taken together, our data suggest that, depending on environmental conditions, chytrid parasitism can impose distinct selection regimes on conspecific phytoplankton populations with similar genotype composition, causing these populations to behave and perhaps to evolve differently.

Simulating water quality and ecological status of Lake Vansjø, Norway, under land-use and climate change by linking process-oriented models with a Bayesian network

Excess nutrient inputs and climate change are two of multiple stressors affecting many lakes worldwide. Lake Vansjø in southern Norway is one such eutrophic lake impacted by blooms of toxic blue-green algae (cyanobacteria), and classified as moderate ecological status under the EU Water Framework Directive. Future climate change may exacerbate the situation. Here we use a set of chained models (global climate model, hydrological model, catchment phosphorus (P) model, lake model, Bayesian Network) to assess the possible future ecological status of the lake, given the set of climate scenarios and storylines common to the EU project MARS (Managing Aquatic Ecosystems and Water Resources under Multiple Stress). The model simulations indicate that climate change alone will increase precipitation and runoff, and give higher P fluxes to the lake, but cause little increase in phytoplankton biomass or changes in ecological status. For the storylines of future management and land-use, however, the model results indicate that both the phytoplankton biomass and the lake ecological status can be positively or negatively affected. Our results also show the value in predicting a biological indicator of lake ecological status, in this case, cyanobacteria biomass with a BN model. For all scenarios, cyanobacteria contribute to worsening the status assessed by phytoplankton, compared to using chlorophyll-a alone.

Historical Planktothrix diversity across seven Norwegian lakes implies environmentally driven niche differentiation

Non-ribosomal synthetase-produced cyanopeptoline oligopeptides enables differentiation of subpopulations of the cyanobacterial genus Planktothrix into chemotypes. It is unknown what influences the population structuring of these chemotypes. Sediment cores from seven lakes in southern Norway allowed temporal reconstruction of chemotype diversity from sites where there is only fragmented historical information. Sediment DNA was amplified using primers designed to specify the chemotype variations found within the cyanopeptoline ociB gene cluster. Findings indicate that of the seven lakes studied, only two lakes had Planktothrix populations containing all four of the most common Norwegian chemotypes. We used Principle Component and Kendall tau analysis to investigate the ability of monitoring data to predict chemotype diversity, and to identify possible biotic or abiotic barriers to chemotype dispersal. The best predictor was a negative relationship between number of chemotypes present in a lake and the concentration of chlorophyll a in the top 0 – 4 meters. At low chlorophyll a concentrations, light penetration is typically deeper, which could allow light tolerant Planktothrix to move deeper into the colder waters. Recent research findings have suggested this allows for a window of opportunity for Planktothrix to escape parasitism. With this added cold, light-constrained niche, more chemotypes might find refuge. The resulting increase in chemotype diversity within Planktothrix populations could present a greater defense against parasitism when conditions varied, such as by seasonal light changes.