Der Großteil von Seen und Flüssen in Deutschland befindet sich nicht in dem von der europäischen Wasserrahmenrichtlinie geforderten guten ökologischen Zustand. Die Ursache hierfür besteht in den meisten Gewässern nach wie vor in zu hohen Nährstoffbelastungen. Dadurch wird besonders im Sommer das Wachstum des Phytoplanktons (Algen) gefördert, das Wasser wird trübe, zeitweise sauerstoffarm und riecht unangenehm. Solche Gewässer stellen für viele Tiere und Pflanzen keinen geeigneten Lebensraum dar und sind für den Menschen unattraktiv. Die Hauptnährstoffe, um die es dabei geht, sind Stickstoff und Phosphor. Dabei galt Phosphor (P) lange Zeit als der begrenzende Faktor der Phytoplanktonbiomasse in Binnengewässern: Je geringer die PKonzentration desto geringer die Biomasse und desto besser die Gewässergüte. Dies ist bis heute Lehrbuchmeinung. In der Praxis wurde und wird daher auf eine Senkung der Phosphorkonzentrationen gesetzt, was in vielen, aber längst nicht allen Gewässern zum Erfolg führte. Deutlich weniger Studien zeigten, dass Stickstoff die Phytoplanktonbiomasse begrenzt, was allerdings auch darauf zurückzuführen ist, dass deutlich weniger Studien zum Einfluss von Stickstoff durchgeführt wurden. Eine systematische Analyse zur Bedeutung von Phosphor im Vergleich zu Stickstoff fehlte bisher. Bis heute wird daher die Bedeutung von Stickstoff als begrenzender Faktor der Phytoplanktonbiomasse weitgehend negiert. In NITROLIMIT I (2011 - 2013) wurde dagegen gezeigt, dass die Algenbiomasse in fast der Hälfte der Seen der Norddeutschen Tiefebene durch N begrenzt wird. In der Praxis wird bislang die gezielte Reduktion von Stickstoffeinträgen abgelehnt, weil man befürchtet, dass dies besonders in Seen durch Stickstofffixierung von Cyanobakterien ausgeglichen werden kann und sinkende Nitratkonzentrationen die Freisetzung von Phosphor aus den Gewässersedimenten steigern. Beides könnte einer Verbesserung der Gewässergüte entgegenwirken. Für diese Argumente fehlte jedoch eine fundierte wissenschaftliche Grundlage. Stickstoff wird sowohl in Seen als auch in Fließgewässern intensiv umgesetzt und kann über verschiedene mikrobielle Umsatzprozesse (insbesondere Denitrifikation) auch wieder aus dem System entfernt werden. Fließgewässer transportieren schließlich die nicht zurückgehaltenen Nährstoffe aus den Einzugsgebieten in die Ästuare, Küstengewässer und Meere, wo in weiten Bereichen Stickstoff der limitierende Nährstoff ist. Über den Umsatz und den Rückhalt von Stickstoff in großen Flüssen bestehen bis heute allerdings große Unsicherheiten. Zur Beantwortung der Frage „Ist Stickstoffreduktion ökologisch sinnvoll?“ bestand daher umfangreicher Forschungsbedarf. Die bisherige Strategie zur Verbesserung der Gewässergüte zielte auf Minderung der Phosphorkonzentration ab. Hierzu existieren Erfahrungswerte zu Wirkung und Kosten von Maßnahmen. Viele Maßnahmen zur Phosphorreduktion gehen zu einem gewissen Teil auch mit Stickstoffreduktion einher. Der Erfolg der Begleiterscheinung „Stickstoffreduktion“ wurde jedoch meist nicht analysiert. Fallstudien zur Verbesserung der Gewässergüte durch gezielte Stickstoffminderung wurden bisher nicht durchgeführt, weshalb Daten und Erfahrungen zu Kosten und Wirksamkeit solcher Maßnahmen fehlen. Unabhängig davon, ob eine Verbesserung der Gewässergüte über Phosphor- oder Stickstoffreduktion angestrebt wurde, fehlte bisher eine Strategie, nach der im Voraus Kosten, Wirksamkeit und Nutzen ermittelt und abgewogen werden. Daher bestand auch zur Beantwortung der Frage, „Ist Stickstoffreduktion wirtschaftlich vertretbar?“ deutlicher Forschungsbedarf.

Der Großteil der bundesdeutschen Binnengewässer wird bis 2015 nicht den guten ökologischen Zustand erreichen, der von der EU-Wasserrahmenrichtlinie gefordert wird. Bisher ging man davon aus, dass die Gewässergüte in erster Linie durch Phosphor bestimmt wird. In jüngster Zeit mehrten sich aber Hinweise, dass in vielen Gewässern auch Stickstoff eine entscheidende Steuergröße der Phytoplanktonentwicklung darstellt. Daher wird die Reduzierung von Stickstoffeinträgen gefordert. Die Kosten für Maßnahmen zur Reduktion der Stickstoffeinträge aus punktuellen (beispielsweise Kläranlagen) und diffusen Quellen (beispielsweise aus der Landwirtschaft) werden um ein Vielfaches höher geschätzt im Vergleich zu Maßnahmen zur Reduktion von Phosphoreinträgen. Ob Maßnahmen zur Stickstoffreduktion ökologisch wirksam werden, kann aufgrund unzureichender Kenntnisse zur Herkunft, Umsetzung und Wirkung von Stickstoff derzeit nicht eingeschätzt werden. Daher fordern öffentliche und wirtschaftliche Maßnahmenträger nachdrücklich eine Klärung des Nutzens von Stickstoffelimination. An diesem Punkt setzt NITROLIMIT an. Es sollte eine fundierte wissenschaftliche Grundlage zur Beurteilung des Einflusses von Stickstoff auf die Gewässergüte geschaffen, die Kosten und Nutzen von Maßnahmen zur Verringerung von Stickstoffeinträgen analysiert und darauf basierend Empfehlungen für eine nachhaltige Gewässerbewirtschaftung erarbeitet werden.

Cyanobacteria of the order Nostocales – native species as well as alien species from tropical regions – were found to increase in many Brandenburg lakes while the formerly dominating microcystin (MC) producing cyanobacteria (Microcystis and Planktothrix) occurred less often and in lower amounts. As a consequence, lower MC concentrations were observed while the toxin cylindrospermopsin (CYN) that is produced by Nostocales was found to be widely distributed and to exceed sometimes the recommended guideline value for drinking-water of 1 µg L-1. Recent data on the occurrence of further neurotoxins (paralytic shellfish poisoningtoxin, PSP and anatoxin, ATX) produced by cyanobacteria of the order Nostocales did not exist. Nostocales are superior competitors under conditions of high light intensity and nitrogen depletion because they can fix molecular nitrogen. Their germination is regulated by temperature and the temporal starting point of the pelagic population determines the population size (the earlier the larger). Therefore, the following working hypothesis has been put forward: Combined effects of declining trophic state and global warming favor the development of Nostocales and cause a shift in the species composition as well as in the occurrences of cyanobacterial toxins. The NOSTOTOX project aimed to determine the present occurrence and future development of Nostocales and their toxins in waterbodies. Special emphasis was paid to answer the question, which Nostocales species and which toxins can be expected under conditions of a proceeding decline in trophic state and increasing water temperature. The outcome of the project aims to contribute to developing recommendations and guidelines for the management of inland waters and drinking water supplies.

Some tropical cyanobacteria have spread to temperate freshwaters during the last decades. To evaluate their further development in temperate lakes, we studied the temperature- and light-dependent growth of three invasive (Cylindrospermopsis raciborskii, Anabaena bergii and Aphanizomenon aphanizomenoides) and three native (Aphanizomenon gracile, Aphanizomenon flos-aquae and Anabaena macrospora) cyanobacterial species (Nostocales) from German lakes. We also included one potentially invasive (Aphanizomenon ovalisporum) Nostocales species. We conducted semi-continuous culture experiments and a microcosm experiment along a natural light gradient. Temperature data were used to design a model to simulate the development of selected species according to three temperature scenarios (past, present and future). Native species had significantly higher growth rates than invasive species and the potential invader A. ovalisporum at low temperatures (<= 10 °C), while the opposite was true at high temperatures (>= 35 °C). Maximum growth rates of A. ovalisporum, A. aphanizomenoides and C. raciborskii were clearly higher than those of A. bergii and the native species. Regarding light-dependent growth, significant differences were found between single species but not between all native and invasive species. The model simulation demonstrates a shift in dominance from the native A. gracile in the historic scenario to C. raciborskii populations in the future scenario, in which also the potential invader A. ovalisporum is able to establish populations in temperate lakes. Our findings suggest that any further temperature increase would promote the growth and development of Nostocales species in general, and that of the invasive species in particular, and would enable a more northward expansion of A. ovalisporum.

The neurotoxin anatoxin-a (ATX), has been detected in several northeast German lakes during the last two decades, but no ATX producers have been identified in German water bodies so far. In 2007 and 2008, we analyzed phytoplankton composition and ATX concentration in Lake tolpsee (NE Germany) in order to identify ATX producers. Sixty-one Aphanizomenon spp. strains were isolated, morphologically and phylogenetically characterized, and tested for ATX production potential by liquid chromatography–tandem mass spectrometry (LC–MS/MS). New primers were specifically designed to identify a fragment of a polyketide synthase gene putatively involved in ATX synthesis and tested on all 61 Aphanizomenon spp. strains from L. Stolpsee and 92 non-ATX-producing Aphanizomenon spp., Anabaena spp. and Anabaenopsis spp. strains from German lakes Langersee, Melangsee and Scharmützelsee. As demonstrated by LC–MS/MS, ATX concentrations in L. Stolpsee were undetectable in 2007 and ranged from 0.01 to 0.12 µg l-1 in 2008. Fifty-nine of the 61 strains isolated were classified as Aphanizomenon gracile and two as Aphanizomenon issatschenkoi. One A. issatschenkoi strain was found to produce ATX at concentrations of 2354 ± 273 µg g-1 fresh weight, whereas the other A. issatschenkoi strain and A. gracile strains tested negative. The polyketide synthase gene putatively involved in ATX biosynthesis was found in the ATX-producing A. issatschenkoi strain from L. Stolpsee but not in the non-ATX-producing Aphanizomenon spp., Anabaena spp. and Anabaenopsis spp. strains from lakes Stolpsee, Langersee, Melangsee, and Scharmützelsee. This study is the first confirming A. issatschenkoi as an ATX producer in German water bodies.

Neurotoxic paralytic shellfish poisoning (PSP) toxins, anatoxin-a (ATX), and hepatotoxic cylindrospermopsin (CYN) have been detected in several lakes in northeast Germany during the last 2 decades. They are produced worldwide by members of the nostocalean genera Anabaena, Cylindrospermopsis, and Aphanizomenon. Although no additional sources of PSP toxins and ATX have been identified in German water bodies to date, the observed CYN concentrations cannot be produced solely by Aphanizomenon flos-aquae, the only known CYN producer in Germany. Therefore, we attempted to identify PSP toxin, ATX, and CYN producers by isolating and characterizing 92 Anabaena, Aphanizomenon, and Anabaenopsis strains from five lakes in northeast Germany. In a polyphasic approach, all strains were morphologically and phylogenetically classified and then tested for PSP toxins, ATX, and CYN by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and enzyme-linked immunosorbent assay (ELISA) and screened for the presence of PSP toxin- and CYN-encoding gene fragments. As demonstrated by ELISA and LC-MS, 14 Aphanizomenon gracile strains from Lakes Melang and Scharmützel produced four PSP toxin variants (gonyautoxin 5 [GTX5], decarbamoylsaxitoxin [dcSTX], saxitoxin [STX], and neosaxitoxin [NEO]). GTX5 was the most prevalent PSP toxin variant among the seven strains from Lake Scharmützel, and NEO was the most prevalent among the seven strains from Lake Melang. The sxtA gene, which is part of the saxitoxin gene cluster, was found in the 14 PSP toxin-producing A. gracile strains and in 11 non-PSP toxin-producing Aphanizomenon issatschenkoi, A. flos-aquae, Anabaena planktonica, and Anabaenopsis elenkinii strains. ATX and CYN were not detected in any of the isolated strains. This study is the first confirming the role of A. gracile as a PSP toxin producer in German water bodies.

Tropical cyanobacteria in German waters: Causes and consequences - Toxic cyanobacteria of tropical origin have spread to water bodies in northern Germany. Here their population size is determined by the onset of germination: the earlier the time of germination, the larger the summer population. Climate-related early increase in water temperatures over the course of the years has promoted their spreading to temperate regions. Toxins known from these species in tropical regions were also found in German lakes. Surprisingly, these toxins are not produced by the invaders but by native species. Thus, the invasion have drawn our attention to an old problem. However, the invaders itself potentially also poses a health risk in German lakes since they produce other – so far unidentified – toxic substances.

Cylindrospermopsis raciborskii, a cyanobacterium of tropical origin, can produce the toxin cylindrospermopsin (CYN). This originally tropical cyanobacterium (bluegreen algae) has now spread to the distant waters of the Berlin area. Cylindrospermopsin has been detected in two lakes in the area, but none of the C. raciborskii strains isolated here so far were found to produce the toxin. The main objectives of the CYLIN project were therefore to analyze the distribution and regulation of C. raciborskii and cylindrospermopsin and to determine which cyanobacteria are producing this toxin in order to establish a basis with which to predict the further course of development of this species and the related health hazards for humans. The CYLIN project was implemented as a three-part program. A screening program was first conducted in 2004 to test regional water bodies for the presence of cylindrospermopsin and potential CYN-producing cyanobacteria in order to obtain an overview of their distribution in the study region. A total of 142 regional water bodies were sampled once each in this qualitative analysis cylindrospermopsin and cyanobacteria. The screening program was followed by a monitoring program designed to generate quantitative data on the concentrations of dissolved CYN, particulate CYN, cyanobacteria and target environmental parameters at 20 selected lakes, which were sampled 3 times each. Last but not least, we investigated the seasonal dynamics of these parameters at two selected lakes in 2004 and 2005. Apart from this we isolated different cyanobacterial strains and conducted chemical and molecular biological analyses of CYN and CYNcoding genes, in order to identify CYN-producing cyanobacteria. The results show that C. raciborskii and CYN are much more widespread than was previously assumed. C. raciborskii was detected in 22 % of the investigated water bodies, and cylindrospermopsin in 52 %. Additionally, two other toxic cyanobacteria of tropical origin were found for the first time in the BerlinBrandenburg region, Anabaena bergii and Aphanizomenon aphanizomenoides. The mean and maximum CYN concentrations were 1 µg L-1 and 12 µg L-1, respectively. Since the particulate CYN fraction did not exceed 0.5 µg L-1, the dissolved CYN fraction was found to be responsible for the high CYN concentrations. The proposed guideline safety value for cylindrospermopsin in drinking water (1 µg L-1) was exceeded 18 times at 8 different lakes. Although Aphanizomenon flos-aquae (Nostocales) has been unequivocally identified as a producer of cylindrospermopsin, the observed cylindrospermopsin concentrations cannot be attributed to this cyanobacterial species alone. Aphanizomenon gracile was also identified as a potential CYN-producing cyanobacterium. Based on the findings of the CYLIN project, we recommend that cylindrospermopsin be included as a risk factor in drinking and bathing water quality assessments. To identify hazard conditions associated with this cyanotoxin, further investigations are needed to identify all cyanobacteria that produce cylindrospermopsin and to elucidate the mechanisms regulating the occurrence of CYN-producing cyanobacteria, CYN synthesis by these organisms, and CYN decomposition in aquatic ecosystems. Our analysis of C. raciborskii population dynamics showed that its germination is temperature-dependent and its population growth light-dependent. Population size was determined by the time of germination, that is, the earlier the time of germination, the bigger the population. Based on these findings, it appears highly likely that the climate-related early rise in water temperatures over the course of the years has promoted the spread of this species to temperate regions. Our hypothesis for the future course of cyanobacterial and cyanotoxin development in German waters is as follows: The combination of trophic decline and global warming works to the general benefit of cyanobacteria of the order Nostocales and leads to a shift in cyanobacterial species and toxin composition. This may ultimately lead to an increase in the incidence of neurotoxins as well as cylindrospermopsin.

Cylindrospermopsis raciborskii, a cyanobacterium of tropical origin, can produce the toxin cylindrospermopsin (CYN). This originally tropical cyanobacterium (blue-green alga) had spread to the waters of the Berlin area. Cylindrospermopsin had been detected in two lakes in the area, but none of the C. raciborskii strains isolated here so far were found to produce the toxin. The main objectives of the CYLIN project were therefore to analyze the distribution and regulation of C. raciborskii and cylindrospermopsin and to determine which cyanobacteria are producing this toxin in order to establish a basis to predict the further course of development of this species and the related health hazards for humans. The CYLIN project was implemented as a three-part program. A screening program was first conducted in 2004 to test regional water bodies for the presence of cylindrospermopsin and potential CYN-producing cyanobacteria in order to obtain an overview of their distribution in the study region. A total of 142 regional water bodies were sampled once each in this qualitative analysis of cylindrospermopsin and cyanobacteria. The screening program was followed by a monitoring program designed to generate quantitative data on the concentrations of dissolved CYN, particulate CYN, cyanobacteria and target environmental parameters at 20 selected lakes, which were sampled 3 times each. Furthermore, we investigated the seasonal dynamics of these parameters at two selected lakes in 2004 and 2005. Apart from this we isolated different cyanobacterial strains and conducted chemical and molecular biological analyses of CYN and CYN-coding genes, in order to identify CYN-producing cyanobacteria. The results show that C. raciborskii and CYN are much more widespread than was previously assumed for the region. C. raciborskii was detected in 22 % of the investigated water bodies, and cylindrospermopsin in 52 %. Additionally, two other toxic cyanobacteria of tropical origin were found for the first time in the Berlin-Brandenburg region, Anabaena bergii and Aphanizomenon aphanizomenoides. The mean and maximum CYN concentrations were 1 µg L-1 and 12 µg L-1, respectively. Since the particulate CYN fraction did not exceed 0.5 µg L-1, the dissolved CYN fraction was found to be responsible for the high CYN concentrations. The proposed guideline value for cylindrospermopsin in drinking water (1 µg L-1; Humpage and Falconer 2003) was exceeded 18 times at 8 different lakes. Although Aphanizomenon flos-aquae (Nostocales) has been unequivocally identified as a producer of cylindrospermopsin, the observed cylindrospermopsin concentrations cannot be attributed to this cyanobacterial species alone. Aphanizomenon gracile was also identified as a potential CYN-producing cyanobacterium. Based on the findings of the CYLIN project, we recommend that cylindrospermopsin be included as in hazard analyisis for drinking and bathing water quality assessments. To identify risk conditions associated with this cyanotoxin, further investigations are needed to identify all cyanobacteria that produce cylindrospermopsin and to elucidate the mechanisms regulating the occurrence of CYN-producing cyanobacteria, CYN synthesis by these organisms, and CYN decomposition in aquatic ecosystems. Our analysis of C. raciborskii population dynamics showed that its germination is temperature-dependent and its population growth light-dependent. Population size was determined by the time of germination, that is, the earlier the time of germination, the bigger the population. Based on these findings, it appears highly likely that the climate-related early rise in water temperatures over the course of the years has promoted the spread of this species to temperate regions. Our hypothesis for the future course of cyanobacterial and cyanotoxin development in German waters is as follows: The combination of trophic decline and global warming works to the general benefit of cyanobacteria of the order Nostocales and leads to a shift in cyanobacterial species and toxin composition. This may ultimately lead to an increase in the incidence of neurotoxins as well as cylindrospermopsin.