FATE OF THE TOXIC CYCLIC HEPTAPEPTIDES,THE MICROCYSTINS,FROM BLOOMS OF MICROCYSTIS (CYANOBACTERIA) IN A HYPERTROPHIC LAKE1

The in situ fate of the toxic cyclic heptapeptides, the microcystins, produced by blooms of Microcystis was examined at two stations in a hypertrophic Japanese lake. Microcystins were detected in all samples of Microcystis with quantities varying seasonally and spatially (230–950 μg · g dry wt^?1 at St. 1 and 160–746 μg · g dry wt^?1 at St. 2) and composed of microcystin-LR, -RR, and-YR. Microcystin-RR was the dominant toxin in most samples. A large amount of microcystin (1.1 μg · L^?1) was detected in only one sample of filtered lake water. Accumulation of microcystin in zooplankton was indirectly estimated from a newly developed equation model. Large amounts of microcystin (75–1387 μg · g dry wt^?1) were accumulated in the zooplankton community, which consisted of two cladocerans, Bosmina fatalis Burckhardt and Diaphanosoma brachyurum Lieve, and a copepod, Cyclops vicinus Uljanin, that co-occurred with the toxic Microcystis blooms. The maximum percent of microcystin content in zooplankton to that in Microcystis was 202%. Among the three species of zooplankton, only B. fatalis seemed to be responsible for accumulation of the microcystins because C. vicinus appeared to avoid contact with Microcystis cells and D. brachyurum did not consume colonies of Microcystis. Microcystins may be transferred to higher trophic levels through B. fatalis.     

Maternal consumption of non‐toxic Microcystis by Daphnia magna induces tolerance to toxic Microcystis in offspring

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Effects of toxic and non-toxic cyanobacteria on the life history of tropical and temperate cladocerans

1. This study compares the effects of four toxic strains of Microcystis aeruginosa on tropical and temperate Cladocera. Survival was tested in acute toxicity experiments using Microcystis alone or in mixtures with the edible green algae Ankistrodesmus falcatus. The effect of chronic exposure on population growth was estimated in life‐table experiments by varying the proportion of Microcystis and the green alga. Nutritional deficiency was assessed using a non‐toxic cyanobacterium in a zooplankton growth experiment. Feeding inhibition was tested using a C‐labelled green alga as a tracer in mixtures with toxic Microcystis.
2. Toxicity varied consistently between Microcystis strains, while sensitivity varied consistently between cladoceran species. However, no relationship was found between sensitivity and geographical origin or cladoceran body size. Two small‐bodied cladocerans from the same tropical lake, Ceriodaphnia cornuta and Moinodaphnia macleayi, were the least sensitive and most sensitive species, respectively.
3. Surprisingly, two small tropical cladocerans survived longer without food than did three large Daphnia species and a third small tropical species.
4. Each of the three tropical Microcystis strains strongly reduced the population growth rate (little ‘r’) and reproductive output of each cladoceran, this reduction being proportional to the percentage of toxic cells in the diet.
5. As the sole food source, the non‐toxic cyanobacterium, Synechococcus elongatus, supported poor growth in M. macleayi. The nutritional deficiency was overcome when Synechococcus was mixed with either Ankistrodesmus or an emulsion rich in omega‐3 fatty acids.
6. Microcystis inhibited the feeding rate of two cladocerans, even when it comprised only 5% of a mixture with the green algae A. falcatus.
7. Differences in sensitivity to the toxic cyanobacterium appear to be associated with differences in life history between the cladoceran species rather than differences between tropical and temperate taxa. Slow‐growing species that are resistant to starvation appear less sensitive to toxic Microcystis than fast‐growing species, which also tend to die more quickly in the absence of food.     

Toxicity to Daphnia of a compound extracted from laboratory and natural Microcystis spp., and the role of microcystins

1. The microcystin content of a variety of Microcystis spp., from both laboratory strains and natural blooms, was analysed by HPLC. The microcystin content of laboratory strains ranged from 1.6 to 4.3μgmg^?1 dry weight. Yearly and seasonal variation was detected in an analysis of bloom material collected from Bautzen Reservoir over a 3-year period. The microcystin concentration in bloom material ranged from undetectable to 1.16 μg ml^?1 dry weight. 2. Toxicity of laboratory and natural Microcystis to Daphnia pulicaria was determined using an established LC₅₀ technique. Partially purified water extracts from different Microcystis samples exhibited a wide range of toxicity. The highest activity was found in natural Microcystis samples, with an LC₅₀ of 36 μgm^?1 dry weight of Microcystis, whereas one strain did not appear toxic at 1600 μg ml^?1. 3. No correlation was found between the concentrations of microcystins of different laboratory and natural Microcystis strains and the toxicity of extracts to Daphnia pulicaria from the same strains. Therefore, we discriminated between hepatotoxic microcystins and the compound(s) that is toxic to Daphnia, here termed DTC (Daphnia-toxic compound), which is independent of microcystins.     

Isolation and characterization of microcystin producing Microcystis from a Central Indian water bloom

Microcystis aeruginosa Kütz, a well-known microcystin (hepatotoxin) producing cyanobacterium was the dominant bloom-forming organism in a mesotrophic lake at Nagpur in Central India, which was isolated and characterized for morphospecies and microcystin content. Compact spherical colonies, formation of daughter colonies, and clathration of older colonies leading to release of solitary cells, were characteristics of laboratory grown M. aeruginosa. Its growth, monitored as increase in optical density (OD) measured at 678 nm (the wavelength selected using dilution curve technique), exhibited a maximum specific growth rate (μ_max) of 0.34 day⁻¹ which, was attained on the 5th day of the experiment with a doubling time of 3.25 days. Though the morphological characters of the M. aeruginosa under field conditions were not retained under laboratory conditions, the microcystin content and type of variants did match with bloom samples. Reverse phase high performance liquid chromatography (RP-HPLC) analyses revealed that the laboratory grown isolate of Microcystis produced microcystin-RR (732 μg g⁻¹ dry weight biomass) and demethylated microcystin-RR (165 μg g⁻¹ dry weight biomass) variants, which are reported to be less toxic when compared to microcystin-LR. LC/ESI/MS further confirmed the presence of these two variants. Geographical distribution of microcystin variants and their prevailing concentrations need to be considered during formulation of guideline values for drinking and recreational waters.     

Influence of ultraviolet radiation, copper, and zinc on microcystin content in Microcystis aeruginosa (Cyanobacteria)

Cyanobacterial toxin production is allied to some unknown trigger resulting in the production of toxins such as microcystin. We hypothesize that microcystins serve as metal ligands to control bioavailability and toxicity of ambient metals. Since ultraviolet radiation (UVR) promotes photo-oxidation of organic metal ligands and influences trace metal bioavailability, the present study aimed to investigate the influence of UVR, Cu, and Zn on specific growth rates, biomass, photosynthetic capacity, and microcystin content in Microcystis aeruginosa. Two toxigenic strains of Microcystis were cultivated using either Lake Erie filtered water or a chemically defined medium, with realistic concentrations of Cu and Zn combined with natural or artificial UVR exposure. Cu was more toxic than Zn on the basis of free ion concentration of trace metals in synthetic medium, although in Lake Erie water total added Zn (10 nM) or Zn plus Cu (10 nM) had a more detrimental effect on biomass and specific growth rate. Natural UVR delivered at 25% ambient levels caused no decrease on the parameters measured (chlorophyll-a, photosynthetic rate), yet artificial levels of UVR (up to 5.9 μmol UVB photons m⁻² s⁻¹) negatively affected biomass and specific growth rate. Cellular levels of microcystin (per unit chlorophyll-a) were concomitant with specific growth rather than being triggered in response either of these stressors (UVR, Zn, and Cu) alone or in combination, in agreement with a purported constitutive production of microcystins.     

Cross-Reactivity and Neutralizing Ability of Monoclonal Antibodies against Microcystins

Monoclonal antibodies (MAbs) against the microcystin-leucine-arginine variant (MCYST-LR), a cyclic peptide toxin of the freshwater cyanobacterium Microcystis aeruginosa, were prepared from cloned hybridoma cell lines. The specificity of the MAbs and their ability to neutralize the toxin were investigated by an indirect enzyme-linked immunosorbent assay (ELISA) and by a neutralizing test in mice, respectively. All MAbs reacted with MCYST-LR and also with the microcystin-arginine-arginine variant (MCYST-RR), 3, 7-didesmethylmicrocystin (MCYST-3, 7-dDMLR) and 7-desmethylmicrocystin (MCYST-7-DMLR). Furthermore, the antibodies reacted with cell-extracts of toxic and non-toxic M. aeruginosa strains. The MAbs can apparently recognize the common configuration, but not the variant-specific structure, in the microcystin molecules. The non-toxic strains apparently contain some substance(s) related antigenically to microcystin. The in vivo toxin-neutralizing ability of MAbs was minimal.     

The involvement of α-proteobacteria Phenylobacterium in maintaining the dominance of toxic Microcystis blooms in Lake Taihu,China

Lake Taihu in China has suffered serious harmful cyanobacterial blooms for decades. The algal blooms threaten the ecological sustainability, drinking water safety, and human health. Although the roles of abiotic factors (such as water temperature and nutrient loading) in promoting Microcystis blooms have been well studied, the importance of biotic factors (e.g. bacterial community) in promoting and meditating Microcystis blooms remains unclear. In this study, we investigated the ecological dynamics of bacterial community, the ratio of toxic Microcystis, as well as microcystin in Lake Taihu. High-throughput 16S rRNA sequencing and principal component analysis (PCA) revealed that the bacteria community compositions (BCCs) clustered into three groups, the partitioning of which corresponded to that of groups according to the toxic profiles (the ratio of toxic Microcystis to total Microcystis, and the microcystin concentrations) of the samples. Further Spearman's correlation network showed that the α-proteobacteria Phenylobacterium strongly positively correlated with the toxic profiles. Subsequent laboratory chemostats experiments demonstrated that three Phenylobacterium strains promoted the dominance of the toxic Microcystis aeruginosa PCC7806 when co-culturing with the non-toxic PCC7806 mcyB⁻ mutant. Taken together, our data suggested that the α-proteobacteria Phenylobacterium may play a vital role in the maintenance of toxic Microcystis dominance in Lake Taihu.     

The effects of temperature and nutrients on the growth and dynamics of toxic and non-toxic strains of Microcystis during cyanobacteria blooms

In temperate latitudes, toxic cyanobacteria blooms often occur in eutrophied ecosystems during warm months. Many common bloom-forming cyanobacteria have toxic and non-toxic strains which co-occur and are visually indistinguishable but can be quantified molecularly. Toxic Microcystis cells possess a suite of microcystin synthesis genes (mcyA–mcyJ), while non-toxic strains do not. For this study, we assessed the temporal dynamics of toxic and non-toxic strains of Microcystis by quantifying the microcystin synthetase gene (mcyD) and the small subunit ribosomal RNA gene, 16S (an indicator of total Microcystis), from samples collected from four lakes across the Northeast US over a two-year period. Nutrient concentrations and water quality were measured and experiments were conducted which examined the effects of elevated levels of temperatures (+4 °C), nitrogen, and phosphorus on the growth rates of toxic and non-toxic strains of Microcystis. During the study, toxic Microcystis cells comprised between 12% and 100% of the total Microcystis population in Lake Ronkonkoma, NY, and between 0.01% and 6% in three other systems. In all lakes, molecular quantification of toxic (mcyD-possessing) Microcystis was a better predictor of in situ microcystin levels than total cyanobacteria, total Microcystis, chlorophyll a, or other factors, being significantly correlated with the toxin in every lake studied. Experimentally enhanced temperatures yielded significantly increased growth rates of toxic Microcystis in 83% of experiments conducted, but did so for non-toxic Microcystis in only 33% of experiments, suggesting that elevated temperatures yield more toxic Microcystis cells and/or cells with more mcyD copies per cell, with either scenario potentially yielding more toxic blooms. Furthermore, concurrent increases in temperature and P concentrations yielded the highest growth rates of toxic Microcystis cells in most experiments suggesting that future eutrophication and climatic warming may additively promote the growth of toxic, rather than non-toxic, populations of Microcystis, leading to blooms with higher microcystin content.     

A PCR‐BASED TEST TO ASSESS THE POTENTIAL FOR MICROCYSTIN OCCURRENCE IN CHANNEL CATFISH PRODUCTION PONDS1,2

Microcystis aeruginosa is a common form of cyanobacteria (blue‐green algae) capable of forming toxic heptapeptides (microcystins) that can cause illness or death. Occasionally, blooms of cyanobacteria have caused toxic fish‐kills in catfish production ponds. We have developed a PCR test that will detect the presence of microcystin‐producing cyanobacteria. Microcystin producers are detected by the presence of the microcystin peptide synthetase B gene (an obligate enzyme in the microcystin pathway), which appears to be present only in toxin‐producing cyanobacteria. These PCR amplifications can be performed in multiplex using purified DNA from pond waters or by two‐stage amplification from native water samples. A synoptic survey of 476 channel catfish production ponds from four states in the southeastern United States revealed that 31% of the ponds have the genetic potential to produce microcystins by toxic algae.     

Variation of Microcystin Content of Cyanobacterial Blooms and Isolated Strains in Lake Grand-Lieu (France)

Abstract Cyanobacterial blooms were sampled at five locations in Lake Grand-Lieu on seven different occasions during May–October 1994. Strains of Microcystis aeruginosa and Anabaena circinalis were isolated from the samples. Microcystins were detected in freeze-dried field samples and the isolated strains by HPLC. The toxins were present in the blooms sampled between June and October. The microcystin content in the blooms varied with site and time, from undetectable concentrations to 0.23 mg g⁻¹. The highest concentrations of microcystin were found in blooms sampled in September. Microcystin-LR and microcystins with retention times close to the retention time of [Dha⁷]microcystin-RR (probably varieties of microcystin-RR) were found in the field samples. Sixteen of the 98 isolated M. aeruginosa strains and 2 of the 24 A. circinalis strains produced microcystins. The total amount of microcystins varied from undetectable concentrations to 5.06 mg g⁻¹ in the M. aeruginosa isolates, and from undetectable concentrations to 1.86 mg g⁻¹ in the A. circinalis strains. Microcystin-LR was the main toxin found in strains of M. aeruginosa, but was not present in strains of A. circinalis. Both microcystin-producing strains and strains that did not produce microcystin coexisted in the bloom samples. Received: 23 January 1997; Accepted: 25 March 1997     

Morphological,biochemical and phylogenetic assessments of water‐bloom‐forming tropical morphospecies of Microcystis (Chroococcales,Cyanobacteria)

Previous polyphasic analyses of five morphospecies of the water‐bloom‐forming cyanobacterial genus Microcystis, Microcystis aeruginosa (Kützing) Lemmermann (=Microcystis aeruginosa (Kützing) Kützing), Microcystis ichthyoblabe Kützing, Microcystis novacekii (Komárek) Compère, Microcystis viridis (A. Braun) Lemmermann, and Microcystis wesenbergii (Komárek) Komárek in Kondratieva, have shown them to be conspecific and they have been proposed to be included under the binomial Microcystis aeruginosa (Kützing) Lemmermann. However, several morphospecies from tropical regions, such as Microcystis bengalensis Banerji, Microcystis panniformis Komárek, Komárková‐Legnerová, Sant'anna, Azevedo & Senna, Microcystis protocystis Crow, Microcystis pseudofilamentosa Crow, Microcystis ramosa Bharadwaya, and Microcystis robusta (Clark) Nygaard, have never been analyzed biochemically or phylogenetically; consequently, their taxonomic status is uncertain. To resolve this issue, we collected 57 strains of Microcystis from Vietnam for taxonomic analysis using a polyphasic approach. Strains were assigned to the six tropical morphospecies listed above or to four morphospecies with cosmopolitan distributions (M. aeruginosa, M. ichthyoblabe, M. novacekii, and M. wesenbergii). Several strains produced colony variants in different culture media; some of these variants had forms that overlapped with those of other morphospecies. Cell diameters varied widely between strains (2.6–9.3 µm) and were unrelated to morphospecies discrimination criteria. Strains of the 10 morphospecies examined had similar fatty acid compositions and closely similar 16S rRNA gene sequences (>99.2% similar). Phylogenetic analyses using 16S rRNA gene and 16S–23S internal transcribed spacer sequences did not identify any clear separations corresponding to morphospecies concepts or microcystin‐producing abilities. Thus, the six tropical morphospecies (M. bengalensis, M. panniformis, M. protocystis, M. pseudofilamentosa, M. ramosa, and M. robusta) are not natural taxonomic units within the genus Microcystis and should be included under M. aeruginosa.     

Effects of unicellular and colonial forms of toxic Microcystis aeruginosa from laboratory cultures and natural populations on tropical cladocerans

Three life-table experiments, two growth experiments and one feedinginhibition experiment, were performed to study the effects of the toxiccyanobacterium Microcystis aeruginosa on the cladoceransofa tropical lagoon (Jacarepaguá Lagoon, Rio de Janeiro, Brazil).Differentexperimental designs were used to estimate toxic effects of both field samplesand laboratory cultures of Microcystis aeruginosa oncladoceran life history parameters and juvenile growth rates. Effects ofnutritional deficiency could be distinguished from toxic effects in experimentswhere green algae in high carbon concentration were mixed withMicrocystis. Our results show that natural assemblages ofMicrocystis caused much less pronounced toxic effects thanlaboratory cultures and that unicellular forms were more toxic than colonialforms, even though both contained high concentrations of toxins. One possibleexplanation is that colonies were too large to be ingested by the smallMoina micrura and Ceriodaphniacornuta. Feeding inhibition by single cells and small colonies seemsto be another mechanism that contributes to the harmful effects ofMicrocystis on cladocerans, both in the laboratory and inthe field. Thus, caution is needed in extrapolating results from the laboratoryto the field. We did find, however, that toxic algae in natural seston caninhibit growth and reproduction of native cladocerans populations.     

Population dynamics and production of cladoceran zooplankton in the highly eutrophic Lake Kasumigaura

The growth rate, birth rate, death rate and production of the cladocera of Lake Kasumigaura were studied. Standing crop of zooplankton seemed to be governed by predation rather than food. Maximum productivity of cladocerans was observed in late August and early September. There were differences in production between sampling stations. The highest production was recorded in the most eutrophic basin, where heavy water blooms of Microcystis aeruginosa occurred. Maximum secondary production coincided with maximum primary production, which was mainly due to M. aeruginosa. Cladocerans probably utilize decomposed or decomposing Microcystis cells and bacteria in summer. Estimates of annual production of cladocerans varied from 4.2 to 13.1 g dry wt · m^–3, and annual P:B ratios ranged from 36 to 108. The production of cladocerans in Takahamairi Bay was 2.7% of gross primary production.     

Responses of cyanobacteria to herbivorous zooplankton across predator regimes: who mows the bloom?

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Effects of arsenate on the growth and microcystin production of Microcystis aeruginosa isolated from Taiwan as influenced by extracellular phosphate

Arsenic pollution and eutrophication are both prominent issues in the aquaculture ponds of Taiwan. It is important to study the effects of arsenic on algal growth and toxin production in order to assess the ecological risk of arsenic pollution, or at least to understand naturally occurring ponds. The sensitivity of algae to arsenate has often been linked to the structural similarities between arsenate and phosphate. Thus, in this study we examined the effects of arsenate (10⁻⁸ to 10⁻⁴ M) on Microcystis aeruginosa TY-1 isolated from Taiwan, under two phosphate regimes. The present study showed that M. aeruginosa TY-1 was arsenate tolerant up to 10⁻⁴ M, and that this tolerance was not affected by extracellular phosphate. However, it seems that extracellular phosphate contributed to microcystin production and leakage by M. aeruginosa in response to arsenate. Under normal phosphate conditions, total toxin yields after arsenate treatment followed a typical inverted U-shape hormesis, with a peak value of 2.25 ± 0.06 mg L⁻¹ in the presence of 10⁻⁷ M arsenate, whereas 10⁻⁸ to 10⁻⁶ M arsenate increased leakage of ∼75% microcystin. Under phosphate starvation, total toxin yields were not affected by arsenate, while 10⁻⁶ and 10⁻⁵ M arsenate stimulated microcystin leakage. It is suggested that arsenate may play a role in the process of microcystin biosynthesis and excretion. Given the arsenic concentrations in aquaculture ponds in Taiwan, arsenate favors survival of toxic M. aeruginosa in such ponds, and arsenate-stimulated microcystin production and leakage may have an impact on the food chain.     

Influence of Cultivation Parameters on Growth and Microcystin Production of Microcystis aeruginosa (Cyanophyceae) Isolated from Lake Chao (China)

Microcystis aeruginosa isolated in 2005 from the shallow eutrophic Lake Chao (Anhui, China) was investigated in terms of growth parameters and microcystin production under varying nutrient concentrations (P, N) and pH values (abiotic factors) as well as under the influence of spent medium of the non-toxic cyanobacterium Synechocystis sp. (biotic factors). Stimulating effects on growth were observed at the alkaline pH value (10.5), whereas toxin production was significantly increased under phosphate-P limitation (0.6 mg L⁻¹ medium). Within a broad range of nitrate–N concentrations (41.2–247.2 mg L⁻¹ medium), no significant influence on cell growth and microcystin production was observed; however, N-starvation resulted in a typical decrease of growth and toxicity. In addition, cryopreservation of M. aeruginosa evidenced the decrease of toxin production by time-dependent exposure with the cryoprotectant dimethyl sulfoxide under thawing conditions without affecting the growth of the cyanobacterial cells.     

广东省水库微囊藻的产毒特征和ITS序列的遗传多样性分析

为了解广东省水库微囊藻的产毒特征和ITS 序列的遗传多样性,从广东省供水水库中分离得到28 株微囊藻(Microcystisspp.),对它们的产毒特征和15 株微囊藻的ITS 序列进行了分析.高效液相色谱(HPLC)和微囊藻毒素合成酶基因mcyE 的检测结果表明,广东省水库中的微囊藻以产毒藻株占优势,微囊藻毒素的主要类型为MC-RR.广东省15 株藻株的ITS 序列相似性大于93.2%,在用相邻法(NJ)构建的系统树上,不同形态的种和不同地理区域的藻株没有区分开,产毒和非产毒藻株没有形成独立分支.这说明微囊藻ITS 序列的遗传多样性较低,ITS 序列和mcyE 存在没有相关性,表型不能够反映藻株的进化关系.因此,有必要将藻类传统分类方法与分子方法结合起来对蓝藻进行重新分类.     

Effects of UV-B irradiated algae on zooplankton grazing

We tested the effects of UV-B stressed algae on grazing rates of zooplankton. Four algal species (Chlamydomonas reinhardtii, Cryptomonas sp., Scenedesmus obliquus and Microcystis aeruginosa) were used as food and fed to three zooplankton species (Daphnia galeata, Bosmina longirostris and Brachionus calyciflorus), representing different taxonomic groups. The phytoplankton species were cultured under PAR conditions, and under PAR supplemented with UV-B radiation at two intensities (0.3 W m^–2 and 0.7 W m^–2, 6 hours per day). Ingestion and incorporation experiments were performed at two food levels (0.1 and 1.0 mg C l^–1) using radiotracer techniques. The effect of food concentration on ingestion and incorporation rate was significant for all three zooplankton species, but the effect of UV-B radiation was more complex. The reactions of the zooplankton species to UV-B stressed algae were different. UV-B stressed algae did not affect Daphnia grazing rates. For Bosmina the rates increased when feeding on UV-B stressed Microcystis and decreased when feeding on UV-B stressed Chlamydomonas, compared with non-stressed algae. Brachionus grazing rates were increased when feeding on UV-B stressed Cryptomonas and UV-B stressed Scenedesmus, and decreased when feeding on UV-B stressed Microcystis, compared with non-stressed algae. These results suggest that on a short time scale UV-B radiation may result in increased grazing rates of zooplankton, but also in decreased grazing rates. Long term effects of UV-B radiation on phytoplankton and zooplankton communities are therefore difficult to predict.     

Nutrient limitation of Microcystis aeruginosa in northern California Klamath River reservoirs

Nutrient limitations were investigated in Copco and Iron Gate Reservoirs, on the Klamath River in California, where blooms of the toxin-producing cyanobacterium Microcystis aeruginosa were first reported in 2005. Nutrient enrichment experiments conducted in situ in June and August, 2007 and 2008, determined responses in phytoplankton biomass, Microcystis abundance and microcystin concentration to additions of phosphorus and different forms of nitrogen (NH₄⁺, NO₃, and urea). Microcystis abundance was determined using quantitative PCR targeting the phycocyanin intergenic spacer cpcBA.Total phytoplankton biomass increased with additions of N both before and during Microcystis blooms, with no primary effects from P, suggesting overall N limitation for phytoplankton growth during the summer season. NH₄⁺ generally produced the greatest response in phytoplankton growth, while Microcystis abundance increased in response to all forms of N. Microcystis doubling time in the in situ experiments was 1.24–1.39 days when N was not limiting growth. The results from this study suggest availability of N during the summer is a key growth-limiting factor for the initiation and maintenance of toxic Microcystis blooms in Copco and Iron Gate Reservoirs in the Klamath River.