EFFECTS OF IRON ON PHYSIOLOGICAL AND BIOCHEMICAL CHARACTERISTICS OF Microcystis wesenbergii (Kom.) Kom. (Cyanobacterium)

Iron is an essential trace element for biological requirements of phytoplankton. Effects of iron on physiological and biochemical characteristics of Microcystis wesenbergii were conducted in this study. Results showed that 0.01 mu M [Fe3+] seriously inhibited growth and chlorophyll synthesis of M. wesenbergii, and induced temporary increase of ATPase activities, however, NR. ACP and ALP activities were restrained by iron limitation. Interestingly, iron addition on day 8 resulted in the gradual restoration of structures and functions of above enzymes and resisted a variety of stresses from iron limitation. M. wesenbergii in 10 mu M [Fe3+] treatment group grew normally. enzymes maintained normal levels, and residual phosphate contents in cultures first sharply decreased, then smoothly as M. wesenbergii has a characteristic of luxury consumption of phosphorus. Above parameters in 100 mu M [Fe3+] treatment group were almost same with those in 10 mu M [Fe3+] treatment group except for NR, ACP and ALP activities. In 100 mu M [Fe3+] treatment group, activities of ACP and ALP had temporary increase because phosphate and ferric iron could form insoluble compound - ferric phosphate (Fe3PO4) through adsorption effect. resulting in lack of bioavailable phosphate in culture media. The experiment suggested that too low or too high iron can affect obviously physiological and biochemical characteristics of M. wesenbergii.

Non-microcystin producing Microcystis wesenbergii (Komarek) Komarek (Cyanobacteria) representing a main waterbloom-forming species in Chinese waters

It is well known that several morphospecies of Microcystis, such as Microcystis aeruginosa (Kutzing) Lemmermann and Microcystis viridis (A. Brown) Lemmermann can produce hepatotoxic microcystins. However, previous studies gave contradictory conclusions about microcystin production of Microcystis wesenbergii (Komarek) Komarek. In the present study, ten Microcystis morphospecies were identified in waterblooms of seven Chinese waterbodies, and Microcystis wesenbergii was shown as the dominant species in these waters. More than 250 single colonies of M. wesenbergii were chosen, under morphological identification, to examine whether M. wesenbergii produce hepatotoxic microcystin by using multiplex PCR for molecular detection of a region (mcyA) of microcystin synthesis genes, and chemical analyses of microcystin content by ELISA and HPLC for 21 isolated strains of M. wesenbergii from these waters were also performed. Both molecular and chemical methods demonstrated that M. wesenbergii from Chinese waters did not produce microcystin. (C) 2007 Elsevier Ltd. All rights reserved.

Effects of iron on growth, pigment content, photosystem II efficiency, and siderophores production of Microcystis aeruginosa and Microcystis wesenbergii

Changes in growth, photosynthetic pigments, and photosystem II (PS II) photochemical efficiency as well as production of siderophores of Microcystis aeruginosa and Microcystis wesenbergii were determined in this experiment. Results showed growths of M. aeruginosa and M. wesenbergii, measured by means of optical density at 665 nm, were severely inhibited under an iron-limited condition, whereas they thrived under an iron-replete condition. The contents of chlorophyll-a, carotenoid, phycocyanin, and allophycocyanin under an iron-limited condition were lower than those under an iron-replete condition, and they all reached maximal contents on day 4 under the iron-limited condition. PS II photochemical efficiencies (maximal PS II quantum yield), saturating light levels (I-k ) and maximal electron transport rates (ETRmax) of M. aeruginosa and M. wesenbergii declined sharply under the iron-limited condition. The PS II photochemical efficiency and ETRmax of M. aeruginosa rose , whereas in the strain of M. wesenbergii, they declined gradually under the iron-replete condition. In addition, I-k of M. aeruginosa and M. wesenbergii under the iron-replete condition did not change obviously. Siderophore production of M. aeruginosa was higher than that of M. wesenbergii under the iron-limited condition. It was concluded that M. aeruginosa requires higher iron concentration for physiological and biochemical processes compared with M. wesenbergii, but its tolerance against too high a concentration of iron is weaker than M. wesenbergii.

微囊藻碳酸酐酶活性在不同环境因素下的调节与适应

测定了3种微囊藻水华中的优势种类,即铜锈微囊藻(Microcystis aeruginosaK櫣tz .) ,绿色微囊藻(Microcystis viridis(A. Br .)Lemm) ,惠氏微囊藻(Microcystis wesenbergii(Kom.)Kom.) ,以及微囊藻573(Microcystissp.573)的碳酸酐酶活性;研究了无机碳、pH、温度、光强、N/P比等环境因素和外源葡萄糖对铜锈微囊藻碳酸酐酶活性的影响,发现微囊藻碳酸酐酶活性受环境中碳酸氢根浓度的调节,故推断碳酸氢根是铜锈

二种淡水微囊藻rDNA16S-235基因间隔区的序列测定与分析

本研究采用ＰＣＲ及序列测定的方法，对我国淡水铜绿微囊藻有毒株（Ｍ８６４１）和另一低毒的种类惠氏微囊藻（Ｍ５７４）ｒＤＮＡ１６Ｓ－２３Ｓ基因间隔区进行了序列的测定和分析，结果表明：ｒＤＮＡ１６Ｓ－２３Ｓ基因间隔区可以作为一个精细且稳定的指标，用于微囊藻的分类和鉴定。并从分子水平提出了铜绿微囊藻与惠氏微囊藻在种系发生上有较近缘的关系。本文首次对微囊藻属Ｍｉｃｒｏｃｙｓｔｉｓ　ｒＤＮＡ基因间隔区全序列作了报道，为微囊藻属的鉴定及系统学研究提供了分子基础。

Mechanism study on the frequent variations of cell-bound microcystins in cyanobacterial blooms in Lake Taihu: Implications for water quality monitoring and assessments

Although Microcystis-based toxins have been intensively studied, previous studies using laboratory cultures of Microcystis strains are difficult to explain the phenomenon that microcystin concentrations and toxin variants in natural blooms differ widely and frequently within a short-term period. The present study was designed to unravel the mechanisms for the frequent variations of intracellular toxins related to the differences in cyanobacterial colonies during bloom seasons in Lake Taihu, China. Monitoring of Microcystis colonies during warm seasons indicated that the variations in microcystins in both concentrations and toxin species were associated with the frequent alteration of Microcystis colonies in Lake Taihu. High concentration of microcystins in the blooms was always associated with two Microcystis colonies, Microcystis flos-aquae and Microcystis aeruginosa, whereas when Microcystis wesenbergii was the dominant colonial type, the toxin production of the blooms was low. Additionally, environmental factors such as temperature and nutrition were also shown to have an effect on the toxin production of the blooms, and may also potentially influence the Microcystis species present. The results of the present study provides insight into a new consideration for quick water quality monitoring, assessment and risk alert in cyanobacterium- and toxin-contaminated freshwaters, which will be beneficial not only for water agencies but also for public health. (C) 2009 Elsevier Ltd. All rights reserved.

Ação alelopática de saxitoxinas e microcistinas em cianobactérias e microalgas verdes em condições controladas

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Identification of triosephosphate isomerase (TIM) genes from Microcystis (Cyanobacteria : Chroococcales) and theoretical analyses of the potential of cyanobacterial TIM as a target for designing specific inhibitors

The genes encoding triosephosphate isomerase (TIM) in three species of Microcystis (M. aeruginosa, M. viridis and M. wesenbergii) were investigated. Reverse transcriptase-polymerase chain reaction indicated that they were transcribed in the cells. Analyses showed that their DNA and deduced amino acid sequences were highly conserved between all the three species, only a single nonsynonymous substitution was seen at position 31, from an Asp in M. aeruginosa and M. viridis to Glu in M. wesenbergii. Sequence alignment of these with 12 other known cyanobacterial TIM sequences showed that all the cyanobacterial TIMs had a very high level of amino acid identity (over 50% between each two). Comparison of the cyanobacterial TIMs with other reported TIMs (from diverse lineages of the three Domains) showed that they possessed common active-site residues and sequence motifs. All cyanobacterial TIMs have two common cysteine residues (Cys127 and Cys176), and the Cys176 is almost cyanobacteria-specific with only one exception in Streptomyces coelicolor. Both secondary structure alignment and comparative modelling of Synechocystis sp. TIM showed that Cys176 was located at the hinge region of the flexible loop-6 and might therefore be critical to the movement of TIM's loop-6, which is important to the function of the enzyme. Thus, the cyanobacterial TIM-specific Cys176 may be a potential site for the discovery of suitable drugs against cyanobacteria, and such drugs may have utility in controlling water blooms due to cyanobacteria.

Taxonomic notes on water bloom forming Microcystis species (Cyanophyta) from China - An example from samples of the Dianchi Lake

Ten common species of Microcystis, based on the examination of water samples from the Dianchi Lake, Yunnan, China, were morphologically described, and their taxonomy was also discussed. They are Microcystis aeruginosa, M botrys, M firma, M flos-aquae, M ichthyoblabe, M novacekii, M pseudofilamentosa, M smithii, M viridis and M wesenbergii. Taxonomic status of other Microcystis species reported in China was also evaluated.

Genetic variability of Brazilian strains of the Microcystis aeruginosa complex (Cyanobacteria/Cyanophyceae) using the phycocyanin intergenic spacer and flanking regions (cpcBA)

The genetic and morphological variability among 15 Brazilian strains of Microcystis aeruginosa (Kütz.) Kütz. collected from four locations was examined and compared with several reference strains of M. aeruginosa, M. viridis (A. Br.) Lemm. and M. wesenbergii (Kom.) Kom. in Kondr. Brazilian strains were classified by morphological features and by comparison of the nucleotide sequences of the cpcBA intergenic spacer and flanking regions. Our results indicate that Brazilian strains classified as M. aeruginosa are phylogenetically diverse compared with reference strains of M. aeruginosa and that the current taxonomy underestimates genetic diversity within M. aeruginosa. The data also demonstrate that morphological criteria alone are inadequate to characterize Microcystis species. Although colonial characters were shown to vary considerably in culture, some genetic lineages demonstrated consistent cellular diameter ranges, indicating that cell size has value as a taxonomic character. The detection of six M. aeruginosa genotypes in a single water body indicates that morphological approaches can also seriously underestimate the diversity of Microcystis bloom populations.

The effects of environmental factors on biomass and microcystin production by the freshwater cyanobacterial genera Microcystis and Anabaena

Several cyanobacterial genera produce the hepatotoxins, microcystins. Microcystins are produced only in cells that have microcystin synthetase gene (mcy) clusters, which encode enzyme complexes involved in microcystin biosynthesis. Microcystin-producing and nonmicrocystin-producing genotypes of single cyanobacterial genus may occur simultaneously in situ. Previously, the effects of environmental factors on the growth and microcystin production of cyanobacteria have mainly been studied by means of isolated cyanobacteria cultures in the laboratory. Studies in the field have been difficult, owing to the lack of methods to identify and quantify the different genotypes. In this study, genus-specific microcystin synthetase E (mcyE) gene primers were designed and a method to identify and quantify the mcyE copy numbers was developed and used in situ. Microcystis and Anabaena mcyE genes were observed in two Finnish lakes. Microcystis appeared to be the most abundant microcystin producer in Lake Tuusulanjärvi and in one basin of Lake Hiidenvesi. Because the most potent microcystin-producing genus of a lake can be identified, it will be possible in the future to design genus-targeted strategies for lake restoration. Effects of P and N concentrations on the biomass of microcystin-producing and nonmicrocystin-producing Microcystis strains and an Anabaena strain were studied in cultures. P and N concentrations and their combined effect increased cyanobacterial biomass of all Microcystis strains. The biomass of microcystin-producing Microcystis was higher than that of nonmicrocystin-producing strains at high nutrient concentrations. The P concentration increased Anabaena biomass, but the effect of N concentration was statistically insignificant for growth yield, probably due to the ability of the genus to fix molecular N2. P and N concentrations and combined nutrients caused an increase in cellular microcystin concentrations of the Microcystis strain cultivated in chemostat cultures. Cyanobacteria are able to hydrolyse nutrients from organic matter through extracellular enzyme activities. Leucine aminopeptidase (LAP) activity was observed in an axenic N2-fixing Anabaena strain grown in batch cultures. The P concentration caused a statistically significant increase in LAP activity, whereas the effect of N concentration was insignificant. The highest LAP activities were observed in the most eutrophic basins of Lake Hiidenvesi. LAP activity probably originated mostly from attached heterotrophic bacteria and less from cyanobacteria.

Avaliação dos efeitos da disponibilidade de luz sobre o crescimento e competição entre cepas de Planktothrix agardhii e Microcystis aeruginosa

As cianobactérias Microcystis aeruginosa e Planktothrix agardhii são espécies formadoras de florações comuns em ecossistemas aquáticos eutrofizados. Nestes ambientes, a disponibilidade de luz é um dos fatores determinantes para o desenvolvimento e estruturação da comunidade fitoplanctônica. O presente estudo teve como objetivo investigar o efeito da luz na fisiologia de cepas de cianobactérias (M. aeruginosa e P. agardhii), avaliando o crescimento, a variabilidade inter e intra-específica e a competição por luz. Para tanto foram realizados cultivos estanques em diferentes intensidades luminosas (10, 40, 60, 100 e 500 mol m-2 s-1) e calculadas as taxas de crescimento e os rendimentos máximos das culturas. O requerimento mínimo de luz de cada cepa foi determinado em experimentos com monoculturas em sistemas de cultivo contínuo (quimiostatos) sob condições de limitação de luz. A competição por luz foi avaliada através de experimentos com biculturas em quimiostatos. Foi observada variabilidade intra e inter-específica das cepas, nas diferentes intensidades luminosas testadas. Em 500 μmol m-2 s-1, as cepas de M. aeruginosa obtiveram maior biomassa do que P. agardhii, corroborando a maior sensibilidade de P. agardhii luz. Embora com rendimento máximo menor, P. agardhii cresceu em intensidades luminosas consideradas elevadas para a espécie, 100 e 500 μmol m-2 s-1. Estes resultados evidenciam a capacidade de P. agardhii ocorrer em ambientes com grandes amplitudes de luminosidade. Na intensidade de 10 μmol m-2 s-1, M. aeruginosa e P. agardhii apresentaram crescimento semelhante, demonstrando a habilidade das duas espécies em crescer com pouca luz. Nas monoculturas em quimiostato, sob condições de limitação de luz, as cepas de M. aeruginosa atingiram maior biomassa durante o equilíbrio (steady-state) do que P. agadhii, refletindo uma capacidade suporte mais elevada, enquanto que os valores de requerimento mínimo de luz foram semelhantes entre as duas espécies. Ao competirem, M. aeruginosa superou P. agardhii imediatamente após o início do experimento. Esse rápido crescimento resultou na dominância de M. aeruginosa em todos os pares de cepas testados e, em dois casos, ocorreu exclusão competitiva de P. agardhii. Quando não ocorreu exclusão, P. agardhii conseguiu manter-se no sistema com uma baixa biomassa (ca.15%). Estes resultados ajudam a entender a co-ocorrência destas espécies no ambiente e a dominância de M. aeruginosa mesmo em condições de baixa luminosidade.

Global transcriptional responses of the toxic cyanobacterium, Microcystis aeruginosa, to nitrogen stress, phosphorus stress, and growth on organic matter

Whole transcriptome shotgun sequencing (RNA-seq) was used to assess the transcriptomic response of the toxic cyanobacterium Microcystis aeruginosa during growth with low levels of dissolved inorganic nitrogen (low N), low levels of dissolved inorganic phosphorus (low P), and in the presence of high levels of high molecular weight dissolved organic matter (HMWDOM). Under low N, one third of the genome was differentially expressed, with significant increases in transcripts observed among genes within the nir operon, urea transport genes (urtBCDE), and amino acid transporters while significant decreases in transcripts were observed in genes related to photosynthesis. There was also a significant decrease in the transcription of the microcystin synthetase gene set under low N and a significant decrease in microcystin content per Microcystis cell demonstrating that N supply influences cellular toxicity. Under low P, 27% of the genome was differentially expressed. The Pho regulon was induced leading to large increases in transcript levels of the alkaline phosphatase phoX, the Pst transport system (pstABC), and the sphX gene, and transcripts of multiple sulfate transporter were also significantly more abundant. While the transcriptional response to growth on HMWDOM was smaller (5–22% of genes differentially expressed), transcripts of multiple genes specifically associated with the transport and degradation of organic compounds were significantly more abundant within HMWDOM treatments and thus may be recruited by Microcystis to utilize these substrates. Collectively, these findings provide a comprehensive understanding of the nutritional physiology of this toxic, bloom-forming cyanobacterium and the role of N in controlling microcystin synthesis.

不同氮磷营养条件下铜绿微囊藻(Microcystis aeruginosa)对正磷酸盐的蓄积效果

铜绿微囊藻是中国湖泊、水库及其它水域生态系统发生、形成富营养化危害的主要藻类。采用了直接显色法对单细胞铜绿微囊藻(Microcystis aeruginosa)蓄积正磷酸盐的浓度进行测定,并与传统测定方法进行比较。直接显色法测定的磷浓度值减去溶液中可溶性磷浓度值即得到铜绿微囊藻蓄积的磷浓度值。对用不同处理方式处理过的铜绿微囊藻藻液进行过滤显色和直接显色测定,结果表明聚磷酸盐(正磷酸盐)在铜绿微囊藻体内不是游离存在,而很可能是与细胞内某一活性部分结合。在确立测定方法的基础上,研究了以修改了的MⅢ培养基为基本

铜绿微囊藻(Microcystis aeruginosa)与三种丝状蓝藻间的相互作用

研究了单细胞铜绿微囊藻和三种丝状蓝藻(水华束丝藻、水华鱼腥藻及土生席藻)间的相互作用,包括以下两个方面的内容:①铜绿微囊藻细胞滤出液对水华束丝藻、水华鱼腥藻及土生席藻生长的影响;②水华束丝藻、水华鱼腥藻及土生席藻细胞滤出液对铜绿微囊藻生长的影响．研究发现,当滤出液浓度为60％(滤出液与BG11的体积比为3:2)时,制绿微囊藻细胞滤出液对水华束丝藻、水华鱼腥藻的生长有显著促进效果,尤其对水华束丝藻的作用更加明屁;对土生席藻的生长却起着微弱的抑制作用,仅表现于100%细胞滤出液中,对铜绿微囊藻而言,土生席藻细