鳢科三种鱼核仁组织者的研究

<正> 1.材料和方法参照Goodpasture的Ag-As法和Bloom,Lau等的Ag法,我们稍加改良应用于研究鱼类的NORs,可获得较好的银染效果.具体方法如下: ①Ag-As法:将干燥后15天左右的片子加4滴AgNO_3溶液(20-30％),盖上长方形的玻片,放在底部铺有润湿滤纸的培养皿中,放于60℃恒温箱中15小时后用自来水

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.

Seasonal variations in stable isotope ratios of two biomanipulation fishes and seston in a large pen culture in hypereutrophic Meiliang Bay, Lake Taihu

This paper reports on seasonal changes in stable carbon and nitrogen isotope ratios of seston and muscle tissue of silver carp and bighead carp during 2004 and 2005, focusing primarily on the carbon sources and trophic relationships among phytoplankton, zooplankton and silver carp and bighead carp in a large fish pen of Meiliang Bay (Lake Taihu, China). delta C-13 showed a minimal value in March 2005 and a maximal value in August 2005 in seston both inside and outside the pen, whereas delta N-15 of seston showed the minimum in winter and the maximum during algal blooms. A positive correlation between delta C-13 of silver carp and that of seston suggested that temporal variation Of delta C-13 in seston was preserved in fish via the food chain. The differences of delta C-13 among seston, zooplankton and muscle tissue of silver carp and bighead carp ranged only 0.2-1.7%, indicating that plankton production was the primary food source of filter-feeding fishes. According to a mass balance model, we estimated that the contributions of zooplankton to the diets of silver carp and bighead carp were 45.7% and 54.3%, respectively, based on the delta N-15 values of zooplankton and planktivorous fishes. (C) 2007 Elsevier B.V. All rights reserved

Rapid quantification of total microcystins in cyanobacterial samples by periodate-permanganate oxidation and reversed-phase liquid chromatography

Microcystins (MCs) comprise a family of more than 80 related cyclic hepatotoxic heptapeptides. Oxidation of MCs causes cleavage of the chemically unique C-20 beta-amino acid (2S, 3S, 8S, 9S)-3-amino-9-methoxy-2,6,8-trimethyl-10-phenyldeca-4,6-dienoic acid (Adda) amino to form 2-methyl-3-methoxy-4-phenylbutanoic acid (MMPB), which has been exploited to enable analysis of the entire family. In the present study, the reaction conditions (e.g. concentration of the reactants. temperature and pH) used in the production of MMPB by oxidation of cyanobacterial samples with permanganate-periodate were optimized through a series of well-controlled batch experiments. The oxidation product (MMPB) was then directly analyzed by high-performance liquid chromatography with diode array detection. The results of this study provided insight into the influence of reaction conditions on the yield of MMPB. Specifically, the optimal conditions, including a high dose of permanganate (>= 50 mM) in saturated periodate solution at ambient temperature under alkaline conditions (pH similar to 9) over 1-4 h were proposed, as indicated by a MMPB yield of greater than 85%. The technique developed here was applied to determine the total concentration of MCs in cyanobacterial bloom samples, and indicated that the MMPB technique was a highly sensitive and accurate method of quantifying total MCs. Additionally, these results will aid in development of a highly effective analytical method for detection of MMPB as an oxidation product for evaluation of total MCs in a wide range of environmental sample matrices, including natural waters, soils (sediments) and animal tissues. (C) 2009 Elsevier B.V. All rights reserved.

Comparative studies on photosynthesis and phosphate metabolism of Cylindrospermopsis raciborskii with Microcystis aeruginosa and Aphanizomenon flos-aquae

The physiological differences for three bloom-forming cyanobacteria (Cylindrospermopsis raciborskii, Microcystis aeruginosa, and Aphanizomenon flos-aquae) were investigated. In comparison with M. aeruginosa and A. flos-aquae, C. raciborskii exhibited a significantly higher concentration of carotenoids, higher values in maximum photosynthesis rate (P-m), apparent photosynthetic efficieny (a), and maximum electron transport rate (ETRmax) during the growth period. In addition, higher extracellular alkaline phosphatase activities and lower light compensation point (I-c) were also detected in C raciborskii (p < 0.05, ANOVA). Therefore, it is suggested that the higher photosynthetic activities, more effective uptake and utilization to phosphate, and low light requirements might play important roles in the occurrence and invasive behavior of C. raciborskii. Crown Copyright (C) 2009 Published by Elsevier B.V. All rights reserved.

Phosphorus in suspended matter and sediments of a hypertrophic lake. A case study: Lake Dianchi, China

From June 2004 to December 2004, Lake Dianchi, which had large scale of cyanobacterial blooms was investigated in order to study P-fractionation in the suspended matter and the sediment. The investigation improves our understanding of phosphorus in Lake Dianchi and the relationship between phosphorus and cyanobacterial blooms. It contributes to the available literature on the behavior of P in hypertrophic lakes. The distribution of P-fractions in Lake Dianchi was not uniform from northwest to south, but was closely related to the trophic status of the whole lake. The concentrations of total phosphorus, labile P (NH4Cl-P), Organic P (NaOH-NRP) and loss on ignition in suspended matter were positively correlated with the strength of cyanobacterial blooms. Total phosphorus in suspended matter was relatively stable for almost half an year and closely related to Chl. a concentration. The main content of organic phosphorus is in the cyanobacterial blooms. The concentrations of phosphorus bound to metal oxides and carbonates (NaOH-SRP and HCl-P) in sediment were similar to NaOH-SRP and HCl-P in the corresponding suspended matter. The latter two forms of P in suspended matter were not affected by cyanobacterial blooms, indicating that the inorganic phosphorus is derived from the sediment after resuspension from the sediment due to wind and wave action. The contribution of the different P-fractions to TP in sediment and in suspended matter indicates that NH4Cl-P in the suspended matter is an important buffer for maintaining dissolved phosphorus in water.

VERTICAL DISTRIBUTIONS OF SIZE-FRACTIONATED IRON IN THREE TYPICAL BAYS OF LAKE DIANCHI (CHINA)

In order to elucidate the vertical distributions of iron in three typical bays (Haigeng bay, Macun bay and Haidong bay) of Lake Dianchi (China), the investigation was conducted on March, 2003. Results showed that the vertical distributions were influenced by monsoon, cyanobacterial bloom and water depth as well as sediment resuspension, which indicated that their translocations and transformations were decided by geographical and physical as well as chemical and biological characteristics.

Feasibility of cyanobacterial inoculation for biological soil crusts formation in desert area

Practical testing of the feasibility of cyanobacterial inoculation to speed up the recovery of biological soil crusts in the field was conducted in this experiment. Results showed that cyanobacterial and algal cover climbed up to 48.5% and a total of 14 cyanobacterial and algal species were identified at the termination of inoculation experiment; biological crusts' thickness, compressive and chlorophyll a content increased with inoculation time among 3 years; moss species appeared in the second year; cyanobacterial inoculation increased organic carbon and total nitrogen of the soil; total salt, calcium carbonate and electrical conductivity in the soil also increased after inoculation. Diverse vascular plant communities composed of 10 and 9 species are established by cyanobacterial inoculation on the windward and leeward surface of the dunes, respectively, after 3 years. The Simpson index for the above two communities are 0.842 and 0.852, while the Shannon-Weiner index are 2.097 and 2.053, respectively. In conclusion, we suggest that cyanobacterial inoculation would be a suitable and effective technique to recover biological soil crusts, and may further restore the ecological system. (C) 2008 Published by Elsevier Ltd.

Identification of Prorocentrum minimum and Takayama pulchella by fluorescence in situ hybridization through epifluorescence microscopy and flow cytometry

Partial rDNA sequences of Prorocentrum minimum and Takayama pulchella were amplified, cloned and sequenced. and these sequence data were deposited in the GenBank. Eight oligonucleotide probes (DNA probes) were designed based on the sequence analysis. The probes were employed to detect and identify P. minimum and T. pulchella in unialgal and mixed algal samples with a fluorescence in situ hybridization method using flow cytometry. Epifluorescence micrographs showed that these specific probes labeled with fluorescein isothiocyanate entered the algal cells and bound to target sequences, and the fluorescence signal resulting from whole-cell hybridization varied from probe to probe. These DNA probes and the hybridization protocol we developed were specific and effective for P. minimum and T. pulchella, without any specific binding to other algal species. The hybridization efficiency of different probes specific to P. minimum was in the order: PM18S02 > PM28S02 > PM28S01 > PM18S01, and that of the probes specific to T. pulchella was TP18S02 > TP28S01 > TP28S02 > TP18S01. The different hybridization efficiency of the DNA probes could also be shown in the fluorescent signals between the labeled and unlabeled cells demonstrated using flow cytometry. The DNA probes PM18S02, PM28S02; TP18S02 and TP28S01, and the protocol, were also useful for the detection of algae in natural samples.

A high throughput Nile red method for quantitative measurement of neutral lipids in microalgae

Isolation of high neutral lipid-containing microalgae is key to the commercial success of microalgae-based biofuel production. The Nile red fluorescence method has been successfully applied to the determination of lipids in certain microalgae, but has been unsuccessful in many others, particularly those with thick, rigid cell walls that prevent the penetration of the fluorescence dye. The conventional "one sample at a time" method was also time-consuming. In this study, the solvent dimethyl sulfoxide (DMSO) was introduced to microalgal samples as the stain carrier at an elevated temperature. The cellular neutral lipids were determined and quantified using a 96-well plate on a fluorescence spectrophotometer with an excitation wavelength of 530 nm and an emission wavelength of 575 run. An optimized procedure yielded a high correlation coefficient (R-2 = 0.998) with the lipid standard triolein and repeated measurements of replicates. Application of the improved method to several green algal strains gave very reproducible results with relative standard errors of 8.5%, 3.9% and 8.6%, 4.5% for repeatability and reproducibility at two concentration levels (2.0 mu g/mL and 20 mu g/mL), respectively. Moreover, the detection and quantification limits of the improved Nile red staining method were 0.8 mu g/mL and 2.0 mu g/mL for the neutral lipid standard triolein, respectively. The modified method and a conventional gravimetric determination method provided similar results on replicate samples. The 96-well plate-based Nile red method can be used as a high throughput technique for rapid screening of a broader spectrum of naturally-occurring and genetically-modified algal strains and mutants for high neutral lipid/oil production. (C) 2009 Published by Elsevier B.V.

Isolation of a novel cyanophage infectious to the filamentous cyanobacterium Planktothrix agardhii (Cyanophyceae) from Lake Donghu, China

A previously unknown cyanophage, PaV-LD (Planktothrix agardhii Virus isolated from Lake Donghu), which causes lysis of the bloom-forming filamentous cyanobacterium P. agardhii, was isolated from Lake Donghu, Wuhan, China. PaV-LD only lysed P. agardhii strains isolated from Lake Donghu and not those isolated from other lakes. The PaV-LD particle has an icosahedral, non-tailed structure, ca. 70 to 85 nm (mean +/- SD = 76 +/- 6 nm) in diameter. PaV-LD was stable at freezing temperature, but lost its infectivity at temperatures >50 degrees C. Lysis of host cells was delayed about 3 d after the PaV-LD treatment with chloroform, and the virus was inactivated by exposure to low pH (<= 4). The latent period and burst size of the PaV-LD were estimated to be 48 to 72 h and about 340 infectious units per cell, respectively. The regrowth cultures of surviving host filaments were not lysed by the PaV-LD suspension. To our knowledge, this is the first isolation and cultivation of a virus infectious to the filamentous bloom-forming cyanobacterium Planktothrix from a freshwater lake.

Grazing on toxic and non-toxic Microcystis aeruginosa PCC7820 by Unio douglasiae and Corbicula fluminea

To explore the potential grazing effects of mussels on Microcystis aeruginosa, a common bloom-forming phytoplankton, Unio douglasiae and Corbicula fluminea were fed with Scenedesmus obliquus, toxic and non-toxic strains of Microcystis aeruginosa as single food and as mixtures in the laboratory. When fed with single foods, U. douglasiae has similar clearance rates on the three algae populations, while C. fluminea has significantly lower clearance rate on toxic M. aeruginosa than those on the other two algae populations. When fed with mixture foods, both the mussels show significantly higher clearance rates than on single foods. The clearance rates of U. douglasiae on the different food mixtures are not significantly different, and C. fluminea has a significantly lower clearance rate on the toxic food mixtures than that on non-toxic food mixtures. Although the relative lower clearance rates of C. fluminea on toxic food, we may still deduce that both the mussels can exert grazing pressure on phytoplankton. The deduction is supported by the composition of the excretion products. The excretion products (faeces and pseudofaeces) of both mussels contained mainly S. obliquus. In both mixed-food treatments, the ratios of S. obliquus to M. aeruginosa in the excrete products are significantly higher than those in the foods. Therefore, it can be concluded that both mussels prefer M. aeruginosa to S. obliquus, and can cause grazing pressure on M. aeruginosa.

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 similar to 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.

EXPERIMENTS ON AQUATIC ECOSYSTEM REGIME SHIFT IN ENCLOSURES NEAR LAKE DIANCHI, CHINA

To discover how a lake converts from a turbid state to clean state, and what drives this process, we constructed controlled enclosure ecosystems and used the ecological remediation method to force ecosystems to convert from the turbid state to the clean state. Our results show that the driving forces include temperature., macrophyte, silver carp and mussel, which form a combined force to drive the controlled ecosystem to switch. There is a threshold existing in treated enclosure ecosystem during the conversion from turbid to clean state. When TP <0.09 mg.L-1, Chl-a <0.036 mg.L-1, transparency >62 cm, TN <2.15 mg.L-1, CODMn <13.7 mg.L-1, tubidity <10, and the number of algal cells <10(6) cells.L-1, the treated ecosystem changes sharply from turbid to clean state. The conversion process can be divided into three phases: turbid state, clean-turbid transitional state as well as clean state, and described with the power function Y = a*X-b (where Y is water parameter, X is time, a and b are constants), which indicates that the shift in the enclosure ecosystem from turbid to clean state is discontinuous.

THE RESPONSES OF Microcystis TO SEDIMENT ENVIRONMENTS AND THE ASSESSMENT FOR ITS OVERWINTERING - A SIMULATION STUDY IN A NOVEL DEVICE

A novel multi-cell device made of organic glass was designed to study morphological and physiological characteristics of Microcystis population trapped in simulated sediment conditions. Changes of colonial morphology and antioxidant activities of the population were observed and measured over the range of 31-day incubation. During the incubation, the antioxidant enzyme activities fluctuated significantly in sediment environments. The activities of catalase (CAT), glutathione peroxidase (GPx) and malondialdehyde (NIDA) reached the highest on the 11(th) day, 6(th) day and 6(th) day. respectively, and then dropped down remarkably in the following days. The ratios of Fv/Fm and the maximal electron transfer rate (ETRm) declined during the initial days (1 similar to 11(th) day), but rebounded on the 16(th) day, which were consistent with the variations of total protein. In the end of incubation. gas vacuoles were hard]), observed and the gelatinous sheath was partly disappeared in the population of Microcystis. Nevertheless, the remaining populations. upon transferred to culture medium, were able to grow though experiencing a longer lag phase of nine days. The results indicated that the sediment environments were able to cause negative effects on M. aeruginosa cells. The cells, however, responded to against the possible damage afterwards. It is thus proposed the acute responses in the population during the early stage of sedimentation could be of importance in aiding the long-term survivor of Microcystis and recruitment in lake sediments. The present study also demonstrated the utility of the device in simulating the sediment environments for further investigation.