Song Lirong

Quantitative assessment of toxic and nontoxic Microcystis colonies in natural environments using fluorescence in situ hybridization and flow cytometry

Toxic cyanobacterial blooms constitute a threat to human safety because Microcystis sp. releases microcystins during growth, and particularly during cell death. Therefore, analysis of toxic and nontoxic Microcystis in natural communities is required in order to assess and predict bloom dynamics and toxin production by these organisms. In this study, an analysis combining fluorescence in situ hybridization (FISH) with flow cytometry (FCM) was used to discriminate between toxic and nontoxic Microcystis and also to quantify the percentage of toxic Microcystis present in blooms. The results demonstrate that the combination of FISH and flow cytometry is a useful approach for studying the ecology of Microcystis toxin production and for providing an early warning for toxic Microcystis blooms.

THE SPATIO-TEMPORAL PATTERN OF PHYTOPLANKTON IN THE NORTH BASIN OF LAKE DIANCHI AND RELATED ENVIRONMENTAL FACTORS

To research spatio-temporal pattern of phytoplankton in the north basin of Lake Dianchi and to find out the key factor that influence the pattern,investigations were conducted 24 times at 6 sampling sites from December 2008 to June 2009.The composition and densities of phytoplankton were measured and canonical correspondence analysis(CCA) was applied to explore the relationship between phytoplankton species and the environmental parameters.By comparing annual monitoring data in Dianchi Lake,chlorophyll a concentration in the north basin started to decline at mid-December,reached the minimum(26.5 μg/L) in mid-February and the maximum(530.5 μg/L) in mid-May.A total of 97 phytoplankton taxon,belonging to 74 genera were identified,consisting of 53 species of green algae,20 species of blue-green algae,17 species of diatoms,2 species of cryptomonad,2 species of both golden algae and euglenoids,and only 1 species of dinoflagellates.Cyanobacterial biomass was dominant in December 2008 and from March 2009 to June 2009.Dominant species of Cyanobacteria were Microcystis viridis in December 2008,Aphanizomenon flos-aquae from March to May 2009,and Microcystis wesenbergii in June 2009.Diatoms peaked in January and dominant species were Melosira granulata var.angustissima.The results showed that the biomass of Microcystis increased rapidly when the water temperature was higher than 14℃;the biomass of Aphanizomenon increased with temperature while decline rapidly when water temperature was higher than 22℃.Phytoplankton distribution at the 6 sampling sites in the north basin of the Dianchi Lake was significantly different.Biomass of Microcystis and Aphanizomenon at D1 and D4 were significantly higher than that of other four sampling sites(P0.01),while biomass of D2 and D5 were significantly higher than that of the other four sampling sites(P0.01).The results indicated that,in the severe eutrophic north basin of Lake Dianchi,water temperature may be the key factor affecting the phytoplankton composition and biomass;the meteorological factors,together with uneven nutrients concentration,were considered to be the main driving forces determining the horizontal distribution of phytoplankton.

Historical changes of phytoplankton functional groups in Lake Fuxian,Lake Erhai and Lake Dianchi since 1960s

Phytoplankton is one of the major primary producers and plays important roles in the aquatic ecosystems. The temporal and spatial variations of the algae could reflect the environmental dynamics. In light of the morphological,physiological and ecological characteristics of algae,phytoplankton functional groups theory was applied first time for these lakes to describe the tolerance and preference of algae in detail. The phytoplankton assemblages were considered to be good indicators of environmental conditions. The data of phytoplankton composition dated from the 1960 s was collected and analyzed for Lake Fuxian,Lake Erhai and Lake Dianchi,respectively. Great changes have taken place in phytoplankton assemblages in the three lakes during the past 60 years. The historical change of dominant phytoplankton functional groups was demonstrated as C,X2,Lo,F,P( 1960s)-H1,C,P( 1980s)- P,C,T( 1990s)-T( after 2000) for Lake Fuxian; J,Lo,MP,C,H1( 1960s)-C,H1( 1980s)-C,H1( 1990s)-C,H1,M( after 2000) for Lake Erhai; J,N,P,MP,Lo( 1960s)-J,P,MP,M( 1980s)-J,M( 1990s)-M( after 2000) for Lake Dianchi. The reduction of low-temperature tolerance species and increase of high-nutrition preference species dated from the 1960 s might give some indication of the environmental changes of warmer temperature and nutrients increases in the three lakes.Phytoplankton is one of the major primary producers and plays important roles in the aquatic ecosystems. The temporal and spatial variations of the algae could reflect the environmental dynamics. In light of the morphological,physiological and ecological characteristics of algae,phytoplankton functional groups theory was applied first time for these lakes to describe the tolerance and preference of algae in detail. The phytoplankton assemblages were considered to be good indicators of environmental conditions. The data of phytoplankton composition dated from the 1960 s was collected and analyzed for Lake Fuxian,Lake Erhai and Lake Dianchi,respectively. Great changes have taken place in phytoplankton assemblages in the three lakes during the past 60 years. The historical change of dominant phytoplankton functional groups was demonstrated as C,X2,Lo,F,P( 1960s)-H1,C,P( 1980s)- P,C,T( 1990s)-T( after 2000) for Lake Fuxian; J,Lo,MP,C,H1( 1960s)-C,H1( 1980s)-C,H1( 1990s)-C,H1,M( after 2000) for Lake Erhai; J,N,P,MP,Lo( 1960s)-J,P,MP,M( 1980s)-J,M( 1990s)-M( after 2000) for Lake Dianchi. The reduction of low-temperature tolerance species and increase of high-nutrition preference species dated from the 1960 s might give some indication of the environmental changes of warmer temperature and nutrients increases in the three lakes.

Improving photosynthesis of microalgae by changing the ratio of light-harvesting pigments

Changing the ratio of light-harvesting pigments was regarded as an efficient way to improve the photosynthesis rate in microalgae, but the underlying mechanism is still unclear. In the present study, a mutant of Anabeana simensis (called SP) was selected from retrieved satellite cultures. Several parameters related with photosynthesis, such as the growth, photosynthesis rate, the content of photosynthetic pigment, low temperature fluorescence spectrum (77K) and electron transport rate, were compared with those of the wild type. It was found that the change in the ratio of light-harvesting pigments in the mutant led to more efficient light energy transfer and usage in mutant than in the wild type. This may be the reason why the mutant had higher photosynthesis and growth rates.

Effects of copper sulfate treatment on eutrophic urban lake phytoplankton communities.

Copper sulphate treatment is widely used as a global and empirical method to remove or control phytoplankton blooms. We tested the acute toxicity of 5 species of familiar microalgae and gave an urban lake copper sulfate treat in order to illuminate how the water body response to copper treat, especially the change of phytoplankton species and density of microcystins. According to OECD Alga growth inhibition test, effect of copper on 5 species of familiar microalgae was tested, and species of Cyanophyta showed much higher sensitivity to copper than species of Chlorophyta and Bacillariophyta. In a eutrophic urban lake, CuSO4 center dot 5H(2)O was used as algicide to control the water bloom caused by cyanobacterial. The copper concentration applied was 102 mu g/L ( as copper). We investigated the lake response to copper sulfate I which showed that the transparence was substantially improved; TN and TP did not change a lot; total algal decreased just after the copper sulfate treatment, and later increased again; at the beginning of the experiment, species of Cyanophyta died and the amount sharply decreased, Bacillariophyta and Chlorophyta turned to be the preponderant species, which could be related to species sensitivity to copper, and species of Cyanophyta began to grow again and took the predominance few days later; density of microcystins in the water rapidly decreased to level lower than that in the treatment before, only in 4d.

An inducible CO2 concentrating mechanism in cyanobacteriumAnabaena sp. strain PCC7120

In order to define its characteristics of the photosynthetic utilization of CO2 and HCO3- when the ambient inorganic carbon changed, HCG (High-CO2-Growing Cells) of cyanobacteriumAnabaena sp. strain PCC7120 were prepared. The growth rate of HCG was higher than that of LCG (low-CO2-growing cells, i.e. air-growing cells). When the HCG cells were transferred from 5% CO2 to air levels of CO2, a series of changes took place: its carbonic anhydrase activity as well as its photosynthetic affinity to the external inorganic carbon significantly increased; the number of the carboxysomes, which is one of the most important components of CCM in cyanobacteria also increased. These facts indicated that the CCM activity ofAnabaena PCC 7120 was induced. When the pH in the medium increased from 6 to 9, the photosynthetic affinity to external inorganic carbon of both HCG and LCG declined, while the apparent photosynthetic affinity to external CO2 increased. In the light of these findings, this inducible CCM in cyanobacteria provided a good model for the study of the photosynthetic Ci utilization in the phototrophic microoganisms.

Physiological response to low nitrogen level in Aphanizomenon flos-aquae from Lake Dianchi

Two strains of Aphanizomenon flos-aquae were isolated from spring cyanobacterial blooms in Haigeng Bay which located in the northern part of Lake Dianchi.Both the growth characteristics and heterocyst formation as well as nitrogen fixation ability of A.flos-aquae were investigated under different low concentrations of sodium nitrate,and in nitrogen-free culture medium,respectively.The results showed that A.flos-aquae strains were able to grow in the BG-11 medium with various low concentrations of nitrate,and the biomass reached a very high level.The biomass increased with the nitrate-nitrogen concentration increasing,but there was no significant difference when nitrate-nitrogen concentration was above 0.5 mg / L.Induction of heterocyst differentiation was rapid when A.flos-aquae was cultured in N-deficient BG-11 medium,and after reaching a maximum of 72% approximately,the filament with heterocyst began to swing lower.The total nitrogen in nitrogen-free medium increased gradually as nitrogen fixation proceed,and nitrogen concentration was close to 30 mg / L at day 43.Two strains of Aphanizomenon flos-aquae were isolated from spring cyanobacterial blooms in Haigeng Bay which located in the northern part of Lake Dianchi.Both the growth characteristics and heterocyst formation as well as nitrogen fixation ability of A.flos-aquae were investigated under different low concentrations of sodium nitrate,and in nitrogen-free culture medium,respectively.The results showed that A.flos-aquae strains were able to grow in the BG-11 medium with various low concentrations of nitrate,and the biomass reached a very high level.The biomass increased with the nitrate-nitrogen concentration increasing,but there was no significant difference when nitrate-nitrogen concentration was above 0.5 mg / L.Induction of heterocyst differentiation was rapid when A.flos-aquae was cultured in N-deficient BG-11 medium,and after reaching a maximum of 72% approximately,the filament with heterocyst began to swing lower.The total nitrogen in nitrogen-free medium increased gradually as nitrogen fixation proceed,and nitrogen concentration was close to 30 mg / L at day 43.

ANALYSIS OF OFF-FLAVORS IN FISH BY MICROWAVE MEDIATED DISTILLATION WITH HEADSPACE SOLID-PHASE MICROEXTRACTION AND GAS CHROMATOGRAPHY-MASS SPECTRUM

Aquaculture the rearing of fish and other aquatic animals is the best approach for meeting the increasing demand for aquaculture products worldwide. However, the problem of off-flavor has been documented as a serious setback to aquaculture. Though there are many off-flavors can result in a fish being objectionable, approximately 80% of the rejected fish have the earthy-musty odor, which is primary caused by the presence of either 2-methylisoborneol (2-MIB) or geosmin. Based on the fact that the detection of earthy-musty odorous compounds in fish mainly relied on the sensory method in China, the present study established the instrumental method to detect 2-MIB and geosmin by microwave mediated distillation with headspace solid-phase microextraction and gas chromatography-mass spectrum (MWDE-SPME-GC-MS). Through investigating the effects of the key parameters, microwave processing time and carrier gas flow rate, in microwave mediated distillation on extracts, it is concluded that 6 ruin of microwave time and 70 mL/min nitrogen flow were the optimized condition on which earthy-musty odorous compounds could be completely distilled from fish flesh. Then the earthy-musty odorous compounds were absorbed in the fiber under HS-SPME, and were finally desorbed at 250 degrees C and determined by GC-MS. The limits of detection for 2-MIB and geosmin in fish were both 0.1 mu g/kg. There were significant linear correlation in the range of 1-20 mu g/kg for the two earthy-musty odorous compounds, with calibration coefficients at 0.987 and 0.995, respectively. Therefore, trace levels of earthy-musty odorous compounds at ppb in fish could be quantified by this method with credible results.

Physiological aspects of a high-co2 requiring mutant and the high-co2 growing cells of the cyanobacteriumSynechococcus pcc7942

A new chemically mutagenic mutant ofSynechococcus PCC7942 named high-CO2 requiring mutant, which could grow at 4% CO2 but could not grow at air levels of CO2, was isolated. Comparative studies on some physiological aspects of the mutant and high-CO2 growing cells (growing at 4% CO2) were conducted. The result showed that the mutant had lower growing rate, about 1/40th photosynthetic affinity to inorganic carbon, 25% lower carbonic anhydrase (CA) activity, lower quenching rate of chlorophyll fluorescence, and about 1/2 alkalinization rate of the medium. The CA activity responses of the two types of cells to different concentration of CO2 were determined. Upon the addition, of inorganic carbon (Ci), the rate of active Ci uptake described by the rate of chlorophyll fluorescence quenching of the mutant was obviously lower compared with that of the high-CO2 growing cells; the size of the internal inorganic carbon pool size detemined by the extent of fluorescence quenching of the mutant was also smaller.

Recent progress in research of the biological function of microcystins

Cyanobacterial bloom has become a frequent and problematic feature of many freshwater bodies over a wide geographical area, especially in China. Microcystis species are widely known for their production of the potent hepatotoxins called microcystins (MCs) . While the toxicity of MCs on animals, including humans, has been well studied, the biological role of MCs on the organ⁃ ism from which they originate has not been clearly elucidated. In this paper, we summarized MCs production, especially MCs bio⁃ logical function with the development of technology in recent years. We reviewed and discussed the biological role of MCs in four as⁃ pects: Serving as allelochemical, involving in photosynthesis, taking part in the wintering and recovery of Microcystis and in the sustaining and formation of Microcystis colonies. In addition, MCs may act as signal molecule inside and outside Microcystis cells, relevant research has also been reviewed. Global climatic change which favour cyanobacterial blooms in eutrophic waters, may also influence the competition between MC⁃producing and non⁃MC⁃producing strains and their dominance. The study on the competition between MC⁃producing and non⁃MC⁃producing strains and the biological role of protein⁃bound MCs should be focused on for further investigation, fast development of variable ⁃omics would favor the investigation.