Effect of Fe3+ on the growth and lipid content of Isochrysis galbana

Inducing lipid accumulation in microalgae cells without suppressing cell growth is vital to the economical production of biodiesel from microalgae. In two experiments, we demonstrate that the cell concentration and lipid content of marine microalgae Isochrysis galbana depend upon the iron concentration in the growth media. In Experiment I, adding chelated FeCl 3 to the medium at the late exponential growth phase prolonged this phase and increased the lipid content in I. galbana cells. The fi nal cell density and lipid content of I. galbana supplemented with chelated FeCl 3 was approximately 2 and 1.65 times higher than that of non-supplemented cultures, respectively. In Experiment II, I. galbana cells in the late exponential phase were collected and re-inoculated into new media containing Fe 3+ at various concentrations. The fi nal cell concentration and lipid content were maximized at the highest iron concentration(38% biomass by dry weight at 1.2×10-5 mol/L Fe 3+). In this study, intracellular neutral lipid storage was evaluated by fl uorescent spectrophotometry using fl uorochrome Nile red, and the measurement conditions were optimized.     

Establishment and characterization of a new cell line from the kidney of spotted halibut <Emphasis Type="Italic">Verasper variegates</Emphasis>

A cell line,SHK,was derived from the kidney of spotted halibut Verasper variegates.The cell line was subcultured more than 40 passages in minimum essential medium(MEM)supplemented with fetal bovine serum(FBS)and 10 ng ml-1 basic fibroblast growth factor(bFGF).Cell morphology from primary culture and subculture was observed continuously by microscopy.The SHK cell line consisted predominantly of fibroblast-like cells.The cell line was able to grow between 20℃ and 30℃ with the optimum growth at 24℃ and with a reduced growth between 12℃ and 20℃.The growth rate of the cells increased as the proportion of FBS increased from 10% to 20% at 28℃ with optimum growth at the concentration of 20%.The doubling time of the cells was determined to be 44.8 h.Chromosome analysis revealed that 52% of the SHK cells maintained a normal diploid chromosome number (2n=46).The cells were successfully transfected with green fluorescent protein(GFP)reporter plasmids and the expression of GFP gene in the cells indicated the possible utility of the cells in gene expression studies.The cells were infected by lymphosystis disease virus(LCDV)and found to be susceptible to the virus in cytopathic effect(CPE)observation.The infection was confirmed by PCR and electron microscopy experiments,which proved the existence of the viral particles in the cytoplasm of the virus-infected cells.     

Substrate selection and seasonal variation in abundance and size composition of isopod Lirceus fontinalis in Ontario streams, Canada

The abundance and size composition of stream isopods Lirceus fontinalis were investigated from April 2001 to August 2002 in 3 streams in southern Ontario, Canada. Effects of current speed was released from the analysis by choosing slow flowing water. The effects of substrate, season and water depth on the abundance, distribution and size composition of the isopods were analyzed. It was found that substrate and season influence isopods the most. The rocky substrate with attached filamentous macro-algae contains an isopod abundance 7.05 times that of bare rock substrate and 14.6 times that of fine-sand and mud substrate. There was a large variation with respect to seasonality in both abundance and size composition of the isopods, with the highest abundance occurring in summer and the lowest in winter and spring; individual isopods also tend to be larger in the winter and spring. In all substrates, shallow areas tend to support higher densities of isopods than deeper areas.     

Production of dimethylsulfide and acrylic acid from dimethylsulfoniopropionate during growth of three marine microalgae

We measured the concentrations of dimethylsulfide （DMS）, acrylic acid （AA）, and dimethylsulfoniopropionate （DMSP） during growth of three microalgae： Prorocentrum micans, Gephyrocapsa oceanica, and Platymonas subcordiformis. The DMSP, AA, and DMS concentrations in culture media varied significantly among algal growth stages, with the highest concentrations in the late stationary growth stage or the senescent stage. In the stationary growth stage, the average DMSP concentration per cell in P. micans （0.066 5 pmol/cell） was 1.3 times that in G. oceanica （0.049 5 pmol/ cell） and 20.2 times that in P. subcordiformis （0.003 29 pmol/cell）. The average concentrations of AA were 0.044 6, 0.026 9, and 0.003 05 pmol/cell in P. micans, G. oceanica, and P. subcordiformis, respectively, higher than the concentrations of DMS （0.272, 0.497, and 0.086 2 fmol/cell, respectively）. There were significant positive correlations between cell density and AA, DMSP, and DMS concentrations. The ratios of DMS/AA and AA/（DMSP＋AA） in the three algae differed significantly over the growth cycle. In all three microalgae, the DMS/AA ratios were less than 25% during the growth period, suggesting that the enzymatic cleavage pathway, which generates DMS, was not the main DMSP degradation pathway. The changes in the DMS/AA ratio indicated that there was a higher rate of enzymatic breakdown of DMSP in the early growth period and a lower rate during senescence. In all three microalgae, the AA/（DMSP＋AA） ratio （degradation ratio of DMSP） decreased during the exponential growth phase, and then increased. The variations in these ratios can approximately indicate the cleavage mechanism of DMSP at different stages of algal growth.     

Size-fractionated phytoplankton biomass in autumn of the Changjiang （Yangtze） River Estuary and its adjacent waters after the Three Gorges Dam construction

A cruise was undertaken from 3rd to 8th November 2004 in Changjiang （Yangtze） River Estuary and its adjacent waters to investigate the spatial biomass distribution and size composition of phytoplankton. Chlorophyll-a （Chl-a） concentration ranged 0.42-1.17 μg L^-1 and 0.41-10.43 μg L^-1 inside and outside the river mouth, with the mean value 0.73 μg L^-1 and 1.86 μg L^-1, respectively. Compared with the Chl-a concentration in summer of 2004, the mean value was much lower inside, and a little higher outside the river mouth. The maximal Chl-a was 10.43 μg L^-1 at station 18 （122.67°E, 31.25°N）, and the region of high Chl-a concentration was observed in the central survey area between 122.5°E and 123.0°E. In the stations located east of 122.5°E, Chl-a concentration was generally high in the upper layers above 5 m due to water stratification. In the survey area, the average Chl-a in sizes of 〉20 μm and 〈20 μm was 0.28 μg L^-1 and 1.40 μg L^-1, respectively. High Chl-a concentration of 〈20 μm size-fraction indicated that the nanophytoplankton and picophytoplankton contributed the most to the biomass of phytoplankton. Skeletonema costatum, Prorocentrum micans and Scrippsiella trochoidea were the dominant species in surface water. The spatial distribution of cell abundance of phytoplankton was patchy and did not agree well with that of Chl-a, as the cell abundance could not distinguish the differences in shape and size of phytoplankton cells. Nitrate and silicate behaved conservatively, but the former could probably be the limitation factor to algal biomass at offshore stations. The distribution of phosphate scattered considerably, and its relation to the phytoplankton biomass was complicated.     

Occurrence and distribution of dissolved tellurium in Changjiang River estuary

With the implementation of the GEOTRACES program, the biogeochemical cycle and distribution of tellurium （Te） in marine environments are becoming increasing environmental concerns. In this study, the concentration of dissolved Te in the Changjiang （Yangtze） River estuary and nearby waters was determined in May 2009 by hydride-generation atomic fluorescence spectrometry to elucidate the abundance, dominant species, distribution, and relationship with environmental factors. Results show that： （1） dissolved Te was low owing to its low abundance in the Earth＇s crust, high insolubility in water, and strong affinity to particulate matter; （2） Te（IV） and Te（VI） predominated in surface water. Te（VI） was the dominant species in bottom water, and Te（IV） was the minor species; （3） Horizontally, resulting from low phytoplankton metabolism and the weak reduction from Te（VI） to Te（IV） in the shore, Te（IV） was concentrated in the central zone instead of the coastal region. However, Te（VI） was abundant near the mouth of the Changjiang River where the Changjiang water is diluted and in the area to the south where the Taiwan Warm Current invaded. In the adsorption-desorption process, Te（IV） was negatively related to suspended paniculate matter （SPM）, indicating that it was adsorbed by particulate matter. While for Te（VI）, the positive correlation with SPM suggested that it was desorbed from the solid phase. In the estuary, dissolved Te had a negative correlation to salinity. However, it deviated from the dilution line in high-salinity regions due to the invasion of the Taiwan Warm Current and the mineralization of organic matter. The relationship between Te（IV） and SPM nutrients indicated that it was more bioavailable and more related to phosphorus than to nitrogen. Progress in the field is slow and more research is needed to quantify the input of Te to the estuary and evaluate the biochemical role of organisms.     

Seasonal Variations in Phytoplankton Community Structure in the Sanggou,Ailian, and Lidao Bays

The seasonal variations in phytoplankton community structure were investigated for the Sanggou Bay （SGB） and the adjacent Ailian Bay （ALB） and Lidao Bay （LDB） in Shandong Peninsula,eastern China.The species composition and cell abundance of phytoplankton in the bay waters in spring （April 2011）,summer （August 2011）,autumn （October 2011）,and winter （January 2012） were examined using the Uterm6hl method.A total of 80 taxa of phytoplankton that belong to 39 genera of 3 phyla were identified.These included 64 species of 30 genera in the Phylum Bacillariophyta,13 species of 8 genera in the Phylum Dinophyta,and 3 species of 1 genus in the Phylum Chrysophyta.During the four seasons,the number of phytoplankton species （43） was the highest in spring,followed by summer and autumn （40）,and the lowest number ofphytoplankton species （35） was found in winter.Diatoms,especially Paralia sulcata （Ehrenberg） Cleve and Coscinodiscus oculus-iridis Ehrenberg,were predominant in the phytoplankton community throughout the study period,whereas the dominance of dinoflagellate appeared in summer only.The maximum cell abundance of phytoplankton was detected in summer （average 8.08 × 103 cells L-1） whereas their minimum abundance was found in autumn （average 2.60 x 103 cellsL-1）.The phytoplankton abundance was generally higher in the outer bay than in the inner bay in spring and autumn.In summer,the phytoplankton cells were mainly concentrated in the south of inner SGB,with peak abundance observed along the western coast.In winter,the distribution of phytoplankton cells showed 3 patches,with peak abundance along the western coast as well.On seasonal average,the Shannon-Wiener diversity indices of phytoplankton community ranged from 1.17 to 1.78 （autumn 〉 summer 〉 spring 〉 winter）,and the Pielou＇s evenness indices of phytoplankton ranged from 0.45 to 0.65 （autumn 〉 spring 〉 summer〉 winter）.According to the results of canonical correspondence analysis,phosphate level w     

A Model Describing the Time Course and Variability in Toxicity of Hydrophobic Chemicals to Aquatic Organisms-Toxicity of chlorinated benzenes to marine algae

The growth of Chlorella marine, Nannochloris oculate, Pyramimonaos sp., Platymonas subcordiformis and Phaeodactylum tricornutum exposed to chlorobenzene, 1,2-dichlorobenzene, 1,2,3,4-tetrachlorobenzene and pentachlorobenzene was tested. The Boltzman equation was used to describe organism growth. The time course for uptake of hydrophobic organic chemicals (HOCs) by aquatic organisms was expressed by incorporating growth and, if desired, the effect of metabolism into the HOC bioconcentration process. The probability of any given concentration of HOCs in the organisms causing a specified toxic endpoint was expressed with a modified Weibull distribution function. The combined bioconcentration and probability equations were tested with data for time course of incubation of algae exposed to chlorinated benzenes (CBs). A set of parameters, including the uptake rate constant k1, the elimination rate constant k2 and thereafter the bioconcentration factor on a dry weight basis, BCFD, the critical HOC concentration in the organism resulting in a specified toxic endpoint, CA^*, and the spread factor, S, could be obtained by fitting only experimental data for percent growth inhibition(%)-time-CB exposure concentration. The average coefficients of variation within CBs were 15.2% for BCFD, 21.0% for k1, 18.3% for k2, 8.1% for CA^* and 9.7% for S. The variability in toxicity (such as EC10, EC50, ECg0) derived from the model equations agreed well with those experimentally observed.     

Characterization and Expression Analysis of Protein Kinase C Gene from Dunaliella salina

Protein kinase C(PKC) has a crucial role in signal transduction for a variety of biologically active substances which activate cellular functions and proliferation. We previously isolated the full-length PKC gene from Dunaliella salina(Ds PKC) using rapid amplification of cDNA ends(RACE) and RT-PCR methods. And we submitted the mRNA sequence of Ds PKC gene to NCBI(Genbank No. JN625213). In the present paper, the Ds PKC gene open reading frame obtained by PCR was cloned into pGS-21 a vector and transformed into Escherichia coli to generate the fusion protein. Bioinformatics analysis revealed that DsPKC gene was a member of serine/threonine kinase with two conserved domains and highly conserved motifs. The Ds PKC was highly expressed upon induction with isopropyl-β-d-thiogalactoside(IPTG) at a final concentration of 0.2 mmol L~(-1) at 37℃. Under salt stress, the fusion protein Green Fluorescent Protein(GFP)-DsPKC was transferred from the cytoplasm to the cell membrane. The expression pattern of Ds PKC gene was analyzed using real-time quantitative PCR, and indicated that Ds PKC gene was up-regulated by 3.0 mol L~(-1) NaCl at 12 h, which was significantly higher than in control values(P 0.05). These results suggest that the Ds PKC gene plays an important role in response to salt stress in D. salina.     

Interannual and Seasonal Variability of Winter-Spring Cohort of Neon Flying Squid Abundance in the Northwest Pacific Ocean During 1995–2011

The neon flying squid, Ommastrephes bartramii, is a species of economically important cephalopod in the Northwest Pacific Ocean. Its short lifespan increases the susceptibility of the distribution and abundance to the direct impact of the environmental conditions. Based on the generalized linear model(GLM) and generalized additive model(GAM), the commercial fishery data from the Chinese squid-jigging fleets during 1995 to 2011 were used to examine the interannual and seasonal variability in the abundance of O. bartramii, and to evaluate the influences of variables on the abundance(catch per unit effort, CPUE). The results from GLM suggested that year, month, latitude, sea surface temperature(SST), mixed layer depth(MLD), and the interaction term(SST×MLD) were significant factors. The optimal model based on GAM included all the six significant variables and could explain 42.43% of the variance in nominal CPUE. The importance of the six variables was ranked by decreasing magnitude: year, month, latitude, SST, MLD and SST×MLD. The squid was mainly distributed in the waters between 40?N and 44?N in the Northwest Pacific Ocean. The optimal ranges of SST and MLD were from 14 to 20℃ and from 10 to 30 m, respectively. The squid abundance greatly fluctuated from 1995 to 2011. The CPUE was low during 1995–2002 and high during 2003–2008. Furthermore, the squid abundance was typically high in August. The interannual and seasonal variabilities in the squid abundance were associated with the variations of marine environmental conditions and the life history characteristics of squid.     

Intraspecific genetic analysis, gamete release performance, and growth of Sargassum muticum （Fucales, Phaeophyta） from China

Sargassum muticum is one of the most abundant and conspicuous native macroalgae species on the northern coasts of China. It often forms large-scale seaweed beds in subtidal zones. This investigation was designed to understand the intraspecific genetic relationships of this alga based on samples collected from four northern coastal sites of China, and to evaluate gamete release and growth capacity in laboratory conditions. The nuclear internal transcribed spacer 2 （ITS2） sequences of 16 samples from four locations were identical. Based on cox3 gene and partial rbcLS operon sequences, intraspecific genetic variability was detected with three and two ribotypes, respectively. Temperature, not irradiance, was shown to significantly affect gamete release and fertilization. Elevated temperature and irradiance enhanced the growth of germlings and vegetative branchlets. Maximum growth rate of germlings was detected at 18-24℃and an irradiance of 60-100 μmol photons/（m^2.s）. Under ambient conditions （12-25℃ and 60-125 gmol photons/ （m2.s））, relative growth rate of young branchlets could reach 7.5%/d.     

Vitreoscilla hemoglobin gene (vgb) improves lutein production in Chlorella vulgaris

Vitreoscilla hemoglobin is an oxygen-binding protein that promotes oxygen delivery and reduces oxygen consumption under low oxygen conditions to increase the efficiency of cell respiration and metabolism. In this study, we introduced a Vitreoscilla hemoglobin gene （vgb） into Chlorella vulgaris by Agrobacterium tumefaciens-mediated transformation （ATMT）. PCR analysis confirmed that the vgb gene was successfully integrated into the Chlorella vulgaris genome. Analysis of biomass obtained in shake flasks revealed transformant biomass concentrations as high as 3.28 g/L, which was 38.81% higher than that of the wild-type strain. Lutein content of transformants also increased slightly. Further experiments recovered a maximum lutein yield of 2.91 mg/L from the transformants, which was 36.77% higher than that of the wild-type strain. The above results suggest that integrated expression of the vgb gene may improve cell growth and lutein yield in Chlorella vulgaris, with applications to lutein production from Chlorella during fermentation.     

Tidal effects on diel variations of picoplankton and viruses in the Changjiang estuary

Spatial distributions and seasonal variations of picoplankton (i.e. Synechococcus spp., Prochlorococcus spp., picoeukaryotes and heterotrophic bacteria) and viruses in the Changjiang estuary have been reported in the past. However, short-term variations (e.g. at a tidal timescale) of these organisms and their regulating factors remain unclear. We determined the time-series of fluctuations of picoplankton and viruses with tide simultaneously in flow cytometry in the Changjiang estuary during a cruise in June 2006, in which a tidal model based rectangle equation was applied. The results indicate that high cell abundances of picoplankton and viruses occurred during flood tide and low cell abundances during ebb tide. The period of the surface cell abundance variations was about 13 h, suggesting the surface cell abundances in the Changjiang estuary were largely regulated by tide. However, cell abundances in middle and bottom waters varied in different periods due to influences of tidal induced physical forces such as resuspension and stratification. Therefore, tidal action is an important factor for the diel variations of picoplankton and viruses in the Changjiang estuary.     

Distributions of Picophytoplankton and Phytoplankton Pigments Along a Salinity Gradient in the Changjiang River Estuary,China

We investigated the abundance of different picophytoplankton groups and the phytoplankton pigment ratio in relation to environmental factors such as nutrients and suspended solids along a salinity gradient in the Changjiang River Estuary.The average numbers of Synechococcus spp.（Syn） and picoeukaryotes （Euk） were （2.7 ±5.1）×l03 and （1.1±1.4）×l03 cells mL-1,respectively.Prochlorococcus spp.（Pro） was only found in the high-salinity brackish water with the concentration of 3.0× 10^3 cells mL-1.Syn and Euk numbers both tended to increase offshore and Syn showed a larger variation in cell abundance than Euk.The contribution of picophytoplankton to total phytoplankton biomass increased with increasing salinity and decreasing nutrient concentrations from the estuary to the open ocean.The response of different picophytoplankton groups to environmental variables was different.Water temperature was more important in its control over Euk than over Syn,while nutrients were more important in their influence over Syn than over Euk.Phytoplankton pigment ratios were different in the three different ecological zones along the salinity gradient （i.e.,freshwater zone with 0-5 range,fresh and saline water mixing zone with 5-20 range,and high-salinity brackish water zone with 20-32 range）,where three different phytoplankton communities were discovered,suggesting that phytoplankton pigment ratios can be considered as a complementary indicator of phytoplankton community structure in the Changjiang River Estuary.