Yu Yuhe

Genetic fingerprinting of plankton community provides new insights into aquatic ecology

Over the past two decades, molecular techniques have been widely used in ecological study and molecular ecology has been one of the most important branches of ecology. Meanwhile, genetic fingerprinting analyses have significantly enhanced our knowledge of the diversity and evolutionary relations of the planktonic organisms. Compared with conventional approaches in ecological study (e. g. morphological classification), genetic fingerprinting techniques are simpler and much more effective. This review provides an overview of the principles, advantages and limitations of the commonly used DNA fingerprinting techniques in plankton research. The aim of this overview is to assess where we have been, where we are now and what the future holds for solving aquatic ecological problems with molecular-level information.

Fish gut microecosystem: a model for detecting spatial pattern of microorganisms

The spatial distribution pattern of organisms is a basic issue in understanding the mechanisms of community assembly. Although the spatial distributions of animals and plants have been well studied, those of microorganisms are still being debated. In this study, we used a fish gut microecosystem to detect the spatial pattern of microbes, because it can provide a relatively unified and stable environment. Results suggest that the turnover of intestinal bacterial assemblages showed a weak but highly significant negative correlation between similarity and distances in the microbial community, in respect of both grass carp intestinal loci distances and the geographical distance between fish sampling sites. Our results also suggest that intestinal bacterial assemblages responded to differences within the external environment and within different parts of the fish themselves. These results show that some, or possibly all, microbes are restricted in their distribution and that environmental factors are also important influences on the structure of intestinal bacterial assemblages. The fish gut microecosystem is useful in promoting study of the spatial distribution patterns of microorganisms.

Preliminary study on applicability of microsatellite DNA primers from parasite protozoa Trypanosoma cruzi in free-living protozoa

In this paper, we took the lead in studying on specificity of the microsatellite DNA loci and applicability of microsatellite DNA primers in protozoa. In order to study characters of microsatellites in free-living protozoa, eight microsatellite loci primers developed from Trypanosoma cruzi (MCLE01, SCLE10, MCLE08, SCLE11, MCLF10, MCLG10, MCL03, MCL05) were employed to amplify microsatellite in four free-living protozoa, including Bodo designis, Euglena gracilis FACHB848, Paramecium bruzise and Tetrahymena thermophila BF1. In the amplification systems of P. bruzise, four loci (SCLE10, SCLE11, MCLF10, MCL03) were amplified successfully, and four amplification fragments were in proper size. In genome of E. gracilis FACHB848, five of eight primers brought five clear amplification bands. In B. designis, three (No.4, 5 and 7) of eight loci produced clear and sharp products without stutter bands, whereas no bands appeared in T. thermophila BF1. Further, eight 300–500 bp amplification fragments were cloned and sequenced. Nevertheless, all sequenced products did not contain corresponding microsatellite sequence, although Bodo is in the same order and has the nearest phylogenetic relation with Trypanosoma among these four species. Thus, the microsatellite DNA primers can not be applied among order or more far taxa, and the specificity of microsatellite DNA is very high in protozoa. The results of this study will contribute to our understanding of microsatellite DNA in protozoa.

Phylogenetic relationships among six species of Epistylis inferred from 18S-ITS1 sequences

Phylogenetic relationships among six species of Epistylis (i. e. E. plicatilis, E. urceolata, E. chrysemydis, E. hentscheli, E. wenrichi, and E. galea) were investigated using sequences of the first internal transcribed spacer region (ITS-1) of ribosomal DNA (rDNA). Amplified rDNA fragment sequences consisted of 215 or 217 bases of the flanking 18S and 5.8S regions, and the entire ITS-1 region (from 145 to 155 bases). There were more than 33 variable bases between E. galea and the other five species in both the 18S region and the ITS-1 region. The affiliation of them was assessed using Neighbor-joining (NJ), maximum parsimony (MP) and maximum likelihood (ML) analyses. In all the NJ, MP and ML analyses E. galea, whose macronucleic position and shape are distinctly different from those of the other five species, was probably diverged from the ancestor of Epistylis earlier than the other five species. The topology in which E. plicatilis and E. hentscheli formed a strongly supported sister clade to E. urceolata, E. chrysemydis, and E. wenrichi was consistent with variations in the thickness of the peristomial lip. We concluded that the macronucleus and peristomial lip might be the important phylogenetic characteristics within the genus Epistylis.

Eutrophication control using modified local soil/sand induced ecological restoration technology: I. Effect and mechanism on short and long term improvement of water quality

2010年10月-2011年9月在太湖梅梁湾围隔内研究了改性当地土壤絮凝除藻及其对水质改善的应急和长期效果,并结合室内实验研究了该技术防控底泥再悬浮和减少底泥二次污染的长效机制.现场围隔实验结果表明,改性当地土壤除藻30 min后,TN、NO3--N、NH4+-N、TP、PO34--P和Chl.a的去除率分别为66%、57%、60%、93%、92%和98%;长期监测结果表明,与对照区域相比,围隔内的TN、NH4+-N、NO3--N、TP和PO34--P在处理后11个月内的平均值分别降低了39.83%、52.30%、48.53%、18.75%和60.00%.室内再悬浮实验结果表明,改性土壤和沙子抗再悬浮能力较未改性土壤分别提高了3和5倍.室内柱培养结果表明改性土壤絮凝除藻和沙土覆盖相结合可有效提高表层沉积物-水界面的氧化还原电位和溶解氧,使沉积物向水体的TP和TN通量从源逆转成汇,PO34--P和NH4+-N通量大幅度降低.改性土壤技术在利用絮凝除藻快速改善水质后,可通过改性沙/土分层底泥调控分别达到对藻絮体再悬浮的物理控制和营养盐再释放的化学控制,通过将亚表层底泥中的藻细胞分解并被沉水植物根系吸收,可实现对底泥中水华蓝藻复苏和水体富营养化的长效生态控制.The effect of algal bloom control and water quality improvement using modified local soil/sand induced ecological restoration technology(MLS-IER) was studied in an enclosure in Meiliang Bay in the northern Lake Taihu from October 2010 to September 2011.Lab experiments were used to study the mechanism of long-term water quality improvement including resuspension and internal loads control at water-sediment interfaces.Field monitoring results indicated that,TN,NO-3-N,NH+4-N,TP and PO3-4-P were reduced by 66%,57%,60%,93%,92% and 98%,respectively,30min after treatment using MLS;and the average concentrations were reduced by 39.83%,48.53%,52.30%,18.75% and 60.00%,respectively,within the following 11 months compared to the control area.The resuspension experiment showed that the anti-resuspension ability of modified soil and sand increased by a factor of 3 and 5 compared to unmodified soil.The sediment-water incubation experiment showed that algae flocculation followed by capping with MLS remarkably increased the ORP and DO at the sediment-water interface,which reversed the source of sediment-water flux of TP and TN into a sink and remarkably reduced the flux of PO3-4-P and NH+4-N from sediment to water column.The results suggested that,after nutrients were quickly removed from the water by flocculating the algal blooms,it is possible to physically reduce the resuspension of the algae flocs and chemically block the release of nutrients from the sediment by MLS capping.The sustainable water quality improvement and reduction of algal bloom recruitment can be achieved when the buried algae banks are decomposed and absorbed by the restoration of submerged vegetations in shallow waters,which is triggered by the MLS-IER technology.

The annual fluctuation of zooplankton community and its relation with environmental fac-tors in Lake Xiliang,Hubei Province

Four zooplanktonic investigations were carried out from November 2007 to October 2008 in Lake Xiliang.Totally,224 species of zooplankton including 112 species of protozoa,90 species of rotifer,15 species of cladocera,and 7 species of copepoda were found.The zooplankton species ranged from 75 to 111,with the lowest value(75) occurring in winter,and the highest one(111) in autumn.The density of zooplankton was the highest in winter(126731ind./L),and the lowest in spring(71356ind./L).The densities of zooplankton in summer and autumn were 109743ind./L and 107650ind./L,respectively,and the abundance of protozoa and rotifer were on predominance.The structure of zooplankton community was analyzed in relation to water quality parameters by canonical correspondence analysis(CCA).Temperature,pH,water depth,and the concentrations of TP,COD and DO were strongly correlated with the zooplankton community structure.The parameters that demonstrated a strong correlation with the zooplankton community structure differed among seasons,suggesting that the seasonal variations of zooplankton community structure were related closely to both the water temperature and water trophic status in Lake Xiliang.

Relationship between DNA fingerprinting structure of plankton community and physicochemical factors in Lake Niushan

Plankton are considered good indicators of water quality because they are highly sensitive to the environmental variation in their short life spans. However, traditional studies on plankton community constructure have depended on taxonomic identification which has historically been a difficult task. Recently, different fingerprinting techniques have been developed and applied successfully to analyze the community structure of bacterial and picoplankton. However, little is known to the whole plankton community. In this study, we analyze DNA polymorphism of plankton communities in five different sampling stations of Lake Niushan by RAPD and PCR-DGGE and explore the relationship between DNA fingerprinting structure of plankton communities and environmental physico-chemical factors. The results were: (i) 9 of 40 screened random primers used in the study amplified a total of 93 observable bands, 58 of which were polymorphic; the mean number of amplified bands at different sampling stations was 67, and station V had the maximum number (74) and station I had the minimum number (61); (ii) a total of 102 bands (56 of 16S rDNA bands and 46 of 18S rDNA bands) were detected by PCR-DGGE, station III, IV, and V had more bands, station I and II had few band; (iii) total phosphorus (TP), chlorophyll-a, chemical oxygen demanded (COD), hardness, and conductance rate at station II were the highest; dissolved oxygen (DO) and pH value were not obviously different among five stations. Based on physico-chemical factors, station I, III, IV, and V were grouped into a cluster, station II grouped into a single cluster. Based on RAPD markers and PCR-DGGE markers, five sampling stations can be grouped into two clusters: station I and II were grouped into a cluster, and station III, IV, and V were grouped into the other cluster. In conclusion, the DNA fingerprinting structure of plankton community was closely related to total phosphorus (TP). DNA based community level analysis is a fast, easy, accurate method to reflect the water quality, and such data accumulation will play an important role in constructing one easy and sensitive molecular system for water quality assessment.

Seasonal variation of plankton communities influenced by environmental factors in an artificial lake

We evaluated the seasonal variation in plankton community composition in an artificial lake. We conducted microscopic analysis and denaturing gradient gel electrophoresis (DGGE) of PCR-amplified partial 16S rRNA and 18S rRNA genes to characterize the plankton community. The clustering of unweighted pair group method with arithmetic mean (UPGMA) was then used to investigate the similarity of these plankton communities. DGGE fingerprinting revealed that samples collected at the different sites within a season shared high similarity and were generally grouped together. In contrast, we did not observe any seasonal variation based on microscopic analysis. Redundancy analysis (RDA) of the plankton operational taxonomic units (OTUs) in relation to environmental factors revealed that transparency was negatively correlated with the first axis (R=−0.931), and temperature and total phosphorus (TP) were positively correlated with the first axis (R=0.736 and R=0.660, respectively). In conclusion, plankton communities in the artificial lake exhibited significant seasonal variation. Transparency, phosphorus and temperature appear to be the major factors driving the differences in plankton composition.

DNA polymorphism and species composition of plankton in an artificial lake for breeding transgenic fish

The present study was designed to explore relationship between DNA polymorphism, as revealed by RAPD and PCR-DGGE fingerprintings, and species composition of plankton community in an artificial lake for breeding transgenic fish. Planktonic samples were collected from five sampling sites in the artificial lake, and their similarity was assessed with cluster analysis. The results were: (1) a total of 44 planktonic taxa were identified, among which there were 13 algae, 11 protozoa, 16 rotifer, 2 Cladocera and Copepoda, respectively; (2) 128 observable bands were totally amplified by the 9 random printers screened in RAPD analysis, among which 61.7% and 19.5% were polymorphic bands and specific bands, respectively; (3) there were 87 distinct bands identified by the method of PCR-DGGE, among which 50 were 16S rRNA bands and 37 were 18S rRNA bands. Despite the analysis of morphological identification and DNA fingerprintings had shown a higher species diversity, the cluster of similarity was different: based on species composition, group 1 consisted of B, C sites, group 2 contained D, E sites, and group 3 only comprised of A site. However, according to the DNA fingerprint analysis, the plankton community of C, D, E were similar, and another two sites-A and B were similar. In summary, our results suggested that the species composition was not closely related to the DNA polymorphism of plankton community, and the latter can describe more species. However, the characteristics of biodiversity revealed front different aspects will provide basic data for elucidating the structure and function of plankton community in the ecosystem.

A preliminary study of single cell PCR in protozoa: take Stentor coeruleus for an instance

Taking Stentor coeruleus for an instance, we have tested single cell PCR for single gene and whole genome. After optimization, we got perfect results. In the single cell PCR for single gene reaction, 40 fresh cells and 40 neutral red stained cells all got 100% success and rich products, and 40 cells kept in the alcohol (95%) for one week got 82.5% success. In the single cell PCR for whole genome reaction, we employed phi 29 DNA Polymerase and random primer for our PCR amplification and got rich products. Using whole genome amplification (10 times diluted) 4 gene loci were amplified, and all of them got objective gene section successfully.