Population genetic structure of an endemic cyprinid fish, Ancherythroculter nigrocauda, in the upper reaches of the Yangtze River and its implication for conservation

Ancherythroculter nigrocauda is a cyprinid fish endemic to the upper reaches of the Yangtze River, which has been reported to have 2 or 3 chambers to its air bladder. Morphological studies showed no differences between individuals with different types of air bladder, but did demonstrate geographical differences from different sources. After the completion of the Three Gorges Dam, it was expected that the population of this species would decrease, but artificial breeding and stocking is under consideration to protect this species from extinction. In the present study, mtDNA cytochrome b gene sequences were determined and analyzed for A. nigrocauda samples of different morphotypes and sources to identify their genetic differentiations, and thereby guide plans for the artificial propagation and conservation of this species. Haplotype diversity index values (h) and nucleotide diversity values (pi) for all the populations were found to be high indicating their high level genetic diversity. An analysis of molecular variance identified no differentiation among the studied populations. Therefore, we suggested that the individuals of different morphological types and geographical sources belong to the same species. To maintain its high level genetic diversity, it mill he necessary to use large and diverse sources of parental fish for artificial reproduction.

Using epilithic diatom communities to assess ecological condition of Xiangxi River system

Assessment method for ecological condition of Xiangxi River system was studied by using 13 candidate metrics of epilithic diatom which can reflect conditions in pH, salinity, nitrogen uptake metabolism, oxygen requirements, saprobity, trophic state, morphological character and pollution tolerant capability etc. By one-way ANOVA, the metrics of relative abundance of acidobiontic algae (ACID), freshwater algae (FRESH), high oxygen requirement (HIGH-O), eutraphentic state (EUTRA) and mobile taxa ( MOBILE) were suitable for distinguishing sites in different conditions. Then, the river diatom index (RDI) composed of these five metrics was used to evaluate ecological condition of the river. The results showed that the healthiest sites were in the Guanmenshan Natural Reserve ( with the mean RDI of 79.73). The sites located in tributary of Jiuchong River also owned excellent state (mean RDI of 78.25). Mean RDI of another tributary - Gufu River and the main river were 70.85 and 68.45 respectively, and the unhealthiest tributary was Gaolan River (with mean RDI of 65.64). The mean RDI for all the 51 sites was 71.40. The competence of RDI was discussed with comparison of evaluation results of DAIpo and TDI, it can be concluded that multimetrics is more competent in assessment task.

Acid volatile sulfide and simultaneously extracted metals in the sediment cores of the Pearl River Estuary, South China

The acid-volatile sulfide (AVS), simultaneously extracted metals (SEM), total metals, and chemical partitioning in the sediment cores of the Pearl River Estuary (PRE) were studied. The concentrations of total metals, AVS, and SEM in the sediment cores were generally low in the river outlet area, increased along the seaward direction, and decreased again at the seaward boundary of the estuary. The amounts of AVS were generally greater in deeper sediments than in surface sediments. SEM/AVS was > 1 in the surface sediments and in the river outlet cores. The ratio was < 1 in the sediments down the profiles, suggesting that AVS might play a major role in binding heavy metals in the deep sediments of the PRE. The SEM may contain different chemical forms of trace metals in the sediments, depending on the metal reaction with 1 M cold HCl in the AVS procedure compared with the results of the sequential chemical extraction. The SEM/AVS ratio prediction may overestimate trace metal availability even in the sediments with high AVS concentrations. (c) 2004 Elsevier Inc. All rights reserved.

Distribution of the macroinvertebrate communities in the Xiangxi River system and relationships with environmental factors

Surveys of macroinvertebrates were carried out in the Xiangxi River system during July of 2001. Among the 121 taxa collected, Ephemeroptera, Trichoptera, and Diptera dominated (41.7, 26.0, and 24.5% of the total relative abundance, respectively). Two-way indictor species analysis and detrended correspondence analysis divided the 49 sites into four groups based on species composition and relative abundance. Canonical correspondence analysis indicated that elevation, SiO2, pH, conductivity, hardness, and NO2-N were significant environmental factors affecting the distribution of macroinvertebrates.

Mobilization potential of heavy metals: A comparison between river and lake sediments

Toxic metals introduced into aquatic environments by human activities accumulation in sediments. A common notion is that the association of metals with acid volatile sulfides (AVS) affords a mechanism for partitioning metals from water to solid phase, thereby reducing biological availability. However, variation in environmental conditions can mobilize the sediment-bound metal and result in adverse environmental impacts. The AVS levels and the effect of AVS on the fate of Cu, Cd, Zn, Ni in sediments in the the Changjiang River, a suboxic river with sandy bottom sediment and the Donghu Lake, a anoxic lake with muddy sediment in China, were compared through aeration, static adsorption and release experiments in laboratory. Sips isotherm equation, kinetic equation and grade ion exchange theory were used to describe the heavy metal adsorb and release process. The results showed that AVS level in the lake sediment are higher than that of the river. Heavy metals in the overlying water can transfer to sediments incessantly as long as the sediment remains undisturbed. The metal release process is mainly related to AVS oxidation in lake sediment while also related to Org-C and Fe-Mn oxyhydroxide oxidation in river sediment. The effect of sulfides on Zn and Ni is high, followed by Cd, and Cu is easy bound to Org-C. AVS plays a major role in controlling metals activity in lake sediment and its presence increase the adsorption capacity both of the lake and river sediments.

Species richness in a riparian plant community along the banks of the Xiangxi River, the Three Gorges region

The distribution of vascular plant species richness along an altitudinal gradient and their relationships with environmental variables, including slope, aspect, bank (flooding) height, and river width of the Xiangxi River, Hubei Province, were examined. Total vascular plant species richness changed with elevation: it increased at lower elevations, reached a maximum in the midreaches and decreased thereafter. In particular, tree and herbaceous species richness were related to altitude. Correlation analysis (Kendall's tau) between species richness and environmental variables indicated that the change in species richness in the riparian zone was determined by riparian environmental factors and characteristics of regional vegetation distribution along the altitudinal gradient. The low species richness at lower elevations resulted from seasonal flooding and human activities - agriculture and fuel collection - and the higher. Species richness ill (he midreaches reflected transitional zones ill natural vegetation types that had had little disturbance. These results oil species distribution in the riparian community could he utilized as a reference for restoration efforts to improve water quality of the emerging reservoir resulting from the Three Gorges Hydroelectric Dam project.

Molecular cloning of TRAF2 binding protein gene and its promoter region from the grass carp Ctenopharyngodon idellus

A tumor necrosis factor receptor-associated factor 2 binding protein (T2BP) gene was isolated from the grass carp (Ctenopharyngodon idellus) by utilizing suppression subtractive hybridization (SSH) and rapid amplification of cDNA ends (RACE). The grass carp T2BP (GT2BP) gene contains an open reading frame of 579 nucleotide(s) (nt), encoding 193 amino acids, with 23 nt 5'-untranslated region and a long 3'-untranslated region of 434 nt including poly (A), 1 AUUUA motif and 4 AUUUUA motifs. No signal peptide has been detected in the predicted GT2BP, but a characteristic forkhead associated domain is present. The GT2BP mRNA shares 83% identity with the zebrafish DNA sequence, and they both have no introns in the genomic DNA. The putative transcription factor binding sites of GT2BP include two C/EBP alpha binding sites, and one c-Jun binding, one AP-1 binding, and one nuclear factor kappa B (NF kappa B) binding sites. Southern blot analysis revealed that the GT2BP was a single-copy gene. Individual difference was observed in GT2BP expression in examined organs of healthy grass carp. However, the expression of GT2BP in all examined organs in a fish with the highest copepod infection level and the significantly higher expression level in spleen and liver in infected fish may indicate its up-regulation with the parasite infection. (c) 2005 Elsevier B.V. All rights reserved.

Characterization of two genes encoding leucine-rich repeat-containing proteins in grass carp Ctenopharyngodon idellus

The cDNAs and genes of two different types of leucine- rich repeat-containing proteins from grass carp ( Ctenopharyngodon idellus) were cloned. Homology search revealed that the two genes, designated as GC-GARP and GC-LRG, have 37% and 32% deduced aminoacid sequence similarities with human glycoprotein A repetitions predominant precursor ( GARP) and leucine-rich alpha2-glycoprotein (LRG), respectively. The cDNAs of GC-GARP and GC-LRG encoded 664 and 339 amino acid residues, respectively. GC-GARP and GC-LRG contain many distinct structural and/or functional motifs of the leucine- rich repeat (LRR) subfamily, such as multiple conserved 11-residue segments with the consensus sequence LxxLxLxxN/CxL ( x can be any amino acid). The genes GC-GARP and GC-LRG consist of two exons, with 4,782 bp and 2,119 bp in total length, respectively. The first exon of each gene contains a small 5'-untranslated region and partial open reading frame. The putative promoter region of GC-GARP was found to contain transcription factor binding sites for GATA-1, IRF4, Oct-1, IRF-7, IRF-1, AP1, GATA-box and NFAT, and the promoter region of GC-LRG for MYC-MAX, MEIS1, ISRE, IK3, HOXA9 and C/EBP alpha. Phylogenetic analysis showed that GC-GARP and mammalian GARPs were clustered into one branch, while GC-LRG and mammalian LRGs were in another branch. The GC-GARP gene was only detected in head kidney, and GC-LRG in the liver, spleen and heart in the copepod ( Sinergasilus major)- infected grass carp, indicating the induction of gene expression by the parasite infection. The results obtained in the present study provide insight into the structure of fish LRR genes, and further study should be carried out to understand the importance of LRR proteins in host - pathogen interactions.

Benthic algae of the Xiangxi River, China

From June 2000 to June 2002, four sites on the mainstem of the Xiangxi River and one site on each of its major tributaries were sampled 16 times each for benthic algae. All total, 223 taxa (most to species and variety levels) were found (193 Bacillariophyta, 20 Chlorophyta, nine Cyanophyta and one Xanthophyta). The diatoms Cocconeis placentula, Achnanthes linearis, and Diatoma vulgare dominated the system, with relative abundance of 33.3%, 18.8%, and 6.4%, respectively. The abundances of all the other taxa were under 5%, and 210 taxa collectively contributed less than 1% of the total abundance. Taxa diversity peaked in winter and reached a minimum in summer. Species richness varied considerably but was not significantly different over time. Maximum algal density occurred in later winter/spring (1.4 x 10(9) ind./m(2)) but was not significantly different from the minimum density, which occurred in August. Chlorophyll a showed similar seasonal fluctuation but also was not significantly different over time. Canonical correspondence analysis demonstrated that water hardness, depth, conductivity, and alkalinity had important influences on variation of epilithic algae in the Xiangxi River system.

Geographic distribution of Pelteobagrus fulvidraco and Pelteobagrus vachelli in the Yangtze River based on mitochondrial DNA markers

Three populations of Pelteobagrus vachelli and Pelteobagrus fulvidraco of the Yangtze River were examined by PCR-RFLP analysis of mitochondrial DNA fragments. ND5/6 and D-loop fragments were digested by 10 restriction endonucleases. Significant geographic variations between upstream and mid-downstream populations in the haplotype frequencies and restriction patterns were revealed. This suggested that the diversity of P. vachelli was high; 11 haplotypes were obtained from all the samples. The upstream population shared seven haplotypes and the middle and downstream populations shared another four haplotypes. Among all of the haplotypes, one haplotype was shared in 30 samples of the populations from middle and downstream, but it was not found in the upstream population. Any haplotype found in the upstream population was not detected in the middle and downstream populations. Genetic diversity of P. fulvidraco was low and only five haplotyes were detected from all 60 samples. Phylogenic relationships also indicated that the fishes from upstream and mid-downstream were apparently divided into two populations.

Identification of two novel interferon-stimulated genes from cultured CAB cells induced by UV-inactivated grass carp hemorrhage virus

Interferon (IFN) exerts its antiviral effect by inducing the expression of a number of IFN-stimulated genes (ISGs) to establish a host antiviral state. Earlier studies identified some important fish IFN system genes from IFN-induced CAB cells (crucian carp Carassius auratus L. embryonic blastulae cells) after treatment with UV-inactivated GCHV (grass carp hemorrhage virus). Herein, the cloning of 2 novel IFN-stimulated genes, termed Gig1 and Gig2, is described for the same cell system. The complete cDNA sequences of Gig1 and Gig2 contain 1244 bp encoding for a 194-amino-acid protein and 693 bp for a 158-amino-acid protein, respectively. A search of public databases revealed that these are 2 novel IFN-stimulated genes, since neither significant homologous genes nor conserved motifs were identified. Active GCHV, UV-inactivated GCHV and CAB IFN-containing supernatant (ICS) induced transcription of these genes and distinct kinetics were observed. An analysis of differences in expression between the 2 genes and the IFN signal factors CaSTAT1 and CaIRF7 indicated that GCHV infection activated different signal pathways for their up-regulation. Upon virus infection, the transcription of Gig1 but not of Gig2 is strongly suppressed by cycloheximide (CHX). In contrast, following treatment with CAB IFN-containing supernatant, CHX does not inhibit either gene transcription. The results suggest that GCHV infection can induce expression of both Gig1 and Gig2 via newly synthesized CAB IFN, most probably through the JAK-STAT signal pathway, and can also directly activate Gig2 transcription without ongoing protein synthesis.

Genetic structure of Chinese sucker population Myxocyprinus asiaticus in the Yangtze River based on mitochondrial DNA marker

The sequencing analysis of the mitochondrial DNA control region (mtCR DNA) was performed to assess the genetic divergence and population structure of the Chinese sucker Myxocyprinus asiaticus (Cypriniformes Catostomidae) using four sample lots from natural populations of the Yangtze River. The mtCR DNA sequences of approximately 920 base pairs were obtained. A total of 223 nucleotide positions were polymorphic, and these defined 39 haplotypes. Of the 39 haplotypes, 37 (90%) were not shared, and among the populations as a whole there was little sharing of haplotypes. The average haplotype diversity (0.958) and the average nucleotide diversity (0.052) indicated a higher level of genetic diversity of Chinese sucker through the river. Analysis of molecular variation (AMOVA) of data revealed significant partitioning of variance (P<0.001) among populations (60.29%), and within populations (39.71%). The topology according to the neighbor joining and maximum parsimony methods showed mosaic composition of the 39 haplotypes, suggesting that the populations wore not completely divergent. The pairwise F statistic values, however, indicated that the population structuring existed to some extent among the geographic populations. There was a positive relationship between the aquatic distance and the genetic distance (Fst) among the populations (P<0.05). Based on our data, it is suggested that genetic drift, gene flow, and stochastic events are the possible factors influencing the population structure and genetic variation.

Differential expression of two Carassius auratus Mx genes in cultured CAB cells induced by grass carp hemorrhage virus and interferon

UV-inactivated GCHV (grass carp hemorrhage virus) is able to induce an antiviral state in cultured CAB cells (crucian carp Carassius auratus blastulae embryonic cells) via the production of interferon (IFN). In the current work, the full-length cDNAs of two Mx genes, termed CaMx1 and CaMx2, have been cloned and sequenced from UV-inactivated GCHV-infected and still IFN-producing CAB cells by suppression subtractive hybridization. Their putative proteins show the characteristically structural features of mammalian IFN-induced Mx proteins, including GTP-binding motif, dynamin family signature and leucine zipper motif. CaMx1 exhibits 85% sequence identity to zebrafish MxA and 72-74% to three Atlantic salmon Mx proteins. CaMx2 is most similar to zebrafish MxE, with 80% identity, and then rainbow trout Mx3, with 52%. Constitutive expression was detected by RT-PCR for CaMx1, but not for CaMx2, in normal CAB cells, but their up-regulations could be induced after treatment with active GCHV, UV-inactivated GCHV and CAB IFN. Distinct kinetics of expression was observed for either CaMx1 or CaMx2 corresponding to the three stimuli, and even between CaMx1 and CaMx2, corresponding to the same stimulus. Upon virus infection, the transcriptional induction was strongly blocked for CaMx2 by cycloheximide (CHX), whereas almost nothing was observed for CaMx1. By contrast, following treatment with CAB IFN, CHX did not inhibit either gene transcription. Collectively, these results suggest that there are very distinct mechanisms for modulating the expression of both CaMx1 and CaMx2 in normal and GCHV-infected CAB cells.

Infection and propagation of lymphocystis virus isolated from the cultured flounder Paralichthys olivaceus in grass carp cell lines

The causative agent of lymphocystis disease that frequently occurs in cultured flounder Paralichthys olivaceus in China is lymphocystis virus (LV). In this study, 13 fish cell lines were tested for their susceptibility to LV. Of these, 2 cell lines derived from the freshwater grass carp Ctenopharyngodon idellus proved susceptible to the LV, and 1 cell line, GCO (grass carp ovary), was therefore used to replicate and propagate the virus. An obvious cytopathic effect (CPE) was first observed in cell monolayers at 1 d post-inoculation, and at 3 d this had extended to about 75% of the cell monolayer. However, no further CPE extension was observed after 4 d. Cytopathic characteristics induced by the LV were detected by Giemsa staining and fluorescence microscopic observation with Hoechst 33258 staining. The propagated virus particles were also observed by electron microscopy. Ultrastructure analysis revealed several distinct cellular changes, such as chromatin compaction and margination, vesicle formation, cell-surface convolution, nuclear fragmentation and the occurrence of characteristic 'blebs' and cell fusion. This study provides a detailed report of LV infection and propagation in a freshwater fish cell line, and presents direct electron microscopy evidence for propagation of the virus in infected cells. A possible process by which the CPEs are controlled is suggested.

Comparative ontogenetic behavior and migration of kaluga, Huso dauricus, and Amur sturgeon, Acipenser schrenckii, from the Amur River

We conducted laboratory experiments with kaluga, Huso dauricus, and Amur sturgeon, Acipenser schrenckii, to develop a conceptual model of early behavior. We daily observed embryos (first life phase after hatching) and larvae (period initiating exogenous feeding) to day-30 (late larvae) for preference of bright habitat and cover, swimming distance above the bottom, up- and downstream movement, and diel activity. Day-0 embryos of both species strongly preferred bright, open habitat and initiated a strong, downstream migration that lasted 4 days (3 day peak) for kaluga and 3 days (2 day peak) for Amur sturgeon. Kaluga migrants swam far above the bottom (150 cm) on only 1 day and moved day and night; Amur sturgeon migrants swam far above the bottom (median 130 cm) during 3 days and were more nocturnal than kaluga. Post-migrant embryos of both species moved day and night, but Amur sturgeon used dark, cover habitat and swam closer to the bottom than kaluga. The larva period of both species began on day 7 (cumulative temperature degree-days, 192.0 for kaluga and 171.5 for Amur sturgeon). Larvae of both species preferred open habitat. Kaluga larvae strongly preferred bright habitat, initially swam far above the bottom (median 50-105 cm), and migrated downstream at night during days 10-16 (7-day migration). Amur sturgeon larvae strongly avoided illumination, had a mixed response to white substrate, swam 20-30 cm above the bottom during most days, and during days 12-34 (most of the larva period) moved downstream mostly at night (23-day migration). The embryo-larva migration style of the two species likely shows convergence of non-related species for a common style in response to environmental selection in the Amur River. The embryo-larva migration style of Amur sturgeon is unique among Acipenser yet studied.