Temporal and Spatial Distributions of Rotifers in Xiangxi Bay of the Three Gorges Reservoir, China

From July 2003 to June 2005, investigations of rotifer temporal and spatial distributions were carried out in a bay of the Three Gorges Reservoir, Xiangxi Bay, which is the downstream segment of the Xiangxi River and the nearest bay to the Three Gorges Reservoir dam in Hubei Province, China. Thirteen sampling sites were selected. The results revealed a high species diversity, with 76 species, and 14 dominant species; i.e., Polyarthra vulgaris, Keratella cochlearis, Keratella valga, Synchaeta tremula, Synchaeta stylata, Trichocerca lophoessa, Trichocerca pusilla, Brachionus angularis, Brachionus calyciflorus, Brachionus forficula forficula, Ascomorpha ovalis, Conochilus unicornis, Ploesoma truncatum and Anuraeopsis fissa. After the first year of the reservoir impoundment, the rotifer community was dominated by ten species; one year later it was dominated by eight species. The community in 2003/2004 was dissimilar to that in 2004/2005, which resulted from the succession of the dominant species. The rotifer community exhibited a patchy distribution, with significant heterogeneity observed along the longitudinal axis. All rotifer communities could be divided into three groups, corresponding to the riverine, the transition and the lacustrine zone, respectively.

Population status, threats and conservation of the Yangtze finless porpoise

The Yangtze finless porpoise (Neophocaena phocaenoides asiaeorientalis) is currently limited to the middle and lower reaches of the Yangtze River from Yichang to Shanghai, China, and the adjoining Poyang and Dongting Lakes. Its population size has decreased remarkably during the last several decades due to the heavy impact of human activities, including overfishing of prey species, water development projects that cause attendant habitat loss and degradation, water pollution, and accidental deaths caused by harmful fishing gear and collisions with motorized vessels. It was estimated that the number of remaining individuals was down to approximately 1800 in 2006, a number that is decreasing at a rate as high as 5% per year. Three conservation measures - in situ and ex situ conservation and captive breeding have been applied to the protection of this unique porpoise since the early 1990s. Seven natural and two "semi-natural" reserves have so far been established. Since 1996, a small group of finless porpoises has been successfully reared in a facility at the Institute of Hydrobiology of the Chinese Academy of Sciences; three babies were born in captivity on July 5, 2005, June 2, 2007 and July 5, 2008. These are the first freshwater cetaceans ever born in captivity in the world. Several groups of these porpoises caught in the main stream of the Yangtze River, or rescued, have been introduced into the Tian'e-Zhou Semi-natural Reserve since 1990. These efforts have proven that, not only can these animals survive in the area, they are also to reproduce naturally and successfully. More than 30 calves had been born in the reserve since then, with one to three born each year. Taking deaths and transfers into account, there were approximately 30 individuals living in the reserve as of the end of 2007. Among eight mature females captured in April 2008, five were confirmed pregnant. This effort represents the first successful attempt at off-site protection of a cetacean species in the world, and establishes a solid base for conservation of the Yangtze finless porpoise. A lesson must be drawn from the tragedy of Chinese River Dolphin (Lipotes vexillifer), which has already been declared likely extinct. Strong, effective and appropriate protective measures must be carried out quickly to prevent the Yangtze finless porpoise from becoming a second Chinese River Dolphin, and save the biodiversity of the Yangtze River as a whole.

Subcellular localization and inductive expression of the dsRNA-dependent protein kinase PKR from Japanese flounder, Paralichthys olivaceus

The double-stranded RNA (dsRNA)-dependent protein kinase (PKR) belongs to the eIF2 alpha kinase family and plays a critical role in interferon (IFN)-mediated antiviral response. Recently, in Japanese flounder (Paralichthys olivaceus), a PKR gene has been identified. In this study, we showed that PoPKR localized to the cytoplasm, and the dsRNA-binding motifs (dsRBMs) played a determinative role in protein localization. In cultured FEC cells, PoPKR was detected at a low level of constitutive expression but was highly induced after treatment with UV-inactivated grass carp hemorrhagic virus, active SMRV and Poly I:C although with different expression kinetics. In flounder, PoPKR was ubiquitously distributed in all tested tissues, and SMRV infection resulted in significant upregulation at mRNA and protein levels. In order to reveal the role of PoPKR in host antiviral response, its expression upon exposure to various inducers was characterized and further compared with that of PoHRI, which is another eIF2 alpha kinase of flounder. Interestingly, expression comparison revealed that all inducers stimulated upregulation of PoHRI in cultured flounder embryonic cells and fish, with a similar kinetics to PoPKR but to a less extent. These results suggest that, during antiviral immune response, both flounder eIF2 alpha kinases might play similar roles and that PoPKR is the predominant kinase. (C) 2009 National Natural Science Foundation of China and Chinese Academy of Sciences. Published by Elsevier Limited and Science in China Press. All rights reserved.

Using drift nets to capture early life stages and monitor spawning of the Yangtze River Chinese sturgeon (Acipenser sinensis)

P>A sampling system for capturing sturgeon eggs using a D-shaped bottom anchored drift net was used to capture early life stages (ELS) of Chinese sturgeon, Acipenser sinensis, and monitor annual spawning success at Yichang on the Yangtze River, 1996-2004, before and just after the Three Gorges Dam began operation. Captured were 96 875 ELS (early life stages: eggs, yolk-sac larvae = eleuthero embryos, and larvae); most were eggs and only 2477 were yolk-sac larvae. Most ELS were captured in the main river channel and inside the bend at the Yichang spawning reach. Yolk-sac larvae were captured for a maximum of 3 days after hatching began, indicating quick dispersal downstream. The back-calculated day of egg fertilization over the eight years indicated a maximum spawning window of 23 days (20 October-10 November). Spawning in all years was restricted temporally, occurred mostly at night and during one or two spawning periods, each lasting several days. The brief temporal spawning window may reduce egg predation by opportunistic predators by flooding the river bottom with millions of eggs. During 1996-2002, the percentage of fertilized eggs in an annual 20-egg sample was between 63.5 to 94.1%; however, in 2003 the percentage fertilized was only 23.8%. This sudden decline may be related to the altered environmental conditions at Yichang caused by operation of the Three Gorges Dam. Further studies are needed to monitor spawning and changes in egg fertilization in this threatened population.

Characterization of the porcine differentially expressed PDK4 gene and association with meat quality

To investigate the differential expression of genes in the skeletal muscle between Yorkshire and Chinese indigenous breed Meishan pigs, suppression subtractive hybridization was carried out and many genes were proved to be expressed significantly different in the two breeds. One gene highly expressed in Meishan but lowly expressed in Yorkshire specific library, shared strong homology with human pyruvate dehydrogenase kinase 4 (PDK4). Using semi-quantity and quantity PCR, We confirmed its differential expression between the two breeds. Temporal and spatial expression analysis indicated that porcine PDK4 gene is highly expressed in skeletal muscle and the highest in neonatal pigs. Complete cDNA cloning and sequence analysis revealed that porcine PDK4 gene contains an open reading frame of 1,221 bp. The deduced amino acid sequence showed conservation in evolution. A G/A mutation in intron 9 was identified and association analysis showed that it was significantly associated with intramuscular fat, muscle water content.

Ultrastructure of the Spermatozoa of the Yangtze Finless Porpoise (Neophocaena phocaenoides asiaeorientalis)

P>Semen sample was collected from two captive adult Yangtze finless porpoises (Neophocaena phocaenoides asiaeorientalis) during physical examination. One individual was aged about 9 years with body length 143 cm (total length) and body weight 46.1 kg in 2003. The age of the other was unknown and its body length was 147 cm and body weight was 43 kg in 2004. Ultrastructure of their spermatozoa was examined using scanning and transmission electron microscope. The sperm concentration was 4.17 x 10(9) spermatozoa per ml by the cytometer. The approximate dimensions of the spermatozoa were as follows: head length, 3.366 +/- 0.140 mu m (mean +/- SE, n = 15); head width, 1.896 +/- 0.099 mu m (n = 15); and neck length, 1.004 +/- 0.074 mu m (n = 10). The tail included midpiece, principal piece and terminal piece. The length of the midpiece was 1.882 +/- 0.077 mu m (n = 9). There is no apparent boundary between the principal piece and the terminal piece, so the length of the principal piece and the terminal piece was 44.612 +/- 3.485 mu m (n = 5). Total length of the spermatozoa was 53.314 +/- 4.580 mu m (n = 10). The acrosome covered approximately 45.8% of the anterior portion of the head.

Origin and evolution of the RIG-I like RNA helicase gene family

Background: The DExD/H domain containing RNA helicases such as retinoic acid-inducible gene I (RIG-I) and melanoma differentiation-associated gene 5 (MDA5) are key cytosolic pattern recognition receptors (PRRs) for detecting nucleotide pathogen associated molecular patterns (PAMPs) of invading viruses. The RIG-I and MDA5 proteins differentially recognise conserved PAMPs in double stranded or single stranded viral RNA molecules, leading to activation of the interferon system in vertebrates. They share three core protein domains including a RNA helicase domain near the C terminus (HELICc), one or more caspase activation and recruitment domains (CARDs) and an ATP dependent DExD/H domain. The RIG-I/MDA5 directed interferon response is negatively regulated by laboratory of genetics and physiology 2 (LGP2) and is believed to be controlled by the mitochondria antiviral signalling protein (MAVS), a CARD containing protein associated with mitochondria. Results: The DExD/H containing RNA helicases including RIG-I, MDA5 and LGP2 were analysed in silico in a wide spectrum of invertebrate and vertebrate genomes. The gene synteny of MDA5 and LGP2 is well conserved among vertebrates whilst conservation of the gene synteny of RIG-I is less apparent. Invertebrate homologues had a closer phylogenetic relationship with the vertebrate RIG-Is than the MDA5/LGP2 molecules, suggesting the RIG-I homologues may have emerged earlier in evolution, possibly prior to the appearance of vertebrates. Our data suggest that the RIG-I like helicases possibly originated from three distinct genes coding for the core domains including the HELICc, CARD and ATP dependent DExD/H domains through gene fusion and gene/domain duplication. Furthermore, presence of domains similar to a prokaryotic DNA restriction enzyme III domain (Res III), and a zinc finger domain of transcription factor (TF) IIS have been detected by bioinformatic analysis. Conclusion: The RIG-I/MDA5 viral surveillance system is conserved in vertebrates. The RIG-I like helicase family appears to have evolved from a common ancestor that originated from genes encoding different core functional domains. Diversification of core functional domains might be fundamental to their functional divergence in terms of recognition of different viral PAMPs.

Population genetic structure and geographical differentiation of the Chinese catfish Hemibagrus macropterus (Siluriformes, Bagridae): Evidence for altered drainage patterns

National Natural Science Foundation of China (NSFC) [2007CB411600, 30530120]

Spring Diatom Blooming Phases in a Representative Eutrophic Bay of the Three-Gorges Reservoir, China

We investigated dynamics of the phytoplankton community and abiotic factors in Xiangxi Bay of the Three-Gorge Reservoir, China, by daily sampling, a specific site during a spring algal bloom (February 23-April 28, 2005). Among the 76 taxa observed, Asterionella formosa and Cyclotella spp. were the dominants, accounting for 47.2% and 29.9% of the total abundance, respectively. We determined the five distinct developing phases of the bloom by analyzing the dissimilarity of physicochemical parameters. Simultaneously, six phytoplankton community groups were distinguished by TWINSPAN classifications. The pattern for algal community succession was similar to that for the bloom phase shift, and the structural complexity of communities significantly decreased over time. Water temperature and silicate were the main factors that related to the development of the bloom and the shifts of the phytoplankton community.

Plasma biochemical responses of the planktivorous filter-feeding silver carp (Hypophthalmichthys molitrix) and bighead carp (Aristichthys nobilis) to prolonged toxic cyanobacterial blooms in natural waters

The planktivorous filter-feeding silver carp (Hypophthalmichthys molitrix) and bighead carp (Aristichthys nobilis) are the attractive candidates for bio-control of plankton communities to eliminate odorous populations of cyanobacteria. However, few studies focused on the health of such fishes in natural water body with vigorous toxic blooms. Blood parameters are useful and sensitive for diagnosis of diseases and monitoring of the physiological status of fish exposed to toxicants. To evaluate the impact of toxic cyanobacterial blooms on the planktivorous fish, 12 serum chemistry variables were investigated in silver carp and bighead carp for 9 months, in a large net cage in Meiliang Bay, a hypereutrophic region of Lake Taihu. The results confirmed adverse effects of cyanobacterial blooms on two phytoplanktivorous fish, which mainly characterized with potential toxicogenomic effects and metabolism disorders in liver, and kidney dysfunction. In addition, cholestasis was intensively implied by distinct elevation of all four related biomarkers (ALP, GGT, DBIL, TBIL) in bighead carp. The combination of LDH, AST activities and DBIL, URIC contents for silver carp, and the combination of ALT. ALP activities and TBIL, DBIL. URIC concentrations for bighead carps were found to most strongly indicate toxic effects from cyanobacterial blooms in such fishes by a multivariate discriminant analysis. (C) 2009 Elsevier B.V. All rights reserved.

Trends of Superoxide Dismutase and Soluble Protein of Aquatic Plants in Lakes of Different Trophic Levels in the Middle and Lower Reaches of the Yangtze River, China

A limnological study was carried out to determine the responses of superoxide dismutase (SOD) activities and soluble protein (SP) contents of 11 common aquatic plants to eutrophication stress. Field investigation in 12 lakes in the middle and lower reaches of the Yangtze River was carried out from March to September 2004. Our results indicated that non-submersed (emergent and floating-leafed) plants and submersed plants showed different responses to eutrophication stress. Both SOD activities of the non-submersed and submersed plants were negatively correlated with their SP contents (P < 0.000 1). SP contents of non-submersed plants were significantly correlated with all nitrogen variables in the water (P < 0.05), whereas SP contents of submersed plants were only significantly correlated with carbon variables as well as ammonium and Secchi depth (SD) in water (P < 0.05). Only SOD activities of submersed plants were decreased with decline of SD in water (P < 0.001). Our results indicate that the decline of SOD activities of submersed plants were mainly caused by light limitation, this showed a coincidence with the decline of macrophytes in eutrophic lakes, which might imply that the antioxidant system of the submersed plants were impaired under eutrophication stress.

Isolation and characterization of Argonaute 2: A key gene of the RNA interference pathway in the rare minnow, Gobiocypris rarus

Argonaute 2 gene plays a pivotal role in RNAi in many species. Herein is the first report of the cloning and characterization of Argonaute 2 gene in fish. The full-length cDNA of Gobiocypris rarus Argonaute 2 (GrAgo2) consisted of 3073 nucleotides encoding 869 amino acid residues with a calculated molecular weight of 98.499 kDa and an estimated isoelectric point of 9.18. Analysis of the deduced amino acid sequence showed the presence of two signature domains, PAZ and Piwi. RT-PCR analysis indicated that GrAgo2 mRNA expression could be detected in widespread tissues. After infection with grass carp reovirus, GrAgo2 expression was up-regulated from 12 h post-injection (p < 0.05) and returned to control levels at 48 h post-injection (p > 0.05). These data imply that GrAgo2 is involved in antiviral defense in rare minnow. (C) 2008 Published by Elsevier Ltd.

First Identification of the Hepatotoxic Microcystins in the Serum of a Chronically Exposed Human Population Together with Indication of Hepatocellular Damage

Hepatotoxic microcystins (MCs) are the most commonly reported cyanotoxins in eutrophic freshwaters. In 1996, human intoxications by MCs caused deaths of 76 patients at Caruaru dialysis centers in Brazil. So far, there have been no direct evidences of MC occurrence in human tissue in consequence of exposure to MC. In this study, we improved cleanup procedures for detecting MCs in serum sample using liquid chromatographymass spectrometry, and confirmed for the first time the presence of MCs in serum samples (average 0.39 ng/ml, which amounts to ca. 1/87 of the concentrations found in tissue samples of the Caruaru victims) of fishermen at Lake Chaohu. Daily intake by the fishermen was estimated to be in the range of 2.2-3.9 mu g MC-LReq, whereas the provisional World Health Organization tolerable daily intake (TDI) for daily lifetime exposure is 0.04 mu g/kg or 2-3 mu g per person. Moreover, statistical analysis showed closer positive relationships between MC serum concentrations and concentrations of alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, and lactate dehydrogenase than between the MC concentrations and other biochemical indicators. Thus, the data raise the question whether extended exposure in the range of the TDI or up to a factor of 10 above it may already lead to indication of liver damage. The results also demonstrate a risk of health effects from chronic exposure to MCs at least for populations with high levels of exposure, like these fishermen.

Genetic Diversity of the Plankton Community and Its Relation to Taxonomic Composition and Environmental Factors in Lake Xiliang

Genetic diversity of the plankton community in Lake Xiliang was depicted by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) fingerprinting. Seventy-seven bands (33 of 16S rDNA and 44 of 18S rDNA) were detected, sixty-two planktonic taxa were identified in six sample stations in November 2007. The most common taxa were Ceratium hirundinella, Bdelloidea, Keratella cochlearis, Polyarthra trigla, and copepod nauplii. Based on environmental factors, taxonomic composition, and PCR-DGGE fingerprinting, unweighted pair-group method using arithmetic averages clustering and principal components analysis were used to analyze habitat similarities. There was distinct spatial heterogeneity in Lake Xiliang, and the genetic diversity of the plankton community was closely related to taxonomic composition and environmental factors.

Effects of adrenergic agonists on the extrahepatic expression of vitellogenin Ao1 in heart and brain of the Chinese rare minnow (Gobiocypris rarus)

Teleost vitellogenins (VTGs) are large multidomain apolipoproteins and traditionally considered as the estrogen responsive precursors of the major egg yolk proteins. We identified five clones encoding VTGs, about 16% of the random EST clones from our constructed cDNA library from Chinese rare minnow liver tissue treated with 17 beta-estradiol (E2). Full-length vtgAo1 has been obtained based on the sequence information of four partial cDNA inserts by RACE. The inducibility of the vtgAo1 expression in liver by E2 was confirmed by RT-PCR. The presence of vtgAo1 transcripts have been observed primarily in liver. However. a significant level of the vtgAo1 was found in an unexpected location, heart, particularly in atrial cells by RT-PCR and whole mount in situ hybridization analyses. The vtgAo1 mRNA expression in heart and liver tissue could be suppressed by both alpha-adrenergic agonist, phenylephrine (PE) and beta-adrenergic agonist, isoproterenol (ISO). The expression of VTG in the heart observed in the present studies suggested it may provide protection from surplus intracellular lipids in fish cardiomyocytes as triglyceride transport proteins do in mammals. The results also indicated that the production of teleost vtg in vivo can be regulated by riot only estrogenic agents, but adrenergic signals as well. (c) 2008 Elsevier B.V. All rights reserved.