Immunoglobulin joining (J) chain and its expression in the Chinese soft-shelled turtle (Pelodiscus sinensis)

The immunoglobulin (Ig) joining (J) chain plays an important role in the formation of polymeric Igs and their transport into secretions. In the present study, the cDNA sequence of J chain has been cloned from the Chinese soft-shelled turtle (Pelodiscus sinensis) by reverse transcription (RT)-PCR and rapid amplification of cDNA ends (RACE). The cDNA sequence is 2347 bp in length and contains an open reading frame of 480 bp encoding 160 aa including the signal sequence. The deduced amino acid sequence has a high degree of homology with that of an already reported turtle J chain (80.7%), and of chicken (71.3%). By using real-time quantitative RT-PCR analysis, a significant up-regulation of J-chain transcripts was observed in spleen, kidney and blood of turtles injected with inactivated Aeromonas hydrophila, indicating the immune role of J chain in response to bacterial infection. (C) 2009 Elsevier B.V. All rights reserved.

Seasonal Variations in Chlorophyll a Concentrations in Relation to Potentials of Sediment Phosphate Release by Different Mechanisms in a Large Chinese Shallow Eutrophic Lake (Lake Taihu)

The relationship between chlorophyll a and fractionation of sediment phosphorus, inorganic phosphate-solubilizing bacteria (IPB), and organic phosphate-mineralizing bacteria (OPB) was evaluated in a large Chinese shallow eutrophic lake (Lake Taihu) and its embayment (Wuli Bay). At the three study sites, the increase of chlorophyll a concentrations in April paralleled those of the iron bound phosphate accounting for major portion of sediment inorganic phosphate, and in June significantly higher OPB and IPB numbers (especially OPB) in sediment were main contributors to the peaks of chlorophyll a concentration. Even though IPB peaked from February to June, it should serve as an unimportant P source due to the irrelevancy with chlorophyll a and soluble reactive phosphorus (SRP). By contrast, at the other site in the embayment, the calcium-bound phosphate was predominant and solid, which was difficult to be released, and neither IPB nor OPB were detectable in the sediment, indicating weak potential for phosphorus release from the sediment, which was reflected in the small seasonal variation in SRP concentration in water column. Hence, the extents to which the three general mechanisms behind phosphate release from sediment (desorption of iron bound phosphate, solubilization by IPB and enzymatic hydrolysis by OPB) operated were different depending on seasons and sites in Lake Taihu, they may jointly drive phosphate release and accelerate the eutrophication processes.

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.

Response Specificity to Advertisement Vocalization in the Chinese Alligator (Alligator sinensis)

Stringency in the identification of conspecific call properties is essential among sympatric species to ensure conspecific mating, as the risk of improper recognition and heterospecific mating is high. In this study we investigated the basic signal structure required for intraspecies communication in the Chinese alligator (Alligator sinensis), a species that has no relatives living in sympatry, by playback of signals modified in the temporal (truncating original bellows with first 1/4, 1/2, 3/4 or last 1/4, 1/2, 3/4 portion) or frequency domain (with low- or high-pass filters at frequencies 100, 250, 500 and 1000 Hz), or by reversal of natural bellows. The playback experiments revealed that relatively large modifications in bellow temporal or frequency structure failed to impair Chinese alligators' bellowing behavior; even reversed bellows effectively evoked a positive response. In general, the first half of the bellow in temporal domain and frequencies below 500 Hz were critical for behavioral induction, while the last half of the bellow in temporal domain and frequencies above 500 Hz failed to produce a single positive response, implying a potential functional signal redundancy. The observed high tolerance to bellow variations in Chinese alligators may be an evolutionary adaptation to (1) the acoustic constraints of propagation imposed by dense vegetative habitats; or (2) a lack of selection pressure due to the low risk of incorrect conspecific recognition and heterospecific mating.

Molecular and expression characterization of two somatostatin genes in the Chinese sturgeon, Acipenser sinensis

Chinese sturgeon (Acipenser sinensis) is a rare and endangered species and also an important resource for the sturgeon aquaculture industry. SMART cDNA was synthesized from the hypothalamus of Chinese sturgeon, and the full-length cDNAs of two somatostatin (SS) genes were cloned and sequenced. The first cDNA (AsSS1) encodes a 116-amino acid protein that contains the SS14 sequence at its C-terminal extremity. AsSS1 shows high identity to that of human and other vertebrates. The second cDNA (AsSS2) encodes a 111-amino acid protein that contains the somatostatin variant [Pro(2)]-SS14 at its C-terminal extremity. Both the two SS mRNAs were expressed in brain and pituitary with different mRNA levels. But in peripheral tissues, AsSS2 was more widely distributed than AsSS1. High mRNA levels of AsSS2 were found in liver, kidney and heart, while low mRNA levels of AsSS2 were also detected in ovary. Throughout embryogenesis and early larval development only AsSS2 mRNAs were detected. Furthermore, in the hypothalamus of one to five year-old Chinese sturgeon, AsSS2 but not AsSS1 maintained stable expression. The mRNA distribution suggests that the Chinese sturgeon AsSS2 products play important physiological functions in adult fish as well as in cell growth and organ differentiation in embryo and larva development. (C) 2009 Elsevier Inc. All rights reserved.

Localization and tracking of phonating finless porpoises using towed stereo acoustic data-loggers

Cetaceans produce sound signals frequently. Usually, acoustic localization of cetaceans was made by cable hydrophone arrays and multichannel recording systems. In this study, a simple and relatively inexpensive towed acoustic system consisting of two miniature stereo acoustic data-loggers is described for localization and tracking of finless porpoises in a mobile survey. Among 204 porpoises detected acoustically, 34 individuals (similar to 17%) were localized, and 4 of the 34 localized individuals were tracked. The accuracy of the localization is considered to be fairly high, as the upper bounds of relative distance errors were less than 41% within 173 m. With the location information, source levels of finless porpoise clicks were estimated to range from 180 to 209 dB re 1 mu Pa pp at 1 m with an average of 197 dB (N=34), which is over 20 dB higher than that estimated previously from animals in enclosed waters. For the four tracked porpoises, two-dimensional swimming trajectories relative to the moving survey boat, absolute swimming speed, and absolute heading direction are deduced by assuming the animal movements are straight and at constant speed in the segment between two consecutive locations.

Diversity and abundance of ammonia-oxidizing bacteria in eutrophic and oligotrophic basins of a shallow Chinese lake (Lake Donghu)

Classical cultivation and molecular methods based on the ammonia monooxygenase gene (amoA) were used to study the abundance and diversity of beta-proteobacterial ammonia-oxidizing bacteria (AOB) in lake sediments. The eutrophic and oligotrophic basins of a Chinese shallow lake (Lake Donghu), in terms of ammonium (NH4+) concentrations, were sampled. The AOB number was significantly lower in the oligotrophic basin, but significantly higher in the eutrophic basin. In addition, using restriction fragment length polymorphism targeting the amoA, ten restriction patterns including six unique ones were found in the eutrophic basin, while five patterns were observed in the oligotrophic basin with only one unique restriction group. Phylogenetic analysis for AOB revealed that Nitrosomonas oligotropha- and Nitrosomonas ureae-related AOB and Nitrosospira-affiliated AOB were ubiquitous; the former dominated in the eutrophic basin (87.2%), while the latter dominated in the oligotrophic basin (65.5%). Furthermore, Nitrosomonas communis-related AOB was only detected in the eutrophic basin, at a small proportion (3.2%). These results indicate significant selection and adaptation of sediment AOB in lakes with differing trophic status. (C) 2009 Elsevier Masson SAS. All rights reserved.

IgM, IgD and IgY and their expression pattern in the Chinese soft-shelled turtle Pelodiscus sinensis

Three Ig isotypes, IgM, IgD, and IgA, were previously known in reptiles. Here, in this report we describe IgM, IgD and a novel immunoglobulin heavy-chain isotype upsilon (IgY) in Chinese soft-shelled turtle (Pelodiscus sinensis). The IgM and IgY constant domains are characteristically similar to their counterparts described in other vertebrates. The expression of IgM and IgD were detected at mRNA level early during embryonic development, and their expression increased during further development. However, the IgY expression was not detected in larval turtles until 90 days after hatching-out. The increase in the transcription of these three Ig molecules was analyzed by using real-time PCR in spleen, kidney and blood following the injection of inactivated Aeromonas hydrophila. The primary increase in the expression of these three Igs was observed I week after the first injection, although not statistically significant, and the second injection 2 weeks after the first injection provoked a significant increase in the expression of these Igs, revealing a pattern of primary and secondary antibody response in the turtle. The present study represents the first report on reptile IgY and the pattern of IgM, IgD and IgY transcription in reptiles. (C) 2009 Elsevier Ltd. All rights reserved.

Molecular characterization and expression profiles in response to bacterial infection of Chinese soft-shelled turtle interleukin-8 (IL-8), the first reptilian chemokine gene

In this study, an IL-8 homologue has been cloned and identified from a reptile, Chinese soft-shelled turtle for the first time. The full-length cDNA of turtle IL-8 was 1188 bp and contained a 312 bp open reading frame (ORF) coding for a protein of 104 amino acids. The chemokine CXC domain, which contained Glu-Leu-Arg (ELR) motif and four cysteine residues, was well conserved in turtle IL-8. The 4924 bp genomic DNA of turtle IL-8 contained four exons and three introns. Phylogenetic analysis showed that the amino acid sequence of turtle IL-8 clustered together with birds. RT-PCR analysis showed that turtle IL-8 mRNA was constitutively expressed liver, spleen, kidney, heart, blood and intestine tissues of control turtles. Real-time quantitative PCR analysis further indicated that the turtle IL-8 mRNA expression was apparent in various tissues at 8 h and up-regulated significantly during 8 h-7 d after Aeromonas hydrophila infection. The present studies will help us to understand the evolution of IL-8 molecule and the inflammatory response mechanism in reptiles. (C) 2009 Elsevier Ltd. All rights reserved.

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]

Simultaneous determination of microcystin contaminations in various vertebrates (fish, turtle, duck and water bird) from a large eutrophic Chinese lake, Lake Taihu, with toxic Microcystis blooms

This is the first to conduct Simultaneous determination of microcystin (MC) contaminations in multi-groups of vertebrates (fish, turtle, duck and water bird) from Lake Taihu with Microcystis blooms. MCs (-RR, -YR, -LR) in Microcystis scum was 328 mu g g(-1) DW. MCs reached 235 mu g g(-1) DW in intestinal contents of phytoplanktivorous silver carp, but never exceeded 0.1 mu g g(-1) DW in intestinal contents of other animals. The highest MC content in liver of fish was in Carassius auratus (150 ng g(-1) DW), followed by silver carp and Culter ilishaeformis, whereas the lowest was in common carp (3 ng g(-1) DW). In livers of turtle, duck and water bird, MC content ranged from 18 to 30 ng g(-1) DW. High MC level was found in the gonad, egg yolk and egg white of Nycticorax nycticorax and Anas platyrhynchos, suggesting the potential effect of MCs on water bird and duck embryos. High MC contents were identified for the first time in the spleens of N. nycticorax and A. platyrhynchos (6.850 and 9.462 ng g(-1) DW, respectively), indicating a different organotropism of MCs in birds. Lakes with deaths of turtles or water birds in the literatures had a considerably higher MC content in both cyanobacteria and wildlife than Lake Taihu, indicating that toxicity of cyanobacteria may determine accumulation level of MCs and consequently fates of aquatic wildlife. (C) 2009 Elsevier B.V. All rights reserved.

Molecular characterization of Chinese sturgeon gonadotropins and cellular distribution in pituitaries of mature and immature individuals

Chinese sturgeon (Acipenser sinensis) is a rare and endangered species, and also an important resource for the sturgeon aquaculture industry. To understand molecular characterization of Chinese sturgeon gonadotropins (GTHs), we cloned the full-length cDNAs of gonadotropin subunits common alpha (GTH-alpha), follicle-stimulating hormone (FSH) and luteinizing hormone (LH) from a pituitary cDNA library of mature female. Two subtypes of GTH-alpha were identified. The nucleotide sequences of A. sinensis common alpha I (AsGTH-alpha I), common alpha II (AsGTH-alpha II), FSH beta (AsFSH beta) and LH beta (AsLH beta) subunit cDNAs are 345, 363, 387 and 414 bp in length, and encode mature peptides of 115, 121, 129 and 138 aa, respectively. Then, three polyclonal antibodies were prepared from the in vitro expressed AsGTH-alpha I, AsFSH beta and AsLH beta mature proteins, respectively. Significant expression differences were revealed between immature and mature sturgeon pituitaries. Western blot detection and immunofluoresence localization revealed the existence of three-gonadotropin subunits (AsGTH-alpha, AsFSH beta and AsLH beta) in mature sturgeon pituitaries, but only AsFSH beta was detected in immature individual pituitaries during early stages in the sturgeon life, and obvious difference was observed between males and females. In males, AsFSH beta was expressed in 4-year-old individuals, whereas in females, AsFSH beta was just expressed in 5-year-old individuals. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

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.

Distribution and depuration of the potentially carcinogenic malachite green in tissues of three freshwater farmed Chinese fish with different food habits

The use of malachite green (MG) in fish farming is prohibited in China due to its potentially toxicological and carcinogenic nature, but it is still illegally used in some places. Uptake, accumulation and deputation of MG in various tissues were studied under laboratory conditions in three common freshwater fish, Parabramis pekinensis (plant-eating fish), Carassius auratus (omnivorous fish) and Ophiocephalus argus (carnivorous fish). The concentrations of MG and its primary metabolite, the reduced and colorless leucomalachite green (LMG), were analyzed by liquid chromatography-mass spectrometry (LC-MS2). Absorption of MG occurred during the waterborne exposure and the MG concentrations in gills of the three fish species all showed a maximum at 0 h after an acute water exposure (6 mg l(-1) MG for 20 min). Afterwards, both MG and LMG declined very rapidly in the blood of the fish. Levels of MG and LMG were still above 0.002 mu g g(-1) in fresh weight muscle at 240 h and may persist for as long as 10 days. Most MG was converted rapidly to LMG in the fish and deputation of LMG was very slow in fat tissue. skin and gonads of the fish. Distribution of LMG was strongly dependent on the fat content in the tissues of the fish, but not related to their different feeding habits. Therefore, it appears that fat tissue, skin and gonads of the fish contaminated by MG and LMG pose the greatest risk for human consumption. (C) 2008 Published by Elsevier B.V.

Relaxed purifying selection of rhodopsin gene within a Chinese endemic cavefish genus Sinocyclocheilus (Pisces: Cypriniformes)

The cyprinid fish genus Sinocyclocheilus, as the most cavefish rich genus, includes many species showing striking adaptation to caves and convergent reduction or even loss of eyes and pigmentation. RH1 is responsible for dim vision. In order to explore the evolution of RH1 gene in this genus, we sequenced the complete gene from 28 individuals of 16 representative species of Sinocyclocheilus, with cave and surface species included. Phylogenetic analyses supported the monophyly of Sinocyclocheilus and polyphyly of the cave species. Codon models implemented in PAML were used to infer the evolution of RH1. We found that Sinocyclocheilus had a significantly higher evolutionary rate for amino acids than other cyprinid fishes compared, which might be the result of relaxation of purifying selection and could be ascribed to cave habit of this genus. In contrast to previous hypotheses, both cave and surface lineages exhibited a similar rate of molecular evolution, so the RH1 of cave species may still be functional, although these species were highly adapted to cave environment. Two amino acid substitutions (D83G and E122V) that were not reported before were found, which may be useful for site-directed mutagenesis in the future.