Molecular characterization and effect of RNA interference of retinoid X receptor (RXR) on E75 and chitinase gene expression in Chinese shrimp Fenneropenaeus chinensis

Retinoid X receptor (RXR)/ultraspiracle (USP) is the heterodimeric partner of ecdysteroid receptor and is required for the molting process of arthropods. To better understand the molecular aspects governing the process of molting in shrimp, the full-length cDNA of two RXRs, named as FcRXR-1 and FcRXR-2 were obtained from Chinese shrimp Fenneropenaeus chinensis which were of 1715 and 1700 bp long, revealed a 1315 and 1300 bp open reading frame (ORF) respectively. Quantitative Real time PCR analysis showed a marked tissue-specific difference in the expression of FcRXR transcript, which revealed that the expression of FcRXR Could be regulated in a tissue-specific manner. Moreover, high expression of FcRXR mRNAs was observed in late pre-molt period (D3) and post molt stages (A-B) of shrimp. Among the two isoforms, FcRXR-2 appeared in a considerably high level in all the stages compared to the FcRXR-1. In addition, we examined the temporal expression of two chitinase genes: FcChitinase (FcChi) and FcChitinase-1 (FcChi-1) during the molt cycle of F chinensis. Both the FcChi and FcChi-1 transcripts were detected in all stages of molting, although considerable fluctuations observed through the molt cycle. Injection of double stranded RXR (dsRXR) into juvenile shrimp resulted in a maximum silencing effect at 48 h post injection. We analyzed the expression levels of FcChi, FcChi-1 and the ecdysone inducible gene E75 (FcE75) in samples of dsRXR injected shrimp. Significant reduction in levels of both FcE75, FcChi and FcChi-1 transcripts Occurred in the silenced shrimp. This correlation suggested that RXR might involve in the downstream regulation of E75 and chitinase gene transcription in the ecdysone signaling pathway of decapod crustaceans. (C) 2009 Published by Elsevier Inc.

Inhibitory effects of the androgenic gland on ovarian development in the mud crab Scylla paramamosain

Isolation and characterization of androgenic hormone in decapod crustaceans depend on an effective bioassay of its action. In the present study, the effect of androgenic gland on ovarian development in the mud crab Scylla paramamosain was investigated with a view to develop a bioassay for androgenic hormone. Ovarian regression with degeneration of oocytes occurred in some female crabs implanted with androgenic gland in vivo. In vitro incubation of ovarian tissues at secondary vitellogenesis in extract of androgenic gland resulted in a significant decrease in amino acid uptake by the tissues. We propose that this inhibitory effect could be established as an effective bioassay for the isolation of androgenic hormone in the mud crab. (c) 2005 Published by Elsevier Inc.

The complete mitochondrial genomes of two common shrimps (Litopenaeus vannamei and Fenneropenaeus chinensis) and their phylogenomic considerations

Complete mitochondrial genomes have proven extremely valuable in helping to understand the evolutionary relationships among metazoans. However, uneven taxon sampling may lead to unclear or even erroneous phylogenetic topologies. The decapod crustaceans are relatively well-sampled, but sampling is still uneven within this group. We have sequenced the mitochondrial genomes of two shrimps Litopenaeus vannamei and Fenneropenaeus chinensis. As seen in other metazoans, the genomes contain a standard set of 13 protein-coding genes, 22 transfer RNA genes, two ribosomal RNA genes and an AT-rich non-coding region. The gene arrangements are consistent with the pancrustacean ground pattern. Both the pattern of gene rearrangements and phylogenomic analyses using concatenated nucleic acid and amino acid sequences of the 13 mitochondrial protein-coding genes strengthened the support that Caridea and Palinura are primitive members of Pleocyemata. These sequences, in combination with two previously published penaeid mitochondrial genomes, suggest that genera within the family Penaeidae have the following relationship: (((Penaeits + Fenneropenaett.) + Litopeiiaelts) + Marsupenaeus). The analyses of nucleic acid and amino acid sequences of the mitochondrial genomes also strongly support the monophyly of Penaeidae, Brachyura and Pleocyemata. In addition, the analyses of the average Ka/Ks in the 13 mitochondrial protein-coding genes of penaeid shrimps indicated a strong purifying selection within this group.

Molecular characterization of an ecdysone inducible gene E75 of Chinese shrimp Fenneropenaeus chinensis and elucidation of its role in molting by RNA interference

Ecdysone inducible gene. E75 is a primary target of ecdysone receptor (EcR). and is found to play a critical role in the molting process of arthropods In this study, a cDNA encoding the E75 of Chinese shrimp Fenneropenaeus chinensis (FcE75) was cloned using RT-PCR and RACE techniques FcE75 cDNA was 3611 bp in length with an ORF of 2394 bp. The deduced amino acid sequence of FcE75 had the highest sequence identity to E75 from a land crab Gecarcinus lateral's and E75 of the shrimp Metapenaeus crisis Quantitative real-time PCR revealed a prominently high expression of FcE75 mRNA in the whole body RNA extract of late premolt period (D3) juvenile shrimp. The role of E75 in the process of shrimp molting was investigated using the RNA interference technique Long double-stranded RNA corresponding to the FcE75 (dsE75) efficiently silenced the FcE75 transcript levels in juvenile F. chinensis. Further, injection with dsE75 completely arrested the molting process in experimental shrimp which eventually caused death Setogenic analysis of the uropods from molt-arrested shrimp, showed defective epidermal retraction, poor development of setae and new cuticle. These results indicate that E75 might be related to the molting process and is essential for proper molting and survival of shrimp This is the first report demonstrating the use of double stranded RNA to elucidate the possible role of E75 in the molting of decapod crustaceans (C) 2010 Elsevier Inc All rights reserved

Controlling factors of spring-summer phytoplankton succession in Lake Taihu (Meiliang Bay, China)

The spring-summer successions of phytoplankton and crustacean zooplankton were examined weekly in Meiliang Bay of the subtropical Lake Taihu in 2004 and 2005. During the study period, the ecosystem of Meiliang Bay was characterized by (i) clearly declined nitrogen compounds (nitrate, TN, and ammonium) and slowly increased phosphorus compounds (TP and SRP), (ii) increased total phytoplankton density and rapid replacement of chlorophyta (mainly Ulothrix) by cyanobacteria (mainly Microcystis), and (iii) rapid replacement of large-sized crustaceans (Daphnia and Moina) by small-sized ones (Bosmina, Limnoithona, and Ceriodaphnia). Results from the CCA and correlation analysis indicate that the spring-summer phytoplankton succession was primarily controlled by abiotic factors. Cyanobacteria were mainly promoted by increased temperature and decreased concentrations of nitrogen compounds. The pure contribution of crustacean was low for the variation of phytoplankton suggesting a weak top-down control by crustacean zooplankton in the subtropical Lake Taihu.

Spatiotemporal heterogeneity of plankton communities in Lake Donghu, China, as revealed by PCR-denaturing gradient gel electrophoresis and its relation to biotic and abiotic factors

The 16S and 18S rRNA genes of planktonic organisms derived from five stations with nutrient gradients in Lake Donghu, China, were studied by PCR-denaturing gradient gel electrophoresis (DGGE) fingerprinting, and the relationships between the genetic diversity of the plankton community and biotic/abiotic factors are discussed. The concentrations of total nitrogen (TN), total phosphorus (TP), NH4-N and As were found to be significantly related (P < 0.05) to morphological composition of the plankton community. Both chemical and morphological analyses suggested that temporal heterogeneity was comparatively higher than spatial heterogeneity in Lake Donghu. Although the morphological composition was not identical to the DGGE fingerprints in characterizing habitat similarity, the two strongest eutrophic stations (I and II) were always initially grouped into one cluster. Canonical correspondence analysis suggested that the factors strongly correlated with the first two ordination axes were seasonally different. The concentrations of TN and TP and the densities of rotifers and crustaceans were generally the main factors related to the DGGE patterns of the plankton communities. The study suggested that genetic diversity as depicted by metagenomic techniques (such as PCR-DGGE fingerprinting) is a promising tool for ecological study of plankton communities and that such techniques are likely to play an increasingly important role in assessing the environmental conditions of aquatic habitats.

Crustacean zooplankton distribution patterns and their biomass as related to trophic indicators of 29 shallow subtropical lakes

A comparative limnological study was carried out to present a snapshot of crustacean zooplankton communities and their relations to environmental factors to test whether there is a consistent relationship between crustacean biomass and trophic indicators among lake groups with similar trophic conditions. The study lakes showed a wide range of trophic status, with total phosphorus (TP) ranging from 0.008 to 1.448mgL(-1), and chlorophyll a from 0.7 to 146.1 mu g L-1, respectively. About 38 species of Crustacea were found, of which Cladocera were represented by 25 taxa (20 genera), and Copepoda by 13 taxa (I I genera). The most common and dominant species were Bosmina coregoni, Moina micrura, Diaphanosoma brachyurum, Cyclops vicinus, Thermocyclops taihokuensis, Mesocyclops notius and Sinocalanus dorrii. Daphnia was rare in abundance. Canonical correspondence analysis showed that except for four species (D. hyalina, S. dorrii, C. vicinus and M. micrura), almost all the dominant species had the same preference for environmental factors. Temperature, predatory cyclopoids and planktivorous fishes seem to be the key factors determining species distribution. TP was a relatively better trophic indicator than chlorophyll a to predict crustacean biomass. Within the three groups of lakes, however, there was no consistent relationship between crustacean biomass and trophic indicators. The possible reason might be that top-down and bottom-up control on crustaceans vary with lake trophic state. The lack of significant negative correlation between crustacean biomass and chlorophyll a suggests that there was little control of phytoplankton biomass by macrozooplankton in these shallow subtropical lakes. (c) 2007 Elsevier GmbH. All rights reserved.

Phytoplankton biomass in relation to nutrients and zooplankton in thirty subtropical lakes adjacent to the Yangtze River, China

A total of 30 shallow lakes, located along the middle and lower reaches of the Yangtze River, were studied to assess the relative importance of nutrients and zooplankton biomass in determining the phytoplankton biomass in subtropical China. Zooplankton biomass and nutrients both varied greatly in these lakes. Factor analysis and multiple linear regression showed that phytoplankton biomass was positively correlated with TN, NH4+, NO3- and TP, while it did not show any negative relationship to zooplankton biomass. Meanwhile, the phytoplankton biomass showed contrary relationships to the mass ratio of TN/TP in spring and summer, suggesting that in nutrient-richer lakes the dominant phytoplankton species have different preferences for TN/TP ratio. The insignificant top-down control of phytoplankton biomass may be attributed to the dominance of small-sized crustaceans and low crustacean biomass resulting from cyanobacterial dominance and planktivorous fish predation as well as other factors. Thus, it is likely that nutrients were more important than zooplankton biomass in explaining the total variance of phytoplaDkton biomass in these subtropical lakes.

Food composition and dietary overlap of macroinvertebrates in a shallow eutrophic lake in China: spatial and temporal variations

The present study was conducted in Lake Donghu, a suburban eutrophic lake arising from the middle reaches of the Yangtze River, China. Food composition of 32 taxa of zoobenthos was analyzed from 1251 gut samples. Macroinvertebrate primary consumers ingested mainly detritus, sand grains and diatoms. The predators primarily preyed on rotifers, crustaceans, oligochaetes and chironomid larvae. The dietary overlap was relatively high among collector taxa but low among other macroinvertebrates. Food composition and dietary overlap of macroinvertebrates changed considerably, both spatially and temporally. Food web structure differed between inshore and offshore regions of Lake Donghu. The inshore web was relatively complex and dynamic whereas the offshore web was simple and stable. Taxon-specific changes of diet seem to have little effect on the benthic food web structure in offshore waters of a eutrophic lake.

Crustacean zooplankton size structure in aquaculture lakes: is larger size structure always associated with higher grazing pressure?

Crustacean zooplankton size structure in 27 aquaculture lakes was studied to test the hypothesis that larger size structure is associated with higher grazing pressure. Mean body length of crustaceans was positively correlated with increasing Chl a (r(2) = 0.40, P = 0.000) and TP (r(2) = 0.38, P = 0.000), contrary to the empirical studies. However, the ratio of zooplankton to phytoplankton biomass decreased significantly with increasing TP (r(2) = 0.27, P = 0.005) and mean body length (r(2) = 0.46, P = 0.000). Meanwhile, size structure showed no significant effect in explaining residual variations of phosphorus-chlorophyll relationship (P = 0.231). These results indicate that larger size structure was not always associated with higher zooplankton grazing pressure. It is likely that in aquaculture lakes crustacean zooplankton size structure was of minor importance in control of phytoplankton biomass, and it was mainly regulated by fish predation. The results showed in our study and the empirical studies might be a reflection of two different stages of lake eutrophication and fish predation intensity.

Influence of selected Indian immunostimulant herbs against white spot syndrome virus (WSSV) infection in black tiger shrimp, Penaeus monodon with reference to haematological, biochemical and immunological changes

Immunostimulants are the substances, which enhance the non-specific defence mechanism and provide resistance against the invading pathogenic micro-organism. In order to increase the immunity of shrimps against the WSSV, the methanolic extracts of five different herbal medicinal plants like Cyanodon dactylon, Aegle marmelos, Tinospora cordifolia, Picrorhiza kurooa and Eclipta alba were selected and mixed thoroughly in equal proportion. The mixed extract was supplemented with various concentrations viz. 100 (A), 200 (B), 400 (C), and 800 (D) mg kg(-1) through artificial diets individually. The prepared diets (A-D) were fed individually to WSSV free healthy shrimp Penaeus monodon with an average weight of 8.0 +/- 0.5 g for 25 days. Control diet (E), devoid of herbal extract was also fed to shrimps simultaneously. After 25 days of feeding experiment, the shrimps were challenged with WSSV, which were isolated and propagated from the infected crustaceans. The shrimps succumbed to death within 7 days when fed on no herbal immunostimulant diet (E). Among the different concentrations of herbal immunostimulant supplemented diets, the shrimps fed on diet D (800 mg kg(-1)) significantly (P < 0.0001) had more survival (74%) and reduction in the viral load. Also the better performance of haematological, biochemical and immunological parameters was found in the immunostimulant incorporated diets fed shrimps. The present work revealed that the application of herbal immunostimulants will be effective against shrimp viral pathogenesis and they can be recommended for shrimp culture. (c) 2006 Published by Elsevier Ltd.

Effects of fish stocking on the zooplankton community structure in a shallow lake in China

Changes in the zooplankton community structure in relation to fishery practices in Lake Donghu, Wuhan, China were examined. The number of Protozoa species increased slightly, whereas the number of rotifers and crustaceans decreased from the 1960s to the 1990s. The total annual average densities of zooplankton increased 15-20 times in the 1990s compared with the 1960s. This increase was largely attributed to Protozoa, which contributed 93.4% by number of the total zooplankton density in 1991. Cladoceran densities decreased markedly from 1987. Changes in densities of rotifers and copepods were not evident. Trends in zooplankton biomass were similar to density. Large changes in zooplankton community structure coincided with markedly changes in concentration of chlorophyll a and transparency in Lake Donghu in 1987. The year 1987 seems to be the threshold year when the zooplankton community structure changed considerably. These changes were related to continuously increasing fish stock biomass in the lake. It was suggested that fish stocking and fish biomass should be a better managed for improvement of the quality of the lake's environment.

The rotifer assemblage in a shallow, eutrophic Chinese lake and its relationships with cyanobacterial blooms and crustacean zooplankton

Rotifer assemblage in the subtropical eutrophic Lake Chaohu was investigated monthly from September 2002 to August 2003. Forty-nine species belonging to 18 genera and 14 families were recorded. The highest densities of rotifer were observed during summer when there were heavy cyanobacterial blooms. There was a significant positive correlation between total rotifer density and the biomass of cyanobacteria. However, no correlations were found between the densities of rotifer and crustacean zooplankton, possibly owing to the paucity of large-bodied planktonic crustaceans. It is likely that the occurrence of cyanobacterial blooms not only caused the shift of dominant crustacean zooplankton from large species to smaller ones but also weakened the negative interaction between crustaceans and rotifers.

Toxicity of cypermethrin on growth, pigments, and superoxide dismutase of Scenedesmus obliquus

Cypermethrin is a synthetic pyrethroid that is particularly toxic to crustaceans. It is therefore applied as a chemotherapeutant in farms for the treatment of pests. The effective concentrations of cypermethrin on the inhibition of Scenedesmus ohliquus growth at 96h (96h EC50) were determined to be 50, 100, 150, 200, and 250mg/L. Algal growth, pigment fractions, and the activity of superoxide dismutase (SOD) in the algal cells were measured in the exponential phase after exposure to cypermethrin. The results show that higher concentration of cypermethrin is inhibitory for growth and other metabolic activities and the 96h EC50 of cypermethrin to S. ohliquus is 112 +/- 9 mg/L; the potential application of SOD activity in S. ohliquus as a sensitive biomarker for cypermethrin exposure is also discussed. (C) 2004 Elsevier Inc. All rights reserved.

Experimental study of trophic cascade effect of silver carp (Hypophthalmichthys molitrixon) in a subtropical lake, Lake Donghu: on plankton community and underlying mechanisms of changes of crustacean community

An enclosure experiment was carried out to test trophic cascade effect of filter-feeding fish on the ecosystem: growth of crustacean zooplankton, and possible mechanism of changes of crustacean community structure. Four fish biomass levels were set as follows: 0, 116, 176 and 316 g m(-2), and lake water ( containing ca. 190 g m(-2) of filter-feeding fishes) was comparatively monitored. Nutrient levels were high in all treatments during the experiment. Lowest algal biomass were measured in fishless treatment. Algal biomass decreased during days 21-56 as a function of fish biomass in treatments of low (LF), medium (MF) and high (HF) fish biomass. Crustaceans biomass decreased with increasing fish biomass. Small-bodied cladocerans, Moina micrura, Diaphanosoma brachyurum and Scapholeberis kingii survived when fish biomass was high whilst, large-bodied cladocerans Daphnia spp. and the cyclopoids Theromcyclops taihokuensis, T. brevifuratus, Mescyclops notius and Cyclops vicinus were abundant only in NF enclosures. Evasive calanoid Sinodiaptomus sarsi was significantly enhanced in LF, but decreased significantly with further increase of fish biomass. Demographic data indicated that M. micrura was well developed in all treatments. Our study indicates that algal biomass might be controlled by silver carp biomass in eutrophic environment. Changes of crustacean community are probably affected by the age of the first generation of species. Species with short generation time were dominant and species with long generation time survived less with high fish biomass. Evasive calanoids hardly developed in treatments with high fish biomass because of the ( bottle neck) effect of nauplii. Species abundance were positively related to fish predation avoidance. Other than direct predation, zooplankton might also be suppressed by filter-feeding fish via competition.