Fenneropenaeus Chinensis Research Articles

Comprehensive identification and profiling of Chinese shrimp (Fenneropenaeus chinensis) microRNAs in response to high pH stress using Hiseq2000 sequencing

pH has a great impact on the distribution, growth, behaviour and physiology in many aquatic animals. MicroRNAs (miRNAs), a class of small non-coding RNAs play essential roles in animal responses to environmental stress. In this study, we constructed two small RNA (sRNA) libraries from control (pH 7.4) and high pH (9.4) treated shrimp to identify high pH stress-responsive miRNAs. Using deep sequencing, we identified 72 (3 conserved and 69 novel) up- and 4 (2 conserved and 2 novel) down-regulated miRNAs from high pH challenged shrimp hepatopancreas. Gene Ontology annotations and Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed that these target transcripts were implicated in a broad range of biological processes and various metabolic pathways. Moreover, we validated the expression patterns of selected miRNAs and their target genes by qRT-PCR. The present work adds new insights into the large-scale identification and characterization of Fenneropenaeus chinensis miRNAs and their potential targets and represents a foundation for underlying mechanisms of animal responses to high pH stress at post-transcriptional level.

Genome-wide identification of Chinese shrimp (Fenneropenaeus chinensis) microRNA responsive to low pH stress by deep sequencing.

pH has a great impact on the distribution, growth, behavior, and physiology in many aquatic animals. Here, we analyzed miRNA expression profiles of Chinese shrimp (Fenneropenaeus chinensis) from control pH (8.2) and low pH (6.5)-treated shrimp. Expression analysis identified 6 known miRNAs and 23 novel miRNAs with significantly different expression between control pH 8.2 and low pH 6.5; the predicted target genes of differentially expressed miRNAs were significantly enriched in organic acid metabolic process, oxidoreductase activity, coenzyme binding, cofactor binding, and collagen trimer. Moreover, target genes were significantly enriched in several Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways including citrate cycle, pyruvate metabolism, cytokine-cytokine receptor interaction, tight junction, carbon metabolism, etc. Our survey expanded the number of known shrimp miRNAs and provided comprehensive information about miRNA response to low pH stress.

Genetic Distances between Two Cultured Penaeid Shrimp (Penaeus chinensis) Populations Determined by PCR Analysis.

Genomic DNA samples were obtained from cultured penaeid shrimp (Penaeus chinensis) individuals such as fresh shrimp population (FSP) and deceased shrimp population (DSP) from Shinan regions in the Korean peninsula. In this study, 233 loci were identified in the FSP shrimp population and 162 in the DSP shrimp population: 33 specific loci (14.2%) in the FSP shrimp population and 42 (25.9%) in the DSP population. A total of 66 (an average of 9.4 per primer) were observed in DSP shrimp population, whereas 55 unique loci to each population (an average of 7.9 per primer) in the FSP shrimp population. The Hierarchical dendrogram extended by the seven oligonucleotides primers indicates three genetic clusters: cluster 1 (FRESH 01, 02, and DECEASED 12, 13, 15, 16, 17, 19, 20, 22) and cluster 2 (FRESH 03, 04, 05, 06, 07, 08, 09, 10, 11, and DECEASED 14, 18, 21). Among the twenty-two shrimp, the shortest genetic distance that exposed significant molecular differences was between individuals 20 and 16 from the DSP shrimp population (genetic distance=0.071), while the longest genetic distance among the twenty-two individuals that established significant molecular differences was between individuals FRESH no. 02 and FRESH no. 04 (genetic distance=0.477). In due course, PCR analysis has revealed the significant genetic distance among two penaeid shrimp populations.

Linking energy metabolism and locomotor variation to osmoregulation in Chinese shrimp Fenneropenaeus chinensis.

Although previous studies have shown that salinity exposure affects shrimp respiration, little is known of the properties and variation of anaerobic and aerobic metabolism in tissues, and in particular of the relationship between energy metabolism and shrimp locomotion. To examine the relationship between energy metabolism and locomotor variation during osmoregulation, Chinese shrimp Fenneropenaeus chinensis were held at 30‰, 25‰, 20‰, 15‰, and 10‰ salinity for 15 days, after which the activities of key enzymes involved in glycolysis, tricarboxylic acid cycle, and electron transport chain in posterior gills, pleopods, and abdominal muscles were determined. Hemolymph osmolality, oxygen consumption rates, swimming and tail-flipping abilities were also obtained. Results showed that hemolymph of F. chinensis was isosmotic to brackish water at 25‰ salinity. The rate of oxygen consumption increased by 27.8% to 126.0% during osmoregulation. Activities of citrate synthase (CS) and cytochrome c oxidase (COX) increased in posterior gill rather than muscles during osmoregulation. In muscles, CS and COX activities decreased by 16.6% to 59.11% at 15‰ and 10‰ salinities, but phosphofructokinase (PFK) and lactate dehydrogenase (LDH) activities decreased by 13.2% to 44.5% at 10‰ salinity. Swimming abilities were positively correlative with CS and COX activities of pleopod muscle. Tail-flipping abilities increased with PFK and LDH activities of abdominal muscles. These findings suggest that shrimp increase reliance on aerobic metabolism for fueling osmoregulation. There is an energetic trade-off between gill and muscles under salinity exposure. The energetic depression of muscles results in the reduction of shrimp locomotion during osmoregulation.

Lower-intensity net-chasing training facilitates the survival and growth of the Chinese shrimp Fenneropenaeus chinensis reared at high stocking densities

Low survival and growth are the major problems in intensive culture of the Chinese shrimp Fenneropenaeus chinensis. We used net-chasing training to simulate predator-like threat to improve the survival and growth of F. chinensis reared at high stocking densities (up to 240 individuals m−2). The shrimps were trained three times a day under four chased intensities, control, 25%-, 50%- and 100%-exhausted, reared at densities of 240 individuals m−2 in each intensity. After 40 days, the biochemistry (lactate concentration, enzyme activities of anaerobic and aerobic metabolism), behavioural indicators (tail-flipping speeds; time spent on stationary, walking and swimming), survival and growth were determined. The net-chasing training increased tail-flipping speeds due to the increased activities of phosphofructokinase, pyruvate kinase and lactate dehydrogenase in abdominal muscles. The shrimps spent less time on swimming and more time on stationary after chasing training. The survival rates were 54.44% in the control group, but ranging from 88.89% to 90.56% in the 25%-, 50%-, and 100%-exhausted groups. Most of dead shrimps lost their pleopods, periodpods and (or) uropods. The 50%- and 100%-exhausted chasing reduced the total food consumption due to high levels of hemolymph lactate, resulting in low growth rates in these groups. The shrimps showed elevated growth rates with no reduced ingestion in the 25%-exhausted group. We conclude that net-chasing increased the tail-flipping ability and suppressed the cannibalistic behaviour, increasing the survival of F. chinensis reared at the high densities. The low-intensity (25%-exhausted) chasing increased the growth rates due to shrimp having more energy available allocating for somatic growth.

Characterization of a Lymphoid Organ Specific Anti-lipopolysaccharide Factor From Shrimp Reveals Structure-Activity Relationship of the LPS-Binding Domain.

Anti-lipopolysaccharide factor (ALF) is a kind of important antimicrobial peptides with broad-spectrum antimicrobial activities. The LPS-binding domain (LBD) contributes to the major antimicrobial activity of ALF. However, LBDs from different ALFs share low sequence similarity. The general character of LBDs needs to be elucidated to understand the molecular mechanism of their function and facilitate LBD-original drug design. Here we identified a lymphoid organ specifically expressed ALF, designated as FcALF8, from the Chinese shrimp Fenneropenaeus chinensis. The synthetic LBD peptide of FcALF8 (LBD8) showed strong antibacterial activities to the pathogenic Vibrio, such as Vibrio alginolyticus, Vibrio harveyi, and Photobacterium damselae with a MIC value of 0.5–1, 1–2, and 1–2 μM, respectively. FcALF8 knock-down using dsRNA led to significant increase of the viable bacteria in the lymphoid organ and hepatopancreas of shrimp upon V. harveyi infection. On the contrary, the proliferation of V. harveyi in the shrimp lymphoid organ and hepatopancreas significantly decreased after infected by LBD8 pre-incubated V. harveyi. Sequence alignments showed that the LBDs from 39 ALFs shared only two identical cysteine residues. However, 17 of the total 22 LBD residues showed high similarity when the amino acids were classified into hydrophobic and hydrophilic ones. A further activity analysis on modified LBD8 peptides showed that the antibacterial activity of LBD8 was lost after linearization and apparently weakened after changing the amino acid property at certain positions. The data indicated that the disulfide bond and amino acid property contributed to the conservation of the functional domain. To the best of our knowledge, this is the first identified ALFs specifically expressed in the lymphoid organ of shrimp with strong antibacterial activity. The present data will give creative instructions for the design of LBD-originated antimicrobial agents.

Screening of the candidate genes related to low-temperature tolerance of Fenneropenaeus chinensis based on high-throughput transcriptome sequencing.

In order to screen the candidate genes of Fenneropenaeus chinensis related to low-temperature tolerance, this research takes juvenile prawns of F. chinensis (P40) in low temperature stress group (4°C) and normal temperature group (18°C) as experimental materials. The results showed that a total of 127,939 Unigenes with average length of 1,190 bp were obtained by assembly, of which 46% were annotated in the Nr database. A total of 1,698 differentially expressed genes were screened by differential gene expression analysis, of which 920 genes showed up-regulated expression and 778 genes showed down-regulated expression. Both GO and KEGG enrichment analysis revealed that differentially expressed genes were enriched in spliceosomes, ribosomes, bile secretion, ABC transport pathways, and cellular nitrogen compound synthesis. A further in-depth analysis obtained 8 genes that may be associated with low-temperature traits of F. chinensis. Five of them displayed up-regulated expression, including ATP-binding cassette protein C, acid ceramidase, glutathione transferase, C-type lectin and heat shock protein HSP70. The remaining three genes, γ-butyl betaine hydroxylase, β-hexosaminidase A and long chain fatty acid-CoA ligase displayed down-regulated expression. Eight differentially expressed genes were randomly selected and the real time RT-PCR verification showed that their expression levels were consistent with the sequencing results, demonstrating the accuracy of the sequencing results. The results of this study provide basic data for revealing the molecular mechanisms of F. chinensis in response to low temperature stress and the molecular assisted breeding of F. chinensis in low temperature.

Transglutaminase-catalysed cross-linking eliminates Penaeus chinensis tropomyosin allergenicity by altering protein structure

ABSTRACT Transglutaminase (TG) is a major food processing enzyme that induces protein cross-linking. Tropomyosin (TM) is a major shellfish allergen that is responsible for at least 80% of the shrimp allergic incidents. The aim of this research was to investigate the effect of TG on TM allergenicity and reveal the involvement of TM structure alternation. After treated with TG for different time, the allergenicity of TM was assessed by indirect ELISA with shrimp allergic patients’ sera, and the structure of TM was characterized by circular dichroism (CD) spectroscopy, ultraviolet (UV) absorption spectroscopy and fluorescence spectroscopy. Results showed that the allergenicity slightly decreased at the early stage of reaction and dramatically decreased at the late stage, which is tightly correlated with the increment of β-turn content and UV absorption. In conclusion, TG modification is an effective and promising food processing method to eliminate TM allergenicity with significant prospect of industrial application.

Behavioural and physiological responses to low- and high-intensity locomotion in Chinese shrimp Fenneropenaeus chinensis.

We explored stroke behaviour, energy sources, and their related metabolic enzymes during multi-intensity swimming and tail-flipping at low- and high-intensity modes in Chinese shrimp Fenneropenaeus chinensis. In swimming, shrimp were encouraged to swim at velocities of 3, 6, 9cms-1 for 200min (low-intensity), and at 12, 15, 18cms-1 until fatigue (high-intensity). In tail-flipping, shrimp were encouraged to tail-flip by tapping cephalothorax at frequencies of 0.020, 0.040, 0.063Hz (one tap every 50, 25, 16s) for 5min (low-intensity), and at 0.083, 0,100, 0.125Hz (one tap every 12, 10, 8s) until no response (high-intensity). Results showed that shrimp increased stroke rates of pleopods and uropods to elevate swimming and tail-flipping ability. For low-intensity locomotion, glycogen was burned in aerobic pathway due to low pleopods beat frequency in swimming; however, glycogen was anaerobically burned due to high uropods beat amplitude in tail-flipping. Anaerobic metabolism occurred in high-intensity locomotion in either swimming or tail-flipping. Critical contents of muscle lactate causing locomotion fatigue might be around threefold of rest condition. Shrimp reduced locomotive time to avoid glycogen exhaustion and lactate accumulation during high-intensity locomotion. These findings highlight our understanding of physiological mechanisms of locomotion activities in shrimp.

Effects of dissolved oxygen, starvation, temperature, and salinity on the locomotive ability of juvenile Chinese shrimp Fenneropenaeus chinensis

Locomotion is considered a fundamental property defining the fitness of animals. Shrimp locomotion includes walking, swimming, and tail-flipping. Little is known about the locomotive ability of juvenile shrimp. We studied the acute effects of key environmental factors on the ability of swimming and tail-flipping of juvenile Chinese shrimp Fenneropenaeus chinensis (1.10 ± 0.07 cm, body length), which was indicated by critical swimming speed (Ucrit) and tail-flipping speed (Utail), respectively. The key environmental factors include dissolved oxygen [DO; 2.2, 3.2, 4.2, 5.2, and 6.2 (control) mg L−1], starvation [1 (control), 3, 5, 7, and 9 days], temperature [9, 13, 17, 21 (control), and 25 °C (degrees Celsius)], and salinity [10, 15, 20, 25, and 30 (control) ppt]. The effect comparisons between these environmental factors were also obtained. Results showed that the changes of DO, starvation, temperature and salinity can affect not only the swimming, but also the tail-flipping of juvenile shrimp. The tail-flipping may be more important than swimming for survival in most shrimps, and therefore, Utail was more stable than Ucrit under hypoxia and starvation exposures. Ucrit and Utail showed similar sensibility to the changes of temperature and salinity. As in the effect comparison, Ucrit of juvenile F. chinensis was most affected by temperature, followed by DO and starvation, and the effect of salinity was the lowermost. These findings can be of value in understanding the physiological ecology of juvenile shrimp, and improving the stock enhancement of F. chinensis.

Molecular isolation and characterization of a spätzle gene from Macrobrachium rosenbergii

Spätzle protein is an extracellular ligand of Toll receptor in Toll signaling pathway involved in the embryonic dorsoventral patterning and in the innate immunity. In this study, a spätzle gene of freshwater prawn, Macrobrachium rosenbergii (MrSpz) was isolated and characterized. The open reading frame of MrSpz consisted of 747 nucleotides encoding 248 amino acid residues containing a signal peptide and C-terminal spätzle activated domain. MrSpz shared high similarity to spätzle of Fenneropenaeus chinensis (FcSpz) at 92% identity and Marsupenaeus japonicus (MjSpz) at 83% identity. Phylogenetic analysis was performed and the results revealed that MrSpz was a member of the clade containing LvSpz3 of Litopenaeus vannamei, FcSpz and Penaeus monodon spätzle protein. The expression distribution at transcriptional level in various tissues of normal prawn revealed that the MrSpz was detected in gills, heart and hepatopancreas while no expression was observed in hemocyte, muscle and stomach. In the Aeromonas caviae challenged prawn, the expression level of MrSpz in hemocyte was increased gradually at 6, 12 and 24 h post-injection. Furthermore, in MrSpz knocked down prawn injected with Aeromonas caviae, the mortality rate were higher than that of non-related dsRNA group and control group. These results suggest that MrSpz protein may play a key role in the innate immunity of M. rosenbergii, especially in response to Gram-negative bacteria A. caviae invasion.

A novel micro‐organism for removing excess ammonia‐N in seawater ponds and the effect of Cobetia amphilecti on the growth and immune parameters of Litopenaeus vannamei

To investigate the feasibility of using micro‐organisms for ammonia‐N removal, six strains were isolated from Chinese white shrimp, Fenneropenaeus chinensis, seawater culture ponds in Dongying (Shandong, China). Of these, strain DY‐01, which exhibited the highest capacity to degrade ammonia‐N, removed 61.7% of the total ammonia‐N (50 mg/L) in 8 hr. An investigation of the factors affecting the removal efficiency indicated the optimum conditions to be 30°C, pH 8.0, and a salinity level of 30 g/L; 16S rDNA gene sequencing and biochemical analysis identified strain DY‐01 as Cobetia amphilecti, which has not previously been reported to degrade ammonia‐N. This strain also boosted the growth of Pacific white shrimp, Litopenaeus vannamei (p < .05), at a concentration of 107 CFU/mL, with no harmful effects on the shrimp's immune system. This study has thus identified a novel aerobic nitrifying bacterium that is potentially an excellent candidate for improving the water quality in mariculture ponds.

Isolation and characterization of a Raf gene from Chinese shrimp Fenneropenaeus chinensis in response to white spot syndrome virus infection

Raf is a member in the Ras/Raf/MAPKK/MAPK signaling transduction pathway. To obtain a better understanding of Raf in the interaction between the Chinese shrimp Fenneropenaeus chinensis and white spot syndrome virus (WSSV), the sequence of cDNA of Raf from F. chinensis (FcRaf) was obtained. The FcRaf gene contained a 2421 bp open reading frame (ORF). The FcRaf shared most characteristic of Raf protein, such as the Raf-like Ras-binding domain (RBD), phorbol esters/diacylglycerol binding domain (C1 domain), and catalytic domain of the serine/threonine kinases, Raf (STKc_Raf). The sequence of functional domains of Raf protein was relatively conserved. The FcRaf mRNA was detected in the tissues of gill, muscle, and hepatopancreas from normal F. chinensis. The mRNA abundance level of FcRaf in the gill was the highest, which was 2.7-fold the level in the hepatopancreas. The expression level of FcRaf was significantly (P < 0.05) up-regulated in the tissues of gill, muscle, and hepatopancreas post WSSV-infection, which suggested that FcRaf might be involved in the interaction between F. chinensis and WSSV. Two SNP loci were identified in the ORF, one of which was a C-T mis-sense mutation, where an Ala was replaced by a Val, and induced the predicted protein secondary structure change. Considering the relatively low MAF (0.07), whether this mis-sense mutation was a detrimental mutation needs further investigation.

Maillard reaction with ribose, galacto-oligosaccharide or chitosan-oligosaccharide reduced the allergenicity of shrimp tropomyosin by inducing conformational changes

Tropomyosin (TM), the predominant shellfish allergen, has induced severe food allergy worldwide, but efficient prevention and cure measures are limited nowadays. In the present study, we introduced an approach to eliminate Penaeus chinensis TM by Maillard reaction with reducing sugars including ribose, galacto-oligosaccharide (GOS) and chitosan-oligosaccharide (COS), which reduced the allergenicity by up to 60%. We also evaluated the conformational changes of TM during reaction by far-UV circular dichroism, and found the reaction-induced alternation from α-helix to β-sheet. We further determined the effect of Maillard reaction on TM allergenic epitopes to investigate the detailed mechanisms, and revealed the structure preference of different sugars. Correlation analysis with experimental data revealed that α-helix mediated the allergenicity elimination by Maillard reaction. There results provided theoretical foundations for the improvement and application of Maillard reaction in allergenic food processing.

The carotenoids of shrimps (Decapoda: Caridea and Dendrobranchiata) cultured in China

We compared the carotenoids in tissues of various organs from four shrimps cultured in China, Penaeus monodon Fabricius, 1978, Fenneropenaeus chinensis Osbeck, 1765, Litopenaeus vannamei Boone, 1931, and Exopalaemon carinicauda Holthuis, 1950. The carotenoid levels vary greatly depending on the species and the tissues sampled, with the highest levels found in the cephalothorax of L. vannamei (295.20 mg kg(-1)), and the lowest in the abdominal muscle of L. vannamei (2.12 mg kg(-1)). Astaxanthin, mainly esterified (47.69-95.56%), but also unesterified (4.445-2.31%), was the predominant carotenoid and contributed 66.64-95.26% of the total carotenoids in all species. beta-carotene, echinenone, canthaxanthin, astacene, zeaxanthin, and some unidentified carotenoids were also detected. All-trans-astaxanthin was the predominant geometrical isomer and 9-cis and 13-cis were the major cis isomers. Two other isomers, 15-cis and di-cis-astaxanthin, were also detected. The stereoisomers (3S, 3'S) and (3S, 3'R)-astaxanthin were only observed in E. carinicauda, where (3'S, 3'S)-astaxanthin contributed more than 96% to the total astaxanthin content. In contrast, all three possible stereoisomers were detected in P. monodon, F. chinensis, and L. vannamei and the relative levels of these stereoisomers were similar, 25.37-39.11% of (3S, 3'S)-astaxanthin, 38.85-52.01% of (3S, 3'R)-astaxanthin, and 14.07-29.89 % of (3R, 3'R)-astaxanthin. The astaxanthin-enriched waste, particularly the cephalothorax generated from L. vannamei, is suggested as a good source of natural astaxanthin available by further processing. These results provide some valuable information on carotenoid characteristics of cultured shrimps, especially the specific configurations of astaxanthin.

Effects of acute and chronic hypoxia on the locomotion and enzyme of energy metabolism in Chinese shrimp Fenneropenaeus chinensis

ABSTRACTTo characterize the locomotor behaviors and their relation with physiological regulation in Chinese shrimp Fenneropenaeus chinensis, animals were held at approximately 6.0 (normoxia), 4.5, and 3.0 mg L-1 dissolved oxygen (DO) for 1 day (acute) and 15 days (chronic), after which the swimming and tail-flipping abilities, and the activities of key enzymes involved in anaerobic and aerobic metabolism in hepatopancreas and pleopod and abdominal muscles were determined. Results showed that hepatopancreas was preferentially powered compared with pleopod and abdominal muscles during hypoxia. Physiological differences in muscles resulted in locomotion differences. F. chinensis presented reduced reliance on anaerobic glycolysis to conserve energy during chronic hypoxia at 3.0 mg L-1 DO, but this physiological regulation reduce the survival of shrimp in the wild due to a reduction in tail-flipping. These findings suggested that when assessing the survival strategy of shrimp during hypoxia, both physiological regulation and behavioral changes should be considered.

Expression of the Shrimp wap gene in Drosophila elicits defense responses and protease inhibitory activity

The wap gene encodes a single whey acidic protein (WAP) domain-containing peptide from Chinese white shrimp (Fenneropenaeus chinensis), which shows broad-spectrum antimicrobial activities and proteinase inhibitory activities in vitro. To explore the medical applications of the WAP peptide, a wap gene transgenic Drosophila melanogaster was constructed. In wap-expressing flies, high expression levels of wap gene (>100 times) were achieved, in contrast to those of control flies, by qRT-PCR analysis. The wap gene expression was associated with increased resistance to microbial infection and decreased bacterial numbers in the flies. In addition, the WAP protein extract from wap-expressing flies, compared with control protein extract from control flies, showed improved antimicrobial activities against broad Gram-positive and Gram-negative bacteria, including the clinical drug resistant bacterium of methicillin-resistant S. aureus (MRSA), improved protease inhibitor activities against crude proteinases and commercial proteinases, including elastase, subtilis proteinase A, and proteinase K in vitro, and improved growth rate and microbial resistance, as well as wound-healing in loach and mouse models. These results suggest that wap-expressing flies could be used as a food additive in aquaculture to prevent infections and a potential antibacterial for fighting drug-resistant bacteria.

Regulatory effects of Shewanella putrefaciens isolated from shrimp Penaeus orientalis on the virulence factors of Vibrio parahaemolyticus and evaluation of the role of quorum sensing in virulence factors regulation.

As an aquatic pathogen widely present in aquatic food, Vibrio parahaemolyticus causes outbreaks of gastroenteritis across the globe. Virulence factors of V. parahaemolyticus increases with the amount of spoilage in aquatic organisms including shrimp, but mechanisms regulating its virulence factors are not well understood. In this study, five spoilage bacteria isolated from shrimp were investigated for their regulatory effects on the virulence factors including haemolysin and biofilm of V. parahaemolyticus. Among these isolates, Shewanella putrefaciens induced haemolytic activity in V. parahaemolyticus in a time-dose-temperature-dependent manner and we found the main component responsible for this effect to be the supernatant or cell-free extract of S. putrefaciens. Total haemolytic activity, expression of the thermostable direct haemolysin gene tdh and biofilm production of V. parahaemolyticus were significantly up-regulated by S. putrefaciens, but also by deletion of quorum-sensing luxM or luxS gene of V. parahaemolyticus. However, this regulation by S. putrefaciens was significantly impaired by deletion of the luxM gene, but not by deletion of the luxS gene. Further study showed that S. putrefaciens exhibited a strong degradation ability on the signalling molecule acylated homoserine lactone (AHL) synthesised by the LuxM enzyme. This study revealed a novel virulence regulatory mechanism that S. putrefaciens can significantly increase the virulence factors of V. parahaemolyticus via interfering with the luxM- type quorum-sensing signalling pathway through its AHL-degradation ability.

Detection and quantification of shrimp hemocyte iridescent virus by TaqMan probe based real-time PCR.

Shrimp hemocyte iridescent virus (SHIV) is a recently reported shrimp virus, which threats the cultured white-leg shrimp Litopenaeus vannamei and can cause huge economic loss in shrimp farming industry in China. A quantitative real time polymerase chain reactio (qPCR) assay was developed using a TaqMan probe to detect and quantify SHIV. A pair of qPCR primers, which amplify a 188 bp DNA fragment, and a TaqMan probe were selected from ATPase gene (ORF114R) of the SHIV genome. The primers and TaqMan probe used in this assay were shown to be specific for SHIV and did not react with White spot syndrome virus (WSSV), Infectious hypodermal and hematopoietic necrosis virus (IHHNV), Hepatopancreatic parvovirus (HPV), Vibrio parahaemolyticus causing acute hepatopancreatic necrosis disease (VPAHPND), and Enterocytozoon hepatopenaei (EHP), or healthy shrimp DNA. The detection limit of the qPCR method was as low as 4 copies per reaction. The diagnostic sensitivity and the diagnostic specificity of the qPCR compared with nested PCR were 95.3% and 99.2%, respectively. The resulting standard curves showed high correlation coefficient values (R2 = 0.998) in the range of 4 × 109 to 4 × 100 DNA copies/reaction. Test of farm samples showed that SHIV was detected in L. vannamei, Fenneropenaeus chinensis and Macrobrachium rosenbergii contained SHIV ranged from 1.21E+02 to 7.95E+07 copies (μg DNA)-1. Quantitative detection of different tissues in artificial infected shrimp showed that haemolymph contained the highest SHIV load and muscle contained lowest SHIV load.

A Putative Insulin-like Androgenic Gland Hormone Receptor Gene Specifically Expressed in Male Chinese Shrimp.

The insulin-like androgenic gland hormone (IAG) is the key regulator in crustacean male sexual differentiation. As a secreted peptide hormone, IAG might perform its biological function through interacting with the membrane receptor. However, the receptor of IAG remains unclear. In the current study, a putative IAG receptor gene (FcIAGR) was identified in Fenneropenaeus chinensis. The deduced amino acid sequence of FcIAGR contained several conserved domains of insulin-like receptor proteins, including two L domains (L1 and L2), a cysteine-rich domain, three fibronectin III domains, a transmembrane domain, and an intracellular tyrosine kinase domain. Tissue distribution and in situ hybridization analysis showed that FcIAGR was predominantly expressed in the androgenic gland and testis in male F. chinensis. Protein colocalization analysis in HEK293 cells showed that FcIAGR could colocalize with both FcIAG1 and FcIAG2, respectively. Yeast two-hybrid assay further confirmed the interactions between FcIAGR and FcIAGs. After a long-term silencing of FcIAGR with double-stranded RNA, most of the germ cells in the testis were arrested at the secondary spermatocytes, whereas those in the control developed into sperm cells. The data indicated that FcIAGR was the receptor of FcIAGs in F. chinensis. The current study provides insight into the mechanism that the insulin-like signaling pathway regulates the male sexual differentiation in Decapoda crustaceans.