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1.
An attempt was made to determine the replication efficiency of hepatopancreatic parvo‐like virus (HPV) of shrimp in different organs of freshwater rice‐field crab Paratelphusa hydrodomous (Herbst) using bioassay, PCR, RT‐PCR, ELISA, Western blot and q‐PCR analyses. Another attempt was made to use this crab as an alternative to penaeid shrimp for the large‐scale production of HPV. This crab was found to be highly susceptible to HPV by intramuscular injection. The systemic HPV infection was confirmed by PCR and Western blot analyses in freshwater crab. The expression of capsid protein gene in different organs of infected crab was revealed by RT‐PCR analysis. Indirect ELISA was used to quantify the capsid protein in different organs of the crab. The copy number of HPV in different organs of the infected crab was quantified by q‐PCR. The results revealed a steady decrease in CT values in different organs of the infected crab during the course of infection. The viral inoculum that was prepared from different organs of the infected crab caused significant mortality in post‐larvae of tiger prawn, Penaeus monodon (Fabricius). The results revealed that this rice‐field crab could be used as an alternative host for HPV replication and also for large‐scale production of HPV.  相似文献   

2.
The potential for developing botanically derived natural products as novel feed‐through repellents for disrupting settlement of the salmon louse, Lepeophtheirus salmonis (Caligidae) upon farmed Atlantic salmon, Salmo salar, was investigated using an established laboratory vertical Y‐tube behavioural bioassay for assessing copepodid behaviour. Responses to artificial sea water conditioned with the odour of salmon, or to the known salmon‐derived kairomone component, α‐isophorone, in admixture with selected botanical materials previously known to interfere with invertebrate arthropod host location were recorded. Materials included oils extracted from garlic, Allium sativum (Amaryllidaceae), rosemary, Rosmarinus officinalis (Lamiaceae), lavender, Lavandula angustifolia (Lamiaceae), and bog myrtle, Myrica gale (Myricaceae), and individual components (diallyl sulphide and diallyl disulphide from garlic; allyl, propyl, butyl, 4‐pentenyl and 2‐phenylethyl isothiocyanate from plants in the Brassica genus). Removal of attraction to salmon‐conditioned water (SCW) or α‐isophorone was observed when listed materials were presented at extremely low parts per trillion (ppt), that is picograms per litre or 10?12 level. Significant masking of attraction to SCW was observed at a level of 10 ppt for diallyl disulphide and diallyl sulphide, and allyl isothiocyanate and butyl isothiocyanate. The potential of very low concentrations of masking compounds to disrupt Le. salmonis copepodid settlement on a host fish has been demonstrated in vitro.  相似文献   

3.
White tail disease (WTD) caused by Macrobrachium rosenbergii nodavirus (MrNV) and extra small virus (XSV) is a serious problem in prawn hatcheries. The gene for capsid protein of MrNV (MCP43) was cloned into pRSET B expression vector. The MCP43 protein was expressed as a protein with a 6‐histidine tag in Escherichia coli GJ1158 with NaCl induction. This recombinant protein, which was used to raise the antiserum in rabbits, recognized capsid protein in different WTD‐infected post‐larvae and adult prawn. Various immunological methods such as Western blot, dot blot and ELISA techniques were employed to detect MrNV in infected samples using the antiserum raised against recombinant MCP43 of MrNV. The dot blot assay using anti‐rMCP43 was found to be capable of detecting MrNV in WTD‐infected post‐larvae as early as at 24 h post‐infection. The antiserum raised against r‐MCP43 could detect the MrNV in the infected samples at the level of 100 pg of total protein. The capsid protein of MrNV estimated by ELISA using anti‐rMCP43 and pure r‐MCP43 as a standard was found to increase gradually during the course of infection from 24 h p.i. to moribund stage. The results of immunological diagnostic methods employed in this study were compared with that of RT‐PCR to test the efficiency of antiserum raised against r‐MCP43 for the detection of MrNV. The Western blot, dot blot and ELISA detected all MrNV‐positive coded samples as detected by RT‐PCR.  相似文献   

4.
White spot syndrome virus (WSSV), an aquatic virus infecting shrimps and other crustaceans, is widely distributed in Asian subcontinents including India. The infection has led to a serious economic loss in shrimp farming. The WSSV genome is approximately 300 kb and codes for several proteins mediating the infection. The envelope proteins VP26 and VP28 play a major role in infection process and also in the interaction with the host cells. A comprehensive study on the viral proteins leading to the development of safe and potent antiviral therapeutic is of adverse need. The novel synthesized compound 3‐(1‐chloropiperidin‐4‐yl)‐6‐fluoro benzisoxazole 2 is proved to have potent antiviral activity against WSSV. The compound antiviral activity is validated in freshwater crabs (Paratelphusa hydrodomous). An in silico molecular docking and simulation analysis of the envelope proteins VP26 and VP28 with the ligand 3‐(1‐chloropiperidin‐4‐yl)‐6‐fluoro benzisoxazole 2 are carried out. The docking analysis reveals that the polar amino acids in the pore region of the envelope proteins were involved in the ligand binding. The influence of the ligand binding on the proteins is validated by the molecular dynamics and simulation study. These in silico approaches together demonstrate the ligand's efficiency in preventing the trimers from exhibiting their physiological function.  相似文献   

5.
Adequate enrichment of live prey like Artemia, naturally deficient of essential highly unsaturated fatty acids (HUFA), such as docosahexaenoic acid (22:6n‐3, DHA), is critical for the rapidly developing tissues, survival, normal development and production of good‐quality fingerlings. The aim of the study was to evaluate the effects of a pulse (10–30 dah) of Shewanella putrefaciens Pdp11 (2.5*107 cfu/ml) using Artemia metanauplii as live vector, on its proper lipid profiles and resultant Solea senegalensis body composition and performance. Probiotic administration significantly increased total lipids and specifically n‐3 HUFA levels in Pdp11‐enriched Artemia. The live prey lipid modulation was also reflected in the total lipid contents and fatty acid profiles of Pdp11 sole specimens, which achieved a higher growth performance. A fatty acid multivariate principal component analysis confirmed a neat separation of two groups corresponding to Control and probiotic fish for each age sampled (23, 56, 87 and 119 dah). In addition, a further SIMPER analysis highlighted that the Pdp11 Artemia effect on sole lipid profile was different for each fatty acid and was gradually diluted with age. Results suggest an ability of Pdp11 strain to produce n‐3 HUFA as an effective tool for fish marine larviculture optimization.  相似文献   

6.
Anguillid herpesvirus 1 (AngHV1) causes a haemorrhagic disease with increased mortality in wild and farmed European eel, Anguilla anguilla (L.) and Japanese eel Anguilla japonica, Temminck & Schlegel). Detection of AngHV1 is currently based on virus isolation in cell culture, antibody‐based typing assays or conventional PCR. We developed, optimized and concisely validated a diagnostic TaqMan probe based real‐time PCR assay for the detection of AngHV1. The primers and probe target AngHV1 open reading frame 57, encoding the capsid protease and scaffold protein. Compared to conventional PCR, the developed real‐time PCR is faster, less labour‐intensive and has a reduced risk of cross‐contamination. The real‐time PCR assay was shown to be analytically sensitive and specific and has a high repeatability, efficiency and r2‐value. The diagnostic performance of the assay was determined by testing 10% w/v organ suspensions and virus cultures from wild and farmed European eels from the Netherlands by conventional and real‐time PCR. The developed real‐time PCR assay is a useful tool for the rapid and sensitive detection of AngHV1 in 10% w/v organ suspensions from wild and farmed European eels.  相似文献   

7.
The codon usage patterns of open reading frames (ORFs) in cyprinid herpesvirus 3 (CyHV‐3) have been investigated in this study. The high correlation between GC12% and GC3% suggests that mutational pressure rather than natural selection is the main factor that determines the codon usage and base component in the CyHV‐3, while mutational pressure effect results from the high correlation between GC3% and the first principal axis of principle component analysis (Axis 1) on the relative synonymous codon usage (RSCU) value of the viral functional genes. However, the interaction between the absolute codon usage bias and GC3% suggests that other selections take part in the formation of codon usage, except for the mutational pressure. It is noted that the similarity degree of codon usage between the CyHV‐3 and goldfish, Carassius auratus (L.), is higher than that between the virus and common carp, Cyprinus carpio L., suggesting that the goldfish plays a more important role than the common carp in codon usage pattern of the CyHV‐3. The study of codon usage in CyHV‐3 can provide some evidence about the molecular evolution of the virus. It can also enrich our understanding about the relationship between the CyHV‐3 and its hosts by analysing their codon usage patterns.  相似文献   

8.
Macrobrachium rosenbergii nodavirus (MrNV) that causes white tail disease (WTD) is an emerging disease that contributes to serious production losses in Macrobrachium hatcheries worldwide. Mosquito cell lines (C6/36) have been reported to support the growth of MrNV and used to observe the cytopathic effects (CPE) in infected cells. This study determined the susceptibility of C6/36 mosquito cells to the Australian isolate of MrNV in order to use fewer animals in further investigations. Different staining methods were used to observe MrNV viral activity in C6/36 cells. Typical cytopathic effects such as vacuolation and viral inclusion bodies were observed in infected C6/36 cells with H&E and Giemsa staining. With acridine orange, it was easier to detect presumptive MrNV messenger ribonucleic acid in the infected cells. Using neutral red staining to measure mitochondrial activity showed light absorption of infected cells maximized at day 4 (O.D. = 0.6) but was significantly lower (chi‐square = 41.265, df = 1, P < 0.05) than control groups (O.D. = 2) which maximized at day 12. Using trypan blue staining to count the number of cells with disrupted cell membranes, the maximum number of presumptively dead cells at day 8 (4 × 105 cells) in infected treatments was higher than the control treatment at day 10 (1.8 × 105 cells). However, TaqMan real‐time PCR did not confirm the replication of MrNV in the cells over 14 days. The mean viral copies and mean cycle times of positive samples were stable at 2.07 × 104 and 24.12, respectively. Limited evidence of viral replication was observed during four serial passages. This study determined the mortality of the C6/36 cell line to the Australian isolate of MrNV but suggests limited patent replication was occurring. Trying different cell lines or adapting the virus to the C6/36 cells may be necessary to successfully replicate Australian MrNV in cell lines.  相似文献   

9.
A range of physical cell disruption techniques have been evaluated to aid the processing of astaxanthin‐rich haematocysts of Haematococcus pluvialis for inclusion in salmonid feeds. Cell disruption by a scalable pressure treatment system was shown to be effective in breaking open the haematocysts without altering the content or isomeric composition of carotenoids in the algal cells. Storage of disrupted cells was optimal at ?20°C in the dark under nitrogen. Disrupted cells were spray‐dried, incorporated into commercial diets and fed to rainbow trout (Oncorhynchus mykiss L.). A marketable level of pigmentation in fish muscle was achieved after 10‐week dietary supplementation. The geometric and optimal isomer composition of the astaxanthin deposited in the muscle was nearly identical to that seen in Haematococcus. Changes were observed in the chirality of the astaxanthin deposited in the skin in comparison with that isolated from both the white muscle and the alga.  相似文献   

10.
A new cell line (TSHC) derived from heart tissues was established from female half‐smooth tongue sole (Cynoglossus semilaevis), an economically important marine fish species in China. The cell line had been subcultured for more than 30 times over a period of 200 days. The cell line was optimally maintained at 24°C in minimum essential medium (MEM) medium containing foetal bovine serum (FBS), 2‐mercaptoethanol (2‐Me), sodium pyruvate, basic fibroblast growth factor (bFGF) and antibiotics. The TSHC cells were mostly composed of fibroblast‐like cells. Chromosome analysis revealed that the TSHC cell line had a normal diploid karyotype with 2n = 42, containing the heterogametic W chromosome. The TSHC cell line was susceptible to infection by flounder Lymphocystis disease virus (LCDV). Although an atypical cytopathic effect and only few of virus particles in the cytoplasm was observed, it provides a research material on the cell–pathogen interaction research about the viral infection of non‐host species.  相似文献   

11.
Vibrio rotiferianus is an important marine pathogen of various aquatic organisms and can be found widely distributed in the marine environment. To further characterize this pathogen, the pathogenic properties and genome of V. rotiferianus SSVR1601 isolated from Sebastes schlegelii with skin ulcer were analysed. SSVR1601 was shown to be short rod‐shaped cell with a single polar flagellum. Different degrees of pathological changes in fish kidney, intestine, gills and liver were observed after SSVR1601 challenge. The SSVR1601 genome consists of two chromosomes and two plasmids with a total of 5,717,113 bp, 42.04%–44.93% GC content, 5,269 predicted CDSs, 134 tRNAs and 40 rRNAs. The common virulence factors including OMPs, haemolysin, flagellin, DNase, entF, algU, tcpI, acfB and rfaD were found in strain SSVR1601. Furthermore, factors responsible for iron uptake (fur, fepC and ccmC) and types II, IV and VI secretion systems were detected, which are likely responsible for the pathogenicity of SSVR1601. The antimicrobial resistance genes, bacA, tet34 and norM, were detected based on Antibiotic Resistance Genes Database. The phylogenetic analysis revealed SSVR1601 to be most closely related to V. rotiferianus strains CAIM577 and B64D1.  相似文献   

12.
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13.
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14.
In this study, we describe the complete mitochondrial genomes of Gyrodactylus brachymystacis and Gyrodactylus parvae infecting rainbow trout (Oncorhynchus mykiss) and the invasive topmouth gudgeon (Pseudorasbora parva), respectively. The two circular genomes have a common genome organization found in other Gyrodactylus species. Comparative analyses of mitochondrial genomes from six Gyrodactylus species were carried out to determine base composition, codon usage, transfer RNA and ribosomal RNA genes, major non‐coding regions, and nucleotide diversity within the genus. We also provide the first universal models of the secondary structures of rrnS and rrnL for this group thereby promoting utilization of these genetic markers. Universal primers provided herein can be used to obtain more mitochondrial information for pathogen identification and may reveal different levels of molecular phylogenetic inferences for this lineage.  相似文献   

15.
The autochthonous microbiota in the foregut, midgut and hindgut of juvenile grouper Epinephelus coioides following the dietary administration of probiotic Bacillus clausii for 60 days were assessed using polymerase chain reaction‐denaturing gradient gel electrophoresis (PCR‐DGGE). A complex and generally similar bacterial composition along the digestive tract of E. coioides was detected in the DGGE profiles, while several bacteria showed regional specialization. Similarity dendrogram revealed that the bacterial composition of the foregut was more similar to the midgut than the hindgut. Samples collected from the probiotic group and the control group showed generally similar DGGE patterns, while no significant difference in the total number of bands and Shannon index were observed between the probiotic group and the control group, suggested that probiotic B. clausii exerted no significant effect on the gut microbiota of E. coioides. However, various potentially beneficial bacteria, such as Enterococcus sp.‐like and Bacillus pumilus‐like bacterium were selectively stimulated by probiotic B. clausii, while some potential harmful species, like Staphylococcus sp.‐like and Vibrio ponticus‐like bacterium were depressed. These indicated that the gut microbiota was modified to some degree by probiotic B. clausii. Sequences analysis showed that the autochthonous gut bacteria of E. coioides could be classified into four groups, i.e. Proteobacteria, Firmicutes, Actinobacteria and unclassified bacteria.  相似文献   

16.
Betanodavirus infection was diagnosed in larvae of farm‐raised tilapia Oreochromis niloticus (L.), in central Thailand. Extensive vacuolar degeneration and neuronal necrosis were observed in histological sections with positive immunohistochemical staining for betanodavirus. Molecular phylogenetic analysis was performed based on the nucleotide sequences (1333 bases) of the capsid protein gene. The virus strain was highly homologous (93.07–93.88%) and closely related to red‐spotted grouper nervous necrosis virus (RGNNV).  相似文献   

17.
Seven marine cyanobacteria were isolated from two regions of the Gulf of Thailand and evaluated by the agar diffusion method for antibacterial activity. Inhibitory compound was purified from the crude methanol extract and its structure was elucidated based on extensive spectroscopic analysis, including IR, 1D and 2D NMR spectra as well as mass spectrometry. A novel antimicrobial compound produced by the marine cyanobacterium Leptolyngbya sp. LT19 was identified to be a 2‐hydroxyethyl‐11‐hydroxyhexadec‐9‐enoate which has so far never been reported in microorganisms. Biological assays revealed that this novel compound exhibited antibacterial activities against the Gram‐negative, persistent shrimp pathogens, Vibrio harveyi and V. parahaemolyticus with minimal inhibitory concentration of 250–1000 and 350–1000 μg mL?1 respectively.  相似文献   

18.
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19.
An attempt was made to determine the replication efficiency of white spot syndrome virus (WSSV) of shrimp in different organs of freshwater rice‐field crab, Paratelphusa hydrodomous (Herbst), using bioassay, PCR, RT‐PCR, ELISA, Western blot and real‐time PCR analyses, and also to use this crab instead of penaeid shrimp for the large‐scale production of WSSV. This crab was found to be highly susceptible to WSSV by intramuscular injection. PCR and Western blot analyses confirmed the systemic WSSV infection in freshwater crab. The RT‐PCR analysis revealed the expression of VP28 gene in different organs of infected crab. The indirect ELISA was used to quantify the VP28 protein in different organs of crab. It was found that there was a high concentration of VP28 protein in gill tissue, muscle, haemolymph and heart tissue. The copy number of WSSV in different organs of infected crab was quantified by real‐time PCR, and the results revealed a steady increase in copy number in different organs of infected crab during the course of infection. The viral inoculum prepared from different organs of infected crab caused significant mortality in tiger prawn, Penaeus monodon (Fabricius). The results revealed that this crab can be used as an alternate host for WSSV replication and production.  相似文献   

20.
When challenged with atypical Aeromonas salmonicida subsp. salmonicida, exposure of the common carp (Cyprinus carpio L.) to different humic‐rich compounds resulted in a significant reduction in infection rates. Specifically, in fish exposed to (i) humic‐rich water and sludge from a recirculating system, (ii) a synthetic humic acid, and (iii) a Leonardite‐derived humic‐rich extract, infection rates were reduced to 14.9%, 17.0% and 18.8%, respectively, as compared to a 46.8% infection rate in the control treatment. An additional set of experiments was performed to examine the effect of humic‐rich components on the growth of the bacterial pathogen. Liquid culture medium supplemented with either humic‐rich water from the recirculating system, the synthetic humic acid or the Leonardite humic‐rich extract resulted in a growth reduction of 41.1%, 45.2% and 61.6%, respectively, as compared to the growth of the Aeromonas strain in medium devoid of humic substances. Finally, in a third set of experiments it was found that while the innate immune system of the carps was not affected by their exposure to humic‐rich substances, their acquired immune system was affected. Fish, immunized against bovine serum albumin, displayed elevated antibody titres as compared to immunized carps which were not exposed to the various sources of humic substances.  相似文献   

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