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1.
RNA‐dependent RNA polymerase (RdRp), B2 and capsid genes of Macrobrachium rosenbergii nodavirus (MrNV) of Indian isolate were polymerase chain reaction amplified, cloned and sequenced. Expression of the MrNV fusion recombinant proteins of RdRp (44.5 kDa), B2 (32.2 kDa) and capsid (58.4 kDa) was confirmed by Western blot analysis using anti‐His mouse monoclonal antibodies. Polyclonal antibodies specific to purified recombinant MrNV capsid protein showed specificity against the capsid protein by Western blot. The protein sequence analysis of the partial RdRp gene of MrNV revealed the signature sequence along with the conserved core residues of the catalytic domain and indicated the presence of active sites, metal ion‐binding site and nucleic acid‐binding site residues. The Indian isolate of MrNV showed high RdRp and capsid gene sequence homology with the other MrNV geographical isolates. However, the Belize isolate was found to be the most distinct among the different geographical prawn nodavirus isolates due to the host specificity. Secondary structure prediction analysis of the MrNV capsid predicted it to be a DNA‐binding protein consisting of α helix (22.91%), extended strand (24.80%), β turn (5.39%) and random coil (46.90%) regions.  相似文献   

2.
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   

3.
The giant freshwater prawn, Macrobrachium rosenbergii, is an economically important and extensively cultured crustacean worldwide. The viral pathogens, Macrobrachium rosenbergii nodavirus (MrNV) and extra small virus (XSV) are responsible for causing severe mortalities in the hatchery and nursery phases. This study investigates the protection of postlarvae of freshwater against white tail disease (WTD) using plant extract derived from Cyanodon dactylon and the modulation of the prawn non‐specific immunity. To determine the immunomodulatory effect of C. dactylon extract, the prawn was injected with plant extract and various immunological parameters were estimated. The immunological parameters such as proPO, SOD, THC and clotting time were found to be significantly higher in the plant extract‐injected prawn when compared with control groups. The results of real time PCR analysis revealed up regulation on the expression proPO, SOD and lysozyme genes in MrNV and XSV challenged prawn postlarvae treated with C. dactylon extract. Infectivity experiment showed high relative per cent survival in MrNV and XSV‐challenged prawn postlarvae treated with C. dactylon extract. These results strongly indicate that the administration of C. dactylon plant extract enhances immunity of the prawn. Based on the results, this study recommends that the immersion of postlarvae in C. dactylon plant extract is a potential prophylactic agent against WTD.  相似文献   

4.
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5.
White tail disease (WTD) is found to cause immense economic losses in hatcheries, with mortalities often reaching 100% within 4 or 5 days. The pathogenic agents have been identified as Macrobrachium rosenbergii nodavirus (MrNV) associated with extra small virus (XSV), which are 27 and 15 nm in diameter respectively. The effects of some chemical disinfectants hydrogen ions (pH), heat and ultraviolet (UV) irradiation on the inactivation of MrNV and XSV were investigated. The viral inoculum exposed to UV irradiation for a period of 5 min and more was totally inactivated and failed to cause mortality in postlarvae of prawn. The viruses were totally inactivated by this high pH (8.5, 9 and 10). The viral suspension treated with sodium hypochloride, formalin, Benzalkonium chloride and Benzethonium chloride at the concentration of 200 ppm caused 100% mortality in postlarvae of prawn. Iodine was found to be effective to inactivate MrNV and XSV at the concentration of 100 ppm or more, whereas the viral suspension treated with iodine at the concentration of 50 ppm or less caused mortality in postlarvae. The infected postlarvae in treated and positive control groups showed positive by RT‐PCR for these viruses.  相似文献   

6.
Haemocyanin is a multi‐subunit protein complex found in the haemolymph and is involved in the immune system of crustaceans. In this study, a haemocyanin gene of Macrobrachium rosenbergii, designated MrHc, was successfully isolated. The MrHc gene contained an open reading frame (ORF) of 1,992 nucleotides, encoding a protein of 663 amino acid residues with a molecular mass of 76.5 kDa. The deduced amino acid sequence contained distinct structural motifs of the haemocyanin superfamily, including an all‐alpha domain, a copper‐containing domain and an immunoglobulin‐like domain. Based on the phylogenetic analysis, the MrHC protein demonstrated a close relationship with the haemocyanins of Palaemon carinicauda and Macrobrachium nipponense. The MrHc gene was expressed in various shrimp tissues, including the hepatopancreas, gill, haemocytes, stomach and muscle. After Macrobrachium rosenbergii nodavirus (MrNV) challenge tests, the MrHc gene was up‐regulated 237‐fold at day 2. A recombinant protein of the MrHc immunoglobulin‐like domain exhibited antibacterial activity against Vibrio vulnificus, V. parahaemolyticus, Aeromonas caviae, A. veronii, A. hydrophila and Bacillus cereus. This study suggested that MrHc may play important roles in the shrimp innate immune response to MrNV infection and bacterial infection.  相似文献   

7.
The freshwater prawn, Macrobrachium rosenbergii naturally lives in the freshwater, though it migrates to the brackish water environment during spawning that claimed to be resistant on a broad range of saline fluxes. However, little is known about the osmoregulatory patterns and the effect of an enzyme glutamine synthetase (GS) in M. rosenbergii under stress. Here, we described the identification and functional characterization of GS from M. rosenbergii (Mr‐GS) at molecular and protein levels. The identified Mr‐GS was comprised of 361 amino acids that phylogenetically shared the highest identity with other crustaceans and predicted to contain Gln‐synt_C and Gln‐synt_N domains at the respective terminal regions. Tissue distribution analysis in M. rosenbergii revealed that the Mr‐GS was highly expressed in muscle, and commonly existed in other examined tissues in the following order gills > heart > stomach > brain > haemolymph. Whereas, the mRNA of Mr‐GS was significantly up‐regulated in the muscle and gill tissues following challenges with either hyper (0 → 13‰), or hypo (13 → 0‰) osmotic stress at 3, 6 and 12 hr. Furthermore, the level of Glutamine concentration was positively correlated with the GS mRNA and protein expression patterns in hyper‐osmotic stress, whereas in hypo‐osmotic stress a slight decrease in the gills and maintained a level in the muscle tissues at 3, 6 and 12 hr post‐treatments. Our findings suggest that Mr‐GS potentially exhibited the osmoregulation responses in the gill and muscle tissues of M. rosenbergii throughout the time of osmotic stress, which will benefit for future study on osmoregulation.  相似文献   

8.
The protective efficacy of a DNA construct containing extra small virus antisense (XSVAS) gene of nodavirus encapsulated with chitosan nanoparticles (NPs) was investigated in giant freshwater prawn Macrobrachium rosenbergii (De Man, 1879). The delivery was carried out using oral and immersion methods. A plasmid concentration of 100 ng μL?1 when conjugated with chitosan NPs was found to be more effective in increasing the survivability of the infected prawn. The particle mean size, zeta potential and loading efficiency percentage were 297 nm, 27 mV and 85%, respectively. The ability of the chitosan to form a complex with the plasmid was studied by agarose gel electrophoresis. The NPs were characterized by atomic force microscopy (AFM). Persistence study showed the presence of the DNA construct up to 30th day post‐treatment. The oral treatment was found to be better than the immersion treatment for delivery of the chitosan‐conjugated DNA construct. This is probably the first report on the delivery of nanoconjugated DNA construct in M. rosenbergii, against nodavirus.  相似文献   

9.
Myostatin (MSTN) is an interesting negative growth‐regulating gene that has been well characterized in vertebrates but scantly described in invertebrates. The current study focuses on the downregulation of the MrMSTN gene and subsequently records any histological changes for giant freshwater prawn, Macrobrachium rosenbergii (Mr). In addition, the study also deals with the MrMSTN gene's influence on other growth‐related genes, which include myosin heavy chain, dystrophin‐dystroglycoprotein complex, tropomyosin, farnesoic acid o‐methyl transferase, arginine kinase, cyclophilin, and acyl CoA desaturase. The preliminary histological analysis following MrMSTN silencing favors muscle regeneration, which supports its functional role as a negative growth regulator and its significant effect on the expression of other growth‐related genes. Overall, our results show that the MrMSTN gene could therefore be a potential target for gene manipulation aimed at enhancing the growth and muscle development of M. rosenbergii, which could be beneficial in increasing the total mass production in the postlarva phase at the hatchery level.  相似文献   

10.
The purpose of this study was to evaluate the resistance to vibriosis, growth, survival and tolerance to stress of the selected prawn, second generation, compared to a non‐selected control. The first generation of selected giant freshwater prawn, which has 10.4% higher of resistance, was used to attain disease resistant freshwater prawn (Macrobrachium rosenbergii) generation through challenge test‐based selection. Resistance test was conducted by infecting the prawn (mean body weight of 10.29 ± 1.40 g) with pathogenic Vibrio harveyi (5 × 105 cfu prawn?1). The growth and survival of the prawn were evaluated by rearing the two populations of prawn in both nursery and grow‐out phases. Stress tolerance test was done by evaluating the viability of postlarvae exposed to environmental stressors, i.e. temperature, salinity, NH3 and formaldehide. Post‐challenge survival of the selected prawn (55.0 ± 5.0%) was about 46% higher than that of the control (37.5 ± 7.5%). The survival of the selected prawn in nursery culture (77.16 ± 0.841%) was significantly higher (p < .05) than that of the control (51.31 ± 2.938%), while the survival in grow‐out culture was similar (p > .05). The growth of selected prawn (4.99 ± 0.03% day?1) was significantly higher than that of the control (4.81 ± 0.05% day?1). There was no difference between treatments on the tolerance level against the tested environmental stressor. Overall data suggested that the selected prawn showed better performance in growth and resistance against vibriosis.  相似文献   

11.
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12.
An 8‐week study was conducted to evaluate the effects of different feeding patterns with dietary Clostridium butyricum supplementation on growth performance, antioxidant and non‐specific immune responses in freshwater prawn, Macrobrachium rosenbergii (0.39 ± 0.001 g). There were four feeding methodologies: feeding basal diet continuously (P1); feeding diet with 500 mg/kg C. butyricum continuously (P2); feeding diet with 500 mg/kg C. butyricum 5 days after 2 days of basal diet (P3) and feeding diet with 500 mg/kg C. butyricum 2 days after 5 days of basal diet (P4). The results revealed that prawns in P3 had the highest weight gain rate (WGR) and lowest feed conversion ratio (FCR) than the other groups. Haemolymph total protein levels and superoxide dismutase activity increased significantly in P2, P3 and P4 groups, while malondialdehyde content and anti‐superoxide anion levels decreased significantly compared to control. The mRNA expression of intestinal dorsal and Toll in P2 and P3 groups decreased significantly compared to control. Prawns in P3 exhibited improved growth performance, increased antioxidant capacity and enhanced immune function. We concluded that feeding diet with 500 mg/kg C. butyricum for 5 days after 2 days of basal diet was recommended for M. rosenbergii.  相似文献   

13.
This study evaluates the effect of probiotics, Zymetin, on the immune efficiency of Macrobrachium rosenbergii juvenile against pathogenic Vibriospp. and Aeromonasspp. The experiment was conducted in glass aquaria with same level of feeding under different treatments, that is, negative control (Cn), positive control with Vibrio spp. (Cv) and Aeromonasspp. (Ca), prawn juveniles fed with probiotics (Zymetin) at 5 g/kg of feed but without pathogen (T1), probiotic fed prawn challenged with Vibrio spp. (T2) and Aeromonasspp. (T3). The results demonstrated that T1 showed higher survival rate, total haemocyte count, non‐granular haemocyte count, phagocytic activity, and clearance efficiency compared to other treatment groups. In contrast, decreased number of small and large granular haemocyte was observed in T1. Despite that, THC was found to be significantly different (p < 0.05) among all the controls and lowest was in Cv, followed by Ca. Besides, phagocytic activity and clearance efficiency against Vibrio spp. and Aeromonasspp. decreased significantly in Cn, Ca, and Cv fed M. rosenbergiijuvenile, while the values were found to be higher in T1 followed by T2 and T3, in that order. In addition, the challenge test showed an increasing trend of total and beneficial bacterial density as well as declining trend of some harmful bacteria in the water and gut of prawn in all the groups tested (p < 0.05).  相似文献   

14.
The effect of dietary supplementation of probiotic bacterium Bacillus licheniformis on the histopathological changes in Macrobrachium rosenbergii juveniles (4.0 ± 0.02 g) challenged with known pathogenic strain of Vibrio alginolyticus are reported. Two isocaloric basal diets supplemented with probiotic bacteria B. licheniformis (1.0 × 109 cfu/g feed) and other without probiotic supplementation were fed to the M. rosenbergii juveniles for 45 days. The histological observations revealed no significant changes in the hepatopancreas and gut tissues of both the experimental and the control groups which indicate that the present bacterium is a safe candidate probiont for the host. Prawns were challenged with V. alginolyticus after 45 days of feeding with probiotic diet. The histopathological studies of the hepatopancreas revealed that M. rosenbergii fed with probiotic‐supplemented diet showed less changes as compared to the prawns fed with control diet on second and fourth day of post‐experimental challenge with V. alginolyticus. Histopathological observations revealed that the gills of the prawns fed with control diet were severely affected in comparison to the prawns fed with probiotic‐supplemented diet after challenging with V. alginolyticus. Results from this study revealed the improved protection by dietary incorporation of B. licheniformis in reducing the histopathological manifestations due to V. alginolyticus infection in freshwater prawn.  相似文献   

15.
The different products of Eichhornia crassipes leaves including dried E. crassipes powder (DEP), hot‐water treated E. crassipe (HTE), hot‐water extract of E. crassipe (ECE) and dreg of hot‐water extract of E. crassipe extract (dECE) were produced and incorporated into the diet of the prawn, Macrobrachium rosenbergii, as an immunostimulant. Results showed that prawn fed the HTE‐, ECE‐ and dECE‐containing diets for 4 months had increased total haemocyte count, different haemocyte count, phenoloxidase activity, respiratory bursts, superoxide dismutase activity, glutathione peroxidase activity especially in HTE and ECE treatments. The phagocytic activity and clearance efficiency against Lactococcus garvieae of prawn fed the HTE‐ and ECE‐containing diets were significantly higher than those of prawn fed the control diet at 2–4 months. The relative percentage survival of prawn fed the DEP‐, HTE‐, ECE‐ and dECE‐containing diets for 4 months following 144 h challenging with L. garvieae were 19.0%, 38.1%, 38.1% and 33.3%. It was concluded that E. crassipes leaves containing an active component which was easily extracted by hot water can enhance innate immunity and resistance against pathogen of M. rosenbergii by dietary long‐term administration, and the administration of HTE in the diet was the best strategy due to the availability and convenience.  相似文献   

16.
Microcystins accumulate in aquatic organisms and can be transferred to higher trophic levels, eventually affecting vector animals and consumers. We examined three levels of an aquatic food chain (Microcystis aeruginosa, Daphnia magna and Macrobrachium rosenbergii) to identify the transfer efficiency and risk of microcystin on prawns. Samples were analysed using ultra performance liquid chromatography‐mass spectrometry (MS)/MS and microcystin‐LR (MC‐LR) distributions in prawn tissues were studied. The results showed that prawns accumulate MC‐LR both directly from M. aeruginosa and indirectly through D. magna which was pre‐exposed to M. aeruginosa. MC‐LR was detected in the gills, digestive tracts and hepatopancreas of the prawns 2 h after exposure. MC‐LR accumulated in prawns to 0.49 ± 0.04 μg g?1 dry weight in hepatopancreas within 24 h, while it was not detected in muscle samples, and rarely appeared in blood samples in such a short period. Although MC‐LR was not detected in muscle, the head including hepatopancreas of the prawns accumulated troublesome amounts of MC‐LR. These results demonstrate that microcystis blooms in prawn farming potentially pose a risk to human consumers, although prawns may be exposed to the bloom for a very short time, hence regular monitoring of blue green algae population is recommended.  相似文献   

17.
A 50‐day growth trial was conducted to evaluate the efficacy of Chlorella vulgaris (Beijerinck) as an ingredient in the diets of giant freshwater prawn Macrobrachium rosenbergii (De Man) postlarvae (PL30). Immune response (total haemocyte count and prophenoloxidase activity) was also assessed by subjecting postlarvae to a challenge test against Aeromonas hydrophila (Chester) for 14 days. Iso‐nitrogenous and iso‐lipidic test diets were prepared using a fishmeal‐based‐positive control diet (D0) and four basal diets with inclusion levels of 2% (D2), 4% (D4), 6% (D6) and 8% (D8) C. vulgaris. Postlarvae of M. rosenbergii were randomly stocked (mean initial body weight of 0.19 ± 0.02 g) in 30‐L tanks in three replicates per dietary treatment for evaluation of growth performance. Another set of postlarvae (mean initial body weight of 1.25 ± 0.02 g) was randomly distributed in 95‐L tanks in three replicates per dietary treatment for the assessment of immune response. Results showed that specific growth rate was significantly higher (P < 0.05) in postlarvae fed D4 and D6. Variations in values for carcass protein, lipid, moisture and ash were also evident. Postlarvae fed diets with Chlorella showed increased prophenol oxidase activity and total haemocyte counts. Moreover, survival rate after challenge with A. hydrophila was significantly increased (P < 0.05). Inclusion of C. vulgaris in diets enhanced immune response and resistance of M. rosenbergii postlarvae against A. hydrophila infection.  相似文献   

18.
We examined the effects of salinity stress and ammonia stress on alteration of the haemocyte count, phenoloxidase activity, expressions of immune‐related genes including prophenoloxidase (proPO), crustin, heat shock protein 70 (HSP70), and expressions of stress‐responsive neurohormone (Bur‐α and Bur‐β) in the thoracic and abdominal ganglia of giant river prawn Macrobrachium. These parameters of prawn that subjected to salinity stress (transferred from 0‰ to 5‰ and 10‰), and subjected to ammonia‐nitrogen (ammonia‐N) stress (transferred from 0 to 0.262 mg/L and 0.786 mg/L) were examined after 0, 3, 6, 24, 72 and 168 hr respectively. During the initial period of 3, 6 and 24 hr, granulocyte haemocyte (granular and semi‐granular hemocyte) count and PO activity significantly decreased, while expressions of Bur‐α and Bur‐β significantly increased. After 24 hr, granulocyte haemocyte count and PO activity significantly increased, whereas expressions of Bur‐α and Bur‐β significantly decreased. The expressions of proPO, crustin and HSP70 were significantly downregulated in the prawn that subjected to salinity stress and ammonia‐N stress at all time periods of 3–168 hr. In conclusion, changes in the granulocyte haemocyte count of M. rosenbergii following salinity stress and ammonia‐N stress are closely associated with the changes of Bur‐α and Bur‐β expressions.  相似文献   

19.
The production performance of genetically improved farmed tilapia (GIFT, Oreochromis niloticus) and freshwater prawn (Macrobrachium rosenbergii) in periphyton‐based systems were studied in farmers' ponds at Mymensingh, Bangladesh. Fifteen ponds (200–300 m2 area and 1.0–1.5 m in depth) were used to compare five stocking ratios in triplicate: 100% GIFT, 75% GIFT plus 25% prawn, 50% GIFT plus 50% prawn, 25% GIFT plus 75% prawn and 100% prawn. Ponds were stocked at a total density of 20 000 GIFT and/or prawn ha?1. Bamboo poles (mean diameter 6.2 cm and 5.5 pole m?2) were posted in pond bottoms vertically as periphyton substrate. Periphyton biomass in terms of dry matter (DM), ash‐free DM and chlorophyll a were significantly higher in ponds stocked with prawn alone than in ponds with different combinations of GIFT and prawn. Survival of GIFT was significantly lower in ponds stocked with 100% GIFT (monoculture) whereas, that of prawn was significantly higher in its monoculture ponds indicating detrimental effects of GIFT on prawn's survival. Individual weight gains for both species were significantly higher in polyculture than in monoculture. The highest total fish and prawn yield (1623 kg GIFT and 30 kg prawn ha?1) over 125–140 days culture period was recorded in ponds with 75% GIFT and 25% prawn followed by 100% GIFT alone (1549 kg ha?1), 50% GIFT plus 50% prawn (1114 kg GIFT and 68 kg prawn ha?1), 25% GIFT plus 75% prawn (574 kg GIFT and 129 kg prawn ha?1) and 100% prawn alone (157 kg ha?1). This combination also gave the highest economic return. Therefore, a stocking ratio of 75% GIFT plus 25% prawn at a total density of 20 000 ha?1 appeared to be the best stocking ratio in terms of fish production as well as economics for a periphyton‐based polyculture system.  相似文献   

20.
Long‐term cryopreservation of the giant freshwater prawn, Macrobrachium rosenbergii, spermatophores using glycerol (Gly) and ethylene glycol (EG) as cryoprotective agents (CPAs) was studied. The tolerance of sperm to cryopreservation was evaluated on the basis of sperm survival and fertilizing ability. The survival of the sperm was determined by trypan blue staining, while the fertilizing ability was assessed from artificial insemination of the cryopreserved spermatophores. The rates of embryo survival on day 5 after spawning and of spermatophores capable of producing embryos survived to hatching were determined. Storage of spermatophores at ?20°C without CPA for a short period of up of 1–5 days decreased the sperm survival significantly and did not preserve fertilizing ability. Preservation at ?20°C in the presence of 10% or 20% Gly or of 10% or 20% EG offered a simple and efficient short‐term storage up to 10 days. For a long‐term storage, cryopreservation in the presence of 20% EG at ?196°C was more efficient than at ?20°C. High sperm survival rates and high fertilizing ability were recorded from those cryopreserved at ?196°C for up to 150 days. High sperm survival rates with moderate levels of fertilizing ability were obtained from those cryopreserved at ?20°C for not more than 30 days. The results indicate that preservation at ?196°C with 20% EG is a suitable procedure for long‐term storage of the giant freshwater prawn spermatophores.  相似文献   

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