A one-step duplex rRT-PCR assay for the simultaneous detection of grass carp reovirus genotypes I and II

Zeng, Weiwei; Wang, Yingying; Liang, Hongru; Liu, Cun; Song, Xinjian; Shi, Chunbin; Shu-qin, WU; Wang, Qing

doi:10.1016/j.jviromet.2014.08.024

Cited by 21 publications

(11 citation statements)

References 7 publications

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“…Over 50 fish species had been described to be susceptible to NNV (Shetty et al ., ). GCRV was a member of genus Aquareovirus within the family Reoviridae and could infect grass carp Ctenopharyngodon idella (Valenciennes 1844), black carp Mylopharyngodon piceus (Richardson 1846), top‐mouth gudgeon Pseudorasbora parva (Temminck & Schlegel 1846) and rare minnow Gobiocypris rarus (Ye & Fu 1983), resulting in severe economic losses in freshwater fish culture (Zeng et al ., ). SGIV, belonging to the genus Ranavirus of Iridoviridae family, was a major pathogen that had caused huge economic losses to the grouper aquaculture industry in Asian, especially in China and Southeast Asian countries (Song et al ., ; Huang et al ., ).…”

Section: Discussionmentioning

confidence: 97%

“…NNV was a pathogen that could cause almost 100% mortality of larval-stage marine and freshwater fish species (Nishizawa et al, 1997). Over 50 fish species had been described to be susceptible to NNV (Shetty et al, 1983), resulting in severe economic losses in freshwater fish culture (Zeng et al, 2014). SGIV, belonging to the genus Ranavirus of Iridoviridae family, was a major pathogen that had caused huge economic losses to the grouper aquaculture industry in Asian, especially in China and Southeast Asian countries (Song et al, 2004;Huang et al, 2011).…”

Section: Discussionmentioning

confidence: 99%

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Establishment of a cell line with high transfection efficiency from zebrafish Danio rerio embryos and its susceptibility to fish viruses

Jin

Chen

et al. 2017

Journal of Fish Biology

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A cell line ZBE3 isolated from a continuous cell culture derived from zebrafish Danio rerio blastomeres by clonal growth was characterized. ZBE3 cells had been subcultured for >120 passages since the initial primary culture of the blastomeres. The ZBE3 cells grow stably at temperature from 20 to 32° C with an optimum temperature of 28° C in ESM2 or ESM4 medium with 15% foetal bovine serum (FBS). The optimum FBS concentration for ZBE3 cell growth ranged from 15 to 20%. Cytogenetical analysis indicated that the modal chromosome number of ZBE3 cells was 50, the same as the diploid chromosome number of D. rerio. Significant cytopathic effect was observed in ZBE3 cells after infection with redspotted grouper nervous necrosis virus, Singapore grouper iridovirus and grass carp reovirus, and the viral replication in the cells was confirmed by real-time quantitative PCR and transmission electron microscopy, indicating the susceptibility of ZBE3 cells to the three fish viruses. After transfected with pEGFP-N3 plasmid, ZBE3 cells showed a transfection efficiency of about 40% which was indicated by the percentage of cells expressing green fluorescence protein. The stable growth, susceptibility to fish viruses as well as high transfection efficiency make ZBE3 cells be a useful tool in transgenic manipulation, fish virus-host cell interaction and immune response in fish.

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Section: Discussionmentioning

confidence: 97%

Section: Discussionmentioning

confidence: 99%

Establishment of a cell line with high transfection efficiency from zebrafish Danio rerio embryos and its susceptibility to fish viruses

Jin

Chen

et al. 2017

Journal of Fish Biology

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“…Specifically, the grass carp reovirus (GCRV), which causes grass carp hemorrhage disease, has received special attention because of the high mortality rate associated with it [ 21 ]. Many researches on GCRV have been reported [ 22 , 23 , 24 , 25 , 26 , 27 , 28 ], but not many effective drugs or vaccines have been developed until now. Moreover, the host–GCRV interaction is poorly understood, and the mechanism underlying GCRV infection is still unknown.…”

Section: Introductionmentioning

confidence: 99%

Deep Circular RNA Sequencing Provides Insights into the Mechanism Underlying Grass Carp Reovirus Infection

Zhang

Xiong

et al. 2017

IJMS

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Grass carp hemorrhagic disease, caused by the grass carp reovirus (GCRV), is a major disease that hampers the development of grass carp aquaculture in China. The mechanism underlying GCRV infection is still largely unknown. Circular RNAs (circRNAs) are important regulators involved in various biological processes. In the present study, grass carp were infected with GCRV, and spleen samples were collected at 0 (control), 1, 3, 5, and 7 days post-infection (dpi). Samples were used to construct and sequence circRNA libraries, and a total of 5052 circRNAs were identified before and after GCRV infection, of which 41 exhibited differential expression compared with controls. Many parental genes of the differentially expressed circRNAs are involved in metal ion binding, protein ubiquitination, enzyme activity, and nucleotide binding. Moreover, 72 binding miRNAs were predicted from the differentially expressed circRNAs, of which eight targeted genes were predicted to be involved in immune responses, blood coagulation, hemostasis, and complement and coagulation cascades. Upregulation of these genes may lead to endothelial and blood cell damage and hemorrhagic symptoms. Our results indicate that an mRNA–miRNA–circRNA network may be present in grass carp infected with GCRV, providing new insight into the mechanism underlying grass carp reovirus infection.

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“…Vaccination and pathogen surveillance are considered to be the most effective measures of protecting grass carp from this disease. At present, GCHD is mainly diagnosed by detecting of the virus specific nucleic acid, including PCR (Zeng, Wang, Wang, Zhang et al., ; Zeng, Wang, Wang, Shi et al., ; Zhang, Luo, Fang, & Wang, ), qPCR (Zeng et al., ), RT‐LAMP (Zeng, Wang, Wang, Zhang et al., ; Zeng, Wang, Wang, Shi et al., ) or NASBA‐ELISA (Zeng, Yao et al., ). Only a few of the immunological methods have been developed, and most are still for detecting virus detection (He et al., ; Jing et al., ; Wang et al., ; Zeng, Wang et al., ) There has been no feasible and effective method for the detection of the antibody level specific to GCRV II until now.…”

Section: Discussionmentioning

confidence: 99%

Development of a VP38 recombinant protein‐based indirect ELISA for detection of antibodies against grass carp reovirus genotype II (iELISA for detection of antibodies against GCRV II)

Zeng

Wang

Guo

et al. 2018

Journal of Fish Diseases

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Currently, serological assays for grass carp reovirus genotype II (GCRV‐II) diagnosis are not available. In this study, an indirect enzyme‐linked immunosorbent assay (ELISA) for the detection of antibodies against GCRV‐II was developed. The structural protein VP38 of GCRV‐II was used as the coating antigen. Monoclonal antibodies (mAb) against IgM of grass carp labelled with HRP were used as a secondary antibody. The antigen concentration and serum dilution were optimized using chess board titration. Furthermore, the specificity of indirect ELISA assay was confirmed by cross check with sera positive for other grass carp pathogens. In comparison with results obtained from indirect immunofluorescence assay (IFA) and Western blot by testing of 60 serum samples to evaluate the sensitivity and specificity of the ELISA, agreement between 90% and 96.7% was reached, respectively. A serological survey was performed using the assay with grass carp field serum samples. The seropositive rate of the 242 serum samples was 69.8%. In conclusion, the developed indirect ELISA is a very specific and sensitive test that will be useful for large‐scale serological surveys to detect indirectly GCRV II infections as well as to monitor the changes of antibody level after immunization.

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A one-step duplex rRT-PCR assay for the simultaneous detection of grass carp reovirus genotypes I and II

Cited by 21 publications

References 7 publications

Establishment of a cell line with high transfection efficiency from zebrafish Danio rerio embryos and its susceptibility to fish viruses

Establishment of a cell line with high transfection efficiency from zebrafish Danio rerio embryos and its susceptibility to fish viruses

Deep Circular RNA Sequencing Provides Insights into the Mechanism Underlying Grass Carp Reovirus Infection

Development of a VP38 recombinant protein‐based indirect ELISA for detection of antibodies against grass carp reovirus genotype II (iELISA for detection of antibodies against GCRV II)

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