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1.
Fish Shellfish Immunol ; 98: 429-437, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31988017

RESUMO

Oxyeleotris marmoratus iridovirus (OMIV) and Oxyeleotris marmoratus rhabdovirus (OMRV) are the two major causative agents of disease leading to massive mortality and severe economic losses in marbled sleepy goby (Oxyeleotris marmoratus) industry. It's urgent to develop an effective vaccine against these fatal diseases. In this study, we developed bivalent inactivated vaccine against OMIV and OMRV and evaluated its protective effect in Oxyeleotris marmoratus. The intraperitoneally vaccinated fish were protected against challenge with OMIV and OMRV with both relative percent survival (RPS) of 100%. In addition, deep RNA sequencing was used to analyze the transcriptomic profiles of the spleen tissues at progressive time points post-vaccination with bivalent inactivated vaccine and challenge with OMIV and OMRV infection. Results showed that adaptive immune response was induced in Oxyeleotris marmoratus injected with bivalent inactivated vaccine. Furthermore, robust adaptive immune responses were also detected in vaccinated fish at 7 d and 2 d post-challenge with OMIV and OMRV. Taken together, these results indicated that bivalent inactivated vaccine activated adaptive immune responses in Oxyeleotris marmoratus, and provided protection against OMIV and OMRV lethal challenge.

2.
Microb Pathog ; 138: 103822, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31669501

RESUMO

The virus inactivation test is a critical skill in inactivated vaccine production. Active viruses produced viral mRNA in susceptible cells or the host can be used to infer whether a DNA virus is replicating by RT-PCR. But it is generally difficult to avoid genomic DNA contamination in the samples. However, the use of primers spanning an intron is an effective alternative for virus inactivation test. Therein, a nested RT-PCR was developed to detect active ISKNV in the inactivated vaccine. At first, the transcriptome analysis of CPB cell infected with ISKNV revealed several gaps in some viral transcripts compared to ISKNV genome. One intron in ORF003L with 80 bp (designated IN-3) was confirmed by PCR and sequencing analysis. Then, two primer sets (primer A and primer B) spanning the IN-3 intron were designed to detect ISKNV transcription. The nested RT-PCR conditions were optimized with 0.4 µM primer A and 0.2 µM primer B, and 68 °C and 55 °C for annealing temperature, respectively. The sensitivity results indicated that the nested RT-PCR could detect one copy of live ISKNV propagating in CPB cells for seven days. The nested RT-PCR method was more sensitive and accurate than the method of blind passages in cells and fish challenge experiments. Together, above results indicate that this assay is a time-saving, labor-extensive and cost-effective for inactivation test of ISKNV in killed vaccine production.

3.
Biomolecules ; 9(9)2019 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-31480692

RESUMO

Glucose is a main carbon and energy source for virus proliferation and is usually involved in the glycolysis, pentose phosphate pathway (PPP), and tricarboxylic acid cycle (TCA cycle) pathways. In this study, we investigated the roles of glucose-related metabolic pathways during the replication of infectious spleen and kidney necrosis virus (ISKNV), which has caused serious economic losses in the cultured Chinese perch (Siniperca chuatsi) industry. We found that ISKNV infection enhanced the metabolic pathways of the PPP and the TCA cycle at the early stage of the ISKNV infection cycle and enhanced the glycolysis pathway at the late stage of the ISKNV infection cycle though the comprehensive analysis of transcriptomics, proteomics, and metabolomics. The advanced results proved that ISKNV replication induced upregulation of aerobic glycolysis at the late stage of ISKNV infection cycle and aerobic glycolysis were required for ISKNV multiplication. In addition, the PPP, providing nucleotide biosynthesis, was also required for ISKNV multiplication. However, the TCA cycle involving glucose was not important and necessary for ISKNV multiplication. The results reported here provide new insights into viral pathogenesis mechanism of metabolic shift, as well as antiviral treatment strategies.

4.
Metabolites ; 9(9)2019 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-31487859

RESUMO

Infectious spleen and kidney necrosis virus (ISKNV) has caused serious economic losses in the cultured mandarin fish (Siniperca chuatsi) industry in China. Host metabolism alteration induced by disease infection may be the core problem of pathogenesis. However, to date, little is known about the disease-induced fish metabolism changes. In this study, we first reported ISKNV, the fish virus, induced metabolism alteration. The metabolomics profiles of Chinese perch brain cells (CPB) post-ISKNV infection at progressive time points were analyzed using the UHPLC-Q-TOF/MS technique. A total of 98 differential metabolites were identified. In the samples harvested at 24 hours post-infection (hpi; the early stage of ISKNV infection), 49 differential metabolites were identified comparing with control cells, including 31 up-regulated and 18 down-regulated metabolites. And in the samples harvested at 72 hpi (the late stage of ISKNV infection), 49 differential metabolites were identified comparing with control cells, including 27 up-regulated and 22 down-regulated metabolites. These differential metabolites were involved in many pathways related with viral pathogenesis. Further analysis on the major differential metabolites related to glucose metabolism and amino acid metabolism revealed that both glucose metabolism and glutamine metabolism were altered and a metabolic shift was determined from glucose to glutamine during ISKNV infection cycle. In ISKNV-infected cells, CPB cells prefer to utilize glucose for ISKNV replication at the early stage of infection, while they prefer to utilize glutamine to synthetize lipid for ISKNV maturation at the late stage of infection. These findings may improve the understanding of the interaction between ISKNV and host, as well as provide a new insight for elucidating the ISKNV pathogenic mechanism.

5.
Microb Pathog ; 135: 103617, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31283962

RESUMO

The bluegill sunfish, Lepomis macrochirus, is an important aquacultural and recreational species in southern China because of its excellent taste, rapid growth rate, and good looks. At present, few pathogens are known to affect the bluegill sunfish. However, an iridovirus-like disease recently caused heavy losses to the bluegill sunfish aquaculture industry in Guangdong, China. We report that a virus, designated BSMIV-SD-20171020, was isolated from diseased bluegill sunfish in China. The isolate was efficiently propagated in a Chinese perch brain (CPB) cell line. The cytopathic effect was observed, the MCP gene PCR amplified, and the virus observed with electron microscopy. Its viral titer in CPB cells reached 104.13 TCID50 mL-1. The mortality rate was 100% when bluegill sunfish were challenged with BSMIV-SD-20171020 at a dose of 103.13 TCID50/fish. A histopathological examination revealed basophilic hypertrophied cells in the intestine, liver, and spleen. A nucleotide sequence alignment and phylogenetic analysis of the major capsid protein revealed that isolate BSMIV-SD-20171020 is the species Infectious spleen and kidney necrosis virus (ISKNV), in the genus Megalocytivirus.


Assuntos
Infecções por Vírus de DNA/veterinária , Infecções por Vírus de DNA/virologia , Doenças dos Peixes/virologia , Iridoviridae/classificação , Iridoviridae/isolamento & purificação , Perciformes/virologia , Animais , Aquicultura , Encéfalo , Proteínas do Capsídeo/classificação , Proteínas do Capsídeo/genética , Linhagem Celular , China , Infecções por Vírus de DNA/patologia , Doenças dos Peixes/patologia , Peixes , Iridoviridae/genética , Iridoviridae/patogenicidade , Rim/patologia , Rim/virologia , Fígado/patologia , Fígado/virologia , Percas , Filogenia , Análise de Sequência de DNA/veterinária , Baço/patologia , Baço/virologia
6.
Microb Pathog ; 129: 146-151, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30731189

RESUMO

To distinguish between three types of Siniperca chuatsi rhabdovirus (SCRV) viral RNA (vRNA, cRNA, and mRNA) and investigate SCRV transcription and replication dynamics in Chinese perch brain CPB cells, a novel, strand-specific, reverse transcriptase quantitative real-time PCR (RT-qPCR) assay was established. The method is based on strand-specific reverse transcription, using tagged primers to add a 'tag' sequence at the 5' end. We used the 'tag' sequence as the forward primer and a strand-specific reverse primer to quantify the three types of RNA. Three types of synthetic viral RNA were used as reference standards for validation and quantification. These assays were optimized to produce a standard curve from 102 to 107 copies/µL, with an efficiency of 91-101% and an R2 value of 0.9949-0.9999. The coefficients of variation for repeatability and reproducibility were less than 2.85% and 5.52%, respectively. Using this method, specific target RNA was detected at a 3500-70,000 fold higher level than other types of RNA. This method was also used to evaluate the dynamics of vRNA, cRNA and mRNA synthesis in CPB cells infected with SCRV. The results indicate that the intracellular dynamics of vRNA, cRNA and mRNA are different. In the earliest phase of SCRV infection, all three types of viral RNA increased very slowly. The copy number of vRNA and mRNA increased exponentially from 4 h post infection, while cRNA increased from 6 h post infection. The amount of cRNA was lower than vRNA and mRNA throughout the infection. The novel, strand-specific RT-qPCR method developed in this study provides critical data to aid the understanding of transcription and replication during SCRV infection.


Assuntos
Doenças dos Peixes/virologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Infecções por Rhabdoviridae/veterinária , Rhabdoviridae/fisiologia , Transcrição Genética , Replicação Viral , Animais , Encéfalo/virologia , Percas , RNA Viral/genética , Reprodutibilidade dos Testes , Rhabdoviridae/genética , Infecções por Rhabdoviridae/virologia , Sensibilidade e Especificidade
7.
Fish Shellfish Immunol ; 84: 1075-1082, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30423456

RESUMO

MicroRNAs are non-coding RNAs, which widely participate in biological processes. In recent years, Siniperca chuatsi rhabdovirus (SCRV) has caused mass mortality in Chinese perch (Siniperca chuatsi). To identify specific miRNAs involved in SCRV infection, deep sequencing of microRNA on Chinese perch brain cell line (CPB) with or without SCRV infection were performed at 6 and 12 h post of infection (hpi). Totally 382 miRNAs were identified, including 217 known miRNA aligned with zebrafish miRNAs and 165 novel miRNAs by MiRDeep2 program. Of which 15 and 35 differentially-expressed miRNAs were determined respectively to 6 and 12 hpi. Nine miRNAs were selected randomly from the differentially-expressed miRNAs and validated by quantitative real-time PCR (qRT-PCR). These results were consistent with the microRNA sequencing results. Besides, target genes of 98 differentially-expressed miRNAs were predicted. Three of miRNAs (miR-122, miR-214, miR-135a) were selected, and its effects were analyzed in CPC cells transfected with appropriate miRNA mimics/inhibitors to evaluate its regulation effects by qRT-PCR and western blot. The results demonstrated that miR-214 inhibited the replication of SCRV, while miR-122 promoted the replication of SCRV and there was no correlation between the miR-135a and SCRV replication. These results will pave a new way for the development of effective strategies against the SCRV infection.


Assuntos
Encéfalo/virologia , Doenças dos Peixes/imunologia , Imunidade Inata/genética , MicroRNAs/genética , Percas , Infecções por Rhabdoviridae/veterinária , Animais , Encéfalo/imunologia , Encéfalo/metabolismo , Doenças dos Peixes/virologia , MicroRNAs/metabolismo , Rhabdoviridae/fisiologia , Infecções por Rhabdoviridae/imunologia , Infecções por Rhabdoviridae/virologia , Análise de Sequência de RNA/veterinária
8.
Fish Shellfish Immunol ; 79: 102-111, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29733959

RESUMO

Infectious spleen and kidney necrosis virus (ISKNV) has caused significant losses in the cultured mandarin fish (Siniperca chuatsi) industry. The molecular mechanisms that underlie interaction between ISKNV and hosts are not fully understood. In this study, the proteomic profile of CPB cells at progressive time points after ISKNV infection was analyzed by isobaric tags for relative and absolute quantitation (iTRAQ). A total of 2731 proteins corresponding to 6363 novel peptides (false discovery rate <0.01) were identified. In the samples harvested 24 h (early-stage) and 72 h (late-stage) post-infection, 232 and 199 differentially expressed proteins were identified comparing with mock-infected cells, respectively. Western-blotting analysis of several proteins as G6PDH, ß-tubulin and RPL11 were done to validate iTRAQ data. Among those differentially expressed proteins, several glucose metabolism-related enzymes, including glucose-6-phosphate dehydrogenase (G6PDH), pyruvate dehydrogenase phosphatase (PDP) and fumarate hydratase (FH), were up-regulated, while pyruvate dehydrogenase kinase (PDK) and enolase (ENO) were down-regulated at 24 h poi, suggesting that ISKNV enhanced glucose metabolism in CPB cells in early-stage infection. Simultaneously, expression of apoptosis-related proteins including Caspase 8, phosphoinositide 3-kinases (PI3Ks), and regulatory-associated protein of mTOR-like isoform X3 changed upon ISKNV infection, indicating that ISKNV induced apoptosis of CPB cells. Autophagy-related proteins including LC3 and PI3Ks were up-regulated at 24 h poi, indicating that ISKNV induced autophagy of CPB cells in early-stage infection. These findings may improve the understanding of ISKNV and host interaction and help clarify its pathogenesis mechanisms.


Assuntos
Apoptose , Autofagia , Doenças dos Peixes/imunologia , Glucose/metabolismo , Perciformes/imunologia , Transdução de Sinais , Animais , Linhagem Celular , Infecções por Vírus de DNA/imunologia , Iridoviridae/fisiologia , Proteômica
9.
Microb Pathog ; 112: 269-273, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28987623

RESUMO

Ranavirus has become a noticeable threat to both farmed and natural populations of fish and amphibians. Herein, we reported that 3 strains of novel viruses, designated as ScRIV-GM-20150902, CmRIV-XT-20150917 and ScRIV-ZS-20151201, were isolated from diseased Chinese perch and snakehead fish in China. Efficient propagation of these isolates were determined in Chinese perch brain (CPB) cell line by the means of cytopathic effect observation, PCR amplification and electron microscopy observation. And their viral titers in CPB cells reached 108.13 TCID50 ml-1, 107.71 TCID50 ml-1 and 107.94 TCID50 ml-1, respectively. While the challenge experiment results showed that 3 isolates resulted in 100% mortality of Chinese perch after virus infection. Electron microscopy analysis showed that two kinds of viral inclusion bodies (intracytoplasmic and intranuclear inclusion body) were observed in infected CPB cells. Sequence alignment and phylogenetic analysis of major capsid protein gene sequences of isolates revealed that these isolates belonged to the species Santee-Cooper Ranavirus.


Assuntos
Infecções por Vírus de DNA/veterinária , Doenças dos Peixes/virologia , Peixes/virologia , Percas/virologia , Ranavirus/classificação , Ranavirus/isolamento & purificação , Ranavirus/patogenicidade , Animais , Ascite/patologia , Ascite/virologia , Sequência de Bases , Encéfalo/patologia , Encéfalo/virologia , Proteínas do Capsídeo/genética , Linhagem Celular , China , Infecções por Vírus de DNA/patologia , Infecções por Vírus de DNA/virologia , DNA Viral , Corpos de Inclusão Viral , Mesentério/patologia , Mesentério/virologia , Microscopia Eletrônica de Transmissão , Filogenia , Ranavirus/genética , Alinhamento de Sequência , Virulência
10.
Fish Shellfish Immunol ; 70: 536-544, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28923524

RESUMO

A number of size variants of the p53 protein have been described in mammal, but little is known about alternative splicing of p53 expression and function in the fish. In our previous study, the immune defense and antiviral responses of p53 had been determined in mandarin fish (Siniperca chuatsi). However, the role of its splicing variants remains unknown. In the present study, the organization of mandarin fish p53 (Sc-p53) genome sequence was determined and a novel splice variant was characterized. The Sc-p53 genomic sequence was composed of 5543 bp, containing 11 exons and 10 introns, which was similar to other species. Then, a 1106 bp full-length cDNA of a novel splice variant p53 from mandarin fish (designed as Sc-p53I6) was cloned and characterized. Quantitative real-time PCR assays revealed that Sc-p53I6 was expressed in all tissues examined, and it was most abundant in the gill, hemocyte and hind kidney. Western blotting analysis revealed that Sc-p53I6 protein was abundant in liver, trunk kidney, hind kidney, stomach and heart. In addition, the regulation of Sc-p53I6 gene expression after virus infection was determined and characterized. The results showed twice rise expression pattern of Sc-p53I6 in CPB cells and spleen of mandarin fish in response to infectious kidney and spleen necrosis virus (ISKNV). However, a different expression pattern, once rise, of Sc-p53I6 in response to Siniperca chuatsi rhabdovirus (SCRV) infection was found. The mRNA expression of Sc-p53I6 was significantly up-regulated in CPB at 4 h and spleen of mandarin fish at 12 h post-infection. These results will shed a new light on antiviral response mechanisms of p53 in mandarin fish.


Assuntos
Doenças dos Peixes/imunologia , Regulação da Expressão Gênica , Imunidade Inata , Perciformes/genética , Perciformes/imunologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/imunologia , Processamento Alternativo , Animais , Linhagem Celular , Infecções por Vírus de DNA/imunologia , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Iridoviridae/fisiologia , RNA Mensageiro/genética , Rhabdoviridae/fisiologia , Infecções por Rhabdoviridae/imunologia , Distribuição Tecidual
11.
Arch Virol ; 162(9): 2829-2834, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28550433

RESUMO

The Rhabdoviridae is a diverse family of negative-sense single-stranded RNA viruses which infects mammals, birds, reptiles, fish, insects and plants. Herein, we reported the isolation and characterization of 6 novel viruses from diseased fish collected from China including SCRV-QY, SCRV-SS, SCRV-GM, CmRV-FS, MsRV-SS, OmbRV-JM. The typical clinical symptom of diseased fish was hemorrhaging. Efficient propagation of these isolates in a Chinese perch brain cell line was determined by means of observation of cytopathic effect, RT-PCR and electron microscopy. Sequence alignment and phylogenetic analysis of the complete G protein sequences revealed that these isolates were clustered into one monophyletic lineage belonging to the species Siniperca chuatsi rhabdovirus.


Assuntos
Doenças dos Peixes/virologia , Variação Genética , Rhabdoviridae/genética , Rhabdoviridae/patogenicidade , Animais , China/epidemiologia , Doenças dos Peixes/epidemiologia , Doenças dos Peixes/patologia , Peixes
12.
J Aquat Anim Health ; 29(2): 89-94, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28379065

RESUMO

Grass Carp reovirus (GCRV) is one of the most pathogenic agents among aquareovirus isolates and has the ability to cause a severe epidemic outbreak of hemorrhagic disease, thus resulting in both a high mortality rate during the culture of Grass Carp Ctenopharyngodon idella and an enormous economic loss. Aptamers have been demonstrated to have strong promising applications in antiviral drug development. In the present study, a complementary DNA fragment encoding the S10 gene of GCRV was cloned. The S10 protein was expressed and purified. Aptamers for S10 protein were selected by the method of selective evolution of ligands by exponential enrichment (SELEX), and their characteristics and antiviral actions were examined. All targeting-selected aptamers formed a similar structure, forming a 5-7 base loop at the terminus. The results show that the aptamers could inhibit the GCRV infection. The most significant inhibitory effect was obsereved when the aptamers were added to the cell culture for 1 h before the cells were infected by GCRV. Our data showed that these novel molecular agents could be considered suitable candidates for anti-GCRV therapy. Received August 23, 2016; accepted February 5, 2017.


Assuntos
Aptâmeros de Nucleotídeos/antagonistas & inibidores , Carpas , Doenças dos Peixes/prevenção & controle , Infecções por Reoviridae/veterinária , Reoviridae/genética , Animais , Infecções por Reoviridae/prevenção & controle
13.
Microb Pathog ; 107: 98-105, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28323153

RESUMO

Infectious spleen and kidney necrosis virus (ISKNV) is one of the major epidemiological agents that had caused great economic loss in Chinese perch (Siniperca chuatsi). In this study, a specific TaqMan real-time PCR was developed using a pair of primers and a TaqMan probe specific to the ORF007 gene of ISKNV to rapidly detect ISKNV copies in Chinese perch samples. This assay was optimized to produce linearity from 8.75 × 108 to 8.75 × 101 copies in standard curve with an efficiency of 98% and a R2 value of 0.9999. Moreover, the minimum detection limit of this assay was 10,000 times more sensitive than that of conventional PCR method. The coefficients of variation of intra- and inter-assay repeatability were less than 2.4% and 3.3%, respectively. The viral distribution in different tissues of diseased Chinese perch was evaluated by TaqMan real-time PCR method and the highest level of viral copies was detected in spleen. Among the 76 diseased Chinese perch clinical samples, 35 and 29 were positive samples based on the TaqMan real-time PCR and conventional PCR methods, respectively, indicating that the TaqMan real-time PCR was more sensitive than conventional PCR. Therefore, the TaqMan real-time PCR should be a useful tool for the early surveillance and quantitation of ISKNV.


Assuntos
Doenças dos Peixes/diagnóstico , Rim/virologia , Necrose/virologia , Perciformes/virologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Baço/virologia , Animais , Linhagem Celular , China , Primers do DNA/genética , Infecções por Vírus de DNA/diagnóstico , Infecções por Vírus de DNA/veterinária , Infecções por Vírus de DNA/virologia , Doenças dos Peixes/virologia , Sensibilidade e Especificidade
14.
Fish Shellfish Immunol ; 60: 25-32, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27856327

RESUMO

Autophagy plays important functions in viral replication and pathogenesis. In this study, we investigated the role of autophagy in the replication of infectious kidney and spleen necrosis virus (ISKNV), an agent that has caused devastating losses in Chinese perch (Siniperca chuatsi) industry. We found that ISKNV infection triggered the complete autophagic process, as demonstrated by microtubule-associated protein 1 light chain 3B II (LC3B-II) conversion, an increased accumulation of punctate GFP-LC3-expressing cells, a higher number of autophagosome-double-membrane vesicles in the cytoplasm, and increased levels of autophagic flux in CPB cells. Then, we investigated the role of autophagy in the process of ISKNV replication. Results showed that inducing autophagy by rapamycin promoted ISKNV replication and proteins synthesis but decreased extracellular virus yields. While, blocking autophagosome-lysosome fusion by chloroquine (CQ) promoted infectious virus yields in culture supernatant. These results offer insight into the complex interactions between ISKNV and host cell, providing new insights into viral pathogenesis and antiviral treatment strategies.


Assuntos
Autofagia , Infecções por Vírus de DNA/veterinária , Doenças dos Peixes/virologia , Iridoviridae/fisiologia , Replicação Viral , Animais , Linhagem Celular/virologia , Infecções por Vírus de DNA/virologia , Percas
15.
Fish Shellfish Immunol ; 56: 286-293, 2016 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-27436517

RESUMO

Co-infection with infectious spleen and kidney necrosis virus (ISKNV) and Aeromonas hydrophila is becoming ever more widespread in Chinese perch (Siniperca chuatsi) aquaculture industry, so that it's necessary to develop the combined vaccine against ISKNV and A. hydrophila disease. The surface display of heterologous on bacteria using anchoring motifs from outer membranes proteins has already been explored as an effective delivery system of viral antigens. In present study, the ISKNV orf086 gene, which is verified as a protective antigen, was inserted into ompA gene cassette of A. hydrophila GYK1 strain by homologous recombination. And an ompA-orf086 fusion A. hydrophila mutant strain K28 was constructed. Then the ISKNV orf086 was verified to express on the surface of A. hydrophila K28 by RT-PCR, western blot and indirect immunofluorescence assay. Next, Chinese perch were intraperitoneally inoculated with formalin inactivated A. hydrophila k28 emulsified with ISA763 adjuvant with a dose of 9 × 10(8) CFU per fish. Transcriptional analysis of non-specific and specific immune related genes revealed that the expression levels of IRF-7, IRAK1, Mx, Viperin, Lysozyme and IgM were strongly up-regulated in Chinese perch post-inoculation. In addition, specific antibodies were detected by ELISA, and the results showed that antibody titer against ISKNV or A. hydrophila reached the highest with 1:800 or 1:1200 on 14dpv, respectively. Lymphocyte proliferation were detected by MTT methods, and the results showed that the SI values of AH-K28 vaccinated group to three different stimulators were significantly higher than those of control group. At last, protective efficacy were determined by challenge trials. The cumulative mortality rates of vaccinated groups were significantly lower than the control one (P < 0.05) after ISKNV or A. hydrophila challenge, and the relative percentage survival (RPS) value was 73.3% and 60%, respectively. This system provides a novel approach to the surface display of heterologous antigenic proteins on A. hydrophila and suggests the possibility to use the recombinant K28 strain as a combined vaccine against ISKNV and A. hydrophila infection.


Assuntos
Aeromonas hydrophila/imunologia , Vacinas Bacterianas/imunologia , Doenças dos Peixes/prevenção & controle , Iridoviridae/imunologia , Perciformes , Vacinas Virais/imunologia , Aeromonas hydrophila/genética , Animais , Coinfecção/imunologia , Coinfecção/microbiologia , Coinfecção/prevenção & controle , Coinfecção/veterinária , Infecções por Vírus de DNA/imunologia , Infecções por Vírus de DNA/prevenção & controle , Infecções por Vírus de DNA/veterinária , Infecções por Vírus de DNA/virologia , Doenças dos Peixes/imunologia , Doenças dos Peixes/microbiologia , Doenças dos Peixes/virologia , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/microbiologia , Infecções por Bactérias Gram-Negativas/prevenção & controle , Infecções por Bactérias Gram-Negativas/veterinária , Organismos Geneticamente Modificados/genética , Organismos Geneticamente Modificados/imunologia , Distribuição Aleatória , Vacinas Combinadas/imunologia , Proteínas Virais/imunologia
16.
Bing Du Xue Bao ; 32(1): 108-20, 2016 Jan.
Artigo em Chinês | MEDLINE | ID: mdl-27295892

RESUMO

Cyprinid herpesvirus 3 (CyHV-3) is the causative agent of an extremely contagious and aggressive disease afflicting common corp Cyprinus carpio L. termed koi herpesvirus disease (KHVD). Since it was first reported in 1997, the virus has spread worldwide rapidly, leading to enormous financial losses in industries based on common carp and koi carp. This review summarizes recent advances in CyHV-3 research on the etiology, epidemiology, pathogenesis, diagnosis, prevention, and control of KHVD.


Assuntos
Doenças dos Peixes/virologia , Infecções por Herpesviridae/veterinária , Herpesviridae/fisiologia , Animais , Doenças dos Peixes/diagnóstico , Peixes/classificação , Peixes/virologia , Herpesviridae/genética , Herpesviridae/isolamento & purificação , Infecções por Herpesviridae/diagnóstico , Infecções por Herpesviridae/virologia
17.
Bing Du Xue Bao ; 32(6): 800-9, 2016 11.
Artigo em Chinês | MEDLINE | ID: mdl-30004655

RESUMO

Viruses "hijack" cellular metabolism to complete their proliferation. Glucose is an important source of energy and carbon in the synthesis of precursor molecules in host cells, and its metabolism is regulated dramatically during virus infection. Here, we reviewed the mechanism of virus infection that alters glucose transport, expression of glucose metabolism-related genes (glycolysis, pentose phosphate pathway, gluconeogenesis) in cells, as well as islet cells and insulin receptors. It provides references for study of virus-altering cellular glucose metabolism.


Assuntos
Glucose/metabolismo , Viroses/metabolismo , Animais , Glicólise , Humanos , Viroses/virologia , Fenômenos Fisiológicos Virais , Vírus/genética
18.
Virol J ; 11: 178, 2014 Oct 07.
Artigo em Inglês | MEDLINE | ID: mdl-25293723

RESUMO

Grass carp reovirus (GCRV) is the causative agent of grass carp hemorrhage and causes significant loss of fingerlings. However, little is known about how the virus is distributed in organs and tissues. The aim of the present study was to investigate the distribution of different GCRV stains in tissues and organs of grass carp. The pathogenicity and tissue distribution of GCRV were monitored after intraperitoneal administration. The study showed a distribution of GCRV in different tissues and organs, particularly in the liver, spleen, kidney, intestine, and muscle, which had a higher number of viral RNA copies during the sixth to ninth days. The kidney had the highest numbers of viral RNA copies, as high as 24000 copies. Until the fourteenth day, nearly no viral RNA copies could be detected. This study defined the virus distribution in different tissues of grass carp inoculated by i.p. and supplied clues for the pathogenesis of GCRV.


Assuntos
Carpas/virologia , Doenças dos Peixes/virologia , Infecções por Reoviridae/veterinária , Reoviridae/patogenicidade , Animais , Rim/virologia , Fígado/virologia , Músculos/virologia , Reoviridae/fisiologia , Infecções por Reoviridae/virologia , Baço/virologia , Virulência
19.
J Virol Methods ; 210: 32-5, 2014 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-25205265

RESUMO

Hemorrhagic disease of grass carp, caused by grass carp reovirus (GCRV), leads to severe economic losses in the grass carp farming industry in China. GCRV has been divided into three genotypes based on genome sequence. Genotypes I and II (GCRV-1 and GCRV-II, respectively) are the dominant genotypes and co-infections of GCRV-I and GCRV-II are common in grass carp aquaculture. A one-step duplex real-time reverse transcriptase polymerase chain reaction (rRT-PCR) assay was developed for simultaneous detection of GCRV-I and GCRV-II. The PCR assay is suitable for early diagnosis of grass carp hemorrhagic disease and for epidemiological surveillance. The detection limit of the assay is 10 copies for both GCRV-I and GCRV-II, which is as high as single-target rRT-PCR and higher than conventional RT-PCR. No cross reactivity with other GCRV subtypes or other viruses was observed. One hundred and twelve samples from grass carp suspected of hemorrhagic disease were collected from South and Central China. Eleven samples were positive for GCRV-I by RT-PCR alone, and fourteen samples were positive by single-target and duplex rRT-PCR. Forty two samples were positive for GCRV-II by RT-PCR alone and forty seven samples were positive by single-target and duplex rRT-PCR. Mixed infections were found in eight samples when analyzed by RT-PCR alone and in ten samples analyzed by single-target and duplex rRT-PCR. The duplex rRT-PCR system provides a sensitive and specific method to detect and differentiate between GCRV-I and GCRV-II in a single sample. This rRT-PCR assay could be a useful tool for the routine diagnosis of these two viruses and for epidemiology studies in grass carp aquaculture.


Assuntos
Carpas/virologia , Doenças dos Peixes/virologia , Infecções por Reoviridae/veterinária , Reoviridae/isolamento & purificação , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Animais , Aquicultura , China , Coinfecção/veterinária , Genótipo , Reoviridae/genética , Infecções por Reoviridae/virologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos
20.
Arch Virol ; 159(9): 2469-73, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24748006

RESUMO

A new rhabdovirus, tentatively designated as hybrid snakehead rhabdovirus C1207 (HSHRV-C1207), was first isolated from a moribund hybrid snakehead (Channa maculata×Channa argus) in China. We present the complete genome sequence of HSHRV-C1207 and a comprehensive sequence comparison between HSHRV-C1207 and other rhabdoviruses. Sequence alignment and phylogenetic analysis revealed that HSHRV-C1207 shared the highest degree of homology with Monopterus albus rhabdovirus and Siniperca chuatsi rhabdovirus. All three viruses clustered into a single group that was distinct from the recognized genera in the family Rhabdoviridae. Our analysis suggests that HSHRV-C1207, as well as MARV and SCRV, should be assigned to a new rhabdovirus genus.


Assuntos
Peixes/virologia , Genoma Viral , RNA Viral/genética , Rhabdoviridae/genética , Análise de Sequência de DNA , Animais , Quimera , China , Análise por Conglomerados , Dados de Sequência Molecular , Filogenia , Rhabdoviridae/isolamento & purificação , Homologia de Sequência
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