Publications by authors named "Dongmin Zhao"

22 Publications

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Transcriptome analysis reveals new insight of duck Tembusu virus (DTMUV)-infected DF-1 cells.

Res Vet Sci 2021 Jul 29;137:150-158. Epub 2021 Apr 29.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Diagnosis, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing, 210014, China.

Duck Tembusu virus (DTMUV) is a newly emerging pathogenic flavivirus that has caused huge economic losses to the duck industry in China since 2010. Moreover, the infection has spread rapidly, resulted in a potential public health concern. To improve our understanding of the host cellular responses to virus infection and the pathogenesis of DTMUV infection, we used RNA-Seq to detect the gene changes in DF-1 cells infected and mock-infected with DTMUV. A total of 663 differentially-expressed genes (DEGs) were identified in DTMUV-infected compared with mock-infected DF-1 cells at 24 h post-infection (hpi), among which 590 were up regulated and 73 were down regulated. Gene Ontology analysis indicated that the DEGs were mainly involved in cellular process, immune system processes, metabolic processes, and signal-organism process. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that the DEGs were mainly involved in several signaling pathways such as Toll-like receptor signaling, Jak-STAT signaling, RIG-I-like receptor signaling and AGE-RAGE signaling pathway. Moreover, some selected DEGs were further confirmed by real-time PCR and the results were consistent with the sequencing data. To our knowledge, this study is the first to analyze the transcriptomic change in DF-1 cells following DTMUV infection. We believe that our research provides useful information in better understanding the host response to DTMUV infection and the inherent mechanism of DTMUV replication and pathogenicity.
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http://dx.doi.org/10.1016/j.rvsc.2021.04.028DOI Listing
July 2021

Nanomaterials for Tumor Hypoxia Relief to Improve the Efficacy of ROS-Generated Cancer Therapy.

Front Chem 2021 19;9:649158. Epub 2021 Mar 19.

State Key Laboratory for Chemistry and Molecular Engineering of Medicinal Resources, School of Chemistry and Pharmaceutical Sciences, Guangxi Normal University, Guilin, China.

Given the fact that excessive levels of reactive oxygen species (ROS) induce damage to proteins, lipids, and DNA, various ROS-generating agents and strategies have been explored to induce cell death and tumor destruction by generating ROS above toxic threshold. Unfortunately, hypoxia in tumor microenvironment (TME) not only promotes tumor metastasis but also enhances tumor resistance to the ROS-generated cancer therapies, thus leading to ineffective therapeutic outcomes. A variety of nanotechnology-based approaches that generate or release O continuously to overcome hypoxia in TME have showed promising results to improve the efficacy of ROS-generated cancer therapy. In this minireview, we present an overview of current nanomaterial-based strategies for advanced cancer therapy by modulating the hypoxia in the TME and promoting ROS generation. Particular emphasis is put on the O supply capability and mechanism of these nanoplatforms. Future challenges and opportunities of design consideration are also discussed. We believe that this review may provide some useful inspiration for the design and construction of other advanced nanomaterials with O supply ability for overcoming the tumor hypoxia-associated resistance of ROS-mediated cancer therapy and thus promoting ROS-generated cancer therapeutics.
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http://dx.doi.org/10.3389/fchem.2021.649158DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8089386PMC
March 2021

Analysis of the microRNA expression profiles of chicken dendritic cells in response to H9N2 avian influenza virus infection.

Vet Res 2020 Oct 17;51(1):132. Epub 2020 Oct 17.

Key Laboratory of Veterinary Biological Engineering and Technology of Ministry of Agriculture, National Center for Engineering Research of Veterinary Bio-products, Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, 50 Zhongling Street, Nanjing, 210014, Jiangsu, China.

MicroRNA (miRNA) plays a key role in virus-host interactions. Here, we employed deep sequencing technology to determine cellular miRNA expression profiles in chicken dendritic cells infected with H9N2 avian influenza virus (AIV). A total of 66 known and 36 novel miRNAs were differently expressed upon H9N2 infection, including 72 up-regulated and 30 down-regulated miRNAs. Functional analysis showed that the predicted targets of these miRNAs were significantly enriched in several pathways including endocytosis, notch, lysosome, p53, RIG-I-like and NOD-like receptor signaling pathways. These data provide valuable information for further investigating the roles of miRNA in AIV pathogenesis and host defense response.
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http://dx.doi.org/10.1186/s13567-020-00856-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7568386PMC
October 2020

Peptide inhibitors of tembusu virus infection derived from the envelope protein.

Vet Microbiol 2020 Jun 7;245:108708. Epub 2020 May 7.

College of Animal Science and Technology, Shandong Agricultural University, Tai'an, Shandong province, PR China; Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, Tai'an, Shandong, PR China; Shandong Provincial Engineering Technology Research Center of Animal Disease Control and Prevention, Tai'an, Shandong, PR China. Electronic address:

The outbreak and spread of Tembusu virus (TMUV) has caused very large losses in the waterfowl-breeding industry since 2010. The viral envelope (E) protein, the principal surface protein of viral particles, plays a vital role in viral entry and fusion. In this study, two peptides derived from domain II (DII) and the stem of the TMUV envelope protein, TP1 and TP2, respectively, were tested for their antiviral activity. TP1 and TP2 inhibited TMUV infection in BHK-21 cells, and their 50% inhibitory concentrations (IC) were 14.19 mg/L and 7.64 mg/L, respectively. Viral inhibition assays in different cell lines of avian origin showed that the inhibitory effects of TP1 and TP2 are not cell type dependent. Moreover, TP2 also exhibited inhibitory activity against Japanese encephalitis virus (JEV) infection. The two peptides inhibited antibody-mediated TMUV infection of duck peripheral blood lymphocytes. Co-immunoprecipitation assays and indirect enzyme-linked immunosorbent assays (ELISAs) indicated that both peptides interact with the surface of the TMUV virion. RNase digestion assays confirmed the release of viral RNA following incubation with TP1, while incubation with TP1 or TP2 interfered with the binding between TMUV and cells. Taken together, these results show that TP1 and TP2 may be developed into antiviral treatments against TMUV infection.
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http://dx.doi.org/10.1016/j.vetmic.2020.108708DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7204726PMC
June 2020

Tembusu virus enters BHK-21 cells through a cholesterol-dependent and clathrin-mediated endocytosis pathway.

Microb Pathog 2020 Oct 12;147:104242. Epub 2020 May 12.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing, China. Electronic address:

Tembusu virus (TMUV) is a newly emerging flavivirus and has caused significant economic loss to the poultry industry in China. To date, the entry of TMUV into host cells remains poorly understood. Here, the mechanism of TMUV entry into BHK-21 cells was investigated. The depletion of cellular cholesterol by methyl-β-cyclodextrin led to a significant decline in the titers and RNA levels of the infectious TMUV. This reduction was restored by supplementation of exogenous cholesterol. Membrane cholesterol depletion mainly blocked viral internalization but not attachment. However, viral infection was unaffected by genistein treatment or caveolin-1 silencing by small interfering RNA. In addition, clathrin-mediated endocytosis might be utilized in TMUV entry given that the viral infection was inhibited by knockdown of clathrin heavy chain and treatment of chlorpromazine (CPZ). Moreover, the number of internalized virus particles decreased under CPZ treatment. Dynasore inhibited TMUV entry suggesting a role for dynamin. Our results reveal that TMUV entry into BHK-21 cells is dependent on cholesterol, clathrin and dynamin but not caveolae.
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http://dx.doi.org/10.1016/j.micpath.2020.104242DOI Listing
October 2020

Global gene expression analysis data of chicken dendritic cells infected with H9N2 avian influenza virus.

Data Brief 2020 Jun 16;30:105430. Epub 2020 Mar 16.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, and Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing, Jiangsu, China.

This data article reports the global gene expression analysis data of chicken DCs infected with H9N2 avian influenza virus (AIV) compared with mock infection. The differentially expressed genes (DEGs), and the data of GO enrichment analysis and KEGG pathway analysis for DEGs were reported here. In addition, some of these DEGs associated with innate immune response and antigen presentation were also verified by qPCR. The replication of H9N2 AIV in DCs, and the viability kinetic of DCs during H9N2 AIV infection, and the primers for qPCR were also reported in this data article. The data presented here was used on the research article entitled "Transcriptomic profile of chicken bone marrow-derive dendritic cells in response to H9N2 avianinfluenza A virus".
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http://dx.doi.org/10.1016/j.dib.2020.105430DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7152653PMC
June 2020

Transcriptomic profile of chicken bone marrow-derive dendritic cells in response to H9N2 avian influenza A virus.

Vet Immunol Immunopathol 2020 Feb 9;220:109992. Epub 2019 Dec 9.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, and Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing, Jiangsu, 50 Zhongling Street, 210014, China. Electronic address:

Avian influenza subtype H9N2 infection is a mild but highly contagious disease that is associated with a decrease in the efficacy of vaccine interventions, and an increase in susceptibility to secondary infections in poultry. However, the immune evasion mechanism of H9N2 avian influenza viruses (AIVs) in chickens is poorly understood. Dendritic cells (DCs) are immune cells of major importance, involved in innate immune responses against viruses, but also in the setting of adaptive immune response due to their high ability to present viral antigen. Therefore, in the present study we used high-throughput RNA-sequencing technology at the transcriptome level to identify the differentially expressed genes (DEGs) between chicken DCs infected with H9N2 virus and mock-infected DCs. We identified 4151 upregulated DEGs and 2138 downregulated DEGs. Further enrichment analysis showed that the upregulated DEGs were enriched in the biological processes mainly involved in signal transduction, transmembrane transport, and innate immune/inflammatory responses. In contrast, the downregulated DEGs were associated with the biological processes mainly including metabolic process, and MHC class I antigen processing and presentation. In addition, 49 of these immune-related DEGs were validated by reverse transcription quantitative PCR (RT-qPCR). Collectively, these data suggest that H9N2 virus infection may enhance the signal transduction, and innate immune responses in chicken DCs, but impair their metabolic functions and antigen-presenting responses, which provide helpful insight into the pathogenesis of H9N2 AIVs in chickens and managing this infection in poultry farms.
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http://dx.doi.org/10.1016/j.vetimm.2019.109992DOI Listing
February 2020

The ubiquitin-proteasome system is necessary for the replication of duck Tembusu virus.

Microb Pathog 2019 Jul 1;132:362-368. Epub 2019 May 1.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, National Center for Engineering Research of Veterinary Bio-products, Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, 225009, PR China. Electronic address:

Duck Tembusu virus (DTMUV) is a newly emerging pathogenic flavivirus that has caused massive economic losses to the duck industry in China. The cellular factors required for DTMUV replication have been poorly studied. The ubiquitin-proteasome system (UPS), the major intracellular proteolytic pathway, mediates diverse cellular processes, including endocytosis and signal transduction, which may be involved in the entry of virus. In the present study, we explored the interplay between DTMUV replication and the UPS in BHK-21 cells and found that treatment with proteasome inhibitor (MG132 and lactacystin) significantly decreased the DTMUV progency at the early infection stage. We further revealed that inhibition of the UPS mainly occurs on the level of viral protein expression and RNA transcription. In addition, using specific siRNAs targeting ubiquitin reduces the production of viral progeny. In the presence of MG132 the staining for the envelope protein of DTMUV was dramatically reduced in comparison with the untreated control cells. Overall, our observations reveal an important role of the UPS in multiple steps of the DTMUV infection cycle and identify the UPS as a potential drug target to modulate the impact of DTMUV infection.
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http://dx.doi.org/10.1016/j.micpath.2019.04.044DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7126904PMC
July 2019

The unfolded protein response induced by Tembusu virus infection.

BMC Vet Res 2019 Jan 22;15(1):34. Epub 2019 Jan 22.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Jiangsu Province, 210014, People's Republic of China.

Background: Tembusu virus (TMUV), classified in the genus Flavivirus, causes reduced egg production and neurological problems in poultry. Flavivirus replication depends on the host endoplasmic reticulum (ER) and induces ER stress that leads to activation of the cellular unfolded protein response (UPR), an important signalling pathway that regulates many biological functions involved in viral pathogenesis and innate immunity. However, the mechanism of TMUV-induced UPR activation remains unclear.

Results: In this study, we systematically investigated the three UPR pathways in TMUV-infected BHK-21 cells. Our results showed that expression of glucose-related protein 78 (GRP78) and GRP94 was upregulated during the course of TMUV infection. We then demonstrated that TMUV activated the PERK pathway in the early stage of infection, resulting in upregulation of ATF4, GADD34 and CHOP, with CHOP induction leading to caspase-3 activation. We also found the IRE1 pathway to be activated, leading to splicing of X box binding protein 1 (XBP1) mRNA and enhanced expression of p58. Finally, we observed increased expression of ATF6 and activity of ER stress-response elements, suggesting stimulation of the ATF6 pathway. In addition, ATF6 pathway activation correlated with the induction of downstream chaperones calnexin, calreticulin, ERp57 and PDI. UPR activity was also observed by the marked elevation in GRP78 and sXBP1 levels in TMUV-infected DF-1 cells.

Conclusions: This is the first report that TMUV infection-induced ER stress activates three branches of the UPR, and these results lay the foundation for elucidating the pathogenesis of TMUV and understanding the inherent mechanism of TMUV infection as well as the host response.
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http://dx.doi.org/10.1186/s12917-019-1781-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6343269PMC
January 2019

Identification and immunogenic evaluation of T cell epitopes based on tembusu virus envelope protein in ducks.

Virus Res 2018 09 18;257:74-81. Epub 2018 Sep 18.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Jiangsu Province, PR China; Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing, Jiangsu Province, PR China. Electronic address:

Newly emerging tembusu virus (TMUV) is a severe threat to poultry industry and causes huge economic losses. Humoral and cell-mediated immunity are both play vital roles in TMUV infection. Up to now, there has been no report on identification of T cell epitopes of the TMUV. In this work, we identified T cell epitopes within TMUV envelope (E) protein using synthesized peptides predicted in silico. A total of ten peptides could stimulate TMUV-specific T cells in murine ELISPOT and duck lymphocyte proliferation assay. Subsequently, DNA vaccine containing these T cell epitopes was constructed (pVAX-T) and the expression of multiepitope protein was confirmed by transfection of BHK-21 cells in vitro. Ducks were administrated intramusclarly to evaluated the immunologic effect of pVAX-T. In ducks immunized with pVAX-T, antibody against TMUV was undetectable, but the expression level of cytokines (IL-2, IL-6, IFN-γ) was upregulated both in peripheral blood lymphocytes and spleen. Furthermore, TMUV challenge revealed that cell-mediated immune response sitmulated by pVAX-T contributed to protection against TMUV infection. The identification of these T cell epitopes will contribute to designing epitope vaccine for preventing infection of TMUV and possibly provide the basis for further studies on cell-mediate immune response activated by TMUV.
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http://dx.doi.org/10.1016/j.virusres.2018.09.008DOI Listing
September 2018

Screening and identification of B-cell epitopes within envelope protein of tembusu virus.

Virol J 2018 09 17;15(1):142. Epub 2018 Sep 17.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, 50 Zhongling Street, Nanjing City, Jiangsu Province, 210014, People's Republic of China.

Background: Tembusu virus is a newly emerging flavivirus that caused egg-drop syndrome in ducks in China. TMUV envelope protein is a major structural protein locates at the surface of tembusu virus particle. During tembusu virus infection, envelope protein plays a pivotal role in induction of neutralizing antibody. However, B cell epitopes within envelope protein have not been well studied.

Method: A series of 13 peptides derived from E protein of tembusu virus were synthesized and screened by Dot blot with tembusu virus-positive duck serum. Potential B-cell epitopes were respectively fused with GST tag and expressed in E. coli. The immunogenicity and protective efficiency of epitopes were assessed in ducks.

Results: Dot blot assay identified the peptides P21 (amino acids 301-329), P23 (amino acids 369-387), P27 (amino acids 464-471) and P28 (amino acids 482-496) as potential B-cell epitopes within the envelope protein of tembusu virus. Immunization of prokaryotically expressed epitopes elicited specific antibodies in ducks and the specific antibody elicited by P21, P27 and P28 could neutralized tembusu virus. In addition, protective test suggested that P21 and P27 could completely protect immunized ducks from TMUV challenge.

Conclusion: Four potential B cell epiotpes within tembusu virus envelope protein were identified and analyzed in vitro and in vivo. It was demonstrated that two of them (P21 and P27) could elicit neutralizing antibodies in ducks and offer complete protection against tembusu virus challenge. This findings will contribute to the development of epitope vaccine for tembusu virus prevention.
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http://dx.doi.org/10.1186/s12985-018-1052-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6142368PMC
September 2018

Identification of Glucose-Regulated Protein 78 (GRP78) as a Receptor in BHK-21 Cells for Duck Tembusu Virus Infection.

Front Microbiol 2018 9;9:694. Epub 2018 Apr 9.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing, China.

Since 2010, outbreak and spread of tembusu virus (TMUV) caused huge losses to the breeding industry of waterfowl in several provinces of China. In this study, we identify the glucose-regulated protein 78 (GRP78) as a receptor in BHK-21 cells for duck TMUV infection. Using cell membrane from BHK-21 cells, a TMUV-binding protein of approximately 70 kDa was observed by viral overlay protein binding assay (VOPBA). LC-MS/MS analysis and co-immunoprecipitation identified GRP78 as a protein interacting with TMUV. Antibody against GRP78 inhibited the binding of TMUV to the cell surface of BHK-21 cells. Indirect immunofluorescence studies showed the colocalization of GRP78 with TMUV in virus-infected BHK-21 cells. We found that GRP78 over-expression increased TMUV infection, whereas GRP78 knockdown by using a specific small interfering RNA inhibited TMUV infection in BHK-21 cells. Taken together, our results indicate that GRP78 is a novel host factor involved in TMUV entry.
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http://dx.doi.org/10.3389/fmicb.2018.00694DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5903163PMC
April 2018

Identification of determinants that mediate binding between Tembusu virus and the cellular receptor heat shock protein A9.

J Vet Sci 2018 Jul;19(4):528-535

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing 210014, China.

Heat shock protein A9 (HSPA9), a member of the heat shock protein family, is a putative receptor for Tembusu virus (TMUV). By using Western blot and co-immunoprecipitation assays, E protein domains I and II were identified as the functional domains that facilitate HSPA9 binding. Twenty-five overlapping peptides covering domain I and domain II sequences were synthesized and analyzed by using an HSPA9 binding assay. Two peptides showed the capability of binding to HSPA9. Dot blot assay of truncated peptides indicated that amino acid residues 19 to 22 and 245 to 252 of E protein constitute the minimal motifs required for TMUV binding to HSPA9. Importantly, peptides harboring those two minimal motifs could effectively inhibit TMUV infection. Our results provide insight into TMUV-receptor interaction, thereby creating opportunities for elucidating the mechanism of TMUV entry.
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http://dx.doi.org/10.4142/jvs.2018.19.4.528DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6070589PMC
July 2018

MC1568 Enhances Histone Acetylation During Oocyte Meiosis and Improves Development of Somatic Cell Nuclear Transfer Embryos in Pig.

Cell Reprogram 2018 02;20(1):55-65

1 Institute of Animal Science , Jiangsu Academy of Agricultural Sciences, Nanjing, P.R. China.

An increasing number of studies have revealed that histone deacetylase (HDAC) mediated histone deacetylation is important for mammalian oocyte development. However, nonselective HDAC inhibitors (HDACi) were applied in most studies; the precise functions of specific HDAC classes during meiosis are poorly defined. In this study, the class IIa-specific HDACi MC1568 was used to reveal a crucial role of class IIa HDACs in the regulation of histone deacetylation during porcine oocyte meiosis. Besides, the functions of HDACs and histone acetyltransferases in regulating the balance of histone acetylation/deacetylation were also confirmed during oocyte maturation. After the validation of nontoxicity of MC1568 in maturation rate, spindle morphology, and chromosome alignment, effects of MC1568 on developmental competence of porcine somatic cell nuclear transfer (SCNT) embryos were evaluated, and data indicated that treatment with 10 μM MC1568 for 12 hours following electrical activation significantly enhanced the blastocyst rate and cell numbers. Moreover, results showed that optimal MC1568 treatment increased the H4K12 acetylation level in SCNT one cells and two cells. In addition, MC1568 treatment stimulated expression of the development-related genes OCT4, CDX2, SOX2, and NANOG in SCNT blastocysts. Collectively, our investigation uncovered a critical role of class IIa HDACs in the regulation of histone deacetylation during oocyte meiosis. Furthermore, for the first time, we showed that MC1568 can improve the in vitro development of porcine SCNT embryos. These findings provide an alternative HDACi for improving animal cloning efficiency and may shed more light on nuclear reprogramming.
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http://dx.doi.org/10.1089/cell.2017.0023DOI Listing
February 2018

Two Genetically Similar H9N2 Influenza A Viruses Show Different Pathogenicity in Mice.

Front Microbiol 2016 4;7:1737. Epub 2016 Nov 4.

Key Laboratory of Veterinary Biological Engineering and Technology, National Center for Engineering Research of Veterinary Bio-products, Institute of Veterinary Medicine, Ministry of Agriculture, Jiangsu Academy of Agricultural SciencesNanjing, China; Jiangsu Key Laboratory of Zoonosis, Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and ZoonosesYangzhou, China.

H9N2 Avian influenza virus has repeatedly infected humans and other mammals, which highlights the need to determine the pathogenicity and the corresponding mechanism of this virus for mammals. In this study, we found two H9N2 viruses with similar genetic background but with different pathogenicity in mice. The A/duck/Nanjing/06/2003 (NJ06) virus was highly pathogenic for mice, with a 50% mouse lethal dose (MLD) of 10 50% egg infectious dose (EID), whereas the A/duck/Nanjing/01/1999 (NJ01) virus was low pathogenic for mice, with a MLD of >10 EID. Further studies showed that the NJ06 virus grew faster and reached significantly higher titers than NJ01 and . Moreover, the NJ06 virus induced more severe lung lesions, and higher levels of inflammatory cellular infiltration and cytokine response in lungs than NJ01 did. However, only 12 different amino acid residues (HA-K157E, NA-A9T, NA-R435K, PB2-T149P, PB2-K627E, PB1-R187K, PA-L548M, PA-M550L, NP-G127E, NP-P277H, NP-D340N, NS1-D171N) were found between the two viruses, and all these residues except for NA-R435K were located in the known functional regions involved in interaction of viral proteins or between the virus and host factors. Summary, our results suggest that multiple amino acid differences may be responsible for the higher pathogenicity of the NJ06 virus for mice, resulting in lethal infection, enhanced viral replication, severe lung lesions, and excessive inflammatory cellular infiltration and cytokine response in lungs. These observations will be helpful for better understanding the pathogenic potential and the corresponding molecular basis of H9N2 viruses that might pose threats to human health in the future.
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http://dx.doi.org/10.3389/fmicb.2016.01737DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5096341PMC
November 2016

Identification of heat shock protein A9 as a Tembusu virus binding protein on DF-1 cells.

Virus Res 2017 01 29;227:110-114. Epub 2016 Sep 29.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing, Jiangsu, China. Electronic address:

This study attempts to identify receptor elements for Tembusu virus (TMUV) on DF-1 cells. Using co-immunoprecipitation and virus overlay protein binding assays, we identified a TMUV-binding protein of approximately 70-kDa on DF-1 cell membranes. Mass spectroscopy identified the protein to be heat shock protein (HSP) A9, which was reconfirmed by an anti-HSPA9 antibody. Indirect immunofluorescence demonstrated a significant degree of colocalization between HSPA9 and TMUV on cell surface. Additionally, an antibody against HSPA9 could inhibit TMUV infection in DF-1 cells in a dose-dependent manner. These results might suggest that HSPA9 is a putative receptor for TMUV.
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http://dx.doi.org/10.1016/j.virusres.2016.09.020DOI Listing
January 2017

Quantitative Proteomic Analysis of Duck Ovarian Follicles Infected with Duck Tembusu Virus by Label-Free LC-MS.

Front Microbiol 2016 31;7:463. Epub 2016 Mar 31.

Key Laboratory of Veterinary Biological Engineering and Technology, National Center for Engineering Research of Veterinary Bio-products, Institute of Veterinary Medicine, Ministry of Agriculture, Jiangsu Academy of Agricultural SciencesNanjing, China; Jiangsu Key Lab of Zoonosis, Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and ZoonosesYangzhou, China.

Duck Tembusu virus (DTMUV) is a newly emerging pathogenic flavivirus that has caused massive economic losses to the duck industry in China. DTMUV infection mainly results in significant decreases in egg production in egg-laying ducks within 1-2 weeks post infection. However, information on the comparative protein expression of host tissues in response to DTMUV infection is limited. In the present study, the cellular protein response to DTMUV infection in duck ovarian follicles was analyzed using nano-flow high-performance liquid chromatography-electrospray tandem mass spectrometry. Quantitative proteomic analysis revealed 131 differentially expressed proteins, among which 53 were up regulated and 78 were down regulated. The identified proteins were involved in the regulation of essential processes such as cellular structure and integrity, RNA processing, protein biosynthesis and modification, vesicle transport, signal transduction, and mitochondrial pathway. Some selected proteins that were found to be regulated in DTMUV-infected tissues were screened by quantitative real-time PCR to examine their regulation at the transcriptional level, western blot analysis was used to validate the changes of some selected proteins on translational level. To our knowledge, this study is the first to analyze the proteomic changes in duck ovarian follicles following DTMUV infection. The protein-related information obtained in this study may be useful to understand the host response to DTMUV infection and the inherent mechanism of DTMUV replication and pathogenicity.
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http://dx.doi.org/10.3389/fmicb.2016.00463DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4815560PMC
April 2016

Design and evaluation of a polytope construct with multiple B and T epitopes against Tembusu virus infection in ducks.

Res Vet Sci 2016 Feb 18;104:174-80. Epub 2015 Sep 18.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, National Center for Engineering Research of Veterinary Bio-products, Nanjing, Jiangsu Province, PR China; Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou 225009, PR China. Electronic address:

Tembusu virus (TMUV) is a newly emerging pathogenic flavivirus that is causing massive economic loss in Chinese poultry industry; until now, there is no effective vaccine or drug for its prevention. Epitope-based vaccination is a promising approach to achieve protective immunity and to avoid immunopathology. In present study, based on in silico epitope selection, we optimized and proposed a polytope DNA vaccine (pVAX1-rTEM) consisting B-cell and T cell epitopes from the TMUV envelope (E) protein. The immunogenicity and protective efficacy of constructed polytope DNA vaccine was assessed by in vitro and in vivo experiments. In in vitro assays, the expressed pVAX1-rTEM showed reactivity with Tembusu positive serum. Its protective efficacy against TMUV infection was evaluated in ducks. The results showed that pVAX1-rTEM was highly immunogenic and could elicit high titer neutralizing antibodies and cell-mediated immune responses. These results indicate that pVAX1-rTEM may be a promising candidate vaccine for prevention of TMUV infection.
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http://dx.doi.org/10.1016/j.rvsc.2015.09.011DOI Listing
February 2016

Domain I and II from newly emerging goose tembusu virus envelope protein functions as a dominant-negative inhibitor of virus infectivity.

Res Vet Sci 2015 Feb 18;98:121-6. Epub 2014 Nov 18.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing, Jiangsu Province, China. Electronic address:

Flavivirus envelope protein locates at the outermost surface of viral particle and mediates virus entry and fusion infection, and domains I and II of E protein play an important role in this process. In this study, we have expressed and purified goose tembusu virus (GTV) E protein domains I and II (DI/II) from E. coli, and tested conceptual approach that purified protein serves as anti-viral reagent. We found that DI/II inhibited GTV JS804 infection in BHK-21 cells in a dose-dependent manner, and this inhibition activity was achieved by binding to cell membrane specifically. Moreover, JS804 treated with DI/II specific anti-serum decreased its infectivity to BHK-21 cells. Taken together, this is first to show that the purified DI/II domain of tembusu virus expressed in E. coli was able to interfere with virus infection, which opens an avenue to develop novel anti-viral regents to prevent and eventually eradicate GTV infection.
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http://dx.doi.org/10.1016/j.rvsc.2014.11.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7172782PMC
February 2015

Complete genome sequence of goose tembusu virus, isolated from jiangnan white geese in jiangsu, china.

Genome Announc 2013 Mar 7;1(2):e0023612. Epub 2013 Mar 7.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing, Jiangsu Province, China.

Avian tembusu virus (TMUV), which was first identified in eastern China, is an emerging virus causing serious economic losses in the Chinese poultry industry. Here, we report the complete genome sequence of goose tembusu virus strain JS804, isolated from Jiangnan white geese with severe neurological signs. The genome of JS804 is 10,990 nucleotides (nt) in length and contains a single open reading frame encoding a putative polyprotein of 3,425 amino acids. Research of the whole sequence of tembusu virus will help us to understand further the molecular and evolutionary characteristics and pathogenesis of this virus.
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http://dx.doi.org/10.1128/genomeA.00236-12DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3593326PMC
March 2013

Identification and molecular characterization of a novel flavivirus isolated from geese in China.

Res Vet Sci 2013 Jun 25;94(3):774-80. Epub 2012 Dec 25.

Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, Nanjing, Jiangsu Province, PR China.

Since April 2010, a novel contagious disease in ducks and geese, with egg drop, feed uptake decline and neurological signs, caused by a newly emerged virus has spread around Eastern China. Dissection conducted on the dead geese demonstrated hemorrhage in brain, lung, liver, heart, ovary, and enlarged and necrotic spleen. A new virus, named Goose/Jiangsu/804/2010 (JS804) virus, was isolated in Jiangsu area from geese. Then the virus was re-isolated from the affected geese and replicated well in duck embryo fibroblasts and Vero cells, causing the cytopathic effect. The virus was identified as an enveloped positive stranded RNA virus with a size of approximately 40-60 nm in diameter. The full-length genome of this isolated virus was determined, showing that it is closely related to Tembusu virus (a mosquito-borne Ntaya group flavivirus) than other members of the Flaviviridae based on the data of phylogenetic analyses. Our systematic studies fulfill Koch's postulates precisely, and therefore, the causative agent of geese occurring in Eastern China is a new flavivirus. This is the first report that flavivirus infects not only egg-laying and breeder ducks but also geese. The findings extend our understanding of how the virus spreads and causes disease.
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http://dx.doi.org/10.1016/j.rvsc.2012.11.014DOI Listing
June 2013

Enantioselective epoxidation of non-functionalized alkenes using carbohydrate based salen-Mn(III) complexes.

Carbohydr Res 2007 Feb 3;342(2):254-8. Epub 2006 Dec 3.

School of Chemical Engineering, Hebei University of Technology, Tianjin 300130, PR China.

Three new salen ligands with carbohydrate moieties were prepared from a salicylaldehyde derivative obtained by reaction of 1,2:5,6-di-O-isopropylidene-alpha-D-glucofuranose with 3-tert-butyl-5-(chloro-methyl)-2-hydroxybenzaldehyde. These ligands were coordinated with Mn(III) to give three chiral salen-Mn(III) complexes. The complexes were characterized and employed in the asymmetric epoxidation of unfunctionalized alkenes. Catalytic results showed that although there are no chiral groups on the diimine bridge, these complexes had some enantioselectivity, which indicates the carbohydrate moiety has an asymmetric inducing effect in the epoxidation reaction.
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http://dx.doi.org/10.1016/j.carres.2006.11.026DOI Listing
February 2007