Publications by authors named "Weiye Chen"

29 Publications

  • Page 1 of 1

An Escherichia coli isolate from hospital sewage carries bla and bla.

Arch Microbiol 2021 Jun 15. Epub 2021 Jun 15.

College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, 210095, China.

Carbapenems, as the "last line of defense" against Gram-negative bacteria, are increasingly being challenged by drug-resistant bacteria, especially Enterobacteriaceae. In this study, a carbapenem-resistant Gram-negative bacterium, named AH001, was isolated from hospital sewage, and a modified Hodge test confirmed that this bacterium can produce carbapenemase. Further analysis revealed that this bacterium exhibits multidrug resistance against an additional seven antibiotics. Whole-genome sequencing and analysis showed that AH001 could not be classified by existing MLST, and its serotype could not be distinguished among O9, O89 or O168 according to O antigen prediction. More attention should be given to the role of environmental sources of Escherichia coli in the development and transfer of drug resistance in the hospital environment.
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http://dx.doi.org/10.1007/s00203-021-02431-2DOI Listing
June 2021

Prediction Models for Prognosis of Cervical Cancer: Systematic Review and Critical Appraisal.

Front Public Health 2021 7;9:654454. Epub 2021 May 7.

Department of Epidemiology and Biostatistics, School of Public Health, Peking University Health Science Center, Beijing, China.

This work aims to systematically identify, describe, and appraise all prognostic models for cervical cancer and provide a reference for clinical practice and future research. We systematically searched PubMed, EMBASE, and Cochrane library databases up to December 2020 and included studies developing, validating, or updating a prognostic model for cervical cancer. Two reviewers extracted information based on the CHecklist for critical Appraisal and data extraction for systematic Reviews of prediction Modeling Studies checklist and assessed the risk of bias using the Prediction model Risk Of Bias ASsessment Tool. Fifty-six eligible articles were identified, describing the development of 77 prognostic models and 27 external validation efforts. The 77 prognostic models focused on three types of cervical cancer patients at different stages, i.e., patients with early-stage cervical cancer ( = 29; 38%), patients with locally advanced cervical cancer ( = 27; 35%), and all-stage cervical cancer patients ( = 21; 27%). Among the 77 models, the most frequently used predictors were lymph node status ( = 57; 74%), the International Federation of Gynecology and Obstetrics stage ( = 42; 55%), histological types ( = 38; 49%), and tumor size ( = 37; 48%). The number of models that applied internal validation, presented a full equation, and assessed model calibration was 52 (68%), 16 (21%), and 45 (58%), respectively. Twenty-four models were externally validated, among which three were validated twice. None of the models were assessed with an overall low risk of bias. The Prediction Model of Failure in Locally Advanced Cervical Cancer model was externally validated twice, with acceptable performance, and seemed to be the most reliable. Methodological details including internal validation, sample size, and handling of missing data need to be emphasized on, and external validation is needed to facilitate the application and generalization of models for cervical cancer.
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http://dx.doi.org/10.3389/fpubh.2021.654454DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8137851PMC
May 2021

Intramolecular CH-migration-controlled cation reactions in the VUV photochemistry of 2-methyl-3-buten-2-ol investigated by synchrotron photoionization mass spectrometry and theoretical calculations.

Phys Chem Chem Phys 2021 May;23(17):10456-10467

National Synchrotron Radiation Laboratory, University of Science and Technology of China, Hefei, Anhui 230029, P. R. China.

2-Methyl-3-buten-2-ol (MBO232) is a biogenic volatile organic compound (BVOC), and has a large percentage of emission into the atmosphere. The vacuum ultraviolet (VUV) photochemistry of BVOCs is of great importance for atmospheric chemistry. Studies have been carried out on several BVOCs but have not extended to MBO232. In the present report, the photoionization and dissociation processes of MBO232 in the energy range of 8.0-15.0 eV have been studied by tunable VUV synchrotron radiation coupled with a time-of-flight mass spectrometer. By measuring the photoionization spectra, the adiabatic ionization energy (AIE) of MBO232 and the appearance energies (AEs) of the eight identified fragment ions (i.e., C4H7O+, C3H7O+, C5H9+, C3H6O+, CH3CO+, CH3O+, C4H5+, and C3H5+) were determined. High-level quantum chemistry calculations suggest that there are 3 direct channels and 5 indirect channels via transition states and intermediates accountable for these fragments. Among the reaction channels, the direct elimination of CH3 is the most dominant channel and produces the resonance-stabilized radical cation. Most interestingly, our results show that the CH3 selectively migrates towards the cation, which leads to the different indirect channels. The CH3 migration is a rare process in the dissociative photoionization of metal-free organic molecules. We explain the process by molecular orbital calculations and electron localization function analysis and explore the non-conventional dissociation channels via the CH3 roaming mechanism. We further perform kinetics analysis using RRKM theory for the channels of interest. The activation barrier, and rate constants are analyzed for the branching fractions of the products. These results provide important implications for the VUV photochemistry of BVOCs in the atmosphere.
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http://dx.doi.org/10.1039/d1cp00490eDOI Listing
May 2021

A seven-gene-deleted African swine fever virus is safe and effective as a live attenuated vaccine in pigs.

Sci China Life Sci 2020 May 1;63(5):623-634. Epub 2020 Mar 1.

State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin, 150001, China.

African swine fever (ASF) is a devastating infectious disease in swine that is severely threatening the global pig industry. An efficacious vaccine is urgently required. Here, we used the Chinese ASFV HLJ/18 as a backbone and generated a series of gene-deleted viruses. The virulence, immunogenicity, safety, and protective efficacy evaluation in specific-pathogen-free pigs, commercial pigs, and pregnant sows indicated that one virus, namely HLJ/18-7GD, which has seven genes deleted, is fully attenuated in pigs, cannot convert to the virulent strain, and provides complete protection of pigs against lethal ASFV challenge. Our study shows that HLJ/-18-7GD is a safe and effective vaccine against ASFV, and as such is expected to play an important role in controlling the spread of ASFV.
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http://dx.doi.org/10.1007/s11427-020-1657-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7223596PMC
May 2020

The Serine/Threonine Kinase AP2-Associated Kinase 1 Plays an Important Role in Rabies Virus Entry.

Viruses 2019 12 30;12(1). Epub 2019 Dec 30.

State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150069, China.

Rabies virus (RABV) invades the central nervous system and nearly always causes fatal disease in humans. RABV enters cells via clathrin-mediated endocytosis upon receptor binding. The detailed mechanism of this process and how it is regulated are not fully understood. Here, we carried out a high-through-put RNAi analysis and identified AP2-associated kinase 1 (AAK1), a serine/threonine kinase, as an important cellular component in regulating the entry of RABV. AAK1 knock-down greatly inhibits RABV infection of cells, and AAK1-induced phosphorylation of threonine 156 of the μ subunit of adaptor protein 2 (AP2M1) is found to be required for RABV entry. Inhibition of AAK1 kinase activity by sunitinib blocked AP2M1 phosphorylation, significantly inhibiting RABV infection and preventing RABV from entering early endosomes. In vivo studies revealed that sunitinib prolongs the survival of mice challenged with RABV street virus. Our findings indicate that AAK1 is a potential drug target for postexposure prophylaxis against rabies.
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http://dx.doi.org/10.3390/v12010045DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7019586PMC
December 2019

Replication and virulence in pigs of the first African swine fever virus isolated in China.

Emerg Microbes Infect 2019 ;8(1):438-447

a State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute , Chinese Academy of Agricultural Sciences , Harbin , People's Republic of China.

African swine fever (ASF) entered China in August 2018 and rapidly spread across the entire country, severely threatening the Chinese domestic pig population, which accounts for more than 50% of the pig population worldwide. In this study, an ASFV isolate, Pig/Heilongjiang/2018 (Pig/HLJ/18), was isolated in primary porcine alveolar macrophages (PAMs) from a pig sample from an ASF outbreak farm. The isolate was characterized by using the haemadsorption (HAD) test, Western blotting and immunofluorescence, and electronic microscopy. Phylogenetic analysis of the viral p72 gene revealed that Pig/HLJ/18 belongs to Genotype II. Infectious titres of virus propagated in primary PAMs and pig marrow macrophages were as high as 10 HAD/ml. Specific-pathogen-free pigs intramuscularly inoculated with different virus dosages at 10-10 HAD showed acute disease with fever and haemorrhagic signs. The incubation periods were 3-5 days for virus-inoculated pigs and 9 days for contact pigs. All virus-inoculated pigs died between 6-9 days post-inoculation (p.i.), and the contact pigs died between 13-14 days post-contact (p.c.). Viremia started on day 2 p.i. in inoculated pigs and on day 9 p.c. in contact pigs. Viral genomic DNA started to be detected from oral and rectal swab samples on 2-5 days p.i. in virus-inoculated pigs, and 6-10 days p.c. in contact pigs. These results indicate that Pig/HLJ/18 is highly virulent and transmissible in domestic pigs. Our study demonstrates the threat of ASFV and emphasizes the need to control and eradicate ASF in China.
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http://dx.doi.org/10.1080/22221751.2019.1590128DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6455124PMC
August 2019

Metabotropic glutamate receptor subtype 2 is a cellular receptor for rabies virus.

PLoS Pathog 2018 07 20;14(7):e1007189. Epub 2018 Jul 20.

State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin, P. R. China.

Rabies virus (RABV) invades the central nervous system and nearly always causes fatal disease in humans. How RABV interacts with host neuron membrane receptors to become internalized and cause rabid symptoms is not yet fully understood. Here, we identified a novel receptor of RABV, which RABV uses to infect neurons. We found that metabotropic glutamate receptor subtype 2 (mGluR2), a member of the G protein-coupled receptor family that is abundant in the central nervous system, directly interacts with RABV glycoprotein to mediate virus entry. RABV infection was drastically decreased after mGluR2 siRNA knock-down in cells. Antibodies to mGluR2 blocked RABV infection in cells in vitro. Moreover, mGluR2 ectodomain soluble protein neutralized the infectivity of RABV cell-adapted strains and a street strain in cells (in vitro) and in mice (in vivo). We further found that RABV and mGluR2 are internalized into cells and transported to early and late endosomes together. These results suggest that mGluR2 is a functional cellular entry receptor for RABV. Our findings may open a door to explore and understand the neuropathogenesis of rabies.
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http://dx.doi.org/10.1371/journal.ppat.1007189DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6070288PMC
July 2018

The Gene of Goatpox Virus Encodes an Inhibitor of NF-κB and Apoptosis and May Serve as an Improved Insertion Site To Generate Vectored Live Vaccine.

J Virol 2018 09 29;92(18). Epub 2018 Aug 29.

State Key Laboratory of Veterinary Biotechnology, Mission of Zoonosis and Exotic Diseases, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin, People's Republic of China

Goatpox virus (GTPV) is an important member of the genus of the Capripoxviruses have large and complex DNA genomes encoding many unknown proteins that may contribute to virulence. We identified that the open reading frame of GTPV is an early gene that encodes an ∼18-kDa protein that is nonessential for viral replication in cells. This protein functioned as an inhibitor of NF-κB activation and apoptosis and is similar to the N1L protein of vaccinia virus. In the natural host, sheep, deletion of the gene from the GTPV live vaccine strain AV41 resulted in less attenuation than that induced by deletion of the gene, a well-defined nonessential gene in the poxvirus genome. Using the gene as the insertion site, a recombinant AV41 strain expressing hemagglutinin of peste des petits ruminants virus (PPRV) was generated and elicited stronger neutralization antibody responses than those obtained using the traditional gene as the insertion site. These results suggest that the gene of GTPV encodes an immunomodulatory protein to suppress host innate immunity and may serve as an optimized insertion site to generate capripoxvirus-vectored live dual vaccines. Capripoxviruses are etiological agents of important diseases in sheep, goats, and cattle. There are rare reports about viral protein function related to capripoxviruses. In the present study, we found that the 135 protein of GTPV plays an important role in inhibition of innate immunity and apoptosis in host cells. Use of the gene as the insertion site to generate a vectored vaccine resulted in stronger adaptive immune responses than those obtained using the locus as the insertion site. As capripoxviruses are promising virus-vectored vaccines against many important diseases in small ruminants and cattle, the gene may serve as an improved insertion site to generate recombinant capripoxvirus-vectored live dual vaccines.
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http://dx.doi.org/10.1128/JVI.00190-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6146686PMC
September 2018

Fusing Benzo[c][1,2,5]oxadiazole Unit with Thiophene for Constructing Wide-bandgap High-performance IDT-based Polymer Solar Cell Donor Material.

Macromol Rapid Commun 2018 Apr 13;39(8):e1700782. Epub 2018 Feb 13.

CAS Key Laboratory of Bio-based Materials, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, 266101, People's Republic of China.

Benzo[c][1,2,5]oxadiazole (BO) moiety is a strong electron-withdrawing unit compared to benzo[c][1,2,5]thiadiazole (BT). It is usually introduced as an acceptor to construct narrow band-gap donor-acceptor (D-A) materials. Herein, the π-extended conjugated moiety dithieno[3',2':3,4″;2,3″:5,6]benzo[1,2-c][1,2,5]oxadiazole (BOT) was adopted as the acceptor moiety to design D-A polymers. Considering the more extended π-conjugated molecular system of BOT compared to the BO unit, a narrower optical band-gap is expected for BOT-based IDT polymer (PIDT-BOT). Unexpectedly, the UV-vis absorption spectra of PIDT-BOT films display a great hypochromatic shift of about 60 nm compared to a BO-based analog (PIDT-BO). The optical band-gaps of the materials are broadened from 1.63 eV (PIDT-BO) to 2.00 eV (PIDT-BOT) accordingly. Although the range of external quantum efficiency (EQE) of PIDT-BOT-based polymer solar cell (PSC) devices is not as wide as for PIDT-BO-based devices, the EQE response intensities of the PIDT-BOT based device are evidently high. As a result, PSC devices based on PIDT-BOT reveal the best power conversion efficiency at 6.08%.
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http://dx.doi.org/10.1002/marc.201700782DOI Listing
April 2018

Establishing a safe, rapid, convenient and low-cost antiviral assay of interferon bioactivity based on recombinant VSV expressing GFP.

J Virol Methods 2018 02 20;252:1-7. Epub 2017 Aug 20.

State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences (CAAS), 678 Haping Road, Harbin 150001, People's Republic of China. Electronic address:

The methods of the quantitative assay of the antiviral activity of interferons (IFNs) (type I, II or III) are very important during carrying out of the research of them, since they were found. Here a recombinant vesicular stomatitis virus expressing green fluorescent protein (GFP) (VSV/GFP) and MDBK cells were used to develop an antiviral assay (AVA) for IFNs. This method was carried out on a 96-well cell culture plate, and the half reduction of virus replication was quantified by assaying GFP. To quantify GFP, cell lysis buffer was directly added to the wells infected with VSV/GFP to lyse cells, the VSV/GFP was then inactivated, and relative fluorescence unit (RFU) of GFP was measured and used to calculate the antiviral activity. This method needed only one step instead of three steps in the staining method with naphthol blue black, medium with phenol red can be used, and it had good reproducibility. The GFP-containing samples could be stored at 4°C in a wet box for at least 1 week without affecting the assay results. In addition, the results obtained with this method were similar to those obtained with the staining method. In conclusion, a safe, rapid, convenient and low-cost AVA of IFN based on recombinant VSV/GFP was established.
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http://dx.doi.org/10.1016/j.jviromet.2017.08.007DOI Listing
February 2018

IFN-lambda preferably inhibits PEDV infection of porcine intestinal epithelial cells compared with IFN-alpha.

Antiviral Res 2017 04 19;140:76-82. Epub 2017 Jan 19.

State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin, China. Electronic address:

In contrast to type I interferons that target various types of cells and organs, interferon lambda (IFN-L) primarily acts on mucosal epithelial cells and exhibits robust antiviral activity within the mucosal surface. Porcine epidemic diarrhea virus (PEDV), which causes high morbidity and mortality in piglets, is an enteropathogenic coronavirus with economic importance. Here, we demonstrated that both recombinant porcine IFN-L1 (rpIFN-L1) and rpIFN-L3 have powerful antiviral activity against PEDV infection of both Vero E6 cells and the intestinal porcine epithelial cell line J2 (IPEC-J2). Both forms of rpIFN-L inhibited two genotypes of PEDV (strain CV777 of genotype 1 and strain LNCT2 of genotype 2). rpIFN-L1 primarily controlled viral infection in the early stage and had less antiviral activity in IPEC-J2 than in rpIFN-L3 cells infected with PEDV. In addition, rpIFN-L1 exhibited greater antiviral activity against PEDV infection of IPEC-J2 cells than that of porcine IFN-alpha. Consistent with this finding, rpIFN-L1 triggered higher levels of certain antiviral IFN-stimulated genes (ISGs) (ISG15, OASL, and MxA) in IPEC-J2 cells than porcine IFN-alpha. Although IPEC-J2 cells responded to both IFN-alpha and lambda, transcriptional profiling of ISGs (specifically ISG15, OASL, MxA, and IFITMs) differed when induced by either IFN-alpha or rpIFN-L. Therefore, our data provide the experimental evidence that porcine IFN-L suppresses PEDV infection of IPEC-J2 cells, which may offer a promising therapeutic for combating PED in piglets.
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http://dx.doi.org/10.1016/j.antiviral.2017.01.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7113730PMC
April 2017

Development of an Immunoperoxidase Monolayer Assay for the Detection of Antibodies against Peste des Petits Ruminants Virus Based on BHK-21 Cell Line Stably Expressing the Goat Signaling Lymphocyte Activation Molecule.

PLoS One 2016 21;11(10):e0165088. Epub 2016 Oct 21.

National Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences (CAAS), Harbin, China.

From 2013 to 2015, peste des petits ruminants (PPR) broke out in more than half of the provinces of China; thus, the application and development of diagnostic methods are very important for the control of PPR. Here, an immunoperoxidase monolayer assay (IPMA) was developed to detect antibodies against PPR. However, during IPMA development, we found that Vero cells were not the appropriate choice because staining results were not easily observed. Therefore, we first established a baby hamster kidney-goat signaling lymphocyte activation molecule (BHK-SLAM) cell line that could stably express goat SLAM for at least 20 generations. Compared with Vero cells, the PPR-mediated cytopathic effect occurred earlier in BHK-SLAM cells, and large syncytia appeared after virus infection. Based on this cell line and recombinant PPR virus expressing the green fluorescent protein (GFP) (rPPRV-GFP), an IPMA for PPR diagnosis was developed. One hundred and ninety-eight PPR serum samples from goats or sheep were tested by the IPMA and virus neutralization test (VNT). Compared with the VNT, the sensitivity and specificity of the IPMA were 91% and 100%, respectively, and the coincidence rate of the two methods was 95.5%. The IPMA assay could be completed in 4 h, compared with more than 6 d for the VNT using rPPRV-GFP, and it is easily performed, as the staining results can be observed under a microscope. Additionally, unlike the VNT, the IPMA does not require antigen purification, which will reduce its cost. In conclusion, the established IPMA will be an alternative method that replaces the VNT for detecting antibodies against PPRV in the field.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0165088PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5074545PMC
June 2017

Characterization of a recombinant Newcastle disease virus expressing the glycoprotein of bovine ephemeral fever virus.

Arch Virol 2017 Feb 18;162(2):359-367. Epub 2016 Oct 18.

State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, 427 Maduan Street, Harbin, 150001, People's Republic of China.

Bovine ephemeral fever (BEF) is caused by the arthropod-borne bovine ephemeral fever virus (BEFV), which is a member of the family Rhabdoviridae and the genus Ephemerovirus. BEFV causes an acute febrile infection in cattle and water buffalo. In this study, a recombinant Newcastle disease virus (NDV) expressing the glycoprotein (G) of BEFV (rL-BEFV-G) was constructed, and its biological characteristics in vitro and in vivo, pathogenicity, and immune response in mice and cattle were evaluated. BEFV G enabled NDV to spread from cell to cell. rL-BEFV-G remained nonvirulent in poultry and mice compared with vector LaSota virus. rL-BEFV-G triggered a high titer of neutralizing antibodies against BEFV in mice and cattle. These results suggest that rL-BEFV-G might be a suitable candidate vaccine against BEF.
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http://dx.doi.org/10.1007/s00705-016-3078-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5306239PMC
February 2017

Extracellular expression and antiviral activity of a bovine interferon-alpha through codon optimization in Pichia pastoris.

Microbiol Res 2016 Oct 19;191:12-8. Epub 2016 May 19.

State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, PR China. Electronic address:

Interferons (IFNs) are the primary line of defense against infectious agents. In particular, IFN-α is an important antiviral cytokine and has a wide range of immune-modulating functions. Porcine and human IFN-α have been successfully prepared and play important roles in the prevention and therapy of viral diseases. To date, there has been limited applied research on bovine IFN-α. To achieve high-level expression of recombinant bovine IFN-α (bIFN-α) in Pichia pastoris for large-scale application, the bIFN-α gene was optimized and synthesized on the basis of codon bias of P. pastoris. Optimized bIFN-α (opti-bIFN-α) was successfully expressed in P. pastoris and directly secreted into the culture supernatant. The amount of extracellular soluble opti-bIFN-α was observed to be 200μg/mL in a shake flask. Expression efficiency of opti-bIFN-α was found to be about three times that of wild-type bIFN-α when the expression yield was compared at the same copies of the targeted gene. In addition, both the original cultural supernatant and purified opti-bIFN-α showed strong antiviral activity in MDBK cells (2×10(6)AU/mL and 1×10(7)AU/mg, respectively) and IBRS-2 cells (3×10(5)AU/mL and 1.5×10(6)AU/mg, respectively) against a recombinant vesicular stomatitis virus expressing the green fluorescence protein. In this study, we demonstrated high-level extracellular expression of opti-bIFN-α by P. pastoris. To the best of our knowledge, the opti-bIFN-α yield observed in this study is the highest to be reported to date. Our results demonstrated that the extracellular opti-bIFN-α with strong antiviral activity could be easily prepared and purified at a low cost and that it may be a potential biological therapeutic drug against bovine viral infections.
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http://dx.doi.org/10.1016/j.micres.2016.05.009DOI Listing
October 2016

Protective efficacy of a recombinant Newcastle disease virus expressing glycoprotein of vesicular stomatitis virus in mice.

Virol J 2016 Feb 24;13:31. Epub 2016 Feb 24.

State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, 427 Maduan Street, Harbin, 150001, People's Republic of China.

Background: Vesicular stomatitis virus (VSV) causes severe losses to the animal husbandry industry. In this study, a recombinant Newcastle disease virus (NDV) expressing the glycoprotein (G) of VSV (rL-VSV-G) was constructed and its pathogenicity and immune protective efficacy in mouse were evaluated.

Results: In pathogenicity evaluation test, the analysis of the viral distribution in mouse organs and body weight change showed that rL-VSV-G was safe in mice. In immune protection assay, the recombinant rL-VSV-G triggered a high titer of neutralizing antibodies against VSV. After challenge, the wild-type (wt) VSV viral load in mouse organs was lower in rL-VSV-G group than that in rLaSota groups. wt VSV was not detected in the blood, liver, or kidneys of mice, whereas it was found in these tissues in control groups. The mice body weight had no significant change after challenge in the rL-VSV-G group. Additionally, suckling mice produced from female mice immunized with rL-VSV-G were partially protected from wt VSV challenge.

Conclusions: These results demonstrated that rL-VSV-G may be a suitable candidate vaccine against vesicular stomatitis (VS).
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http://dx.doi.org/10.1186/s12985-016-0481-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4765107PMC
February 2016

Genetically modified rabies virus ERA strain is safe and induces long-lasting protective immune response in dogs after oral vaccination.

Antiviral Res 2015 Sep 18;121:9-15. Epub 2015 Jun 18.

Key Laboratory of Veterinary Public Health of Ministry of Agriculture, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China. Electronic address:

Oral immunization in free-roaming dogs is one of the most practical approaches to prevent rabies for developing countries. The safe, efficient and long-lasting protective oral rabies vaccine for dogs is highly sought. In this study, rabies virus (RABV) Evelyn-Rokitnicki-Abelseth (ERA) strain wild-type (rERA) and a genetically modified type (rERAG333E) containing a mutation from arginine to glutamic acid at residue 333 of glycoprotein (G333E) were generated by reverse genetic. The recombinant virus rERAG333E retained growth properties of similar to the parent strain rERA in BHK-21 cell culture. The G333E mutation showed genetic stability during passage into neuroblastoma cells and in the brains of suckling mice and was significantly reduced the virulence of rERA in mice. rERAG333E was immunogenic in dogs by intramuscular inoculation. Mice orally vaccinated with rERAG333E induced strong and one year longer virus neutralizing antibodies (VNA) to RABV, and were completely protected from challenge with lethal street virus at 12months after immunization. Dogs received oral vaccination with rERAG333E induced strong protective RABV VNA response, which lasted for over 3years, and moderate saliva RABV-specific IgA. Moreover, sizeable booster responses to RABV VNA were induced by a second oral dose 1year after the first dose. These results demonstrated that the genetically modified ERA vaccine strain has the potential to serve as a safe and efficient oral live vaccine against rabies in dogs.
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http://dx.doi.org/10.1016/j.antiviral.2015.06.011DOI Listing
September 2015

Establishment of MDCK Stable Cell Lines Expressing TMPRSS2 and MSPL and Their Applications in Propagating Influenza Vaccine Viruses in Absence of Exogenous Trypsin.

Biotechnol Res Int 2015 30;2015:402628. Epub 2015 Mar 30.

State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, 427 Maduan Street, Harbin 150001, China.

We established two Madin-Darby canine kidney (MDCK) cell lines stably expressing human airway transmembrane protease: transmembrane protease, serine 2 (TMPRSS2) and mosaic serine protease large form (MSPL) which support multicycle growth of two H5 highly pathogenic avian influenza viruses (HPAIV) recombinant vaccines (Re-5 and Re-6) and an H9 avian influenza virus (AIV) recombinant vaccine (Re-9) in the absence of trypsin. Data showed that the cell lines stably expressed TMPRSS2 and MSPL after 20 serial passages. Both MDCK-TMPRSS2 and MDCK-MSPL could proteolytically cleave the HA of Re-5, Re-6, and Re-9 and supported high-titer growth of the vaccine without exogenous trypsin. Re-5, Re-6, and Re-9 efficiently infected and replicated within MDCK-TMPRSS2 and MDCK-MSPL cells and viral titer were comparable to the virus grown in MDCK cells with TPCK-trypsin. Thus, our results indicate a potential application for these cell lines in cell-based influenza vaccine production and may serve as a useful tool for HA proteolytic cleavage-related studies.
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http://dx.doi.org/10.1155/2015/402628DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4396729PMC
April 2015

Induction of protective immune response against both PPRV and FMDV by a novel recombinant PPRV expressing FMDV VP1.

Vet Res 2014 Jun 4;45:62. Epub 2014 Jun 4.

Department of Microbiology and Immunology, College of Veterinary Medicine, China Agricultural University, Beijing 100094, China.

Peste des petits ruminants (PPR) and foot-and-mouth disease (FMD) are both highly contagious diseases of small domestic and wild ruminants caused by the PPR virus (PPRV) and the FMD virus (FMDV). In this study, a recombinant PPRV expressing the FMDV VP1 gene (rPPRV/VP1) was generated and FMDV VP1 expression did not impair replication of the recombinant virus in vitro and immunogenicity in inducing neutralizing antibody against PPR in goats. Vaccination with one dose of rPPRV/VP1 induced FMDV neutralizing antibody in goats and protected them from challenge with virulent FMDV. Our results suggest that the recombinant PPRV expressing the FMDV VP1 protein is a potential dual live vectored vaccine against PPRV and FMDV.
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http://dx.doi.org/10.1186/1297-9716-45-62DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4059095PMC
June 2014

Recombinant lentogenic Newcastle disease virus expressing Ebola virus GP infects cells independently of exogenous trypsin and uses macropinocytosis as the major pathway for cell entry.

Virol J 2013 Nov 9;10:331. Epub 2013 Nov 9.

State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, 427 Maduan Street, Harbin 150001, People's Republic of China.

Background: Using reverse genetics, we generated a recombinant low-pathogenic LaSota strain Newcastle disease virus (NDV) expressing the glycoprotein (GP) of Ebola virus (EBOV), designated rLa-EBOVGP, and evaluated its biological characteristic in vivo and in vitro.

Results: The introduction and expression of the EBOV GP gene did not increase the virulence of the NDV vector in poultry or mice. EBOV GP was incorporated into the particle of the vector virus and the recombinant virus rLa-EBOVGP infected cells and spread within them independently of exogenous trypsin. rLa-EBOVGP is more resistant to NDV antiserum than the vector NDV and is moderately sensitive to EBOV GP antiserum. More importantly, infection with rLa-EBOVGP was markedly inhibited by IPA3, indicating that rLa-EBOVGP uses macropinocytosis as the major internalization pathway for cell entry.

Conclusions: The results demonstrate that EBOV GP in recombinant NDV particles functions independently to mediate the viral infection of the host cells and alters the cell-entry pathway.
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http://dx.doi.org/10.1186/1743-422X-10-331DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3826533PMC
November 2013

Newcastle disease virus-vectored Nipah encephalitis vaccines induce B and T cell responses in mice and long-lasting neutralizing antibodies in pigs.

Virology 2012 Oct 21;432(2):327-35. Epub 2012 Jun 21.

State Key Laboratory of Veterinary Biotechnology and Animal Influenza Laboratory of the Ministry of Agriculture, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, People's Republic of China.

Nipah virus (NiV), a member of the Paramyxoviridae family, causes deadly encephalitis in humans and huge economic losses to the pig industry. Here, we generated recombinant avirulent Newcastle disease virus (NDV) LaSota strains expressing the NiV G and F proteins respectively (designated as rLa-NiVG and rLa-NiVF), and evaluated their immunogenicity in mice and pigs. Both rLa-NiVG and rLa-NiVF displayed growth properties similar to those of LaSota virus in chicken eggs. Co-infection of rLa-NiVG and rLa-NiVF caused marked syncytia formation, while intracerebral co-inoculation of these viruses in mice showed they were safe in at least one mammalian species. Animal immunization studies showed rLa-NiVG and rLa-NiVF induced NiV neutralizing antibody responses in mice and pigs, and F protein-specific CD8+ T cell responses in mice. Most importantly, rLa-NiVG and rLa-NiVF administered alone or together, induced a long-lasting neutralizing antibody response in pigs. Recombinant rLa-NiVG/F thus appear to be promising NiV vaccine candidates for pigs and potentially humans.
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http://dx.doi.org/10.1016/j.virol.2012.06.001DOI Listing
October 2012

Rescue of recombinant peste des petits ruminants virus: creation of a GFP-expressing virus and application in rapid virus neutralization test.

Vet Res 2012 Jun 2;43:48. Epub 2012 Jun 2.

College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, 210095, People's Republic of China.

Peste des petits ruminants virus (PPRV) causes high mortality in goats and sheep and the disease has shown a greatly increased geographic distribution over the last 15 years. It is responsible for serious socioeconomic problems in some of the poorest developing countries. The ability to create recombinant PPRV would provide a useful tool for investigating the biology of the virus and the pathology of disease, as well as for developing new vaccines and diagnostic methods. Here we report the first successful rescue of recombinant PPRV from a full-length cDNA clone of the virus genome. Successful recovery of PPRV was achieved by using a RNA polymerase II promoter to drive transcription of the full-length virus antigenome. We have used this technique to construct a virus expressing a tracer protein (green fluorescent protein, GFP). The recombinant virus replicated as well as the parental virus and could stably express GFP during at least 10 passages. The newly established reverse genetics system for PPRV provides a novel method for constructing a vaccine using PPRV as a vector, and will also prove valuable for fundamental research on the biology of the virus. We found that our recombinant virus allowed more rapid and higher throughput assessment of PPRV neutralization antibody titer via the virus neutralization test (VNT) compared with the traditional method.
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http://dx.doi.org/10.1186/1297-9716-43-48DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3412694PMC
June 2012

Establishment of a stable CHO cell line with high level expression of recombinant porcine IFN-β.

Cytokine 2011 Jun 1;54(3):324-9. Epub 2011 Apr 1.

National Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences (CAAS), 427 Maduan Street, Harbin 150001, People's Republic of China.

A CHO cell clone (CHO-PoIFN-β) with stable porcine IFN-β expression under control of CMV promoter was selected under G418 pressure. In a 25cm(2) cell culture flask (5 ml culture medium), the cumulative protein yield of recombinant PoIFN-β reached 2.3×10(6) IU/ml. This cells clone maintained stable expression for at least 20 generations even in the absence of G418 selection pressure. The expressed recombinant PoIFN-β could induce the expression of porcine Mx protein in PK15 cells, and activate the chicken Mx promoter-controlled luciferase reporter gene expression, confirming that the recombinant PoIFN-β has the biological activity of natural porcine type-I interferon. In addition, the recombinant PoIFN-β fully protected PK15 cells against 1000 TCID(50) of porcine transmissible gastroenteritis virus and pseudo-rabies virus infection, demonstrating its high potential in therapeutic applications. This is the first report of establishing a mammalian cell line with stable expression of porcine IFN-β.
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http://dx.doi.org/10.1016/j.cyto.2010.12.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7128424PMC
June 2011

A goat poxvirus-vectored peste-des-petits-ruminants vaccine induces long-lasting neutralization antibody to high levels in goats and sheep.

Vaccine 2010 Jul 14;28(30):4742-50. Epub 2010 May 14.

Key Laboratory of Veterinary Public Health of Ministry of Agriculture and State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150001, PR China.

Recombinant capripoxvirus (CPV) is a promising candidate differentiating infected from vaccinated animals (DIVA) vaccine against peste-des-petits-ruminants (PPR). In order for recombinant CPV to be successfully used in the field, there should exist dependable indicators for quality control of vaccine products, surveillance and vaccination evaluation. Viral neutralization antibody (VNA) is correlated to protection against PPR and is a technically feasible indicator for this purpose. The immunogenicity of this vectored vaccine in goats and sheep, however, has not been fully evaluated. In this study, we generated two recombinant CPV viruses, rCPV-PPRVH and rCPV-PPRVF, that express PPR virus (PPRV) glycoproteins H and F, respectively. Vaccination studies with different dosages of recombinant viruses showed that rCPV-PPRVH was a more potent inducer of PPRV VNA than rCPV-PPRVF. One dose of rCPV-PPRVH was enough to seroconvert 80% of immunized sheep. A second dose induced significantly higher PPRV VNA titers. There was no significant difference in PPRV VNA responses between goats and sheep. Subcutaneous inoculation also induced a significant PPRV VNA response. PPRV VNA could be detected for over 6 months in more than 80% of vaccinated goats and sheep. Boost vaccination at 6-month intervals induced significant re-boost efficacy of PPRV VNA in goats and sheep. More over, two doses of rCPV-PPRVH could completely overcome the interference caused by pre-existing immunity to the CPV vaccine backbone in animals. Vaccination with rCPV-PPRVH also protected goats from virulent CPV challenge. Our results demonstrate that VNA can serve as a dependent indicator for effective vaccination and immune protection of animals in the field. The recombinant CPV vaccine used in our studies could be a practical and useful candidate DIVA vaccine in countries where PPR newly emerges or where stamp-out plans are yet to be implemented.
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http://dx.doi.org/10.1016/j.vaccine.2010.04.102DOI Listing
July 2010

[Recombinant goat pox virus expressing PPRV H protein].

Sheng Wu Gong Cheng Xue Bao 2009 Apr;25(4):496-502

State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150001, China.

The purpose of the study is to construct recombinant goat pox virus (GPV) expressing Peste des petits ruminants virus (PPRV) H protein, and to evaluate the immunization effect. Recombinant GPV containing PPRV H gene (rGPV-PPRV-H) was selected and purified by gpt and eGFP utilizing plaque purification, and the final selected recombinant GPV was proved to be purified by PCR. Immunofluorescence and Western blotting showed that the recombinant virus could express H protein of PPRV while infecting lamb testis cells. Six goats were immunized with 2 x 10(6) PFU rGPV-PPRV-H through intradermal injection, and were immunized for the second time at 28 days with the same dose recombinant virus after first immunization. Serum was collected after immunization, and was analyzed for the neutralization antibodies. 21 days after first immunization, the neutralization antibodies of GPV were 40, 80, > or = 80, > or = 80, 40, > or = 80 in turn, and neutralization antibodies of PPRV were 80, 80, 80, 80, 40, 40, 10 in turn; 14 days after second immunization, the neutralization antibodies of GPV were all > or = 80, and the neutralization antibodies of PPRV were > 80, 80, > 80, 80, 80 and 40 in turn. This study established a foundation for the industrialization of the PPRV recombinant GPV vaccine.
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April 2009

Expression of biologically active recombinant equine interferon-gamma in Escherichia coli.

Comp Immunol Microbiol Infect Dis 2010 Jul 10;33(4):333-42. Epub 2009 Mar 10.

Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, PR China.

Interferon gamma (IFN-gamma) is a pleiotropic cytokine that is recognized as an important modulator of the immune response. To date, there is no report that prokaryocyte-derived recombinant equine IFN-gamma has antiviral activity. In this report, the gene coding equine IFN-gamma (EIFN-gamma) mature protein was cloned into pET-28a (+) and the recombinant EIFN-gamma was expressed in Escherichia coli (E. coli). The antiviral activity of expressed recombinant EIFN-gamma was evaluated by using a recombinant Vesicular Stomatitis Virus expressing green fluorescence protein (rVSV-GFP) system in the equine fetal kidney-78 cell line (EFK-78). The GFP expression in the EFK-78 cells dramatically decreased in the cells treated with EIFN-gamma in a dose-dependent manner, comparing with the mock-treated cells. The titer of antiviral activity was 1 x 10(3)AU/ml. These results demonstrated that the EIFN-gamma expressed in this study had good biological activity. Pure forms and sufficient quantities of biologically active IFN-gamma could facilitate the study of its activities in modulating immune responses both in vivo and in vitro.
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http://dx.doi.org/10.1016/j.cimid.2008.12.004DOI Listing
July 2010

Variance of DDAH/PRMT/ADMA pathway in atrial fibrillation dogs.

Biochem Biophys Res Commun 2008 Dec 23;377(3):884-8. Epub 2008 Oct 23.

Department of Cardiology, The First Affiliated Hospital of Harbin Medical University, 23 Youzheng Street, Nangang District, Harbin 150001, PR China.

Atrial fibrillation (AF) may cause thrombus formation in the left atrial appendage (LAA). Thrombus formation is associated with LAA endocardial dysfunction. Because asymmetrical dimethylarginine (ADMA) can cause endothelial dysfunction by decreasing nitric oxide (NO) formation, we investigated plasma ADMA and nitrite/nitrate (NO(X)) levels and myocardial dimethylarginine dimethylaminohydrolase-2 (DDAH-2), protein arginine methyltransferase-1 (PRMT-1), and endothelial NO synthase (eNOS) protein contents from AF dogs. The results displayed that plasma ADMA level significantly increased, and plasma NO(X) concentration significantly decreased. Compared with normal heart, DDAH-2 expression was unchanged in the fibrillating atria. However, the DDAH activity was significantly decreased in the fibrillating atria. PRMT-1 expression significantly increased in the LAA and in the left atrium (LA). ENOS expression significantly decreased in the LA. ENOS and PRMT-1 expressions were unchanged in the right atria. Our results suggested that the DDAH-PRMT-ADMA system maybe play a pivotal role in regulating endothelial function in AF.
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http://dx.doi.org/10.1016/j.bbrc.2008.10.080DOI Listing
December 2008

[Antiviral activity determination of recombinant equine interferon-gamma and identification inhibited antiviral activity of monoclonal antibodies].

Sheng Wu Gong Cheng Xue Bao 2008 Jul;24(7):1258-62

National Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, CAAS, Harbin 150001, China.

Equine interferon-gamma (eIFN-gamma) expressed both in E. coli and baculovirus were evaluated for antiviral activity against recombinant Vesicular Stomatits Virus expressing green fluorescence protein (rVSV-GFP) in EFK-78 cells. The assays were conducted in 96-well plate. Virus infectivity was measured by quantifying GFP-positive cells, instead of quantifying the CPE reduction. Prior to infection of EFK-78 cells with rVSV-GFP, the cells were incubated with eIFN-gamma. The GFP expression in the EFK-78 cells dramatically decreased in the cells treated with eIFN-gamma in a dose-dependent manner, comparing with the mock-treated cells. The titers of antiviral activity were 1 x 10(3) AU/mL and 1 x 10(5) AU/mL of eIFN-gamma expressed from E. coli and baculovirus, respectively. The antiviral activities of the recombinant eIFN-gamma were highly efficient and specific, as it was blocked by mAbs against eIFN-gamma.
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July 2008

Modified BIGH3 with an RGDRGD motif promotes human corneal epithelial cell adhesion and migration in vitro.

Curr Eye Res 2008 Mar;33(3):215-23

Eye Hospital, The First Clinical Hospital, Harbin Medical University, Harbin, China.

Purpose: BIGH3 protein plays an important role in mediating human corneal epithelial (HCE) cell adhesion and migration. The aim of this study was to investigate the effects of native and modified BIGH3 protein containing an Arg-Gly-Asp-Arg-Gly-Asp (RGDRGD) motif on the adhesion and migration of HCE cells.

Methods: A modified human BIGH3 gene containing an RGDRGD motif was obtained by rapid site-directed mutagenesis. Recombinant human native and modified BIGH3 proteins were then obtained and purified. The effects of the native and the modified version on the adhesion and migration of HCE cells were examined in the presence or absence of anti-alpha3beta1 antibody or anti-BIGH3 antibody or RGD peptide in vitro.

Results: Recombinant native and modified BIGH3 proteins were successfully obtained and significantly promoted the adhesion and migration of human HCE cells in vitro, and the construct with the RGDRGD motif was more effective. The enhanced adhesion and migration were blocked by anti-alpha3beta1antibody or anti-BIGH3 antibody or RGD peptide.

Conclusion: BIGH3 promotes HCE cell adhesion and migration; modified RGDRGD-BIGH3 was more effective than native BIGH, and this is mediated by the Arg-Gly-Asp (RGD) motif and alpha3beta1integrin in a RGD-dependent manner.
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http://dx.doi.org/10.1080/02713680801911218DOI Listing
March 2008

A naturally occurring deletion in its NS gene contributes to the attenuation of an H5N1 swine influenza virus in chickens.

J Virol 2008 Jan 17;82(1):220-8. Epub 2007 Oct 17.

Harbin Veterinary Research Institute, 427 Maduan Street, Harbin 150001, People's Republic of China.

In 2001 and 2003, we isolated two H5N1 viruses, A/swine/Fujian/1/01 (SW/FJ/01) and A/swine/Fujian/1/03 (SW/FJ/03), from pigs in Fujian Province, southern China. Genetically, these two viruses are similar, although the NS gene of the SW/FJ/03 virus has a 15-nucleotide deletion at coding positions 612 to 626. The SW/FJ/01 virus is highly lethal for chickens, whereas the SW/FJ/03 virus is nonpathogenic for chickens when administrated intravenously or intranasally. To understand the molecular basis for the difference in virulence, we used reverse genetics to create a series of single-gene recombinants of both viruses. We found that a recombinant virus containing the mutated NS gene from the SW/FJ/03 virus in the SW/FJ/01 virus background was completely attenuated in chickens. We also found that viruses expressing the mutant NS1 protein of SW/FJ/03 did not antagonize the induction of interferon (IFN) protein. Conversely, only the recombinant virus containing the wild-type SW/FJ/01 NS gene in the SW/FJ/03 background was lethal in chickens and antagonized IFN protein levels. Further, we proved that the NS1 genes of the two viruses differ in their stabilities in the host cells and in their abilities to interact with the chicken cleavage and polyadenylation specificity factor. These results indicate that the deletion of amino acids 191 to 195 of the NS1 protein is critical for the attenuation of the SW/FJ/03 virus in chickens and that this deletion affects the ability of the virus to antagonize IFN induction in host cells.
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http://dx.doi.org/10.1128/JVI.00978-07DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2224367PMC
January 2008
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