1.Construction of an infectious cDNA clone derived from foot-and-mouth disease virus O/QYYS/s/06.
Shousheng LU ; Qizu ZHAO ; Xiangtao LIU ; Yanwei SUN ; Tao REN ; Guihong ZHANG ; Wenbao QI ; Yunfeng ZHA ; Lingchen KONG ; Han ZHANG ; Huiying FAN ; Ming LIAO
Chinese Journal of Biotechnology 2009;25(7):982-986
After sequencing, we amplified and cloned foot-and-mouth disease virus (FMDV) O/QYYS/s/06 whole genome by three fragments. These three fragments were cloned into vector P43 one by one to construct recombinant plasmid P43C, which carried the full-length cDNA of FMDV O/QYYS/s/06. Then, plasmid P43C and plasmid T7 expressing T7 RNA polymerase were co-transfected into BHK-21 cells. After 48 h, we harvested the culture broth from transfected BHK-21 cells and inoculated into 2-3 day-old sucking mice. After four generation passage, the virus harvested from sucking mice was confirmed to be type O FMDV by the indirect hemagglutination test, sucking mice's neutralization test and sequencing. The results showed that we have successfully constructed the full-length cDNA clone of FMDV O/QYYS/s/06 strain.
Animals
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Animals, Newborn
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Cloning, Molecular
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DNA, Complementary
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genetics
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DNA, Viral
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biosynthesis
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genetics
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Foot-and-Mouth Disease
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virology
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Foot-and-Mouth Disease Virus
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classification
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genetics
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pathogenicity
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Mice
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Transcription, Genetic
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Transfection
2.Establishment of colloidal gold-immunochromatography assay strip for detection of type Asia1 foot-and-mouth disease virus.
Tong LIN ; Junjun SHAO ; Guozheng CONG ; Junzheng DU ; Shandian GAO ; Huiyun CHANG ; Qingge XIE
Chinese Journal of Biotechnology 2009;25(5):767-772
To establish a sensitive, rapid and simple gold immunochromatography assay (GICA) for detecting Asia1 type of foot-and-mouth disease virus (FMDV) from the field samples. The purified anti-FMDV type Asia1 monoclonal antibody labeled with colloidal gold and the goat anti-Guinea pig IgG were wrapped onto nitrocellulose membrane as the test line (T line) and the control line (C line), respectively. The strip was then further optimized. A total of 87 field samples were detected. The results indicated a correct rate of 98.8% for detecting FMDV Asia1 type. No cross reaction was found with swine vesicular disease (SVD) and FMDV O, A and C type antigen. The sensitivity of the strip can reach to 10(-4) (TCID50 6.25). It had the same results for positive and negative specimens tested in three times. This strip could be stored at 4 degrees C for three months. In this study, the established gold immunochromatographic strip test kit is simple, rapid, sensitive and specific for detecting FMDV type Asial, and is potentially useful for the for pen-side diagnosis.
Animals
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Antibodies, Monoclonal
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Chromatography
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methods
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Foot-and-Mouth Disease
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diagnosis
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Foot-and-Mouth Disease Virus
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classification
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immunology
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isolation & purification
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Gold Colloid
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Immunoassay
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methods
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Immunoglobulin G
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immunology
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Reagent Strips
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Sensitivity and Specificity
3.The expressions of HSP70 and alphaB-crystallin in myocarditis associated with foot-and-mouth disease virus in lambs.
Mustafa Yavuz GULBAHAR ; Yonca Betil KABAK ; Mehmet Onder KARAYIGIT ; Murat YARIM ; Tolga GUVENC ; Unal PARLAK
Journal of Veterinary Science 2011;12(1):65-73
This study describes the expression of heat shock protein70 (HSP70) and alpha-basic-crystallin (alpha-BC) and their association with apoptosis and some related adaptor proteins in the pathogenesis of foot-and-mouth disease virus (FMDV)-induced myocarditis in lambs. HSP70 was generally overexpressed in the myocardial tissues and inflammatory cells of FMDV-induced myocarditis with differential accumulation and localization in same hearts when compared to non-foot-and-mouth disease control hearts. alpha-BC immunolabeling showed coarse aggregations in the Z line of the cardiomyocytes in FMDV-infected hearts in contrast to control hearts. Overall, the results of this study show that the anti-apoptotic proteins, HSP70 and alpha-BC, were overexpressed with increased apoptosis in FMDV-infected heart tissues. Both proteins failed to protect the cardiomyocytes from apoptosis as defense mechanisms to the FMDV during the infection, suggesting that the virus is able to increase apoptosis via both downregulation and/or upregulation of these anti-apoptotic proteins.
Animals
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Apoptosis Regulatory Proteins/metabolism
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Foot-and-Mouth Disease/*complications/*virology
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Foot-and-Mouth Disease Virus/*classification
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Gene Expression
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HSP70 Heat-Shock Proteins/*metabolism
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Myocarditis/complications/pathology/*veterinary/virology
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Myocardium/pathology
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Sheep
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Sheep Diseases/*virology
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Turkey
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alpha-Crystallin B Chain/*metabolism
4.Rescue of bovine Asia 1 serotype foot-and-mouth disease virus from a full-length cDNA clone.
Shuang LI ; Runxiang ZHANG ; Ge SONG ; Mingchun GAO ; Xiangtao LIU ; Junwei WANG
Chinese Journal of Biotechnology 2009;25(11):1621-1626
After sequencing the Asia 1 foot-and-mouth disease virus (FMDV) (As01 strain), we amplified the two fragments covering the whole genome by overlapping PCR and long PCR. The 5' fragment was 1.8 kb in length including 15Cs, and the 3' fragment was 6.7 kb in length. The two fragments were cloned into the pBluescript SK vector to construct recombinant plasmid pBSAs carrying the full-length cDNA of FMDV As01 strain. The RNA transcript was synthesized in vitro using T7 polymerase and transfected into BHK-21 cells. We observed the typical CPE caused by rescued FMDV. The harvested virus was confirmed to be Asia 1 FMDV by RT-PCR, indirect immunofluorescence assay (IFA) and electron microscope observation. The rescued virus showed a similar pathogenicity in suckling mouse (LD50) compared to its wild-type virus. The infectious cDNA clone of the FMDV As01 strain laid a new ground for further investigation of FMDV virulence determinants and development of novel vaccines against FMD.
3' Untranslated Regions
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5' Untranslated Regions
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Animals
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Base Sequence
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Cattle
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Cell Line
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Cloning, Molecular
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DNA, Complementary
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genetics
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Foot-and-Mouth Disease
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metabolism
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virology
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Foot-and-Mouth Disease Virus
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classification
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genetics
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growth & development
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Genome, Viral
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Mice
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Molecular Sequence Data
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Plasmids
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RNA, Viral
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genetics
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Transfection
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Virulence
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Virus Replication
5.Molecular characterization of a 13-amino acid deletion in VP1 (1D) protein and novel amino acid substitutions in 3D polymerase protein of foot and mouth disease virus subtype A/Iran87.
Majid ESMAELIZAD ; Saber JELOKHANI-NIARAKI ; Khadije HASHEMNEJAD ; Morteza KAMALZADEH ; Mohsen LOTFI
Journal of Veterinary Science 2011;12(4):363-371
The nucleotide sequence of the VP1 (1D) and partial 3D polymerase (3Dpol) coding regions of the foot and mouth disease virus (FMDV) vaccine strain A/Iran87, a highly passaged isolate (~150 passages), was determined and aligned with previously published FMDV serotype A sequences. Overall analysis of the amino acid substitutions revealed that the partial 3Dpol coding region contained four amino acid alterations. Amino acid sequence comparison of the VP1 coding region of the field isolates revealed deletions in the highly passaged Iranian isolate (A/Iran87). The prominent G-H loop of the FMDV VP1 protein contains the conserved arginine-glycine-aspartic acid (RGD) tripeptide, which is a well-known ligand for a specific cell surface integrin. Despite losing the RGD sequence of the VP1 protein and an Asp26-->Glu substitution in a beta sheet located within a small groove of the 3Dpol protein, the virus grew in BHK 21 suspension cell cultures. Since this strain has been used as a vaccine strain, it may be inferred that the RGD deletion has no critical role in virus attachment to the cell during the initiation of infection. It is probable that this FMDV subtype can utilize other pathways for cell attachment.
Amino Acid Sequence
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Amino Acid Substitution
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Antigens, Viral/chemistry/*genetics/metabolism
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Capsid Proteins/chemistry/*genetics/metabolism
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Cloning, Molecular
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Foot-and-Mouth Disease Virus/classification/*genetics/*metabolism
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Gene Expression Regulation, Viral
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Molecular Sequence Data
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Phylogeny
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Viral Nonstructural Proteins/chemistry/*genetics/metabolism