ABSTRACT
Two giant pandas (Ailuropoda melanoleuca) died of unknown causes in a Chinese zoo. The clinical disease profile suggested that the pandas may have suffered a viral infection. Therefore, a series of detection including virus isolation, electron microscopy, cytobiological assay, serum neutralization and RT-PCR were used to identify the virus. It was determined that the isolated virus was a canine coronavirus (CCV), on the basis of coronavirus, neutralization by canine anti-CCV serum, and 84.3% to 100% amino acid sequence similarity with CCV. The results suggest that the affected pandas had been infected with CCV.
Subject(s)
Animals , Female , Male , Amino Acid Sequence , Animal Diseases/virology , Animals, Zoo/virology , Coronaviridae Infections/veterinary , Coronavirus, Canine/genetics , Fatal Outcome , Molecular Sequence Data , Sequence Alignment , Sequence Homology, Amino Acid , Ursidae/virology , Viral Proteins/chemistryABSTRACT
HA and M2 genes derived from human highly pathogenic avian influenza H5N1 virus (A/Anhui/ 1/2005) isolated from China, were amplified and cloned into the DNA vaccine expression vector pVRC. In order to improve the expression of hemagglutinin, the human codon usage preference was made and the whole length of HA gene of H5NI (A/Anhui/1/2005) influenza virus was synthesized,named HA/YH/K, and inserted to pVRC vector, the expression of HA/YH/K protein in eukaryotic cells was significantly improved according to internal control of actin protein. Furthermore, the M2 and HA/YH/K genes were cloned into bicistronic eukaryotic expressing vector pIRES to yield the recombinant plasmid pIRES-HA/ YH/K-M2/YS/K, which could expressed HA and M2 protein simultaneously by transfection of one plasmid. Western blot and IFA showed that the recombinant pIRES-HA/YH/K-M2/YS/K plasmid was successfully expressed in several mammalian cells (Hela, MDCK and 293FT). The above results may help to identify the function and pathogenic mechanism of HA, M2 genes derived from HPAI H5N1 (Anhui strain) and pave a way for the development of novel bivalent vaccines against human highly pathogenic avian influenza virus and for preparedness for influenza pandemic.
Subject(s)
Animals , Humans , Cell Line , Gene Expression , Genetic Engineering , Genetic Vectors , Genetics , Metabolism , Hemagglutinin Glycoproteins, Influenza Virus , Genetics , Metabolism , Influenza A Virus, H5N1 Subtype , Genetics , Metabolism , Influenza, Human , Virology , Recombinant Fusion Proteins , Genetics , Metabolism , Viral Matrix Proteins , Genetics , MetabolismABSTRACT
In order to survey the infectious situation of canine coronavirus (CCV) in giant panda population, a virus neutralization test detecting specific antibodies against CCV in giant panda's sera was established by using two-fold dilutions of serum and 100 TCID50 of the virus. The 62 sera samples of giant pandas, which were gathered from zoos and reserve region of Sichuan Province, China were detected. The neutralization antibody titer of 1:4 was recognized as the positive criterion, 8 sera samples were detected to be positive, and the positive rate was 12.9%. The titers of neutralizing antibody ranged from 1:8 to 1:32. It was the first comprehensive investigation on neutralization antibodies against CCV in giant panda population in China. The results of study showed that the infection of CCV in giant panda population was universal, which has posed a threat to the health of giant panda. Therefore, it is incumbent on us to study safe and effective vaccines to protect giant panda against CCV infection.
