Isolation And Biological Characterization Of Canine Parainfluenza Virus

Canine parainfluenza virus(CPIV)is the main cause of canine cough,which was isolated from dogs suffering from respiratory diseases for the first time in 1967,and its main clinical manifestations are cough,runny nose and other respiratory diseases.At present,canine parainfluenza virus is distributed all over the world,which has brought huge economic losses to the dog breeding industry.Therefore,isolating the current epidemic strain of canine parainfluenza virus is conducive to understanding the epidemic situation of the virus,as well as the prevention and control of the disease.This research work is mainly carried out in three parts:1?this experiment established a multiplex PCR method for rapid detection of canine parainfluenza virus,canine distemper virus,canine parvovirus and canine adenovirus type II.The results showed that the specificity and sensitivity were good.Thirty clinical samples of infected dogs were detected by the multiplex PCR,and the positive rate of CPIV was 0%(0/30),CDV 63.33%(19/30),CPV 33.33%(10/30)and CAV-2 6.66%(2/30).The established PCR method can be used for clinical diagnosis.2?We collected the lung tissues of sick dogs with symptoms of respiratory diseases such as cough and runny nose,and detected by the established multiplex PCR method.The single infected with CPIV was prepared into samples,and inoculated on Vero cell to cultivate.The virus was identified by RT-PCR,virus electron microscopy and hemagglutination test.)The results RT-PCR assay showed a 534 bp specific target fragment.The ultrastructural observation of the virus was circular,with a diameter of80-200 nm and envelop,which was the same as Jin Changde's report.The virus is capable of agglutinating 1% of pig and chicken red blood cells,consistent with the characteristics of the parainfluenza virus.The ORF sequence of the CPIV NP gene was cloned and compared with the parainfluenza virus on GenBank,and the homology was over 97%.The isolated virus was identified as canine parainfluenza virus and named CPIV-BJ01.3?In this study,a one-step growth curve was measured for the eighth and ninth generations of CPIV-BJ01.The results showed that the virus proliferated rapidly at12-48 h,and the virus titer in the cell culture solution rose sharply and became stable after 60 h.The virus was subjected to a thermal test,an ultraviolet irradiation test,anda full-length sequence analysis.The results showed that the virus was sensitive to heat and prolonged uv irradiation would kill the virus.The full-length sequence analysis showed that the virus was 99% homologous with PIV5 1168-1(login number:KC237064.1)and PIV5 ZJQ-221(login number: KX100034.1),and it was in the same branch with a relatively close genetic relationship.