Isolation And Identification Of Feline Panleukopenia Virus And Evolution Analysis Of VP2 Gene In Yinchuan Of Ningxia Hui Autonomous Region

Feline panleukopenia virus(FPV),also known as feline distemper virus and feline parvovirus,is a virus that causes the main clinical features of kittens with vomiting,diarrhea,bidirectional fever and severe leukopenia.The causative agent of sexually transmitted diseases.The disease is widely distributed all over the world,and the fatality rate is increasing year by year,which has brought huge economic losses to the cat industry.The full length of the FPV VP2 gene is 1755bp,encoding 584 amino acids.The VP2 gene determines the host specificity,hemagglutination and antigenicity of the FPV virus.The VP2 structural protein is the main structural protein of FPV and plays an important role in the infection and replication process of the virus.It can induce the animal body to produce neutralizing antibodies,play a role in self-protection,and also has the ability to assemble virus-like particles,and is also an important immunogenic protein.1.In order to understand the infection of domestic cats and cats with panleukopenia in Yinchuan,Ningxia,the conserved sequence of FPV VP2 gene(190/73 strains)included in Genbank was selected,and a pair of specific primers for detecting FPV virus were designed.From March to November 2021,22 diarrhea fecal samples of domestic cats and 3 suspected FPV-infected cats were collected from Chongyijia Animal Hospital in Xingqing District,Yinchuan and some pet hospitals and clinics in Xixia District and Jinfeng District.In the eye,mouth and nose secretion samples,10 strains of FPV were isolated and identified from 25 samples using CRFK cells,and the 10 strains of FPV cell cultures showed cytopathic effect(Cytopathic effect,CPE)phenomenon of atrophy,rounding and shedding at different times.The 10 FPV strains were further identified by molecular biology identification,virus morphological observation,Indirect immunofluorescence assay(IFA),TCID50 detection,and electron microscope negative staining.By designing primers for the full-length gene of FPV virus VP2,the VP2 gene of 10 FPV isolates was amplified and sequenced.2.As the main coding gene of FPV structural protein,VP2 gene contains all neutralizing antigenic sites of structural protein,so VP2 gene is selected as the target gene of this experiment.The results of the VP2 gene sequence analysis of the 10 isolates showed that the nucleotide identity of the VP2 gene between the 10 isolates and the primer reference strain(190/73 strains)was 98.8%-99.3%,and the nucleotide identity of the 10 isolates was 98.8%-99.3%with the standard(FPV-CU4 strain)VP2 The nucleotide identity of the gene was 98.8%-99.2%,and the amino acid identity of the isolated strain,primer reference strain and standard strain were 86.9%-97%.3.Compared with the nucleotide sequence of the VP2 gene of the reference primer strain(190/73 strain),it was found that there were 13 gene loci changes,7 synonymous mutations,and the remaining 6 scattered mutations.the same direction of genetic evolution.The mutated amino acids in the isolates were located at positions(200-250),which were the mutation of Gln→Thr at position 213 and the mutation of Val→Gln at position 232,respectively.The mutation of Ser→Pro at position 344 of FPV 12 is unique among the isolates.The results of genetic phylogenetic tree analysis showed that of the 10 FPV isolates,only FPV21 and FPV24 and the Chinese(Nanjing)CPV-2c strains MH476592 and MF805797 were recently clustered into a small branch,and the remaining strains were closely related to the current domestic FPV strains.The prevalent strains(Changchun,Shanghai,Jilin)are relatively closely related and are in the same branch,but there is no obvious Chinese branch.In summary,this study analyzed the FPV infection of domestic cats in Yinchuan,Ningxia,explained the prevalence and genetic evolution of FPV in Yinchuan,Ningxia at the molecular level,and clarified the relationship between FPV isolates and other domestic and foreign epidemic strains.The genetic relationship and variation situation provide certain data support for the prevention and control of panleukopenia in domestic cats and the formulation of reasonable immunization procedures,and also provide a reference for the follow-up FPV research in this region.