Epidemiological Investigation And Pathogenicity Analysis Of Canine Distemper Virus In Part Areas Of Henan And Hebei

Canine distemper virus(CDV)has emerged as a significant disease of wildlife,which is highly contagious and readily transmitted between susceptible hosts.Although CDV was initially described as an infectious disease of domestic dogs,it has increasingly become known as a worldwide multi-host pathogen,infecting and causing mass mortalities in a wide range of carnivore species.Previous studies have shown that CDV outbreaks have an increasing impact on wildlife populations and potentially threaten the survival of endangered wild species.Other studies have shown that CDV may even become a zoonotic pathogen.In recent years,the canine distemper(CD)epidemic often occurs in China,and CD tends to be highly infection and mortality rate.Consequently,it is extremely important to investigate the variation and pathogenicity of the current epidemic strain.To understand the popular characteristics of CDV,we first collected a total of459 samples of dogs,minks,raccoon dogs and foxes from Henan and Hebei provinces during 2019 and 2020.We used RT-PCR(Reverse transcription polymerase chain reaction)to identify the CDV-positive samples.The results showed that 87 subjects of the 459 samples(18.96%)were CDV-positive.In parts of Henan and Hebei provinces,CDV was detected in animal samples of different species.The incidence rate of canine distemper(CD)in dogs was highest in autumn(37.77%)and the lowest in winter(11.10%),and was highest in fur animals in July(Martes:33.33%;Raccoon dog:36.36%;Fox:36.84%).Young animals showed significantly higher levels of CDV positive than adult animals.The experimental results showed that the infection rate of CDV was distinctively different in species,regions,seasons and ages of animals.To understand the genetic evolution of CDV,a CDV strain was secondly isolated from lung samples of a dog that was infected with CDV from a pet hospital in Hebei province.After identification of RT-PCR,morphology observation,and indirect immunofluorescence assay,the isolate was proved to be a CDV wild strain,and named HB19-1.The hemagglutinin(H)gene of HB19-1 was amplificated by RT-PCR,cloned,and sequenced.Next,we performed a phylogenetic analysis based on the H gene nucleotide sequences.Sequence analysis of H genes indicated that HB19-1 belongs to the Asia-1 genotype.Phylogenetic analysis of the viral H gene showed that HB19-1 has the closest nucleotide similarity and amino acid with KJ994343 from Gen Bank and the farthest nucleotide similarity and amino acid with AF378705.Finally,we used puppies without CDV-positive serum to clarify the pathogenicity of the HB19-1 strain.The results showed that the puppies were sensitive to the HB19-1 strain.The puppies developed fever,severe conjunctivitis,pathological lesions,and high viral RNA loads within 3-6 days post-infection(dpi),culminating in75%(3/4)mortality within 21 dpi.Compared with the control group puppies,the temperature rise showed fever,but temperature change had no “dual phase heat”characteristics.The quantitative real-time PCR(q PCR)method was used to detect the CDV load in all tissues of infected puppies.The results showed that the lymph nodes had the highest levels of the virus.This study revealed that the HB19-1 strain caused a serious infection in the lymph nodes,and led the host to death.In this study,we isolated and obtained a CDV strain,HB19-1,during the investigation of CDV in parts of Henan and Hebei from 2019 to 2020.The sequence analysis of H genes of HB19-1 indicated that HB19-1 belongs to the Asia-1 genotype and has low homology with the vaccine strain.The results of the pathogenicity study showed that puppies were sensitive to the HB19-1 strain,and this strain had the strongest ability to infect lymphatic tissue.The results are helpful for the prevention and control of CDV in parts of Henan and Hebei.