Isolation,Identification And Genome Analysis Of Pigeonpox Virus

Pigeonpox is a highly infectious viral disease caused by pigeonpox virus.The symptoms are pox on the body surface skin,eyelid edge,oral mucosa,feet and other parts.In late July 2020,our laboratory carried out an epidemiological survey on pigeon stalls in Xi’an,Shaanxi Province,and found that pigeons suspected of being infected with PGPV existed in each pigeon stall.In this study,the diseased skin tissue materials of pigeons suspected to be infected with pigeonpox virus in a pigeon racing shed in Xi’an,Shaanxi Province were collected.The virus was isolated by inoculating 11 day old SPF chicken embryo chorioallantoic membrane,and verified by a series of identification methods such as animal pathogenicity test,histopathological observation,virus liquid electron microscope observation,PCR method and ha test.The results were consistent with the biological characteristics of Pigeonpox virus.The p4 b gene of the isolate was amplified by PCR,and the genetic evolution analysis was carried out after sequencing verification.The results showed that the isolate belonged to A2 subtype and was named PGPV/SX-20.By inoculating chicken embryo,the half infection amount（EID50）of chicken embryo of PGPV/SX-20 isolate was 10-5.5/0.2ml;By inoculating pigeons,the minimum infection amount of PGPV/SX-20 isolate was 100EID50;Through animal pathogenicity test,it was found that PGPV/SX-20 isolate infected pigeons,but not chickens.The genome sequence of PGPV/SX-20 was obtained by high-throughput Illumina sequencing technology,with a total length of 282182 bp.It is mainly located in the central coding region of the whole PGPV genome,and the content of GC is 29.5%.Comparing the genome sequence of PGPV/SX-20 with that of six avian poxviruses,it was found that it had the highest similarity with Fe P2 genome sequence of PGPV strain in South Africa.Blast（V2.10.0 +）alignment was conducted between PGPV/SX-20 genome sequence and virus-NT database to determine the candidate reference sequence with the closest evolutionary relationship.Combined with the candidate sequence,Prokka（V1.14.5）supporting software was used for gene function annotation,268 genes were identified and 268 proteins were predicted to be encoded.The corresponding genes were analyzed according to the characteristics of these predicted coding proteins.By comparing the nucleotide sequence consistency between PGPV/SX-20 genome sequence and REV（NC₀06934.1）genome,it was found that there was no Rev genome fragment integrated in PGPV/SX-20 genome.To sum up,this study provided a scientific basis for PGPV related epidemiological investigation,development of attenuated vaccine,antigenicity analysis,development of live virus vector,etc.Through the isolation and identification of PGPV and whole genome sequencing of PGPV infected pigeons in a racing pigeon shelter in Xi’an.