Epidemiological Investigation Of Marek's Disease In The Pearl River Delta Region From 2016 To 2017

Marek's disease virus(MDV)is a herpes virus that causes tumors in chickens to replicate in lymphatic and epithelial tissues.In the past decades,MD has been popular among chickens in our country.The pathogenicity of wild strains of Marek's disease seems to have gradually increased.CVI988/Rispens vaccine has been widely used in the prevention and control of MD in China.During 2016-2017,many poultry farms in the Pearl River Delta region of Guangdong Province reported suspected MD cases.This study intends to carry out MD epidemiological survey in the Pearl River Delta region during this period,and attempts to analyze the immune effect through the copy number of CVI988/Rispens vaccine in the feather pulp(FP),in order to provide reference for prevention and control of MD in this region.This study investigated the epidemiology of suspected MD cases and the immune effect of MD vaccine by clinical autopsy,virus isolation,histopathology and indirect immunofluorescence assay(IFA);and 6 isolated MDV were identified;they were named HD1 D,HD7E,HZM01,HZQ03,HZY04 and HZS03 M strains;PCR identification results can be seen,6 strains of virus can be amplified to pp38 fragment(873bp),Meq fragment(1200bp)and vIL-8 fragment(886bp)and so on,indicating 6 strains of isolates is serotype-?MDV.IFA results of 6 isolates showed that HD1 D,HD7E,HZM01,HZQ03 and HZY04 isolates could be recognized by MDV-1 specific monoclonal antibody BA4 and monoclonal antibody H19,indicating that the five isolates were virulent MDV wild-type strain rather than vaccine strains.To further understand the genetic derivatization characteristics of the major pathogenic genes associated with each MDV isolate,the sequences of Meq,pp38 and vIL-8 from each isolate were amplified by PCR and sequenced.Sequence analysis showed that,the amino acid sequences of Meq gene of HD1 D,HZM01,HZQ03 and HZY04 isolates were found to be highly homologous to the very virulent strains in China(GX0101)of K77 E,D80Y,V115 A,T139A,P176 R,and P217 A.It showed the same mutations at 217(P-A)position as the foreign super-virulent strains(584A),there are mutations at 176 sites but the bases are different.The isolates of HD1 D,HZQ03,HZM01,HZY04,HD7 E and virulent strain(MD5,GX0101)were found to be glutamine(Q)at the 107 th amino acid of pp38 gene,the 109 th amino acid of pp38 gene and the CVI988/Rispens strains were the same glycines(G);The amino acids encoded by the vIL-8 genes of HZM01,HZQ03 and HZY04 isolates showed the same mutation as GX0101 at C38 R,L110P,and D123 G sites.Genetic evolutionary tree analysis based on Meq,pp38 and vIL8 showed that the homology of HZS03 M isolate and CVI988/Rispens was 100%;the homology of isolates HD7 E and 814 strains was 100%;4 isolates(HD1D,HZM01,HZQ03 and HZY04)were on the same branch as domestically isolated virulent strains(GX0101,etc.).From the above,it can be speculated that the HD1 D,HZM01,HZQ03 and HZY04 isolates should be vMDV or vvMDV.In order to investigate the effect of chickens immunized with CVI988/Rispens vaccine,a total of 1031 FP samples in the amount of CVI988/Rispens replication were detected by Q-PCR method in different fields(including A,B,C,D,E,F,G,H,I,J and K fields)in Guangdong Province.The results showed that the replication of CVI988/Rispens between different fields had differences or significant differences in either the 14-day-old or 21-day-old FP samples,especially among different varieties,the difference was extremely significant,it is shown that the average replication amount of CVI988/Rispens between different fields or different varieties is uneven.The detection rate of 565 samples of aged feather tissue of 21 days was 0 based on the wild strain Q-PCR method.However,the same method was used to detect the FP sample of 24 MD chickens,of which 79.17% of the samples could detect the effective replication of MDV wild strains;The 14-21 day old FP sample is suitable for the detection of the replication of the CVI988/Rispens vaccine strain,the 565 samples submitted were not infected with MDV wild strains.The study indicate that there has been virulent strains MDV endemic in the Pearl River Delta region of Guangdong Province over past several years.The prevention and control of MD should continue to be done.In particular,attention should be paid to the immune effect of CVI988/Rispens vaccine.