| Marek's disease(MD)is a highly infectious disease of chicken,which is characterized by lymphoid hyperplasia and tumor transformation.The pathogen of MD is Marek's disease virus(MDV).MDV infection can directly lead to the death and the decline in production performance of the host,and indirectly lead to immune suppression.MDV is popular all over the world,has caused enormous economic losses to the poultry industry.The main strategy of MD prevention and control is vaccination,but MD vaccine did not induce sterile immunity,MDV field strain and vaccine strain can coexist in the host,and allow viral replication and transmission,lead to the evolution of MDV field strain.Although universal vaccination is made in China,MD often occurred across the country in recent years,causing great harm to the poultry industry.Therefore,it is necessary to understand the epidemic situation of MDV in China,and to isolate and identify the MDV field strain.These studies are of great significance for the basic research in MDV and the prevention and control of MD.In this study,489 clinical samples suspected MD positive were received from 15 provinces during August 2015 to July 2016.Detection results showed that the positive rate of MDV reached 42.7%,and mixed infection rates with avian reticuloendotheliosis virus(REV),avian infectious anemia virus(CIAV)and avian leukemia virus(ALV)in MDV positive samples were 12.6%,17.5% and 1.8%,respectively.The isolation and identification of the pathogen of some MDV positive samples were performed,and 4 MDV strains were isolated and identified,which were named BC/15,HLD/16,JL/16 and SY/15,respectively.By PCR detection,BC/15 and HLD/16 were identified as MDV field strains,while JL/16 and SY/15 were MDV vaccine or attenuated strains.Genetic analysis based on the meq gene showed that BC/15 and HLD/16 were in the same evolutionary branch with most Chinese virulent MDV strains,but the two further divided into a new separated branche,while JL/16 and SY/15 were in the same branch with the early Chinese MDV isolates J-1,as well as the vaccine strains CVI988 and 814 and mild strain CU-2.These results indicate that MDV strains with different characteristics are prevalent in China,and the MDV is still evolving.In 2015,a MDV field strain BS/15 was isolated from a severely diseased Chinese chicken flock previously vaccinated with CVI988 vaccine,which located in Jilin,China.By PCR and IFA detection,MDV BS/15 has the characteristics of MDV field strains.Furthermore,four pathogenicity-related genes meq,vIL-8,pp38 and ICP4 was sequenced,and sequence comparison was performed,the results showed that MDV BS/15 included many unique mutations which characteristics are in line with most of the Chinese isolates.To study the pathogenicity of BS/15 and evaluate the protection effects of the existing vaccines,we compared the pathogenicity of MDV BS/15 and MDV reference strain vvMDV Md5 in SPF chickens,and evaluated the protective effects of CVI988 and 814 vaccines,which is most widely used in China,to the two strains.The results showed that MDV BS/15 and Md5 could lead to the occurrence of MD in 100% SPF chickens during the 90-day experimental period.Mortality rates were similar at 85.7% and 80%,respectively.The tumor rate of MDV BS/15 challenged group was 64.3%,which was higher than that of Md5 challenged group(40.0%)However,compared with Md5,the survival time of BS/15 infected chickens was longer,and the damage to the growth and development was slighter and the damage to the immune organs was similar.These results indicated that the virulence of MDV BS/15 was comparable to that of Md5,but its pathogenic characteristics were changed.In addition,the protective index of CVI988 and 814 for the BS/15 were 33.3 and 66.7,respectively,while the protective index of the to the Md5 was 92.9 and 100,respectively.These results indicated that the two vaccines do not provide adequate protection for BS/15,especially the CVI988 vaccine.Taken together,these data suggested that BS/15 has the characteristics of “late virulence” and high vaccine resistance.These studies suggested the existent of MDV variant strains in China and these strains may be one reason for the MD vaccination failure.These studies can help us to understand the pathogenicity and evolutionary characteristics of MDV strains which are epidemic in China.In this study,we investigated the effects of MDV infection on the expression of interferons in the central immune organs of chickens,and studied the effects of different MDV strains(BS/15 and Md5)on the differential expression of host interferons for the first time.It was found that MDV infection could significantly inhibit the expression of IFN-? and IFN-? in the bursa and thymus of SPF chickens.In the early stage of MDV infection,the expression of IFN-? was significantly up-regulated,then was significantly down-regulated.These results indicated that MDV could affects the innate immune system and inhibits the expression of interferons to avoide the antiviral effects of the host.In addition,the effects of different MDV strains on the differential expression of host interferons were significantly different,which may be related to the pathogenic characteristics and vaccine resistance of MDV strains.To further analyze the infection process of the different MDV strains,the fluorescence in situ hybridization method(FISH)was established to detect integration of MDV genome into the host chromosome.In vivo infection model,we detected the integration of the MDV genome in the host chromosome.We performed the initial analysis of the host integration rules of different MDV strains(BS/15 and Md5)in the host.The results showed the integration rule of different MDV strains in the host chromosome is different,which may be related to the different pathogenicity of MDV strains.In summary,our study performed detection,isolation,identification and analysis of part of epidemic MDV strains in China,and systematically studied the biological characteristics of Chinese MDV isolates BS/15.The differential expression of host infections induced by MDV infection was analyzed,as well as its integration rules in the host chromosomes.These results help us to understand the prevalence,pathogenicity and evolution of Chinese MDV strains,which is of great guiding significance for the prevention and control of MD in China,and provides support for the basic research of MDV. |
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