Effect Of Vaccinal Immunity To Marek's Disease By Infection With Chicken Infectious Anemia Virus

Marek's disease?MD?is a lymphoproliferative disease of domestic chickens caused by Marek's disease virus?MDV?.Vaccination has been successfully used to control MD from causing serious outbreaks.Current vaccines may fail to provide sufficient protection even though the chickens were vaccinated with CVI988/Rispens or 814 strains of MD.It is found that the co-infection with other viruses are very common in the infected chickens,especially immunosuppressive viruses such as chicken infectious anemia virus?CIAV?,avian reticuloendotheliosis virus?REV?.Immunosuppression diseases can lead to growth retardation,which causes serous economic losses in chickens to the poultry industry.Marek's Disease by Infection with CIAV has drawn increasing attention.Chicken infectious anemia?CIA?that caused by CIAV,is characterized by aplastic anemia and generalized lymphoid atrophy with concomitant immunosuppression with secondary viral,bacterial or fungal infections in chickens,which can be spreadhorizontally.As early as 1880s,CIAV was found in MD infected chickens.CIAV can be passaged and cause lesions in the chicken MDV tumor cell line MDCC-MSB1 cells.In this study,we analyzed the effects of congenital CIAV on the immunization of commercial MDV vaccine CVI988/Rispens.This study reveals that CIAV affects the immune effect of MDV vaccine from different perspectives.It provides scientific theoretical basis for prevention of poultry diseases.1.Infection of CIAV-SD15 significantly reduces the immune protection of Marek's virulent strains with Marek's virus vaccineTwo-hundred chicken embryo fibroblast?SPF?chickens were randomly divided into five groups with 40 per group at 1 day old and were housed separately in a negative-pressure isolator.All chickens from groups 1,2,3,and 4 were intra-abdominally infected at 1 day old with CVI988/Rispens.Groups 2 and 3 were inoculated intraoral in addition with a dosage of 400 EID₅₀0 of CIAV-SD15.Chickens from groups 3,4,and 5 were challenged with 2000PFU of GX0101on day 5 posthatch.Chickens that died during 90 days post-challenges with GX0101were removed from the isolators and recorded and also necropsied.Marek's disease vaccine CVI988/Ripens can provide better protection against SPF chickens infected with2000 PFU GX0101 at 5 days of age.CVI988/Rispens can reduce the pathogenicity of GX0101 infected chickens.The mortality rates of groups 1,2,3,4,and 5 were 0,14.3%,42.9%,5.71%,and 31.4%.The MDV vaccine cannot prevent MDV infection,but it can effectively reduce the proliferation of MDV.CIAV infection reduces the immune effect of MDV vaccine and enhances the pathogenicity of MDV wild virus.The body weight of the chickens in different groups was measured at 0,5,9,16,23,30,37,and 44 days post-infection?dpi?with GX0101 to evaluate the effect of viral infection on growth rates.All chickens from each treatment group were vaccinated with Newcastle Disease Virus?NDV?and Avian Influenza Virus?AIV?–H9 inactivated vaccines according to the previously described procedure at 10 days old.On days 21,28,and 35 post-vaccination,serum samples were randomly collected from chickens of each group.After 9 and 16 dpi,five chickens per group were used to evaluate the immune organs indices,the immune organs indices were determined by the relative weight of the thymus,spleen,and bursa to the whole body.The results showed that compared with CVI988/Rispens immunization group,CVI988/Rispens immunized with GX0101 challenge group had no significant difference in body weight at different time?P>0.05?,while CVI988/Rispens immunization group GX0101and CIAV infection group were significantly decreased?P<0.05?.Infection of SD15 strain of CIAV significantly reduced the weight and antibody titers to AIV/NDV inactivated vaccines of chickens immunized with CVI988/Rispens,and resulted in atrophy of thymus/bursa and enlargement of spleen.2.Infection with CIAV-SD15 significantly reduces the prophylactic effect of virulent Marek's virus in vivo replicationThe MDV-specific primers were designed to be specific for the unique molecular marker of REV LTR in GX0101.Blood samples in anticoagulants were collected from six chickens of each of the GX0101-infected groups?groups 3,4,and 5?at 5,9,16,23,and 30 dpi.DNA from peripheral blood lymphocytes?PBLs?were extracted using standard procedures.GX0101 DNA in PBLs was quantified with real-time quantitative PCR?RT-qPCR?according to the previous method.The results showed that CVI988/Ripens inhibited the ability of GX0101 to replicate in chickens.In the GX0101 infected group,the copy number of lymphocytes reached the peak on the 23 day after the challenge,while the CVI988/Rispens-immunized GX0101 infected group infected the GX0101 on the 16th day after the challenge.The number reached the peak,CVI988/Rispens immunization with GX0101,CIAV infection group increased the number of copies of lymphocytes GX0101 until30 days after infection,GX0101 copy number of different detection time was significantly higher in the GX0101 infection group than the CVI988/Rispens immune GX0101,CIAV infection group,higher than CVI988/Rispens-immunized GX0101-infected group?P<0.05?.Total RNA was extracted from PBLs collected from six chickens of each group?groups 3 and4?at 0,5,9,16,and 23 dpi with GX0101.The production of cytokine mRNA of interleukin-6?IL-6?,IL-18,and gamma interferon?IFN-??at different stages was quantified by RT-quantitative reverse transcription PCR.The mRNA expression increased significantly after5 days of GX0101 challenge and then decreased again.Decreased protective efficacy of the MDV vaccine may be associated with decreased levels of IL-18 and IFN-?mRNA at a later stage.When infected with CIAV,the lack of chicken IL-6 may also lead to increased MDV titers and decreased vaccine immunity to MD,revealing from the cell transcript level that infection with CIAV affects the immune effect of MDV vaccine CVI988/Rispens on MDV.Overall,infection of CIAV significantly reduced the protective effects of the CVI988/Rispens vaccine against MDV,results in aggravation of the immunosuppression,viremia of GX0101in CVI988/Rispens-immunized/GX0101-challenged chickens.Implying that infection with CIAV or other immunosuppressive viruses may be a major contributor in the frequent attacks of MD in China in recent years.