DETECTION OF RETICULOENDOTHELIOSIS VIRUS IN LIVE VIRUS VACCINES OF POULTRY

Authors: Fadly, Aly; GARCIA, MARICARMEN - UNIVERSITY OF GEORGIA

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 10/3/2005
Publication Date: 10/3/2005
Citation: Fadly, A.M., Garcia, M. 2005. Detection of reticuloendotheliosis virus in live virus vaccines of poultry [abstract]. New Diagnostic Technology: Applications in Animal Health and Biologics Controls. p.65.

Interpretive Summary:

Technical Abstract: Reticuloendotheliosis virus (REV) is an avian oncornavirus that is structurally and antigenically unrelated to the leukosis-sarcoma group of viruses. All REV isolates are antigenically related to each other. However, using monoclonal antibodies, REV isolates can be classed into three different subtypes, A, B and C. REV infects chickens, turkeys, ducks, geese, pheasants, quail, and probably many other avian species. The most common clinical diseases induced by REV are chronic lymphomas (reticular cell tumor, B- or T-cell lymphoma) and an immunosuppressive runting disease. Although losses in REV affected flocks can be significant due to tumor mortality and immunosuppression, the principal economic concerns of REV infection are as contaminants of live-virus vaccines of poultry or as a barrier to export of breeding stock to certain countries. Because REV can be transmitted vertically from dams to offspring, embryos and tissue cultures prepared from such embryos may harbor REV; therefore these embryos or cells could serve as a source of REV contamination of poultry and other vaccines produced in such ingredients. Accidental contamination of live virus poultry vaccines such Marek’s disease virus (MDV) and fowlpox virus (FPV) with REV has been reported, sometimes resulting in major economic consequences. The fact that partial or complete insertion of REV genome in large DNA viruses such as MDV and FPV can occur also poses a problem. Most insertions consist of a solitary long terminal repeat (LTR), sometimes with partial deletions. However, full-length, infectious REV genomes have been detected in turkey herpesvirus, and a nearly full-length, infectious REV provirus has also been detected in certain strains of FPV. In vitro and in vivo assays have been used for detection of REV in live-virus poultry vaccines. The presence of REV is confirmed by the demonstration of viral antigen or provirus in chicken embryo fibroblasts (CEFs) or in pathogen-free (SPF) chickens inoculated with suspect vaccines. To avoid or minimize cytopathic effects induced by vaccine virus in CEFs, filtration, neutralization or other methods to rid the inoculum of the vaccine virus are used prior to testing vaccines for REV. Using REV polyclonal or monoclonal antibodies, CEFs inoculated with vaccines can be examined for REV by immunofluorescence or immunoperoxidase staining methods. Cell lysates from such inoculated CEFs can also be used for detection of REV major group-specific antigen (p30) by an enzyme-linked immunoassay. It is worth noting that if in vivo assays of vaccines are used for detection of REV contamination, the assay should include a virus detection test, as negative antibody test can be misleading. Detection of proviral DNA by polymerase chain reaction (PCR) assays that amplifies the 291 base pairs product of REV LTR has been shown to be a sensitive and specific method for detection of various strains REV in infected CEFs as well as in blood of SPF chickens inoculated with contaminated FPV vaccines. Reverse transcriptase PCR (RT-PCR) can also be used in detection of REV in contaminated vaccines. Most recently, using PCR tests that amplify REV envelope and REV 3’ LTR sequences provided a more accurate assessment of the inserted REV provirus than PCR assays that amplify the REV 5’ LTR. Although vaccine manufacturers are required to produce vaccines in CEFs or embryos obtained from flocks that are know to be free from various avian pathogens including REV, there is no standardized method that is used for screening commercial vaccines for REV contamination. Furthermore, quality control procedures to exclude REV from licensed poultry biologics are not uniformly effective. This presentation is primarily aimed at methods that can be used for evaluation of commercial live virus poultry vaccines for possible contamination with REV.