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United States Department of Agriculture

Agricultural Research Service

Title: Comparative Evaluation of Three Methods for Detection of Reticuloendotheliosis Virus As a Contaminant in a Live-Virus Vaccine of Poultry

Authors
item Fadly, Aly
item Witter, Richard

Submitted to: Avian Diseases
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: February 5, 1997
Publication Date: N/A

Interpretive Summary: Accidental contamination of commercial vaccines with reticuloendotheliosis virus (REV) is an important source of virus infection and has been implicated in induction of runting disease, immunosuppression as well as tumors in vaccinated chickens. Until recently, routine testing of live-virus poultry vaccines for contamination with REV was not required by regulatory agencies in the United States. However, vaccine manufacturers are required to produce vaccines in substrates (cell cultures or chicken embryos) obtained from flocks that are known to be free from various infectious agents including REV. Although several tests can be used for detection of REV using in-vitro (cell cultures) and in-vivo (chicken inoculation) assays of contaminated vaccines, the sensitivity of such tests has not been compared. Data from this study suggest that the sensitivity of all tests for detection of REV contamination in vaccines depends on the concentration of REV in the vaccine, whether or not the vaccine is filtered before testing, and whether in-vitro or in-vivo assays were used. Further, our data indicate that in-vivo assays of vaccines for contamination with REV should include a test for virus, as a negative antibody test may be misleading. The data obtained from this study will assist the biologics industry and regulatory agencies in the development of a mutually agreeable standard requirement that will ensure that live-virus vaccines of poultry are free from REV contamination prior to licensure or prelicense field testing.

Technical Abstract: Indirect immunofluorescence (IFA), polymerase chain reaction (PCR), and enzyme-linked immunosorbent assay (ELISA) were used for detection of reticuloendotheliosis virus (REV) as a contaminant in a live-virus fowl pox (FP) vaccine of poultry. A FP vaccine known to be contaminated with REV was tested by in-vitro and in-vivo assays in chicken embryo fibroblasts (CEFs) and day-old specific pathogen-free (SPF) chicks, respectively. Using in-vitro assays, IFA and PCR were more sensitive than ELISA in detection of REV in CEFs inoculated with REV-contaminated FP vaccine. However, when the vaccine was tested by in-vivo assays using SPF chickens, the sensitivity of ELISA was comparable to that of IFA and PCR. Further, antibody to REV was not detected in SPF chickens up to 4 weeks postinoculation with contaminated vaccine at hatch. Filtration of vaccine before testing in CEFs, to rid inoculum of vaccine virus, resulted in significant reduction in frequency of REV detection by PCR or IFA. The data suggest that the sensitivity of IFA, PCR, and VI-ELISA depends on the concentration of REV in the vaccine, and that in-vivo assays of vaccines for contamination with REV should include a test for virus, as a negative antibody test may be misleading.

Last Modified: 9/1/2014
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