|Garcia, Maricarmen - UNIVERSITY OF GEORGIA|
|Reed, Willie - MICHIGAN STATE UNIVERSITY|
Submitted to: Avian Diseases
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: November 18, 2002
Publication Date: January 1, 2003
Citation: GARCIA, M., NARANG, N., REED, W.M., FADLY, A.M. 2003. MOLECULAR CHARACTERIZATION OF RETICULOENDOTHELIOSIS VIRUS INSERTIONS IN THE GENOME OF FIELD AND VACCINE STRAINS OF FOWLPOX VIRUS. AVIAN DISEASES. 47:343-354. Interpretive Summary: Fowlpox is an economically important disease of domestic chickens and turkeys caused by a virus known as fowlpox virus (FPV). The disease is and has been controlled by vaccines. These vaccines are vulnerable to contamination with another virus termed Reticuloendotheliosis virus (REV) that can be present in chicken embryos or cells used to prepare the vaccine. This research developed highly sensitive DNA-based methods that can be used to determine whether part of or the complete REV is present in the FPV. Using this technology, five field isolates of FPV and one commercial vaccine were shown to have the complete REV. The information is useful to scientists in academia and industry, particularly vaccine manufacturers.
Technical Abstract: Evidence of the wide spread occurrence of reticuloendotheliosis virus (REV) sequence insertions in fowlpox virus (FPV) genome of field isolates and vaccine strains has increased in recent years. However only those strains carrying a near intact REV provirus are more likely to cause problems in the field. Detection of the intact provirus, or REV protein expression from FPV stocks has proven to be technically difficult. The objective of the present study was to evaluate current and newly developed REV and FPV PCR assays to detect the presence of REV provirus in FPV samples. The second objective was to characterize REV insertions among recent "variant" FPV field isolates and vaccine strains. Using REV, FPV and heterologous REV-FPV primers, five FPV field isolates and four commercial vaccines were analyzed by PCR and nucleotide sequence analysis. Intact and truncated REV 5' LTR sequences were detected in all FPV field isolates and vaccine strains indicating heterogeneous REV genome populations. However only truncated 3' LTR and envelope sequences were detected among field isolates and in one vaccine strain. Amplifications of the REV envelope and 3' LTR provided strong evidence to indicate that these isolates carry a full-length REV genome. Three of the four FPV vaccine strains analyzed carry in solo intact or truncated 5' LTR sequences indicating that a complete REV provirus was not present. Comparison of PCR assays indicated that REV envelope and REV 3' LTR PCR provided a more accurate assessment of REV provirus than PCR assays that amplify the 5' LTR. Therefore, to differentiate FPV strains that carry complete REV provirus from those that carry solo 5' LTR sequences, positive PCR results with primers that amplify the 5' LTR should be confirmed with more specific PCR assays that either amplify REV structural genes, as the envelope, or REV-FPV heterologous primers as the pair used to amplify the REV 3' LTR.