Detection of avian leukosis virus in albumen of chicken eggs using reversetranscription polymerase chain reaction

A reverse transcriptase polymerase chain (RT-PCR) assay was developed to de tect avian leukosis retrovirus (ALV) in egg albumen. Eggs of Single Comb Wh ite Leghorns were from a commercial breeder (stock F) and from a pathogen-f ree hock (stock N). RT-PCR was undertaken on isolated RNA from 20 unfertili zed egg samples using seven sets of primers that correspond to the ALV gp85 envelope glycoprotein which determines the ALV subgroup classification. An ELISA assay for ALV gs antigen of egg albumen was positive for all stock F birds tested and negative for all stock N birds. Virus isolation was under taken by inoculating egg albumen, feather pulp, or blood from five stock F chickens onto cultures of chicken embryo fibroblasts (C/E). IFA analysis of the inoculated C/E cultures indicated that all stock F birds tested contai ned infectious ALV. For the virus-positive stock F chickens, RT-PCR analyse s using primers designed to detect all ALV subgroups detected ALV in 15/15 (100%) egg albumen samples, while primers designed to detect subgroup A ALV were positive for 12/15 (80%) egg albumen samples. RT-PCR products were no t detected from five egg albumen samples from five stock N chickens by any primer sets. Direct sequencing using primers specific for subgroup A ALV ve rified the viral subgroup in the RT-PCR amplification products. The combine d use of RT-PCR and direct sequencing of the RT-PCR product provides a new approach for identifying ALV-infected poultry. (C) 1999 Elsevier Science B. V. All rights reserved.