| Marek's disease (MD) is a lymphoproliferative disease of chickens caused by an acute-transforming, cell-associated herpesvirus, Marek's disease virus (MDV). This work details the construction and characterization of several recombinant attenuated serotype 1 MDVs that have been mutagenized by the insertion of the lacZ gene of E. coli. Recombinant strains were constructed that had lacZ insertions into MDV homologues to herpes simplex virus type 1 US1, US6 and US10 genes. In addition, a deletion mutant was constructed lacking US1, US2 and US10 genes and three MDV-specific genes (Sorfs 1-3). A recombinant strain containing an identical deletion was also constructed in an oncogenic MDV strain, RB1B. Recombinant MDVs were characterized according to changes in virus genome structure, gene expression and growth properties in chicken embryo fibroblasts (CEF) and in chickens. The results indicate: (1) the US1, US2, US6, US10 MDV homologues, as well as MDV-specific genes Sorf1, Sorf2 and Sorf3 are nonessential for the growth of an attenuated or oncogenic MDV in CEF, (2) MDVs lacking a functional US1 gene show impaired growth in established CEF, suggesting the presence of a US1-compensatory factor in dividing or serum-stimulated CEF, (3) MDV US1, US2, US6, US10, Sorf1, Sorf2 and Sorf3 genes are nonessential for the early cytolytic infection of the chicken spleen, (4) immunohistochemical analysis of spleen sections from chickens infected with GAatt, GAatt-derived recombinants, or oncogenic RB1B and T-King MDVs revealed that infected cells could be detected only in chickens infected with the oncogenic viruses. Furthermore, T-King-infected chicken spleens contained non-B-cell infected lymphocytes at five days post-infection, suggesting that, at least for T-King MDV, T-cell infection could precede or be coincident with B-cell infection. The construction of the first recombinant RB1B provides a method for the direct analysis of determinants of MDV tissue tropism and oncogenesis. |
