This study was aimed at investigating the genes that control host responses to Marek's disease virus (MDV). Spleen tissues from infected and age-matched uninfected control chickens were examined 4, 7, 14, and 21 d postinfection for gene expression differences, using both microarray and quantitative real-time polymerase chain reaction (PCR) methodologies. Up to 51% of genes assayed during microarray analysis showed a significant change (p < or = 0.05) in expression after MDV infection, of which cell surface molecules, transcription and signal transduction molecules, housekeeping and metabolism mediators, and cytokines and cytokine receptors were most commonly differentially expressed. Setting a fold change threshold (> or =2), 14 of 84 genes showed differential expression over time. In addition, some genes showed differential expression at more than one time point. For example, the granzyme-A homolog gene remained upregulated in infected chickens, with fold changes of 7.98, 13.91, and 9.07 at 7, 14, and 21 d postinfection, respectively. Other genes that were differentially expressed at more than one time point were invariant chain, IgM, and CD3. Quantitative real-time PCR analysis was used to validate microarray results for a subset of genes showing a :2-fold change in expression. Expression of all but one gene (CD28) was confirmed. Identification of genetic mechanisms initiated by in vivo infection with MDV expands the current understanding of immune response to the virus in addition to host response elements associated with viral pathogenesis.