Human cytomegalovirus (HCMV) is a herpesvirus that infects a majority of the world’s population, where it persists as a life-long infection in a non-replicating latent state. Periodically, the virus can reactivate from latency, resulting in new infectious virus. Primary and reactivated infections are usually mild or asymptomatic in immunocompetent individuals. However, virus reactivation in immunocompromised haematopoietic stem cell transplant (HSCT) recipients occurs frequently, and this is one of the most serious post-transplant complications that can be life-threatening. Diagnosis of reactivation in HSCT patients relies upon detection of a rise in viral genome copies in blood, at which point the virus has already replicated extensively, and this can limit the effectiveness of anti-viral therapies. We utilized a high-throughput HCMV gene-specific qRT-PCR array approach to interrogate a majority of the HCMV transcriptome in allogeneic HSCT recipients with reactivated HCMV. RNA from blood drawn on a weekly basis before, during and after reactivation was screened for expression of ~130 HCMV mRNAs. This approach detected multiple viral transcripts that preceded the rise in viral genome copiess by several weeks, providing evidence that patterns of viral gene transcription may serve as a useful means for early detection and/or prediction of virus reactivation. In addition to analysis of viral gene expression, we also sought to better understand why some HCMV seropositive HSCT recipients suffer from reactivation, whereas others do not. Analysis of 84 cytokine mRNAs by qRT-PCR revealed distinct expression patterns in HCMV-seropositive HSCT recipients who either did or did not undergo HCMV reactivation. For example, upregulation of IL-5, leukaemia inhibitory factor, Fas ligand and TNF superfamily member 11 was strongly associated with HCMV reactivation, whereas the parallel comparison in HCMV-seropositive recipients in which the virus did not reactivate, revealed a different subset of differentially regulated cytokines, including upregulated CCL24, IFN-γ, IL-15, IL-2 and CD40 ligand. This analysis indicates that reactivation is linked to a unique cytokine signature. Together, analysis of viral and cellular gene expression profiles in naturally infected HSCT recipients has enabled the first identification of potential novel markers and/or predictors of virus reactivation.