During an immune response to viral infection the draining lymph nodes (LN) undergo significant expansion to accommodate the recruitment of lymphocytes and dendritic cells (DC). LN Stromal cell (LNSC) networks support the LN architecture and control DC and T cell migration within the lymph node via production of chemokines, such as CCL21. These include the fibroblastic reticular cells (FRC), lymphatic endothelial cells (LEC) and blood endothelial cells (BEC) (including high endothelial venules, HEV). Considerable changes in the appearance of these stromal cell networks during infection along with previous data showing transient down-regulation of lymphoid chemokines in response to other viral infections, led us to examine how these stromal cells contribute to hypertrophy of the LN following infection. Following cutaneous herpes simplex virus type 1 (HSV-1) or vaccinia virus infection in mice, immune responses coincided with increased numbers and proliferation of FRC, LEC and BEC that persisted for more than 30 days after infection. Global transcriptional profiling of LNSC after HSV infection revealed substantial changes in gene expression, and many unique pathways activated in each subset. Early expansion of FRC was dependent on type I interferon, yet not T cells. Re-infection with either LCMV or Vaccinia virus induced reduced LNSC expansion despite large T and B cell responses. Contrary to current models, these data suggest that LNSC responses to infection can occur independent of local lymphocyte recruitment and may modulate LN hypertrophy by integrating a range of inflammatory and environmental signals.