Acyclovir (ACV) has been used for more than 15 years in the management of herpes simplex virus (HSV) and varicella-zoster virus (VZV) disease. The present survey was undertaken to assess the level of ACV resistance in the population. More than 2,000 HSV isolates from both immunocompetent and immunocompromised patients in northwest England were collected over a 2-year period and tested for sensitivity to ACV. These studies suggested a prevalence of resistance of approximately 0.1 to 0.6% in immunocompetent individuals, with no apparent difference in prevalence between treated and untreated groups. In line with previous studies, the prevalence of resistance in treated immunocompromised individuals was approximately 6%.