Herpes simplex virus (HSV) commonly reactivates after stem-cell transplantation (SCT), despite acyclovir prophylaxis. Whether HSV-seropositive recipients with HSV-seronegative or type-discordant donors had more frequent and severe HSV infections than those with HSV type-concordant donors was explored. Banked serum samples from HSV-positive SCT recipients and their donors were tested for the presence of HSV antibodies. HSV-1-positive SCT recipients from HSV-1-negative donors had more frequent and longer episodes than HSV-1-positive SCT recipients from HSV-1-positive donors; the proportion of patients receiving antiviral treatment for >10% of follow-up days was 27.4% versus 7.2% (P<.001). Both HSV-1 visceral infection (9.8% vs. 2.2%; P=.001) and acyclovir resistance (5.8% vs. 1.8%; P=.03) were more common in type-discordant than -concordant patients, respectively; these associations were confirmed in multivariable models. Serological testing of donors can identify patients who are at highest risk for HSV-related morbidity, for whom prolonged prophylaxis or donor vaccination (once available) could be considered.