Telomere length and telomerase activity have received increased attention as markers of cellular aging, but the determinants of inter-individual variation in these markers are incompletely understood. Cytomegalovirus (CMV) infection may be particularly important for telomere and telomerase dynamics due to its dramatic impact on peripheral blood lymphocyte composition, i.e., increasing the number and proportions of highly differentiated T cells that are characterized by shorter telomere length (TL) and lowered telomerase activity (TA). However, the possible relationship between CMV infection and leukocyte TL and TA has not been well-examined in vivo. This study examined the associations of CMV seropositivity and CMV IgG antibodies with leukocyte (TL) and (TA) in a sample of 434 healthy individuals (ages 53–76) from the Whitehall II cohort. Positive CMV serostatus was significantly associated with lower TA among women, and higher CMV IgG antibody levels were associated with lower TA in the overall sample. However, neither CMV seropositivity nor CMV IgG antibody levels (reflecting subclinical reactivation) among the seropositive were significantly associated with TL. These associations were robust to adjustment for age, employment grade, BMI, and smoking status. The results demonstrate that CMV seropositivity and subclinical reactivation predict lower TA. Future longitudinal studies should test whether the association of CMV with lower TA contributes to accelerated telomere shortening over time.