Human herpesvirus 6 (HHV-6) frequently reactivates in human immunodeficiency virus 1 (HIV-1) infected patients, and is thought to be a cofactor in AIDS progression. Macrophages are targets and reservoirs of HIV-1 and HHV-6; hence, they have an important role in dissemination and pathogenesis of these viruses. The present study examined the effects of HHV-6 A variant on replication of R5 variants of HIV-1 in macrophages. For this purpose, HIV-1 replication was investigated in macrophages infected with HIV-1 alone or along with HHV-6A. Our results demonstrated that HHV-6A significantly suppressed HIV-1 replication in coinfected cultures. HHV-6A infection resulted in increased secretion of RANTES and IL-8. Experiments with exogenous RANTES and IL-8 revealed that these chemokines also significantly suppressed HIV-1 replication in infected macrophages. RANTES is able to induce desensitization and internalization of CCR5, the chemokine coreceptor of R5 variants. In addition, IL-8 receptor activation results in cross-desensitization and cross-internalization of CCR5. We found that CCR5 sensitivity and expression level is diminished in HHV-6A-infected macrophage cultures compared with uninfected cells. Taken together, our results indicate that HHV-6A infection decreases the susceptibility of macrophages to R5 variants of HIV-1 in which the HHV-6A induced RANTES and IL-8 may have importance.