Subsets of CD16-positive monocytes produce proinflammatory cytokines and expand during chronic infection with the human immunodeficiency virus type 1 (HIV). HIV-infected macrophage in tissues may be long lived and contribute to the establishment and maintenance of the HIV reservoir. We found that the (intermediate) CD14(++)CD16(+) and (nonclassical) CD14(+)CD16(++) monocyte subsets are significantly expanded during infection of Rhesus macaques with pathogenic SIV(mac251) but not during infection of sooty mangabeys with the nonpathogenic isolate SIVSM. In vitro glucocorticoid (GC) treatment of peripheral blood mononuclear cells (PBMCs) from uninfected or SIV(mac251)-infected Rhesus macaques and HIV-infected patients treated or not with antiretroviral therapy (ART) resulted in a significant decrease in the frequency of both CD16-positive monocyte subsets. Short-term in vivo treatment with high doses of GC of chronically SIV(mac251)-infected macaques resulted in a significant decrease in the CD14(+)CD16(++) population and, to a lesser extent, in the CD14(++)CD16(+) monocytes, as well as a significant decrease in the number of macrophages in tissues. Surprisingly, treatment of SIV(mac251)-infected macaques with ART significantly increased the CD14(++)CD16(+) population and the addition of GC resulted in a significant decrease in only the CD14(+)CD16(++) subset. No difference in SIV DNA levels in blood, lymph nodes, gut, and spleen was found between the groups treated with ART or ART plus GC. Thus, it appears that high doses of GC treatment in the absence of ART could affect both CD16-positive populations in vivo. Whether the efficacy of this treatment at higher doses to decrease virus levels outweighs its risks remains to be determined.