Dengue virus (DV) primarily infects blood monocytes (MO) and tissue macrophages (Mφ). We have shown in the present study that DV can productively infect primary human MO/Mφ regardless of the stage of cell differentiation. After DV infection, the in vitro-differentiated MO/Mφ secreted multiple innate cytokines and chemokines, including tumor necrosis factor alpha, alpha interferon (IFN-α), interleukin-1β (IL-1β), IL-8, IL-12, MIP-1α, and RANTES but not IL-6, IL-15, or nitric oxide. Secretion of these mediators was highlighted by distinct magnitude, onset, kinetics, duration, and induction potential. A chemokine-to-cytokine hierarchy was noted in the magnitude and induction potential of secretion, and a chemokine-to-cytokine-to-chemokine/Th1 cytokine cascade could be seen in the production kinetics. Furthermore, we found that terminally differentiated MO/Mφ cultured for more than 45 days could support productive DV infection and produce innate cytokines and chemokines, indicating that these mature cells were functionally competent in the context of a viral infection. In addition, DV replication in primary differentiated human MO/Mφ was enhanced and prolonged in the presence of lipopolysaccharide (LPS), and LPS-mediated synergistic production of IFN-α could be seen in DV-infected MO/Mφ. The secretion of innate cytokines and chemokines by differentiated MO/Mφ suggests that regional accumulation of these mediators may occur in various tissues to which DV has disseminated and may thus result in local inflammation. The LPS-mediated enhancement of virus replication and synergistic IFN-α production suggests that concurrent bacterial infection may modulate cytokine-mediated disease progression during DV infection.