<i>Aedes aegypti</i> Mosquito Probing Enhances Dengue Virus Infection of Resident Myeloid Cells in Human Skin

The most prevalent arthropod-borne viruses, including the dengue viruses, are primarily transmitted by infected mosquitoes. However, the dynamics of dengue virus (DENV) infection and dissemination in human skin following <i>Aedes aegypti</i> probing remain poorly understood. We exposed human skin explants to adult female <i>Ae. aegypti</i> mosquitoes following their infection with DENV-2 by intrathoracic injection. Skin explants inoculated with a similar quantity of DENV-2 by a bifurcated needle were used as controls. Quantitative in situ imaging revealed that DENV replication was greatest in keratinocytes in the base of the epidermis, accounting for 50–60% of all infected cells regardless of the route of inoculation. However, DENV inoculation by <i>Ae. aegypti</i> probing resulted in an earlier and increased viral replication in the dermis, infecting twice as many cells at 24 h when compared to needle inoculation. Within the dermis, enhanced replication of DENV by <i>Ae. aegypti</i> infected mosquitoes was mediated by increased local recruitment of skin-resident macrophages, dermal dendritic cells, and epidermal Langerhans cells relative to needle inoculation. An enhanced but less pronounced influx of resident myeloid cells to the site of mosquito probing was also observed in the absence of infection. <i>Ae. aegypti</i> probing also increased recruitment and infection of dermal mast cells. Our findings reveal for the first time that keratinocytes are the primary targets of DENV infection following <i>Ae. aegypti</i> inoculation, even though most of the virus is inoculated into the dermis during probing. The data also show that mosquito probing promotes the local recruitment and infection of skin-resident myeloid cells in the absence of an intact vasculature, indicating that influx of blood-derived neutrophils is not an essential requirement for DENV spread within and out of skin.