Abstract Development of a cytomegalovirus (CMV) vaccine is a major public health priority due to the risk of congenital infection. Clinical strains of human CMV (HCMV) differ from lab adapted virus as the clinical strains express a pentameric complex (PC) and retain the ability to infect epithelial and endothelial cells unlike lab strains which are limited to fibroblast cells. The mechanism of virus entry into epi/endothelial cells is different from the gB pathway of entry into fibroblast cells as it requires viral proteins gH/gL/UL128-131 forming an endocytic complex (PC) to enable viral cell entry. Importantly, transplacental CMV infection is highly dependent upon viral epithelial/endothelial tropism since the virus has to infect endothelial cells of blood vessels (maternal and fetal) as well as a layer of epithelial trophoplast cells that constitute part of the placenta. Neutralizing antibodies to the PC are potentially more important than antibodies to the immunodominant gB glycoprotein, since anti- PC antibodies are more effective in neutralizing virus on epi/endothelial cells compared to gB. Importantly, in convalescent patients high anti-PC titers are thought to be effective in preventing congenital CMV. Human CMV cannot be directly studied in an animal model because of species specificity. The guinea pig is the only small animal model that allows the study of congenital CMV but requires the use of species specific guinea pig CMV (GPCMV). In the guinea pig model for congenital CMV, we have defined a homolog pentameric complex as necessary for epithelial tropism, virus dissemination and congenital infection by GPCMV. Furthermore, the GPCMV homolog PC is highly immunogenic and generates antibodies that neutralize virus infection on epithelial cells. This pre-clinical translational study seeks to develop GPCMV PC specific vaccine strategies (either subunit or defective adenovirus vector based) to determine if a PC or PC subcomplex vaccine strategy can prevent congenital CMV in the guinea pig model. Importantly, as part of these studies we will evaluate the ability of PC vaccines to cross protect against congenital CMV from different GPCMV strains. Additionally, the synergy effect of gB and PC vaccine strategies will be evaluated. Finally, since in HCMV both gB and gH induce a CD4 T cell response, the protective impact of the T cell response will be evaluated.