Circulating monocytes differentiate into tissue macrophages and dendritic cells that provide immune defense against microbial pathogens. Monocytes in the bloodstream consist of subpopulations with distinct differentiation potentials and trafficking patterns. Reconstitution of circulating monocyte subsets varies between patients following allogeneic hematopoietic stem cell transplantation (allo-HSCT) and we postulate that this has implications for susceptibility to infection. The goal of our studies is to characterize the contribution of distinct monocyte subsets to defense against Aspergillus fumigatus infection in a murine model of fungal infection and ip humans following allogeneic hematopoietic stem cell transplantation (allo- HSCT). Our first aim is to characterize circulating and tissue inflammatory monocytes in mice following allo- HSCT. The contribution of MCP-1to CCR2-mediated monocyte repopulation of tissues will be investigated following transplantation and the impact of GVHD on this process will be determined. We will determine the susceptibility of transplanted mice to pulmonary infection with Aspergillus fumigatus and correlate susceptibility with the presence and activation status of inflammatory monocytes. Our second aim is to characterize the ability of inflammatory monocytes to mediate protection against invasive fungal infection in the allo-HSCT setting. We will generate murine monocyte subsets in vitro from bone marrow precursors and investigate their ability to mediate protection against Aspergillus fumigatus infections. Recombinant chemokines and growth factors will be administered to transplanted mice to accelerate and enhance monocyte reconstitution. The impact of innate immune receptor mediated signaling on monocyte mediated antifungal defense in the allo-transplant setting will be investigated. Our third aim is to prospectively investigate monocyte subset frequencies and function in allo-HSCT recipients and correlate these with development of fungal infections. In addition, we will investigate bone marrow and peripheral blood stem cell allografts for the presence of monocyte subsets. Monocyteswill be isolated from grafts and from allo-HSCT patients and their ability to inactivate A. fumigatus conidia and hyphae will be investigated. Relevance: The results of these studies may provide a foundation for future clinical protocols to investigate the infusion of differentiated monocytes into allogeneic stem cell transplant patients to enhance resistance to fungal pathogens..