Efforts to identify wildlife reservoirs for tick-borne pathogens are frequently limited by poor understanding of tick - host interactions and potentially transient infectivity of hosts under natural conditions. To identify reservoir hosts for lone star tick (Amblyomma americanum)-associated pathogens, we used a novel technology. In field-collected ticks, we used PCR to amplify a portion of the 18S rRNA gene in remnant blood meal DNA. Reverse line blot hybridization with host-specific probes was then used to subsequently detect and identify amplified DNA. Although several other taxa of wildlife hosts contribute to tick infection rates, our results confirm that the white-tailed deer (Odocoileus virginianus) is a reservoir host for several A. americanum - associated pathogens. Identification of host blood meal frequency and reservoir competence can help in determining human infection rates caused by these pathogens.