Patients presenting at the Juba Teaching Hospital, either with fever of undetermined origin or with a clinical cause of fever, gave evidence of exposure to a wide range of viral and rickettsial agents. Serological tests showed high antibody levels to flaviviruses (56.9%) and alphaviruses (29.2%), with lesser levels of bunyamweraviruses (3.8%), Rift Valley fever (2.3%), and sandfly fever (0.75%). Flavivirus exposure was significantly associated with clinical evidence of liver disease; repeated exposure to flaviviruses was particularly prevalent in those with poor sanitation and who had received previous injections. A significant focus of Ebola and Marburg exposure in Juba has been identified. Clinical evidence of liver disease was evident in 37% of patients studied, and 24.6% were HBsAg positive. The first 2 HIV-positive individuals from the southern Sudan are reported, including one with clinical AIDS. A high prevalence of positive antibodies to Rickettsia typhi in the population indicated that murine typhus was common locally. This study indicates the need for further public health measures in the southern Sudan to control the spread of these infections.