The present study was conducted to investigate the presence of Arenaviruses from rodents, shrews and elephant shrews captured in selected wildlife-human interfaces in Tanzania. The study involved six sites with high potential for contact between wildlife and humans namely; Ruaha, Kilombero, Mtwara, Mbeya, Mbinga and Mikumi. A total of 121 animals comprising 111 rodents, 3 shrews and 7 elephant shrews were screened for Arenaviruses using conventional Polymerase Chain Reaction (PCR). The genetic relatedness of Arenaviruses was evaluated by conducting phylogenetic analysis of partial sequences of the S gene. The association between age and sex with the presence of Arenaviruses was assessed. Of the 121 animals, 7 (5.8 %) were shedding Arenavirus. All positive animal samples were obtained from the Ruaha site at crop raiding and peridomestic interfaces. Eighty six percent of the infected animals were Mastomys sp. and 14 % were Arvicanthis sp. Age and sex of the animals were not significantly associated with occurrence of Arenaviruses in rodents, shrews and elephant shrews (P > 0.05). Additionally, Arenavirus detection in fecal specimens was not significantly different from the detection in oropharyngeal specimens (P > 0.05), clearly indicating that both specimens are useful for Arenavirus testing. Phylogenetic analysis showed that isolates obtained from this study were related to the Old World Arenaviruses, and include strains of Morogoro virus and a novel strain of Arenavirus. In conclusion, the present study has confirmed the presence of Arenaviruses closely related to other known Old World Arenaviruses in the Ruaha ecosystem.Southern African Center for Infectious Disease Surveillance (SACIDS)