A relapsing fever Borrelia and spotted fever Rickettsia in ticks from an Andean valley, central Chile Abstract In humans, emerging infectious diseases are mostly zoonoses with ticks playing an important role as vectors. Tick-borne relapsing fever Borrelia and spotted fever Rickettsia occur in endemic foci along tropical and subtropical regions of the globe. However, both are widely neglected etiologic agents. In this study, we performed molecular analyses in order to assess the presence of Borrelia and Rickettsia DNA in ticks infesting small-mammals within a National Reserve located in the Andes Mountains, central Chile. While hard ticks were negative for the presence of both agents, sequences of four rickettsial (gltA, htrA, ompA, ompB) and two borrelial (16S rRNA and flaB) genes were obtained from larvae of an Ornithodoros sp. morphologically related with Ornithodoros atacamensis. Phylogenetic analyses indicated that the detected Borrelia and Rickettsia spp. belong to the relapsing fever and spotted fever groups, respectively. Moreover, the agents formed monophyletic clades with Rickettsia amblyommatis and “Candidatus Borrelia johnsonii.” As positive ticks parasitize rodents within a highly visited National Reserve where outdoor activities are of common practice, the risk for human parasitism should not be discarded.