Background: Recent works identified a novel healthcare problem in Africa: acute Rickettsia felis-induced fever that mimics malaria. The epidemiology of these diseases remains to be characterized. Indeed, the natural reservoirs of R. felis are not known. Since the first clinical descriptions of R. felis-associated fever, the cat and dog fleas Ctenocephalides felis and C. canis were implicated as themost probable vectors inmany countries, but not in Senegal. Methods & Materials: Arthropods (ticks, mites, fleas, tsetse flies, biting midges, mosquitoes, bed bugs) and environmental samples (dust and water samples) were collected in the villages of Dielmo and Ndiop where R. felis-associated acute fevers contribute up to 8% of all acute fevers with the incidence of 6.7 per 100 person-years. DNA extracted from all samples was tested by Rickettsia genus-specific qPCR and by two different R. felis-specific qPCR. Positive environmental samples were subjected to PCR with the universal eukaryotic cytochrome oxydase I-based primers followedbycloningof theampliconswithpGemvectors. Isolationwas performed in XTC cell line using the shell-vial technique. Characterization of the isolated strainswas done by sequencing of gltA and 16S rRNA genes. Results: Arthropods (ticks, mites, fleas, tsetse flies, biting midges, mosquitoes, bed bugs) and environmental samples (dust and water samples) were collected in the villages of Dielmo and Ndiop where R. felis-associated acute fevers contribute up to 8% of all acute fevers with the incidence of 6.7 per 100 person-years. DNA extracted from all samples was tested by Rickettsia genusspecific qPCR and by two different R. felis-specific qPCR. Positive environmental samples were subjected to PCR with the universal eukaryotic cytochrome oxydase I (COI)-based primers followed by cloning of the amplicons with pGem vectors. Isolation was performed in XTC cell line using the shell-vial technique. Characterization of the isolated strainswas done by sequencing of gltA and 16S rRNA genes. Conclusion: This is the first evidence of the presence of R. felis in arthropods in the endemic focus or R. felis infection in Senegal. The roles of Siteroptidae mites in the hosting and transmission of the R. felis is to be further investigated