Borrelia miyamotoi is a tick-borne zoonotic agent that causes hard tick-borne relapsing fever, an emerging disease in humans. Some small mammalian and bird species are reported to be reservoirs of B. miyamotoi. This study aims to examine Borrelia species present in rodents captured from rural areas of Turkey. Blood samples of rodents were initially screened with Borrelia 16S rRNA qPCR. The Borrelia flaB gene was subsequently amplified by conventional PCR, after which all positive samples were sequenced. Borrelia miyamotoi was observed in nine out of 536 blood samples (1.7%) collected from wild rodents. Phylogenetic analysis showed that all positive samples belonged to the European genotype clade of B. miyamotoi. PCR positivity was 5.3%, 3.7%, and 1.8% in Apodemus uralensis, Apodemus flavicollis, and Myodes glareolus, respectively. Borrelia burgdorferi sensu lato that causes Lyme borreliosis in humans could not be detected in the rodents. In this study, presence of B. miyamotoi DNA is reported for the first time in rodents in Turkey.