| Summary: | Lyme disease is a tick-borne zoonotic disease caused by <i>Borrelia burgdorferi</i> sensu lato (s. l.) via transmission cycles involving competent tick vectors and vertebrate reservoirs. Here, we determined the prevalence and distribution of <i>Borrelia</i> genospecies in 738 ticks of at least three species from wild rodents in nine regions of the Republic of Korea (ROK). Ticks were analyzed using nested PCR targeting partial flagellin B gene sequences, followed by sequence analysis. The prevalence of <i>Borrelia</i> infection was 33.6%, and the most common genospecies were <i>B. afzelii</i> (62.5%), <i>B. valaisiana</i> (31.9%), <i>B. yangtzensis</i> (2.4%), <i>B. garinii</i> (1.6%), and <i>B. tanukii</i> (1.6%). <i>Borrelia afzelii</i> was found in all regions except Jeju Island; this predominant genospecies was found in the northern and central sampling regions. <i>Borrelia valaisiana</i>, <i>B. yangtzensis</i>, and <i>B. tanukii</i> were found only in the southern regions with <i>B. valaisiana</i> being the most common, whereas <i>B. yangtzensis</i> and <i>B. tanukii</i> were only found on Jeju Island. Our study is the first to describe the nationwide prevalence of <i>B. burgdorferi</i> s. l. in ticks from wild rodents in the ROK. Continuous surveillance in ticks, animals, humans, and different regions is required to avoid disease distribution and possible transmission to humans in the ROK.
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