Hemorrhagic fever with renal syndrome caused by hantaviruses has long been a serious public health issue in Yunnan Province. Hantaviruses exhibit a high extent of biodiversity in their natural hosts, particularly in mammalian hosts. This study was conducted to screen for hantaviruses in bats and rodents in Yunnan Province and elucidate their genetic characteristics and possible zoonotic disease risk. Hantaviruses were detected in 202 bats and 372 rodents with the positive rates 27.49% and 1.25% respectively. A novel lineage (named Lineage 10) of the Seoul virus (SEOV) from rodents and the geographic clustering of hantavirus in bats were identified using phylogenetic analyses of the full-length M- and S-segments. Our study suggest a high cross-species transmissibility of hantaviruses in bats and existence of a new lineage of SEOV in rodents differing significantly from other SEOVs. These results provide data to support the prevention and control of hantavirus-associated diseases in Yunnan Province. Hantavirus is an important pathogen that can cause hemorrhagic fever with renal syndrome (HFRS) and hantavirus cardiopulmonary syndrome (HCPS). HFRS has long been a serious public health problem in Yunnan Province. However, the molecular epidemiology and host characteristics of hantavirus in Yunnan Province are still unclear. Therefore, we screened hantaviruses in bats and rodents in selected areas of Yunnan from 2018 to 2022 and found the 10th SEOV lineage in rodents, as well as new geographic clusters of Xuan son virus (XSV) and novel hantavirus in bats. We proposed the possibility of the 10th SEOV lineage in rodents as an ancestral lineage and explained the related host-switching events in bat hantaviruses. These results provide data to reveal a greater risk of zoonotic disease and support the prevention and control of hantavirus-associated diseases and in Yunnan Province.