Objective To investigate the enterovirus ( EV)-carrying status and the circulating se-rotypes in healthy children from inner and border areas of Yunnan Province in 2014 and 2015 and to analyze the genetic characteristics of echovirus 6 (ECHO6), ECHO25 and ECHO11 strains. Methods Stool sam-ples were collected from children less than 15 years old living in 6 to 7 counties of 3 inner prefectures/cities and 8 to 9 counties of border prefectures/cities. Altogether 921 samples were collected including 453 sam-ples in 2014 (213 samples in inner counties and 240 samples in border counties) and 468 samples in 2015 (195 samples in inner counties and 273 samples in border counties). Viruses were isolated from the stool samples and their serotypes were identified by gene sequencing. Results The numbers of EV strains isola-ted from the samples collected in inner counties and border counties in 2014 were 20 ( isolating rate:9.39%, 20/213) and 16 (isolating rate: 6. 67%, 16/240), respectively. The overall isolating rate for 2014 was 7. 95% (36/453). The predominant species was enterovirus B, accounting for 88. 89% of all iso-lated strains (32/36), followed by enterovirus A species (11. 11%, 4/36). No strains of enterovirus spe-cies C (including poliovirus) and D was detected in 2014. In total, 46 EV strains were isolated in 2015 with an overall isolating rate of 9. 83% (46/468), including 13 strains in inner counties (isolating rate:6. 67%, 13/195) and 33 strains in border counties (isolating rate:12. 09%, 33/273). Most of the strains were enterovirus B species, accounting for 78. 26% (36/46), followed by enterovirus C species (19. 57%, 9/46) and enterovirus A species (2. 17%, 1/46). Altogether 82 EV strains were isolated in 2014 and 2015 with an isolating rate of 8. 90% (82/921), of which 33 strains were isolated in inner counties (8. 09%, 33/408) and 49 strains were isolated in border counties (9. 55%, 49/513). Among the 82 EV strains, 9 strains were polioviruses (0. 98%, 9/921) and all of them were Sabin-like polioviruses. The rest of the strains were non-polio enterovirus (7. 93%, 73/921). Conclusion In 2014, the EV isolating rate in inner counties (9. 39%) was higher than that in border counties (6. 67%). However, the EV isolating rate in border counties (12. 09%) was higher than that in inner counties (6. 67%) in 2015. Enterovirus B was the predominant species in both 2014 and 2015. No wild type polioviruses and enterovirus D species were detec-ted. Polio-free status was maintained well in Yunnan Province.