Non-tuberculous mycobacteria (NTM) are ubiquitous microorganisms that have the potential to cause disease in both humans and animals. Recently, NTM infections have rapidly increased in South Korea, especially in urbanized areas. However, the distribution of species and the antibiotic resistance profile of NTM in environmental sources have not yet been investigated. Therefore, we analyzed the distribution of species and the antibiotic resistance profile of NTM in soil within urban areas of South Korea. A total of 132 isolates of NTM were isolated from soil samples from 1 municipal animal shelter and 4 urban area parks. Among the 132 isolates, 105 isolates were identified as slowly growing mycobacteria (SGM) and 27 isolates as rapidly growing mycobacteria (RGM) based on the sequences of the rpoB and hsp65 genes. The antibiotic resistance patterns of NTM isolates differed from species to species. Additionally, a mutation in the rrs gene found in this study was not associated with aminoglycoside resistance. In conclusion, our results showed that NTM isolates from South Korean soil exhibit multidrug resistance to streptomycin, amikacin, azithromycin, ethambutol, isoniazid, and imipenem. These results suggest that NTM may pose a public threat.