Abstract:[Objective] To isolate new actinomycetes for discovering compounds of pharmaceutical importance. [Methods] We collected 280 soil samples from virgin forest in Wuling Mountain. In total 1134 actinomycetes were isolated by culture-dependent method. Of them 30 strains were selected and then characterized by phylogenetic analysis based on sequences of 16S rRNA gene. Antimicrobial activities were determined by agar well diffusion method, and genes of typeⅠandⅡpolyketide synthases (PKSⅠ, PKSⅡ), nonribosomal peptide synthase (NRPS) and polyene cytochrome P450 hydroxylase (CYP) were screened by PCR. [Results] Thirty strains belonged to 8 families, 13 genera: Streptomyces (>70%), Micromonospora, Dactylosporangium, Catellatospora, Sphaerisporangium, Streptosporangium, Actinomadura, Nonomuraea, Nocardia, Rhodococcus, Arthrobacter, Microbacterium, and Pseudonocardia. There are 5 novel species candidates. Thirty strains showed different antimicrobial activities against one to three bacteria and pathogenic fungi. [Conclusion] Abundant diversity of actinomycetes existed in the virgin forest of Wuling Mountain which has many new taxa, and Streptomyces is the predominant population. The strains showed high antimicrobial activities and can be further researched for exploiting new compounds of pharmaceutical.