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The genus Opisthopappus consists of two species, O. longilobus and O. taihangensis. Both of them are important economic and ornamental plants but endangered in China. In this study, genetic variation in four populations of O. longilobus and nine populations of O. taihangensis was estimated by chloroplast microsatellites (cpSSR). Low genetic diversity was detected in O. longilobus and O. taihangensis populations. The percentages of polymorphic loci (PPL), Nei’s gene diversities ( h), and Shannon’s indices ( I) was 53.56 %, 0.074, and 0.139, respectively, for O. longilobus, while PPL = 44.44 %, h = 0.018, I = 0.048 for O. taihangensis. Thirteen chloroplast haplotypes were identified from 153 individuals analyzed of two species. Ten haplotypes were found in O. longilobus, while seven in O. taihangensis. Haplotype 1 was the common and ancestral one. The level of haplotype diversity was also low (Hd O. longilobus = 0.4875, Hd O. taihangensis = 0.3452). AMOVA analysis revealed that the majority of the genetic differences was partitioned within populations of O. longilobus and O. taihangensis, as confirmed by PCoA and cluster analysis. Inter-population genetic distances among population of O. longilobus and O. taihangensis correlated insignificantly with geographic distances ( r = 0.286, P = 0.128 for O. longilobus vs. r = 0.356, P = 0.058 for O. taihangensis). The self-compatibility, effective gene flow, narrow geographical distribution, and historical factors may be the main factors causing differentiation in the genetic structure of O. longilobus and O. taihangensis populations. [ABSTRACT FROM AUTHOR]