Hesperiidae is one of the largest families of butterflies. Our knowledge of the higher systematics on hesperiids from China is still very limited. We infer the phylogenetic relationships of the subfamilies of Chinese skippers based on three mitochondrial genes (cytochrome b (Cytb), the NADH dehydrogenase subunit 1 (ND1) and cytochrome oxidase I (COI)). In this study, 30 species in 23 genera were included in the Bayesian and maximum likelihood analyses. The subfamily Coeliadinae, Eudaminae, Pyrginae and Heteropterinae were recovered as a monophyletic clade with strong support. The subfamily Hesperiinae formed a clade, but support for monophyly was weak. Our results imply that the five subfamilies of Chinese Hesperiidae should be divided into: Coeliadinae, Eudaminae, Pyrginae, Heteropterinae and Hesperiinae. The relationships of the five subfamilies should be as follows: Coeliadinae + (Eudaminae + (Pyrginae + (Heteropterinae + Hesperiinae))).Reconstruction of the phylogenetic relationship of organisms plays an essential role in better understanding their evolution and diversification 1 . Lepidoptera, as the second largest order of insects with more than 157,000 species, are of particular interest in systematic research 2,3 . The skipper butterfly (Hesperiidae) which include around 4000 species is one of the most diverse groups of butterflies 4,5 . Although Hesperidae has been well defined, historically there exists disagreement at the subfamily and tribe levels.The higher-level classification of Hesperiidae was established in the late 19th century. Watson divided Hesperiidae into three subfamilies (Pyrrhopyginae, Hesperiinae and Pamphilinae) based on the morphological characteristics of 201 genera 6 . The family was further arranged into six subfamilies by Evans 7 . Evans placed 130 genera in 4 subfamilies and 13 generic groups (equivalent to the current tribes), which shaped the current system for higher-level classification and interrelationships of the Hesperiidae. Studies on more detailed morphological characteristics have further advanced our knowledge important for classification and construction of phylogenetic relationships of the members in Hesperiidae [8][9][10] . Chou 11,12 proposed three families in Hesperioidea (Euschemonidae, Megathymidae and Hesperiidae) and added three subfamilies (Coediadinae, Pyrginae and Hesperiinae) of Hesperiidae identified in China.With information from molecular systematics studies in the past two decades, Warren et al. 13 proposed the recent classification of Hesperiidae, to include five subfamilies: Coeliadinae, Pyrginae, Heteropterinae, Trapezitinae and Hesperiinae. With combined molecular and morphological data, Warren et al. 14 subsequently revised the classification of Hesperiidae to include seven subfamilies: Coeliadinae, Euschemoninae, Eudaminae, Pyrginae, Heteropterinae, Trapezitinae and Hesperiinae. Warren's molecular phylogeny included approximately 200 genera, representing about 35% of the skipper genera in the world. The skipper butterflies distributed in t...