As a genus within the Clavicipitaceae, Metarhizium exhibits rich morphological and ecological diversity, with a wide distribution and a variety of hosts. Currently, sixty-eight species of Metarhizium have been described. A new species of Metarhizium, M. puerense (Hong Yu bis), was described in Pu'er City, Yunnan Province, south-western China. Based on morphological characteristics and multilocus phylogenetic analyses, Metarhizium puerense was confirmed to be phylogenetically related to M. album, but was clearly separated and formed a distinct branch. In contrast, the host of Metarhizium album was plants and leafhoppers and that lepidopteran larvae were the host of M. puerense. The diagnostic features of M. puerense were solitary to multiple stromata and smooth-walled, cylindrical with rounded apices conidia.