Simplicillium species have a wide host range and an extensive distribution. Some species are associated with rusts, as well as other plant pathogenic fungi and play an important role in biological control. In this study, two specimens of Simplicillium were collected from Chiang Mai Province, Thailand. Simplicillium formicae sp. nov. was isolated from an infected ant and S. lanosoniveum from Ophiocordyceps unilateralis which is a new host record. Species were initially identified using ITS gene sequences and confirmed using morphology coupled with phylogenetic analyses of a combined nrLSU, nrSSU, TEF and RPB1 dataset. Simplicillium formicae differs from other species in the genus by the presence of flask-shaped synnemata and phialides with intercalary nodes. Simplicillium lanosoniveum resembles other collections of the species by its completely solitary, tapering phialides and globose to ellipsoidal conidia which adhere in a slimly head. A key to species of Simplicillium is also provided.