The first cases of gynandromorphy in the non-biting midge genus Polypedilum Kieffer, 1912 (Diptera: Chironomidae) are reported. Gynandromorphs of two species, P. (Tripodura) unifascium (Tokunaga, 1938) and P. (Uresipedilum) bingoparadoxum Kawai, Inoue & Imabayashi, 1998, from China are described and illustrated. They were determined through comparison of mitochondrial cytochrome c oxidase subunit 1 DNA barcodes with those of determined Polypedilum adult males. Occurrence of gynandromorphs, which have a feminine head, thorax and abdomen, but a masculine hypopygium, in the Chironomidae is discussed.