Two closely-related endoparasitoids of Cotesia plutellae and C. glomerata parasitize the diamondback moth, Plutella xylostella. The parasitized hosts by either parasitoid species exhibited the extended larval period and died without further metamorphosis to pupal stage. However, two parasitoid species exhibited significantly different parasitic capacity and developmental rate, in which C. plutellae showed higher parasitism and faster development in the parasitized P. xylostella. To discriminate these two similar species, morphological and molecular differences were analyzed. Three dichotomous morphological characters including antennal flagellum, hind-leg femur, and terminal abdominal terga were determined. Based on the presumptive polydnaviral particles found in the ovarian calyx of C. glomerata, three genes similar to C. plutellae bracoviral genes were cloned in the C. glomerata genome and compared in their cDNA and the deduced amino acid sequences. Several polymorphic sites were detected to be applicable to design molecular markers to discriminate these two species.