To monitor aphid parasitism by Aphelinus hordei (Kurdjumov) and Aphidius colemani Viereck, we developed specific ribosomal DNA markers to distinguish them from several other cereal aphid parasitoid species and two important host species, the Russian wheat aphid, Diuraphis noxia (Mordvilko), and the greenbug, Schizaphis graminum (Rondani). Ribosomal DNA sequences for the internal transcribed spacer 2 (ITS2) were first cloned and sequenced from A. hordei, A. albipodus Hayat & Fatima, A. asychis Walker, A. varipes (Foerster), A. colemani, D. noxia, and S. graminum. We designed specific primers based on the ITS2 sequences. Polymerase chain reaction (PCR) amplification of wasp and aphid DNA using these primers, followed by agarose gel electrophoresis, successfully distinguishes A. hordei and A. colemani from all three other Aphelinus species and two aphid species. A 411-bp nucleotide fragment and a 571-bp fragment were amplified only from A. hordei and from A. colemani, respectively, and no such fragments were amplified from any other wasp species or aphids. DNA could be detected at a level as low as 10⁻³ adult wasp equivalent for A. hordei and 5 × 10⁻⁴ adult wasp equivalent for A. colemani. The DNA of both species was detectable in parasitized D. noxia 24 h after initial contact with adult parasitoid pairs.