Molecular markers for Peristenus spp. (Hymenoptera: Braconidae) parasitoids associated with Lygus spp. (Hemiptera: Miridae)

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Abstract
Molecular markers for identifying Peristenus spp. parasitoids to species level and preliminary molecular markers to distinguish two groups of Lygus spp. common to the Canadian prairies were developed. Peristenus species-specific polymerase chain reaction (PCR) primers were developed based on DNA sequence data from a 1600-bp region of the internal transcribed spacer region between the 5.8S and 18S nuclear rRNA genes (ITS2). These primers were able to distinguish Peristenus digoneutis Loan, Peristenus stygicus Loan, and Peristenus pallipes (Curtis). Their ability to identify to species-level parasites dissected from field-collected Lygus spp. nymphs was examined by analysis of DNA from 100 parasite samples. Of those samples showing positive PCR amplification with both control (ITS2) and species-specific primers, all were positive for P. pallipes; none of the samples amplified appropriately sized products with P. digoneutis specific or P. stygicus specific primers. These findings were validated using restriction enzyme digests of amplified regions of the Peristenus spp. cytochrome oxidase 1 gene. Both methods were consistent with earlier studies that showed P. pallipes to be the only species of the genus Peristenus to be associated with Lygus spp. on the Canadian prairies. PCR primers based on DNA sequence data from a 550-bp region of the mitochondrial 16S rRNA gene were designed to discriminate Lyguslineolaris (Palisot de Beauvois) from Lygus borealis (Kelton), and Lygus elisus (Van Duzee). These PCR primers were used to identify field-collected nymphs, with most being identified as either L. borealis/L. elisus (72–82%) orL. lineolaris (14–18%). These estimates of species composition closely reflected those of subsequent adult population surveys from the same fields.