SMALL SCALE POST-RELEASE EVALUATION OF A GONATOCERUS MORRILLI PROGRAM IN CALIFORNIA AGAINST THE GLASSY-WINGED SHARPSHOOTER: UTILITY OF DEVELOPED MOLECULAR DIAGNOSTIC TOOLS Project Leaders

Previously we discovered a cryptic species complex in Gonatocerus morrilli (Howard) and developed molecular diagnostic markers that distinguished the two cryptic species. In the current study we tested the utility of the two developed molecular diagnostic markers to evaluate the establishment of G. morrilli in California. In the two cryptic species, the size of the internal transcribed spacer 2 region (ITS2) varies by about 212 base pairs; the Texas G. morrilli species is associated with a size of about 851-853 base pairs and the California G. morrilli (nov.) species with a size of about 1063-1067 base pairs. Secondly, the two cryptic species do not share any inter-simple sequence-polymerase chain reaction (ISSR-PCR) bands or markers. Initially releases were made from what was thought to be a Mexico culture, but contamination was suspected to have occurred from a Texas culture and therefore, the culture was name 'TX/MX". Post-released collections from years 2002 and 2003 were made from the following locations: San Juan Capistrano, Glen Ivy, Pauma, Temecula, and San Marcos. Amplification of the ITS rDNA fragments demonstrated that all or 100% of the randomly chosen individuals (125 total) were of the California ITS2 genotype and none were of the Texas ITS2 genotype. ISSR-PCR DNA fingerprinting of the TX/MX colony along with native California and Texas G. morrilli species demonstrated that the TX/MX ISSR-PCR banding pattern was superimposable to that of the California G. morrilli (nov.) species. The results demonstrated that the TX/MX colony was contaminated with the California species, indicating that what was being released in California was California's own native species. Therefore, this is why screening with the ITS2 fragment detected only the California ITS2 genotype. The present results confirm the utility of the two developed molecular diagnostic methods in monitoring the success of the G. morrilli biological control program in California. In addition, this molecular technology will allow us to monitor egg parasitoid colonies to eliminate unwanted species.