Rapid identification of Ips sexdentatus (Boerner, 1766) (Curculionidae) from adults and frass with real-time PCR based on probe technology

Abstract

A molecular tool has been developed for the molecular identification of Ips sexdentatus (Börner 1776) (Coleoptera Curculionidae Scolytidae), the well-known six thooted bark beetle, widely distributed in Eurasia, where it infests several species of the genus Pinus and occasionally a few conifer species of the genera Abies, Larix and Picea. The developed test can be useful both in countries where I. sexdentatus is handled as a quarantine species and, to greater reason, in Europe to discriminate biological traces of this commonly found beetle from those produced by regulated pests. The protocol is based on real-time PCR with TaqMan probe technology and has been developed on whole insect bodies (adults) as well as on artificial frass contaminated by DNA of the beetle. The molecular test developed here for both direct and indirect identification of I. sexdentatus has proven effective in terms of analytical specificity, analytical sensitivity, reliability and reproducibility. The recommended protocol is a practical diagnostic tool allowing a rapid identification of the six toothed bark beetle in the presence of any biological trace of other xylophagous pests collected at points of entry during phytosanitary surveys.