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Molecular identification of Anoplophora glabripennis (Coleoptera: Cerambycidae) and detection from frass samples based on real-time quantitative PCR

Affiliation
Plant Health Laboratory, Entomology and Invasive Plants Unit, French Agency for Food, Environmental and Occupational Health & Safety (ANSES), Montferrier-sur-Lez Cedex, France
Taddei, Andrea;
GND
1205678026
Affiliation
Julius Kühn-Institute (JKI), Institute for national and international plant health, Germany
Becker, Matthias;
GND
1175986992
Affiliation
Julius Kühn-Institute (JKI), Institute for national and international plant health, Germany; Julius Kühn-Institute (JKI), Institute of Plant Protection in Field Crops and Grassland, Germany
Berger, Beatrice;
Affiliation
Department of Agriculture, Food and Environment, University of Pisa, Via del Borghetto 80, Pisa, Italy
Da Lio, Daniele;
GND
1238231918
Affiliation
Julius Kühn-Institute (JKI), Institute for national and international plant health, Germany
Feltgen, Stephanie;
GND
143103393
Affiliation
Julius Kühn-Institute (JKI), Institute for national and international plant health, Germany
König, Stephan;
GND
1139391372
Affiliation
Julius Kühn-Institute (JKI), Institute for national and international plant health, Germany
Hoppe, Björn;
Affiliation
Laboratory of Phytopathological Diagnostics and Molecular Biology, Plant Protection Service of Tuscany, Via Ciliegiole 99, Pistoia, Italy
Rizzo, Domenico

Anoplophora glabripennis (Motschulsky 1853) (Coleoptera: Cerambycidae), the Asian Longhorned Beetle, is native to temperate and subtropical areas of China and the Korean peninsula. Due to its wide range of host plants, it is considered among the most economically important invasive plant pests. The morphological identification of A. glabripennis larvae can be confirmed by DNA barcoding, but obtaining the specimens from infested trees can be a demanding and challenging task. Therefore, non-invasive diagnostic tools based on DNA extracted from frass samples can be of key importance in phytosanitary surveys. In this study, an in silico generated real-time quantitative PCR test was developed for the detection of A. glabripennis DNA from frass material, which is naturally extruded from larval tunnels through cracks in the bark. Specificity was confirmed against a wide range of other wood-boring insect species frequently encountered during phytosanitary surveys and inclusivity was demonstrated for different populations of A. glabripennis from all main European outbreak areas. The test proved sensitive and reliable in detecting A. glabripennis DNA extracted from woody frass material of Acer saccharinum and Aesculus hippocastanum at least up to the 100-fold dilution. Furthermore, the test allowed the molecular identification of any life stage of the insect, including eggs and young larvae, whose morphological identification is impossible or very challenging. This study provides a reliable and sensitive molecular tool to detect A. glabripennis DNA in woody frass material, thus allowing a non-invasive sampling approach.

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