Development of a loop-mediated isothermal amplification (LAMP) assay for the identification of the invasive wood borer Aromia bungii (Coleoptera: Cerambycidae) from frass

The red-necked longhorn beetle Aromia bungii (Faldermann, 1835) (Coleoptera: Cerambycidae) is native to east Asia, wherit is a major pest of cultivated and ornamental species of the genus Prunus. Morphological or molecular discrimination ofadults or larval specimens is required to identify this invasive wood borer. However, recovering larval stages of the pest from trunks and branches causes extensive damage to plants and is timewasting. An alternative approach consists in applying noninvasive molecular diagnostic tools to biological traces (i.e., fecal pellets, frass). In this way, infestations in host plants canbe detected without destructive methods. This paper presents a protocol based on both real-time and visual loop-mediatedisothermal amplification (LAMP), using DNA of A. bungii extracted from fecal particles in larval frass. Laboratory validations demonstrated the robustness of the protocols adopted and their reliability was confirmed performing an inter-lab blindpanel. The LAMP assay and the qPCR SYBR Green method using the F3/B3 LAMP external primers were equally sensitive,and both were more sensitive than the conventional PCR (sensitivity > 10 to the same starting matrix). The visual LAMPprotocol, due to the relatively easy performance of the method, could be a useful tool to apply in rapid monitoring of Abungii and in the management of its outbreaks. 3