Molecular Identification of an Invasive Wood-Boring Insect Lyctus brunneus (Coleoptera: Bostrichidae: Lyctinae) Using Frass by Loop-Mediated Isothermal Amplification and Nested PCR Assays.

Abstract

Lyctus brunneus (Stephens) is one of the most destructive and worldwide invasive pests of seasoned woods for wooden products. This and other pest Lyctus species have had their distribution expanded by international and domestic human transportation of infested wood and wood products. Rapid detection and accurate identification of Lyctus species are effective tools for helping to eradicate them in new introduction sites. The accurate species-level identification of adults requires expert knowledge about their morphology. However, it takes much time and effort to recover suitable adult specimens because they are borers inside wood. Frass of Lyctus species can easily be detected and recovered in and around infested wood. Thus, frass was tested to see if it was a suitable sample to allow development of a rapid and technically easy molecular detection and identification method for L.brunneus. Species-specific primers were designed from the cytochrome c oxidase subunit I region of L.brunneus and used in development and testing of methods for successfully identifying them from their frass using the loop-mediated isothermal amplification (LAMP) or species-specific nested polymerase chain reaction (PCR) assays. The LAMP assay was faster and more sensitive for detecting the presence of DNA derived from L.brunneus in their frass than the nested PCR assay. These methodologies will be applicable for the rapid detection and identification of other wood-boring invasive pests in regulatory applications.