In-Field Capable Loop-Mediated Isothermal Amplification Detection of Spodoptera frugiperda (Lepidoptera: Noctuidae) Larvae Using a Rapid and Simple Crude Extraction Technique

Abstract

Abstract Fall armyworm, Spodoptera frugiperda J.E. Smith (Lepidoptera: Noctuidae), is an economically important pest worldwide and has recently been identified in Australia. Morphological identification of S. frugiperda at early larval stages can be difficult often requiring expert microscopy analysis. Rapid and accurate in-field diagnosis is vital for management decision support and there are no tools currently available for this purpose. In this study, a sensitive, specific, and in-field capable loop-mediated isothermal amplification (LAMP) assay was developed to detect S. frugiperda larvae. A primer set based on a highly conserved region of the S. frugiperda cytochrome oxidase subunit 1 (COX1) gene provided detection within 30 min from both total DNA and crude extractions. The crude extraction technique of crushing 10 mg of S. frugiperda material in 50 µl ddH2O and further diluting the homogenate in ddH2O is rapid, simple, and does not require heat blocks, centrifuges, or special buffers increasing its utility as a field-based technique. The primer set detected as little as 24 pg of S. frugiperda DNA and did not cross-react with any other of the lepidopteran species tested that are easily confused with S. frugiperda in Australia. Therefore, this assay could be used in-field to correctly identify the presence of S. frugiperda and thereby greatly assist with timely management decisions.