Multiple loop-mediated isothermal amplification assays for the identification of Spodoptera eridania and Spodoptera...

The southern armyworm (SAW) Spodoptera eridania (Stoll) and the yellow-striped armyworm (YSAW) Spodoptera ornithogalli (Guenée) (Lepidoptera: Noctuidae) are designated as regulated quarantine pests in South Korea and quarantine pests in Europe because of their potential to cause serious economic losses when introduced outside their native ranges. In this study, we developed rapid loop-mediated isothermal amplification (LAMP) assays with in-field applicability for the first time to detect SAW and YSAW. To design these assays, we selected 5 and 6 genomic regions specific to SAW and YSAW, respectively, from the whole-genome sequences of the 2 target species, along with those of 12 nontarget species (including 5 Spodoptera species and 7 lepidopteran species that infect similar hosts). We assessed the specificity of these regions to each target species. The developed markers consistently detected SAW and YSAW using 1 antenna, 1 leg, and 1/16 of a thoracic muscle. The detection time was a maximum of 30 min for SAW and 35 min for YSAW, including the crude DNA extraction time. Consistent positive reactions were noted for all markers using crude DNA extracted from the 3 aforementioned tissue types, even when employing a simplified incubation-only method omitting the tissue chopping step. The LAMP reaction mixture, which was pre-mixed to streamline the in-field process, remained stable for up to 20 d at -20 °C. The 5 and 6 LAMP markers developed for SAW and YSAW, respectively, could serve as reliable in-field diagnostic tools individually or in combination to enable cross-verification and increase diagnostic confidence.