Loop-Mediated Isothermal Amplification for the Diagnostic Detection of Meloidogyne chitwoodi and M. fallax

Abstract

Meloidogyne chitwoodi is a root-knot nematode that parasitizes a broad range of plants. In the Pacific Northwest (PNW) of the United States, M. chitwoodi is a major potato pest. The nematodes infect roots and tubers; blemishes caused by the nematodes on the tubers significantly affect potato marketability. M. chitwoodi is a quarantine pathogen by many regulatory agencies, limiting potato trade opportunities when it is present. A loop-mediated isothermal amplification (LAMP) assay was developed to amplify the intergenic spacer (IGS2)-18S region of the ribosomal rDNA of M. chitwoodi. Using the LAMP assay, we could detect the presence of M. chitwoodi from infected Washington State soil samples. The LAMP primers showed specificity for DNA from M. chitwoodi and the closely related species M. fallax. There was no cross reaction of the LAMP primers with DNA from tropical nematodes M. incognita, M. arenaria, and M. javanica, or the Northern root-knot nematode M. hapla. The LAMP assays can be completed within 45 min, and they were 100 times more sensitive in nematode detection than conventional PCR. The LAMP assay will facilitate detection of potato nematodes M. chitwoodi and M. fallax. Knowledge of potato nematodes, particularly M. chitwoodi in PNW soils, will aid management decisions.