Evaluation of Portable DNA-Based Technologies for Identification of Ralstonia solanacearum Race 3 Biovar 2 in the Field

Abstract

Abstract. We have previously reported several new technologies for selective and sensitive detection of quarantine race 3 biovar 2 strains of the bacterial wilt pathogen Ralstonia solanacearum, including new highly specific PCR and LAMP primers, immunomagnetic separation (IMS) for improved isolation of the pathogen from contaminated materials, and two handheld platforms to implement the LAMP-based assay in the field. Here we report on field testing of these new technologies in a naturally infested potato field in the highlands of Guatemala and compare the results to those obtained by standard reference methods, including plating on SMSA media, use of immunostrips, and quantitative PCR. The new LAMP-based assay was demonstrated to be equivalent to quantitative PCR, as sample classifications using the respective methods were absolutely consistent when directly assaying soil and plant tissue extracts. Isolation of DNA from the extracts using IMS enabled more sensitive detection in samples that otherwise appeared negative. These results suggest that the new LAMP tools are indeed viable approaches for rapid, selective, and sensitive field-based detection of race 3 biovar 2 strains of Ralstonia solanacearum, and that IMS is a useful tool for enhancing the isolation and detection of target pathogens in the field.