Early and on-site detection of Ganoderma-induced basal stem rot in oil palm: using recombinase polymerase amplification-lateral flow assay.

Abstract

Basal stem rot disease, caused by Ganoderma spp., is the major economic menace of oil palm cultivation. The disease begins with an asymptomatic phase, progresses to ambiguous foliar abnormalities, and culminates in stress malign fructification stage, the only conclusive visual evidence of infection. At this stage, the pathogen is well-established and resistant to curative measures, highlighting the critical need for early detection. The current study deployed recombinase polymerase amplification-lateral flow assay (RPA-LFA) for onsite and early detection of Ganoderma at the asymptomatic phase. The RPA reaction conditions were standardised with respect to the concentration of magnesium acetate and betaine concentration, incubation temperature as well as time. The assay was validated by analysing pure fungal DNA, pure plant DNA, and crude DNA extracted from palms showing varying degrees of disease severity collected from diverse sampling sources, including soil, stem, and roots. The detection system could detect Ganoderma with crude DNA extracted from asymptomatic palm roots. The method was highly sensitive, detecting as little as 10 pg/µL of Ganoderma DNA at 41 °C for 30 min. This assay, highly specific to Ganoderma, was validated across ten different species of Ganoderma. Further, there was no cross-reaction with ten other oil palm-related microbes/pathogens. To our knowledge, this is the first report to establish an RPA-LFA for the detection of Ganoderma-induced rots. The kit enables rapid and early detection of BSR samples at the point of care, at the asymptomatic stage, and is supportive of prompt and effective management strategies.