Nanoclay-based dsRNA delivery: a novel approach to control Globodera rostochiensis

BACKGROUND: Globodera rostochiensis, the golden cyst nematode, is a major pest affecting potato production worldwide. The ams-1 gene, which encodes a secreted amphid-associated protein essential for host recognition, is an ideal target for controlling this pest through RNA interference (RNAi). However, the exogenous application of double-stranded RNA (dsRNA) is challenging due to its low stability under open environmental conditions. METHODS AND RESULTS: This study aimed to reduce the G. rostochiensis infestation in the popular Indian potato cultivar Kufri Jyoti by targeting the ams-1 gene using RNAi technology. The in vitro assay showed a clear dose- and time-dependent reduction in mobility of juveniles (J2s), with the highest concentration (2.5 mg/ml) reducing mobility from 91% at 5 hours to 58% at 10 hours and 34% at 15 hours. Fluorescence microscopy showed fluorescein isothiocyanate (FITC) signals in the dsRNA-treated J2s, confirming dsRNA uptake, while untreated J2s showed no fluorescence and maintained normal mobility. In the plant bioassay, dsRNA-treated J2s showed significantly fewer infection points (33%) compared to untreated J2s (78%), along with marked downregulation of the ams-1 expression. TEM and SEM imaging revealed that kaolinite formed a protective coating around the dsRNA, which is crucial for enhancing its stability under field conditions. The dsRNA-kaolinite formulation reduced nematode infestation by 45% compared to the control. Furthermore, drenching with the dsRNA-kaolinite formulation proved to be most effective in reducing the cyst population, with a notably low reproduction factor (Rf) of 0.5, indicating that kaolinite encapsulation enhances dsRNA stability and efficacy against G. rostochiensis. CONCLUSION: This study demonstrates the effectiveness of RNAi-mediated silencing of the ams-1 gene in G. rostochiensis and, for the first time, reports the use of kaolinite nanoclay for stable dsRNA delivery. The enhanced stability and efficacy of the dsRNA-kaolinite formulation provide an environmentally friendly strategy for the management of G. rostochiensis in potato.