High-throughput Root Dip Assay for Rapid Pathogenicity Testing of Ralstonia solanacearum in Tomato Seedlings

Abstract

Ralstonia solanacearum, a devastating vascular pathogen of tomato, causes bacterial wilt leading to significant crop losses globally. Rapid and reliable screening methods are essential to assess the pathogenic potential of different isolates for resistance breeding and epidemiological studies. This study presents a high-throughput root dip inoculation assay to evaluate the virulence of R. solanacearum strain RsT-7 in tomato (cv. Sahoo and Aaryaman). Six distinct treatments involving bacterial broth, sterile water and bacterial suspension (OD₆₀₀ = 0.6) were compared in 6–7-day-old seedlings. Among all treatments, direct root exposure to bacterial suspension (S‑1) resulted in 100% wilting within 48 hours post-inoculation, exhibiting rapid and consistent symptom development including turgor loss, drooping and seedling collapse. Other treatments involving broth or delayed suspension addition showed significantly lower wilt incidence (0–80%). The assay was successfully adapted to 30-day-old seedlings, which developed progressive wilting and stem collapse. Pathogen colonization was confirmed via ooze test and decreased lignification in vascular tissues was visualized using phloroglucinol staining. The results indicate that direct root dip in bacterial suspension is a reliable, reproducible and resource-efficient method for pathogenicity screening of R. solanacearum in tomato. This approach enables rapid disease assessment in early growth stages and can significantly accelerate screening pipelines in plant pathology research.