UNLABELLED: Soybean yellow mottle mosaic virus (SYMMV) is a newly emerging seed borne plant virus responsible for mosaic and mottling disease in mungbean and soybean. The present study aimed to evaluate the response of nine widely cultivated mungbean and 15 newly developed soybean genotypes to SYMMV-Mb infection under controlled environment conditions using agro-inoculation, followed by serological and molecular detection. The genotypes exhibited differential local symptoms at 13-17 dpi and systemic symptoms at 21-29 days post-inoculation (dpi), ranging from mild chlorosis to severe necrotic local spots with a PDI of 60-95% in mungbean and 30-50% in soybean genotypes. Among the mungbean genotypes, Sona Moong was found to be moderately susceptible (MS), six were susceptible and Pusa Vishal and SML-668 were highly susceptible. Among the soybean genotypes, nine were found to be immune, four were moderately resistant and two were MS. DAC-ELISA and RT-PCR confirmed the presence of SYMMV-Mb in all infected genotypes with higher absorbance values and amplification of the 1064 bp coat protein (CP) region. RT-qPCR analysis of whole seeds harvested from agro-inoculated plants revealed virus accumulation only in mungbean genotypes ranging from 6.33 × 10 SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-025-04347-w.