Bispyribac-sodium which is an acetolactate synthase inhibiting herbicide is a frequently used herbicide for the control of weeds in rice fields of India including the most common noxious weed barnyardgrass. However, recently rice growers reported reduced control of barnyardgrass with bispyribac-sodium. Hence, a large-scale survey was carried out and collected 37 putative barnyardgrass biotypes for bispyribac-sodium resistance in two rice growing states Chhattisgarh and Kerala. Open field pot experiments were conducted for two years to confirm the resistance in those biotypes. Of the tested 37 biotypes, 30% of the biotypes (11) survived recommended label rate of bispyribac-sodium (25 g ai ha-1). The effective rate of bispyribac-sodium required to achieve 50% control (ED50) of putative resistant biotypes ranged from 18 to 41 g ha-1, whereas it was about 10 g ha-1 in susceptible biotypes. This suggests that putative biotypes were 2 to 4 times resistant to bispyribac-sodium. At 6 days after herbicide application, in-vitro enzyme assay showed higher ALS enzyme activity in putative resistant biotypes (66 to 75%) compared to susceptible biotypes (48 to 52%). This indicates insensitive ALS enzyme to bispyribac-sodium in those biotypes and target site mutation as possible mechanism for resistance. Whole-plant bioassay also suggested that the resistance problem is more widespread in Chhattisgarh than in Kerala state. This study confirmed the first case of evolved resistance in barnyardgrass to bispyribac-sodium in rice fields of India.