Abstract: In order to investigate the resistance level and target-based resistant molecular mechanism of Amaranthus retroflexus L. populations in summer corn fields of Hebei Province to nicosulfuron, whole-plant bioassay experiments were conducted to evaluate the susceptibility of 52 field populations of A. retroflexus collected from Hebei Province between 2021 and 2022. The resistance mechanisms were further investigated through the in vitro acetolactate synthase (ALS) inhibition assay and ALS gene sequence analysis. The results indicated that, within the 52 populations of A. retroflexus in summer corn fields of Hebei Province, 48 (92.3%) exhibited varying degrees of resistance to nicosulfuron, with 67.3% showing high-level resistance (HR), 13.5% demonstrating moderate resistance (MR), and 11.5% displaying low-level resistance (LR). The A. retroflexus populations gathered from Shijiazhuang and Handan both demonstrated high level of resistance. In the three cities of Langfang, Baoding, and Xingtai, high-resistance populations exceeded 60%. The resistant population (BD-R, 49) with the highest resistance level originated from Nanyi Village, Beiyang Town, Boye County, Baoding City, possessed a GR₅₀ value (50% growth reduction )of 338.4 g/hm² and a resistance index (RI) of 715.4. The in vitro ALS activity assay indicated that the IC₅₀ value of nicosulfuron for ALS activity in the BD-R population was 0.3690 mg/L, achieving a resistance index of 31.54. The ALS gene sequence analysis revealed that the tryptophan (Trp) at the position 574 was replaced by leucine (Leu) (Trp574Leu). The above research findings indicated that A. retroflexus in summer corn fields of Hebei Province had generally developed a high level of resistance to nicosulfuron, and the Trp574Leu amino acid mutation in the ALS gene may be one of the primary factors contributing to such resistance.