The health of the tributary rivers in Chongqing profoundly influences the ecological and environmental security of the Three Gorges Reservoir Area and the Yangtze River watershed. In April to May of 2024 and 2025, benthic macroinvertebrate in 243 evaluated reaches of 46 tributary rivers in Chongqing was sampled to assess river health using Benthic Index of Biotic Integrity (B-IBI). The macroinvertebrate Integrity in different regions was compared and the impacts of environmental variables on B-IBI as well as the mechanism behind was explored. The results indicated that the B-IBI for the tributary rivers was composed of four key indexes: total richness at sampling sites, relative abundance of EPT (Ephemeroptera, Plecoptera, Trichoptera) species, Hilsenhoff biotic index, and relative abundance of dominant taxon. The overall health status of the 46 tributary rivers was dominated by fair (43.5%) and good (36.9%) condition. At the reach scale, the proportion of reaches in the poor condition was the highest (27.6%), followed by those in the excellent (23.1%) and fair (18.5%) condition. B-IBI values exhibited a consistent spatial pattern at both river and reach scales, with the highest values observed in Northeast Chongqing, followed by Southeast Chongqing and then the main urban area. Habitat Quality Assessment (HQA) scores indicated a significant linear positive correlation with B-IBI (P<0.05). Significant differences in B-IBI were also found among reaches with different HQA grades (P<0.05). Specifically, the average B-IBI scores of reaches with excellent and good HQA grades corresponded to good and fair condition categories, respectively, while those of reaches with other HQA grades fell into the poor condition category. B-IBI values of near-natural reaches were mostly classified as excellent, whereas those of reservoir reaches were significantly lower than for reaches with other types of hydrological disturbance (P<0.05). Shapley additive explanations (SHAP) analysis indicated that hydrological disturbance level was the dominant environmental factor influencing the B-IBI of tributary rivers in Chongqing, followed by HQA, sulfate, and chloride. These environmental factors exhibited clear threshold effects on B-IBI. Random forest analysis revealed that hydrological factors contributed the highest relative importance to B-IBI (57.9%), followed by habitat factors (29.3%) and water quality factors (12.8%). The relative contribution of water quality factors to B-IBI in the main urban area was higher than that in Northeast Chongqing and Southeast Chongqing. At the river scale, the proportion of reservoir reaches showed a highly significant linear negative correlation with B-IBI (P<0.001), while the proportion of near-natural reaches exhibited a highly significant linear positive correlation (P<0.001). This study demonstrates that hydrological disturbance is the dominant factor affecting the benthic macroinvertebrate integrity of the tributary rivers in Chongqing, and the construction and operation of reservoirs significantly constrain B-IBI levels.