The Macrobenthic Integrity Index (B-IBI) was constructed based on the macrobenthic community data in Lake Dongting for more than 30 years (1988-2021) in order to assess the ecological quality of Lake Dongting, capture the status and change trend of aquatic biological integrity in Lake Dongting, and support the assessment of ecological condition in the Yangtze River Basin. Based on the integrity assessment results, the main environmental factors and contribution rates of the B-IBI in Lake Dongting over the past 30 years were analysed using the multiple stepwise regression method. The reference state of the benthic characteristics was set by the data during the low-pressure period (1988-1994). The B-IBI consisted of five core parameters: total taxon number, crustacean and mollusk taxon number, Shannon diversity index, total density and BI. At the same time, the expected values of the five parameters were determined and the criteria for B-IBI scoring were established: B-IBI≥6.34, excellent; 4.75≤B-IBI<6.34, good; 3.17≤B-IBI<4.75, average; 1.58≤B-IBI<3.17, poor; B-IBI<1.58, very poor. Box plot analysis of the B-IBI in the low- and high-pressure groups further verified the discriminatory power and applicability of the B-IBI. According to the B-IBI scoring criteria, the B-IBI decreased from 6.99 (excellent) in 1988 to 2.97 (poor) in 2021, indicating that the integrity of the benthic fauna in Lake Dongting showed a significant decline. The analysis of related environmental factors showed that the comprehensive nutritional status index, total nitrogen and chlorophyll-a in Lake Dongting showed a significant upward trend. The analysis showed that B-IBI was negatively correlated with total nitrogen and chlorophyll-a significantly. Multiple stepwise regression analysis showed that total nitrogen explained 56.1% of the variation in macrobenthic integrity, and 61.3% when combined with chlorophyll-a, indicating that the change in macrobenthic integrity was mainly driven by the increase in nutrients. The B-IBI assessment standard showed good applicability in Lake Dongting, which could provide a valuable scientific basis for long-term monitoring of changes in lake integrity and analysis of changes in ecological status. In the context of the ecological protection of important water bodies in the Yangtze River basin, it is suggested to pay more attention to the influence of nitrogen pollution in the continuous control of phosphorus, to strengthen the control of total nitrogen, and to include total nitrogen in the national and local water ecological environment control systems.