The gradual improvement of traffic network in urban agglomeration plays an important role in the construction of national land development environment, construction of national land development structure, and optimization of functional space system. At the same time, it also causes serious disturbance to the structure and function of ecosystem and the health status of ecosystem, and seriously threatens the sustainable development of urban agglomeration. How to promote the high-quality development of urban agglomerations and alleviate the disturbance and damage of urban agglomerations to ecosystem health has become an urgent problem to be answered and solved in the current development process of urban agglomerations. To scientifically reveal the spatial relationship between traffic accessibility and ecosystem health in urban agglomerations, and to clarify the impact mechanism of traffic accessibility on ecosystem health is an effective way to solve this issue. Based on multi-source data, this study measured the traffic accessibility and ecosystem health level in the middle reaches of the Yangtze River Urban Agglomerations (MRYRUA) from 1995 to 2015, and estimated the influence mechanism of traffic accessibility on ecosystem health with bivariate spatial autocorrelation and an integrated spatial regression models from the global and local perspectives. The results showed that:(1) the overall ecosystem health in the MRYRUA exhibited a decreasing trend, and the ecosystem health level in the mountainous areas were significantly higher than those of the plain areas during the study period. (2) The bivariate spatial autocorrelation results showed that there was a significantly spatial dependence between traffic accessibility and ecosystem health. The spatial relationships between them mainly included low traffic accessibility-low ecosystem health level, high traffic accessibility-low ecosystem health level, and low traffic accessibility-high ecosystem health level. (3) The spatial regression results showed that traffic accessibility development would lead to the deterioration of ecosystem health. The impact of traffic accessibility on ecosystem health had significantly spatial heterogeneity. The results show that the ecosystem health is not only affected by the ecosystem elements of the local unit, but also by the health status of the surrounding units or distant units and other factors. Therefore, the trans-regional joint governance is particularly important in urban agglomeration. Secondly, the results show that the impact of traffic accessibility on ecosystem health has significant spatial heterogeneity, and local differentiated management and control measures for regional future traffic construction and ecosystem health protection are necessary for the sustainable development of urban agglomerations. The findings can provide a scientific basis for the formulation of macro-control policies for ecosystem health protection and differentiated control policies for urban agglomerations in the MRYRUA.