This study aimed to spatially assess the ecological state of the upper Siret River basin in Ukraine, along with its main tributaries, using physicochemical and microbiological indicators. Additionally, we evaluated the impact of anthropogenic activities on surface water quality in the region. The research spanned one year, with nine sampling points ranging from the Siret River sources in the Pokutsko-Bukovynian Carpathians to the Romania border crossing area. Notably, a significant portion of the upper basin lies within the Vyzhnytskyi National Nature Park. Physicochemical analysis involved measuring pH, water turbidity, and concentrations of NH4+, NO3-, and CI- ions. Microbiological analysis focused on total and fecal bacteria, specifically E. coli species. Results revealed a significant correlation between population density, the absence of centralized sewage treatment facilities in large settlements, and the level of surface water pollution in the Ukrainian segment of the Siret River basin. The upper part of the river network, where the Vyzhnytskyi National Nature Park is located, displayed the cleanest waters. Downstream, surface water pollution increased, particularly near certain points outside large settlements and tourist complexes (Myhove, Berehomet, and Storozhynets). Our findings highlight the importance of using microbiological indicators to monitor the ecological state of Danube sources in the Eastern Carpathians.