Gold mining pollution has long-lasting effects on the environment, particularly through acid mine drainage (AMD) and heavy metal contamination. Monitoring and assessing the impact of this pollution is crucial, as well as evaluating the effectiveness of remediation efforts. In our study, conducted in the gold mining area of Zlatna (GMAZ), western Romania, we utilised on-site measurements of water temperature, pH, electrical conductivity, and dissolved oxygen, along with the quantification of culturable aerobic bacteria and microfungi using ready-to-use media plates. We also examined the taxonomic richness of water invertebrates (TRWI) and the environmental features of the sites. Our study found significant negative impacts on the water biota in mining areas, with microbial abundance proving to be a reliable indicator of AMD pollution. While water invertebrates can also serve as indicators of mining effects, their abundance alone may not always accurately reflect pollution levels at every site. This multiple-factor analysis highlights the influences of water type, geological characteristics, air temperature, and precipitation on the structure of the aquatic biota. We observed a natural attenuation of mining pollution in the GMAZ in the last seven years. This study demonstrates that the quantification of microbiota, along with TRWI and basic physicochemical parameters, can offer a cost-effective alternative to expensive monitoring methods for assessing mining pollution.