Water quality assessments provide essential information for protecting aquatic habitats and stakeholders downstream of mining sites. Moreover, mining companies must comply with environmental quality standards and include public participation in water quality monitoring (WQM) practices. However, overarching challenges beyond corporate environmental responsibility are the scientific soundness, political relevance and harmonization of WQM practices. In this study, a mountainous watershed supporting large-scale gold mining in the headwaters, besides urban and agricultural landuses at lower altitudes, is assessed in the dry season. Conventional physicochemical and biological (Biological Monitoring Water Party-Colombia index) freshwater quality parameters were evaluated, including hydromorphological and land-use characteristics. According to the indicators used, water quality deterioration by mining was absent, in contrast to the effects of urban economic activities, hydromorphological alterations and (less important) agricultural pollutants. We argue that mining impacts are hardly captured due to the limited ecological knowledge of high-mountain freshwaters, including uncharacterized mining-specific bioindicators, environmental baselines and groundwater processes, as well as ecotoxicological and microbial freshwater quality components. Lessons for overcoming scientific and operational challenges are drawn from joint efforts among governments, academia and green economy competitiveness. Facing a rapid development of extractive industries, interinstitutional and multidisciplinary collaborations are urgently needed to implement more integrated freshwater quality indicators of complex mining impacts.