Concentrations of pollutants were measured in water, sediment and algal samples collected along a longitudinal gradient from a stretch of the Olifants River, South Africa, that receives AMD from the Klipspruit River. The effects of AMD were determined through macroinvertebrate biotic indices (SASS5) and multivariate analysis of macroinvertebrate communities. The acidic Klipspruit River caused increased concentrations of total Al, Fe and Mn in the Olifants River. Upon mixing of the Klipspruit with that from the alkaline Olifants River, Al and Fe precipitate rapidly, leading to lower concentrations in the dissolved phase and higher concentrations in the suspended phase and in sediment at sites in close proximity to the confluence. Similarly filamentous algae accumulated high concentrations of Al, Fe and Zn immediately after the confluence. Mn remains in the dissolved phase and sediment and algal concentrations increase with increasing distance downstream. Metal speciation analysis indicate that Al is rapidly converted from more toxic forms (e.g., Al 3+ and Al(OH) 2+ ) to less toxic forms (e.g., Al(OH) 3 (aq) and Al(OH) 4-). In contrast, Mn remains in the soluble Mn 2+ form.Macroinvertebrate metrics and community structure showed clear signs of deterioration in water quality in the Olifants River downstream of the point of AMD input. While total TDS concentrations at all sites fall within ranges likely to affect macroinvertebrates, the relative composition of major ions changes as a result of AMD input, which may also account for the observed changes in macroinvertebrate communities. Further downstream, the Wilge River discharges into the Olifants River, and significantly improves water quality downstream of the confluence. Future mining and development activities in the Wilge catchment should be carefully managed and monitored so as to ensure sufficient flows of acceptable quality to prevent further deterioration of water quality in the Olifants River and downstream reservoirs.