Hexavalent chromium (Cr 6+ ) contamination from a dolomite stone mine in Limpopo Province, South Africa, has resulted in extensive groundwater contamination. In order to circumvent any further negative environmental impact at this site, an effective and sustainable treatment strategy for the removal of up to 6.49 mg/ℓ Cr 6+ from the groundwater was developed. Laboratory-scale, continuous up-flow bioreactors were constructed to evaluate reduction of Cr 6+ , with a residence time of 24 h, an efficiency porosity of 44% and a flow rate of 1.5 mℓ/min. Stoichiometrically balancing terminal electron acceptors in the feed water with a selected electron donor, directed reactor balance for complete Cr 6+ reduction. The microbial community shifted in relative dominance during operation to establish an optimal metal-reducing community, including Enterobacter cloacae, Flavobacterium sp. and Ralstonia sp., which achieved 100% reduction. Evaluation after reactor termination with SEM-EDX and XRD confirmed the establishment of biofilm on the reactor matrix, as well as trivalent chromium (Cr 3+ ) precipitation within the reactor. Due to gravitational force, high concentrations of Cr 3+ were found in the bottom third of the reactor. Based on the results from the laboratory investigation, a 24 000 ℓ fixed-film pilot bioreactor was designed and constructed at this site. Influent flow rates, electron donor injection and automated sampling were remotely controlled by a programmable logic controller (PLC). Similar to the laboratory column study, steady state conditions could be achieved and successful Cr 6+ reduction was evident. This is the first up-scaled, effective demonstration of a biological chromium(VI) bioremediation system in South Africa.