This study explores the potential for synchronous extraction of Cu, Cr, Ni and Zn during sewage sludge bioleaching processes, using three types of bacterial cultures: a pure culture of Acidithiobacillus ferrooxidans (A. ferrooxidans); a pure culture of Acidithiobacillus thiooxidans (A. thiooxidans); and a mixed culture of A. ferrooxidans and A. thiooxidans. Variable operating parameters included initial pH, solids concentration, sulfur concentration and ferrous iron concentration, with optimization via Box-Behnken design of response surface methodology. Results indicate that the mixed culture of A. ferrooxidans and A. thiooxidans, was the most effective at bioleaching heavy metals from sewage sludge. The optimal operating conditions were as follows: an initial pH of 2.0, with concentrations of 3% solids, 6.14 g L−1 sulfur and 4.55 g L−1 ferrous iron. Maximum extraction efficiencies obtained after 14 days of bioleaching under optimal conditions, were 98.54% Cu, 57.99% Cr, 60.06% Ni and 95.60% Zn. Bioleaching kinetics were effectively simulated using a shrinking core model to explain the leaching reaction, with modelling results suggesting that the rate was determined by the diffusion step.