BACKGROUND: A bioleaching process could offer the advantage of higher metal recovery in a sustainable manner even from lithium-ion battery (LIB) samples with very low metal concentrations. In recent years, there has been a significant increase in the use of secondary resources such as LIBs for various purposes including transportation, large-scale energy storage and use in portable devices. RESULTS: The adaptation of a mixed culture of acidophilic microorganism (lab stock culture) to a representative LIB sample allowed the setting of 0.5% of the pulp density under lab scale conditions. The maximum metal dissolution by bioleaching in a 1-L bioreactor for the as-received and thermally treated samples was found to be Li (67% & 49%), cobalt (81% & 86%), nickel (99% & 87%) and manganese (86% & 75%). Likewise, on the 10-L scale, the dissolutions observed were: Li (80% & 67%), Co (75%), Ni (91% & 88%) and Mn (63% & 75%) for the as-received and heat-treated samples, respectively. CONCLUSION: Parameters such as particle size, leaching time, pH and iron ions (Fe2+) affect the efficiency of acidophilic bioleaching of Li, Co, Ni and Mn from spent LiBs. © 2024 Society of Chemical Industry (SCI).