Transcriptional regulatory factor EpsR mediates adaptation mechanism of extremophile Acidithiobacillus caldus to copper stress environment and performance in bioleaching of low-grade copper ore by enhancing "contact" mechanism.

Bioleaching is a critical approach for processing low-grade copper ores, relying on the "contact" mechanism mediated by microbial extracellular polymeric substance (EPS) secretion and biofilm formation. In the typical leaching microorganism Acidithiobacillus caldus, there is a transcriptional regulatory factor EpsR that regulates EPS secretion. It has a typical PilZ domain that binds cyclic diguanylate (c-di-GMP) and can sense Cu to relieve its own repression. Here, we overexpressed EpsR in A. caldus via conjugation transfer, demonstrating that the engineered strain exhibited a 78.0 % increase in maximum specific growth rate under 3 g/L CuSO stress. Based on TEM and CLSM analysis, it was found that the overexpressed strain had strong aggregation and increased EPS secretion. Quantitative results showed that the biofilm formation ability of the overexpressed strain increased by 43.5 %, and the exopolysaccharide content increased by 180.8 %. 3D-EEM and HPLC analysis showed that the proline, glycine, aspartic acid, arginine, histidine and humic acid in the EPS of the overexpression strain were increased to varying degrees. Bioleaching experiments revealed 13.6 % and 55.0 % elevations in planktonic and attached cell populations, respectively, with significant 36.8 % improvement in Cu bioleaching efficiency. All the above research results indicate that overexpression of EpsR can enhance the leaching efficiency by strengthening the "contact" mechanism. This study shows the adaptation strategy against environmental Cu stress by enhancing biofilm formation, which also provides an engineering pathway to raise the copper extraction efficiency.