Oxidative leaching is an inexpensive alternative to using chemical cyanide extraction methods for gold from low-grade gold sulfide. Oxidation of finely ground gold-bearing ore by Acidithiobacillus ferrooxidans was evaluated in terms of cell density, pH, and leaching efficiency of Fe and Au in shake flask experiments. The compositional and elemental analyses of the beneficiated ore were analyzed using XRD and EDXRF spectroscopy. The ore’s primary constituents are gold (4.356 mg/L), silicon, iron, and sulfur (62.456, 15.441, and 7.912 wt%, respectively). XRD spectra, the main phases of the concentrated ore, showed that the major components of the ore were quartz, syn, silicon sulfide, pyrite, and polymetallic elements such as silderenrite, gismondine, siderenikite, hematite, and syn. The experimental results, with Acidithiobacillus ferrooxidans bacteria and blank, were evaluated. The pH of the blank remained nearly constant, and the pH of the bioleached was occasionally lowered. The A. ferrooxidans strain always grew better throughout the bioleaching process. For the A. ferrooxidans strain, the cell density of cells reached a maximum of 90.00 × 106 cells/mL after the 11th week and decreased to 87.00 × 106 cells/mL after the 12th week. The decrease in cell density may be due to the presence of polymetallic elements such as Al, Cr, Ti, and Ni, leading to reduced metal tolerance of the A. ferrooxidans strain. In the A. ferrooxidans leaching process, the maximum total iron and gold extraction reached 92.16% (14.23 mg/L) and 99.97% (4.355 ppm), respectively, after the 11th week, and leaching tends to decrease up to 14 weeks, which may be due to the formation of secondary minerals. More research will be performed to optimize the procedure and leaching kinetic, examine the impact of metal content, and take into account the potential for bioleaching process pollution in addition to the amount of gold recovered.