Staphylococcus aureus (S. aureus) and Staphylococcus epidermidis (S. epidermidis) are major skin commensals involved in acne pathogenesis and are increasingly associated with antibiotic resistance. This study aimed to determine the prevalence and resistance patterns of isolated Staphylococcus spp., identify associated risk factors, and evaluate the antibacterial activity of Ganoderma lucidum (G. lucidum) extract as a potential natural therapeutic alternative. Acne lesion samples from 135 volunteers were analyzed, yielding 128 S. aureus and 135 S. epidermidis isolates confirmed by biochemical tests. Antibiotic susceptibility was assessed against commonly used drugs. The antibacterial efficacy of G. lucidum extract was tested on Mueller-Hinton agar. Bioinformatics analysis, including molecular docking and ADMET profiling, predicted interactions of G. lucidum bioactives with resistance-related proteins (ermC, mecA, and β-lactamase). Gentamicin and rifampin showed 100% susceptibility in both species. However, erythromycin resistance was 93.3% in S. aureus and 60% in S. epidermidis, while amoxicillin resistance was highest in S. epidermidis (86.7%). Significant risk factors included age, gender, BMI, oily skin, family history, diet, and psychological stress. G. lucidum extract exhibited strong dose-dependent antibacterial activity against both species. Docking and dynamic simulations revealed lucidenic acid D1 strongly and stably binding ermC, while ganoderlactone B and E showed dual binding to mecA and β-lactamase, suggesting inhibition of erythromycin and amoxicillin resistance. ADMET analysis confirmed favorable pharmacokinetics and low toxicity. The study highlights high prevalence of antibiotic-resistant staphylococci in acne and demonstrates the potential of G. lucidum as an effective natural antibacterial and resistance-modifying agent, offering a promising alternative to conventional antibiotics.