We and others have postulated that the intracellular persistence of Staphylococcus aureus (1) may underlie the high failure rate of current gold-standard treatments for Periprosthetic Joint Infection (PJI) associated with joint replacement surgery. As such, the characterisation of the interactions between S. aureus and osteocytes, the most abundant cell type in bone, is of primary importance for understanding not only mechanisms of treatment failure and the development of chronic infection but also new modalities through which clinical cure can be achieved. An additional axis requiring consideration is how antibiotics influence this relationship, as chronic or high dose antibiotic treatments are known to cause adverse host cell effects. Autophagy, the physiological process by which intracellular material is targeted for phagolysosomal degradation, is a first-line innate immune mechanism against intracellular pathogens. In this study, the effects of modulation of autophagy on the clearance or persistence of intracellular S. aureus in a human osteocyte model (2) were assessed. Additionally, we examined the influence of clinically used antibiotics on this relationship. Interestingly, whilst both promotion and blockade of autophagic flux affected intracellular bacterial culturability in both an acute and chronic infection model, neither of these treatments affected bacterial burden in terms of DNA levels. Concerningly, treatment with vancomycin or rifampicin resulted in a modest but significant blockade of autophagic flux and an increase in intracellular viable bacterial DNA quantity. In conclusion, whilst autophagic flux was demonstrated to have a significant impact on the growth phenotype of the resident bacteria, its modulation was not capable of affecting the number of viable bacteria. Additionally, whilst antibiotics were capable of blocking autophagy, the paradoxical increase in the number of viable bacteria inside host osteocytes suggests the involvement of additional host or bacterial mechanisms resulting in failure to clear these S. aureus intracellular infections.