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bioRxiv · 10.1101/2023.10.27.564475

Failure to clear intra-osteocytic Staphylococcus aureus is not the result of antibiotic-mediated autophagy dysregulation

Gunn, N. J.·Zelmer, A. R.·Kidd, S. P.·Solomon, L. B.·Yang, D.·Roscioli, E.·Atkins, G. J.
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Abstract

BackgroundStaphylococcus aureus is a major causative pathogen of osteomyelitis. The intracellular infection of osteocytes and related bone cells can persist despite application of gold-standard clinical interventions. The mechanisms by which intracellular S. aureus persists during antibiotic therapy are unknown. In this study, we apply S. aureus to an in vitro model of differentiated osteocytes to investigate whether antibiotic-mediated dysregulation of autophagy contributes to this phenomenon. MethodsHuman osteocyte-like cells were exposed to combinations of rifampicin, vancomycin and modulators of autophagy, in the presence or absence of S. aureus. Intracellular bacterial growth characteristics were assessed through CFU analysis, viable bacterial DNA abundance and the rate of escape into antibiotic-free medium, in parallel with measures of host cell autophagic flux. ResultsRifampicin, alone or in combination with vancomycin, caused a rapid decrease in the culturability of the intracellular bacterial community, concomitant with stable or increased absolute bacterial DNA levels. Both antibiotics significantly inhibited autophagic flux. However, while the modulation of autophagic flux affected bacterial culturability, this modulation did not affect viable bacterial DNA levels. ConclusionsAutophagy was shown to be a factor in the host-pathogen relationship in this model, as its modulation affected the growth state of intracellular S. aureus with respect to both their culturability and propensity to escape the intracellular niche. Whilst rifampicin and vancomycin treatments moderately suppressed autophagic flux acutely, this did not explain the paradoxical response of antibiotic treatment in decreasing S. aureus culturability while failing to clear bacterial DNA and hence intracellular bacterial load. Thus, whilst rifampicin and vancomycin exhibited off-target effects that modulated autophagy in osteocyte-like cells, this could not explain the persistent infection observed for S. aureus.

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Gunn, N. J., Zelmer, A. R., Kidd, S. P., Solomon, L. B., Yang, D., Roscioli, E., Atkins, G. J.. 2023-10-28. Failure to clear intra-osteocytic Staphylococcus aureus is not the result of antibiotic-mediated autophagy dysregulation. https://doi.org/10.1101/2023.10.27.564475

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