Staphylococcus aureus is a skin commensal microorganism commonly colonizing healthy humans. Nevertheless, S. aureus can also be responsible for cutaneous infections and contribute to flare-up of inflammatory skin diseases such as atopic dermatitis (AD), which is characterized by dysbiosis of the skin microbiota with S. aureus as the predominant species. However, the role of major virulence factors of this pathogen such as phenol-soluble modulin (PSM) toxins in epidermal inflammation remains poorly understood. Stimulation of primary human keratinocytes with sublytic concentrations of synthetic and purified PSM α3 resulted in upregulation of a large panel of pro-inflammatory chemokine and cytokine gene expression, including CXCL1, CXCL2, CXCL3, CXCL5, CXCL8, CCL20, IL-1α, IL-1β, IL-6, IL-36γ and TNF-α, while inducing the release of CXCL8, CCL20, TNF-α and IL-6. In addition, using S. aureus culture supernatant from mutants deleted from genes encoding either α-type PSMs or all PSM production, PSMs were shown to be the main factors of S. aureus secretome responsible for pro-inflammatory mediator induction in human keratinocytes. On the other hand, α-type PSM-containing supernatant triggered an intense induction of pro-inflammatory mediator expression and secretion during both topical and basal layer stimulation of an ex vivo model of human skin explants, a physiologically relevant model of pluristratified epidermis. Taken together, the results of this study show that PSMs and more specifically α-type PSMs are major virulence factors of S. aureus inducing a potent inflammatory response during infection of the human epidermis and could thereby contribute to AD flare-up through exacerbation of skin inflammation.

The inoculum effect (i.e., reduction in antimicrobial activity at large starting inoculum) is a phenomenon described for various pathogens. Since limited data exist regarding inoculum effect of Acinetobacter baumannii , we evaluated killing of A. baumannii by polymyxin B, a last-resort antibiotic, at several starting inocula and developed a PKPD model to capture this phenomenon. In vitro static time-kill experiments were performed using polymyxin B at concentrations ranging from 0.125 to 128 mg/L against a clinical A. baumannii isolate at four starting inocula from 10 5 to 10 8 CFU/mL. Samples were collected up to 30 h to quantify the viable bacterial burden and were simultaneously modeled in the NONMEM software program. The expression of polymyxin B resistance genes ( lpxACD , pmrCAB and wzc ), and genetic modifications were studied by RT-qPCR and DNA sequencing experiments, respectively. The PKPD model included a single homogeneous bacterial population with adaptive resistance. Polymyxin B effect was modelled as a sigmoidal E max model and the inoculum effect as an increase of polymyxin B EC 50 with increasing starting inoculum using a power function. Polymyxin B displayed a reduced activity as the starting inoculum increased: a 20-fold increase of polymyxin B EC 50 was observed between the lowest and the highest inoculum. No effects of polymyxin B and inoculum size were observed on the studied genes. The proposed PKPD model successfully described and predicted the pronounced in vitro inoculum effect of A. baumannii on polymyxin B activity. These results should be further validated using other bacteria/antibiotic combinations and in vivo models.

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