Methicillin-resistant Staphylococcus aureus (MRSA), also known as oxacillin-resistant S. aureus, is a leading cause of community and hospital associated infections globally. In this work, we found that deletion of the arlRS two-component system genes in the USA300 and USA500 strains resulted in increased susceptibilities to oxacillin (8–16-fold decrease in minimal inhibitory concentrations). In USA300Δ arlRS, transcriptional levels of mecA or blaZ showed no obvious change, while mRNA levels of spx showed a 4-fold decrease at 4 h and a 6.3-fold decrease at 10 h. Overexpression of spx in Δ arlRS restored oxacillin resistance to a similar level in USA300. In addition, gel shift assay showed that the recombinant ArlR bound to spx promoter region. Furthermore, silencing of spx led to a significant increase of oxacillin susceptibility in multiple MRSA isolates. Our results indicate that ArlRS plays a strong role in regulating oxacillin resistance in MRSA strains, which involves direct modulation of spx expression. Moreover, oritavancin showed inhibition to ATPase activity of the recombinant histidine kinase ArlS (IC 50 = 5.47 μM). Oritavancin had synergy effect on oxacillin activity against the MRSA strains in both planktonic and biofilm state. Our data suggest that ArlRS is an attractive target for breaking antimicrobial resistance of MRSA.

Staphylococcus epidermidis is a common bacterial colonizer of human skin and mucous membranes, yet it has emerged as an important nosocomial pathogen largely due to its ability to form biofilms. Tobacco smoke has been demonstrated as a contributor to various infection diseases by improving the biofilm formation of multiple bacterial species; however, the association between tobacco smoke and S. epidermidis biofilm is still unclear. In this study, we tested the effect of nicotine, one of the most active components of tobacco, on S. epidermidis biofilm formation, and we studied the underlying mechanisms. Our results showed that nicotine promoted the biofilm formation of S. epidermidis 1457 strain (SE1457) and enhanced its initial attachment to a polyethylene surface as well as polysaccharide intercellular adhesin (PIA) production. In addition, an increased extracellular DNA release and a higher autolysis rate of SE1457 was detected after nicotine treatment, which was consistent with the increased ratio of dead cells in nicotine-treated SE1457 biofilm observed with confocal laser-scanning microscopy. Furthermore, the effect of nicotine on several autolysis-related and biofilm-related gene knockout mutants of SE1457 was tested. It showed that in ΔsaeRS, ΔlytSR, and ΔsceD, nicotine induced increase in biofilm formation was similar to that in SE1457; but in ΔarlRS, ΔatlE, and ΔicaC, the effect was obviously impaired. Consistently, the increase of the bacterial autolysis rate in ΔarlRS and ΔatlE induced by nicotine was not as significant as that in SE1457. Meanwhile, the growth inhibition of nicotine on SE1457 was observed, and it was much less on ΔarlRS and restored by the arlRS complementation. The arlRS transcription in SE1457 was inhibited by nicotine during cultivation as indicated by a promoter reporter assay using green fluoresent protein. Taken together, our study indicates that nicotine improves S. epidermidis biofilm formation by promoting its initial attachment and intercellular accumulation; the arlRS, atlE, and ica genes mediating bacterial autolysis and PIA production play an important role in this process.