The sortase A substrates FnbpA, FnbpB, ClfA and ClfB antagonize colony spreading of Staphylococcus aureus

Abstract

Staphylococcus aureus is an important human pathogen that is renowned both for its rapid transmission within hospitals and the community, and for the formation of antibiotic resistant biofilms on medical implants. Recently, it was shown that S. aureus is able to spread over wet surfaces. This motility phenomenon is promoted by the surfactant properties of secreted phenol-soluble modulins (PSMs), which are also known to inhibit biofilm formation. The aim of the present studies was to determine whether any cell surface-associated S. aureus proteins have an impact on colony spreading. To this end, we analyzed the spreading capabilities of strains lacking non-essential components of the protein export and sorting machinery. Interestingly, our analyses reveal that the absence of sortase A (SrtA) causes a hyper-spreading phenotype. SrtA is responsible for covalent anchoring of various proteins to the staphylococcal cell wall. Accordingly, we show that the hyper-spreading phenotype of srtA mutant cells is an indirect effect that relates to the sortase substrates FnbpA, FnbpB, ClfA and ClfB. These surface-exposed staphylococcal proteins are known to promote biofilm formation, and cell-cell interactions. The hyper-spreading phenotype of srtA mutant staphylococcal cells was subsequently validated in Staphylococcus epidermidis. We conclude that cell wall-associated factors that promote a sessile lifestyle of S. aureus and S. epidermidis antagonize the colony spreading motility of these bacteria.

Figure 1. Hyper-spreading phenotype of srtA mutant S. aureus strains.

From an overnight culture, an aliquot of 2 µl was spotted in the middle of a TSA plate, which was then incubated overnight at 37°C. The analyses include the laboratory strains S. aureus SH1000 and NCTC8325 (both labeled WT), as well as their srtA mutant derivatives (labeled srtA) and srtA mutants complemented with a plasmid pCN51-borne copy of S. aureus srtA (labeled srtA-pCN51). The spreading areas of the investigated mutant and parental strains were determined by ImageJ. The graphs show the areas covered in arbitrary units (AU) and respective standard deviations.

Figure 2. The influence of fnbpA, fnbpB, clfA and clfB mutations on colony spreading of S. aureus.

Spreading motility of S. aureus SH1000-derived fnbpA, fnbpB, clfA and/or clfB mutant strains or the S. aureus Newman srtA mutant strain was assayed as described for Figure 1.

Figure 3. Hyper-spreading phenotype of a srtA mutant of S. epidermidis 1457.

Spreading motility of S. epidermidis 1457 (WT), a srtA mutant derivative of this strain (srtA), and a complemented derivative of the srtA mutant (srtA^Se-pCN51) was assayed as described for Figure 1.