Construction and biological phenotype characterization of vraSR-srrAB deletion mutant of Staphylococcus epidermidis

Abstract:

[Background] Biofilm formation is the major pathogenic factor of Staphylococcus epidermidis. The two-component regulatory system (TCS) composed of VraSR and SrrAB is involved in the growth, biofilm formation and the other biological phenotypes of bacteria. The deletion of vraSR leads to thinner cell wall and decreased biofilm formation, and the deletion of srrAB results in lagged growth before the stationary phase and decreased biofilm formation of S. epidermidis 1457 (SE1457). Both VraSR and SrrAB modulate biofilm formation in an ica-dependent manner. The ica operon is a key regulatory element of biofilm formation in S. epidermidis, which is composed of four genes (icaADBC) and a transcriptional repressor (icaR). [Objective] To explore the synergistic regulation of TCS-VraSR and SrrAB on the growth and biofilm formation of S. epidermidis, so as to lay a foundation for the prevention and control of persistent infection caused by S. epidermidis. [Methods] The recombinant plasmid pKOR1-ΔvraSR was constructed, modified by E. coli DC10B, and transformed into ΔsrrAB. The vraSR gene was deleted from the genome of ΔsrrAB by homologous recombination. The suspected ΔvraSR-srrAB was verified by PCR, RT-PCR, and sequencing. The growth, biofilm formation, and drug susceptibility of ΔvraSR-srrAB were examined. [Results] ΔvraSR-srrAB was successfully constructed. Compared with SE1457, ΔvraSR, and ΔsrrAB, ΔvraSR-srrAB exhibited retarded growth, increased drug susceptibility, and decreased biofilm formation. The RT-qPCR showed that the deletion of vraSR or srrAB down-regulated the transcriptional level of icaA by 13%–17% and up-regulated that of icaR by 5–9 folds compared with that in SE1457. The deletion of both vraSR and srrAB down-regulated the transcriptional level of icaA by 6% and up-regulated that of icaR by 14 folds compared with that in SE1457. [Conclusion] VraSR and SrrAB may cooperatively regulate the biofilm formation of S. epidermidis through the ica pathway, and the mechanism of VraSR and SrrAB in modulating the responses to stress remains to be studied.

Key words:VraSR;SrrAB;two-component regulatory system;biofilm;Staphylococcus epidermidis