Biofilm production by staphylococci is an important virulence determinant mediated by the icaADBC-encoded polysaccharide intercellular adhesin (PIA) or by surface and extracellular proteins. Deletion of the Staphylococcus accessory regulator sarX significantly reduced biofilm-forming capacity in Staphylococcus epidermidis CSF41498, whereas multicopy sarX complemented the sarX mutant and increased wild-type biofilm production. In Staphylococcus aureus, SarX negatively regulates the accessory gene regulator (Agr) system, which in turn has strain-specific effects on biofilm regulation. Here we found that purified S. epidermidis SarX protein bound specifically to the agr P3 promoter. However RT-PCR analysis revealed that both mutation of sarX and multicopy sarX activated RNAIII transcription, making it difficult to correlate sarX-mediated biofilm regulation with altered agr activity. In contrast, RT-PCR and immunoblot analysis revealed that icaA transcription and PIA expression were decreased in the sarX mutant, whereas multicopy sarX increased ica and PIA expression. Furthermore, multicopy sarX did not promote biofilms in an icaC mutant. Finally, purified SarX protein bound specifically to the ica operon promoter. Taken together, these data reveal that the S. epidermidis SarX protein regulates the transcriptional activity of the agr and ica loci and controls the biofilm phenotype, primarily by regulating icaADBC transcription and PIA production.
BeenkenK. E.,
MrakL. N.,
GriffinL. M.,
ZielinskaA. K.,
ShawL. N.,
RiceK. C.,
HorswillA. R.,
BaylesK. W.,
SmeltzerM. S.2010; Epistatic relationships between sarA and agr in Staphylococcus aureus biofilm formation. PLoS ONE 5:e10790
CoelhoL. R.,
SouzaR. R.,
FerreiraF. A.,
GuimarãesM. A., Ferreira-CarvalhoB. T.,
FigueiredoA. M.2008; agr RNAIII divergently regulates glucose-induced biofilm formation in clinical isolates of Staphylococcus aureus . Microbiology 154:3480–3490
ConlonK. M.,
HumphreysH.,
O'GaraJ. P.2002a; icaR encodes a transcriptional repressor involved in environmental regulation of ica operon expression and biofilm formation in Staphylococcus epidermidis . J Bacteriol 184:4400–4408
CramtonS. E.,
GerkeC.,
SchnellN. F.,
NicholsW. W.,
GötzF.1999; The intercellular adhesion ( ica ) locus is present in Staphylococcus aureus and is required for biofilm formation. Infect Immun 67:5427–5433
GillS. R.,
FoutsD. E.,
ArcherG. L.,
MongodinE. F.,
DeboyR. T.,
RavelJ.,
PaulsenI. T.,
KolonayJ. F.,
BrinkacL.other authors2005; Insights on evolution of virulence and resistance from the complete genome analysis of an early methicillin-resistant Staphylococcus aureus strain and a biofilm-producing methicillin-resistant Staphylococcus epidermidis strain. J Bacteriol 187:2426–2438
HeilmannC.,
HussainM.,
PetersG.,
GötzF.1997; Evidence for autolysin-mediated primary attachment of Staphylococcus epidermidis to a polystyrene surface. Mol Microbiol 24:1013–1024
HennigS.,
Nyunt WaiS.,
ZiebuhrW.2007; Spontaneous switch to PIA-independent biofilm formation in an ica -positive Staphylococcus epidermidis isolate. Int J Med Microbiol 297:117–122
HollandL. M.,
O'DonnellS. T.,
RyjenkovD. A.,
GomelskyL.,
SlaterS. R.,
FeyP. D.,
GomelskyM.,
O'GaraJ. P.2008; A staphylococcal GGDEF domain protein regulates biofilm formation independently of cyclic dimeric GMP. J Bacteriol 190:5178–5189
KreiswirthB. N.,
LöfdahlS., BetleyM. J.,
O'ReillyM.,
SchlievertP. M.,
BergdollM. S.,
NovickR. P.1983; The toxic shock syndrome exotoxin structural gene is not detectably transmitted by a prophage. Nature 305:709–712
MackD.,
FischerW.,
KrokotschA.,
LeopoldK.,
HartmannR.,
EggeH.,
LaufsR.1996; The intercellular adhesin involved in biofilm accumulation of Staphylococcus epidermidis is a linear beta-1,6-linked glucosaminoglycan: purification and structural analysis. J Bacteriol 178:175–183
ManiN.,
TobinP.,
JayaswalR. K.1993; Isolation and characterization of autolysis-defective mutants of Staphylococcus aureus created by Tn 917 - lacZ mutagenesis. J Bacteriol 175:1493–1499
MannaA. C.,
CheungA. L.2006; Expression of SarX, a negative regulator of agr and exoprotein synthesis, is activated by MgrA in Staphylococcus aureus . J Bacteriol 188:4288–4299
MorfeldtE.,
TaylorD.,
von GabainA.,
ArvidsonS.1995; Activation of alpha-toxin translation in Staphylococcus aureus by the trans -encoded antisense RNA, RNAIII. EMBO J 14:4569–4577
MorfeldtE.,
Panova-SapundjievaI.,
GustafssonB.,
ArvidsonS.1996; Detection of the response regulator AgrA in the cytosolic fraction of Staphylococcus aureus by monoclonal antibodies. FEMS Microbiol Lett 143:195–201
O'GaraJ. P.2007; ica and beyond: biofilm mechanisms and regulation in Staphylococcus epidermidis and Staphylococcus aureus . FEMS Microbiol Lett 270:179–188
O'NeillE.,
PozziC.,
HoustonP.,
SmythD.,
HumphreysH.,
RobinsonD. A.,
O'GaraJ. P.2007; Association between methicillin susceptibility and biofilm regulation in Staphylococcus aureus isolates from device-related infections. J Clin Microbiol 45:1379–1388
OshidaT.,
SugaiM.,
KomatsuzawaH.,
HongY. M.,
SuginakaH.,
TomaszA.1995; A Staphylococcus aureus autolysin that has an N -acetylmuramoyl-l-alanine amidase domain and an endo-beta- N -acetylglucosaminidase domain: cloning, sequence analysis, and characterization. Proc Natl Acad Sci U S A 92:285–289
QinZ.,
OuY.,
YangL.,
ZhuY.,
Tolker-NielsenT.,
MolinS.,
QuD.2007; Role of autolysin-mediated DNA release in biofilm formation of Staphylococcus epidermidis . Microbiology 153:2083–2092
SugaiM.,
KoikeH.,
HongY. M.,
MiyakeY.,
NogamiR.,
SuginakaH.1989; Purification of a 51 kDa endo-beta- N -acetylglucosaminidase from Staphylococcus aureus . FEMS Microbiol Lett 52:267–272
VuongC.,
SaenzH. L.,
GötzF., OttoM.2000b; Impact of the agr quorum-sensing system on adherence to polystyrene in Staphylococcus aureus . J Infect Dis 182:1688–1693
VuongC.,
GerkeC.,
SomervilleG. A.,
FischerE. R.,
OttoM.2003; Quorum-sensing control of biofilm factors in Staphylococcus epidermidis . J Infect Dis 188:706–718
YarwoodJ. M.,
SchlievertP. M.2000; Oxygen and carbon dioxide regulation of toxic shock syndrome toxin 1 production by Staphylococcus aureus MN8. J Clin Microbiol 38:1797–1803