Staphylococcus aureus Pathogenesis - Gene exchanges - Gene regulation - Gene products - Gene product cooperation Brussels 07
Some Staphylococcal Species from Mammals Host Species Coagulase Clumping factor Virulence Human ++ ++ +++ S. aureus and other S. epidermidis - - + primates S. capitis - - (+) - - (+) S. caprae S. saccharolyticus + - - S. warneri - - - - - - S. pasteuri S. haemolyticus - - + S. hominis - - (+) - + + S. lugdunensis S. auricularis - - (+) - S. saprophyticus - + - - - S. cohnii S. xilosus - - - S. simulans - - - + + + S. schleiferi Carnivores S. intermedius + - ++ ++ S. felis - - Brussels 07
Whole genomes of S. aureus MW2, N315 and Mu50 MW2 N315 Mu50 - GC content - Gene comparison - Codon usage - Genometrics Baba T et al. Lancet 2002;359:1819 Brussels 07
From Linguistics to Genometrics theicaacisagreatconférenceparcequellegathermanygreatscientists theicaacisagreatconférenceparcequellegathermanygreatscientists http://www2.unil.ch/comparativegenometrics/ CA Roten and L. Guy Brussels 07
Guy Lionel, Thesis 2007 Brussels 07 S. aureus Genometric Analysis
Whole genomes of S. aureus MW2, N315 and Mu50 Methicillin resistance MW2 N315 Mu50 tst1 set Provirus encoding PVL toxin Baba T et al. Lancet 2002;359:1819 Brussels 07
Define Pathogenicity Islands - DNA region (up to 200 kb) of foreign origin - Contain pathogenic genes - May have a different GC content - Flanked by direct repats of IS - Associated with tRNA genes - Contain relics of integrase/excisase genes or of phage genes Hacker J et al. Mol Microbiol 1997; 23:1089-97 Brussels 07
Brussels 07 - Transduction (generalized) How do they move ? - Transformation - Conjugation
Brussels 07 Moving Genes Around att int att The example of SaPI1
Brussels 07 Moving Genes Around att xis int att The example of SaPI1
Brussels 07 Moving Genes Around att xis int att The example of SaPI1
Brussels 07 Moving Genes Around The example of SaPI1
Brussels 07 Moving Genes Around att int att The example of SaPI1
Brussels 07 Moving SaPI1 with Phage 80 α Novick, Schlievert, Ruzin. Microbes and Infect 2001; 3:585
Staphylococcus aureus Pathogenesis - Gene exchanges - Gene regulation - Gene products - Gene product cooperation Brussels 07
agr-Regulation of Virulence Determinants During in vitro Growth of S. aureus (1) Growth Transcription Transcription of adhesins of exoproteins Time Adapted from R.P. Novick ASM press, 2000 Brussels 07
Virulence Gene Regulation by Peptides in S. aureus (1) AIP AgrC AgrB P AgrA δ A C D B P2 P3 RNAIII AIP = auto-inducing peptide Novick et al. Curr. Op. Microbiol. 1999;2:40-45 Brussels 07
Virulence Gene Regulation by Peptides in S. aureus (2) AIP AgrC AgrB P AgrA AgrA Activation of Tx δ A C D B P2 P3 RNAIII Translation of exoproteins AIP = auto-inducing peptide Novick et al. Curr. Op. Microbiol. 1999;2:40-45 Brussels 07
Regulation of S. aureus super-antigens Action of regulatory genes Gene Product Activity/function agr Sae- rot sarA sarS sarT tst RS Superantigens sea Enterotoxin A Food poisoning, TSS 0 0 seb Enterotoxin B Food poisoning, TSS + b - sec Enterotoxin C Food poisoning, TSS + sed Enterotoxin D Food poisoning, TSS + eta Exfoliatin A Scalded skin syndrome + etb Exfoliatin B Scalded skin syndrome + tst Toxic shock toxin-1 Toxic shock syndrome + - b - 0, no effect of gene on expression; +, upregulated; -, downregulated. b. Controversial. Brussels 07
Conditions for Developing Toxic Shock Syndrome Host: - Lack of anti-TSST antibodies Environment: - High protein concentration - Neutral pH Altered - High pCO2 Ecology ! - High pO2 Brussels 07
Staphylococcus aureus Pathogenesis - Gene exchanges - Gene regulation - Gene products - Gene product cooperation Brussels 07
Gene Protein Function Implication in disease spa Protein A Binds Fc fragment Sepsis, osteoarthritis clfA Clumping factor A Binding to fibrinogen Endocarditis clfB Clumping factor B Binding to fibrinogen - cna Collagen binding protein Binding to collagen Osteomyelitis fnA Fn-binding protein A Binding to fibronectin Endocarditis, persistence fnB Fn-binding protein B Binding to fibronectin - sdrC Serine-aspartate repeat prot Binding to fibrinogen - sdrD Serine-aspartate repeat prot Binding to fibrinogen ? - sdrE Serine-aspartate repeat prot Binding to fibrinogen ? - pls Plasmin-sensitive protein Binding to nasal mucosa Nasal colonization? fmtB Methicillin resistance Putative cell-wall building Expression of methi-R sasA S. aureus surface protein A Undetermined - sasB S. aureus surface protein B Undetermined - sasC S. aureus surface protein C Undetermined - sasE S. aureus surface protein E Undetermined - sasF S. aureus surface protein F Undetermined - sasG S. aureus surface protein G Binding to nasal mucosa Invasive disease sasH S. aureus surface protein H Undetermined Invasive disease sasI S. aureus surface protein I Undetermined - sasJ S. aureus surface protein J Undetermined - sasK S. aureus surface protein K Undetermined -
Understanding Pathogenic Factors Coa - Coa - /ClfA - Coa - /ClfA - /Fnb - Wild Type ? Platelet ? ? ? Factors ? ? ? ? Coagulation ? ? ? Tissue factor Vegetations Brussels 07
Transfer or Staphylococcal Pathogenic genes into less Virulent Bacteria S. aureus donor Lactococcus Lactococcus (Coa + /ClfA + /Fnb + ) recipient (ClfA - ) recombinant (ClfA + ) 1: Transfer of S.aureus clfA gene 2: Select clumping-positive into viridans group streptococcus recombinants Que et al. I&I 2000; 68:3516-22 Brussels 07
Adherence of L. lactis to PMMA coverslips coated with fibronectin 100 Adherent CFU (x10 3 /coverslip) 80 60 8325-4 pIL253 40 FnbpA (+) 8325-4 20 0 0.0 0.25 0.50 0.75 Soluble concentration of Fn (µg/ml) Que et al, I&I 2001; 69:6296-302 Brussels 07
Brussels 07 Experimental Endocarditis
Infectivity of L. lactis Recombinants Binding to Fibrinogen or Fibronectin L. lactis parent ClfA+ FnBPA+ 100 % Infected Vegetations 80 60 40 20 0 10 4 10 5 10 6 10 7 10 4 10 5 10 6 10 7 Inoculum = 10 4 10 5 10 6 10 7 Que et al. Infect. Immun. 2001 Brussels 07
Dynamic of L. lactis Endocarditis L. lactis parent ClfA+ FnBPA+ 10 Log CFU/g Vegetations 8 6 4 2 0 Time after infection (h) = 2 12 48 72 2 12 48 72 2 12 48 72 Que et al. J. Exp. Med . 2005 Brussels 07
Brussels 07 Experimental Endocarditis
Neighboring Aorta of Valves Infected with ClfA-positive Lactococci /Gram Stain Que YA, J. Exp. Med 2005;201:1627 Brussels 07
Brussels 07 Que YA, J. Exp. Med 2005;201:1627
Binding Domains of FnBPA Fibronect. Fibronect. Fibronect. Fibrinogen Leader M-LPXTG W D1-4 Du C B A L Binding to Fibrinogen Fibonectin ClfA +++ - FnBPA ++ +++ Membrane Peptidoglycan Que YA, J. Exp. Med 2005;201:1627 and Piroth et al. 2007, submitted Brussels 07
Binding Domains of FnBPA Fibronect. Fibronect. Fibronect. Fibrinogen Leader M-LPXTG W D1-4 Du C B A L M-LPXTG W L M-LPXTG W D1-4 Du C L M-LPXTG W A L Membrane Peptidoglycan Que YA, J. Exp. Med 2005;201:1627 and Piroth et al. 2007, submitted Brussels 07
Binding Domains of FnBPA Surrogate recombinant Binding Infection Internalization Persistence Fg +++ --- --- Fn --- +++ --- cis Fn/Fg +++ +++ +++ trans Fn/Fg +++ +++ +++ Que YA, J. Exp. Med 2005;201:1627 and Piroth et al. 2007, submitted Brussels 07
- S. aureus needs both fibrinogen- and fibronectin- binding domains to be fully infective - Fibronectin-binding protein represents an example of intra-molecular domain cooperation - Fibronectin- and fibrinogen-binding represent an example of inter-molecular domain cooperation Brussels 07
E. Widmer et al. I&I , submitted Brussels 07
Brussels 07 Que YA, J. Exp. Med 2005;201:1627
Brussels 07 Que YA, J. Exp. Med 2005;201:1627
LPXTG-proteins Toxins 21 > 30 S. aureus S. epidermidis > 10 No 6 No S. haemolyticus S. saprophyticus 1 No But all have prophages and genomic islands, including the SCC staphylococcal K7 chromosome Brussels 07 Que YA, J. Exp. Med 2005;201:1627
Staphylooccus aureus a paradigm of host-parasite evolution YA Que N. Widmer L Piroth M. Giddey CA. Roten J. Entenza L. Guy S. Piu F. Collyn J. Vouillamoz
Subcutaneous Infection of Mice with 10 8 CFU of AIP I S. aureus supplemented or not with AIP II Strain 6390B, agr+ 1000 Strain 6911 , agr- Lesion size in mm 2 6390B + 10 μ g AIP II 800 6390B + 5 μ g AIP II 600 400 200 0 n = 4 4 3 4 4 4 3 4 Day 1 Day 5 Mayville et al. PNAS 1999; 96:1218-1223 Brussels 07
Is agr a Virulence Factor ? - No, not sensu stricto - All Staphylococcus spp. carry an agr - agr is also required for regulation of house-keeping genes Brussels 07
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