RÉSUMÉ
Atopic dermatitis (AD) is an inflammatory skin condition with a childhood prevalence of up to 25%. Microbial dysbiosis is characteristic of AD, with Staphylococcus aureus the most frequent pathogen associated with disease flares and increasingly implicated in disease pathogenesis. Therapeutics to mitigate the effects of S. aureus have had limited efficacy and S. aureus-associated temporal disease flares are synonymous with AD. An alternative approach is an anti-S. aureus vaccine, tailored to AD. Experimental vaccines have highlighted the importance of T cells in conferring protective anti-S. aureus responses; however, correlates of T cell immunity against S. aureus in AD have not been identified. We identify a systemic and cutaneous immunological signature associated with S. aureus skin infection (ADS.aureus) in a pediatric AD cohort, using a combined Bayesian multinomial analysis. ADS.aureus was most highly associated with elevated cutaneous chemokines IP10 and TARC, which preferentially direct Th1 and Th2 cells to skin. Systemic CD4+ and CD8+ T cells, except for Th2 cells, were suppressed in ADS.aureus, particularly circulating Th1, memory IL-10+ T cells, and skin-homing memory Th17 cells. Systemic γδ T cell expansion in ADS.aureus was also observed. This study suggests that augmentation of protective T cell subsets is a potential therapeutic strategy in the management of S. aureus in AD.
Sujet(s)
Eczéma atopique , Infections cutanées à staphylocoques , Staphylococcus aureus , Eczéma atopique/immunologie , Eczéma atopique/microbiologie , Humains , Staphylococcus aureus/immunologie , Enfant , Femelle , Infections cutanées à staphylocoques/immunologie , Infections cutanées à staphylocoques/microbiologie , Mâle , Enfant d'âge préscolaire , Peau/microbiologie , Peau/immunologie , Peau/anatomopathologie , Chimiokine CXCL10/immunologie , Chimiokine CXCL10/métabolisme , Lymphocytes auxiliaires Th1/immunologie , Lymphocytes auxiliaires Th2/immunologie , Cellules Th17/immunologie , Théorème de Bayes , Lymphocytes T CD8+/immunologie , Interleukine-10/métabolisme , Interleukine-10/immunologie , Lymphocytes intra-épithéliaux/immunologie , Antigènes de différenciation des lymphocytes T , Glycoprotéines membranairesRÉSUMÉ
Staphylococcus aureus and Staphylococcus epidermidis are frequently associated with medical device infections that involve establishment of a bacterial biofilm on the device surface. Staphylococcal surface proteins Aap, SasG, and Pls are members of the Periscope Protein class and have been implicated in biofilm formation and host colonization; they comprise a repetitive region ("B region") and an N-terminal host colonization domain within the "A region," predicted to be a lectin domain. Repetitive E-G5 domains (as found in Aap, SasG, and Pls) form elongated "stalks" that would vary in length with repeat number, resulting in projection of the N-terminal A domain variable distances from the bacterial cell surface. Here, we present the structures of the lectin domains within A regions of SasG, Aap, and Pls and a structure of the Aap lectin domain attached to contiguous E-G5 repeats, suggesting the lectin domains will sit at the tip of the variable length rod. We demonstrate that these isolated domains (Aap, SasG) are sufficient to bind to human host desquamated nasal epithelial cells. Previously, proteolytic cleavage or a deletion within the A domain had been reported to induce biofilm formation; the structures suggest a potential link between these observations. Intriguingly, while the Aap, SasG, and Pls lectin domains bind a metal ion, they lack the nonproline cis peptide bond thought to be key for carbohydrate binding by the lectin fold. This suggestion of noncanonical ligand binding should be a key consideration when investigating the host cell interactions of these bacterial surface proteins.
Sujet(s)
Protéines bactériennes , Modèles moléculaires , Domaines protéiques , Staphylococcus aureus , Humains , Adhésines bactériennes/génétique , Adhésines bactériennes/métabolisme , Protéines bactériennes/composition chimique , Protéines bactériennes/métabolisme , Lectines/composition chimique , Lectines/métabolisme , Infections à staphylocoques/microbiologie , Staphylococcus epidermidis/composition chimique , Staphylococcus epidermidis/génétique , Staphylococcus epidermidis/métabolisme , Domaines protéiques/physiologie , Structure tertiaire des protéines , Liaison aux protéines , Staphylococcus aureus/composition chimique , Staphylococcus aureus/génétique , Staphylococcus aureus/métabolisme , Protéines recombinantes/composition chimique , Protéines recombinantes/génétique , Protéines recombinantes/métabolisme , Escherichia coli , Cellules épithéliales/microbiologieRÉSUMÉ
Staphylococcus aureus infections have become a major challenge in health care due to increasing antibiotic resistance. We aimed to design small molecule inhibitors of S. aureus surface proteins to be developed as colonization inhibitors. We identified allantodapsone in an initial screen searching for inhibitors of clumping factors A and B (ClfA and ClfB). We used microbial adhesion assays to investigate the effect of allantodapsone on extracellular matrix protein interactions. Allantodapsone inhibited S. aureus Newman adhesion to fibrinogen with an IC50 of 21.3 µM (95% CI 4.5-102 µM), minimum adhesion inhibitory concentration (MAIC) of 100 µM (40.2 µg/mL). Additionally, allantodapsone inhibited adhesion of Lactococcus lactis strains exogenously expressing the clumping factors to fibrinogen (L. lactis ClfA, IC50 of 3.8 µM [95% CI 1.0-14.3 µM], MAIC 10 µM, 4.0 µg/mL; and L. lactis ClfB, IC50 of 11.0 µM [95% CI 0.9-13.6 µM], MAIC 33 µM, 13.3 µg/mL), indicating specific inhibition. Furthermore, the dapsone and alloxan fragments of allantodapsone did not have any inhibitory effect. Adhesion of S. aureus Newman to L2v loricrin is dependent on the expression of ClfB. Allantodapsone caused a dose dependent inhibition of S. aureus adhesion to the L2v loricrin fragment, with full inhibition at 40 µM (OD600 0.11 ± 0.01). Furthermore, recombinant ClfB protein binding to L2v loricrin was inhibited by allantodapsone (P < 0.0001). Allantodapsone also demonstrated dose dependent inhibition of S. aureus Newman adhesion to cytokeratin 10 (CK10). Allantodapsone is the first small molecule inhibitor of the S. aureus clumping factors with potential for development as a colonization inhibitor. IMPORTANCE S. aureus colonization of the nares and the skin provide a reservoir of bacteria that can be transferred to wounds that can ultimately result in systemic infections. Antibiotic resistance can make these infections difficult to treat with significant associated morbidity and mortality. We have identified and characterized a first-in-class small molecule inhibitor of the S. aureus clumping factors A and B, which has the potential to be developed further as a colonization inhibitor.
Sujet(s)
Kératines/métabolisme , Infections à staphylocoques , Staphylococcus aureus , Adhésines bactériennes/métabolisme , Adhérence bactérienne/physiologie , Fibrinogène/métabolisme , Humains , Protéines membranaires/métabolisme , Protéines recombinantes/composition chimique , Protéines recombinantes/métabolisme , Infections à staphylocoques/traitement médicamenteux , Infections à staphylocoques/microbiologie , Staphylococcus aureus/métabolismeRÉSUMÉ
Colonisation of humans by Staphylococcus aureus is a major risk factor for infection, yet the bacterial and host factors involved are not fully understood. The first step during skin colonisation is adhesion of the bacteria to corneocytes in the stratum corneum where the cornified envelope protein loricrin is the main ligand for S. aureus. Here we report a novel loricrin-binding protein of S. aureus, the cell wall-anchored fibronectin binding protein B (FnBPB). Single-molecule force spectroscopy revealed both weak and ultra-strong (2 nN) binding of FnBPB to loricrin and that mechanical stress enhanced the strength of these bonds. Treatment with a peptide derived from fibrinogen decreased the frequency of strong interactions, suggesting that both ligands bind to overlapping sites within FnBPB. Finally, we show that FnBPB promotes adhesion to human corneocytes by binding strongly to loricrin, highlighting the relevance of this interaction to skin colonisation.
Sujet(s)
Infections à staphylocoques , Staphylococcus aureus , Adhésines bactériennes/composition chimique , Adhérence bactérienne , Fibronectines/métabolisme , Humains , Protéines membranaires , Liaison aux protéines , Infections à staphylocoques/microbiologie , Staphylococcus aureus/métabolismeRÉSUMÉ
Staphylococcus aureus colonizes the skin of the majority of patients with atopic dermatitis (AD), and its presence increases disease severity. Adhesion of S. aureus to corneocytes in the stratum corneum is a key initial event in colonization, but the bacterial and host factors contributing to this process have not been defined. Here, we show that S. aureus interacts with the host protein corneodesmosin. Corneodesmosin is aberrantly displayed on the tips of villus-like projections that occur on the surface of AD corneocytes as a result of low levels of skin humectants known as natural moisturizing factor (NMF). An S. aureus mutant deficient in fibronectin binding protein B (FnBPB) and clumping factor B (ClfB) did not bind to corneodesmosin in vitro. Using surface plasmon resonance, we found that FnBPB and ClfB proteins bound with similar affinities. The S. aureus binding site was localized to the N-terminal glycine-serine-rich region of corneodesmosin. Atomic force microscopy showed that the N-terminal region was present on corneocytes containing low levels of NMF and that blocking it with an antibody inhibited binding of individual S. aureus cells to corneocytes. Finally, we found that S. aureus mutants deficient in FnBPB or ClfB have a reduced ability to adhere to low-NMF corneocytes from patients. In summary, we show that FnBPB and ClfB interact with the accessible N-terminal region of corneodesmosin on AD corneocytes, allowing S. aureus to take advantage of the aberrant display of corneodesmosin that accompanies low NMF in AD. This interaction facilitates the characteristic strong binding of S. aureus to AD corneocytes.
Sujet(s)
Eczéma atopique/microbiologie , Protéines et peptides de signalisation intercellulaire/métabolisme , Staphylococcus aureus/métabolisme , Adhésines bactériennes/métabolisme , Adhérence bactérienne/physiologie , Coagulase/métabolisme , Eczéma atopique/métabolisme , Épiderme , Cellules épithéliales/métabolisme , Humains , Microscopie à force atomique , Peau/métabolisme , Peau/microbiologie , Infections à staphylocoques/microbiologie , Staphylococcus aureus/pathogénicitéRÉSUMÉ
Staphylococcus aureus expresses a number of cell wall-anchored proteins that mediate adhesion and invasion of host cells and tissues and promote immune evasion, consequently contributing to the virulence of this organism. The cell wall-anchored protein clumping factor B (ClfB) has previously been shown to facilitate S. aureus nasal colonization through high affinity interactions with the cornified envelope in the anterior nares. However, the role of ClfB during skin and soft tissue infection (SSTI) has never been investigated. This study reveals a novel role for ClfB during SSTIs. ClfB is crucial in determining the abscess structure and bacterial burden early in infection and this is dependent upon a specific interaction with the ligand loricrin which is expressed within the abscess tissue. Targeting ClfB using a model vaccine that induced both protective humoral and cellular responses, leads to protection during S. aureus skin infection. This study therefore identifies ClfB as an important antigen for future SSTI vaccines.
Sujet(s)
Adhésines bactériennes/métabolisme , Infections cutanées à staphylocoques/microbiologie , Staphylococcus aureus/immunologie , Vaccins/immunologie , Facteurs de virulence/métabolisme , Virulence , Adhésines bactériennes/génétique , Adhésines bactériennes/immunologie , Animaux , Adhérence bactérienne , Femelle , Protéines membranaires/métabolisme , Souris , Souris de lignée BALB C , Infections cutanées à staphylocoques/immunologie , Infections cutanées à staphylocoques/métabolisme , Vaccins/administration et posologie , Facteurs de virulence/génétique , Facteurs de virulence/immunologieRÉSUMÉ
Clumping factor A (ClfA), a cell-wall-anchored protein from Staphylococcus aureus, is a virulence factor in various infections and facilitates the colonization of protein-coated biomaterials. ClfA promotes bacterial adhesion to the blood plasma protein fibrinogen (Fg) via molecular forces that have not been studied so far. A unique, yet poorly understood, feature of ClfA is its ability to favor adhesion to Fg at high shear stress. Unraveling the strength and dynamics of the ClfA-Fg interaction would help us better understand how S. aureus colonizes implanted devices and withstands physiological shear stress. By means of single-molecule experiments, we show that ClfA behaves as a force-sensitive molecular switch that potentiates staphylococcal adhesion under mechanical stress. The bond between ClfA and immobilized Fg is weak (â¼0.1 nN) at low tensile force, but is dramatically enhanced (â¼1.5 nN) by mechanical tension, as observed with catch bonds. Strong bonds, but not weak ones, are inhibited by a peptide mimicking the C-terminal segment of the Fg γ-chain. These results point to a model whereby ClfA interacts with Fg via two distinct binding sites, the adhesive function of which is regulated by mechanical tension. This force-activated mechanism is of biological significance because it explains at the molecular level the ability of ClfA to promote bacterial attachment under high physiological shear stress.
Sujet(s)
Adhérence bactérienne/physiologie , Coagulase/métabolisme , Fibrinogène/métabolisme , Infections à staphylocoques/microbiologie , Staphylococcus aureus/physiologie , Sites de fixation , Phénomènes biomécaniques , Cellules cultivées , Coagulase/génétique , Fibrinogène/génétique , Simulation de dynamique moléculaire , Liaison aux protéinesRÉSUMÉ
Staphylococcus aureus skin infection is a frequent and recurrent problem in children with the common inflammatory skin disease atopic dermatitis (AD). S. aureus colonizes the skin of the majority of children with AD and exacerbates the disease. The first step during colonization and infection is bacterial adhesion to the cornified envelope of corneocytes in the outer layer, the stratum corneum. Corneocytes from AD skin are structurally different from corneocytes from normal healthy skin. The objective of this study was to identify bacterial proteins that promote the adherence of S. aureus to AD corneocytes. S. aureus strains from clonal complexes 1 and 8 were more frequently isolated from infected AD skin than from the nasal cavity of healthy children. AD strains had increased ClfB ligand binding activity compared to normal nasal carriage strains. Adherence of single S. aureus bacteria to corneocytes from AD patients ex vivo was studied using atomic force microscopy. Bacteria expressing ClfB recognized ligands distributed over the entire corneocyte surface. The ability of an isogenic ClfB-deficient mutant to adhere to AD corneocytes compared to that of its parent clonal complex 1 clinical strain was greatly reduced. ClfB from clonal complex 1 strains had a slightly higher binding affinity for its ligand than ClfB from strains from other clonal complexes. Our results provide new insights into the first step in the establishment of S. aureus colonization in AD patients. ClfB is a key adhesion molecule for the interaction of S. aureus with AD corneocytes and represents a target for intervention.