yduosnue Ww souny yduosnue w sou;ny yduosnuew souny yduosnuew sowny -~-y HHS Public Access Author manuseript Nat Rev Microbiol. Author manuscript, available in PMC 2015 October 29. Published in final edited form as: ‘Nat Rev Mcrobiol. 2015 September ; 13(9): 529-543. doi 10.1038/nmicro3521 ‘Staphylococcal manipulation of host immune responses Vilasack Thammavongsa’, Hwan Keun Kim, Dominique Missiakas, and Olaf Schneewind Department of Microbiology, University of Chicago, 920 East 58* Street, Chicago, Ilinois 60637, USA Abstract Staphylococcus aureus, a bacterial commensal of the human nares and skin, is @ frequent cause of soft tissue and bloodstream infections. A hallmark of staphylococcal infections is their frequent recuence, even when treated with antibiotics and sugical intervention, which demonstrates the bbacteritum’s ability to manipulate innate and adaptive immune responses. In this Review, we highlight how 5 aureus virulence factors inhibit complement activation, block and destroy phagocytic cells and modify host B and T cell responses, and we discuss how these insights might ‘be useful for the development of novel therapies against infections with antibiotic resistant strains such as methicillin resistant S| aureus. Apguoximately 30% ofthe human population is continuously colonized with Staphylococcus aureus, whereas some individuals are hosts for intermittent colonization!. S| aureus typically resides inthe nares but is also found onthe skin andi the gastrointestinal tract Although colonization is nota prerequisite for staphylococcal disease, colonized individuals ‘more frequently acquire infections!, Skin and soft tissue infections (SSTIs) are the most frequent disease form ofS aurens, and these infections can jrogzes to bacteremia and invasive disease, ie, bloodstream infection, endocanits or sepsis? Inadelition, 5 aureus can cause pheunonie, osteomyelitis, infectious arthntis, abscesses in many ongen tastes and infections of surgical wounds or prosthetic materials?. Annual attack rates for aureus disease range between 1.3% and vary with age, ethnicity and geographical location of ‘human populations? At elevated risk for staphylococcal infection are low-birth-weight infents, children, elderly and patients with indwelling catheters, endotracheal intubation, Garepaninceo 05. xd Godctcagy st, Sent dass" REGENERON Shannsceubcas, 755 OM Se Mil River Reed, Taxytorm, NY 10591 Competing interests ‘The ers declare conflict of teres as iventars of patents der cence for conmarcial development of Sepiplococes aureus ‘Subject categeries ‘Bologiel scinces / Microbiology / Bacteria Bacterial mane evtson [URE /631/5261410534) Biblogil scinces / Microbiology /Bactra Bacterial pebogenesis [URE 6315261410531) Biologil scinces / Merobiolgy /Bactera /Buctralhostrespase [URE 631/5261410533) ‘Biologuel scinces / Merobislogy /Bactra /Baceriltoxins (URE /631/5261411319) Biological scimces / Microbiology Petogens TURre315261421)yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal Page? medical implantation of foreign bodie, trauma, sugicel procedures, hemodialysis, diabetes snd immonosupgtessive or cancer therapy? A key feature of aureus disease i its recurence, which for SSTI and bloodstream infections, occu for 8-337 of cases. Prior disease does not elicit protection against subsequent 5’ aureus infection? [Neutrophils playa cental put in protecting humans against S aureus infection Staphylococcal entry and replication in host tasues leads to the release of bacterial products (onnyl-peptides,ligoproteins or peptidoglycan) and to damaged tissues that produce inflammatory signals, ie. chemoattractants and cytokines‘. Immune cells perceive staphylococcal products via Tol-ike receptors and G-protein coupled receptors, while cyfokines activate cognate immune receptors, Neutrophils answer this call, extavasate from, blood vessels, and migrate towards the site of infection to phagocytose and all bacteria orto sammobilize and damage the pathogen thnough NETosis~the release of neutropil extracellular taps (NETS) comprinng DNA and antimicrobial peptidest, The importance of xeutrophil in contolling 5 aurens infection has been documented through the study of immune defects. Mutations in genes encoding NADPH oxidase, the enzyme generating bactericidal superorile in phagocytes, cause chronic granulomatous disease (CGD), which is associated with defects in phagocytic killing of S* aureus and frequent infection’. Individuals with inborn errors of STATI/STATS signalling of immune cells are pertubed forlL-17 cytokine pathways, which diminishes mucocutaneous immunity and promotes S. ‘aureus infection®, IL-17-dependent T cell signalling is a key activator of neutrophils and of ‘anti-staphylococcal defenses’. Finally, cancer patients with diminished blood neutrophil counts are highly susceptible to S' aureus infection’. ‘Nevertheless, the vast majority of S, aureus disease occuss in immune-competent individuals without defects in phagocyte fimction. To achieve this, . aureus deploys an arsenal of immune evasive strategies that together prevent phagocytosis and killing by neutrophils Further, the pathogen’s ability to cause recurrent disease implies the presence of ‘mechanisms that effectively block the development of adaptive immune responses. Here, we review recent work on the immune evasive attributes of St aureus, including the subversion of the innate and adaptive immune systems and the killing of immune cells, along with epidemiological features of the conesponing genes. We also discuss how the characterization of bacterial immune evasive factors can have translational impact inthe therapy of eutoimmune diseases or the development of vaccines and immunotherapeutics against 5. aureus infection. Subversion of innate immune responses Neutrophil extravasation and chemotaxis Pro-inflammatory signals promote neutrophil adhesion and extravasation across capillary cendothelia, relying on reciprocal interactions between endothelial receptoxs (P-(E-selectins, ICAML1, hyalwonan) and ligands on neutrophil swfaces (PSGL-1, LFA-1, Mac-1, CD44) Although neutrophils seek to migrate towards bacterial invaders, 5\ aureus can interfere with neutrophil extravasation and chemotaxis through the secretion of staphylococcal superentigen-like proteins (SSLs), phenol-soluble moctulins (PSIMs), chemotaxis inhibitory [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal Page protein of 5. aureus (CHIPS), formyl peptide receptor-like 1 inhibitor (FLIPr) and its homologue FLIPr-like (FLIPr-L) SSLs area family of secreted proteins with structural homology to staphylococcal ssuperantigens!™!?. The ssl genes are aanged as tandem repeats in genomic island a (Gla, 30 proteins with broad functions in host defense against microbes, inflammation, hemostasis and wound repair. Complement is activated via any one of three routes, the classical pathway (CP, antibody and Clq deposition on the staphylococcal surface), lectin pathway (LP, MBL/ [ASPs association with staphylococcal caibohytiates) and eltemative pathway (AP; spontaneous breakdown of the complement protein C3 in serum), which converge in the assembly of C3 convertase (C4b2a for CP and LP and C3oBb for AP). C3 convertase cleaves C3 into 3a and C3b; C3b becomes covalently linked to the staphylococcal surface (opsonization) and C3 is released as @ chemoattractant for phagocytes*”. At high local concentrations of Cb, the C5 convertase cleaves C5 into C5, another chemoattractant, and C5b, whose surface deposition promotes membrane attack complex (MAC) formation?7 MAC is effective at killing Gram-negative bacteria; however, itis not effective for S ‘aureus, which has a thick peptidoglycan layer that prevents access to the bacterial membrane In addition tothe thick peptidoglycan layer, many clinical S. aureus strains express genes for the production of one of two types of capsular polysaccharide, type 5 or type 82°. in wtro ‘hagocytosis assays suggested that capsule expression protects staphylococci from ‘eutrophil phagocytosis in the presence of opsonins and that capsule expression contributes to the pathogenesis of 5 aureus infection in mice®. Capsule-induced protection from phagocytosis maybe strain specific, as capsule mutations in other 5 aureus isolates do not affect the pathogenesis of bloodstream infections in mice?S. USA300, the cuent pandemic clone of community-acquired MRSA infections, carries a mutation that abrogates capsule expression*® (Table 1), [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal Pages S aureus secretes several proteins that interfere withthe deposition of complement on the bacterial surface (Fig. Ib). Aureolysin, a secreted Zn-dependent metalloprotease, cleaves C3 to generate functionally active C3a and C3. Complement factors I (f) and H (fH) degrade orbind C3b, which prevents its accumulation on the staphylococcal surface *2. The aureolysin gene, aur, is polymorphic and specific alleles are associated with different ‘aureus lineages. Anin vivo phenotype for S aureus aur mutants has not yet been described ‘Staphylococcal complement inhibitor (SCIN) associates with and inhibits C3 convertase (C30Bb), thereby preventing the production of C3a, C3b and C5a and interfering with complement activation*?(Fig. 1b). The structural gene for SCIN, scn, is also located on hlb- converting phages together with chp and sak (staphylokinase, see below). Two polymorphic homologs of SCIN, designated SCIN-B and SCIN-C, are encoded by genes in the IEC2 locus, scm as well as senBlscnC are found in many, but not all, human clinica isolates (Box 1),SCIN, SCIN-B and SCIN-C associate with C3 convertase from humans but not with that of other vertebrates*? ‘The genes encoding extracellular fibrinogen-binding protein (Efb) and its homologue, ‘extracellular complement-binding protein (Ec), are also located on IEC2. Both Efb and Ec bind C3d, a cleavage product of C3b that activates innate and adaptive responses by nding to complement receptor 2 (CR2), and inhibit C3bBb and the C5 convertases! 4(Fig. 1b) Ech associates with both fH ancl C3b to facilitate the complement inhibitory atrbutes of factor H'®. Efp also binds fibrinogen and prevents fibrinogen interaction with aMIf2, an integrin on neutrophils that activates proinflammatory responses, as well as fibrinogen- rediated platelet activation*®#” Efy and Ecb inhibitory activities have been observed for ‘human as well as mouse convertases and fibrinogen. In the mouse intravenous challenge model, the S' aureus Aefb Aech mutant displayed reduced time-to-death and increased survival as well as diminished abscess formation in organ tissues*®, The ecb gene is found in all $ aureus genomes sequenced to date, whereas ¢f is found in many, but not all human clinical isolates! ‘SSLs also interfere with complement activation and phagocytosis. For example, SSL7 binds ‘human IgA and complement C5, interfering with IgA binding to FeaRI binding an the ‘roduction of C5a and the oxidative burst of phagocytes in vitro; the in wvo contributions of SSL1 towards S aureus pathogenesis are not known'®. $SL10 binds to human and non- ‘human primate IgGl, but not to immunoglobulins of lower vertebrates, and inhibits IgGl binding to Fey receptors and the in vttro phagocytosis of IgGl-opsonized bacteria by jmmune cells!#1 ‘Staphylococcal binder of immunoglobulin (Sbi) is @ secreted protein with two ‘immunoglobulin binding domains (IgBDs, designated Sbi-I and Sbi-l), which are triple telicel bundles that associate withthe Fey-domain of human and vertebrate immunoglobulin (Box 2). Sbi- and Sbi-II interfere with Clq binding to immunoglobulin and block the classical complement pathway*™*%(Fig. 1b). The SbilIl and Sbi-V domains associate with C3 and factor H to form tripartite complexes that inhibit the eltemative pathway (Fig 1b). The sbi gene is located inthe sbi/hlg locus ofthe core genome of el isolates"? Staphylokinase forms enzymatically active complexes with plasminogen, cleaving fibrin, [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal Pages defensins, human IgG, C3b and iC3b on bacterial surfaces, thereby blocking complement activation’®59(Fig. 1b). Collagen adhesin (Cna), e surface protein expressed! by some S ‘aureus isolates, binds Clq and interferes with CP activation, blocking the association between Clq and Cir? (Once phagocytosed, staphylococci are exposed to a variety of toxic products that kill and degrade the engulfed bacteria’ antimicrobial peptides, reactive nitrogen (NO), reactive oxygen species (ROS - hytrogen peroxide, superoxide, hytroxyl radicals), cell wall, Ihytholases, and proteolytic enzymes*. However, S aureus has evolved a number of strategies to survive inthis environment (Fig. 1c). Peptidoglycan acetylation (OatA), D- alanyiation of teichoic acids (DIABCD), and lysyi- or alanyl-phosphatidyiglycerol synthesis (MprF) provide for staphylococcal resistance against lysozyme- and antimicrobial-peptide ‘mediated killing by blocking enzyme (lysozyme) or peptide binding to its envelope target®©2, Staphyloxenthin, a carotenoid pigment synthesized by ell aureus isolates? ‘rovitles resistance against hytrogen peroxide/hydroxy/ radicals, the bactericidal compounds of neutrophils (this is not the case for CC75 isolates but we consider these to belong to a separate species, Staphylococcus argenteus (Box 1)). Similan, two superoxide cismutases (SodA and SodM), fulfill overlapping functions in eliminating neutrophil ssuperoxide®, whereas catalase (KatG) and alkyihyeropsroxide reductase (AhpC) protect staphylococci against hytrogen peroxide®® In response to nitrosative stress, S aureus expresses flavohemoglobin (Hp), which etoxifies nitric oxide, and L-lactate dehydrogenase, which maintains redox-hemostasis anc survival within neutrophils by producing L-lactate®? In addition to its role in inhibiting complement activation, Eap and two structural homologs, EapHil and EapH? (which do not inhibit complement) promote S: aureus survival by inhibiting neutrophil serine proteases (lastase, cathepsin G and proteinase 3)6®. In the inevenous mouse challenge model, the 5 ‘aureus Aeap mutant displays @ moderate virulence defect®6, however the Aeqp Aeap HI ‘Aeqp#? mutant displays reduced bacterial load and increased mouse survivalS® ‘Staphylococcal killing of host cells In addition to its ability to inhibit phagocyte-mediated killing, $ curens also manipulates innate immune responses by inducing the killing of innate immune cells via PSMs and clifferent toxins (Fig. Id). The PSMa locus encodes psmal-4, wherees PSMp encodes ‘psmpl and psm/2. psmfl is found in only some, but not all S aureus strains!?, Peptides similar o PSMal-4 and PSM are expressed by Staphylococcus epidermis, a commensal of the human skin that cannot cause abscess lesions or bloodstream infections in immune competent individuals, Mutations that delete pmal-é and psmyll-2 interfere within vtro biofilm formation of S¢ aureus mutants and with the expression of virulence factors, including a-hemolysin?®. 5 aureus Apsma-4 mutants are attenuated in the mouse bloodstream infection model®, @ phenotype that maybe due to defects in biofilm formation, ‘virulence gene expression and/or contributions of PSMal-4 towards lysis of immune cells, resumebly via membrane insertion and pore formation”? [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal f-bamel pore-forming toxins (j-PFT) are secreted as soluble monomers and, upon association with receptors on cell surfaces, assemble into multimeric pore structures, ‘penetrating the lipid bilayer to invoke alterations in the physiology of injured cells or their outright lysis”! q-hemolysin Hla), the prototype [PFT of S' aureus, is encoded by the hla gene, which is located within IEC2. Although conserved among all S\ aureus isolates, some lineages of * aureus carry a nonsense mutation that blocks ila expression”?. Hla binds to its receptor, ADAMIO, and assembles into @ heptameric pore; through the metalloprotease activity of ADAMIO, Hla modulates the function of immune cells, including neutrophils, or triggers lysis of epithelial cells”>7*. $\ aureus hla mutents display defects in disease severity ‘in mouse models for lethal pneumonia, bacteremia and SSTI, albeit that hla is not required for the establishment of 5 aureus abscess lesions”S7”. Based on ADAMIO expression on the surface of myeloid cells, organ epithelia and the vascular endothelium, Hla causes @ wide spectrum of global as well as organ-specific changes that affect physiological host responses to S aureus infection”* Several other f-PFTs secreted by St aureus are designated leukocidins (Fig. 1d), Following leukocidin association with receptors on myeloid cells and exythrocyies, these toxins assemble fiom two different subunits (F an S) info an octameric pore structure”®, All $ ‘qureus stains produce at least three leukocidins, HigAB, HigCB and LukAB (LukGH), whereas other strains may also secrete Panton-Valentine leucocidin (PVL) and LukED or LukMF?? (Box 1). The genes encoding iukAB (IukGH) are located immediately adjacent to ‘hb, whereas those encoding y-hemolysin (hg4BC) are part of the si/hlg locus. LukAB (LukGH) binds to the I domain of human, but not mouse, CD11b (integrin cM) on myeloid cells®_ Purified LukAB (LukGH) can trigger human neutrophils to release NETS that, at least temporarily, ensnare staphylococci®!, LukAB (LukGH) has also been reported to promote S' aureus escape fiom the phagosome of neutrophils®?, Purified HigAB y- ‘hemolysin, but not purified HigCB y-hemolysin, is able to lyse human and rabbit ed blood, cells HigAB binds chemokine receptors CKCRI, CXCR2 and CCR2, whereas HigCB utilizes complement receptors C5aR and C5L? to associate with target cells. Following staphylococcal inoculation into human blood, hlgABCis upregulated 34-145 fold®® and the S aureus AhlgABC mutant displays reduced survival, presumably because HigAB and [HIgCB promote release of iron-compounds from erythrocytes, thereby enabling bacterial acquisition of this essential nutrient®?. Both purified HigAB and HigCB promote lysis of| xeulrophils, monocytes and macrophages from humans as well as non-human primates, and toa lesser degree rabbits and mice®?. Ine mouse intravenous challenge model, animals infected with a St aureus ukGH (IukAB) mutant displayed increased time-to-death and survival, Using subcutaneous inoculation in mice or rabbits, the S| qurens hukGH (IukAB) mutant did not display defects in skin abscess formation®S. The AllgAB mutant displayed a ‘virulence defect in the intraperitoneal challenge model in mice® LikEDis present in the GI locus of ebout 10% of clinical 5. aureus isolates"? (Box 1) Purified LukED triggers lysis of macrophages, dendritic cells and T lymphocytes from many different vertebrates, asthe toxin binds to the chemokine receplors CCR5, CXCRI and CXCR2°7#8. For 5 aureus Newman, which harbors GIp, the AlukED mutation increased the time-to-death and survival of mice following intravenous challenge with mutant [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal Pages staphylococci®. Panton-Valentine leukocidin, also designated PVL or LukPV, is secreted byS aureus lysogenized with PVL bacteriophage, PVL binds to the C5aR on neutrophils, ‘monocytes and macrophages but is activity is restricted towards human and rabbit cells? Byvvirtue of binding C5aR, PVL not only exerts its lytic activity on target host cells but can also facilitate the priming of human PMNs by proinflammatory stimuli, for exemple formyi- ‘peptides, Injection of punfied recombinant PVL leads to increased immune cell recruitment and increased architectural destruction of the lung, owing to toxin-mediated recruitment and subsequent lysis of immune cells®. Only about 2% of S| aureus isolates secrete PVL, however community-acquired MRSA isolates fiequently harbor PVL becteriophage ancl PVL expression is also associated with necrotizing eumonia®! 5. aureus AlukPV variants display defects in the pathogenesis of skin and soft tissue infections and lung infections in bits, but not in mice, which seems to be due to neutrophi-mediated inflammatory responses and tissue distruction’®9?. IukMF, genes for another bacteriophage-encoded leucocidin, are found in S aureus isolates associated with bovine masttis!? ‘Staphylococcal agglutination Coagulation, the conversion of fibrinogen to a crosslinked fibrin meshwork by activated thrombin, is an innate defense of all vertebrates, which immobilizes microbial invaders and attracts immune cells for phagocytic clearance of bacteria. Therefore, every successful bacterial pathogen must evolve mechanisms for escape from fibrin entrapment anc subsequent phagocytosis by infiltrating immune cells. A hallmark of all. aureus isolates is the secretion of two coagulases, coagulase (Coa) and von Willebrand Factor-binding protein (AWop)®3. Coa and vWbp associate with prothrombin, a zymogen, to generate enzymatically active staphylothombin, which cleaves the A ancl B peptides of fibrinogen to generate fibrin fibrls*(Fig. 2). As staphylothrombin does not cleave other substrates of thrombin, it avoids the activation of clotting and inflammatory factors that ordinarily accompany fibrin polymerization®S. Staphylothrombin activity is not subject to feedback inhibition through host antithrombin, However, staphylothvombin is blocked by dabigetran and other direct thrombin inhibitors of the same family®®. The staphylothnombin-generated fibrin meshwork protects 5 aureus from phagocytes and contributes to the formation of staphylococcal abscess lesions and lethal bacteraemia in mice®”. Activation of prothrombin is mediated by the N-terminal D1-D2 domain of Coa and is blocked by specific antibodies, which provide rotection fiom 5. aureus bloodstream infection inthe mouse model®®. Pethaps owing to ‘purifying selection, coc is one of the most variable genes in the core genome of S* aureus with >50% sequence variation inthe coding sequence for its D1-D? domains and 14 distinct isoforms (Teble 1)%2.vWbp also has a conserved D1-D? domain for association with prothrombin, but this complex generates fibrin at a reduced rate and contributes to abscess formation without affecting staphylococcal escape fiom phagocytosis®®. The gene encoding ‘vWop, vb, displays limited sequence variability? 5 qureus agglutinates with congulase-derived fibrin fibrils, which requires clumping factor A (CIEA), a glycosylated, sortase-anchored surface protein whose immunoglobulin-like comains bind to the C-terminal end of the y-chain in fibrinogen/fibrin (D domain)®6100,01(Fig. 2). Thus, CIEA acts synergistically with CoahWbp in protecting [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal Pages staphylococci from opsonophagocytic killing and Ac/fA mutants display defects in the ‘pathogenesis of lethal bloodstream infections in mice Four other sortase-anchored surface proteins use their immunoglobulin-like domains to bind fibrinogen/fibrin: CHB (c-chain), fibronectin binding protein A (FnBPA) and B (FnBPB, C- terminal end of y-chain), and bone sialoprotein binding protein (Bop(SaiE isoform, o- chain)!°°(Fig. 2), These surface proteins display functional redundancy for the 5: aureus ‘agglutination pathway and contribute to the pathogenesis of bloodstream infections!2 Purified, recombinant SSL10 has also been reported to bind human fibrinogen and fibronectin as well as porcine prothrombin and factor Xa*!. The association of SSL10 with prothrombin and Xa occurs via the Gla (y-carboxyjic acid) domain and interferes with calcium-activated blood clotting but not with staphylotivombin (congulase)-mediated fibrin formation’ (Fig. 2). ss!7 and ssl 0 are not found in ell $ aureus isolates!? Staphylokinase activates human, but not murine, plasminogen andl may solubilize coagulase- induced fibrin deposits, thereby aiding S° aureus in generating purulent lesions for issemination to new hosts (Fig. 1b). Of note, binding of surface proteins by fibrinogen! fibrin deposits has also been reported to influence staphylococcal interference with platelet ‘aggregation and innate immune functions!°25. Tn addition to their role in binding to fibrin! fibrinogen, some surface proteins have been reported to bind additional host ligands, at least in vitro, Binding to these additional host ligends, which include complement factor I (by the surface protein CIEA), fibronectin (by the surface proteins FnBPA and FnBPB), keratin 10, (by the surface protein CHB) and loricrin (by the surface protein CIfB), may contribute to staphylococcal immune evasion, invasion of host cells or colonization of squamous, cepithelia!®?_frbpA and fnbpB, which are components of the 5° aureus core genome, display sequence polymorphisms and seven isotypes with discrete antigenicity have been described! Box 1), Adenosine/deoxyadenosine signalling and NETosis Adenosine is a potent mediator of immune responses and, under physiological conditions, is synthesized following hypoxia, exposure to reactive oxygen species (ROS) and cell lysis, associated with tissue damage. Adenosine elicits its biological effects bybinding to one or rmore of four G protein coupled receptors!”, Al A2A, AB and A3. Adenosine receptor interaction triggers ant-inflammatory signalling cascades that inhibit platelet aggregation, ‘eutrophil supsroxide burst, neutrophil degrantlation, T cell activation ancl release of the cytokines IL-1 and IL-10™. $ aureus increases the concentrations of extracellular adenosine during infection by expressing AdsA, a sortase-anchored protein that catalyses the cephosphorylation of adenosine mono-, di- and triphosphates!°°(Fig. 3a). Both ex vtvo andl during mouse infection, the AadsA mutation increases killing of staphylococci by blood. neutrophils, while decreasing extracellular adenosine!°°, Thus, AdsA-mediated synthesis of| adenosine promotes survival of 5: aureus within neutrophils, presumably by inhibiting superoxide burst and/or degranulation!®°. Further, adenosine decreases MHC-II expression ‘in macrophages and dendritic cells and dampens IL-12 production, a pivotal stimulus for ‘Thl-type immune responses!” Staphylococcal enhancement of adenosine production may [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Tamumaavengss et Page 10 therefore interfere with T cell effector mechanisms and adaptive immune responses in infected hosts!0? Adsh activity aso modulates immune responses following the degradation of NETs. Duzing bloodstream infection in mice, S| aureus disseminates to many different orgen tissues to establish abscess lesions. These lesions are composed of a bacterial nid, designated as the staphylococcal abscess community (SAC), encased within a preudocepsule of fibrin deposits, and sunounded by layers of immune cells°”. In spite of lange numbers of infiltrated neutrophils, mice ere unable fo eliminate staphylococci from abscess lesions and eventually succumb to persistent infection. Although neutrophils use NETosis to entangle staphylococci, NETS are degraded by staphylococcal nuclease (Nuc) and thereby fail to exert bactericidal activities! (Fig. 3b). Nuclease digestion of NETs releases 5? and 3” ‘monophosphate nucleotides that are converted by AdsA into deoxyedenosine (dAdo) 4(Fig. 3). dAdo production triggers caspase-3 induced apoptosis of macrophages and prevents ‘phagocyte entry into the SAC, the core of staphylococcal abscess lesions, thereby promoting bacterial survival within the lesion! ulation of adaptive immune responses B cell responses 5 aureus is capable of manipulating B cell survival and function, especially via the activity of SpA, which is sortase-anchored surface protein with high affinity for vertebrate ‘immunoglobulin, including human IgA, IgD, IgG1-4, IgM and Ig? Spa is initially deposited in the staphylococcal envelope and subsequently released by cell wall hytolases (LytM"3_ pa is expressed by ll clinical $ aureus isolates, the immunoglobulin binding domains are conserved in the genomes of these isolates but region X, the cell wall spanning domain of SpA, is « highly polymonphic sequence!*15 Box 1) ‘The immunosuppressive attributes of SpA have been ascribed to two distinct binding activities: association with the Fey domain and withthe Feb domains of entibodies!16217 ‘SpA binding to the Fey domain of IgG blocks phagocytosis of staphylococci", whereas ‘SpA binding to Feb and crosslinking of V3 clan IgM promotes B cell superantigen activity!9(Fig. 4a). Of note, SpA binds specifically o Vix3 clan IgM antibodies, which mediate the predominant antibody responses to infection and immunization, but not to other clan antibodies. In the inkavenous challenge model ofS. aureus infected mice, spa ‘expression suppresses antibody responses against many different staphylococcal antigens ‘and provides antiphagocytic attributes, promoting staphylococcal survival in blood Infection of mice with 5: aureus spa variants that cannot bind immunoglobulin is associated with attenuated disease and with antibody responses against many different antigens that can protect animals against subsequent lethal challenge with other S. aureus isolates! Mice harbor a limited repertoire of V23* B cells, whereas humans possess lage populations of Vy" B cells, yetboth species cannot develop SpA-neutralizing antibodies during infection’. 5 aureus infection in humans triggers expansions of V3 idiotypic plasmablests (-90% of blood plasmablasts), whose antibodies (B cell receptors) associate ‘vi their Feb domains with SpA but do not display pathogen specific binding activities!” [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal Pg (Fig. 4), When mice are treated with purified SpA, crosslinking of V3 clonal B cells triggers proliferation and apoptotic collapse of expanded populations of B cells!2? It is not clear, however, whether apopiotic collapse of expanded lymphocyte populations occurs during S¢ aureus infection in mice or in humans. Non-toxigenic Sp, designated SpAxacaa, Was engineered by substituting twenty amino acid residues essential for its association with Ig Fey and Feb'3. Although SpAsacaa has twenty amino acid substitutions, this antigen elicits antibodies that neutralize SpA when injected into animals!?3. The SpAxxaa-detived polyclonal entbodies promote phagocytosis of staphylococci and display adjuvant attributes by suppressing staphylococcal B cell superantigen activity and promoting humoral immune responses against a wide spectrum of antigens!2, Studies with mouse monoclonal antibodies (SpAxacaa-MAbs) comoborate this concept’? T cell responses Outlook ‘Staphylococcal T cell superantigens bind to MHC class II molecules on the surface of antigen-presenting cells, providing entigen independent crosslinking with T-cell receptors on T helper cells! (Fig. 4c). 5. aureus strains have been shown to express 23 diferent enterotoxins and T cell superantigens!25. Three superantigens are most frequently associated with human disease - toxic-shock-syncrome toxin I (TSST-1), staphylococcal enterotoxin B (GEB) and SEC - each providing high-affinity interactions with distinct subsets of V chain Tell receptors! In humans with toxic shock syndrome, S aureus secretion of TSST-1 or other enterotoxins trigger expansions of cognate T cell populations, up to 30% of blood lymphocytes and non-specific release of cytokines, preventing a focused adaptive immune response?” Depending on the site and severity of S| aureus infectionlintoxication, superantigen-mediated activation of T cell responses maybe associated with cytokine storms and toxic shock syndrome pathology!2®. Staphylococcal superantigens are also thought to interfere with antigen-specific proliferation of T cells and with antibody responses against specific subsets of staphylococcal antigens, including staphylococcal superantigens!?9. [tis not yet known whether superantigens play critical role in the suppression of T cell responses in mice that are observed during S¢ aureus bloodstream infections! S aureus can also manipulate T cell responses by promoting T cell lysis. For example, §- toxin (Hid or S-hemolysin), a member of the PSMa family, can lyse T cells!" and has also been reported to trigger mast cell degranulation, which could be a key factor in the ‘exacerbation of S aureus infected atopic dermatitis lesions, where histamine release is otherwise triggered by antigen-induced crosslinking of IgE bound to FceRI receptor!#?, Hid is encrypted within the agr-regulated RNA III molecule, the regulatory arm of staphylococcal quorum-sensing 2 S aureus strains secrete a plethora of immune evasive molecules and, when placed under selective pressure, acquire mobile genetic elements with additional factors so that staphylococci meet the demands for invasion of host species and replication in specific [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal rp 2 ‘anatomical niches!3. These strategies are accomplished through factors that block phagocyte chemotaxis, complement activation, phagocytic upake and oxidative ling, often redirecting host defenses suchas fibrin formation o: NETosis to favour pathogen plication Staphylococcal infection is als associated with pertubations of edaptive responses, including the disruptive proliferation of Band T cells, which prevents the establishment of rotectve immune responses. Why do S. aureus isolates acquite so many different immune cvasion factors when other bacterial pathogens make do with only small number? -Addiessing this question, one should consider that aureus anti if-long asociation with ts human hosts, colonizing and aeteratively invading large segments ofthe population ‘We know of no other bacteval pathogen able to sustain snularfestyle. Staphylococcal capeblities of causing recuent and reitemative infections robebly rely on two mechanisms Fist, aureus manipulation of B and T cell esponses mustbe successful, as increased age snot associated with decreased incidence, Second, mobile genetic elements enable scquisition endor exchange of immune evasive titsbetween S aureus sins and orizonfal gene transfer may implement disease in individuals who did mount succesful ‘mune responses aguinst imine evasion determinants. I presse o predict the future of 5S aureuss it evolves with population whose lifespan continues to increase, we would forecast more invasion by mobile genetic elements and more inuune evasion determinants Considering the formidable weapons of the pathogen against the host’s immune defenses, development of vaccines against 5. aureus isa daunting task. Conventional approaches for ‘vaccine development follow the mantra of eliciting specific antibodies that trigger pathogen [ulling in vttro andl disease protection in preclinical (animal) models of staphylococcal disease. The simplest means of achieving these goals are whole-cell vaccines, either killed or attenuated preparations. Indeed, autologous vaccines for individuals with recuurent 5 ‘aureus infection have been practised for many years! This approach can elicit pathogen- specific antibodies; however, it has not been demonstrated to raise protective immunity! Live-attemuated vaccines have been studied in animal models with variable success but not in humans. [fone considers thatthe immune-evasive strategies are often species specific, it seems risky to derive claims on human protective immune responses against 5. aureus from experiments with animals. Whats tru for whole-cell vaccines certainly applies to subunit ‘vaccines. Antibodies against capsular polysaccharide, CAA, IsdB anc lipoteichoie acid bind to the surface ofthe pathogen, which enables phagocyte-mediated killing in vttro ancl ‘provides protection from infection in specific animal models, however, the comesponding ‘vaccines or antibodies didnot achieve efficacy in clinical rials}952. Tt occued to us that in vitro essays for phagocytic killing ofS. aureus often do not consider key evasion strategies of the pathogen and the corresponding defenses of humans. We believe this canbe addressed with Lancefield’s assay for antibody mediated killing of bacteria in fresh anti- coagulated human blood™”, The Lancefield assay can also be used for prospective studies in Inumans, assessing antibody titers, status of immunity (bactericidal activity of blood) andl probability of disease. This approach may identify criteria for protective immunity, stratify patients at risk for $ aureus disease and guide applications for immune-therapy or ‘vaccination that reduce the incidence of disease. Previous work tageted S. aureus B cell and cell superantigens as vaccine antigens to enable the development of broad spectrum immune responses during each encounter of the host with tis pathogen’23.4®, Another [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnuew sowny yduosnuew sowny osnue jy JOyINy du! osnue | JOUINY qdu ‘Toummarvengye etal Ap D ‘promising approach exploited the structural relatedness of immune evasion factors to target rultiple molecules with entbodies that recognize conserved structural features!*? (Box 2) Nonetheless, the efficacy of these approaches has not yetbeen assessed in human clinical trials Display items Box 1 Variability of Staphylococcus aureus immune evasion determinants Genome sequencing of Staphylococcus aurens isolates from humans and animals has ‘ovitled insights into the origin, diversification and spread of the pathogen. Over the ‘st 10,000 years, 5 aureus evolved as colonizer and pathogen of humans anc their lifestock", generating lineages with unique genetic traits and discrete host ranges!*2 Staphylococcal evolution was accompanied by the loss of genes encoding the CRISPR- cas system, which protect the genome against bacteriophage and mobile genetic elements. 5 aureus relies on horizontal gene transfer mediated by these elements for adaptation, and preserves its identity through restriction modification systems and satellite phage-encoded pathogenicity islands that block bacteriophage replication’? ‘When placed under selection in different hosts, S aureus acquires mobile genetic clements that contain genes for antibiotic resistance, immune evasion and adhesion to specific anatomic niches. Analysis of large genome datasets described the core genome, which is common to all & aureus isolates, and found that these core genes contribute to colonization, tissue invasion, establishment of abscess lesions, dissemination, immune evasion and the pathogenesis of reiteative disease ®. Variable genes are associated with ‘S aureus colonization or invasion of specific host species or maybe present in subsets of strains associated with increased virulence or specific disease, for example enterotoxin- mediated gestroenterits®?. As a rule of thumb, capsular polysaccharide and cell wall- anchored surface proteins are components of the core genome and contribute to immune evasion by synthesizing adenosine (adenosine synthase A), binding fibrinogen or fibrin (clumping factors A and B and fibronectin binding proteins A and B) and ‘immunoglobulin (staphylococcal protein A). Several secreted proteins are components of the core genome: proteases cleaving host factors (aureolysin and staphopain), coagulases activating prothrombin (coagulase and von- Willebrand factor binding protein), toxins lysing immune cells (y-hemolysin ABC and leukocidin AB), inhibitors of host proteases (Staphylococcal extracellular adherence protein and its homologues) and phenol-soluble ‘odulins, peptides that pertu host cell membranes and trigger neutrophil chemotaxis, Genetic determinants that interfere with neutrophil chemotexis, phagocytosis, complement activation, promote lysis of immune cells or activate T lymphocytes often represent constituents of the variable genome. Table 1 summarizes core genome and ‘variable immune evasion factors contributing to staphylococcal disease pathogenesis. Isolates of the CC75 lineage are predominantly found in the South-West Pacific and were originally isolated from superficial skin lesions of individuals from the indigenous communities of Australial*. C75 strains lack the staphyloxanthin gene cluster, retain the CRISPR.cas system and lack pathogenicity islands (SaPls), yet ae endowed with [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnuew sowny yduosnuew sowny osnue jy JOyINy du! osnue | JOUINY qdu ‘Toummarvengye etal ripe genomic islands q and fa unique coa gene andl a unique spa sequence type, which are ¢lements important for staphylococcal evasion from innate and adaptive immune responses" 4, These strains, withthe species designation Staphylococcus argenteus, may represent an early and terminal branch in the development of aureus in which mobile genetic elements were not incorporated into the genome because the retained CRISPR- cas system prevented horizontal gene transfer! Box2 ‘Structural features of immune evasion factors Crystallographic analysis of S| aurens immune evasion determinants revealed five discrete structural domains that enable specific interactions with the host’s immune system: oligonucleotide-binding (OB) fold, p-grasp domain, triple-helical bundle (THB), P-pore forming toxin (PFT), and immunoglobulin-like fold (Ig). Varying the amino acid sequence for these domains has created panoplies of ligands that interact with the defence molecules of infected hosts at the places that matter most**5. ‘Thus, the study of ‘5 aureus immune evasion factors laid bare the most intricate workings ofthe human ‘immune system and identified new avenues forthe therapy of autoimmune and inflammatory diseases. Examples for immune evasion factors with OB fold and p-grasp domains include the staphylococcal T cell superantigens (SEA, SEB, SEC1-3, SED, SEE, ‘SEG, SHE, SEI, SEI, SEK, SEL, SEM, SEN, SEO, SEP, SEO, SER, SEU, TSST-1/2 and ‘Se1X) and the staphylococcal superantigen-like family (SSL1-13)!25, CHIPS, FLIPr, FLIPt-L, Eap, EapHll, and EapH? have f-grasp domains but not an OB fold®®146 Immune evasion factors with triple helical bundles include Ecb, Efo, SpA, Sbi, SCIN, SCIN-B and SCIN-C”, whereas HigABC, LukAB (LukGH), LukED, LukMF, PVL and Hla are members of the f-PFT family”®. Surface proteins with IgG-like domains include ‘the immune evasion factor CICA and its relatives CMB, FnBPA and FnBPBY®? (Table 1) Acknowledgements Wark on saplococce mame evasion was supperted by gs fom the Netional stu of Allergy and Infections Diseases AID38807 (0.5), AIOS2747 (0.5) md ANL10037 (DM). V.T admowledges suppor from the ‘Americ Heat Association (P574580023). We apologise to authors whose wor was eihernotreferenced ot discussed ovmngto spice consents Biographies Vilasack Thammavongsa was educated atthe University of Michigan, Ann Arbor, where he received graduate training in immunology in the laboratory of Professor Malini Raghavan. He then undertook a posttloctoral fellowship in bacterial pathogenesis under the guidance of Professor Olaf Schneewind atthe University of Chicago, studying the immune evasive properties of Staphylococcus aureus. He is curentlya scientist at REGENERON Pharmaceuticals in the Department of Discovery Sciences (Target Information), developing ew therapies for human diseases [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal Pig is Hien Keun Kim eamed his PhD. in the Department of Microbiology at the University of Chicago. During doctoral training in the laboratory of Professor Olaf Schnee wind, his thesis focused on identifying, characterizing and developing bacterial antigens into vaccines against Staphylococcus aureus. Hwan Keun Kim now investigates the immune-modulatory mechanisms of 5 aureus in preclinical models for staphylococcel disease Dominique Missiekas was educated at the University of Montpellier, France and received a PhD. in enzymology fiom the University of Paris (Orsay), studying in the laboratory of Professor Jeannine Yon, Following fellowship training with Professor Costa Georgopoulos at the University of Utah (Salt Lake City, USA) and the University of Geneva, Switzerland, Dominique Missiekas worked at the CNRS in Marseille, France and was subsequently appointed Assistant Professor of Microbiology at the University of California, Los Angeles. In 2001, Dominique Missiakas moved to the University of Chicago, where she is curently Professor in the Department of Microbiology. Dominique Missiakas’ laboratory studies staphylococcal agglutination and type VII secretion as well as S-layer assembly in Bacillus anthracis. Olaf Schneewind was educated at the University of Cologne, Germany Following postdoctoral fellowship at the Rockefeller University in the laboratory of Professor Vincent A Fischetti, he joined the faculty at the University of California, Los Angeles. In 2001, Olaf ‘Schneewind moved to the University of Chicago, where he founded the graduate program in ‘Microbiology (Committee on Microbiology) and the Department of Microbiology, He is currently the Louis Block Professor and Chai of the Department of Microbiology. Olaf ‘Schneewind’s laboratory studies molecular mechanisms of disease pnthogenesis for several ricrobes, including Bacillus anthracts, Staphylococcus aureus and Yersinia pestis Glossary Recurrence ‘The propensity of Staphylococcus aureus infections to reoccur when surgery and/or antibiotic therapy are initially effective Abscess ‘The pathological product of S aureus infection, harboring a staphylococcal abscess community within a pseudocapsule of fibrin deposits that is surounded by layers of infiltrating immune cells destroying physiological organ tissue. Amaphylatoxin Protein fragments generated during complement activation of C3a ‘and Ca that trigger immune responses via C3a and Ce receptors con immune ces Dabigatran. ‘A small molecule that directly binds and inhubits thrombin as well as staphylothvombin, the complex formed between Coa or vWop and prothrombin Fab domain ‘The portion of antibodies dedicated to antigen binding. Fey domain ‘The portion of antibodies dedicated to Clq complement and Fe receptor activation. [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal Autologous Opsonization Fibrinogen, ‘Vag clan IgM FeaRI ‘Leukocidins Factor Xa Sortase Hiotype Fibronectin Core genome REFERENCES ‘Molecules that crosstink B cell receptors (IgM) or T cell receptors ‘and major histocompatiblity complexes to trigger lymphocyte proliferation, thereby diverting edaptive immune responses Immature B cell inblood that secretes antibodies ‘Staphylococcal superantigen that crosslinks MHC class II molecules ‘and T-cell receptors, thereby triggering T cell proliferation, nergy ‘and cytokine storms, ‘A whole-cell killed S aureus vaccine administered to an infected, individual that was derived fiom the patient's isolate Deposition of complement components on bacterial surfaces to ‘promote recognition, phagocytosis and killing by host phagocytes. An ebundant glycoprotein of vertebrates that, when cleavedlby thrombin (factor Ila) or staphylothomibin, self-essemibles into fibrin clots. [Immunoglobulin M derived from one of thee clans of Viz genes whose products provide the scaffold for the antigen binding determinants of antibodies. ‘The immunoglobulin A Fe receptor, which regulates mucosal ‘immune responses in humans. Bacterial secreted toxins teugeting white blood cells (leukocytes) for destruction ‘The activated serine protease, also designated thrombokinase, which cleaves prothrombin o activate the cloting cascade of vertebrates. ‘The bacterial transpeptidase responsible for anchoring swface ‘proteins to the cell wall envelope. A setof epitopes on the V region ofan antibody molecule High molecular weight glycoprotein of the extracellular matrix of ‘vertebrates that associates with integrins on cll surfaces. ‘The portion ofthe genome shared by all members of bacterial species. 1. van Bellum A, etal. 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(PubMed: 18197165] [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnuew sowny yduosnuew sowny osnue jy JOyINy du! osnue | JOUINY qdu ‘Toummarvengye etal Papas Online summary (© Staphylococcus aurens evades innate and adaptive immune responses to cause localized or systemic infections in humans. Because the development of protective immunity is prevented, S| aureus infections reoccur even with antibiotic or surgical therapy. ‘Genome sequencing of 5: aureus isolates fiom humans or domesticated animals revealed that only some immune evasion genes are conseived among all strains. Even conserved genes display sequence polymorphisms, which presents a formidable challenge for the design of S| aureus vaccines. Panoplies of immune evasion factors endow staphylococcal strains with unique virulence attributes and with the ability for epidemic spread ‘© Mechanisms have been revealed whereby S' aureus secreted products interfere with neutrophil chemotaxis, complement activation, opsonization and phagocytic Jklling of bacteria. Immune evasion determinants can interact with host factors from Jnumans, but not with counterparts fiom other vertebrates, which presents a challenge for animal model development ‘OHellmatk of 5. aureus isthe secretion of coagulases that associate with rothrombin to generate fibrin clots. Through the fbrinogen/fibrin binding attributes of staphylococcal surface proteins, the pathogen shields itself from host phagocytes, whichis a prerequisite for abscess formation in infected tissues, (OS aureus AdsA generates adenosine from adenine nucleotides during infection, thereby suppressing innate end adaptive immune responses through adenosine receptor signalling. Staphylococcal AdsA and muclease collaborate to covert neutrophil NETs, the released DNA of host neutrophils, into deoxyadenosine, thereby restricting macrophage access to S. aureus abscess lesions ‘© Staphylococcal protein A (SpA) crosslinks B cell receptors (IgM) and triggers proliferative expansion of V333 clonal B cells and the secretion of antibodies that fail to recognize 5. aureus antigens. SpA blocks host antibody responses that ae required for the establishment of protective immunity (© T cell superantigens (S Ags or enterotoxins) crosslink MHC class II molecules of antigen presenting cells with the T cell receptor of T lymphocytes, promoting lymphocyte proliferation, anergy and the release of cytokines (cytokine storm). SAgs varybetween 5 aurens strains and activate distinct subsets of Vij chain T cell receptors, endowing staphylococcal isolates with unique T cell avoidance attributes, [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny (@) Neutrophil extravasation and chemotaxis is inhibited by Staphylococcus aureus through the secretion of staphylococcal superantigen-like (SSL) molecules. SSL3 inhibits Toll-like receptor heterodimers; SSL5, SSL11 and SELX inhibit PSGL-1 signalling; and SSL6 inhibits the G-protein coupled receptor CD47. Other secreted proteins include chemotaxis inhibitory protein of 5. aureus (CHIPS), which inhibits the complement receptor C5aR and the formyi- peptide receptors 1 and 2 (FPR1/2); formyl peptide receptor-like 1 inhibitor (FLIP) and FLIPr-like (FLIPr-L), which inhibit FPR12, and staphopain, which inhibits signaling from the chemokine receptor CXCR2., (6) Complement activation and phagocytosis of staphylococci are blocked through the secretion of inhibitory factors to interfere with opsonization. Cha blocks the association of complement factor C1q bound to ‘immunoglobulin with complement receptor Clr, SpA and Sbi binding to immunoglobulin blocks its association with Clq; Sbi, SpA, SSL7 and SSL10 sequester immunoglobulins to block ther abilitiy to promote complement activation; Sbi (when associated with the host {factors C3d and fH) and SSL17 also inactivate the complement factors C3 and C5, respectively, Sak associates with plasminogen (PLG) and activates the zymogen to cleave complement factor C3b and immunoglobulin; Efp, Ecb, Bap and SCIN inhibit C3 convertases; and aureolysin cleaves the complement factor C3, which compromises opsonization because the cleavage product C3b is degraded by a complex of the host ‘proteins factor I (fl) and co-fector H (fH). (¢) 5. aureus inhibits neutrophil-mediated killing of phagocytosed bacteria by expressing several enzymes and inhibitors. The adenosine- synthesizing enzyme AdsA enables the inhibition of granulation via adenosine receptor (AdoR) signalling; staphyloxanthin, superoxide dismutase (SodA/SodM/), the catalase Kat and alkylhydropsroxide reductase (AhpC) are antioxidants that reduce oxidative stress caused by phagosomel reactive oxygen species (ROS) and H;0> generation, aureolysin (Aww) cleaves antimicrobial peptides; DItA-D promote D-elanyl esterification of teichoic acids to protect staphylococci fiom antimicrobial peptides; MxF modifies ‘phosphatidyiglycerol with alanine or lysine, another mechanism to protect staphylococci ‘against antimicrobial peptides; Lah and Hmp inhibit nitrosative stress; Eap, EapHl and [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal Pag a7 EapH? inhibit neutrophil serine proteases; and OatA. O-acetylates peptidoglycan, which events its lysozymal degradation. (@) Secreted p-bamel pore forming toxins (j-PFTS), bind specific receptors on immune cells to impair immune cell functions or promote cell lysis. ‘These f-PFTS include LukED, which binds to neutrophils, T cells and macrophages; HigAB, which binds to neutrophils, macrophages and red blood cells, HigCB and PVL, which bind to neutrophils and macrophages; and LukAB and Hla, which bind to neutrophils. Phenol- soluble modulin « (PSMa), which is another factor secreted by 5 aureus but nota p-PFT, can also lyse white blood cells. [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal _. ag By * ane Flag Fy Ok foinagen “zy ‘ets ovo ‘ete fibrinopeptides ¥* €>--D fibrin-dimer je, ota FnBPA FnBPB staphylothrombin Figure 2 Staphslococcus aureus agghutination with Sin provides protection against phagpeytes Physiological host defenses immobilize bacteria through the activation of the serine protease _zymogens prothrombin (also known as factor II) and factor X (not shown). In the contact activation pathway, surface contact results in the autocleavage of prothrombin (also known, as factor ID), thereby generating thrombin (also known as factor Ila). The Staphylococcus ‘ureus superantiger-like protein SSL0 inhibits prothrombin autoactivation, whereas the 5. ‘aureus coagulases Coa and vWbp convert prothrombin to staphylotivombin. Both thrombin and staphylotiombin cleave fibrinopsptides A andl B fiom fibrinogen to generate fibrin, which selfassembles and polymerizes into cable structures that immobilize bacteria ‘Thrombin activation results inthe activation of additional haemostasis factors that facilitate the simultaneous attraction of phagocytes to immobilized bacteria, which is thus inhibited by SSL10 secretion, However, staphyiothrombin cleaves fibrinopeptides from fibrinogen without activation of other haemostasis factors and promotes fibrin polymer assembly on the staphylococcal surface, where it protects the bacterium from neutrophils andl phagocytic clearance. Fibrin agglutination on the staphylococcal surface also involves the S. aureus surface proteins CICA, FrbpB and FrbpB, which bind tothe fibrinogen y-chains [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal Andee A} ating me al ‘ey e- faa’ Figure 3. Staphylococcus aureus AdA pertuibs adenosine and deoxyadenosine signalling (@) Staphylococcus aureus infection and its associated inflammatory damage promote the release of ATP, which is converted by adenosine synthase A (AdsA) info the immune suppressive signalling molecule adenosine (A). Adenosine inhibits activation of B cells, T cells, macrophages and denchritic cells via adenosine receptor (AdoR) signalling by acting on four different receptors (AdoRinanpas). Under physiological conditions, CD39 and CD73 generate adenosine signals to limit inflammatory responses; CD39 and CD73 are also responsible for the adenosine halo surounding immune cells and for immune suppressive states involving regulatory'T cells (T cells expressing the Foxp3* marker protein (not shown)). (6) 5 aureus induced NETosis of infiltrating neutrophils leads to nuclease mediated degradation of the DNA fibres that are the major components of neutrophil extracellular taps (NETs) and AdsA-mediated conversion of 5”-monophosphate- deoxyadenosine into deoxyedenosine, which promotes autocleavage ofthe apoptosis factor ro-capsase 3 to caspase 3. Cespase 3 induces macrophage death, thereby protecting S aureus ageinst professional phagocytes [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnue w sou;ny yduosnuew souny yduosnuew sony yduosnue w sowny ‘Toummarvengye etal a@ Gs. Figure 4. Staphylococcus aureus manipulates B and T cell responses (@) Staphylococcus aureus eleases SpA into host tissues, where itbinds to and crosslinks ‘V3 clan B cell receptors. In BI cells, marginal zone (IZ) B cells and B2 cells, SpA. crosslinking is associated with proliferative expansion and apoptotic collapse. The death of these cells impedes the development of adaptive immunity during 5 aureus infections. (b) In V3 plasmablasts, SpA crosslinking promotes somatic hypermutation and class switching from IgM antibodies to IgG antibodies, followed by the secretion of antibodies that are not specific for the S aureus antigen. (€) S aureus secretes T cell superantigen (SAg), which crosslinks major-histocompatibility complex class Il antigens (IMHCII) on the surface of antigen-presenting cells and T-cell receptors (TCR) on the surface of helper T (Tx) cells, triggering T cell expansion and anergy and causing cytokine storms. As result, T cells specific for S aureus antigens are not produced. [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnuew souny yduosnuew souny yduosnuew souny yduosnuew souny ‘Toummarvengye etal Tablet Staphylococcus aureus immune evasion determinants, their function end epidemiology Pagal ame Gee | Coane | Bepesed Ration Tart ‘alas ‘Admeste srduse [atid | cor nae apes ‘Ala, io ess T Aral ao [ewe Boose C3) 1 Copal maw [cae Bgogtoss ani 2 ca ae oe RL CAR, T CFA ca [ewe agocyoss nbn wed T OB ce [eae Mera Gekewh Wisc [1 oo aa [ aie | Colkgnediesim Clabading | Cla T Congise ca [ewe Baggies ain Hil Fe w Hom ia__[ewe as cellectneon a T Be op few Bagot iiingniin | ICAML C#,ehstse, ra cabepsin G protease 3 ‘Bee pH [cae Bepoelgaiiin | caaes ches T BpiD opie [cae Repoulaigaianin | eases ce T ee Ee Tedispawign viTCR T Benwonc [a | St Tedispawtign vpTcR 1 a ae Tedinpawtgn PGE 7 Be 23 | tem) | Conpkmet nha C3athm) 7 me | Brawn) | Coapkmet nba C38 M2 am) z ety fpr | Tecan) | Cumctecs hin eR e Hel fipri_[Teta(en) | Cencecs nha LID e Ey ‘ipA_| cae Tagotas shin anim | Fem 7 Ey fine [cae asin eee emt 7 EAB AB | stihig(con) | Bagssane sce CORI, CRD, COR 3 gee TigeB_| tihig (can) | Bagssone wee Ca a 38 TEAR (GEGH) | mia | ibmnAs | PNM Etec wtveten | CDLID a Tae aD | Giiem) _| PATe CORE, CORT CORD a Tan man | Gio) | PAOTie palo PMal Paval | coe Chara, POI RD 1 PEM Pad [ewe Chana POI ay T PENG panes [cae Chama, PRON 7 T Pat pone [cor Chana, POI RD T PENAL ingl_[ PSN (em) | Chencs, PONS RD 1 PENG Pau | PSN (re) | Checics, PONE ay T Patan Vaan | GRAS | PUL ping | PNNs ca T Jewordin (PVE) Se oe oe Pheiogn ORS | [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.yduosnuew souny yduosnuew souny yduosnuew souny yduosnuew souny ‘Toummarvengye etal Pag 32 ame Gee | Goan | Bepesed Ration Tart ‘alas wi Bi__| aihig (con) | Bigectoc Rain eR CE 7 scat sa | Bti (re) | Comphmee nib C3 GH) = a) iaB_| tive) | Caplets CRB OH) 7 SeRee ia_| tive) | Conpkmet shan CB) 7 Er me few Bagotewhhnin Bell | igh ig hb (VES) SE SPARES) spent ssi3 ais | Glave) | ThRsgaling nbn 7 5 Ssis S| Gate) | CemansMacktniitin | PGI, GPCRs GPb,, GPU | 5 sss 16 | Giatwa) | Chencends xibiion PGE 7 ssi a7 | Gate) | Bagectass anti ACS 7 ssLi0 30 | Giatee) | Bageotass aati KOM |e ssl ait_| Giatcan) | Cemands shin PSG 0 Sanyo ia_[ ewe Chee shin xo T ast [Sei _| Tellapentien VaTOR MHC ass | ike wo [ewe Bgegtosk abit il, Fe BIL AY z [Nee Rey Microbiol, Author muses, evelbble in PMC 2015 Octaber 2.