New Weapons to Fight against Staphylococcus aureus Skin Infections.

The treatment of Staphylococcus aureus skin and soft tissue infections faces several challenges, such as the increased incidence of antibiotic-resistant strains and the fact that the antibiotics available to treat methicillin-resistant S. aureus present low bioavailability, are not easily metabolized, and cause severe secondary effects. Moreover, besides the susceptibility pattern of the S. aureus isolates detected in vitro, during patient treatment, the antibiotics may never encounter the bacteria because S. aureus hides within biofilms or inside eukaryotic cells. In addition, vascular compromise as well as other comorbidities of the patient may impede proper arrival to the skin when the antibiotic is given parenterally. In this manuscript, we revise some of the more promising strategies to improve antibiotic sensitivity, bioavailability, and delivery, including the combination of antibiotics with bactericidal nanomaterials, chemical inhibitors, antisense oligonucleotides, and lytic enzymes, among others. In addition, alternative non-antibiotic-based experimental therapies, including the delivery of antimicrobial peptides, bioactive glass nanoparticles or nanocrystalline cellulose, phototherapies, and hyperthermia, are also reviewed.

Neutralization of Staphylococcus aureus Protein A Prevents Exacerbated Osteoclast Activity and Bone Loss during Osteomyelitis.

Osteomyelitis caused by Staphylococcus aureus is an important and current health care problem worldwide. Treatment of this infection frequently fails not only due to the increasing incidence of antimicrobial-resistant isolates but also because of the ability of S. aureus to evade the immune system, adapt to the bone microenvironment, and persist within this tissue for decades. We have previously demonstrated the role of staphylococcal protein A (SpA) in the induction of exacerbated osteoclastogenesis and increased bone matrix degradation during osteomyelitis. The aim of this study was to evaluate the potential of using anti-SpA antibodies as an adjunctive therapy to control inflammation and bone damage. By using an experimental in vivo model of osteomyelitis, we demonstrated that the administration of an anti-SpA antibody by the intraperitoneal route prevented excessive inflammatory responses in the bone upon challenge with S. aureus. Ex vivo assays indicated that blocking SpA reduced the priming of osteoclast precursors and their response to RANKL. Moreover, the neutralization of SpA was able to prevent the differentiation and activation of osteoclasts in vivo, leading to reduced expression levels of cathepsin K, reduced expression of markers associated with abnormal bone formation, and decreased trabecular bone loss during osteomyelitis. Taken together, these results demonstrate the feasibility of using anti-SpA antibodies as an antivirulence adjunctive therapy that may prevent the development of pathological conditions that not only damage the bone but also favor bacterial escape from antimicrobials and the immune system.

Protein A Modulates Neutrophil and Keratinocyte Signaling and Survival in Response to Staphylococcus aureus.

The type 1 TNF-α receptor (TNFR1) has a central role in initiating both pro-inflammatory and pro-apoptotic signaling cascades in neutrophils. Considering that TNFR1 signals Staphylococcus aureus protein A (SpA), the aim of this study was to explore the interaction of this bacterial surface protein with neutrophils and keratinocytes to underscore the signaling pathways that may determine the fate of these innate immune cells in the infected tissue during staphylococcal skin infections. Using human neutrophils cultured in vitro and isogenic staphylococcal strains expressing or not protein A, we demonstrated that SpA is a potent inducer of IL-8 in neutrophils and that the induction of this chemokine is dependent on the SpA-TNFR1 interaction and p38 activation. In addition to IL-8, protein A induced the expression of TNF-α and MIP-1α highlighting the importance of SpA in the amplification of the inflammatory response. Protein A contributed to reduce neutrophil mortality prolonging their lifespan upon the encounter with S. aureus. Signaling initiated by SpA modulated the type of neutrophil cell death in vitro and during skin and soft tissue infections (SSTI) in vivo triggering the apoptotic pathway instead of necrosis. Moreover, SpA induced pro-inflammatory cytokines in keratinocytes, modulating their survival in vitro and preventing the exacerbated necrosis and ulceration of the epithelium during SSTI in vivo. Taken together, these results highlight the importance of the inflammatory signaling induced by protein A in neutrophils and skin epithelial cells. The ability of protein A to modulate the neutrophil/epithelial cell death program in the skin is of clinical relevance considering that lysis of neutrophils and epithelial cells will promote an intense inflammatory response and contribute to tissue damage, a non-desirable feature of complicated SSTI.

The good side of inflammation: Staphylococcus aureus proteins SpA and Sbi contribute to proper abscess formation and wound healing during skin and soft tissue infections.

Staphylococcus aureus is the most prominent cause of skin and soft tissue infections (SSTI) worldwide. Mortality associated with invasive SSTI is a major threat to public health considering the incidence of antibiotic resistant isolates in particular methicillin resistant S. aureus both in the hospital (HA-MRSA) and in the community (CA-MRSA). To overcome the increasing difficulties in the clinical management of SSTI due to MRSA, new prophylactic and therapeutic approaches are urgently needed and a preventive vaccine would be welcome. The rational design of an anti-S. aureus vaccine requires a deep knowledge of the role that the different bacterial virulence factors play according to the type of infection. In the present study, using a set of isogenic deficient mutants and their complemented strains we determined that the staphylococcal surface proteins SpA and Sbi play an important role in the induction of inflammatory cytokines and chemokines in the skin during SSTI. SpA and Sbi initiate signaling cascades that lead to the early recruitment of neutrophils, modulate their lifespan in the skin milieu and contribute to proper abscess formation and bacterial eradication. Moreover, the expression of SpA and Sbi appear critical for skin repair and wound healing. Thus, these results indicate that SpA and Sbi can promote immune responses in the skin that are beneficial for the host and therefore, should not be neutralized with vaccine formulations designed to prevent SSTI.

TNFR1 Signaling Contributes to T Cell Anergy During Staphylococcus aureus Sepsis.

Early research on sepsis has focused on the initial hyper-inflammatory, cytokine mediated phase of the disorder whereas the events that govern the concomitant and subsequent anti-inflammatory compensatory response are not completely understood. In this context, the putative participation of TNFR1-mediated signaling in the immunosuppressive phase of Staphylococcus aureus sepsis has not been elucidated. The aim of this study was to determine the role of TNFR1 in directing the immune dysfunction during S. aureus sepsis and the potential contribution of MDSC to this process. Using a model of sepsis of peritoneal origin and tnfr1-/- mice, we demonstrated that during staphylococcal sepsis CD4+ T cell anergy is significantly dependent on TNFR1 expression and that signaling through this receptor has an impact on bacterial clearance in the spleen. MDSC played a major role in the generation of anergic CD4+ T cells and their accumulation in the spleen during S. aureus sepsis correlated with IL-6 induction. Although TNFR1 signaling was not required for MDSC accumulation and expansion in the spleen, it determined the in vivo expression of Arginase 1 and iNOS, enzymes known to participate in the suppressive function of this population. Moreover, our data indicate that TNFR1-mediated IL-10 production may modulate MDSC function during staphylococcal sepsis. Taken together these results indicate that TNFR1 plays a critical role on T cell dysfunction during S. aureus sepsis by regulating immunomodulatory mediators in MDSC. The role of TNFR1-mediated signaling during the immunosuppressive phase of staphylococcal sepsis should be considered when designing novel alternative therapeutic approaches.

Mutation of Agr Is Associated with the Adaptation of Staphylococcus aureus to the Host during Chronic Osteomyelitis.

Selection pressures exerted on Staphylococcus aureus by host factors may lead to the emergence of mutants better adapted to the evolving conditions at the infection site. This study was aimed at identifying the changes that occur in S. aureus exposed to the host defense mechanisms during chronic osteomyelitis and evaluating whether these changes affect the virulence of the organism. Genome assessment of two S. aureus isolates collected 13 months apart (HU-85a and HU-85c) from a host with chronic osteomyelitis was made by whole genome sequencing. Agr functionality was assessed by qRT-PCR. Isolates were tested in a rat model of osteomyelitis and the bacterial load (CFU/tibia) and the morphometric osteomyelitic index (OI) were determined. The ability of the isolates to trigger the release of proinflammatory cytokines was determined on macrophages in culture. Persistence of S. aureus within the host resulted in an agrC frameshift mutation that likely led to the observed phenotype. The capacity to cause bone tissue damage and trigger proinflammatory cytokines by macrophages of the agr-deficient, unencapsulated derivative (HU-85c) was decreased when compared with those of the isogenic CP8-capsulated parental strain (HU-85a). By comparison, no significant differences were found in the bacterial load or the OI from rats challenged with isogenic Reynolds strains [CP5, CP8, and non-typeable (NT)], indicating that lack of CP expression alone was not likely responsible for the reduced capacity to cause tissue damage in HU-85c compared with HU-85a. The production of biofilm was significantly increased in the isogenic derivative HU-85c. Lack of agr-dependent factors makes S. aureus less virulent during chronic osteomyelitis and alteration of the agr functionality seems to permit better adaptation of S. aureus to the chronically infected host.

Staphylococcus aureus Induces Shedding of IL-1RII in Monocytes and Neutrophils.

Interleukin 1 (IL-1) ß is a critical cytokine that orchestrates host defenses against Staphylococcus aureus and is crucial for the eradication of bacteria. The production and action of IL-1ß are regulated by multiple control pathways. Among them, IL-1RII (the type II IL-1 receptor) acts as a decoy receptor and has been shown to regulate the biological effects of IL-1ß. High levels of soluble IL-1RII are present in septic patients; however, the stimuli that regulate the expression and release of IL-1RII in pathological conditions are incompletely elucidated. In the present study, we demonstrated the ability of S. aureus and protein A to induce IL-1RII shedding in myeloid cells. The positive modulation of IL-1RII expression and cleavage was associated with the failure to detect IL-1ß in response to S. aureus both in vitro and in vivo, suggesting that the soluble form of the receptor could be masking the availability of IL-1ß. The absence of detectable IL-1ß was associated with low levels of inflammatory cytokines and chemokines known to be regulated by IL-1ß and with increased bacterial persistence. Modulation of decoy receptors during systemic S. aureus infection is proposed as a new strategy used by this bacterium to evade the immune response.

Treatment in vitro with PPARα and PPARγ ligands drives M1-to-M2 polarization of macrophages from T. cruzi-infected mice.

Trypanosoma cruzi, the etiological agent of Chagas' disease, induces a persistent inflammatory response. Macrophages are a first line cell phenotype involved in the clearance of infection. Upon parasite uptake, these cells increase inflammatory mediators like NO, TNF-α, IL-1ß and IL-6, leading to parasite killing. Although desired, inflammatory response perpetuation and exacerbation may lead to tissue damage. Peroxisome proliferator-activated receptors (PPARs) are ligand-dependent nuclear transcription factors that, besides regulating lipid and carbohydrate metabolism, have a significant anti-inflammatory effect. This is mediated through the interaction of the receptors with their ligands. PPARγ, one of the PPAR isoforms, has been implicated in macrophage polarization from M1, the classically activated phenotype, to M2, the alternatively activated phenotype, in different models of metabolic disorders and infection. In this study, we show for the first time that, besides PPARγ, PPARα is also involved in the in vitro polarization of macrophages isolated from T. cruzi-infected mice. Polarization was evidenced by a decrease in the expression of NOS2 and proinflammatory cytokines and the increase in M2 markers like Arginase I, Ym1, mannose receptor and TGF-ß. Besides, macrophage phagocytic activity was significantly enhanced, leading to increased parasite load. We suggest that modulation of the inflammatory response by both PPARs might be due, at least in part, to a change in the profile of inflammatory macrophages. The potential use of PPAR agonists as modulators of overt inflammatory response during the course of Chagas' disease deserves further investigation.