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Objective: To identify the role of Staphylococcus aureus (SA) or its intracellular small-colony variant phenotype (SCV) with co-trimoxazole (CTX) or ANCA-associated vasculitis (AAV) activity., Methods: All consecutive AAV patients with granulomatosis with polyangiitis (GPA), eosinophilic GPA or microscopic polyangiitis, followed at the French National Vasculitis Referral Center (09/2012-05/2013), and hospitalized non-AAV controls, exclusively for SA/SCV carriage comparisons, were enrolled. All had bilateral anterior nasal swab cultures. Nasal SA or SCV carriage was determined and associations with relapse(s), BVAS, ANCA-positivity, anti-staphylococcal and immunosuppressant use, were analysed ⩾4 years post-inclusion., Results: Nasal SA carriage rates did not differ among AAVs (P = 0.53): GPA (24/80; 30%), EGPA (7/28; 25%) and microscopic polyangiitis (3/11; 27.3%); and the rate was less frequent in controls than in GPA patients not taking CTX (P = 0.04). AAV patients taking CTX prophylaxis had less nasal SA carriage (8.7% vs 36.2%; P = 0.02). Nasal SA carriage or CTX use did not modify relapse rates, BVAS or ANCA-positivity at inclusion or during follow-up. Nasal SCV carriage, found in 15/207 (7.2%) patients, was similar for GPA (10/24; 41.7%), microscopic polyangiitis (2/7; 28.6%) and eosinophilic GPA (2/3; 66.7%), but higher (P = 0.02) than controls (1/14; 7.1%). SCV carriage by AAV groups did not modify relapse rates or ANCA positivity at inclusion or during follow-up; a trend towards higher BVAS was observed only for anti-PR3 ANCA patients., Conclusion: Nasal SA or SCV carriage was comparable among AAVs but more frequent than in controls. Nasal SA or SCV carriage and CTX use did not modify AAV relapse rates., (© The Author(s) 2019. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

The small vessel vasculitides granulomatosis with polyangiitis (GPA) and microscopic polyangiitis are associated with autoantibodies to neutrophil cytoplasm antigens (ANCA), principally proteinase-3 (PR3) and myeloperoxidase (MPO). There is an association between GPA and nasal carriage of Staphylococcus aureus. The recent finding that S. aureus produces proteins that bind tightly to and block the function of both PR3 and MPO suggests a mechanism for ANCA formation. The bacterial protein-autoantigen conjugate is recognised by B cells with ANCA specificity, internalised, and the bacterial protein processed and presented to T cells with specificity for bacterial peptides. The T cell can then provide help to the B cell, allowing class switching, affinity maturation and the production of pathogenic ANCA. This mechanism predicts that T cells with this specificity will be found in patients, and that the bacterial protein-autoantigen conjugate will be particularly efficient at eliciting ANCA production., (Crown Copyright © 2018. Published by Elsevier Ltd. All rights reserved.)

Objectives: Nasal carriage of Staphylococcus aureus and its superantigens (SAg) seem to be a risk factor disease exacerbation in granulomatosis with polyangiitis (GPA). We investigated the association between the presence of SAg in nasal swabs and activity of disease in GPA patients also taking into account correlation with an antimicrobial treatment., Methods: In a prospective study of a total of 150 GPA patients hospitalised in the period 2009-2016, nasal swabs were examined for the presence of Staphylococcus aureus and SAg. Subsequently, the association with disease activity was assessed., Results: Of 362 Staphylococcus aureus-positive nasal swab cultures from 115 of the 150 patients, the presence of at least one SAg in 126 samples (34.8%) from 56 patients (48.7%) was found. Among the 17 patients with limited to subglottic stenosis (SGS) disease, SAg were detected in 6 cases (35.3%). We did not find a significant correlation between the presence of SAg and disease activity (p=0.986), although when individual SAg were analysed separatively, SED and TSST-1 were more frequently present in active disease. Additionally, the results of the analysis demonstrated a protective effect of trimethoprim/sulfamethoxazole (T/S) treatment (0R 0.52, p<0.0092) in GPA patients. Interestingly, GPA limited to SGS appeared as an unfavourable factor associated with disease activity (0R 1.84, p=0.05)., Conclusions: The association between staphylococcal SAg in nasal swabs and GPA activity is not evident. Multiple mechanisms that may lead to disease activation still need to be investigated.

The proteinase 3 (PR3)-positive anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitis (AAV) granulomatosis with polyangiitis (GPA) has been associated with chronic nasal S. aureus carriage, which is a risk factor for disease relapse. The present study was aimed at comparing the genetic make-up of S. aureus isolates from PR3-ANCA-positive GPA patients with that of isolates from patients suffering from myeloperoxidase (MPO)-ANCA-positive AAV, and isolates from healthy controls. Based on a DNA microarray-based approach, we show that not only PR3-ANCA-positive GPA patients, but also MPO-ANCA-positive AAV patients mainly carried S. aureus types that are prevalent in the general population. Nonetheless, our data suggests that MPO-ANCA-associated S. aureus isolates may be distinct from healthy control- and PR3-ANCA-associated isolates. Furthermore, several genetic loci of S. aureus are associated with either PR3-ANCA- or MPO-ANCA-positive AAV, indicating a possible role for pore-forming toxins, such as leukocidins, in PR3-ANCA-positive GPA. Contrary to previous studies, no association between AAV and superantigens was detected. Our findings also show that a lowered humoral immune response to S. aureus is common for PR3-ANCA- and MPO-ANCA-positive AAV. Altogether, our observations imply that the presence or absence of particular virulence genes of S. aureus isolates from AAV patients contributes to disease progression and/or relapse.

Objectives: Nasal S. aureus carrier rates are significantly higher in patients with Wegener's granulomatosis (WG) compared to healthy controls (HC), and nasal colonisation is a risk-factor for relapse. Antimicrobial peptides (AMP) are important defence molecules maintaining an intact barrier function. It is the aim of this study to see if there is a possible link between the nasal AMP pattern and S. aureus colonisation, a link which has not been investigated so far., Methods: ELISA was applied to quantify LL-37 and hBD-3 concentrations in nasal secretions (14 WG patients, 13 HC) with and without nasal S. aureus colonisation. Immunohistochemistry was used to detect the cellular sources of AMP in the nasal mucosa. Functional analyses of primary nasal epithelial cell cultures (NEC) of these groups stimulated with S. aureus were performed., Results: LL-37 was found in significantly higher concentrations in colonised individuals (WG: p=0.001; HC: p=0.014).Using immunohistochemistry, local cellular sources for AMP could be demonstrated. After stimulation with S. aureus, significantly higher concentrations of LL-37 and hBD-3 could be detected in the supernatant of NEC of WG patients (LL-37: p=0.001; hBD-3: p=0.001) and HC (LL-37: p=0.019; hBD-3: p=0.001). HBD-3 concentrations were significantly lower in the supernatant of stimulated NEC of WG patients compared to the NEC of HC (p=0.032), and the dynamic range of the hBD-3 answer was significantly smaller in WG compared to HC (p=0.016)., Conclusions: The dynamic response towards challenges with microbes is dysregulated in WG, and this might be one reason for higher S. aureus colonisation rates in WG.

Objectives: Nasal colonisation with Staphylococcus aureus (S. aureus) has been implicated in Wegener's granulomatosis (WG) disease activity. In this study, the frequency of nasal colonisation with S. aureus in WG was compared to healthy and disease control groups for the first time. Moreover, endonasal activity was correlated to colonisation., Patients and Methods: Nasal carriage of S. aureus of a well-defined group of 89 patients with WG was compared to 40 patients with chronic rhinosinusitis with nasal polyps (CRS), 35 patients with rheumatoid arthritis (RA), 50 hospital staff members and 25 subjects without regular hospital contact and correlation analysis of nasal carriage and endonasal activity of WG was performed., Results: WG patients showed significantly higher rates (72%) of nasal colonisation with S. aureus compared to CRS patients (28%) and healthy subjects without regular hospital contact (25%, 95%-CI), but not to RA patients (46%) and hospital staff members (58%). WG patients with nasal carriage of S. aureus had significantly higher endoscopically proven endonasal activity (p=0.01), significantly more often first manifestation of WG in the upper respiratory tract (p=0.02) and higher relapse-rates (p=0.052) than WG patients without such carriage., Conclusions: Endonasal activity in WG is associated with higher nasal S. aureus colonisation rates and subsequent higher relapse rates. The higher frequency of S. aureus colonisation could be a consequence of a recently shown mucosal barrier defect in WG and facilitate chronic inflammation and granuloma formation in the upper respiratory tract.

Rationale: Wegener granulomatosis (WG) has previously been associated with increased nasal carriage of Staphylococcus aureus, but no studies have investigated the occurrence of pathogen growth in the lower airways., Objectives: To culture bronchoalveolar lavage fluid (BALF) from patients with WG, patients with idiopathic pulmonary fibrosis (IPF) and normal controls., Methods: 33 patients with WG, 22 with IPF and 8 normal controls underwent bronchoscopy and bronchoalveolar lavage. Quantitative culture established bacterial levels in the lower airways. Culture experiments were designed to investigate whether BALF is a supportive environment for S aureus growth. BALF cytokines were measured by ELISA., Results: Pathogens were commonly grown from BALF of patients with WG and those with IPF. S aureus was particularly associated with patients with WG both in relapse and in remission. BALF levels of interleukin 1 receptor antagonist (IL1ra) were statistically significantly elevated in those patients who grew a pathogen from lavage fluid. BALF from patients with WG and IPF stimulated S aureus growth compared with normal lavage fluid., Conclusions: Pathogens are more commonly isolated from BALF from patients with WG than from that of patients with IPF or normal controls, and with a different culture profile. IL1ra was associated with pathogen growth in WG and IPF. WG BALF is a trophic environment for S aureus growth. Pulmonologists treating patients with acute or relapsing WG should consider bronchoscopic microbiological sampling and consider antibiotics with antistaphylococcal activity.

Objectives: Nasal carriage of Staphylococcus aureus constitutes a risk factor for disease exacerbation in Wegener's granulomatosis (WG). We hypothesized that staphylococcal superantigens (SAg) are a determinant of S. aureus-related risk for disease relapse in WG., Methods: In a retrospective longitudinal cohort study in 62 WG patients, we investigated the presence of the staphylococcal SAg genes sea, seb, sec, sed, see, tsst-1 and eta in S. aureus strains isolated from WG patients during an observation period of seven years. Subsequently, we assessed whether relapses of WG were associated with the presence of SAg-positive staphylococci., Results: Of 1718 swab cultures analysed, 709 (41.2%) were S. aureus-positive. Fifty-one patients carried S. aureus, of whom 37 (72.5%) patients carried at least one SAg-positive S. aureus strain. Of the 709 S. aureus-positive cultures, 326 (46%) contained at least one SAg gene. Except for see, all assessed SAg genes were detected. sea was found most frequently, followed by sec, tsst-1 and eta and finally, by sed and seb. Using a multivariate, time-dependent Cox regression analysis we found that the presence of S. aureus was associated with relapses of WG (RR 3.2; 95% CI 1.2-8.4). The risk for relapse was modulated by the presence and type of SAg, with tsst-1 being associated with an increased risk for relapse (RR 13.3, 95% CI 4.2-42.6)., Conclusion: The risk for relapse of WG increases with the presence of tsst-1-positive S. aureus. Eradication of tsst-1-positive S. aureus in WG may show whether disease relapses can be prevented.

To allow rapid identification of toxic shock syndrome toxin-1 (TSST-1)-producing Staphylococcus aureus strains, a real-time PCR assay for the detection of the tst gene, which encodes TSST-1, was developed. The assay was applied to S. aureus isolates from patients with Wegener's Granulomatosis (WG), as well as isolates that were classified as either community- (CA) or hospital-acquired (HA). No significant difference in the percentage of tst-positive strains was observed between isolates from WG patients and CA isolates (24% and 25%, respectively). In contrast, only 14% of the HA isolates were tst-positive (p<0.05). Investigation of the clonal relationship between tst-positive CA and HA strains could indicated the recent emergence of a virulent S. aureus clone in the community.

To date, in the investigation of the role of S. aureus in WG, we face a paradoxical situation. On the one hand, clinical results obtained from treatment of WG patients with co-trimoxazole and studies assessing the impact of S. aureus on disease relapses strongly suggest that this bacterium contributes to disease pathophysiology. On the other hand, laboratory investigation of the possible mechanisms by which S. aureus is involved in WG is scarce, despite the fact that knowledge and tools to study this microorganism are abundant. In the present review, we discuss recent works investigating the possible pathophysiologic contribution of S. aureus to WG. Moreover, we propose a number of possibly relevant pathways of interaction of this bacterium with lymphoid and nonlymphoid cells of the WG host.

The majority of patients with Wegener's granulomatosis (WG) are chronic nasal carriers of Staphylococcus aureus. Chronic nasal carriage of S. aureus is associated with an increased risk of developing a relapse of the disease. The mechanism by which this occurs is still unknown. We hypothesized that a cationic protein of S. aureus, staphylococcal acid phosphatase (SAcP), acts as a planted antigen and initiates glomerulonephritis and vasculitis in patients with WG. In order to test the hypothesis that SAcP can act as a planted antigen in WG, we studied the ability of SAcP to bind to human umbilical vein endothelial cells (HUVEC) and human glomerular endothelial cells. We also studied whether this binding can be prevented by preincubation with an anionic protein, and whether binding of SAcP activates endothelial cells. We also evaluated whether antibodies in sera of patients with WG are able to bind to endothelial cell-bound SAcP. The results show that SAcP can act as a planted antigen by binding to both types of endothelial cells in a concentration-dependent manner. Binding of concentrations as low as 4 microg/ml can be detected on HUVEC within 5 min of incubation. Binding of SAcP to endothelial cells was charge-dependent but did not activate endothelial cells. Finally, endothelial cell-bound SAcP was recognized by sera of patients with WG. The data suggest a possible pathogenic role for SAcP by acting as a planted antigen thereby initiating glomerulonephritis and vasculitis in patients with WG.

The initial stage of Wegener's Granulomatosis (WG) is often marked by symptoms of infection and it has been postulated that a bacterial infection could be the aetiologic factor of this disease. The objective of our work was to investigate T-cell-mediated immunity in WG by testing proliferative responses on bacterial antigens and particularly Staphylococcus aureus. We investigated the bulk proliferative response of peripheral blood lymphocytes (PBL) from patients with clinically active WG to gram-positive bacteria and purified proteinase 3 (PR-3), the major target antigen of c-ANCA. We generated S. aureus-specific PBL-derived T-cell lines and T-cell clones (TLC). In two WG patients 27 TLC were characterized in terms of reactivity to bacterial antigens/PR-3, phenotype, HLA class II restriction and pattern of cytokine secretion. Compared to coagulase-negative Staphylococci and beta-haemolytic Streptococci A, reactivity to S. aureus was significantly increased in all patients with WG. Using purified PR-3, we found a PBL proliferation in five out of 25 WG patients. The TLC were S. aureus-specific and did not cross-recognize Streptococci or coagulase-negative Staphylococci. The S. aureus-specific TLC were of the alphabeta-TCR+ CD4+ phenotype and HLA-DR-restricted. These TLC predominantly showed a Th2-type of cytokine secretion. Interestingly, seven of the S. aureus-reactive TLC also recognized the PR-3 antigen. From these data we conclude that Staphylococci-specific HLA-DR-restricted CD4+ T cells may play a key role in the initial triggering of immune responses in WG.