Sandra Schneider

Simultaneous cytometric analysis of (auto)antigen-reactive T and B cell proliferation

The detection and characterization of (auto)antigen-specific lymphocytes, both B and T cells, is essential to investigate immunopathologic mechanisms. Our aim was to perform a CFSE (Carboxyfluorescein diacetate succinimidyl ester)-based cytometric analysis of peripheral blood mononuclear cells (PBMC) proliferating in response to antigenic provocation. CFSE-labeled PBMC were stimulated with a superantigen (SEB), a recall antigen (tetanus toxoid), an allergen (grass pollen) and an autoantigen (nucleosomes) and stained after cultivation with CD4-, CD8- and CD19-antibodies. Proliferated cells were identified cytometrically by the decrease of the CFSE fluorescence intensity due to cell division. Antigen-reactive, proliferated B cells were further analysed phenotypically, antigen-specific proliferated Th cells were further characterized functionally regarding their cytokine secretion pattern after polyclonal restimulation. Using this technique, antigen-specific proliferated B and Th cells were detected even at low frequencies. Analyzing the cytokine secretion pattern of allergen-reactive proliferated Th cells after polyclonal restimulation we found differences in the expression of IL-13 and IL-4 between an atopic and a healthy donor. After stimulation of PBMC from TT-vaccinated donors TT-specific proliferated B cells were detected in high frequencies and showed a plasmablast-typical CD20(low) CD27(high) phenotype with only low frequencies expressing CD138 (= Syndecan-1). Proliferation of nucleosome-reactive Th cells and B cells was observed in both patients and healthy controls. We have optimized here the cytometric analysis of reactive cell proliferation based on CFSE offering various facilities of application on the further characterization of both antigen-specific B and T cells.

T cell reactivity against the SmD183–119 C terminal peptide in patients with systemic lupus erythematosus

Background: The SmD183–119 peptide is a major target of the B cell response in patients with systemic lupus erythematosus (SLE). Objective: To investigate the T cell response directed against this peptide, its disease specificity, and possible impact on SLE pathogenesis. Methods: Peripheral blood mononuclear cells derived from 28 patients with SLE and 29 healthy and disease controls were stimulated by the SmD183–119 and the recombinant (r)SmD1 protein, and [3H]thymidine incorporation was measured. Patients with SLE were simultaneously tested for autoantibodies, disease activity, clinical symptoms, and medical treatments. Results: T cell reactivity against the SmD183–119 peptide was detected in 11/28 (39%) patients with SLE and against the rSmD1 protein in 10/28 (36%) patients. In contrast, only 2/29 (7%) controls exhibited SmD1 reactivity. An analysis of proliferation kinetics showed that SmD1 reactive T cells are activated in vivo, as additionally confirmed by cytometric analysis. Addition of mammalian dsDNA to rSmD1 enhanced the rSmD1-specific T cell response. SmD183–119-specific T cell reactivity was significantly more common in patients with cardiac and pulmonary symptoms. No correlation between T and B cell responses and disease activity was seen. Conclusion: SmD183–119 is a major T cell epitope of SmD1, commonly recognised by T cells from patients with SLE and much less commonly found by healthy or disease controls. This strong T cell reactivity as well as the high frequency and specificity of anti-SmD183–119 antibodies in SLE suggest a possible role in SLE pathogenesis, at least in a subset of patients.

Time-series analysis to observe the impact of a centrally organized educational intervention on the prevention of central-line-associated bloodstream infections in 32 German intensive care units

BACKGROUND Prevention measures reduce central-line-associated bloodstream infections (CLABSIs) but are not always implemented. AIM To investigate the effect of a central educational programme in German intensive care units (ICUs) on CLABSI rates. METHODS Thirty-two German ICUs with CLABSI rates greater than or equal to the national average were compared with two control groups containing 277 and 67 ICUs. Processes and CLABSI rates were surveyed before, during and two years after the implementation of a year-long intervention programme. Segmented regression analysis of interrupted time series using generalized linear models was performed to estimate the association between the number of CLABSIs per month and time, intervention and other confounders, with the clustering effect within an ICU taken into account. FINDINGS In total, 508 cases of CLABSI were observed over 266,471 central line (CL)-days. At baseline, the pooled mean CLABSI rate was 2.29 per 1000 CL-days, and this decreased significantly to 1.64 per 1000 CL-days in the follow-up period. Compared with baseline, the relative risk for CLABSI was 0.88 [95% confidence interval (CI) 0.70-1.11] for the intervention period and 0.72 (95% CI 0.58-0.88) for the follow-up period. No changes were observed in either control group. Following successful implementation of the programme, ICUs showed a significant decrease in CLABSI rates. Although rates were already decreasing prior to implementation of the intervention, the invitation to participate in the study, and increased general awareness of CLABSI prevention through use of the comprehensive multi-modal training materials may have had a beneficial effect on practice.

Development and resolution of secondary autoimmunity after autologous haematopoietic stem cell transplantation for systemic lupus erythematosus: competition of plasma cells for survival niches?

Haematopoietic stem cell transplantation (HSCT) is an effective treatment for severe autoimmune diseases such as systemic lupus erythematosus (SLE).1 However, it is increasingly recognised that these patients have an added propensity to develop secondary autoimmune disorders.2 ,3 Here, we report on a 21-year-old male patient who received a CD34-selected autologous HSCT following conditioning with antithymocyte-globulin and cyclophosphamide (CYC) after written informed consent for refractory, severe SLE with renal, haematological, mucocutaneous and musculoskeletal manifestations (SLEDAI 19).1 Clinical remission was achieved for SLE within 3 months after HSCT and anti-double-stranded DNA (anti-dsDNA) antibodies disappeared despite immunosuppressive drug withdrawal. Eight months after HSCT, the patient presented with spontaneous joint and skin bleeding and was diagnosed with factor VIII (FVIII) inhibitor haemophilia with an activated partial thromboplastin time >100 s, FVIII activity <1% and a FVIII inhibitor titre of 435 Bethesda units (figure 1A). At that time point, flow cytometric analyses revealed a drastic increase in B cell numbers, expansion of circulating plasmablasts and a predominance of CD45RO memory CD4 T cells with oligoclonal T cell receptor Vβ expression (table 1), but clinical and laboratory tests showed no evidence of lupus activity. FVIII haemophilia was refractory to methylprednisolone, plasmapheresis, intravenous immunoglobulin (IVIG), intravenous CYC, rituximab and extracorporeal …

Perceptions and attitudes regarding antibiotic resistance in Germany: a cross-sectoral survey amongst physicians, veterinarians, farmers and the general public

Background Drivers of antibiotic (AB) resistance (ABR) include outpatient treatment, hospital care and animal husbandry. During the first phase of the One Health project RAI (Responsible Antibiotic Use via Information and Communication) surveys were conducted in these sectors. Objectives To compare perceptions and attitudes towards ABR among general practitioners (GPs), hospital physicians, veterinarians, pig farmers and the general public. Methods Cross-sectional questions on AB use and ABR were integrated in group-specific surveys of GPs, hospital physicians, veterinarians, pig farmers and the German general population. Results A total of 1789 participants (340 GPs, 170 hospital physicians, 215 pig farmers, 60 veterinarians and 1004 members of the public) responded. Each group tended to identify drivers of ABR as being from outside its own area of activity. Guidelines were shown to be an important information source for AB therapy for all prescriber groups, but the frequency of routine use differed (39% of GPs, 65% of hospital physicians and 53% of veterinarians). Regarding further information sources, hospital physicians preferred smartphone apps and e-learning, GPs preferred non-sponsored training and veterinarians preferred multidisciplinary networks and e-learning. Farmers were predominantly satisfied with existing solutions. Farmers had three times better basic knowledge of ABR and knew twice as many people with MDR organism problems than the general public. They also received information on ABR more often from their veterinarians than patients did from their doctors. Conclusions This study reveals considerable differences in perceptions and attitudes to ABR among the groups investigated. The results can help to tailor future interventions. Furthermore, they promote mutual understanding and thus support the One Health approach.

Antimicrobial prescription behavior in 16 German intensive care units: room for improvement in pneumonia therapy

Antimicrobial surveillance in German hospitals is mainly based on consumption monitoring. Sparse data exist on prescription culture.

O050: Antibiotics in 70 German intensive care units: risk factors for high overall consumption

Due to increasing burden of multidrug resistant organisms and the lack of new antimicrobial substances prudent use of antibiotics (AB) becomes more essential than ever before. Surveillance of antimicrobial use is an important component of AB stewardship.