Comprehensive infectious disease screening in a cohort of unaccompanied refugee minors in Germany from 2016 to 2017: A cross-sectional study.

BACKGROUND: Information regarding the prevalence of infectious diseases (IDs) in child and adolescent refugees in Europe is scarce. Here, we evaluate a standardized ID screening protocol in a cohort of unaccompanied refugee minors (URMs) in a municipal region of southwest Germany. METHODS AND FINDINGS: From January 2016 to December 2017, we employed a structured questionnaire to screen a cohort of 890 URMs. Collecting sociodemographic information and medical history, we also performed a standardized diagnostics panel, including complete blood count, urine status, microbial stool testing, tuberculosis (TB) screening, and serologies for hepatitis B virus (HBV) and human immunodeficiency virus (HIV). The mean age was 16.2 years; 94.0% were male, and 93.6% originated from an African country. The most common health complaints were dental problems (66.0%). The single most frequent ID was scabies (14.2%). Of the 776 URMs originating from high-prevalence countries, 7.7% and 0.4% tested positive for HBV and HIV, respectively. Nineteen pathogens were detected in a total of 119 stool samples (16.0% positivity), with intestinal schistosomiasis being the most frequent pathogen (6.7%). Blood eosinophilia proved to be a nonspecific criterion for the detection of parasitic infections. Active pulmonary TB was identified in 1.7% of URMs screened. Of note, clinical warning symptoms (fever, cough >2 weeks, and weight loss) were insensitive parameters for the identification of patients with active TB. Study limitations include the possibility of an incomplete eosinophilia workup (as no parasite serologies or malaria diagnostics were performed), as well as the inherent selection bias in our cohort because refugee populations differ across Europe. CONCLUSIONS: Our study found that standardized ID screening in a URM cohort was practicable and helped collection of relevant patient data in a thorough and time-effective manner. However, screening practices need to be ameliorated, especially in relation to testing for parasitic infections. Most importantly, we found that only a minority of infections were able to be detected clinically. This underscores the importance of active surveillance of IDs among refugees.

Development of prevalence and incidence of non-tuberculous mycobacteria in German laboratories from 2016 to 2020.

Numbers of non-tuberculous mycobacteria (NTM) pulmonary diseases (PD) have been repeatedly reported as increasing over the last decades, particularly in Europe. Sound epidemiological data are however missing for most European regions. This study calculated prevalence and incidence of NTM recovered from patients' lungs in Germany, the largest Central European country, over a five-year period. It furthermore determined regional particularities of NTM species and results from susceptibility testing. 22 German NTM laboratories provided their mycobacteriological diagnostic data of 11,430 NTM isolates recovered from 5998 pulmonary patients representing 30% of all notified NTM-PD cases of Germany from 2016 to 2020. NTM incidence and prevalence were calculated for every study year. The presented epidemiological indicators are particularly reliant as TB surveillance data were used as a reference and TB notification reaches almost 100% in Germany. Laboratory incidence and prevalence of NTM recovered from respiratory samples ranged from 4.5-4.9 and from 5.3-5.8/100,000 for the population of Germany, respectively, and did not change over the five-year study period. Prevalence and incidence were stable also when stratifying for facultative pathogenic NTM, M. avium/intracellulare complex (MAIC), and M. abscessus/chelonae complex (MABSC). The proportion of NTM with drug susceptibility testing (DST) increased from 27.3% (2016) to 43.8% (2020). The unchanging laboratory NTM prevalence/incidence in Germany represents a "ceiling" of possible NTM-PD notification when diagnostic strategies do not change in the coming years. A notable increase in NTM-DST may indicate better notification of NTM-PD and/or awareness of new clinical guidelines but still remains below clinical needs.

T cells are required for the production of blister-inducing autoantibodies in experimental epidermolysis bullosa acquisita.

Epidermolysis bullosa acquisita is a prototypical organ-specific autoimmune disease caused by autoantibodies against type VII collagen of the dermal-epidermal junction. Although mechanisms of autoantibody-induced blister formation were extensively characterized, the initiation of autoantibody production in autoimmune blistering diseases is still poorly defined. In the current study, we addressed the role of T cells for the production of blister-inducing autoantibodies in mice immunized with type VII collagen. To detect autoreactive type VII collagen-specific T cells, lymph node cells from immunized SJL mice were stimulated in vitro with recombinant Ag, and their proliferation was measured by radioactive thymidine incorporation and flow cytometry analysis of CFSE-labeled cells. Interestingly, using synthetic peptides of the immunogen, partly different T and B cell epitopes in mice immunized with type VII collagen were demonstrated. In contrast to wild-type mice, immunization with type VII collagen of SJL athymic nude mice lacking T cells did not induce an autoimmune response and blistering phenotype. Importantly, SJL nude mice repleted with T cells from immunized wild-type mice showed a robust and durable autoantibody production resulting in subepidermal blistering disease in the recipients. Our present results demonstrate that T cells are required for the initiation of autoimmunity against type VII collagen in experimental epidermolysis bullosa acquisita and provide a basis for developing T cell-directed immunomodulatory strategies for this and related autoimmune diseases.

Neonatal Fc receptor deficiency protects from tissue injury in experimental epidermolysis bullosa acquisita.

Epidermolysis bullosa acquisita (EBA) is an autoimmune blistering disease caused by autoantibodies against type VII collagen. The neonatal Fc receptor (FcRn) regulates immunoglobulin G (IgG) homeostasis and thus controls serum levels of antibodies. In this study, we investigated the effects of FcRn deficiency on the levels of autoantibodies against type VII collagen and blistering in EBA. For this purpose, rabbit IgG against murine type VII collagen was injected into FcRn-deficient and wild-type (n = 10 per group) mice. Enzyme-linked immunosorbent assay levels of serum IgG against type VII collagen were significantly lower in mutant compared with wild-type mice. Analysis of serum levels of specific autoantibodies induced in FcRn-deficient and wild-type mice (n = 10 per group) by immunization with type VII collagen showed significantly lower serum levels of IgG against type VII collagen in FcRn-deficient mice compared with wild-type animals. Importantly, the extent of blistering disease after injection of IgG against type VII collagen was significantly reduced in FcRn-deficient mice compared to wild-type controls. Our data demonstrate that FcRn maintains levels of pathogenic autoantibodies and thereby promotes tissue injury in experimental EBA. Therefore, modulation of FcRn function using inhibitors may reduce pathogenic IgG levels, offering therapeutic benefit in patients with antibody-mediated diseases.

Decreased platelet reactivity identified by whole blood flow cytometry in Fanconi anaemia patients.

Fanconi anaemia (FA) is a rare inherited chromosome instability disorder characterized by congenital anomalies and a high risk for bone marrow failure and cancer. Bleeding is a frequent complication in FA, leading to substantial morbidity and mortality. Thrombocytopenia is a major factor leading to this complication, but the bleeding tendency of FA patients often exceeds what one might expect based on their platelet counts. We therefore investigated if alterations of platelet function contribute to the bleeding tendency of FA patients. We assessed platelet function in 11 FA patients and 23 controls with whole blood flow cytometry. We analyzed the expression of platelet membrane glycoprotein receptors, reactivity of platelets to physiologic agonists and the proportion of young platelets. In FA patients platelet reactivity was decreased: Expression of P-selectin and binding of PAC-1 after stimulation with thrombin receptor activating peptide (TRAP) and adenosine diphosphate (ADP) were 15-70% lower than in controls. We found no or only minor differences of platelet glycoprotein receptor expression between groups. While the proportion of reticulated platelets was not different, the absolute number of reticulated platelets was markedly lower in FA patients. Our data show that FA is associated with reduced platelet reactivity, which may contribute to the high bleeding tendency in FA patients. Whole blood flow cytometry is a suitable method for analysis of platelet function in FA patients.

Chorioamnionitis is associated with increased CD40L expression on cord blood platelets.

Chorioamnionitis (CA) is a severe infection responsible not only for premature birth but also for many severe neonatal diseases. The aim of the present study was to investigate the expression of CD40L and P-selectin on platelets and the plasma levels of their soluble forms in the umbilical cord blood in infants with documented chorioamnionitis. Umbilical cord blood samples were obtained from 10 healthy term newborns, 10 non-infected preterm infants, 10 preterm infants with premature rupture of membranes and 9 preterm infants with clinical and histological CA. The expression of CD40L and P-selectin on platelets was analyzed by flow cytometry. Soluble P-selectin (sCD62P), soluble CD40L (sCD40L) and interleukine-6 (IL-6) were measured in plasma by ELISA assays. Neonates with CA had significantly higher percentages of platelets expressing CD40L in basal conditions (5.3 +/- 2.9% vs. 1.6 +/- 0.7% and in non-infected preterm infants p < 0.05), while the percentages of P-selectin positive platelets were similar among all groups. In contrast, the level of sP-selectin was higher in infants with CA (222 +/- 128 ng/ml vs. 104 +/- 71 ng/ml in non-infected preterm infants, p < 0.05) but no differences were found in the levels of sCD40L. As expected, the levels of IL-6, a pro-inflammatory cytokine were significantly higher in samples obtained from preterm neonates whose mothers had also elevated inflammatory parameters. Our observations suggest that platelets are involved in the complex inflammatory pathogenesis of CA. Neither P-selectin expression on cord blood platelets nor plasma sP-selectin or sCD40L were suitable platelet markers in CA, whereas CD40L was significantly elevated in histologically proven CA.

Allogeneic core amino acids of an immunodominant allopeptide are important for MHC binding and TCR recognition.

The indirect alloimmune response seems to be restricted to a few dominant major histocompatibility complex (MHC)-derived peptides responsible for T-cell activation in allograft rejection. The molecular mechanisms of indirect T-cell activation have been studied using peptide analogues derived from the dominant allopeptide in vitro, whereas the in vivo effects of peptide analogues have not been well characterized yet. In the present study, we generated allochimeric peptide analogues by replacing the three allogeneic amino acids 5L, 9L, and 10T in the sequence of the dominant MHC class I allopeptide P1. These allochimeric peptide analogues were used to define the allogeneic amino acids critical for the MHC binding and TCR recognition. We found that position 5 (5L) of the dominant allopeptide acts as an MHC-binding residue, while the other two allogeneic positions, 9 and 10, are important for the T-cell receptor (TCR) recognition. A peptide containing the MHC-binding residue 5L, as the only different amino acid between donor (RT1.A(u)) and recipient (RT1.A(l)) sequences, did not induce proliferation of lymph node cells primed with the dominant peptide and prevented dominant peptide-induced acceleration of allograft rejection. Identification of MHC and TCR contact residues should facilitate the development of antigen-specific therapies to inhibit or regulate the indirect alloimmune response.

Hierarchical immunogenicity of donor MHC class I peptides in allotransplantation.

The recognition of major histocompatibility complex (MHC) allopeptides by recipient MHC class II-restricted CD4(+) T cells via indirect pathway is a prerequisite for the generation of an immune response to the allograft. We tested 13-mer to 24-mer peptides from the MHC class I molecule for their possible immunogenicity in a fully MHC-mismatched rat strain combination. Our results confirm the hierarchical distribution of the immunogenicity of donor MHC class I peptides in the T cell alloactivation via indirect pathway. In addition, we show that allopeptide-induced immune response is critical for acute rejection of heart allografts. Among the seven allopeptides tested, peptide P1 was identified as immunodominant; it induced the greatest T cell proliferation and cytokine production in vitro as well as a significant reduction in allograft survival time. The TCR repertoire of T cells involved in the in vitro and in vivo responses induced by the dominant allopeptide P1 was found to be limited to the Vbeta10 and Vbeta 19 gene families. The identification of dominant allopeptides should greatly facilitate characterization of the specific T cell population responsible for allograft rejection and may be used to modulate the alloimmune response through antigen-specific therapy.