Focus groups are becoming increasingly popular in research, especially in parent and child research. Focus group interviews allow participants to tell their own stories, express their opinions, and even draw pictures without having to adhere to a strict sequence of questions. This method is very suitable for collecting data from children, youths, and parents. However, focus group interviews must be carefully planned and conducted. The literature on focus group interviews with adult participants is extensive, but there are no current summaries of the most important issues to consider when conducting focus group interviews with children, youths, or parents. This article outlines the use of focus groups in child, youth, and parent research and the important factors to be considered when planning, conducting, and analyzing focus groups with children, youths, or parents.

Gastric infection with Helicobacter heilmannii (previously known as Gastrospirillum hominis) is invariably linked with the presence of chronic gastritis and the risk of developing low-grade mucosa-associated lymphoid tissue lymphoma in humans. In contrast to Helicobacter pylori, various H. heilmannii species colonize the stomachs of domestic animals, which might be a reservoir for transmission to humans (zoonosis). To identify the number and prevalence of different H. heilmanni types in humans, we analyzed 89 gastric biopsy samples histologically identified as H. heilmannii positive by fluorescence in situ hybridization. Of these gastric specimens, 84 (94.4%) contained a single H. heilmannii type. In five samples, however, two different H. heilmannii types were detected. The most prevalent species in monoinfected samples is H. heilmannii type 1, found in 78.5% (66 of 84) of the specimens, followed by a novel H. heilmannii-like organism (HHLO), HHLO type 4, identified in 9.6% (8 of 84) of tissue sections. H. heilmannii type 2 and a further HHLO type not described before, type 3, were found in 8.3% (7 of 84) and 1.2% (1 of 84) of the monoinfected samples, respectively. Additionally, HHLO type 5 with a 16S ribosomal DNA sequence identical to that of Helicobacter salomonis was found with a prevalence of 2.4% (2 of 89). Thirteen of these biopsy samples were also investigated by a PCR approach developed for this study that allows a Helicobacter-specific amplification of a variable portion of the 16S rRNA gene and subsequent sequencing. In total, five different types of HHLOs could be identified within these samples. We conclude that humans can be infected by at least five different HHLO types, which presumably have their origin in animal species like dogs, cats, and pigs.

Stimulation of CD4+ helper T lymphocytes by antigen-presenting cells requires the degradation of exogenous antigens into antigenic peptides which associate with major histocompatibility complex (MHC) class II molecules in endosomal or lysosomal compartments. B lymphocytes mediate efficient antigen presentation first by capturing soluble antigens through clonally distributed antigen receptors (BCRs), composed of membrane immunoglobulin (Ig) associated with Ig-α/Ig-β heterodimers which, second, target antigens to MHC class II–containing compartments. We report that antigen internalization and antigen targeting through the BCR or its Ig-α–associated subunit to newly synthesized class II lead to the presentation of a large spectrum of T cell epitopes, including some cryptic T cell epitopes. To further characterize the intracellular mechanisms of BCR-mediated antigen presentation, we used two complementary experimental approaches: mutational analysis of the Ig-α cytoplasmic tail, and overexpression in B cells of dominant negative syk mutants. Thus, we found that the syk tyrosine kinase, an effector of the BCR signal transduction pathway, is involved in the presentation of peptide– MHC class II complexes through antigen targeting by BCR subunits.

Phenotypic susceptibility testing for clarithromycin by E-test and disk diffusion of 109 cultured Helicobacter pylori isolates was compared with the genotypic susceptibility determination by fluorescent in situ hybridization (FISH). No discrepancies were found between these three methods. However, FISH has the advantage of providing results after 3 h.

Fluorescence in situ hybridisation (FISH) targeted to ribosomal RNA is well established for studies in environmental microbiology. Initial applications of this technique in the field of medical microbiology showed that FISH is also a suitable means for the rapid, reliable and cultivation-independent identification of bacterial pathogens. In particular, for infectious diseases that follow a fulminant live-threatening course, such as sepsis or necrotising fasciitis (NF), a fast and reliable detection technique is of great importance. This study describes the development of an rRNA-targeted oligonucleotide set covering more than 95% of the pathogens associated with NF. These probes were tested with a broad collection of target and non-target organisms and found to be highly specific. Subsequently, the FISH approach was applied for the direct detection of bacterial pathogens in clinical samples. Two cases of NF and one case of streptococcal toxic shock syndrome (STSS) were analysed. FISH correctly identified almost all pathogens present in the samples examined within 2-3 h. However, Proteus mirabilis, which was identified in one sample by conventional methods was detected as a rod-shaped bacteria but could not be identified by FISH, since no specific probe was available for this particular organism. In contrast, identification of pathogens in these samples by conventional laboratory methods took 48-72 h. Furthermore, in one patient with pre-sampling antimicrobial therapy bacteria could not be grown from any of the samples. FISH unequivocally revealed the presence of Streptococcus pyogenes in affected tissue samples from this patient. In an experimental setting we demonstrated that FISH readily identifies S. pyogenes cells rendered non-cultivable by antibiotic treatment.

Polymyxin resistance among common CF pathogens is not rare, thus underlining the necessity of accurate susceptibility testing. When compared with the agar dilution method, it was found that the microdilution method is a valid, rapid and cost effective alternative for the determination of polymyxin activity. The performance of the microdilution method was most reliable after prolonged incubation (48 h) at a susceptibility breakpoint of < or =4 mg/L according to the BSAC guidelines (specificity 91%, sensitivity 89%, 1.5% very major errors).

Background: Recently gene therapy with onasemnogene abeparvovec has been approved for the treatment of spinal muscular atrophy (SMA). As the experience from clinical trials is limited, there are still uncertainties for which patient population the treatment can be considered safe and effective. Methods: We report our experience with eight consecutive patients with SMA who were treated with the standard dose of onasemnogene abeparvovec (1.1×1014 vg/kg) at the University Hospital Bonn, Germany. All patients received prophylactic immunosuppression with 1 mg/kg/d prednisolone for four weeks starting on the day before gene therapy. Results: We treated eight patients (4 male, 4 female, age range 10–37 months) with a body weight between 7.1 and 11.9 kg. All patients had 2 or 3 copies of the SMN2-gene and were previously treated with nusinersen. Following treatment with onasemnogene abeparvovec all patients showed a temporary increase of the body temperature and an increase of transaminase levels. In all but one patient it was necessary to increase or prolong the standard steroid dose to control the immune response. In one severe case, liver damage was associated with impaired liver function. This patient received a steroid pulse therapy for five days. Blood counts revealed asymptomatic thrombocytopenia (<150×109/L) in 6/8 patients and a significant increase of monocytes following gene therapy. Liver values and blood counts returned to almost normal levels during the post-treatment observation period. Troponin I increased above normal limit in 4/8 patients but was not associated with any abnormalities on cardiac evaluation. Conclusions: In a broader spectrum of patients, treatment with onasemnogene abeparvovec was associated with a higher rate of adverse events. In our cases it was possible to control the immune response by close monitoring and adaptation of the immunosuppressive regimen. Further research is needed to better understand the immune response following gene therapy and ideally to identify patients at risk for a more severe reaction.

AimsA novel concept for the treatment of heart failure is the neutralization of antibodies against the b 1 -adrenergic receptor (anti-b 1 AR-ab). In a rat model of autoimmune cardiomyopathy, the cyclic peptide COR-1 (given i.v. once monthly) neutralized anti-b 1 AR-abs and prevented anti-b 1 AR-ab-induced myocardial damage, and completely reverted cardiac dysfunction over 3-6 months. Methods and resultsA clinical phase I trial was designed as a single-blinded, placebo-controlled study. Fifty human volunteers received COR-1 or matching placebo as a single i.v. administration with ascending doses (10 -240 mg). Primary endpoints were safety and tolerability, while the pharmacokinetic profile of COR-1 was assessed as a secondary endpoint. All five investigated dose groups were well tolerated; no drug-related side effects occurred. Pharmacokinetics revealed a favourable profile with an almost complete plasma clearance within 60 min after administration. Pharmacodynamic investigation showed dose-dependent efficacy with almost complete scavenging of pathological antib 1 AR-abs ex vivo at the two highest doses. No anti-COR-1 autoantibodies occurred. No other effects on the immune system (such as an increase of crucial cytokines) were observed up to 43 days after drug administration, nor upon incubation of anti-b 1 AR-ab-positive patient blood samples with COR-1 ex vivo. ConclusionsCOR-1 was shown to be safe after i.v. administration in vivo; no relevant side effects occurred. Efficacy was estimated from ex vivo investigation of the potency to neutralize specific anti-b 1 -AR-abs.