[Adaption of the "Empty Nose 6 Item Questionnaire" (ENS6Q) into German language]. / Erstellung einer deutschen Version des "Empty Nose Syndrome 6 Item Questionnaire" (ENS6Q).

OBJECTIVE: The English "Empty-Nose-6-Item-Questionnaire" (ENS6Q) is a validated tool to assess subjective and disease-specific complaints of patients suffering from empty nose syndrome. The aim of this study was to create a validated German adapted version (GAV) of the ENS6Q. MATERIAL AND METHODS: The ENS6Q was translated into German language using a multistage process. Subsequently the translated questionnaire was completed twice by a group of patients suffering from empty nose syndrome (n = 36), a healthy control group (n = 77) and a group of patients with a symptomatic pathology of the nasal septum (n = 78). Additionally, the NOSE© questionnaire was completed twice by all groups. The ENS6Q was then examined regarding internal consistency, test-retest reliability, discriminant validity, sensitivity and specificity. RESULTS: The ENS6Q-GAV shows high internal consistency with Cronbachs α = 0,760 and α = 0,795. It also shows good test-retest reliability with an interclass correlation coefficient of 0,947 [0,930 - 0,961]. The ENS6Q-GAV discriminated significantly between empty nose syndrome patients and the control group. The receiver-operating characteristics curve showed a sensitivity of 83,3 % and a specificity of 88,3 % using a cut-off point of 8,5. The area-under-the-curve threshold score was 0,914 ± 0,029 [0,856 - 0,971]. CONCLUSIONS: The ENS6Q-GAV is the first validated disease-specific questionnaire in German to help identify patients suspected of developing ENS more reliably.

The iSplit GFP assay detects intracellular recombinant proteins in Bacillus subtilis.

BACKGROUND: Bacillus subtilis is one of the most important microorganisms for recombinant protein production. It possesses the GRAS (generally recognized as safe) status and a potent protein secretion capacity. Secretory protein production greatly facilitates downstream processing and thus significantly reduces costs. However, not all heterologous proteins are secreted and intracellular production poses difficulties for quantification. To tackle this problem, we have established a so-called intracellular split GFP (iSplit GFP) assay in B. subtilis as a tool for the in vivo protein detection during expression in batch cultures and at a single-cell level. For the iSplit GFP assay, the eleventh ß-sheet of sfGFP is fused to a target protein and can complement a detector protein consisting of the respective truncated sfGFP (GFP1-10) to form fluorescent holo-GFP. RESULTS: As proof of concept, the GFP11-tag was fused C-terminally to the E. coli ß-glucuronidase GUS, resulting in fusion protein GUS11. Variable GUS and GUS11 production levels in B. subtilis were achieved by varying the ribosome binding site via spacers of increasing lengths (4-12 nucleotides) for the GUS-encoding gene. Differences in intracellular enzyme accumulation were determined by measuring the GUS11 enzymatic activity and subsequently by adding the detector protein to respective cell extracts. Moreover, the detector protein was co-produced with the GUS11 using a two-plasmid system, which enabled the in vivo detection and online monitoring of glucuronidase production. Using this system in combination with flow cytometry and microfluidics, we were able to monitor protein production at a single-cell level thus yielding information about intracellular protein distribution and culture heterogeneity. CONCLUSION: Our results demonstrate that the iSplit GFP assay is suitable for the detection, quantification and online monitoring of recombinant protein production in B. subtilis during cultivation as well as for analyzing production heterogeneity and intracellular localization at a single-cell level.