Specific molecular peak analysis by ion mobility spectrometry of volatile organic compounds in urine of COVID-19 patients: A novel diagnostic approach.

INTRODUCTION: SARS-CoV-2 is usually diagnosed from naso-/oropharyngeal swabs which are uncomfortable and prone to false results. This study investigated a novel diagnostic approach to Covid-19 measuring volatile organic compounds (VOC) from patients' urine. METHODS: Between June 2020 and February 2021, 84 patients with positive RT-PCR for SARS-CoV-2 were recruited as well as 54 symptomatic individuals with negative RT-PCR. Midstream urine samples were obtained for VOC analysis using ion mobility spectrometry (IMS) which detects individual molecular components of a gas sample based on their size, configuration, and charge after ionization. RESULTS: Peak analysis of the 84 Covid and 54 control samples showed good group separation. In total, 37 individual specific peaks were identified, 5 of which (P134, 198, 135, 75, 136) accounted for significant differences between groups, resulting in sensitivities of 89-94% and specificities of 82-94%. A decision tree was generated from the relevant peaks, leading to a combined sensitivity and specificity of 98% each. DISCUSSION: VOC-based diagnosis can establish a reliable separation between urine samples of Covid-19 patients and negative controls. Molecular peaks which apparently are disease-specific were identified. IMS is an additional non-invasive and cheap device for the diagnosis of this ongoing endemic infection. Further studies are needed to validate sensitivity and specificity.

Repurposing CPAP machines as stripped-down ventilators.

The worldwide shortage of medical-grade ventilators is a well-known issue, that has become one of the central topics during the COVID-19 pandemic. Given that these machines are expensive and have long lead times, one approach is to vacate them for patients in critical conditions while patients with mild to moderate symptoms are treated with stripped-down ventilators. We propose a mass-producible solution that can create such ventilators with minimum effort. The central part is a module that can be attached to CPAP machines and repurpose them as low-pressure ventilators. Here, we describe the concept and first measurements which underline the potential of our solution. Our approach may serve as a starting point for open-access ventilator technologies.

Characterization of proximal femoral anatomy in the skeletally-immature patient.

PURPOSE: The morphology of the proximal femur has been extensively studied in the adult population. However, no literature providing a comprehensive evaluation of the anatomy in paediatric patients exists. The current study aims to characterize such anatomy in skeletally-immature patients, examine potential differences between genders, and analyze how these anatomical parameters change with age. METHODS: Cadaveric femurs from the Hamann-Todd Osteological Collection were examined. Specimens with open physes and no skeletal disease or deformity were included for analysis. Age and gender were recorded for each specimen. Each femur was photographed in standardized modified axial and anteroposterior views. In all, 14 proximal femoral anatomical parameters were measured from these photographs. Comparisons between genders and age were calculated. RESULTS: A total of 43 femurs from ages four to 17 years met inclusion criteria. The majority were female (56%); no difference existed in age between genders (p = 0.62). The specimens had a neutral mean neck-shaft angle (130.7º) and anteversion (12.8º), and the sphericity of the ossified femoral heads was symmetrical. Male specimens had significantly higher alpha angles (p = 0.01), posterior offset (p = 0.02), neck width (p = 0.04) and head-neck length ratio (p = 0.02) values than female specimens. Strong positive correlations exist between length/size parameters and age, while negligible correlations were noted for angular measurements. CONCLUSIONS: This study establishes reference values for a comprehensive list of anatomical parameters for the skeletally-immature ossified proximal femur. It highlights gender differences in morphology and demonstrates that angular characteristics remain relatively stable while length parameters generally increase with age. LEVEL OF EVIDENCE: Level III Diagnostic.

[Multifocal Pulmonary Neuroendocrine Tumours: Genesis, Diagnostics and Treatment]. / Multifokale neuroendokrine Lungentumoren: Entstehung, Diagnostik und Behandlung.

Multifocal neuroendocrine lung tumour is a rare diagnosis. Multiple lung foci of different sizes are usually apparent on chest CT scans. It is assumed that multifocal neuroendocrine lung tumours originally develop from diffuse idiopathic pulmonary neuroendocrine cell hyperplasia (DIPNECH). This results in cell aggregations formed by proliferation of neuroendocrine cells that are already physiologically present in the bronchial system. If these cell proliferations break through the bronchial basement membrane, they are considered to constitute tumourlets if they measure ≤ 5 mm and carcinoid tumours if they are larger than 5 mm. The speed of proliferation of the cell hyperplasias appears to vary. Many of the patients are completely asymptomatic, the multifocal neuroendocrine lung tumours being diagnosed by chance. However, other patients complain of breathlessness, reduced physical capacity and cough. There may also be reduction of lung function. In these cases, chest HRCT often reveals peribronchial fibrosis or bronchiectasis in addition to the lung foci. Bronchoscopy is usually not helpful. Surgical lung biopsy is considered to be the diagnostic gold standard. Histological examination typically shows a mixture of cell hyperplasias, tumourlets and carcinoid tumours. There is no consensus on the treatment of multifocal neuroendocrine tumours. Taking the clinical situation and the chest HRCT findings as our starting point, we developed a stepwise approach that is guided by the success of the individual therapeutic procedures. The most favourable prognosis is found in affected people without clinical symptoms whose lung foci all measure less than 5 mm. In these cases the 5-year survival rate is over 90%.

[Pulmonary allergic reactions]. / Allergische Reaktionen der Lunge.

Allergic diseases of the lungs may affect the airways, the pulmonary parenchyma and the pulmonary vessels. The most relevant representatives are allergic asthma, hypersensitivity pneumonitis, bronchopulmonary aspergillosis and the Churg-Strauss syndrome. The type of allergic reaction and the pathophysiological consequences vary considerably between these entities. New drugs target specific mechanisms based on new insights into the pathogenetic processes of the underlying disease.

Complex gel permeation assays for screening combinatorial libraries.

Gel permeation methods have been commonly used to screen combinatorial libraries synthesized on a solid support. We report here three screens of combinatorial libraries using gel permeation assays. These include a simple enzymatic assay to identify inhibitors of the influenza enzyme neuraminidase, and two more complex assays designed to screen for inhibitors of the interleukin-8 (IL-8)-IL-8 receptor and the urokinase-urokinase receptor interactions, respectively. The IL-8 ligand-receptor assay makes use of IL-8 receptor-expressing cells attached to a membrane, thus enabling washing steps as part of the assay. The urokinase ligand-receptor assay employs an enzyme-linked immunosorbent assay-type format, previously thought to be amenable only to well-based assays. The results of these three screens are reported here, including the discovery of a novel series of acyclic inhibitors of neuraminidase. The development of complex assays in a gel permeation format allows for the routine screening of combinatorially as well as noncombinatorially made compound collections against virtually any kind of target, and is being widely used in our high throughput screening operations.

Novel inhibitors of Erm methyltransferases from NMR and parallel synthesis.

The Erm family of methyltransferases confers resistance to the macrolide-lincosamide-streptogramin type B (MLS) antibiotics through the methylation of 23S ribosomal RNA. Upon the methylation of RNA, the MLS antibiotics lose their ability to bind to the ribosome and exhibit their antibiotic activity. Using an NMR-based screen, we identified a series of triazine-containing compounds that bind weakly to ErmAM. These initial lead compounds were optimized by the parallel synthesis of a large number of analogues, resulting in compounds which inhibit the Erm-mediated methylation of rRNA in the low micromolar range. NMR and X-ray structures of enzyme/inhibitor complexes reveal that the inhibitors bind to the S-adenosylmethionine binding site on the Erm protein. These compounds represent novel methyltransferase inhibitors that serve as new leads for the reversal of Erm-mediated MLS antibiotic resistance.

Structural and functional characterization of potent antithrombotic oligonucleotides possessing both quadruplex and duplex motifs.

We report the results of a selection for single-stranded DNA oligonucleotide ligands to the serine protease thrombin using recently developed methods. This selection yielded a family of DNA sequences that conform to a consensus structure comprised of a unimolecular quadruplex motif and complementary flanking sequences capable of forming an additional Watson-Crick duplex motif. This novel quadruplex/duplex structure was not reported in a previous selection for DNA molecules which bind to thrombin [Bock et al. (1992) Nature 355, 564-566]. All quadruplex/duplex molecules tested bound to thrombin with higher affinity than quadruplex structures lacking the duplex structure. However, binding affinities did not always correlate with inhibitory potency since some molecules with high affinity were not potent inhibitors in vitro. 1H NMR spectroscopy studies demonstrated that the complementarity of bases in the duplex portion of a selected sequence allows it to form multimolecular structures. Constraining these molecules to the unimolecular quadruplex/duplex structure by bridging the 5' and 3' ends of the duplex motif with either triethylene glycol or disulfide bonds improved their thrombin inhibitory activity. All bridged quadruplex/duplex molecules were more potent inhibitors than molecules with only a quadruplex motif. Bridging the ends of these structures not only increased thrombin inhibition but also improved resistance to nucleases in serum more than 40-fold over the unbridged quadruplex. In addition, we have found that both the length and sequence of the duplex motif are important for inhibition.

In vitro evolution of intrinsically bent DNA.

DNA fragments which are intrinsically bent or curved migrate anomalously during electrophoresis through polyacrylamide gels. Starting with an initial population of approximately 10(12) unique DNA sequences, DNA which exhibited the kind of anomalous mobility associated with DNA bending was selected and enriched using a variation of the SELEX procedure. After seven rounds of selection and amplification, the vast majority of the remaining population of DNA fragments migrated as bent DNA. Cloning and sequencing of 30 individual sequences from this population has yielded information regarding the relationship between DNA sequence and bending. Some of the previous conclusions on DNA bending have been confirmed while others have been modified, by the results presented here. In addition, the dinucleotide base step CA/TG, which had not been thought to be a major factor in DNA bending, appears to be important.

E. coli promoter spacer regions contain nonrandom sequences which correlate to spacer length.

The -10 and -35 regions of E. coli promoter sequences are separated by a spacer region which has a consensus length of 17 base-pairs. This region is thought to contribute to promoter function by correctly positioning the two conserved regions. We have performed a statistical evaluation of 224 spacer sequences and found that spacers which deviate from the 17 base-pair consensus length have nonrandom sequences in their upstream ends. Spacer regions which are shorter than 17 base-pairs in length have a significantly higher than expected frequency of purine-purine and pyrimidine-pyrimidine homo-dinucleotides at the six upstream positions. Spacer regions which are longer than 17 base-pairs in length have a significantly higher than expected frequency of purine-pyrimidine and pyrimidine-purine hetero-dinucleotides at these positions. This suggests that the nature of the purine-pyrimidine sequence at the upstream end of spacer regions affect promoter function in a manner which is related to the spacer length. We examine the spacer sequences as a function of spacer length and discuss some possible explanations for the observed relationship between sequence and length.