SARS-CoV-2 inactivation in laboratory animal tissues with 4% formaldehyde or 5% glutaraldehyde for transfer to biosafety level 1 laboratories.

The SARS-CoV-2 pandemic required the immediate need to transfer inactivated tissue from biosafety level (BSL)-3 to BSL-1 areas to enable downstream analytical methods. No validated SARS-CoV-2 inactivation protocols were available for either formaldehyde (FA)-fixed or glutaraldehyde (GA)-fixed tissues. Therefore, representative tissue from ferrets and hamsters was spiked with 2.2 × 106 tissue culture infectious dose 50% per ml (TCID50/ml) SARS-CoV-2 or were obtained from mice experimentally infected with SARS-CoV-2. SARS-CoV-2 inactivation was demonstrated with 4% FA or 5% GA at room temperature for 72 hours by a titer reduction of up to 103.8 TCID50/ml in different animal tissues with a maximum protein content of 100 µg/mg and a thickness of up to 10 mm for FA and 8 mm for GA. Our protocols can be easily adapted for validating the inactivation of other pathogens to allow for the transfer of biological samples from BSL-3 areas to BSL-1 laboratories.

CYP19A1 mediates severe SARS-CoV-2 disease outcome in males.

Male sex represents one of the major risk factors for severe COVID-19 outcome. However, underlying mechanisms that mediate sex-dependent disease outcome are as yet unknown. Here, we identify the CYP19A1 gene encoding for the testosterone-to-estradiol metabolizing enzyme CYP19A1 (also known as aromatase) as a host factor that contributes to worsened disease outcome in SARS-CoV-2-infected males. We analyzed exome sequencing data obtained from a human COVID-19 cohort (n = 2,866) using a machine-learning approach and identify a CYP19A1-activity-increasing mutation to be associated with the development of severe disease in men but not women. We further analyzed human autopsy-derived lungs (n = 86) and detect increased pulmonary CYP19A1 expression at the time point of death in men compared with women. In the golden hamster model, we show that SARS-CoV-2 infection causes increased CYP19A1 expression in the lung that is associated with dysregulated plasma sex hormone levels and reduced long-term pulmonary function in males but not females. Treatment of SARS-CoV-2-infected hamsters with a clinically approved CYP19A1 inhibitor (letrozole) improves impaired lung function and supports recovery of imbalanced sex hormones specifically in males. Our study identifies CYP19A1 as a contributor to sex-specific SARS-CoV-2 disease outcome in males. Furthermore, inhibition of CYP19A1 by the clinically approved drug letrozole may furnish a new therapeutic strategy for individualized patient management and treatment.

Canine real-time detection of SARS-CoV-2 infections in the context of a mass screening event.

INTRODUCTION: Previous research demonstrated that medical scent detection dogs have the ability to distinguish SARS-CoV-2 positive from negative samples with high diagnostic accuracy. To deploy these dogs as a reliable screening method, it is mandatory to examine if canines maintain their high diagnostic accuracy in real-life screening settings. We conducted a study to evaluate the performance of medical scent detection dogs under real-life circumstances. METHODS: Eight dogs were trained to detect SARS-CoV-2 RT-qPCR-positive samples. Four concerts with a total of 2802 participants were held to evaluate canines' performance in screening individuals for SARS-CoV-2 infection. Sweat samples were taken from all participants and presented in a line-up setting. In addition, every participant had been tested with a SARS-CoV-2 specific rapid antigen test and a RT-qPCR and they provided information regarding age, sex, vaccination status and medical disease history. The participants' infection status was unknown at the time of canine testing. Safety measures such as mask wearing and distance keeping were ensured. RESULTS: The SARS-CoV-2 detection dogs achieved a diagnostic specificity of 99.93% (95% CI 99.74% to 99.99%) and a sensitivity of 81.58% (95% CI 66.58% to 90.78%), respectively. The overall rate of concordant results was 99.68%. The majority of the study population was vaccinated with varying vaccines and vaccination schemes, while several participants had chronic diseases and were under chronic medication. This did not influence dogs' decisions. CONCLUSION: Our results demonstrate that SARS-CoV-2 scent detection dogs achieved high diagnostic accuracy in a real-life scenario. The vaccination status, previous SARS-CoV-2 infection, chronic disease and medication of the participants did not influence the performance of the dogs in detecting the acute infection. This indicates that dogs provide a fast and reliable screening option for public events in which high-throughput screening is required.

Reversible Glutamate Coordination to High-Valent Nickel Protects the Active Site of a [NiFe] Hydrogenase from Oxygen.

NAD+-reducing [NiFe] hydrogenases are valuable biocatalysts for H2-based energy conversion and the regeneration of nucleotide cofactors. While most hydrogenases are sensitive toward O2 and elevated temperatures, the soluble NAD+-reducing [NiFe] hydrogenase from Hydrogenophilus thermoluteolus (HtSH) is O2-tolerant and thermostable. Thus, it represents a promising candidate for biotechnological applications. Here, we have investigated the catalytic activity and active-site structure of native HtSH and variants in which a glutamate residue in the active-site cavity was replaced by glutamine, alanine, and aspartate. Our biochemical, spectroscopic, and theoretical studies reveal that at least two active-site states of oxidized HtSH feature an unusual architecture in which the glutamate acts as a terminal ligand of the active-site nickel. This observation demonstrates that crystallographically observed glutamate coordination represents a native feature of the enzyme. One of these states is diamagnetic and characterized by a very high stretching frequency of an iron-bound active-site CO ligand. Supported by density-functional-theory calculations, we identify this state as a high-valent species with a biologically unprecedented formal Ni(IV) ground state. Detailed insights into its structure and dynamics were obtained by ultrafast and two-dimensional infrared spectroscopy, demonstrating that it represents a conformationally strained state with unusual bond properties. Our data further show that this state is selectively and reversibly formed under oxic conditions, especially upon rapid exposure to high O2 levels. We conclude that the kinetically controlled formation of this six-coordinate high-valent state represents a specific and precisely orchestrated stereoelectronic response toward O2 that could protect the enzyme from oxidative damage.

Modification of Skin Flaps for Single-Stage Reconstruction of the Nasal Ala Using All-Layer Suture: A Prospective Study.

BACKGROUND AND OBJECTIVES: After local flaps, it may be necessary to reconstruct the contour of the nasal ala. This is possible with a single-stage all-layer shaping suture. In the present study, the functional and aesthetic results after single-stage reconstruction of the nasal ala were prospectively evaluated. PATIENTS AND METHODS: Patients who underwent surgery for skin tumors of the nose between 06/2019 and 06/2020 who required reconstruction of the nasal ala as part of the defect closure and had an all-layer suture used were prospectively included in the study. A standardized evaluation of aesthetic and functional outcome was conducted by the patient and a physician at discharge as well as 4 weeks later. Patients additionally underwent a follow-up survey 6 months later. RESULTS: Thirty-seven patients were included in the study. Four weeks postoperatively, all flaps were found to be fully healed and vital. Aesthetic outcome at 4 weeks was rated as very good or good by physicians in 73% and by patients in 78.4%. Persistent complications due to reduced blood flow were not observed. CONCLUSION: The reshaping of the nasal ala as part of the defect reconstruction with an all-layer suture demonstrates very good aesthetic as well as functional results and can be performed in a single-stage procedure. .

Outcomes and costs of ureteroscopy, extracorporeal shockwave lithotripsy, and percutaneous nephrolithotomy for the treatment of urolithiasis: an analysis based on health insurance claims data in Germany.

PURPOSE: Comparisons of ureteroscopy (URS), extracorporeal shockwave lithotripsy (SWL), and percutaneous nephrolithotomy (PCNL) for urolithiasis considering long-term health and economic outcomes based on claims data are rare. Our aim was to analyze URS, SWL, and PCNL regarding complications within 30 days, re-intervention, healthcare costs, and sick leave days within 12 months, and to investigate inpatient and outpatient SWL treatment as the latter was introduced in Germany in 2011. METHODS: This retrospective cohort study based on German health insurance claims data included 164,203 urolithiasis cases in 2008-2016. We investigated the number of complications within 30 days, as well as time to re-intervention, number of sick leave days and hospital and ambulatory health care costs within a 12-month follow-up period. We applied negative binomial, Cox proportional hazard, gamma and two-part models and adjusted for patient variables. RESULTS: Compared to URS cases, SWL and PCNL had fewer 30-day complications, time to re-intervention within 12 months was decreased for SWL and PCNL, SWL and PCNL were correlated with a higher number of sick leave days, and SWL and particularly PCNL were associated with higher costs. SWL outpatients had fewer complications, re-interventions and lower costs than inpatients. This study was limited by the available information in claims data. CONCLUSION: URS cases showed benefits in terms of fewer re-interventions, fewer sick leave days, and lower healthcare costs. Only regarding complications, SWL was superior. This emphasizes URS as the most frequent treatment choice. Furthermore, SWL outpatients showed less costs, fewer complications, and re-interventions than inpatients.

Clinical pharmacy services in Germany: a national survey.

OBJECTIVES: Clinical pharmacy services in German hospitals appear to be underdeveloped compared with other European countries. However, recent developments have increased the interest in expanding these services. Detailed data about the current state of clinical pharmacy services in Germany are lacking. This survey establishes the current level of pharmacy services in Germany and the barriers to implementation. METHODS: An online survey conducted in 2017 was distributed to directors of all 389 German hospital pharmacies. The survey contained 26 questions addressing hospital and pharmacy characteristics, clinical pharmacy services provided, the number of clinical pharmacists and the frequency as well as the quality assurance of these services. RESULTS: There were 133 responses (34%). Of these, 84 (63%) pharmacies provided some form of clinical pharmacy services. Based on the 389 contacted pharmacies, a clinical pharmacy service is available in at least 22% of hospital pharmacies in Germany. On average there are 2.4 full-time equivalent (FTE) clinical pharmacists per hospital employed, although there is a wide variation in numbers (0.3-22 FTE) and service provision between hospitals. Clinical pharmacy services are generally provided on a daily or weekly basis, with a principal focus on general surgery, critical care and general medicine wards. CONCLUSIONS: This is the first survey providing a detailed picture of clinical pharmacy services in Germany. There is wide variation in clinical service provision among hospitals, with some hospitals having developed a comprehensive range of clinical services. Compared with other countries, particularly the UK where the focus has shifted to provision of 7-day clinical services, the gap in clinical pharmacy services remains large. The focus should be turned to refining clinical pharmacy services in hospital admissions and discharge planning while also improving Health IT, the opportunities for specialisation and aligning education in accordance with the EAHP common training framework.

Post-operative follow-up of 150 patients with sentinel lymph node biopsy under tumescence local anesthesia.

BACKGROUND: Sentinel lymph node biopsy (SLNB) is useful for staging of patients with melanoma. Although SLNB is mostly performed under general anesthesia (GA), tumescence local anesthesia (TLA) can also be used. However, less data are available regarding feasibility of SLNB under TLA. Here we present a post-operative follow-up of 150 patients. PATIENTS AND METHODS: We prospectively analyzed data from 150 patients with primary cutaneous malignant melanoma. We assessed pain, post-operative complications and patients' satisfaction after SLNB under TLA. RESULTS: 32 % of the patients reported post-operative pain within the first 48 h after SLNB. Seroma was the most frequent complication, as 29 seromas after SLNB were observed. Wound infection was observed in 3.3 % of the patients. 98.7 % of the patients were satisfied with SLNB under TLA. CONCLUSIONS: SLNB under TLA is a safe and feasible option and should be considered for patients with melanoma. Especially with multimorbid or elderly patients, the risks of GA can be avoided.

Local Recurrence Rates of Skin Tumors After Resection With Complete Circumferential Peripheral and Deep Margin Assessment-Identification of High-Risk Entities.

BACKGROUND: The local recurrence rates of malignant skin tumors after micrographic controlled surgery using complete circumferential peripheral and deep margin assessment (CCPDMA) are reported to be low. However, in daily practice, tumor entities with a significantly higher recurrence rate are found. OBJECTIVE: The aim of the investigation was to identify these high-risk tumors to develop approaches for risk stratification. PATIENTS AND METHODS: We included different malignant tumors that were surgically treated and examined with CCPDMA using paraffin sections and H&E staining. Re-excisions were performed until the tumor was completely removed. RESULTS: Ninety-nine thousand three hundred seventy-two tumors were included in the study; the follow-up period was 4 years (median). Eight tumor entities were identified as high-risk entities, showing a significantly higher local recurrence rate of over 20%: desmoplastic squamous cell carcinoma, desmoplastic melanoma, Merkel cell carcinoma, microcystic adnexal carcinoma, pleomorphic sarcoma, liposarcoma as well as angiosarcoma, and extramammary Paget's disease. CONCLUSION: Complete circumferential peripheral and deep margin assessment allows complete control of the resection margins and enables skin-sparing resections with low recurrence rates for basal cell carcinomas, nondesmoplastic squamous cell carcinomas, lentiginous melanomas, and dermatofibrosarcoma protuberans. However, other tumors show significantly higher recurrence rates and therefore need to be classified as high-risk tumors.

Sentinel lymph node biopsy for high-thickness cutaneous squamous cell carcinoma.

Squamous cell carcinomas are among the most common skin tumors and show a risk of metastasis depending on various factors such as tumor thickness, localization, histological subtype and immune status of the patient. Sentinel lymph node biopsy (SLNB) SLNB represents a possibility for assessing the locoregional lymph node status. In this study, the role of the SLNB in lymph node status and survival was analyzed. Retrospectively, 720 patients with high-risk squamous cell carcinoma (tumor thickness > 5 mm) were examined. 150 patients agreed to SLNB, 570 patients did not undergo histologic confirmation of lymph node status and were included directly in follow-up. In 101 patients, a sentinel lymph node was successfully marked and extirpated, followed by regular follow-up examinations.A total of 11.11% of the patients showed lymph node metastasis in the course of their treatment, with no difference in the proportion of patients in the SLNB group (11.9%) and the observation group (11.4%) (p = 0.873). The proportion of distant metastasis also did not differ between the groups (p = 0.898). In 3.96% of the patients in the SLNB group, a metastasis was found in the sentinel lymph node. Tumor-specific death was observed in 7.14% of the patients in the SLNB group and 4.74% in the observation group (p = 0.269). Although SLNB is a principally suitable method for determining lymph node status, the available data do not provide any benefit regarding further metastasis or tumor-specific survival.

Impact of morbidity on care need increase and mortality in nursing homes: a retrospective longitudinal study using administrative claims data.

BACKGROUND: A growing number of older people are care dependent and live in nursing homes, which accounts for the majority of long-term-care spending. Specific medical conditions and resident characteristics may serve as risk factors predicting negative health outcomes. We investigated the association between the risk of increasing care need and chronic medical conditions among nursing home residents, allowing for the competing risk of mortality. METHODS: In this retrospective longitudinal study based on health insurance claims data, we investigated 20,485 older adults (≥65 years) admitted to German nursing homes between April 2007 and March 2014 with care need level 1 or 2 (according to the three level classification of the German long-term care insurance). This classification is based on required daily time needed for assistance. The outcome was care level change. Medical conditions were determined according to 31 Charlson and Elixhauser conditions. Competing risks analyses were applied to identify chronic medical conditions associated with risk of care level change and mortality. RESULTS: The probability for care level change and mortality acted in opposite directions. Dementia was associated with increased probability of care level change compared to other conditions. Patients who had cancer, myocardial infarction, congestive heart failure, cardiac arrhythmias, renal failure, chronic pulmonary disease, weight loss, or recent hospitalization were more likely to die, as well as residents with paralysis and obesity when admitted with care level 2. CONCLUSION: This paper identified risk groups of nursing home residents which are particularly prone to increasing care need or mortality. This enables focusing on these risk group to offer prevention or special treatment. Moreover, residents seemed to follow specific trajectories depending on their medical conditions. Some were more prone to increased care need while others had a high risk of mortality instead. Several conditions were neither related to increased care need nor mortality, e.g., valvular, cerebrovascular or liver disease, peripheral vascular disorder, blood loss anemia, depression, drug abuse and psychosis. Knowledge of functional status trajectories of residents over time after nursing home admission can help decision-makers when planning and preparing future care provision strategies (e.g., planning of staffing, physical equipment and financial resources).

[Surgical techniques in the treatment of basal cell carcinoma-a prospective investigation]. / Chirurgische Techniken in der Therapie des Basalzellkarzinoms ­ eine prospektive Untersuchung.

BACKGROUND: Basal cell carcinomas are the most frequent epithelial skin tumors and a frequent indication for dermatological interventions. Despite the establishment of drug treatment options, surgery is still the treatment of choice. Various options are available ranging from curettage to complex dermatosurgical procedures. In addition to the main aspect of low local recurrence rates, esthetic factors and the number of procedures are also important for the choice of treatment. METHODS: In this study 347 patients with 398 basal cell carcinomas (nodular type, diameter up to 10 mm) were prospectively examined. The patients were randomized into two treatment arms: in one group the tumor was treated by ring curettage and in the other group the tumor was excised. Patients who underwent 3D histologically controlled surgery due to basal cell carcinoma during the same investigation period served as controls. RESULTS: The highest local recurrence rate was observed after curettage (14.0%), whereas the group with 3D histology had the lowest recurrence rate (0.9%, p < 0.001). In the 3D group, more re-excisions were required to achieve complete tumor clearance compared to the group who underwent excisions with serial section histology. Patients rated the esthetic outcome best after curettage. The median follow-up was 3.9 years. CONCLUSION: The choice of surgical treatment for small nodular basal cell carcinomas depends on individual circumstances. Excisions controlled by 3D histology with wound closure after complete tumor clearance showed the lowest recurrence rate in our study; however, curettage is also a possible surgical treatment option with minimal effort and an acceptable recurrence rate, which can lead to good esthetic results.

Scent dog identification of samples from COVID-19 patients - a pilot study.

BACKGROUND: As the COVID-19 pandemic continues to spread, early, ideally real-time, identification of SARS-CoV-2 infected individuals is pivotal in interrupting infection chains. Volatile organic compounds produced during respiratory infections can cause specific scent imprints, which can be detected by trained dogs with a high rate of precision. METHODS: Eight detection dogs were trained for 1 week to detect saliva or tracheobronchial secretions of SARS-CoV-2 infected patients in a randomised, double-blinded and controlled study. RESULTS: The dogs were able to discriminate between samples of infected (positive) and non-infected (negative) individuals with average diagnostic sensitivity of 82.63% (95% confidence interval [CI]: 82.02-83.24%) and specificity of 96.35% (95% CI: 96.31-96.39%). During the presentation of 1012 randomised samples, the dogs achieved an overall average detection rate of 94% (±3.4%) with 157 correct indications of positive, 792 correct rejections of negative, 33 incorrect indications of negative or incorrect rejections of 30 positive sample presentations. CONCLUSIONS: These preliminary findings indicate that trained detection dogs can identify respiratory secretion samples from hospitalised and clinically diseased SARS-CoV-2 infected individuals by discriminating between samples from SARS-CoV-2 infected patients and negative controls. This data may form the basis for the reliable screening method of SARS-CoV-2 infected people.

Regional variation of care dependency after hip fracture in Germany: A retrospective cohort study using health insurance claims data.

OBJECTIVE: To investigate variation of care dependency after hip fracture across German regions based on the assessment by the German statutory long-term care insurance. DATA SOURCES/STUDY SETTING: Patient-level statutory health and long-term care insurance claims data from 2009-2011 and official statistical data from Germany. STUDY DESIGN: We performed a retrospective cohort study. Investigated multinomial outcome categories were increase in care dependency (new onset or a higher care dependency than pre-fracture), no change as reference and death as competing risk in the quarterly period following hip fracture (follow-up 3 months). Regional variation was operationalized with the variance of regional-level random intercepts based on generalized linear mixed models. We adjusted for patient and regional characteristics. PRINCIPAL FINDINGS: The study included 122,887 hip fracture patients in 95 German postal code regions. Crude outcomes were 30.87% increase in care dependency and 14.35% death. Results indicated modest variation on regional level. Male sex, increasing age, increasing comorbidity, pertrochanteric and subtrochanteric fracture site compared to femoral neck, time from hospital admission to surgery of 3 or more days, as well as increasing inpatient length of stay, non-participation in rehabilitation and regions with lower hospital density were positively associated with an increase in care dependency. CONCLUSIONS: Several characteristics on patient and regional level associated with the outcome were identified. Variation in the increase in care dependency after hip fracture appeared to be attributable primarily to patient characteristics. Variation on regional level was only modest.

Treatment utilization and treatment barriers in individuals with body dysmorphic disorder.

BACKGROUND: Although effective treatments are available, most individuals with body dysmorphic disorder (BDD) do not receive an appropriate diagnosis or treatment. We aimed to examine treatment utilization and barriers to treatment, and to identify associated socio-demographic and clinical characteristics. METHODS: German individuals completed an online self-report survey of appearance concerns. A sample of N = 429 individuals met criteria for BDD. We examined the frequency of treatment utilization and barriers, analyzed comparisons between treated and untreated individuals and assessed the relationships of socio-demographic and clinical features with mental health treatment utilization and treatment barriers, respectively. RESULTS: Only 15.2% of the individuals with BDD had been diagnosed with BDD, and lifetime rates of mental health treatment were low (39.9%). Individuals endorsed multiple barriers to mental health treatment, especially shame, low perceived need and a preference for cosmetic and medical treatments. Associated features were identified, including age, a BDD diagnosis, body dysmorphic symptom severity, a likely major depressive disorder, prior cosmetic surgery, and insight. CONCLUSIONS: The results of this largest study to date highlight that BDD is still underrecognized and undertreated even in a country with extensive mental health care and few financial barriers. We discuss modifiable factors and strategies to foster awareness of BDD in sufferers and professionals to improve treatment dissemination and to reduce treatment barriers.

Camelids and Cattle Are Dead-End Hosts for Peste-des-Petits-Ruminants Virus.

Peste-des-petits-ruminants virus (PPRV) causes a severe respiratory disease in small ruminants. The possible impact of different atypical host species in the spread and planed worldwide eradication of PPRV remains to be clarified. Recent transmission trials with the virulent PPRV lineage IV (LIV)-strain Kurdistan/2011 revealed that pigs and wild boar are possible sources of PPRV-infection. We therefore investigated the role of cattle, llamas, alpacas, and dromedary camels in transmission trials using the Kurdistan/2011 strain for intranasal infection and integrated a literature review for a proper evaluation of their host traits and role in PPRV-transmission. Cattle and camelids developed no clinical signs, no viremia, shed no or only low PPRV-RNA loads in swab samples and did not transmit any PPRV to the contact animals. The distribution of PPRV-RNA or antigen in lymphoid organs was similar in cattle and camelids although generally lower compared to suids and small ruminants. In the typical small ruminant hosts, the tissue tropism, pathogenesis and disease expression after PPRV-infection is associated with infection of immune and epithelial cells via SLAM and nectin-4 receptors, respectively. We therefore suggest a different pathogenesis in cattle and camelids and both as dead-end hosts for PPRV.

Evaluation of an IGM-specific ELISA for early detection of bluetongue virus infections in domestic ruminants sera.

Competitive-ELISA (c-ELISA) is the most widely used serological test for the detection of Bluetongue virus (BTV) viral protein 7 (VP7) antibodies (Ab). However, these BTV c-ELISAs cannot to distinguish between IgG and IgM. IgM Ab are generated shortly after the primary immune response against an infectious agent, indicating a recent infection or exposure to antigens, such as after vaccination. Because the BTV genome or anti-VP7 Ab can be detected in ruminant blood months after infection, BTV diagnostic tools cannot discriminate between recent and old infections. In this study, we evaluated an IgM-capture ELISA prototype to detect ruminant anti-BTV VP7 IgM on 1,650 serum samples from cattle, sheep, or goats. Animals were BTV-naive, infected, or/and vaccinated with BTV-1, -2, -4, -8, -9, -16, or -27, and we also included 30 sera from cattle infected with the Epizootic haemorrhagic disease virus (EHDV) serotype 6. Results demonstrated that this ELISA kit is specific and can detect the presence of IgM with satisfactory diagnostic specificity and sensitivity from 1 to 5 weeks after BTV infection in domestic ruminants (for goats and cattle; for sheep, at least up to 24 days). The peak of anti-VP7 IgM was reached when the level of infectious viruses and BTV RNA in blood were the highest. The possibility of detecting BTV-RNA in IgM-positive sera allows the amplification and sequencing of the partial RNA segment 2 (encoding the serotype specific to VP2) to determine the causative BTV serotype/strain. Therefore, BTV IgM ELISA can detect the introduction of BTV (or EHDV) in an area with BTV-seropositive domestic animals regardless of their serological BTV status. This approach may also be of particular interest for retrospective epidemiological studies on frozen serum samples.