FungalTraits vs. FUNGuild: Comparison of Ecological Functional Assignments of Leaf- and Needle-Associated Fungi Across 12 Temperate Tree Species.

Recently, a new annotation tool "FungalTraits" was created based on the previous FUNGuild and FunFun databases, which has attracted high attention in the scientific community. These databases were widely used to gain more information from fungal sequencing datasets by assigning fungal functional traits. More than 1500 publications so far employed FUNGuild and the aim of this study is to compare this successful database with the recent FungalTraits database. Quality and quantity of the assignment by FUNGuild and FungalTraits to a fungal internal transcribed spacer (ITS)-based amplicon sequencing dataset on amplicon sequence variants (ASVs) were addressed. Sequencing dataset was derived from leaves and needles of 12 temperate broadleaved and coniferous tree species. We found that FungalTraits assigned more functional traits than FUNGuild, and especially the coverage of saprotrophs, plant pathogens, and endophytes was higher while lichenized fungi revealed similar findings. Moreover, ASVs derived from leaves and needles of each tree species were better assigned to all available fungal traits as well as to saprotrophs by FungalTraits compared to FUNGuild in particular for broadleaved tree species. Assigned ASV richness as well as fungal functional community composition was higher and more diverse after analyses with FungalTraits compared to FUNGuild. Moreover, datasets of both databases showed similar effect of environmental factors for saprotrophs but for endophytes, unidentical patterns of significant corresponding factors were obtained. As a conclusion, FungalTraits is superior to FUNGuild in assigning a higher quantity and quality of ASVs as well as a higher frequency of significant correlations with environmental factors.

Epidemiology, healthcare resource use, and mortality in patients with probable Dravet syndrome: A population-based study on German health insurance data.

OBJECTIVE: Ten-year retrospective study to assess burden of illness in patients with probable Dravet syndrome (DS) identified from German healthcare data. METHODS: In the absence of an International Classification of Diseases code, patients with probable DS were identified using a selection algorithm considering diagnoses and drug prescriptions. Primary analyses were prevalence and demographics; secondary analyses included healthcare costs, annual hospitalization rate (AHR) and length of stay (LOS), medication use, and mortality. RESULTS: In the final study year, 64 patients with probable DS (mean [range] age: 33.2 [3-82] years; male: 48%) were identified. Prevalence: 4.7 per 100,000 people. During the study, 160 patients with probable DS were identified and followed up for 1,261 patient-years. Mean cost of healthcare was €11,048 per patient-year (PPY), mostly attributable to inpatient care (47%), medication (26%), and services and devices (19%). Annual healthcare costs were significantly greater for those with prescribed rescue medication (15% of patient-years) vs. without (€16,123 vs. €10,125 PPY, p < 0.001). Mean (standard deviation [SD]) AHR and LOS were 1.1 (1.7) and 17.5 (33.5) days PPY. AHR was significantly greater in patients with prescribed rescue medication vs. without (1.6 [2.0] vs. 1.0 [1.6] PPY, p < 0.001). Mean (SD) number of antiseizure medications prescribed was 2.6 (1.2) PPY and 5.0 (2.5) over the entire observable time for each patient. Mortality rate was significantly higher for probable DS vs. matched controls (11.88% [19 events] vs. 1.19% [172 events], p < 0.001). CONCLUSION: Probable DS is associated with substantial healthcare costs in Germany.

Epidemiology, healthcare resource use, and mortality in patients with probable Lennox-Gastaut syndrome: A population-based study on German health insurance data.

OBJECTIVE: This retrospective study examined patients with probable Lennox-Gastaut syndrome (LGS) identified from German healthcare data. METHODS: This 10-year study (2007-2016) assessed healthcare insurance claims information from the Vilua Healthcare research database. A selection algorithm considering diagnoses and drug prescriptions identified patients with probable LGS. To increase the sensitivity of the identification algorithm, two populations were defined: all patients with probable LGS (broadly defined) and only those with a documented epilepsy diagnosis before 6 years of age (narrowly defined). This specific criterion was used as LGS typically has a peak seizure onset between age 3 and 5 years. Primary analyses were prevalence and demographics; secondary analyses included healthcare costs, hospitalization rate and length of stay (LOS), medication use, and mortality. RESULTS: In the final year of the study, 545 patients with broadly defined probable LGS (mean [range] age: 31.4 [2-89] years; male: 53%) were identified. Using the narrowly defined probable LGS definition, the number of patients was reduced to 102 (mean [range] age: 7.4 [2-14] years; male: 52%). Prevalence of broadly defined and narrowly defined probable LGS was 39.2 and 6.5 per 100,000 people. During the 10-year study, 208 patients with narrowly defined probable LGS were identified and followed up for 1379 patient-years. The mean annual cost of healthcare was €22,787 per patient-year (PPY); greatest costs were attributable to inpatient care (33%), home nursing care (13%), and medication (10%). Mean annual healthcare costs were significantly greater for those with prescribed rescue medication (45% of patient-years) versus those without (€33,872 vs. €13,785 PPY, p < 0.001). Mean (standard deviation [SD]) annual hospitalization rate was 1.6 (2.0) PPY with mean (SD) annual LOS of 22.7 (46.0) days. Annual hospitalization rate was significantly greater in those who were prescribed rescue medication versus those who were not (2.2 [2.3] vs. 1.1 [1.6] PPY, p < 0.001). The mean (SD) number of different medications prescribed was 11.3 (7.3) PPY and 33.8 (17.0) over the entire observable time per patient (OET); antiepileptic drugs only accounted for 2.1 (1.1) of the medications prescribed PPY and 3.8 (2.0) OET. Over the 10-year study period, mortality in patients with narrowly defined probable LGS was significantly higher than the matched control population (six events [2.88%] vs. oneevent [0.01%], p < 0.001). CONCLUSION: Annual healthcare costs incurred by patients with probable LGS in Germany were substantial, and mostly attributable to inpatient care, home nursing care, and medication. Patients prescribed with rescue medication incurred significantly greater costs than those who were not. Patients with narrowly defined probable LGS had a higher mortality rate versus control populations.

An Evaluation of the Swissmedic Regulatory Framework for New Active Substances.

BACKGROUND: Swissmedic is a major regulatory agency that has been benchmarking its timelines for 20 years. To better understand the Swissmedic review times and to examine whether measures introduced to accelerate the process were effective, a retrospective analysis was undertaken. The objective was to provide a breakdown of where time is spent in the phases of Swissmedic's approval process (validation, scientific assessment, authorisation) and how this compared to other major authorities. METHODS: Data on Swissmedic, EMA and FDA product approvals were collected from websites or through direct communication, using a standardised CIRS method and milestones previously identified, focusing on new active substances approved 2019-2021. RESULTS: In 2019, 2020, and 2021, Swissmedic median approval times were 520, 470, and 392 days, respectively. The decrease over this time was mainly observed in the Authorisation Phase and can be attributed to lower proportions of applications with multiple "labelling loops", in addition to shorter times for final label negotiation. While Swissmedic had the longest overall approval time (447 days) compared to EMA (428) and FDA (244), the timelines were more comparable when considering only the agency's time spent on the scientific assessment, with Swissmedic at 194 days, EMA at 218 days, and FDA at 184 days. CONCLUSIONS: These observations represent an important analysis of Swissmedic regulatory activity timelines, demonstrate the impact of process improvements, and emphasise the importance of measuring timelines. Swissmedic will continue to expedite its processes also by promoting international collaborations with like-minded authorities.

In Vivo Engineering and Transplantation of Axially Vascularized and Epithelialized Flaps in Rats.

The arteriovenous loop (AVL) model allows the in vivo engineering of axially vascularized flaps, the so-called AVL flaps. Although AVL flaps can be transplanted microsurgically to cover tissue defects, they lack an epithelial layer on the surface. Therefore, the objective of this study was to engineer axially vascularized AVL flaps with an accompanying epithelial layer for local defect reconstruction. In this study, AVLs were established in 20 male Lewis rats. Minimally invasive injection of keratinocytes onto the surface of the AVL flaps was performed on postoperative day (POD) 21. AVL flaps were explanted from 12 rats on POD 24 or POD 30, then the epithelium formed by the keratinocytes on the surface of the flaps was evaluated using immunofluorescence staining. In six other rats, the AVL flap was locally transposed to cover a critical defect in the rats' leg on POD 30 and explanted for analysis on POD 40. In two control rats, sodium chloride was applied instead of keratinocytes. These control flaps were also transplanted on POD 30 and explanted on POD 40. Our results revealed that 3 days after keratinocyte application, a loose single-layered epithelium was observed histologically on the AVL flaps surface, whereas after 9 days, a multilayered and structured epithelium had grown. The epithelium on the transplanted AVL flaps showed its physiological differentiation when being exposed to an air-liquid interface. Histologically, a layered epithelium identical to the rats' regular skin was formed. In the sodium chloride control group, no epithelium had been grown. This study clearly demonstrates that axially vascularized AVL flaps can be processed in the subcutaneous chamber by minimally invasive injection of keratinocytes. Thus, AVL flaps with an intact epithelial layer were engineered and could be successfully transplanted for local defect coverage in a small animal model.

Medication Adherence and Healthcare Costs in Chronically Ill Patients Using German Claims Data.

BACKGROUND AND OBJECTIVE: Despite the importance of medication adherence for chronically ill patients and the vast literature on its relationship to costs, this field suffers from methodological limitations. These are caused, amongst others, by the lack of generalizability of data sources, varying definitions of adherence, costs, and model specification. We aim to address this with different modeling approaches and to contribute evidence on the research question. METHODS: We extracted large cohorts of nine chronic diseases (n = 6747-402,898) from German claims data of stationary health insurances between 2012 and 2015 (t0-t3). Defined as the proportion of days covered by medication, we examined the relationship of adherence using several multiple regression models at baseline year t0 with annual total healthcare costs and four sub-categories. Models with concurrent, and differently time-lagged measurements of adherence and costs were compared. Exploratively, we applied non-linear models. RESULTS: Overall, we found a positive association between the proportion of days covered by medication and total costs, a weak association with outpatient costs, positive with pharmacy costs, and frequently negative with inpatient costs. There were major differences by disease and its severity but little between years, provided adherence and costs were not measured concurrently. The fit of linear models was mainly not inferior to that of non-linear models. CONCLUSIONS: The estimated effect on total costs differed from most other studies, which highlights concerns about generalizability, although effect estimates in sub-categories were as expected. Comparison of time lags indicates the importance of avoiding concurrent measurement. A non-linear relationship should be considered. These methodological approaches are valuable in future research on adherence and its consequences.

Risk Factors for Fear of Recurrence in Head and Neck Cancer Patients.

OBJECTIVE: Fear of recurrence (FoR) affects the quality of life of head and neck cancer survivors. Identification of factors predisposing to FoR may help to recognize and treat patients at risk. MATERIALS AND METHODS: For this exploratory study, 101 disease-free head and neck cancer survivors completed a cross-sectional survey in 2017 that included the FoR questionnaire at a random point in time during their follow-up. Additionally, the patients were asked to choose their favorite among four follow-up schedules with or without systematic imaging and varying frequency of visits. RESULTS: Elevated FoR was present in 36.6% of patients. Females and patients ≤65 years showed significantly higher FoR overall scores than males (score difference 3.40; CI 0.49-6.32; p = 0.022) and patients >65 years (score difference 4.25; CI 1.58-6.92; p = 0.002). A history of cancer recurrence or second primary malignancy increased the relative risk (RR) for elevated FoR (RR 1.7; CI 1.01-2.86; p = 0.046). Tumor stage and treatment modality were not significantly associated with elevated FoR or FoR overall score. Higher FoR overall scores were recorded in patients who favored intensive follow-up plans (mean overall FoR score 18 vs. 15; SD 7.7; p = 0.076) and systematic imaging in follow-up (17 vs. 13, SD 7.1; p = 0.034). CONCLUSION: Fear of recurrence in head and neck cancer patients is associated with female sex, younger age, and history of a past recurrence or second primary malignancy. Due to its high prevalence, it should be addressed in clinical practice and future research. LEVEL OF EVIDENCE: NA Laryngoscope, 133:1630-1637, 2023.

Acellular Human Placenta Small-Diameter Vessels as a Favorable Source of Super-Microsurgical Vascular Replacements: A Proof of Concept.

In this study, we aimed to evaluate the human placenta as a source of blood vessels that can be harvested for vascular graft fabrication in the submillimeter range. Our approach included graft modification to prevent thrombotic events. Submillimeter arterial grafts harvested from the human placenta were decellularized and chemically crosslinked to heparin. Graft performance was evaluated using a microsurgical arteriovenous loop (AVL) model in Lewis rats. Specimens were evaluated through hematoxylin-eosin and CD31 staining of histological sections to analyze host cell immigration and vascular remodeling. Graft patency was determined 3 weeks after implantation using a vascular patency test, histology, and micro-computed tomography. A total of 14 human placenta submillimeter vessel grafts were successfully decellularized and implanted into AVLs in rats. An appropriate inner diameter to graft length ratio of 0.81 ± 0.16 mm to 7.72 ± 3.20 mm was achieved in all animals. Grafts were left in situ for a mean of 24 ± 4 days. Decellularized human placental grafts had an overall patency rate of 71% and elicited no apparent immunological responses. Histological staining revealed host cell immigration into the graft and re-endothelialization of the vessel luminal surface. This study demonstrates that decellularized vascular grafts from the human placenta have the potential to serve as super-microsurgical vascular replacements.

Tree mycorrhizal type regulates leaf and needle microbial communities, affects microbial assembly and co-occurrence network patterns, and influences litter decomposition rates in temperate forest.

Background: Tree mycorrhizal types (arbuscular mycorrhizal fungi and ectomycorrhizal fungi) alter nutrient use traits and leaf physicochemical properties and, thus, affect leaf litter decomposition. However, little is known about how different tree mycorrhizal species affect the microbial diversity, community composition, function, and community assembly processes that govern leaf litter-dwelling microbes during leaf litter decomposition. Methods: In this study, we investigated the microbial diversity, community dynamics, and community assembly processes of nine temperate tree species using high-resolution molecular technique (Illumina sequencing), including broadleaved arbuscular mycorrhizal, broadleaved ectomycorrhizal, and coniferous ectomycorrhizal tree types, during leaf litter decomposition. Results and discussion: The leaves and needles of different tree mycorrhizal types significantly affected the microbial richness and community composition during leaf litter decomposition. Leaf litter mass loss was related to higher sequence reads of a few bacterial functional groups, particularly N-fixing bacteria. Furthermore, a link between bacterial and fungal community composition and hydrolytic and/or oxidative enzyme activity was found. The microbial communities in the leaf litter of different tree mycorrhizal types were governed by different proportions of determinism and stochasticity, which changed throughout litter decomposition. Specifically, determinism (mainly variable selection) controlling bacterial community composition increased over time. In contrast, stochasticity (mainly ecological drift) increasingly governed fungal community composition. Finally, the co-occurrence network analysis showed greater competition between bacteria and fungi in the early stages of litter decomposition and revealed a contrasting pattern between mycorrhizal types. Conclusion: Overall, we conclude that tree mycorrhizal types influence leaf litter quality, which affects microbial richness and community composition, and thus, leaf litter decomposition.

Treatment Costs of Colorectal Cancer by Sex and Age: Population-Based Study on Health Insurance Data from Germany.

Objective: Evidence on the cost-effectiveness of screening for colorectal cancer (CRC) in the German general population remains scarce as key input parameters, the costs to treat CRC, are largely unknown. Here, we provide detailed estimates on CRC treatment costs over time. Methods: Using insurance claims data from the Vilua healthcare research database, we included subjects with newly diagnosed CRC and subjects who died of CRC between 2012 and 2016. We assessed annualized CRC-related inpatient, outpatient and medication costs for up to five years after first diagnosis and prior to death, stratified by sex and age. Findings: We identified 1748 and 1117 subjects with follow-up data for at least 1 year after diagnosis and prior to death, respectively. In those newly diagnosed, average costs were highest in the first year after diagnosis (men, EUR 16,375−16,450; women, EUR 10,071−13,250) and dropped steeply in the following years, with no consistent pattern of differences with respect to age. Costs prior to death were substantially higher as compared to the initial phase of care and consistently on a high level even several years before death, peaking in the final year of life, with strong differences by sex and age (men vs. women, <70 years, EUR 34,351 vs. EUR 31,417; ≥70 years, EUR 14,463 vs. EUR 9930). Conclusion: Once clinically manifest, CRC causes substantial treatment costs over time, particularly in the palliative care setting. Strong differences in treatment costs by sex and age warrant further investigation.

Identification of target groups and individuals for adherence interventions using tree-based prediction models.

Background: In chronically ill patients, medication adherence during implementation can be crucial for treatment success and can decrease health costs. In some populations, regression models do not show this relationship. We aim to estimate subgroup-specific and personalized effects to identify target groups for interventions. Methods: We defined three cohorts of patients with type 1 diabetes (n = 12,713), type 2 diabetes (n = 85,162) and hyperlipidemia (n = 117,485) from German claims data between 2012 and 2015. We estimated the association of adherence during implementation in the first year (proportion of days covered) and mean total costs in the three following years, controlled for sex, age, Charlson's Comorbidity Index, initial total costs, severity of the disease and surrogates for health behavior. We fitted three different types of models on training data: 1) linear regression models for the overall conditional associations between adherence and costs, 2) model-based trees to identify subgroups of patients with heterogeneous adherence effects, and 3) model-based random forests to estimate personalized adherence effects. To assess the performance of the latter, we conditionally re-estimated the personalized effects using test data, the fixed structure of the forests, and fixed effect estimates of the remaining covariates. Results: 1) our simple linear regression model estimated a positive adherence effect, that is an increase in total costs of 10.73 Euro per PDC-point and year for diabetes type 1, 3.92 Euro for diabetes type 2 and 1.92 Euro for hyperlipidemia (all p ≤ 0.001). 2) The model-based tree detected subgroups with negative estimated adherence effects for diabetes type 2 (-1.69 Euro, 24.4% of cohort) and hyperlipidemia (-0.11 Euro, 36.1% and -5.50 Euro, 5.3%). 3) Our model-based random forest estimated personalized adherence effects with a significant proportion (4.2%-24.1%) of negative effects (up to -8.31 Euro). The precision of these estimates was high for diabetes type 2 and hyperlipidemia patients. Discussion: Our approach shows that tree-based models can identify patients with different adherence effects and the precision of personalized effects is measurable. Identified patients can form target groups for adherence-promotion interventions. The method can also be applied to other outcomes such as hospitalization risk to maximize positive health effects of an intervention.

Association of Multiprofessional Preoperative Assessment and Information for Patients With Head and Neck Cancer With Postoperative Outcomes.

IMPORTANCE: Multidisciplinary perioperative assessment for patients undergoing complex oncologic head and neck cancer (HNC) surgery is widely implemented. However, to our knowledge, the association of multiprofessional preoperative assessment, information, and briefing with postoperative outcomes has not been evaluated. OBJECTIVE: To compare postoperative complications, length of hospital stay (LOS), readmissions, mortality, and costs per case among patients undergoing complex oncologic HNC surgery before and after the implementation of a comprehensive preoperative multiprofessional assessment and information day (MUPAID). DESIGN, SETTING, AND PARTICIPANTS: This retrospective, single-center case-control study was conducted at a tertiary referral head and neck anticancer center/university cancer institute and compared patients with HNC who were undergoing complex oncological surgeries between January 2012 and July 2018 before (control group) and after (intervention group) implementation of the institutional MUPAID. Data analysis was conducted between 2019 and 2020. The intervention group comprised patients who participated in the MUPAID beginning in February 2015. These patients were assessed by a multiprofessional team and provided with structured and comprehensive information on the surgical procedure and its functional, social, financial, and psychological effects, as well as the postoperative care, rehabilitation, and follow-up period. Patients in the control group had also undergone complex oncologic HNC surgery and were selected through surgical procedure codes. MAIN OUTCOMES AND MEASURES: The end points were postoperative rate and severity of complications, LOS, readmissions, mortality, and costs per case. RESULTS: The study included 161 patients, 81 in the intervention (25 women [30.9%]) and 80 in the control group (18 women [22.5%]). The groups showed no relevant differences in sociodemographic, disease, and procedural characteristics. The intervention cohort presented with fewer major local and systemic complications (Clavien-Dindo score, III-V: 34.6% vs 52.5%; difference proportion, -0.179; 95% CI, -0.33 to -0.03), shorter median LOS (12 days [IQR, 10-16 days] vs 16 days [IQR, 11-20] days; effect size, 0.482; 95% CI Cohen d, 0.152-0.812) and decreased median charge per case ($50 848 [IQR, $42 510-$63 479] vs $69 602 [IQR, $45 631-$96 280]; effect size, 0.534; 95% CI Cohen d, 0.22-0.85). CONCLUSIONS AND RELEVANCE: The results of this case-control study suggest that MUPAID for patients who are undergoing complex oncologic HNC surgery is associated with shortened LOS and costs per case as well as decreased complications severity. These results are promising on a patient level in the potential to minimize individual treatment burden, as well as on an institutional and health care system level in the potential significant optimization of surgical outcomes and financial aspects.

The Free Myocutaneous Tensor Fasciae Latae Flap-A Workhorse Flap for Sternal Defect Reconstruction: A Single-Center Experience.

INTRODUCTION: Deep sternal wound infections (DSWI) after cardiac surgery pose a significant challenge in reconstructive surgery. In this context, free flaps represent well-established options. The objective of this study was to investigate the clinical outcome after free myocutaneous tensor fasciae latae (TFL) flap reconstruction of sternal defects, with a special focus on surgical complications and donor-site morbidity. METHODS: A retrospective chart review focused on patient demographics, operative details, and postoperative complications. Follow-up reexaminations included assessments of the range of motion and muscle strength at the donor-site. Patients completed the Quality of Life 36-item Short Form Health Survey (SF-36) as well as the Lower Extremity Functional Scale (LEFS) questionnaire and evaluated aesthetic and functional outcomes on a 6-point Likert scale. The Vancouver Scar Scale (VSS) and the Patient and Observer Scar Assessment Scales (POSAS) were used to rate scar appearance. RESULTS: A total of 46 patients (mean age: 67 ± 11 years) underwent sternal defect reconstruction with free TFL flaps between January 2010 and March 2021. The mean defect size was 194 ± 43 cm2. The mean operation time was 387 ± 120 min with a flap ischemia time of 63 ± 16 min. Acute microvascular complications due to flap pedicle thromboses occurred in three patients (7%). All flaps could be salvaged without complete flap loss. Partial flap loss of the distal TFL portion was observed in three patients (7%). All three patients required additional reconstruction with pedicled or local flaps. Upon follow-up, the range of motion (hip joint extension/flexion (p = 0.73), abduction/adduction (p = 0.29), and internal/external rotation (p = 0.07)) and muscle strength at the donor-sites did not differ from the contralateral sides (p = 0.25). Patient assessments of aesthetic and functional outcomes, as well as the median SF-36 (physical component summary (44, range of 33 to 57)) and LEFS (54, range if 35 to 65), showed good results with respect to patient comorbidities. The median VSS (3, range of 2 to 7) and POSAS (24, range of 18 to 34) showed satisfactory scar quality and scar appearance. CONCLUSION: The free TFL flap is a reliable, effective, and, therefore, valuable option for the reconstruction of extensive sternal defects in critically ill patients suffering from DSWIs. In addition, the TFL flap shows satisfactory functional and aesthetic results at the donor-site.

A poisoned apple: First insights into community assembly and networks of the fungal pathobiome of healthy-looking senescing leaves of temperate trees in mixed forest ecosystem.

Despite the abundance of observations of foliar pathogens, our knowledge is severely lacking regarding how the potential fungal pathobiome is structured and which processes determine community assembly. In this study, we addressed these questions by analysing the potential fungal pathobiome associated with the senescing leaves and needles of 12 temperate tree species. We compared fungal plant pathogen load in the senescing leaves/needles and demonstrated that healthy-looking leaves/needles are inhabited by diverse and distinct fungal plant pathogens. We detected 400 fungal plant pathogenic ASVs belonging to 130 genera. The fungal plant pathogenic generalist, Mycosphaerella, was found to be the potential most significant contributor to foliar disease in seedlings. The analyses of assembly process and co-occurrence network showed that the fungal plant pathogenic communities in different tree types are mainly determined by stochastic processes. However, the homogenising dispersal highly contributes in broadleaf trees, whereas ecological drift plays an important role in coniferious trees. The deterministic assembly processes (dominated by variable selection) contributed more in broadleaf trees as compared to coniferous trees. We found that pH and P level significantly corresponded with fungal plant pathogenic community compositions in both tree types. Our study provides the first insight and mechanistic understanding into the community assembly, networks, and complete taxonomy of the foliar fungal pathobiome in senescing leaves and needles.

More than you can see: Unraveling the ecology and biodiversity of lichenized fungi associated with leaves and needles of 12 temperate tree species using high-throughput sequencing.

Currently, lichen surveys are generally based on the examination of fruiting bodies. Lichens in the mycelial stage, in spores, or awaiting conditions for fruiting body formation are usually overlooked, even though they are important for maintaining biodiversity and ecosystem functions. This study aimed to explore the lichenized fungal community composition and richness associated with leaves and needles of 12 temperate tree species using Illumina MiSeq-based amplicon sequencing of the internal transcribed spacer (ITS) 2 region. Picea abies harbored the highest richness and number of lichenized fungal species. We found that the lichenized fungus Physcia adscendens dominated the leaves and needles of the most temperate tree species. Eleven lichenized fungal species detected in this study were recorded for the first time on leaves and needles. In addition, we identified Athallia cerinella, Fellhanera bouteillei, and Melanohalea exasperata that are on the German national red lists. Lichenized fungal richness was higher in conifer compared to broadleaf trees. Overall, tree species (within coniferous trees) and tree types (broadleaved vs. coniferous trees) harbored significantly different lichenized fungal community compositions pointing out the importance of host species. Diversity and community composition patterns of lichenized fungi were correlated mainly with tree species. Our study demonstrates that the diversity of foliicolous lichens associated with leaves and needles of 12 temperate tree species can be appropriately analyzed and functionally assigned using the ITS-based high-throughput sequencing. We highlighted the importance of conifers for maintaining the biodiversity of foliicolous lichens. Based on the discovery of many red list lichens, our methodological approach and results are important contributions to subsequent actions in the bio-conversation approaches.

Epidemiology, healthcare resource use, and mortality in patients with tuberous sclerosis complex: A population-based study on German health insurance data.

PURPOSE: 10-year retrospective study to assess burden of illness in individuals with tuberous sclerosis complex (TSC) identified from German healthcare data. METHODS: Patients with TSC were identified by International Classification of Diseases code Q85.1. Patients with epilepsy were identified by epilepsy diagnosis or antiseizure medication (ASM) prescription after TSC diagnosis. RESULTS: Using data from 2016 (final study year), 100 patients with TSC were identified (mean [range] age: 38 [1-86] years; male: 40%); prevalence: 7.9 per 100,000 (TSC), 2.2 per 100,000 (TSC with epilepsy). During the 10-year study period (2007-2016), 256 patients with TSC were identified and followed up for 1,784 patient-years (epilepsy: 36%, 616 patient-years). TSC manifestations/comorbidities (apart from epilepsy) were identified more frequently in patients with epilepsy than without. Mean annual healthcare costs for patients with TSC were €6,139 per patient-year (PPY), mostly attributable to medication (35%) and inpatient care (29%). Patients with epilepsy incurred costs more than double those without. Mean (standard deviation [SD]) annual hospitalisation rate (AHR) and length of stay (LOS) PPY: 0.5 (1.0) and 5.9 (18.6) days for TSC. AHR and LOS were greater in patients with epilepsy than without. Mean (SD) number of ASMs prescribed (TSC with epilepsy): 3.0 (2.3) over the entire observable time per patient. Mortality rates (vs. control): 5.08% (vs. 1.69%, p<0.001) for TSC, 7.53% (vs. 0.98%, p<0.001) for TSC with epilepsy, 3.68% (vs. 2.03%, p = 0.003) for TSC without epilepsy. CONCLUSION: Healthcare costs, resource utilisation, and mortality were greater in patients with TSC and epilepsy than those without epilepsy.

More than experience: a post-task reflection intervention among team members enhances performance in student teams confronted with a simulated resuscitation task-a prospective randomised trial.

Background: Teams that regularly step back from action and deliberately reflect on their performance and strategies show higher performance. Ad hoc emergency teams with changing team composition cannot develop such habits but may engage in short postaction reflection to discuss shortcomings of past performance and potential adaptations of their strategies for future similar tasks. This study aimed to test the effect of a short postaction self-led reflective team briefing on resuscitation performance in a simulator setting in terms of three performance parameters: hands-on time, coordination between chest compression and ventilation, and defibrillation. Methods: We performed a randomised controlled trial including 56 ad hoc formed teams of three fourth-year medical students each. All groups performed a resuscitation task, followed by a self-guided reflective briefing, based on a general instruction (n=28 teams), or an unrelated discussion session (control condition; n=29), followed by a second resuscitation task in the same team composition. Results: Adjusted for performance in the first task, teams in the reflection condition showed higher performance gain in the second resuscitation than teams in the control condition (6.21 percentage points (95% CI 1.31 to 11.10, p<0.001)) for basic hands-on performance; 15.0 percentage points (95% CI 2 to 28, p<0.001) for coordinative performance but non-significantly lower performance for defibrillation (-9%, 95% CI -27% to -9%, p=0.312). Conclusion: Even very short self-led postaction reflective briefings enhance basic resuscitation performance in ad hoc groups but may not influence more complex aspects of the task. We recommend including short self-led team debriefings as part of simulator training.

Amino acid variants of the HybB membrane subunit of Escherichia coli [NiFe]-hydrogenase-2 support a role in proton transfer.

[NiFe]-hydrogenase (Hyd) 2 of Escherichia coli has been proposed to generate proton motive force during H2 -oxidation, which it is dependent on if cells are incubated anaerobically with glycerol to drive reverse H2 -production. The integral membrane subunit HybB is required for proton transfer (PT) by Hyd-2 but has no cofactor. To provide evidence for PT by HybB, we analyzed the roles of conserved amino acid residues in a predicted proton channel. Exchange of conserved residues identified residues Y99, E133, H184, and E228 as mandatory for PT from the cytoplasm and quinol oxidation. In contrast, exchange of W54, D58, or R89 rendered Hyd-2 uni-directional and influenced the equilibrium. Our findings show that HybB is the key subunit in PT.

Application of a Dual Internally Quenched Fluorogenic Substrate in Screening for D-Arginine Specific Proteases.

The application of D-stereospecific proteases (DSPs) in resolution of racemic amino acids and in the semisynthesis of proteins has been a successful strategy. The main limitation for a broader application is, however, the accessibility of suitable DSPs covering multiple substrate specificities. To identify DSPs with novel primary substrate preferences, a fast specificity screening method using the easily accessible internally quenched fluorogenic substrate aminobenzoyl-D-arginyl-D-alanyl-p-nitroanilide was developed. By monitoring both UV/vis-absorbance and fluorescence signals at the same time it allows to detect two distinct D-amino acid substrate specificities simultaneously and separately with respect to the individual specificities. In order to identify novel DSP specificities for synthesis applications, DSPs specific for D-arginine were of special interest due to their potential ability as catalysts for substrate mimetics-mediated peptide and protein ligations. D-alanine in the substrate served as positive control and reference based on its known acceptance by numerous DSPs. In silico analysis suggested that DSPs are predominantly present in gram-positive microorganisms, therefore this study focused on the bacilli strains Bacillus thuringiensis and Bacillus subtilis as potential hosts of D-Arg-specific DSPs. While protease activities toward D-alanine were found in both organisms, a novel and so far unknown D-arginine specific DSP was detected within the culture supernatant of B. thuringiensis. Enrichment of this activity via cation exchange and size exclusion chromatography allowed isolation and further characterization of this novel enzyme consisting of a molecular mass of 37.7 kDa and an enzymatic activity of 8.3 U mg-1 for cleaving the D-Arg|D-Ala bond in the detecting substrate. Independent experiments also showed that the identified enzyme shows similarities to the class of penicillin binding proteins. In future applications this enzyme will be a promising starting point for the development of novel strategies for the semisynthesis of all-L-proteins.

Patients' preferences concerning follow-up after curative head and neck cancer treatment: A cross-sectional pilot study.

BACKGROUND: Evidence of the oncological benefit of scheduled follow-up in head and neck squamous cell carcinoma is weak; symptom-oriented self-referral may be an alternative. Patients' preferences regarding follow-up remain poorly investigated. METHODS: We conducted a cross-sectional survey among patients undergoing follow-up at a tertiary outpatient clinic, focusing on their preferences, correlating factors, fear of recurrence, and willingness for participation in a randomized trial on follow-up. RESULTS: Of 101 patients, 89.1% preferred scheduled follow-up to self-referral, 57% favored fewer visits than the current standard, and 85.1% endorsed regular imaging. Recurrence or second primary was associated with preference of intensive follow-up schedules (P = 0.02). There were trends for women and patients with high fear of recurrence score to favor intensive follow-up. Two-third of the participants declared willingness to participate in a randomized controlled trial. CONCLUSIONS: Patients' preferences only partially correspond to current follow-up guidelines. Recruitment for randomized controlled studies evaluating the value of follow-up seems feasible.