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Unlike the native surface of the implant material (Ti6Al4V), oxidation with H2O2 leads to increased binding of the effective antimicrobial agent poly(hexamethylene) biguanide [PHMB]. However, treating with NaOH instead results in an even higher PHMB mass coverage. After oxidation with H2O2, strong differences in the PHMB adsorption capability between polished and corundum-blasted surfaces appear, indicating a roughness dependence. After NaOH treatment, no such effect was observed. The wetting properties of specimens treated with either H2O2 or NaOH prior to PHMB exposure clearly varied. To unravel the nature of this interaction, widespread in silico and in vitro experiments were performed. Methods: By X-ray photoelectron spectroscopy, scanning electron microscopy, water contact angle measurements and MD simulations, we characterized the interplay between the polycationic antimicrobial agent and the implant surface. A theoretical model for PHMB micelles is tested for its wetting properties and compared to carbon contaminated TiO2. In addition, quantitation of anionic functional group equivalents, the binding properties of PHMB with blocked amino end-group, and the ability to bind chlorhexidine digluconate (CHG) were investigated. Ultimately, the capability of osteoblasts to build calcium apatite, and the activity of alkaline phosphatase on PHMB coated specimens, were determined. Results: Simulated water contact angles on carbon contaminated TiO2 surfaces and PHMB micelle models reveal little influence of PHMB on the wetting properties and point out the major influence of remaining and recovering contamination from ambient air. Testing PHMB adsorption beyond the critical micelle concentration and subsequent staining reveals an island-like pattern with H2O2 as compared to an evenly modified surface with NaOH. Both CHG and PHMB, with blocked amino end groups, were adsorbed on the treated surfaces, thus negating the significant influence of PHMB’s terminal groups. The ability of osteoblasts to produce calcium apatite and alkaline phosphatase is not negatively impaired for PHMB mass coverages up to 8 μg/specimen. Conclusion: Differences in PHMB adsorption are triggered by the number of anionic groups and carbon contaminants, both of which depend on the specimen pre-treatment. With more PHMB covering, the implant surface is protected against the capture of new contamination from the ambient air, thus building a robust antimicrobial and biocompatible surface coating.
Purpose: To (1) describe the prevalence of abnormal sleep quality in patients with hip abductor tears (HAT), to (2) determine whether sleep quality improves after open HAT repair, and to (3) to report clinical short-term outcomes in patients undergoing open HAT repair. Methods: The data of 28 patients (29 hips) who underwant open HAT repair were prospectively analyzed at midterm follow-up. The Pittsburgh Sleep Quality Index (PSQI), modified Harris Hip Score (mHHS), the University of California, Los Angeles activity scale (UCLA), and Visual Analog Scale (VAS) for pain were determined via questionnaire. Paired t-tests were applied to compare preoperative and post-operative Patient-reported Outcome Measures (PROMs). Logistic regression was performed to determine the association between PSQI improvement achievement and demographic variables (laterality, sex, age, body-mass-index (BMI), and preoperative mHHS). The minimal clinically important difference (MCID) was calculated for the mHHS. Results: A total of 28 patients were included. Four patients (14.3%) suffered post-operative complications after open HAT repair. The predominance of patients was female (77.4%), with a mean age of 60 ± 13 years. The average follow-up was 30.35 ± 16.62 months. Preoperatively, 27 (96.4%) patients experienced poor sleep quality (PSQI > 5); at follow-up, 7 (25%) patients experienced poor sleep quality. Univariate logistical regression analysis demonstrated no significant association between preoperative demographic data and achieving postoperative PSQI < 5. The MCID of mHHS was calculated to be 12.5. Overall, 90% of patients achieved MCID for mHHS. Conclusion: Preoperative sleep quality was impaired in 96.4% of HAT patients (PSQI > 5). However, these patients showed an improvement in sleep disturbances after open HAT repair in the early postoperative period. Ninety percent of patients showed significant improvements in mHHS and achieved the corresponding MCID. Level of Evidence: Case series; Level IV.
Background:
Arthroscopic treatment of femoroacetabular impingement syndrome (FAIS) has become a common procedure. However, meaningful long-term clinical outcomes have not been defined.
Purpose:
To define the minimal clinically important difference (MCID), substantial clinical benefit (SCB), and patient acceptable symptomatic state (PASS) for the modified Harris Hip Score (mHHS) at a minimum 10-year follow-up in patients undergoing arthroscopic treatment for FAIS and identify preoperative predictors for achievement of the MCID, SCB, and PASS.
Study Design:
Case-control study; Level of evidence, 3.
Methods:
A consecutive series of patients undergoing arthroscopic treatment for FAIS between 2007 and 2009 with a minimum 10-year follow-up was analyzed. Patient data included patient characteristics, radiographic parameters, and the pre- and postoperative mHHS and visual analog scale (VAS) for pain score. Paired t tests were used to compare the patient-reported outcome measures (PROMs). The MCID was determined by calculating half of the standard deviation, and SCB and PASS were calculated by the anchor method. Correlation and logistic regression analyses were conducted to identify predictors for the achievement of the MCID, SCB, and PASS.
Results:
A total of 44 patients (27 men, 17 women) were included. The mean age and body mass index were 42.2 years (range, 16-67 years) and 22.3 kg/m2 (range, 16.76-29.78 kg/m2), respectively. The MCID, absolute SCB, net change SCB, and PASS of the mHHS were calculated to be 19.6, 90.1, 31.5, and 84.4 points, respectively. Preoperative symptom duration was identified as an independent predictor for the achievement of meaningful clinical outcomes. The median symptom durations for patients who achieved the MCID, absolute SCB, net change SCB, and PASS were 11.7, 9.1, 9.0, and 10.8 months, respectively. The median symptom duration for patients who did not achieve the MCID, absolute SCB, net change SCB, and PASS were 15.8, 17.4, 17.3, and 18.4 months, respectively. No other statistically significant correlations were found.
Conclusion:
The preoperative duration of symptoms was identified as an independent predictor for achievement of the MCID, SCB, and PASS. These findings can be helpful in accelerating the transition to surgical treatment of FAIS.
Adaptation mechanisms within the B cell composition for successful human and murine pregnancies.
(2021)
Introduction
A well-balanced immune maternal status is essential for favourable outcome of pregnancy. Due to their complexities, not all immune adaptations that promote tolerance during pregnancy are known. To understand the adaptation of the B cell compartment, we analysed and compared B cell lymphopoiesis in different lymphoid tissues in a number of murine models.
Furthermore, we focused on the humoral immune response during pregnancy. We analysed immunoglobulin profiles in human subjects and mice during pregnancy.
These cellular alterations are subject to the influence of chemokines, among others. Therefore, we assessed serum levels of B cell activation factor to clarify its effects during pregnancy.
Methods
For analysis of the human peripheral B cell compartment, peripheral blood samples from age-matched non-pregnant and pregnant women without pregnancy complications, immunological disease or acute/chronic inflammation were collected and sub-classified into four different groups: non-pregnant, and first, second, or third trimester of pregnancy. The experiments, based on a mouse model, were performed with 8-week-old female mice: clinically healthy non-pregnant (CBA/J (H2k)), pregnant mice with normal gestation (BALB/c (H2d) x CBA/J (H2k)), and mice with pregnancy loss (DBA/2J (H2d) x CBA/J (H2k)). Subsequently, peripheral blood mononuclear cells from blood and lymphatic organs were isolated following standard protocols. The B cell analysis was performed by flow cytometry. The immunoglobulin serum levels of the human and murine subgroups were quantitated using Bio-Plex isotyping assay and analysed by a Bio-Plex reader. To quantify B cell activating factor (BAFF) in serum of pregnant and non-pregnant mice a BAFF enzyme-linked immunosorbent assay was used. The concentrations were determined by using a FLUOstar OPTIMA microplate reader. All statistical analyses were performed using the Kruskal–Wallis test with Dunn’s post-test in GraphPad Prism software. P values of < 0.05 were considered statistically significant.
Results
We were able to demonstrate B cell lymphopenia in mice bone marrow downstream of pre-pro B cells, irrespective of pregnancy outcome. The mature bone marrow B cells did not show this adjustment mechanism during normal gestation.
Closer inspection of the splenic tissue revealed expansion and activation of marginal zone B cells in mice with a normal pregnancy. However, this was not observed in mice suffering from pregnancy disturbances. Natural antibodies secreted from marginal zone B cells were also present at higher concentrations in serum of pregnant mice, compared to non-pregnant animals.
We also found significantly higher levels of natural antibodies in serum of pregnant women compared to non-pregnant age-matched controls. Analysis showed significantly lower levels of BAFF in mice with normal pregnancy as compared to non-pregnant mice.
Conclusions
We are able to show mechanisms within the B cell compartment as well as the change within the natural antibodies that might be crucial for successful pregnancy in both humans and mice. Furthermore, BAFF seems to play a central role as a mediator of peripheral B cell compartment and B cell lymphopoiesis in the bone marrow for successful pregnancy.
Endothelial dysfunction (ED) comes with age, even without overt vessel damage such as that which occurs in atherosclerosis and diabetic vasculopathy. We hypothesized that aging would affect the downstream signalling of the endothelial nitric oxide (NO) system in the vascular smooth muscle (VSM). With this in mind, resistance mesenteric arteries were isolated from 13-week (juvenile) and 40-week-old (aged) mice and tested under isometric conditions using wire myography. Acetylcholine (ACh)-induced relaxation was reduced in aged as compared to juvenile vessels. Pretreatment with L-NAME, which inhibits nitrix oxide synthases (NOS), decreased ACh-mediated vasorelaxation, whereby differences in vasorelaxation between groups disappeared. Endothelium-independent vasorelaxation by the NO donor sodium nitroprusside (SNP) was similar in both groups; however, SNP bolus application (10−6 mol L−1) as well as soluble guanylyl cyclase (sGC) activation by runcaciguat (10−6 mol L−1) caused faster responses in juvenile vessels. This was accompanied by higher cGMP concentrations and a stronger response to the PDE5 inhibitor sildenafil in juvenile vessels. Mesenteric arteries and aortas did not reveal apparent histological differences between groups (van Gieson staining). The mRNA expression of the α1 and α2 subunits of sGC was lower in aged animals, as was PDE5 mRNA expression. In conclusion, vasorelaxation is compromised at an early age in mice even in the absence of histopathological alterations. Vascular smooth muscle sGC is a key element in aged vessel dysfunction.
ABSTRACT
The Upper Pleistocene geoarchives in the south‐eastern Carpathian Basin are represented predominantly by loess–palaeosol records. In 2015, a 10 m sediment core composed of clay‐rich lacustrine sediments was recovered by vibracoring a dry lake basin located between the Vršac Mountains (Serbia) and the Banat Sands in the south‐eastern Carpathian Basin; a location relevant for placing regional archaeological results in a palaeoenvironmental context. Here, we present results from geoelectrical prospection and a lithostratigraphic interpretation of this sequence supported by a detailed granulometric study supplemented by ostracod analysis. An age model based on luminescence dating is discussed against sedimentological proxy data and its implication for palaeoenvironmental change. The cores show a stratigraphy of lighter ochre‐coloured and darker greyish sediment, related to the deposition of clay and silt trapped in an aquatic environment. Geophysical measurements show ~20 m thick lacustrine sediments. The grain‐size distributions including the variability in fine clay are indicative of a lacustrine environment. Fine particles were brought into the depositional environments by aquatic input and settled from suspension; also, direct dust input is constrained by grain‐size results. Riverine input and aeolian dust input interplayed at the locality.
Technological advances in light microscopy have always gone hand in hand with unprecedented biological insight. For microbiology, light microscopy even played a founding role in the conception of the entire discipline. The ability to observe pathogens that would otherwise evade human observation makes it a critical necessity and an indispensable tool to infectious disease research. Thus, the aim of this thesis was to optimize, extend, and functionally apply advanced light microscopy techniques to elucidate spatio-temporal and spatio-morphological components of bacterial and viral infection in vitro and in vivo.
Pathogens are in a constant arms race with the host’s immune system. By finding ways to circumvent host-mediated immune responses, they try to evade elimination and facilitate their own propagation. The first study (publication I) demonstrated that the obligate intracellular pathogen Coxiella burnetii is not just able to infect natural killer (NK) cells, but is actually capable of surviving the harsh degradative conditions in the cytotoxic lymphocyte’s granules. Using live-cell imaging of reporter-expressing Coxiella burnetii, the transient NK cell passage was closely monitored to provide detailed spatio-temporal information on this dynamic process in support of a range of static analyses. Bacterial release from NK cells was pinpointed to a time frame between 24 to 48 hours post-infection and the duration of release to about 15 minutes.
The second approach (publications II-V) aimed at shedding light on the greater spatio-morphological context of virus infection. Thus far, most studies investigating the distribution or tropism of viruses in vivo have used conventional immunohistochemistry in thin sections. Omitting the native spatial context of the infection site in vivo inherently bears the risk of incomplete description. While the microscopic tools and sample preparation protocols needed for volumetric 3D immunofluorescence imaging have recently been made available, they had not gained a foothold in virus research yet. An integral part of this thesis was concerned with the assessment and optimization of available tissue optical clearing protocols to develop an immunofluorescence-compatible 3D imaging pipeline for the investigation of virus infection inside its intact spatio-morphological environment (publication II). This formed the basis for all subsequent volumetric analyses of virus infection in vivo presented here. Consequently, this thesis provided a valuable proof of concept and blueprints for future virus research on the mesoscopic scale of host-pathogen interactions in vivo (publications II-V), using rabies virus (RABV; publications II-IV) and the newly-emerged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2; publication V) as infection models for the nervous system and the respiratory tract, respectively.
Applying and further improving this volumetric 3D imaging workflow enabled unprecedented insights into the comprehensive in vivo cell tropism of RABV in the central (CNS) (publication III) and peripheral nervous system (PNS) (publication IV). Accordingly, differential infection of CNS-resident astrocytes by pathogenic and lab-attenuated RABV was demonstrated (publication III). While either virus variant showed equal capacity to infect neurons, as demonstrated by quantitative image analysis, only pathogenic field RABVs were able to establish non-abortive infection of astrocytes via the natural intramuscular inoculation route. A combined 3D LSFM-CLSM workflow further identified peripheral Schwann cells as a relevant target cell population of pathogenic RABV in the PNS (publication IV). This suggested that non-abortive infection of central and peripheral neuroglia by pathogenic RABV impairs their immunomodulatory function and thus represents a key step in RABV pathogenesis, which may contribute significantly to the establishment of lethal rabies disease.
Finally, utilizing the full volumetric acquisition power of LSFM, a further refined version of the established 3D imaging pipeline facilitated a detailed mesoscopic investigation of the distribution of SARS-CoV-2 in the respiratory tract of the ferret animal model (publication V). Particularly for this newly-emerged pathogen of global concern, in-depth knowledge of host-pathogen interactions is critical. By preserving the complete spatio-morphological context of virus infection in the ferret respiratory tract, this thesis provided the first specific 3D reconstruction of SARS-CoV-2 infection and the first report of 3D visualization of respiratory virus infection in nasal turbinates altogether. 3D object segmentation of SARS-CoV-2 infection in large tissue volumes identified and emphasized a distinct oligofocal infection pattern in the upper respiratory tract (URT) of ferrets. Furthermore, it corroborated a preferential replication of SARS-CoV-2 in the ferret URT, as only debris-associated virus antigen was detected in the lower respiratory tract of ferrets, thus providing crucial information on the spatial distribution of SARS-CoV-2.
Objective: To compare the effectiveness and complications of intraligamentary anesthesia
(ILA) with conventional inferior alveolar nerve block (IANB) during injection and dental
treatment of mandibular posterior teeth.
Materials and Methods: In this randomized, prospective clinical trial, 72 patients (39 males, 33
females) patients scheduled for dental treatment of mandibular posterior teeth, were randomly
allocated to ILA group (n=35) received ILA injection or IANB group (n=37) received the
conventional IANB. Our primary outcome was to assess pain and stress (discomfort) during the
injection and dental treatment, using the Numeric Rating Scale (NRS) from 0 to 10 (0 = no
pain, 10= the worst pain imaginable). Whereas; recording 24 hours postoperative complications
were our Secondary outcomes.
Results: Patients in ILA group reported significantly less pain during injection when compared
with IANB group (p=0.03). While pain during dental treatment was similar in both groups
(p=0.2). Patients in both groups also reported similar low values of discomfort during treatment
(p= 0.7). Although no signs of nerve contact or any other postoperative complications were
observed, five patients in IANB group (none in ILA group) reported temporary irritations
Conclusion: This study showed equivalent effectiveness of both intraligamentary anesthesia
and conventional inferior alveolar nerve block, for pain control during routine dental treatment
of mandibular posterior teeth. Nevertheless, ILA showed significantly less pain during
injection. No major postoperative complications in both groups were observed.
Clinical Relevance: ILA could be considered as an effective alternative for routine dental
treatment.
Polyethylene terephthalate (PET) is a mass-produced petroleum-based non-biodegradable plastic that contributes to the global plastic pollution. Recently, biocatalytic degradation has emerged as a viable recycling approach for PET waste, especially with thermophilic polyester hydrolases such as a cutinase (LCC) isolated from a leaf-branch compost metagenome and its variants. To improve the enzymatic PET hydrolysis performance, we fused a chitin-binding domain (ChBD) from Chitinolyticbacter meiyuanensis SYBC-H1 to the C-terminus of the previously reported LCCICCG variant, demonstrating higher adsorption to PET substrates and, as a result, improved degradation performance by up to 19.6% compared to with its precursor enzyme without the binding module. For compare hydrolysis with different binding module, the catalytic activity of LCCICCG-ChBD, LCCICCG-CBM, LCCICCG-PBM and LCCICCG-HFB4 were further investigated with PET substrates of various crystallinity and it showed measurable activity on high crystalline PET with 40% crystallinity. These results indicated that fusing a polymer-binding module to LCCICCG is a promising method stimulating the enzymatic hydrolysis of PET.
Sturgeons are among the most ancient linages of actinopterygians. At present, many sturgeon species are critically endangered. Surrogate production could be used as an affordable and a time-efficient method for endangered sturgeons. Our study established a method for identifying and isolating type A spermatogonia from different developmental stages of testes using flow cytometric cell sorting (FCM). Flow cytometric analysis of a whole testicular cell suspension showed several well-distinguished cell populations formed according to different values of light scatter parameters. FCM of these different cell populations was performed directly on glass slides for further immunocytochemistry to identify germ cells. Results showed that the cell population in gate P1 on a flow cytometry plot (with high forward scatter and high side scatter parameter values) contains the highest amount of type A spermatogonia. The sorted cell populations were characterized by expression profiles of 10 germ cell specific genes. The result confirmed that setting up for the P1 gate could precisely sort type A spermatogonia in all tested testicular developmental stages. The P2 gate, which was with lower forward scatter and side scatter values mostly, contained type B spermatogonia at a later maturing stage. Moreover, expressions of plzf, dnd, boule, and kitr were significantly higher in type A spermatogonia than in later developed germ cells. In addition, plzf was firstly found as a reliable marker to identify type A spermatogonia, which filled the gap of identification of spermatogonial stem cells in sterlet. It is expected to increase the efficiency of germ stem cell culture and transplantation with plzf identification. Our study thus first addressed a phenotypic characterization of a pure type A spermatogonia population in sterlet. FCM strategy can improve the production of sturgeons with surrogate broodstock and further the analysis of the cellular and molecular mechanisms of sturgeon germ cell development.
Background: Gastrointestinal hormones (GIHs) are crucial for the regulation of a variety of physiological functions and have been linked to hunger, satiety, and appetite control. Thus, they might constitute meaningful biomarkers in longitudinal and interventional studies on eating behavior and body weight control. However, little is known about the physiological levels of GIHs, their intra-individual stability over time, and their interaction with other metabolic and lifestyle-related parameters. Therefore, the aim of this pilot study is to investigate the intra-individual stability of GIHs in normal-weight adults over time. Methods: Plasma concentrations of ghrelin, leptin, GLP-1 (glucagon-like-peptide), and PP (pancreatic polypeptide) were assessed by enzyme-linked immunosorbent assay (ELISA) in 17 normal-weight, healthy adults in a longitudinal design at baseline and at follow-up six months later. The reliability of the measurements was estimated using intra-class correlation (ICC). In a second step, we considered the stability of GIH levels after controlling for changes in blood glucose and hemoglobin A1 (HbA1c) as well as self-reported physical activity and dietary habits. Results: We found excellent reliability for ghrelin, good reliability for GLP1 and PP, and moderate reliability for leptin. After considering glucose, HbA1c, physical activity, and dietary habits as co-variates, the reliability of ghrelin, GLP1, and PP did not change significantly; the reliability of leptin changed to poor reliability. Conclusions: The GIHs ghrelin, GLP1, and PP demonstrated good to excellent test–retest reliability in healthy individuals, a finding that was not modified after adjusting for glucose control, physical activity, or dietary habits. Leptin showed only moderate to poor reliability, which might be linked to weight fluctuations, albeit small, between baseline and follow-up assessment in our study sample. Together, these findings support that ghrelin, GLP1, and PP might be further examined as biomarkers in studies on weight control, with GLP1 and PP serving as anorexic markers and ghrelin as an orexigenic marker. Additional reliability studies in obese individuals are necessary to verify or refute our findings for this cohort.
Pentathiepins are polysulfur-containing compounds that exert antiproliferative and cytotoxic activity in cancer cells, induce oxidative stress and apoptosis, and inhibit glutathione peroxidase (GPx1). This renders them promising candidates for anticancer drug development. However, the biological effects and how they intertwine have not yet been systematically assessed in diverse cancer cell lines. In this study, six novel pentathiepins were synthesized to suit particular requirements such as fluorescent properties or improved water solubility. Structural elucidation by X-ray crystallography was successful for three derivatives. All six underwent extensive biological evaluation in 14 human cancer cell lines. These studies included investigating the inhibition of GPx1 and cell proliferation, cytotoxicity, and the induction of ROS and DNA strand breaks. Furthermore, selected hallmarks of apoptosis and the impact on cell cycle progression were studied. All six pentathiepins exerted high cytotoxic and antiproliferative activity, while five also strongly inhibited GPx1. There is a clear connection between the potential to provoke oxidative stress and damage to DNA in the form of single- and double-strand breaks. Additionally, these studies support apoptosis but not ferroptosis as the mechanism of cell death in some of the cell lines. As the various pentathiepins give rise to different biological responses, modulation of the biological effects depends on the distinct chemical structures fused to the sulfur ring. This may allow for an optimization of the anticancer activity of pentathiepins in the future.
The genus Capripoxvirus of the family Poxviridae consists of the species lumpy skin disease virus, sheeppox virus and goatpox virus that affect cattle, sheep and goats, respectively. Whereas lumpy skin disease virus (LSDV) is transmitted mainly mechanically via blood-feeding insects and possibly hard ticks, the major transmission routes of sheeppox virus (SPPV) and goatpox virus (GTPV) are via direct contact and aerosols. Affected animals develop fever and display clinical signs such as ocular and nasal discharge, lymphadenopathy and characteristic lesions of the skin. Severe clinical course, especially in combination with respiratory signs, can result in the death of the affected animals. In endemic regions, mortality of capripox virus-induced diseases is low (1-10%). However, mortalities of up to 75% have been reported for LSDV and up to 100% for SPPV and GTPV in exotic breeds and high-producing dairy or beef animals. The loss of quality of the leather, reduced weight gain and milk yield as well as complete loss of affected animals have severe impact on national and global economies. Therefore, capripox virus-induced diseases have significant impact on both the affected individual animal as well as on the existence of small-scale farmers and large agricultural enterprises. However, until now, only live attenuated vaccines are commercially available. These attenuated vaccines are not authorized in the European Union and their administration would comprise the disease-free status of the respective country. Thus, reliable diagnostic tools for the detection and characterization of capripox viruses as well as safe and efficient control measures are of high importance.
The objectives of the present thesis were the development, validation and comparison of diagnostic tools, the establishment of challenge infection models and the performance of pathogenesis studies for all three capripox virus species, and the development and testing of different inactivated prototype vaccine candidates against LSDV.
First, new real-time quantitative polymerase chain reaction (qPCR) assays for robust detection and differentiation of LSDV field strains, LSDV vaccine strains, SPPV and GTPV were developed and extensively validated. In the following, two single assays were combined to duplex assays, one for the differentiation between LSDV field strains and LSDV vaccine strains, and the second for discrimination of SPPV and GTPV. Finally, a diagnostic workflow based on these new duplex assays in combination with already published methods was established. This workflow enables time-saving, robust and reliable detection, species-specific identification and genetic and phylogenetic characterization of all three capripox virus species. In addition, already existing serological examination methods (serum neutralization assay and commercial enzyme-linked immunosorbent assay) were compared regarding their sensitivity and specificity. Furthermore, pathogenesis studies with different capripox virus isolates were performed in the respective target species, and the suitability of selected virus isolates as challenge viruses for future vaccine studies was analyzed. Pathogenesis studies with isolates GTPV-“V/103” and LSDV-“Macedonia2016” revealed that both are proper candidates for challenge models. Finally, three different SPPV isolates (SPPV-“V/104”, SPPV-“India/2013/Surankote” and SPPV-“Egypt/2018”) were tested in sheep regarding their virulence to find a suitable challenge model for SPPV, and SPPV-“India/2013/Surankote” was chosen for future vaccine studies.
Once appropriate challenge models were established, different inactivated prototype vaccines against LSDV were developed, and vaccine safety as well as vaccine efficacy were tested in cattle. Eventually, a Polygen-adjuvanted inactivated LSDV-vaccine candidate was selected that is able to fully prevent cattle from any LSDV-related clinical signs after severe challenge infection. Furthermore, molecular and serological data indicate that this inactivated prototype vaccine is even able to induce a kind of “sterile immunity” against LSDV in those cattle. It has to be mentioned that a commercially available vaccine similar to this prototype vaccine would be a great advance for the control of LSDV.
In the future, additional studies addressing diagnostics and optimized control of capripox viruses should be performed. Firstly, probe-based real-time qPCR assays for the differentiation of SPPV and GTPV vaccine strains from their respective virulent field strains should be developed and included into the diagnostic workflow. Secondly, further tests of the inactivated prototype vaccine, e.g. determination of the minimum protective dose and the possibility of cross-protection in sheep and goats against SPPV and GTPV, respectively, should be performed.
The aims of this study were to 1) determine if continuous eruption occurs in the maxillary teeth, 2) assess the magnitude of the continuous eruption, and 3) evaluate the effects of continuous eruption on the different periodontal parameters by using data from the population-based cohort of the Study of Health in Pomerania (SHIP). The jaw casts of 140 participants from the baseline (SHIP-0) and 16-y follow-up (SHIP-3) were digitized as 3-dimensional models. Robust reference points were set to match the tooth eruption stage at SHIP-0 and SHIP-3. Reference points were set on the occlusal surface of the contralateral premolar and molar teeth, the palatal fossa of an incisor, and the rugae of the hard palate. Reference points were combined to represent 3 virtual occlusal planes. Continuous eruption was measured as the mean height difference between the 3 planes and rugae fix points at SHIP-0 and SHIP-3. Probing depth, clinical attachment levels, gingiva above the cementoenamel junction (gingival height), and number of missing teeth were clinically assessed in the maxilla. Changes in periodontal variables were regressed onto changes in continuous eruption after adjustment for age, sex, number of filled teeth, and education or tooth wear. Continuous tooth eruption >1 mm over the 16 y was found in 4 of 140 adults and averaged to 0.33 mm, equaling 0.021 mm/y. In the total sample, an increase in continuous eruption was significantly associated with decreases in mean gingival height (B = −0.34; 95% CI, −0.65 to −0.03). In a subsample of participants without tooth loss, continuous eruption was negatively associated with PD. This study confirmed that continuous eruption is clearly detectable and may contribute to lower gingival heights in the maxilla.
Drainage has commonly been a pre-requisite for the productive use of peatlands. The biased focus on agriculture, forestry and peat extraction has long ignored the destructive effects of drainage and the successive degradation of ecosystem functions of wet peatlands. Accelerated by the climate crisis, the finite nature of drainage-based peatland use is increasingly recognised. Consequently, productive land use options for wet or rewetted peatlands (paludiculture) are required as sustainable alternatives. A wide range of paludiculture plants and options of biomass utilisation are identified as suitable and promising. Despite the growing interest, experiences with and research on the economic viability of paludiculture are still rare.
This thesis addresses the lack of knowledge on paludiculture in terms of practical feasibility, costs and benefits at the farm level, market prospects and framework conditions. I selected the two currently most advanced paludicultural practices in Europe: a) Harvesting natural reed beds as a traditional ‘low-input’ paludiculture, i. e. the utilisation of existing ‘wild’ vegetation stands; b) ‘Sphagnum farming’ as a novel ‘high-input’ paludiculture including stand establishment and water management required for the active transformation from drainage-based peatland use to paludiculture. In both cases, I investigate three different biomass utilisation avenues. This thesis adds to the fields of problem-driven sustainability and land-use science. Procedures and costs of paludiculture were studied in transdisciplinary research projects in close cooperation with practitioners. Due to the novelty of the topic, I put special emphasis on the triangulation of methods and data sources: pilot trials, field measurements, semi-structured expert interviews, structured questionnaires, secondary data from trade statistics and literature. To account for uncertainty related to costs and revenues, I conduct stochastic scenario analysis (Monte Carlo simulation) for the extended contribution margin accounting of harvesting reeds and sensitivity analysis for the investment appraisal of Sphagnum farming.
Paludiculture on fens: harvesting reeds
Paper I investigates harvesting procedures for reed-dominated (Phragmites australis) vegetation stands. In many European countries special-purpose tracked machinery is applied for large-scale conservation management and the commercial harvest of thatching reed. Stochastic scenario analysis reveals a wide range of possible economic outcomes (ca. € -1000 to € 1500 ha-1 a-1) and identifies material use of reed superior to its use as a source of energy. Winter harvest of high-quality thatching reed in bundles is the most profitable option. Winter harvest of bales for direct combustion is suitable for low-quality stands and has a limited risk of loss. In the case of summer harvest, revenues for green chaff for biogas production cannot cover harvesting costs but non-market income via subsidies and agri-environmental payments may ensure profitability. While biomass for energy generation is limited to a local market, thatching reed is traded as an international commodity. The market situation for thatching reed is investigated for Europe (Paper II) and Germany (Paper III). The major reed consuming countries in Western Europe (Netherlands, Germany, UK, Denmark) rely on imports of up to 85 % of the national consumption, with reed being imported from Eastern and Southern Europe and since 2005 also from China. The total market volume for reed for thatching in Northern Germany is estimated with 3 ± 0.8 million bundles of reed with a monetary value at sales prices of € 11.6 ± 2.8 million. Most of the thatchers (70 %) did not promote reed of regional origin to their customers due to insufficient availability in the first place and a lack in quality as second reason. The cultivation of reed in paludiculture may improve quantity and quality of domestic thatching reed. An area of 6000 ± 1600 ha with an average yield of 500 bundles per hectare would allow covering the current total demand of 3 million bundles of the German thatching reed market (Paper III).
Paludiculture on bogs: Sphagnum farming
Sphagnum farming provides an alternative to peatland degradation in two ways: Firstly, Sphagnum mosses can be cultivated as new agricultural crops on rewetted peatlands. Secondly, the produced Sphagnum biomass is a high-quality raw material suitable to replace peat in horticultural growing media (Paper V). Pilot trials have demonstrated the practical feasibility of establishing Sphagnum cultures on former bog grassland, cut-over bogs and mats floating on acidic waters bodies; Paper IV compares for the three types of production sites the specific procedures, costs and area potential in Germany. Water-based Sphagnum farming is not recommended for large-scale implementation due to highest establishment costs, major cultivation risks and limited area potential. For soil-based Sphagnum farming, the most important cost positions were Sphagnum shoots to set up pilots, investment for water management and regular weed management. Bog grassland has the highest area potential, i. e. 90,000 ha in NW Germany. Paper V assesses the profitability of Sphagnum farming on former bog grassland based on extrapolating five years of field experience data (establishment ņ management ņ harvest) to a total cultivation time of twenty years. Cultivating Sphagnum biomass as founder material for Sphagnum farming or restoration was profitable even in pessimistic scenarios with high costs, high bulk density and low yields. Selling Sphagnum for orchid production was economically viable in the case of medium to high yields with a low bulk density. Cost-covering prices for Sphagnum biomass substituting peat seem achievable if end consumers pay a surcharge of 10 % on the peat-free cultivated horticultural end-product. An area of 35,000 ha of Sphagnum farming suffices to meet the annual demand of the German growing media industry for slightly decomposed Sphagnum peat.
Framework conditions affecting feasibility of paludiculture
The relation of revenues from selling biomass to its production costs is an important piece of the paludiculture feasibility puzzle. Further aspects effecting the economic viability and competitiveness of paludiculture encompass the market demand, the availability of mature technology, legal restrictions, the eligibility for agricultural subsidies, a remuneration of external benefits and the opportunity costs of present farming activities (Paper I, V). Legal and policy regulations are of major importance for land use decisions on peatlands – both for keeping up drainage and for shifting to paludiculture.
Conclusion and Outlook
This thesis provides a first assessment of the costs and profitability of large-scale harvesting of reeds and Sphagnum farming based on real-life data. The paludicultural practices investigated may be a solution for a minor share of the more than 1 million ha of peatlands drained for agriculture in Germany. Future research should also address other biomass utilisation options and other crops. Large-scale pilots are required to improve technical maturity of procedures and machinery, gather reliable data to replace assumptions on costs and revenues and study long-term effects on economics and ecosystem services. The micro-economic perspective needs to be complemented by the societal perspective quantifying and monetising external effects of peatland restoration, paludiculture and drainage-based peatland use. There is a high need for intensified research, large-scale implementation and accelerated adaption of the policy and legal framework to develop paludiculture as an economically viable option for degraded peatlands.
The ubiquitin-proteasome system (UPS) and the autophagy-lysosomal pathway (ALP) are the main proteolytic systems involved in cellular homeostasis. Since cardiomyocytes, as terminally differentiated cells, lack the ability to share damaged proteins with their daughter cells, they are especially reliant on these protein degradation systems for their proper function. Alterations of the UPS and ALP have been reported in a wide range of cardiac diseases, including cardiomyopathies. In this study, we determined whether the UPS and ALP are altered in a mouse model of eccentric left ventricular (LV) hypertrophy expressing both cyclin T1 and Gαq under the control of the cardiac-specific α-myosin heavy chain promoter (double transgenic; DTG). Compared to wild-type (WT) littermates, DTG mice showed higher end-diastolic (ED) LV wall thicknesses and diameter with preserved ejection fraction (EF). The cardiomyopathic phenotype was further confirmed by an upregulation of the fetal gene program and genes associated with fibrosis as well as a downregulation of genes involved in Ca2+ handling. Likewise, higher NT-proBNP levels were detected in DTG mice. Investigation of the UPS showed elevated steady-state levels of (poly)ubiquitinated proteins without alterations of all proteasomal activities in DTG mice. Evaluation of ALP key marker revealed a mixed pattern with higher protein levels of microtubule-associated protein 1 light chain 3 beta (LC3)-I and lysosomal-associated membrane protein-2, lower protein levels of beclin-1 and FYVE and coiled-coil domain-containing protein 1 (FYCO1) and unchanged protein levels of p62/SQSTM1 in DTG mice when compared to WT. At transcriptional level, a > 1.2-fold expression was observed for Erbb2, Hdac6, Lamp2, Nrg1, and Sqstm1, while a < 0.8-fold expression was revealed for Fyco1 in DTG mice. The results related to the ALP suggested overall a repression of the ALP during the initiation process, but an induction of the ALP at the level of autophagosome-lysosome fusion and the delivery of ubiquitinated cargo to the ALP for degradation.
Intestinal transporter proteins are known to affect the pharmacokinetics and in turn the efficacy and safety of many orally administered drugs in a clinically relevant manner. This knowledge is especially well-established for intestinal ATP-binding cassette transporters such as P-gp and BCRP. In contrast to this, information about intestinal uptake carriers is much more limited although many hydrophilic or ionic drugs are not expected to undergo passive diffusion but probably require specific uptake transporters. A transporter which is controversially discussed with respect to its expression, localization and function in the human intestine is the organic cation transporter 1 (OCT1). This review article provides an up-to-date summary on the available data from expression analysis as well as functional studies in vitro, animal findings and clinical observations. The current evidence suggests that OCT1 is expressed in the human intestine in small amounts (on gene and protein levels), while its cellular localization in the apical or basolateral membrane of the enterocytes remains to be finally defined, but functional data point to a secretory function of the transporter at the basolateral membrane. Thus, OCT1 should not be considered as a classical uptake transporter in the intestine but rather as an intestinal elimination pathway for cationic compounds from the systemic circulation.
Objectives: Clear guidelines on when to remove an implant are missing. This study aimed to evaluate the amount of peri-implant bone loss at explantation by specialists.
Material and Methods: Implantology specialists were asked to provide implants explanted due to peri-implantitis with related clinical information. Questionnaires inquired age, sex, smoking habit, implant location, usage of bone substitutes, and implant brand. Early failures (survival time <12 months) were analysed separately. Explants were measured and bone loss and type of bone loss were assessed using radiographs. Bivariate analysis was used for the type of bone loss, and covariate-adjusted mixed-effects models were evaluated for the amount of bone loss and survival time.
Results: Twelve dental offices provided 192 explants from 161 patients with 99 related radiographs. Most implants were affected by vertical bone loss (51.1%), followed by combined horizontal and vertical bone loss (22.3%), peri-implant gap (11.7%), horizontal bone loss (10.6%), and only a few by apical inflammation (4.3%). Thirty-three (17.2%) explants were early failures. Type of bone loss was significantly associated with survival time and implant brand. Implant brand also showed a significant correlation with early/late implant failure. Excluding early failures, combined horizontal and vertical bone loss was additionally significantly associated with smoking, and the location when grouped to incisor, canine, premolar, and molar showed a significant association with the type of bone loss. Further, the average survival time was 9.5 ± 5.8 years with absolute and relative bone loss of 7.0 ± 2.7 mm and 66.2 ± 23.7%, respectively. Late failures were removed at a mean bone loss of 50.0% with 5.44 mm residual alveolar bone in the posterior maxilla and 73.8% with 2.89 mm residual alveolar bone in other locations. In fully adjusted mixed-effects models, only the age at implantation (B=-0.19; 95% CI: -0.27 to -0.10) remained a significant factor for survival time. Implants exhibited significantly more relative bone loss if they were positioned in the mandible (B=17.3; 95% CI: 3.91 to 30.72) or if they were shorter (B=-2.79; 95% CI: -5.50 to -0.08).
Conclusions: Though the mean bone loss (66.2%) at which implants were explanted was in accordance with the literature, its wide variation and differentiation between the posterior maxilla and other locations showed that the profession has no universally accepted threshold beyond which an implant cannot be preserved.
Ebolaviruses are dependent on host cell proteins for almost all steps in their viral life cycle. While some cellular factors with crucial roles in the ebolavirus life cycle have been identified, many of them remain to be identified or fully characterised. This thesis focuses on the characterisation and identification of host cell interactions of the highly pathogenic Ebola virus (EBOV), probing host-virus interaction at various stages of the viral life cycle. Beginning with viral budding, the function of a recently proposed late domain motif within the EBOV matrix protein VP40 was examined using an EBOV transcription and replication-competent virus-like particle (trVLP) system. Although this motif has been suggested to interact with the endosomal sorting complex required for transport (ESCRT), we could show that this late domain motif does not contribute to EBOV budding.
While many host cell proteins have been identified so far that are important for viral budding, only a few proteins are known that are necessary for EBOV RNA synthesis. Thus, to identify host proteins that are involved in viral replication and transcription, we performed a genome-wide siRNA screen in the context of an EBOV minigenome assay. Using this approach, we identified several proteins that appear to be important for viral RNA synthesis or protein expression. Two of the most prominent hits in our screen were CAD (Carbamoyl-phosphate synthetase 2, aspartate transcarbamylase and dihydroorotase) and NXF1 (nuclear RNA export factor 1). CAD catalyses the first three steps in the de novo pyrimidine biosynthesis, while NXF1 is the main nuclear export protein for cellular mRNAs. In subsequent characterisation studies, using a range of life cycle modelling systems as well as molecular analyses, we could demonstrate that the canonical function of CAD during the pyrimidine biosynthesis is necessary for EBOV replication and transcription. In contrast to this, for NXF1 we discovered a so-far unknown function: Again, by applying different life cycle modelling alongside with molecular assays, we provided evidence that the EBOV nucleoprotein recruits NXF1 into inclusion bodies, the site of EBOV RNA synthesis, where it binds viral mRNAs to export them from these structures. Importantly, for both CAD and NXF1 we were able to recapitulate key data in the context of live EBOV infection, confirming their roles in the viral life cycle.
Both of these identified host factors are promising targets for antiviral therapies and indeed de novo pyrimidine synthesis is emerging as a possible antiviral target for a number of viruses. Similarly, as we could show NXF1 to be important in the life cycle of the highly pathogenic Junín virus, this raises the possibility that disruption of this interaction may result in broad-spectrum antiviral activity. Moreover, for an increasing number of negative-sense RNA viruses inclusion bodies as site of viral RNA synthesis are described to have a liquid organelle character. Therefore, our findings on NXF1 also provide an intriguing model to explain how negative-sense RNA viruses in general overcome this obstacle and export viral mRNAs from inclusion bodies.
The full genome of a Methanomassiliicoccales strain, U3.2.1, was obtained from enrichment cultures of percolation fen peat soil under methanogenic conditions, with methanol and hydrogen as the electron acceptor and donor, respectively. Metagenomic assembly of combined long-read and short-read sequences resulted in a 1.51-Mbp circular genome.
The objectives of this study were to ascertain the fecal ESBL/AmpC-E. coli prevalence and to detect risk factors for their occurrence in young pre-weaned calves and their dams on large dairy farms in Germany. From 2018–2019 we investigated 2816 individual fecal samples from pre-weaned dairy calves and their dams, representing seventy-two farms (mean = 667 milking cows) from eight German federal states. To assess possible risk factors associated with ESBL/AmpC-E. coli prevalence in calves and dams, a questionnaire was performed, collecting management data. We observed an ESBL/AmpC-E. coli prevalence of 63.5% (95% CI: 57.4–69.5) among the sampled calves and 18.0% (95% CI: 12.5–23.5) among the dams. On all farms, at least one positive sample was obtained. To date, this is the highest ESBL/AmpC-E. coli prevalence observed in dairy herds in Europe. Feeding with waste milk was identified as a significant risk factor for a high prevalence of ESBL/AmpC-E. coli in calves. Many calves at large dairies in Germany are fed with waste milk due to the large amounts generated as a result of antibiotic dry-off routines and mastitis treatment with antibiotics. Other notable risk factors for high ESBL/AmpC-E. coli in calves were the general fitness/health of dams and calves, and the quality of farm hygiene. Taken together, these findings suggest that new or improved approaches to animal health management, for example, antibiotic dry cow management (selective dry cow therapy) and mastitis treatment (high self-recovery), as well as farm hygiene, should be researched and implemented.
Chagas’ disease (CD), caused by the hemoflagellate protozoan, Trypanosoma cruzi, is endemic in most countries of Latin America. Heart failure (HF) is often a late manifestation of chronic CD, and is associated with high morbidity and mortality. Inflammatory processes mediated by cytokines play a key role in the pathogenesis and progression of CD. Keeping in view the inflammatory nature of CD, this study investigated the possible role of 21 different inflammatory cytokines as biomarkers for prediction and prognosis of CD. The plasma concentration of these cytokines was measured in a group of patients with CD (n = 94), and then compared with those measured in patients with dilated cardiomyopathy (DCM) from idiopathic causes (n = 48), and with control subjects (n = 25). Monovariately, plasma levels of cytokines such as stem cell growth factor beta (SCGF beta), hepatocyte growth factor (HGF), monokine induced by interferon gamma (CXCL9), and macrophage inhibitory factor (MIF) were significantly increased in CD patients with advanced HF compared to control group. None of the cytokines could demonstrate any prognostic potency in CD patients, and only MIF and stromal derived factor-1 alpha (CXCL12) showed significance in predicting mortality and necessity for heart transplant in DCM patients. However, multivariate analysis prognosticated a large proportion of CD and DCM patients. In CD patients, HGF and Interleukin-12p40 (IL-12p40) together separated 81.9% of 3-year survivors from the deceased, while in DCM patients, CXCL12, stem cell factor (SCF), and CXCL9 together discriminated 77.1% of survivors from the deceased. The significant increase in plasma concentrations of cytokines such as HGF and CXCL9 in CD patients, and the ability of these cytokines to prognosticate a large proportion of CD and DCM patients multivariately, encourages further studies to clarify the diagnostic and prognostic potential of cytokines in such patients.
Aging is an independent risk factor for hypertension, cardiovascular morbidity, and mortality. However, detailed mechanisms linking aging to cardiovascular disease are unclear. We studied the aging effects on the role of perivascular adipose tissue and downstream vasoconstriction targets, voltage-dependent KV7 channels, and their pharmacological modulators (flupirtine, retigabine, QO58, and QO58-lysine) in a murine model. We assessed vascular function of young and old mesenteric arteries in vitro using wire myography and membrane potential measurements with sharp electrodes. We also performed bulk RNA sequencing and quantitative reverse transcription-polymerase chain reaction tests in mesenteric arteries and perivascular adipose tissue to elucidate molecular underpinnings of age-related phenotypes. Results revealed impaired perivascular adipose tissue-mediated control of vascular tone particularly via KV7.3–5 channels with increased age through metabolic and inflammatory processes and release of perivascular adipose tissue-derived relaxation factors. Moreover, QO58 was identified as novel pharmacological vasodilator to activate XE991-sensitive KCNQ channels in old mesenteric arteries. Our data suggest that targeting inflammation and metabolism in perivascular adipose tissue could represent novel approaches to restore vascular function during aging. Furthermore, KV7.3–5 channels represent a promising target in cardiovascular aging.
Terrestrial surface waters and submarine ground water discharge (SGD) act as a source of dissolved substances for coastal systems. Solute fluxes of SGD depend on the ground water composition and the water-solid-microbe interactions close to the sediment-water interface. Thus, this study aims to characterize and evaluate the hydrogeochemical gradients developing in the fresh-salt water mixing zone of the Wismar Bay (WB), southern Baltic Sea, Germany. Sampling campaigns covering the WB, the fresh-salt water mixing zone at the beach of the WB shoreline, terrestrial surface and ground waters near the WB as well sediments pore water were carried out. In these different waters, the distribution of dissolved inorganic carbon, nutrients, major ions, trace elements, stable isotopes (H, O, C, S), and radium isotopes have been investigated. Enhanced concentrations of radium isotopes together with dissolved manganese, barium in the surface waters of the eastern WB indicated benthic-pelagic coupling via the exchange between pore water and the water column. Salinity, stable isotopes, and major ions in sediment pore water profiles identified the presence of fresh ground water below about 40 cmbsf in the central part of the bay. Geophysical acoustic techniques revealed the local impact of anthropogenic sediment excavation, which reduced the thickness of a sediment layer between the coastal aquifer and the bottom water, causing, therefore, a ground water upward flow close to the top sediments. The fresh impacted pore water stable isotope composition (δ18O, δ2H) plot close to the regional meteoric water line indicating a relatively modern ground water source. The calculated organic matter mineralization rates and the dissolved inorganic carbon sediment-water fluxes were much higher at the fresh impacted site when compared to other unimpacted sediments. Therefore, this study reveals that different fresh water sources contribute to the water balance of WB including a SGD source.
Analysis of volatile organic compounds (VOCs) is a novel approach to accelerate bacterial culture diagnostics of Mycobacterium avium subsp. paratuberculosis (MAP). In the present study, cultures of fecal and tissue samples from MAP-infected and non-suspect dairy cattle and goats were explored to elucidate the effects of sample matrix and of animal species on VOC emissions during bacterial cultivation and to identify early markers for bacterial growth. The samples were processed following standard laboratory procedures, culture tubes were incubated for different time periods. Headspace volume of the tubes was sampled by needle trap-micro-extraction, and analyzed by gas chromatography-mass spectrometry. Analysis of MAP-specific VOC emissions considered potential characteristic VOC patterns. To address variation of the patterns, a flexible and robust machine learning workflow was set up, based on random forest classifiers, and comprising three steps: variable selection, parameter optimization, and classification. Only a few substances originated either from a certain matrix or could be assigned to one animal species. These additional emissions were not considered informative by the variable selection procedure. Classification accuracy of MAP-positive and negative cultures of bovine feces was 0.98 and of caprine feces 0.88, respectively. Six compounds indicating MAP presence were selected in all four settings (cattle vs. goat, feces vs. tissue): 2-Methyl-1-propanol, 2-methyl-1-butanol, 3-methyl-1-butanol, heptanal, isoprene, and 2-heptanone. Classification accuracies for MAP growth-scores ranged from 0.82 for goat tissue to 0.89 for cattle feces. Misclassification occurred predominantly between related scores. Seventeen compounds indicating MAP growth were selected in all four settings, including the 6 compounds indicating MAP presence. The concentration levels of 2,3,5-trimethylfuran, 2-pentylfuran, 1-propanol, and 1-hexanol were indicative for MAP cultures before visible growth was apparent. Thus, very accurate classification of the VOC samples was achieved and the potential of VOC analysis to detect bacterial growth before colonies become visible was confirmed. These results indicate that diagnosis of paratuberculosis can be optimized by monitoring VOC emissions of bacterial cultures. Further validation studies are needed to increase the robustness of indicative VOC patterns for early MAP growth as a pre-requisite for the development of VOC-based diagnostic analysis systems.
Quality of Life in Young Adults With Cerebral Palsy: A Longitudinal Analysis of the SPARCLE Study
(2021)
Introduction: While most people with cerebral palsy (CP) will have a life expectancy similar to that of the general population, international research has primarily focused on childhood and adolescence; and knowledge about the quality of life (QoL) of young adults with CP, its trajectories, and associated factors remains scarce.
Methods: This longitudinal study included young adults with CP living in five European regions and who had previously participated in the SPARCLE cohort as children and/or adolescents. Their QoL in the psychological well-being and social relationships domains was estimated using age-appropriate validated instruments (KIDSCREEN-52 in childhood/adolescence and WHOQOL-Bref in young adulthood). We used generalized linear mixed-effect models with random intercept to estimate long-term trajectories of QoL in both domains and to investigate whether severity of impairment, pain, and seizure influenced these trajectories. We sought to identify potentially different trajectories of QoL from childhood to adulthood using a shape-based clustering method.
Results: In total, 164 young adults with CP aged 22–27 years participated in the study. Psychological well-being linearly decreased by 0.78 points (scale 0–100) per year (95% confidence interval (CI) −0.99 to −0.56) from childhood to young adulthood, whereas QoL in the social relationships domain increased (β coefficient 1.24, 95% CI 0.92–1.55). Severity of impairment was associated with reduced QoL in all life periods of the study (childhood, adolescence, and young adulthood): motor impairment with social relationships, and more nuancedly intellectual disability with psychological well-being and social relationships. At all periods, frequent pain significantly reduced psychological well-being, and seizures were associated with lower QoL in the social relationships domain. In both domains, we identified a group of individuals with CP who presented a reverse trajectory compared with the general QoL trajectory.
Conclusion: Identification of QoL trajectories and their associated factors yields improved knowledge about the experience of individuals with CP until young adulthood. Further studies are needed to better understand the determinants that have the greatest influence on the different shapes of long-term trajectories of QoL.
Abstract
White spot disease (WSD) is one of the most devastating viral infections of crustaceans caused by the white spot syndrome virus (WSSV). A conserved sequence WSSV131 in the DNA genome of WSSV was found to fold into a polymorphic G‐quadruplex structure. Supported by two mutant sequences with single G→T substitutions in the third G4 tract of WSSV131, circular dichroism and NMR spectroscopic analyses demonstrate folding of the wild‐type sequence into a three‐tetrad parallel topology comprising three propeller loops with a major 1 : 3 : 1 and a minor 1 : 2 : 2 loop length arrangement. A thermodynamic analysis of quadruplex formation by differential scanning calorimetry (DSC) indicates a thermodynamically more stable 1 : 3 : 1 loop isomer. DSC also revealed the formation of additional highly stable multimeric species with populations depending on potassium ion concentration.
The thyroid gland is both a thyroid hormone (TH) generating as well as a TH responsive
organ. It is hence crucial that cathepsin-mediated proteolytic cleavage of the precursor thyroglobulin
is regulated and integrated with the subsequent export of TH into the blood circulation, which is
enabled by TH transporters such as monocarboxylate transporters Mct8 and Mct10. Previously, we
showed that cathepsin K-deficient mice exhibit the phenomenon of functional compensation through
cathepsin L upregulation, which is independent of the canonical hypothalamus-pituitary-thyroid axis,
thus, due to auto-regulation. Since these animals also feature enhanced Mct8 expression, we aimed
to understand if TH transporters are part of the thyroid auto-regulatory mechanisms. Therefore,
we analyzed phenotypic differences in thyroid function arising from combined cathepsin K and
TH transporter deficiencies, i.e., in Ctsk-/-/Mct10-/-
, Ctsk-/-/Mct8-/y, and Ctsk-/-/Mct8-/y/Mct10-/-
.
Despite the impaired TH export, thyroglobulin degradation was enhanced in the mice lacking Mct8,
particularly in the triple-deficient genotype, due to increased cathepsin amounts and enhanced cysteine peptidase activities, leading to ongoing thyroglobulin proteolysis for TH liberation, eventually
causing self-thyrotoxic thyroid states. The increased cathepsin amounts were a consequence of
autophagy-mediated lysosomal biogenesis that is possibly triggered due to the stress accompanying
intrathyroidal TH accumulation, in particular in the Ctsk-/-/Mct8-/y/Mct10-/- animals. Collectively,
our data points to the notion that the absence of cathepsin K and Mct8 leads to excessive thyroglobulin
degradation and TH liberation in a non-classical pathway of thyroid auto-regulation.
Universal products provide an axiomatic framework to study noncommutative independences general enough to include, besides the well known "single-faced" case (i.e., tensor, free, Boolean, monotone and antimonotone independence), also more recent "multi-faced" examples like bifree independence. Questions concerning classification have been fully answered in the single-faced case, but are in general still open in the multi-faced case. In this thesis we discuss how one can use insights in the relation between universal products and their associated moment-cumulant formula as a starting point towards a combinatorial approach to (multi-faced) universal products. We define certain classes of partitions and discuss why the defining axioms are sufficient to associate to each of them a multi-faced universal product. For the two-faced case we present our result that every positive and symmetric universal product can be produced in this fashion and we outline how these results might contribute to a classification of positive and symmetric universal products.
Permafrost-affected soil stores a significant amount of organic carbon. Identifying the biological constraints of soil organic matter transformation, e.g., the interaction of major soil microbial soil organic matter decomposers, is crucial for predicting carbon vulnerability in permafrost-affected soil. Fungi are important players in the decomposition of soil organic matter and often interact in various mutualistic relationships during this process. We investigated four different soil horizon types (including specific horizons of cryoturbated soil organic matter (cryoOM)) across different types of permafrost-affected soil in the Western Canadian Arctic, determined the composition of fungal communities by sequencing (Illumina MPS) the fungal internal transcribed spacer region, assigned fungal lifestyles, and by determining the co-occurrence of fungal network properties, identified the topological role of keystone fungal taxa. Compositional analysis revealed a significantly higher relative proportion of the litter saprotroph Lachnum and root-associated saprotroph Phialocephala in the topsoil and the ectomycorrhizal close-contact exploring Russula in cryoOM, whereas Sites 1 and 2 had a significantly higher mean proportion of plant pathogens and lichenized trophic modes. Co-occurrence network analysis revealed the lowest modularity and average path length, and highest clustering coefficient in cryoOM, which suggested a lower network resistance to environmental perturbation. Zi-Pi plot analysis suggested that some keystone taxa changed their role from generalist to specialist, depending on the specific horizon concerned, Cladophialophora in topsoil, saprotrophic Mortierella in cryoOM, and Penicillium in subsoil were classified as generalists for the respective horizons but specialists elsewhere. The litter saprotrophic taxon Cadophora finlandica played a role as a generalist in Site 1 and specialist in the rest of the sites. Overall, these results suggested that fungal communities within cryoOM were more susceptible to environmental change and some taxa may shift their role, which may lead to changes in carbon storage in permafrost-affected soil.
Rice husk, one of the main side products in the rice production, and its sustainable management represent a challenge in many countries. Herein, we describe the use of this abundant agricultural bio‐waste as feedstock for the preparation of silver‐containing carbon/silica nano composites with antimicrobial properties. The synthesis was performed using a fast and cheap methodology consisting of wet impregnation followed by pyrolysis, yielding C/SiO2 composite materials doped with varying amounts of silver from 28 to 0.001 wt %. The materials were fully characterized and their antimicrobial activity against ESKAPE pathogens, namely E. faecium, S. aureus, K. pneumoniae, A. baumannii, P. aeruginosa, and E. coli, and the pathogenic yeast C. albicans was investigated. Sensitivities of these strains against the prepared materials were demonstrated, even with exceptional low amounts of 0.015 m% silver. Hence, we report a straightforward method for the synthesis of antimicrobial agents from abundant sources which addresses urgent questions like bio‐waste valorization and affordable alternatives to increasingly fewer effective antibiotics.
Abstract
Aerated topsoils are important sinks for atmospheric methane (CH4) via oxidation by CH4‐oxidizing bacteria (MOB). However, intensified management of grasslands and forests may reduce the CH4 sink capacity of soils. We investigated the influence of grassland land‐use intensity (150 sites) and forest management type (149 sites) on potential atmospheric CH4 oxidation rates (PMORs) and the abundance and diversity of MOB (with qPCR) in topsoils of three temperate regions in Germany. PMORs measurements in microcosms under defined conditions yielded approximately twice as much CH4 oxidation in forest than in grassland soils. High land‐use intensity of grasslands had a negative effect on PMORs (−40%) in almost all regions and fertilization was the predominant factor of grassland land‐use intensity leading to PMOR reduction by 20%. In contrast, forest management did not affect PMORs in forest soils. Upland soil cluster (USC)‐α was the dominant group of MOBs in the forests. In contrast, USC‐γ was absent in more than half of the forest soils but present in almost all grassland soils. USC‐α abundance had a direct positive effect on PMOR in forest, while in grasslands USC‐α and USC‐γ abundance affected PMOR positively with a more pronounced contribution of USC‐γ than USC‐α. Soil bulk density negatively influenced PMOR in both forests and grasslands. We further found that the response of the PMORs to pH, soil texture, soil water holding capacity and organic carbon and nitrogen content differ between temperate forest and grassland soils. pH had no direct effects on PMOR, but indirect ones via the MOB abundances, showing a negative effect on USC‐α, and a positive on USC‐γ abundance. We conclude that reduction in grassland land‐use intensity and afforestation has the potential to increase the CH4 sink function of soils and that different parameters determine the microbial methane sink in forest and grassland soils.
Extracellular vesicles (EVs) are reminiscent of their cell of origin and thus represent a
valuable source of biomarkers. However, for EVs to be used as biomarkers in clinical practice, simple,
comparable, and reproducible analytical methods must be applied. Although progress is being
made in EV separation methods for human biofluids, the implementation of EV assays for clinical
diagnosis and common guidelines are still lacking. We conducted a comprehensive analysis of
established EV separation techniques from human serum and plasma, including ultracentrifugation
and size exclusion chromatography (SEC), followed by concentration using (a) ultracentrifugation,
(b) ultrafiltration, or (c) precipitation, and immunoaffinity isolation. We analyzed the size, number,
protein, and miRNA content of the obtained EVs and assessed the functional delivery of EV cargo.
Our results demonstrate that all methods led to an adequate yield of small EVs. While no significant
difference in miRNA content was observed for the different separation methods, ultracentrifugation
was best for subsequent flow cytometry analysis. Immunoaffinity isolation is not suitable for
subsequent protein analyses. SEC + ultracentrifugation showed the best functional delivery of
EV cargo. In summary, combining SEC with ultracentrifugation gives the highest yield of pure
and functional EVs and allows reliable analysis of both protein and miRNA contents. We propose
this combination as the preferred EV isolation method for biomarker studies from human serum
or plasma.
Significant alterations of cambial activity might be expected due to climate warming, leading to growing season extension and higher growth rates especially in cold-limited forests. However, assessment of climate-change-driven trends in intra-annual wood formation suffers from the lack of direct observations with a timespan exceeding a few years. We used the Vaganov-Shashkin process-based model to: (i) simulate daily resolved numbers of cambial and differentiating cells; and (ii) develop chronologies of the onset and termination of specific phases of cambial phenology during 1961–2017. We also determined the dominant climatic factor limiting cambial activity for each day. To asses intra-annual model validity, we used 8 years of direct xylogenesis monitoring from the treeline region of the Krkonoše Mts. (Czechia). The model exhibits high validity in case of spring phenological phases and a seasonal dynamics of tracheid production, but its precision declines for estimates of autumn phenological phases and growing season duration. The simulations reveal an increasing trend in the number of tracheids produced by cambium each year by 0.42 cells/year. Spring phenological phases (onset of cambial cell growth and tracheid enlargement) show significant shifts toward earlier occurrence in the year (for 0.28–0.34 days/year). In addition, there is a significant increase in simulated growth rates during entire growing season associated with the intra-annual redistribution of the dominant climatic controls over cambial activity. Results suggest that higher growth rates at treeline are driven by (i) temperature-stimulated intensification of spring cambial kinetics, and (ii) decoupling of summer growth rates from the limiting effect of low summer temperature due to higher frequency of climatically optimal days. Our results highlight that the cambial kinetics stimulation by increasing spring and summer temperatures and shifting spring phenology determine the recent growth trends of treeline ecosystems. Redistribution of individual climatic factors controlling cambial activity during the growing season questions the temporal stability of climatic signal of cold forest chronologies under ongoing climate change.
The human pathogen Clostridioides difficile has evolved into the leading cause of nosocomial diarrhea. The bacterium is capable of spore formation, which even allows survival of antibiotic treatment. Although C. difficile features an anaerobic lifestyle, we determined a remarkably high oxygen tolerance of the laboratory reference strain 630Δerm. A mutation of a single nucleotide (single nucleotide polymorphism [SNP]) in the DNA sequence (A to G) of the gene encoding the regulatory protein PerR results in an amino acid substitution (Thr to Ala) in one of the helices of the helix-turn-helix DNA binding domain of this transcriptional repressor in C. difficile 630Δerm. PerR is a sensor protein for hydrogen peroxide and controls the expression of genes involved in the oxidative stress response. We show that PerR of C. difficile 630Δerm has lost its ability to bind the promoter region of PerR-controlled genes. This results in a constitutive derepression of genes encoding oxidative stress proteins such as a rubrerythrin (rbr1) whose mRNA abundance under anaerobic conditions was increased by a factor of about 7 compared to its parental strain C. difficile 630. Rubrerythrin repression in strain 630Δerm could be restored by the introduction of PerR from strain 630. The permanent oxidative stress response of C. difficile 630Δerm observed here should be considered in physiological and pathophysiological investigations based on this widely used model strain.
IMPORTANCE The intestinal pathogen Clostridioides difficile is one of the major challenges in medical facilities nowadays. In order to better combat the bacterium, detailed knowledge of its physiology is mandatory. C. difficile strain 630Δerm was generated in a laboratory from the patient-isolated strain C. difficile 630 and represents a reference strain for many researchers in the field, serving as the basis for the construction of insertional gene knockout mutants. In our work, we demonstrate that this strain is characterized by an uncontrolled oxidative stress response as a result of a single-base-pair substitution in the sequence of a transcriptional regulator. C. difficile researchers working with model strain 630Δerm should be aware of this permanent stress response.
Duckweeds comprise a distinctive clade of pleustophytic monocots that traditionally has been classified as the family Lemnaceae. However, molecular evidence has called into question their phylogenetic independence, with some authors asserting instead that duckweeds should be reclassified as subfamily Lemnoideae of an expanded family Araceae. Although a close phylogenetic relationship of duckweeds with traditional Araceae has been supported by multiple studies, the taxonomic disposition of duckweeds must be evaluated more critically to promote nomenclatural stability and utility. Subsuming duckweeds as a morphologically incongruent lineage of Araceae effectively eliminates the family category of Lemnaceae that has been widely used for many years. Instead, we suggest that Araceae subfamily Orontioideae should be restored to family status as Orontiaceae, which thereby would enable the recognition of three morphologically and phylogenetically distinct lineages: Araceae, Lemnaceae, and Orontiaceae.
Urm1: A Non-Canonical UBL
(2021)