Two-dimensional speckle tracking echocardiography in fetuses with critical aortic stenosis before and after fetal aortic valvuloplasty.

BACKGROUND: Critical aortic stenosis (AS) in fetuses may progress to hypoplastic left heart syndrome (HLHS) with need for postnatal single ventricular (SV) palliation. Fetal aortic valvuloplasty (FAV) is performed to achieve postnatal biventricular (BV) circulation. However, the impact of FAV on fetal myocardial function is difficult to measure. Prediction of postnatal circulatory status and, therefore, counseling is challenging. METHODS: Retrospective study of fetuses with critical AS who underwent FAV. Global Longitudinal Peak Systolic Strain (GLPSS) of the left ventricle (LV) and right ventricle (RV) were retrospectively analyzed before and after intervention. Fisher's Exact Test and Mann-Whitney-U Test were used for univariant statistical analysis. RESULTS: 23 fetuses with critical AS were included. After intervention fetuses demonstrated more negative LV-GLPSS mean values post- vs. pre-intervention (- 5.36% vs. - 1.57%; p < 0.05). RV-GLPSS was decreased in all fetuses, there was no peri-interventional change. 20 fetuses were born alive. Postnatally, 10 had BV and 10 SV circulation. Improved post-interventional LV-GLPSS strain values correlated with BV outcome (p < 0.05). Pre-interventional continuous LV-GLPSS values correlated with postnatal SV vs. BV outcome (p < 0.05). CONCLUSION: In some fetuses, LV myocardial function assessed by speckle tracking echocardiography (STE) improves after FAV. Improved post-interventional LV-GLPSS correlates with biventricular postnatal outcome. Furthermore, pre-interventional LV- and RV-GLPSS correlate with postnatal outcome. Further studies are needed to asses, if pre-interventional STE parameters might predict which fetuses will benefit from FAV with postnatal BV circulation.

Fetal NT-proBNP levels and their course in severe anemia during intrauterine treatment.

PURPOSE: In adults and fetuses, N-terminal pro-B-type natriuretic peptide (NT-proBNP) is a marker of cardiac failure and myocardial remodelling. We examined the effect of anemia and intrauterine transfusion (IUT) on NT-proBNP concentrations in fetuses with anemia and established gestational age-dependent reference values of a control group. METHODS: We analyzed NT-proBNP levels in anemic fetuses that underwent serial intrauterine transfusions (IUT), focusing on different causes and severity of anemia and comparing the results to a non-anemic control group. RESULTS: In the control group, the average NT-proBNP concentration was 1339 ± 639 pg/ml, decreasing significantly with increasing gestational age (R = - 74.04, T = - 3.65, p = 0.001). Subjects had significantly higher NT-proBNP concentrations before initiation of IUT therapy (p < 0.001), showing fetuses with parvovirus B19 (PVB19) infection having the highest concentrations. Hydropic fetuses also showed an increased NT-proBNP concentration compared to non-hydropic fetuses (p < 0.001). During the course of therapy, NT-proBNP concentration before subsequent IUT decreased significantly from pathologically high levels, while MoM-Hb and MoM-MCA-PSV remained pathological. CONCLUSION: NT-pro BNP levels in non-anemic fetuses are higher than in postnatal life, decreasing with ongoing pregnancy. Anemia is a hyperdynamic state and its severity correlates with circulating NT-proBNP levels. Highest concentrations occur in fetuses with hydrops and with PVB19 infection, respectively. Treatment by IUT leads to a normalisation of NT-proBNP concentrations, so the measurement of its levels may be useful in therapy monitoring.

Aorto-Left Ventricular Tunnel - Prenatal Diagnosis and Outcome.

PURPOSE: Aorto-left ventricular tunnel (ALVT) is an extremely rare, albeit prenatally detectable, extracardiac channel that connects the ascending aorta to the cavity of the left ventricle. MATERIALS AND METHODS: All ALVTs diagnosed prenatally (2006-2020) in five tertiary referral centers were retrospectively assessed for prenatal ultrasound findings, intrauterine course, postnatal outcome, and surgical treatment. We focused on the size of the tunnel and alterations of perfusion of the left ventricular outflow tract and aortic arch. RESULTS: 11 fetuses were diagnosed with ALVT at a mean gestational age of 24.8 weeks. All cases were associated with severe dilatation of the left ventricle and a to-and-fro flow in the left outflow tract. Signs of congestive heart failure were present in five fetuses, four of which were terminated and one of which died in the neonatal period. One fetus died in utero at 34 weeks without prior signs of cardiac failure. Of the five survivors, two underwent the Ross procedure. In both cases the prenatal left ventricular outflow was exclusively via a large tunnel. The remaining three neonates underwent patch closure of the tunnel. In these cases, the prenatal outflow of the left ventricle was via the aortic valve and simultaneously over the tunnel. CONCLUSION: Prenatal diagnosis of ALVT should be considered in the presence of left ventricular hypertrophy, dilatation of the aortic root, and to-and-fro flow in the aortic outflow tract. Signs of heart failure are associated with an unfavorable outcome. Large tunnels, particularly in combination with the absence of flow over the aortic valve, may be an unfavorable predictor of surgical repair.

Effect of using different component combinations for orthodontic bracket bonding with self-etch primers.

PURPOSE: To evaluate bonding quality for orthodontic bracket bonding with different component combinations of self-etch primers in vitro. METHODS: Metallic brackets were bonded to bovine lower incisors and assigned to groups. Group 1: comparison of self-etch (Transbond™ Plus, 3M™ Unitek, Neuss, Germany, n = 30; BrackFix® primer SE, VOCO®, Cuxhaven, Germany, n = 20) and etch-and-rinse bonding systems (Transbond™ XT, n = 20; BrackFix®, n = 20); group 2: comparison of different self-etch primer (Transbond™ Plus; BrackFix® primer SE) and adhesive (Transbond™ XT, n = 20; BrackFix®, n = 20) product combinations; group 3: testing cyclic fatigue bond strength of self-etch bonding systems (Transbond™ Plus, n = 20; BrackFix® primer SE, n = 20). All teeth were tested for shear bond strength according to the DIN-13990 standard, the adhesive remnant index (ARI) and enamel fractures were determined microscopically (10 נmagnification). RESULTS: The mean shear bond strength of the self-etch (Transbond™ Plus: 16.38 ± 3.68 MPa; BrackFix® primer SE: 16.24 ± 1.73 MPa) and etch-and-rinse bonding systems (Transbond™ XT: 18.45 ± 2.56 MPa; BrackFix®: 17 ± 5.2 MPa) were of a clinically adequate order of magnitude (≥ 6-10 MPa) and were not statistically different. The component combination BrackFix® primer SE/Transbond™ XT adhesive led to a significantly lower shear bond strength (11.99 ± 3.68 MPa). There were no significant differences between static and fatigue shear bond strengths of self-etch bonding systems. Mean ARI scores mostly ranged between 4 and 5. The combination of the self-etch primer Transbond™ Plus with the BrackFix® adhesive led to a significantly increased enamel fracture rate. CONCLUSIONS: Based on the present findings bond strength of self-etch primers was equal to etch-and-rinse primers for bracket bonding. Combining different self-etch bonding systems might alter the clinical performance.

Evaluation of an OSCE's implementation and a two-step approach for a theoretical and practical training program in Obstetrics and Gynecology.

Objective structured clinical examination (OSCE) is a well-known assessment method to evaluate clinical skills and competence in healthcare. Following the recently reformed National Competence-Based Catalog of Learning Objectives in Medicine, the implementation of this assessment method in the training program for medical students is now obligatory in Germany. This major change requires a reorganization not only of the training programs but also of the students themselves and the way they learn. We performed a poll evaluating the students' opinions regarding these major changes and the implementation of the OSCE with a new training program. To implement this assessment method and to evaluate the OSCE, Kern's six-step approach comprising (1) problem identification and general needs assessment, (2) needs assessment of the targeted learners, (3) goals and objectives, (4) educational strategies, (5) implementation, and (6) evaluation and feedback was applied. To evaluate and gather feedback, a poll was used to analyze the student's opinions regarding OSCE in gynecology and obstetrics and OSCE in general, in addition to the regular analysis of the students' results. To reform the educational strategy, a two-step approach was developed: First, the students completed the regular training program and a written examination, and second, they participated in a 1-week clerkship, in small group teaching, and in the OSCE. The OSCE stations were developed primarily based on the National Competence-Based Catalog and the German Catalog of Learning Objectives in Medicine, as well as on the feedback of experts reflecting their expectations for physicians beginning their careers. The students performed well in the OSCE and gave positive feedback regarding this examination method. Furthermore, they welcomed the upcoming changes by considering OSCE a valuable assessment tool, and they showed appreciation for the two-step approach by supporting the combination of an OSCE and a written examination. Thus, this article presents the implementation of an OSCE and a strategy for the adaptation of the curriculum to fulfill the new OSCE requirements and-to our knowledge-reveals students' primary opinions regarding the changes in their medical training program for the first time.

Intrauterine Valvuloplasty in Severe Aortic Stenosis-A Ten Years Single Center Experience.

Objective: To assess the course and outcome of fetal aortic valvuloplasty (FAV) in fetuses with severe aortic stenosis (SAS) in a single center. Methods: All fetuses with a prenatal diagnosis of SAS with subsequent FAV were retrospectively collected in one tertiary center for fetal medicine over a period of 10 years. In the study, period fetuses with SAS were considered suitable for FAV in the presence of markedly elevated left ventricular pressures (maximum velocity of mitral regurgitation (MR Vmax) >250 cm/s and/or maximum velocity of aortic stenosis (AS Vmax) >250 cm/s), retrograde flow in the transverse aortic arch and a left ventricular length Z-score >−1. Results: In the study period 29 fetuses with AS were treated with 38 FAV. If reinterventions are included 82.7% of fetuses received a technically successful FAV. Procedure related death occurred in three (10.3%) cases, spontaneous fetal death in 2 (6.9%), and termination of pregnancy was performed in 3 cases (10.3%). Among the 21 live births (72.4%), four died in infancy. Among the remaining survivors, 8/17 (47.1%) had a biventricular outcome at the age of one year, 8/17 (47.1%) were univentricular and one infant (5.9%) is biventricular at the age of eight months. Fetuses with biventricular outcome had significantly greater left ventricular (LV) length Z-scores (p = 0.031), and lower tricuspid to mitral valve (TV/MV) ratios (p = 0.003). Conclusions: FAV has a high technical success rate and a low rate of procedure related mortality if performed in experienced hands. The success rate of biventricular circulation at the age of one year is moderate and seems to depend rather on the center's experience and postnatal surgical strategies than solely on prenatal selection criteria. In the absence of randomized controlled trials, FAV remains an experimental intervention.

Intrauterine Thoracoamniotic Shunting of Fetal Hydrothorax with the Somatex Intrauterine Shunt: Intrauterine Course and Postnatal Outcome.

(1) Background: Severe fetal hydrothorax can be treated by intrauterine thoracoamniotic shunting (TAS). The aim of this study was to assess perinatal outcome and complication rates of TAS with a novel Somatex intrauterine shunt. (2) Methods: This is a single-center retrospective study of all fetuses with hydrothorax treated with TAS using a Somatex shunt between 2014 and 2020. (3) Results: A total of 39 fetuses were included in the study. Mean gestational age at first intervention was 27.4 weeks (range 19-33). Of these, 51% (n = 20) of fetuses had fetal hydrops, which resolved in 65% (13/20) before delivery. The live birth rate was 97% (n = 38), and 74% (n = 29) survived the neonatal period. The rate of postnatal pulmonary complications was high, with 88% of neonates requiring any kind of ventilatory support. There were 23% (n = 9) genetic abnormalities (trisomy 21 and Noonan syndrome). (4) Conclusions: TAS with a Somatex shunt has a high technical success rate, leading to high neonatal survival rates. Pregnancy and neonatal outcome is comparable to TAS for fetal hydrothorax using different shunt types.

Outcome of Monochorionic Pregnancies after Selective Feticide with Bipolar Cord Coagulation: A German Single Center Experience.

OBJECTIVES: To review the outcome of complicated monochorionic pregnancies after fetoscopic selective feticide with bipolar cord coagulation in an experienced German center. METHODS: All cases that underwent selective feticide using fetoscopic bipolar umbilical cord occlusion (and simultaneous dissection in monoamniotics) at the University of Bonn in the past 10 years were retrospectively analyzed for antenatal and neonatal course and outcome. An adverse outcome was defined as either intrauterine death (IUD), neonatal death (NND), preterm prelabour rupture of membranes (PPROM), or preterm delivery (PTD) before 32.0 weeks of gestation. RESULTS: We diagnosed 56 monochorionic pregnancies, including 43 diamniotic and 8 monoamniotic twins, as well as 5 triplets, complicated by discordant fetal anomalies (n = 10), selective intrauterine growth restriction (n = 29), twin-to-twin transfusion syndrome (n = 13), twin reversed arterial perfusion sequence (n = 3), or severe early twin anemia polycythemia sequence (n = 1), that underwent fetoscopic selective feticide in the 10 years study period. Selective feticide was performed by bipolar cord coagulation at a median gestational age of 21.2 weeks. PPROM occurred in 11 cases, 7 (12.5%) before 32.0 weeks and 4 (7.1%) between 34.0 and 36.0 weeks, respectively. There were five (8.9%) co-twins IUDs at a median of 2 weeks after the intervention. We observed 12 (21.4%) PTDs before 32.0 weeks of gestation and 2 (3.6%) NNDs. Mean gestational age at delivery was 37.1 weeks, with an overall survival of the co-twin of 87.5%. CONCLUSION: In experienced hands, fetoscopic selective feticide is an effective treatment in complicated monochorionic pregnancies. By sacrificing a sick fetus that jeopardizes the entire pregnancy, a higher survival rate of the co-twin can be achieved.

Prenatal Diagnosis and Outcome of Scimitar Syndrome: A Case Series of Six Patients.

Scimitar syndrome is a rare disease characterized by hypoplasia of the right lung and partial anomalous pulmonary venous drainage to the inferior vena cava. All cases with a prenatal diagnosis of scimitar syndrome with or without associated malformations in an 18-year period (2000-2018) in two large tertiary referral centers (University of Bonn and University of Cologne, Germany) were retrospectively reviewed for the intrauterine course and postnatal outcome. Six cases were diagnosed in the study period. All presented with hypoplasia of the right lung, right-sided mediastinal shift, and abnormal pulmonary venous drainage to the inferior vena cava. Systemic arterial blood supply to the right lung, albeit postnatally present in all cases, could not be detected prenatally. Major associated anomalies were present in all cases and included atrial septal defect (n = 5), coarctation (n = 3), diaphragmatic hernia (n = 2), and VACTERL association (n = 1). Half of the cohort died within 6 months after birth and all three survivors suffer from long-term pulmonary sequelae. The primary hint to the prenatal diagnosis of scimitar syndrome is the abnormal position of the heart in the chest. If searched for, abnormal venous drainage can be identified prenatally and confirms the diagnosis. The prognosis depends on the presence of associated major anomalies and the need for neonatal intervention.

Management and outcome of prenatal absent pulmonary valve syndrome.

OBJECTIVE: The purpose of this study was to explore the outcome in fetuses with prenatal diagnosis of absent pulmonary valve syndrome (APVS) on ultrasound imaging. METHODS: A manual web scraping technique was utilized, where MEDLINE and EMBASE were searched along the combination with other relevant medical subject headings such as "absent pulmonary valve syndrome", "prenatal APVS" and "APVS/outcome". The observed outcomes encompassed the rate of chromosomal abnormalities, associations and malformations linked to APVS and fetuses with APVS. A quality assessment of the included studies was also performed. We used meta-analyses of proportions to combine data and fixed or random-effects models according to the heterogeneity of the results. RESULTS: Seven studies including 199 fetuses with APVS were included in the analysis. The median gestational age at referral to the tertiary center was 24.8 weeks. An association to tetralogy of Fallot (TOF) could be seen in 84.4% of all cases. In total 140 out of 199 cases underwent invasive testing, with a total number of 55 abnormal karyotypes [39.3% (95% CI 31.1-47.9%)]. 35.2% of the patients opted for termination of pregnancy (95% CI 28.5-42.3%). CONCLUSION: The analysis underlines the distribution of fetuses with APVS, with 84.4% of cases presenting with TOF/APVS and only 12.6% having APVS/intact ventricular septum (IVS). Larger and more prospective study analyses is now needed, especially focusing on long-term follow-up periods of fetuses and children with APVS. Particularly as the postnatal course shows great variety depending on prenatal diagnosis.

Outcome of TRAP Sequence Treated in the First Trimester - A Ten-Year Single-Center Experience.

PURPOSE: To evaluate the outcome of first trimester intervention by intrafetal laser (IFL) in pregnancies complicated by twin reversed arterial perfusion (TRAP). MATERIALS AND METHODS: For a 10-year study period, all patients with TRAP diagnosed < 14.0 weeks of gestation were retrospectively analyzed for intrauterine course and outcome. Monoamniotic pregnancies were excluded. Patients were offered either intervention by IFL in the first trimester, expectant management, or termination of pregnancy (TOP). Adverse outcome was defined as either intrauterine death (IUD), neonatal death, or preterm birth. RESULTS: In 45 cases TRAP was diagnosed. 17 monoamniotics were excluded. The cohort was divided into two groups according to management. Group A: 24 cases underwent IFL and group B: 4 cases were managed expectantly. No patient opted for TOP. In group A, 70.8 % of pump twins were born alive, including one preterm delivery, and 29.2 % died within four days after the intervention. All 4 expectantly managed cases in group B had an adverse outcome (1 preterm delivery, 3 IUDs < 15.0 weeks). There were no neonatal deaths. In cases treated by IFL, a comparison of survivors and non-survivors identified no significant differences in gestational age at IFL or any of the assessed biometrical and functional parameters. There was a trend towards better outcome in the second half of the study period. CONCLUSION: IFL in first trimester TRAP sequence is technically feasible but is associated with significant mortality, albeit less than previously reported. No risk stratification is possible using the investigated parameters. However, there seems to be a learning curve.

Outcome of Monochorionic Monoamniotic Twin Reversed Arterial Perfusion Sequence Diagnosed in the First Trimester.

INTRODUCTION: The aim of this study is to evaluate the outcome of pregnancies complicated by monochorionic monoamniotic twin reversed arterial perfusion sequence (MOMA TRAP) diagnosed in the first trimester. METHODS: All patients diagnosed with MOMA TRAP sequence <14.0 weeks of gestation in a 10-year study period were retrospectively analyzed for intrauterine course and outcome. All patients were offered either expectant management or intrauterine intervention. Adverse outcome was defined as either intrauterine death (IUD), neonatal death or preterm birth <34.0 weeks of gestation. RESULTS: In the study period, 17 cases with MOMA TRAP sequence were diagnosed. Of these, 2 couples opted for termination of pregnancy. The remaining 15 were divided into 2 groups depending on the management: group A (n = 8) with expectant management and group B (n = 7) with intrauterine intervention. All fetuses in group A died before 20 weeks. Survival in group B was significantly better with 4/7 (57.1%) life births at a median of 39.6 weeks of gestation (p = 0.0256). The reasons for IUD in the 3 cases in group B were hemodynamic, strangulation, and bleeding complications during intervention. CONCLUSIONS: Intrauterine intervention in MOMA TRAP pregnancies significantly improves neonatal survival, although it is still associated with a substantial risk for IUD by hemodynamic complications or entanglement.

Point-of-care ultrasound in obstetrics and gynecology.

BACKGROUND: The rapid technical development and portability of ultrasound systems over recent years has had a profound impact on the area of point-of-care-ultrasound (POCUS), both in general medicine and in obstetrics and gynecology. The use of POCUS enables the clinician to perform the ultrasound scan either at the medical office or the patient's bedside and used as an extension of the physical examination. Real-time images can immediately be correlated with the patient's symptoms, and any changes in a (critical) patient's condition can be more rapidly detected. POCUS IN OBGYN: POCUS is also suitable for time-critical scenarios, and depending on the situation and its dynamics, the course and results of any therapy may be observed in real time. POCUS should be considered to be a routine extension of practice for most OB/GYN clinicians as it can give immediate answers to what could be life-threatening situations for the mother and/or baby. With its proven usefulness, the applications and use of POCUS should be incorporated in teaching programs for medical students, OBGYN residents and emergency physicians.

Lung Sonography in Obstetrics during COVID-19.

In the current coronavirus SARS-CoV-2 pandemic, certain patients are becoming seriously ill. Lung pathologies are common, and some patients even go on to develop acute respiratory distress syndrome (ARDS), which requires intubation and artificial respiration of the critically ill patient. Imaging of the lung is absolutely necessary to obtain a diagnosis, assess the course of disease and for treatment. Particularly in gynecology and obstetrics (OBGYN), ultrasound scans of the lung can be a useful additional tool when caring for pregnant patients in the delivery room. As obstetricians use ultrasound imaging a lot in routine clinical practice, in the current pandemic setting, routine prenatal imaging screening could be expanded by the addition of ultrasound scans of the lung. Lung sonography can offer important additional information, particularly in obstetrics where the indications for radiation-emitting imaging are particularly restrictive. If there is a sonographic suspicion of lung involvement, then, depending on the symptoms and the morphological extent of the ultrasound findings, it may be necessary to consider admitting the patient to hospital for close fetal and maternal monitoring.

Analysing the potential of hydrophilic adhesive systems to optimise orthodontic bracket rebonding.

INTRODUCTION: Bond failure during fixed orthodontic treatment is a frequently occurring problem. As bracket rebonding is associated with reduced shear bond strength, the aim of the present investigation is to analyse the effect of different innovative rebonding systems to identify optimised rebonding protocols for orthodontic patient care. METHODS: Metallic brackets were bonded to the frontal enamel surfaces of 240 bovine lower incisors embedded in resin bases. Teeth were randomly divided into two major experimental groups: in group 1 a hydrophilic primer (Assure™ PLUS) was compared to commonly used orthodontic adhesives (Transbond XT™, BrackFix®, Grengloo™) and a zero control. In group 2 different rebonding systems were analysed using a hydrophilic primer (Assure™ PLUS), a methyl methacrylate-consisting primer (Plastic Conditioner) and a conventional adhesive (Transbond XT™). All teeth were tested for shear bond strength according to the DIN-13990 standard, the Adhesive Remnant Index and enamel fracture rate. RESULTS: The hydrophilic primer enhanced shear bond strength at first bonding (Assure™ PLUS 20.29 ± 4.95 MPa vs. Transbond XT™ 18.45 ± 2.57 MPa; BrackFix® 17 ± 5.2 MPa; Grengloo™ 19.08 ± 3.19 MPa; Meron 8.7 ± 3.9 MPa) and second bonding (Assure™ PLUS 16.76 ± 3.71 MPa vs. Transbond XT™ 13.06 ± 3.19 MPa). Using Plastic Conditioner did not seem to improve shear bond strength at rebonding (13.57 ± 2.94). When enamel etching was left out, required shear bond strength could not be achieved (Plastic Conditioner + Assure™ PLUS 8.12 ± 3.34 MPa; Plastic Conditioner: 3.7 ± 1.95 MPa). Hydrophilic priming systems showed decreased ARI-scores (second bonding: 2.63) and increased enamel fracture rates (first bonding: 55%; second bonding 21,05%). CONCLUSIONS: Based on the present study we found that rebonding strength could be compensated by the use of hydrophilic priming systems. The additional use of a methyl methacrylate-consisting primer does not seem to enhance shear bond strength. No etching approaches resulted in non-sufficient bond strength.

Genetic architecture of inherited retinal degeneration in Germany: A large cohort study from a single diagnostic center over a 9-year period.

We aimed to unravel the molecular genetic basis of inherited retinal degeneration (IRD) in a comprehensive cohort of patients diagnosed in the largest center for IRD in Germany. A cohort of 2,158 affected patients from 1,785 families diagnosed with IRD was analyzed by targeted next-generation sequencing (NGS). Patients with single-gene disorders (i.e., choroideremia and retinoschisis) were analyzed by Sanger sequencing and multiplex ligation-dependent probe amplification. Our study cohort accounts for ∼7% of the estimated 30,000 patients with IRD in Germany, thereby providing representative data for both the prevalence of IRDs and the mutation spectrum of IRD genes for the population in Germany. We achieved a molecular diagnostic rate of 35-95%, depending on the clinical entities, with a high detection rate for achromatopsia, retinoschisis, and choroideremia, and a low detection rate for central areolar choroidal dystrophy and macular dystrophy. A total of 1,161 distinct variants were identified, including 501 novel variants, reaffirming the known vast genetic heterogeneity of IRD in a mainly outbred European population. This study demonstrates the clinical utility of panel-based NGS in a large and highly heterogeneous cohort from an outbred population and for the first time gives a comprehensive representation of the genetic landscape of IRDs in Germany. The data are valuable and crucial for the scientific community and healthcare providers, but also for the pharmaceutical industry in the progressing field of personalized medicine and gene therapy.

Forces during cellular uptake of viruses and nanoparticles at the ventral side.

Many intracellular pathogens, such as mammalian reovirus, mimic extracellular matrix motifs to specifically interact with the host membrane. Whether and how cell-matrix interactions influence virus particle uptake is unknown, as it is usually studied from the dorsal side. Here we show that the forces exerted at the ventral side of adherent cells during reovirus uptake exceed the binding strength of biotin-neutravidin anchoring viruses to a biofunctionalized substrate. Analysis of virus dissociation kinetics using the Bell model revealed mean forces higher than 30 pN per virus, preferentially applied in the cell periphery where close matrix contacts form. Utilizing 100 nm-sized nanoparticles decorated with integrin adhesion motifs, we demonstrate that the uptake forces scale with the adhesion energy, while actin/myosin inhibitions strongly reduce the uptake frequency, but not uptake kinetics. We hypothesize that particle adhesion and the push by the substrate provide the main driving forces for uptake.

Lattice Shrinkage by Incorporation of Recombinant Starmaker-Like Protein within Bioinspired Calcium Carbonate Crystals.

The biological mediation of mineral formation (biomineralization) is realized through diverse organic macromolecules that guide this process in a spatial and temporal manner. Although the role of these molecules in biomineralization is being gradually revealed, the molecular basis of their regulatory function is still poorly understood. In this study, the incorporation and distribution of the model intrinsically disordered starmaker-like (Stm-l) protein, which is active in fish otoliths biomineralization, within calcium carbonate crystals, is revealed. Stm-l promotes crystal nucleation and anisotropic tailoring of crystal morphology. Intracrystalline incorporation of Stm-l protein unexpectedly results in shrinkage (and not expansion, as commonly described in biomineral and bioinspired crystals) of the crystal lattice volume, which is described herein, for the first time, for bioinspired mineralization. A ring pattern was observed in crystals grown for 48 h; this was composed of a protein-enriched region flanked by protein-depleted regions. It can be explained as a result of the Ostwald-like ripening process and intrinsic properties of Stm-l, and bears some analogy to the daily growth layers of the otolith.

Silica Nanoparticles for Intracellular Protein Delivery: a Novel Synthesis Approach Using Green Fluorescent Protein.

In this study, a novel approach for preparation of green fluorescent protein (GFP)-doped silica nanoparticles with a narrow size distribution is presented. GFP was chosen as a model protein due to its autofluorescence. Protein-doped nanoparticles have a high application potential in the field of intracellular protein delivery. In addition, fluorescently labelled particles can be used for bioimaging. The size of these protein-doped nanoparticles was adjusted from 15 to 35 nm using a multistep synthesis process, comprising the particle core synthesis followed by shell regrowth steps. GFP was selectively incorporated into the silica matrix of either the core or the shell or both by a one-pot reaction. The obtained nanoparticles were characterised by determination of particle size, hydrodynamic diameter, ζ-potential, fluorescence and quantum yield. The measurements showed that the fluorescence of GFP was maintained during particle synthesis. Cellular uptake experiments demonstrated that the GFP-doped nanoparticles can be used as stable and effective fluorescent probes. The study reveals the potential of the chosen approach for incorporation of functional biological macromolecules into silica nanoparticles, which opens novel application fields like intracellular protein delivery.

Isolation of 'Candidatus Nitrosocosmicus franklandus', a novel ureolytic soil archaeal ammonia oxidiser with tolerance to high ammonia concentration.

Studies of the distribution of ammonia oxidising archaea (AOA) and bacteria (AOB) suggest distinct ecological niches characterised by ammonia concentration and pH, arising through differences in substrate affinity and ammonia tolerance. AOA form five distinct phylogenetic clades, one of which, the 'Nitrososphaera sister cluster', has no cultivated isolate. A representative of this cluster, named 'Candidatus Nitrosocosmicus franklandus', was isolated from a pH 7.5 arable soil and we propose a new cluster name:'Nitrosocosmicus' While phylogenetic analysis of amoA genes indicates its association with the Nitrososphaera sister cluster, analysis of 16S rRNA genes provided no support for a relative branching that is consistent with a 'sister cluster', indicating placement within a lineage of the order Nitrososphaerales 'Ca.N. franklandus' is capable of ureolytic growth and its tolerances to nitrite and ammonia are higher than in other AOA and similar to those of typical soil AOB. Similarity of other growth characteristics of 'Ca.N. franklandus' with those of typical soil AOB isolates reduces support for niche differentiation between soil AOA and AOB and suggests that AOA have a wider physiological diversity than previously suspected. In particular, the high ammonia tolerance of 'Ca.N. franklandus' suggests potential contributions to nitrification in fertilised soils.