Genotyping of European Toxoplasma gondii strains by a new high-resolution next-generation sequencing-based method.

PURPOSE: A new high-resolution next-generation sequencing (NGS)-based method was established to type closely related European type II Toxoplasma gondii strains. METHODS: T. gondii field isolates were collected from different parts of Europe and assessed by whole genome sequencing (WGS). In comparison to ME49 (a type II reference strain), highly polymorphic regions (HPRs) were identified, showing a considerable number of single nucleotide polymorphisms (SNPs). After confirmation by Sanger sequencing, 18 HPRs were used to design a primer panel for multiplex PCR to establish a multilocus Ion AmpliSeq typing method. Toxoplasma gondii isolates and T. gondii present in clinical samples were typed with the new method. The sensitivity of the method was tested with serially diluted reference DNA samples. RESULTS: Among type II specimens, the method could differentiate the same number of haplotypes as the reference standard, microsatellite (MS) typing. Passages of the same isolates and specimens originating from abortion outbreaks were identified as identical. In addition, seven different genotypes, two atypical and two recombinant specimens were clearly distinguished from each other by the method. Furthermore, almost all SNPs detected by the Ion AmpliSeq method corresponded to those expected based on WGS. By testing serially diluted DNA samples, the method exhibited a similar analytical sensitivity as MS typing. CONCLUSION: The new method can distinguish different T. gondii genotypes and detect intra-genotype variability among European type II T. gondii strains. Furthermore, with WGS data additional target regions can be added to the method to potentially increase typing resolution.

Invasive electrical stimulation in the treatment of avascular osteonecrosis of the femoral head - mid-term results.

The study aimed to evaluate the outcomes of osteonecrosis of the femoral head (ONFH) in adults after surgical treatment including invasive electromagnetic osteostimulation (E-Stim). Further, the influence of disease stage and several comorbidities on the joint preservation rate should be examined. Sixty patients (66 hip joints) with ONFH were included in this retrospective cross-sectional analysis (mean follow-up: 58 months, 19-110 months). Potential ONFH risk factors and comorbidities (ONFH stage, age, sex, alcohol, smoking, cortisone medication, chemotherapy) were recorded. The influence of specific parameters on the joint preservation rates was evaluated by a multivariate logistic regression analysis. Finally, patients with preserved hip joints underwent an assessment of their last available X-rays. The joint preservation rate depended on the initial ONFH Steinberg stage (I+II: 82.8%, III: 70.8%, ≥ IVa: 38.5%). Initially collapsed ONFH (p ≤ 0.001) and cortisone therapy (p = 0.004) significantly decreased the joint preservation rates. In case of progressed ONFH, the presence of ≥ 2 risk factors resulted in higher THA conversion rates (stage III: OR 18.8; stage ≥IVa: OR 12). In 94% of the available X-rays, the ONFH stage improved or did not progress. No complications could be attributed to the E-Stim device or procedure. The present surgical protocol including minimally invasive E-Stim revealed high joint preservation rates for non-collapsed ONFH after mid-term postoperative follow-up. Especially in progressed ONFH, the-risk profile seems to be crucial and hence, for joint preserving surgery, careful patient selection is recommended.

[Saturated fatty acids and cardiovascular risk : Is a revision of the recommendations on nutrition indicated?] / Gesättigte Fettsäuren und kardiovaskuläres Risiko : Ist eine Revision der Ernährungsempfehlungen angezeigt?

The "fat hypothesis of coronary heart disease", according to which saturated fatty acids (SFA) increase the concentration of low-density lipoprotein-cholesterol (LDL-C) and consequently increase the risk for cardiovascular diseases, influenced the nutritional recommendations over the last 60 years, initially in the USA and later also in Europe. Over the years there accumulated a growing body of evidence from epidemiology and controlled clinical studies that the consumption of SFA per se was not associated with an increased cardiovascular risk and the limitation of consumption of SFA did not show a preventive effect. The focus on the SFA content negated the biologically heterogeneous and sometimes biologically favorable effects of various SFAs. In addition, it was neglected that SFAs in foodstuffs are bound in a variety of complex matrices, which are composed of dozens of nutrients with different structures and concomitant substances and therefore each triggers different biological responses and metabolic effects. Accordingly, such nutrient-based recommendations are principally not very productive and also difficult to realize. In addition, LDL­C is not a suitable marker to assess the effect of lifestyle interventions, such as nutrition or physical activity, on the global cardiovascular risk.

Outcome of Surgical and Conservative Treatment of Patients with Shoulder Impingement Syndrome - a Prospective Comparative Clinical Study.

PURPOSE OF THE STUDY Subacromial impingement is one of the most common reasons for shoulder pain. The surgical management of this condition has recently become the focus of criticism because of the rising number of surgical procedures and the lack of superiority of surgical over conservative treatment. In this prospective comparative study, we compared standardised conservative care with surgical treatment and placed special emphasis on the patients' ability to work. MATERIAL AND METHODS A total of 106 patients (25 women, 81 men; mean age: 45.4 12.3 years) were included in this prospective comparative clinical study. Patients in the non-operative arm (n = 42) received standardised physiotherapy. Patients in the surgical arm (n = 38) underwent arthroscopic subacromial decompression. All patients were followed up at 3, 6 and 12 months. Shoulder function (Constant score), pain (Numerical Rating Scale), and the duration of inability to work were assessed. RESULTS Shoulder function and pain improved significantly with both kinds of treatment. At no time of follow up we detected significant differences between the two treatment options. An analysis of the patients' ability to work showed that conservative treatment was superior to surgical treatment at 3-month follow-up (0.3 versus 5.0 weeks; p < 0.001) and between 4 and 6-month after intervention (0.2 versus 1.6 weeks; p = 0.032). DISCUSSION In the study presented here, significant improvements in function (Constant score) and pain (NRS) were achieved in both the non-operative and the surgical arm. There were no significant differences between the two groups at any time point. These results are similar to those reported by other authors in recent studies. Unlike other research work, however, our study demonstrated a major difference in the development and duration of inability to work. CONCLUSIONS Conservative and surgical treatment of subacromial impingement syndrome led to similar outcomes for shoulder pain and function at 3, 6 and 12 months after intervention. However, patients who were managed conservatively returned to work significantly earlier than patients who underwent surgery. Key words: subacromial impingement, shoulder, constant score, pain, ability to work.

Cross-cultural validity of the German version of the Pediatric Evaluation of Disability Inventory (PEDI-G)-a Rasch model application.

AIM: The aim of this study was to evaluate the cross-cultural validity of the German version of the Pediatric Evaluation of Disability Inventory (PEDI-G) when used in Austria, Germany and Switzerland. METHOD: A total of 118 girls and 144 boys participated in this study; 198 of the children (75.6%) had a developmental disability and 64 (24.4%) were without a known disability. The mean age was four years (range 11 months to 10 years and six months, SD 1.91). Item goodness of fit, differential item functioning (DIF) and differential test functioning (DTF) were evaluated by use of a Rasch model. RESULTS: Twenty-four (11.6%) out of 206 items of the Functional Skills Scale and one (5%) out of 20 items of the Caregiver Assistance Scale demonstrated misfit according to the Rasch model. Thirty-four (16.5%) out of 206 items of the Functional Skills Scale and no item from the Caregiver Assistance Scale demonstrated DIF. Almost half (46%) of the items demonstrating misfit also demonstrated DIF, indicating an association between them. The DIF by country only demonstrated a minimal impact on the person measures of the PEDI-G. INTERPRETATION: Even though some items did not meet the statistical and clinical criteria set, the PEDI-G can be used, on a preliminary basis as a valid tool to measure activities of daily living of children with and without a disability in these countries. Further larger studies are needed to evaluate more psychometric item properties of the PEDI-G in relation to context.

[Therapy of a gunshot fracture to the midfoot]. / Therapie einer Schussfraktur am Mittelfußknochen.

We report on a soldier with a gunshot wound to the midfoot. The diaphysis of MT 5 was pounded to pieces and there was a fracture in the diaphysis of MT 4. After debridement and fixation of MT 4 and MT 5 to MT 2 and 3 via two Kirschner wires we conducted a second look procedure with reconstruction of MT 5 via a piece of iliac crest. After eight weeks of unloading and removal of the last wire we started to put successively heavier weight on the foot.

High and novel genetic diversity of Francisella tularensis in Germany and indication of environmental persistence.

In Germany tularemia is a re-emerging zoonotic disease. Therefore, we investigated wild animals and environmental water samples for the presence and phylogenetic diversity of Francisella tularensis in the poorly studied Berlin/Brandenburg region. The phylogenomic analysis of three isolates from wild animals revealed three new subclades within the phylogenetic tree of F. tularensis [B.71 from a raccoon dog (Nyctereutes procyonoides); B.74 from a red fox (Vulpes vulpes), and B.75 from a Eurasian beaver (Castor fiber albicus)]. The results from histological, PCR, and genomic investigations on the dead beaver showed that the animal suffered from a systemic infection. Indications were found that the bacteria were released from the beaver carcass into the surrounding environment. We demonstrated unexpectedly high and novel phylogenetic diversity of F. tularensis in Germany and the fact that the bacteria persist in the environment for at least one climatic season. These findings support a broader host species diversity than previously known regarding Germany. Our data further support the assumption derived from previous serological studies of an underestimated frequency of occurrence of the pathogen in the environment and in wild animals. F. tularensis was isolated from animal species not previously reported as natural hosts in Germany.

Electrical Injuries in Animals: Causes, Pathogenesis, and Morphological Findings.

Electrical injuries in animals occur most often accidentally. They comprise contact to various forms of currents, including alternating, rotary, or direct currents. Depending on various parameters of the current (including the type of circuit, voltage, current and duration of exposure) and conditions of the animal (such as wet or dry hair coat and pathway of current through the body), lesions may be absent or may include early or localized development of rigor mortis, signs of acute circulatory failure, or severe thermoelectrical burns. Such burns may present as external current marks, singed hair or feathers, metallization of the skin, or occasionally internal electroporation injury resulting in muscle necrosis, hemolysis, vascular damage with thrombosis, injury to brain and spinal cord, or skeletal fractures. Furthermore, lightning strikes occur regularly in grazing animals, which have greater risk of death from step potentials (ground current) in addition to direct strike and contact injury. Such cases may have no lesions, external signs of linear or punctate burns, keraunographic markings, or exit burns on the soles of the hooves or the coronary bands. Besides detailed information about the circumstances at the location where the animal was found, electrical injuries in animals require a thorough morphological workup, including additional investigations in conjunction with certain knowledge about the possible lesion spectrum.

Thermal Injuries in Veterinary Forensic Pathology.

Localized thermal injuries in animals may be caused by exposure to fire and radiant heat, contact with hot items including hot liquids or steam, inhalation of hot air, and exposure to cold temperatures. In addition, animal fire victims may have intoxications caused by smoke gas. This article reviews the causes, pathogenetic aspects, morphological findings, additional investigations, differential diagnoses, and causes of death in various forms of thermal injuries. Since these cases do not occur frequently in diagnostic pathology, they represent a challenging task in general but also with respect to forensic or criminal aspects, such as whether a lesion represents an accidental or nonaccidental effect. Besides detailed information about the circumstances at the location, thermal injuries in animals require a thorough morphological evaluation, including additional investigations in conjunction with a profound knowledge about the possible lesion spectrum and suitable additional investigations.

IL7RA haplotype-associated alterations in cellular immune function and gene expression patterns in multiple sclerosis.

Interleukin-7 receptor alpha (IL7RA) is among the top listed candidate genes influencing the risk to develop multiple sclerosis (MS), an inflammatory demyelinating disease of the central nervous system. Soluble IL-7RA (sIL-7RA) protein and mRNA levels vary among the four common IL7RA haplotypes. Here we show and confirm that protective haplotype carriers have three times lower sIL-7RA serum levels than the other three haplotypes. High sIL-7RA concentrations significantly decrease IL-7-mediated STAT5 phosphorylation in CD4(+) T cells. Transcriptome analysis of unstimulated and stimulated CD4(+) T cells of MS patients carrying the different IL7RA haplotypes revealed complex and overlapping patterns in genes participating in cytokine signaling networks, apoptosis, cell cycle progression and cell differentiation. Our findings indicate that genetic variants of IL7RA result in haplotype-associated differential responsiveness to immunological stimuli that influence MS susceptibility not exclusively by varying levels of sIL-7RA.

Probing the energy barriers and magnetization reversal processes of nanoperforated membrane based percolated media.

Magnetization reversal processes in Co/Pt multilayers prepared on nanoperforated templates are probed by magnetization relaxation measurements. The signature of pinning controlled domain wall movement as expected for percolated media is identified. This contrasts with the nucleation-type reversal mechanism of a Co/Pt reference film prepared on a smooth substrate. A zero field energy barrier of 93kBT is determined by fluctuation field measurements and is elucidated by micromagnetic calculations using the nudged elastic band method. This value is sufficiently large to qualify the material as a promising percolated medium.

Tularaemia seroprevalence of captured and wild animals in Germany: the fox (Vulpes vulpes) as a biological indicator.

A total of 2475 animals from Germany, both captive and wild, were tested for antibodies against Francisella tularensis to obtain more knowledge about the presence of this pathogen in Germany. An indirect and a competitive ELISA served as screening methods, positive and inconclusive samples were confirmed by Western blot. Of the zoo animals sampled between 1992 and 2007 (n = 1122), three (0·3%) were seropositive. The seroconversion of a hippopotamus in Berlin Zoo was documented. From 1353 serum samples of wild foxes (Vulpes vulpes), raccoon dogs (Nyctereutes procyonoides) and wild boars (Sus scrofa), collected between 2005 and 2009 in the federal state of Brandenburg (surrounding Berlin), a total of 101 (7·5%) tested positive for antibodies to F. tularensis lipopolysaccharide. Our results indicate a higher seroprevalence of F. tularensis in wildlife in eastern Germany than commonly assumed. Furthermore, we found foxes and raccoon dogs to be biological indicators for tularaemia.

No proinflammatory signature in CD34+ hematopoietic progenitor cells in multiple sclerosis patients.

Autologous hematopoietic stem cell transplantation (aHSCT) has been used as a therapeutic approach in multiple sclerosis (MS). However, it is still unclear if the immune system that emerges from autologous CD34+ hematopoietic progenitor cells (HPC) of MS patients is pre-conditioned to re-develop the proinflammatory phenotype. The objective of this article is to compare the whole genome gene and microRNA expression signature in CD34+ HPC of MS patients and healthy donors (HD). CD34+ HPC were isolated from peripheral blood of eight MS patients and five HD and analyzed by whole genome gene expression and microRNA expression microarray. Among the differentially expressed genes (DEGs) only TNNT1 reached statistical significance (logFC=3.1, p<0.01). The microRNA expression was not significantly different between MS patients and HD. We did not find significant alterations of gene expression or microRNA profiles in CD34+ HPCs of MS patients. Our results support the use of aHSCT for treatment of MS.

Magnetic films on nanoperforated templates: a route towards percolated perpendicular media.

We present a study on the magnetization reversal in Co/Pt multilayer films with an out-of-plane easy axis of magnetization deposited onto substrates with densely distributed perforations with an average period as small as 34 nm. Deposition of magnetic Co/Pt multilayers onto the nanoperforated surface results in an array of magnetic nanodots surrounded by a continuous magnetic film. Following the evolution of the magnetic domain pattern in the system, we suggest that domain walls are pinned on structural inhomogeneities given by the underlying nanoperforated template. Furthermore, a series of micromagnetic simulations was performed in order to understand the modification of the pinning strength of domain walls due to the magnetic interaction between nanodots and the surrounding film. The results of the simulations show that magnetic exchange coupling between the nanodots and the surrounding film strongly influences the pinning behavior of the magnetic domain walls which can be optimized to provide maximal pinning.

Peripheral versus central cannulation for extracorporeal membrane oxygenation: a comparison of limb ischemia and transfusion requirements.

BACKGROUND: Extracorporeal membrane oxygenation (ECMO) can be instituted centrally, through the right atrium and ascending aorta, or peripherally, most commonly using the femoral artery and vein. We sought to investigate the impact of the mode of cannulation on the incidence of limb ischemia, perfusion and overall morbidity. METHODS: A retrospective analysis of 50 consecutive patients over 5 years who underwent ECMO by central or peripheral cannulation was performed. RESULTS: There was no difference in the incidence of limb ischemia and end-organ perfusion when peripheral and central cannulation cohorts were compared. Central cannulation was associated with a higher incidence of bleeding from the cannulation site (64% vs. 18%, P = 0.002), blood product utilization and reoperation (66% vs. 14%, P < 0.0001). 30-day mortality was similar in both cohorts (46% peripheral, 50% central, P = 0.8). CONCLUSION: Our results suggest that there is comparable tissue perfusion and limb ischemia with both cannulation techniques. Central cannulation is associated with a higher incidence of bleeding, higher transfusion rates, a greater need for reoperation and greater resource utilization. Therefore, peripheral cannulation is safe and may be advantageous in certain clinical scenarios.

Establishment of a cross-European field site network in the ALARM project for assessing large-scale changes in biodiversity.

The field site network (FSN) plays a central role in conducting joint research within all Assessing Large-scale Risks for biodiversity with tested Methods (ALARM) modules and provides a mechanism for integrating research on different topics in ALARM on the same site for measuring multiple impacts on biodiversity. The network covers most European climates and biogeographic regions, from Mediterranean through central European and boreal to subarctic. The project links databases with the European-wide field site network FSN, including geographic information system (GIS)-based information to characterise the test location for ALARM researchers for joint on-site research. Maps are provided in a standardised way and merged with other site-specific information. The application of GIS for these field sites and the information management promotes the use of the FSN for research and to disseminate the results. We conclude that ALARM FSN sites together with other research sites in Europe jointly could be used as a future backbone for research proposals.

p120, a p120-related protein (p100), and the cadherin/catenin complex.

Cadherins and catenins play an important role in cell-cell adhesion. Two of the catenins, beta and gamma, are members of a group of proteins that contains a repeating amino acid motif originally described for the Drosophila segment polarity gene armadillo. Another member of this group is a 120-kD protein termed p120, originally identified as a substrate of the tyrosine kinase pp60src. In this paper, we show that endothelial and epithelial cells express p120 and p100, a 100-kD, p120-related protein. Peptide sequencing of p100 establishes it as highly related to p120. p120 and p100 both appear associated with the cadherin/catenin complex, but independent p120/catenin and p100/catenin complexes can be isolated. This association is shown by coimmunoprecipitation of cadherins and catenins with an anti-p120/p100 antibody, and of p120/p100 with cadherin or catenin antibodies. Immunocytochemical analysis with a p120-specific antibody reveals junctional colocalization of p120 and beta-catenin in epithelial cells. Catenins and p120/p100 also colocalize in endothelial and epithelial cells in culture and in tissue sections. The cellular content of p120/p100 and beta-catenin is similar in MDCK cells, but only approximately 20% of the p120/p100 pool associates with the cadherin/catenin complex. Our data provide further evidence for interactions among the different arm proteins and suggest that p120/p100 may participate in regulating the function of cadherins and, thereby, other processes influenced by cell-cell adhesion.

Virological monitoring of white storks (Ciconia ciconia) for avian influenza.

Between 2003 and 2008, more than 600 white stork (Ciconia ciconia) nestlings in the German federal state of Brandenburg were ringed and examined for influenza A viruses. With the spread of highly pathogenic avian influenza virus (HPAIV) of subtype H5N1 among wild birds in Germany in spring 2006, dead wild birds, including 88 white storks, were tested for infection with HPAIV. Furthermore, fresh fecal samples were examined by RT-PCR to monitor the occurrence of HPAIV in adult storks. While the monitoring of nestlings and adult white storks failed to yield evidence of influenza A virus infections in these birds, two storks found dead in April 2006 in the same location tested positive for HPAIV H5N1. Sequence analysis revealed that the virus isolated from one of the storks belonged to clade 2.2, which was commonly found in wild birds in the north of Germany and other European countries during the epidemic in 2006. Despite these two cases, white storks seemed to serve as neither a vector nor as a reservoir for HPAIV in Germany. The risk of white storks transmitting HPAIV to domestic poultry and humans is low.