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As a zoonotic pathogen, the hepatitis E virus (HEV) leads to numerous infections in humans with different clinical manifestations. Especially genotype 3, as causative agent of a foodborne zoonosis, is transmitted to humans by ingestion of undercooked or raw meat containing liver from HEV‐infected animals. Although the virus' prevalence and dissemination in hosts like wild boar and pig have been well characterized, HEV is greatly understudied on a molecular level and reliable cell culture models are lacking. For this reason, the present study concentrated on the isolation and subsequent characterization of porcine HEV from tissue samples derived from wild boar and domestic pigs: 222 wild boars hunted in Northern Germany were investigated for the presence of HEV RNA with a detection rate of 5.9%. Three additional HEV‐positive wild boar liver samples as well as an HEV‐positive spleen and a positive kidney from domestic pigs were included. After inoculation of positive samples onto the human hepatoma cell line PLC/PRF/5, cells were grown for several weeks. Successful isolation was confirmed by RT‐qPCR, virus passage, immunofluorescence staining and titration. Overall, 15 strains from a total of 18 RNA‐positive organ samples could be obtained and viral loads >109 RNA copies/ml were measured in cell culture supernatants. Accordingly, 83.3% of the HEV RNA‐positive samples contained infectious hepatitis E viral particles and therefore must be considered as a potential source for human infections. Phylogenetic analyses revealed that all isolated strains belong to genotype 3. Further genetic characterization showed a high degree of sequence variability, but no sequence insertions, in the hypervariable region within the open reading frame 1. [ABSTRACT FROM AUTHOR]

Current meat production places high costs on the environment. However, only a small portion of consumers are willing to opt for meat substitutes or a vegetarian diet. Cultured meat may contribute to solve this dilemma. In this journal, Bryant and Barnett recently reviewed current attitude research and summarized objections perceived by consumers concerning cultured meat. However, no research from Germany was available. Thus, we conducted a survey of German participants, including attitudes previously found to be important in the literature. With a panel sample of 713 consumers, attitudes were found to structure in three dimensions: ethics (e.g., animal welfare, ecological) was the strongest positive driver and depended on pre-knowledge available for 38% of participants; emotional objections (e.g., unnatural) were the second strongest predictor but unrelated to pre-knowledge and demographics; and the third attitudinal dimension expresses concern over the global diffusion of cultured meat. A path model summarizes the results. In conclusion, Germany shows itself to be only moderately prepared to accept cultured meat., Competing Interests: Declaration of Competing Interest The authors declare that there are no conflicts of interests., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Molecular and cellular research modalities for the study of liver pathologies have been tremendously improved over the recent decades. Advanced technologies offer novel opportunities to establish cell isolation techniques with excellent purity, paving the path for 2D and 3D microscopy and high-throughput assays (e.g., bulk or single-cell RNA sequencing). The use of stem cell and organoid research will help to decipher the pathophysiology of liver diseases and the interaction between various parenchymal and non-parenchymal liver cells. Furthermore, sophisticated animal models of liver disease allow for the in vivo assessment of fibrogenesis, portal hypertension and hepatocellular carcinoma (HCC) and for the preclinical testing of therapeutic strategies. The purpose of this review is to portray in detail novel in vitro and in vivo methods for the study of liver cell biology that had been presented at the workshop of the 8th meeting of the European Club for Liver Cell Biology (ECLCB-8) in October of 2018 in Bonn, Germany., Competing Interests: The authors have no conflicts of interest.

Over the last 20 years, the highly interdisciplinary field of tissue engineering (TE) has become an established subspecialty in research facilities all over the world. Numerous methods and protocols are available for various research intentions and aims, but there are no data indicating which of these methods and resources are generally used. This study is an overview of the resources and methods that are commonly applied in TE research in general, and in the field of oral and maxillofacial surgery (OMFS) in Germany, Austria and Switzerland. The DÖSAK collaborative group for TE developed a detailed questionnaire and collected information from participating university hospitals in these three countries. We evaluated the availability of research facilities, in vitro realisation and in vivo designs for animal studies in these departments. 11 units who replied, out of 35 we contacted, conducted research on bone regeneration in interdisciplinary research facilities. 10 departments used xenogeneic and alloplastic scaffolds for in vitro and in vivo applications. In this case, the most commonly utilised trademarks were Bio-Oss(®) and CERASORB(®). 9 units used osteoblasts (73%) and 10 proliferation assays in vitro, whereas rats served as the standard animal model for histology/immunohistochemistry in 6. All research units were interested in establishing a platform for research exchange and communication. This study shows that tissue engineering is well established and highly accepted in most participating university hospitals and research facilities. The presented data, together with data published in a foregoing paper will help arrange more readily available standardised procedures for further investigations., (Copyright © 2014 European Association for Cranio-Maxillo-Facial Surgery. Published by Elsevier Ltd. All rights reserved.)

A short description of the history of the 3Rs concept is given, which was developed as the scientific concept to refine, reduce and replace animal experiments by Russel and Burch more than 40 years ago. In addition, the legal framework in Europe for developing alternatives to animal experiments is given and the current status of in vitro systems in pharmacology and toxicology is described including an update on metabolising systems. The decrease in experimental animal numbers during the past decade in Europe is illustrated by the situation in Germany and the contribution of international harmonisation of test guidelines on reducing animal numbers in regulatory testing is described. A review of the development of the principles of experimental validation is given and the 3T3 NRU in vitro phototoxicity test is used as an example for a successful validation study, which led to the acceptance of the first in vitro toxicity test for regulatory purposes by the OECD. Finally, the currently accepted alternative methods for standardisation and safety testing of drugs, biologicals and medical devices are summarised.

We have compared in extracts of activated sludge the number of enteroviruses detectable with buffalo green monkey (BGM) cell-cultures versus the number of enteroviral genomes determined by reverse-transcription quantitative real-time PCR (RT-qPCR). In order to find conditions adequate for quantifying enteroviral RNA isolated from (waste)water we have investigated affinity capture of RNA with polystyrene beads (Dynabeads). The capture efficiency strongly depended on the genomic region chosen for the affinity binding. Capture of the RNA by its 3'-tail was most efficient (almost 100%); other regions within the genome yielded variable but lower results. Indirect capture (first hybridization of the RNA to the oligonucleotides, then attachment of the duplex molecules to the beads) was much more efficient than direct capture (attachment of the oligonucleotides to the beads first, then binding of the RNA), and resulted in RNA capture of maximally 60-80%. At least partly, this was due to incomplete hybridization of the RNA to the complementary oligonucleotides. No correlation was found between the number of cytopathic effects (CPE) determined by cell culture and the number of genomes quantified by RT-qPCR; RT-qPCR values were consistently much higher than the number of CPE. This points to overestimation of infectious enteroviruses by RT-qPCR and/or underestimation by the cell culture approach.

The degradation of chlorobenzene was investigated with the specially chosen strain Methylocystis sp. GB 14 DSM 12955, using 23 ml headspace vials and in a soil column filled with quaternary aquifer material from a depth of 20 m. A long-term experiment was carried out in this column, situated in a mobile test unit at a contaminated location in Bitterfeld (Germany). Groundwater polluted by chlorobenzene was continuously fed through the column, through which a mixture comprising 4% CH(4) and 96% air was bubbled. Chlorobenzene was oxidized by up to 80% under pure culture conditions in the model experiments and was completely degraded under the mixed culture conditions of the column experiments. Over a period of 4 months, the stability of the biological system was monitored regularly by analyzing the sMMO activity as well as by classical microbiological and molecular biological methods.

Background: Cytokines play an important role in mediating immunoglobulin switch, the secretion of protective mucosal immunoglobulins, and the development of allergic diseases. This study investigates whether B cells from allergic and healthy children have different capacities to secrete immunoglobulins after stimulation with IL-4, IL-6, IL-10, IL-11, and IL13., Methods: We analyzed the peripheral venous blood of 44 healthy probands and of 109 allergic patients with a mean age of 13 years, allergic to grass pollen, birch pollen, and house dust mites. Lymphocytes were isolated by a density gradient and B cells were enriched by using a Magnetic Activated Cell Separator (MACS) and anti-CD19 microbeads. B Cells were co-cultured with human CDw32 (Fc gammaRII) expressing mouse Ltk fibroblasts and mouse anti-human CD40 monoclonal antibodies (CD40 system). The interleukins IL-4, IL-6, IL-10, IL-11, and IL-13 were supplemented in various combinations. After 14 days, concentrations of IgE, IgG, IgA, and IgM were measured in the supernatants with ELISA., Results: Suppression of IgA-, IgG, and IgM- synthesis was induced by stimulation of B cells with IL-4. After additional application of IL-10, IgA, IgG, and IgM synthesis was significantly increased. When cultures stimulated with IL-4 were additionally supplemented with IL-10, IgA, and IgG synthesis of B cells obtained from allergic individuals was significantly decreased compared to nonallergic individuals. IgE-secretion of B cells from allergic individuals was significantly increased compared to nonallergic individuals after stimulation with IL-4., Conclusion: Our results implicate that IL-4 is essential for the regulation of immunoglobulin class switch to IgE and that IL-4 is an important cytokine for the development of allergic diseases. The capacity of B cells in allergic children to produce less IgA and IgG in response to additional stimulation with IL-10 of cultures supplemented with IL-4 could play an important role in mediating a mucosal immune system vulnerable to allergens. This phenomenon could contribute to the pathogenesis of allergic diseases.

Unlabelled: From December 1990 to November 1993 nasopharyngeal specimens were obtained for culture from 50 children (mean 4.9 +/- 3.3 months of age) who had died suddenly. Bordetella pertussis was not isolated. Subsequently, nasopharyngeal specimens for polymerase chain reaction (PCR) analysis were obtained from another 51 victims of sudden death (mean 5.4 +/- 4.4 months of age); nine (18%) were B. pertussis positive., Conclusion: Our findings support previous epidemiological studies which noted an association between epidemic pertussis and sudden infant death syndrome. Further PCR studies with both internal and external controls should be performed.