Friedrich Loeffler Institute FLI

Greifswald, Germany

Friedrich Loeffler Institute FLI

Greifswald, Germany

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Adlhoch C.,Centers for Disease Control and Prevention | Gossner C.,Centers for Disease Control and Prevention | Gossner C.,Maastricht University | Koch G.,Wageningen University | And 5 more authors.
Eurosurveillance | Year: 2014

Since the beginning of November 2014, nine outbreaks of highly pathogenic avian influenza virus (HPAIV) A(H5N8) in poultry have been detected in four European countries. In this report, similarities and differences between the modes of introduction of HPAIV A(H5N1) and A(H5N8) into Europe are described. Experiences from outbreaks of A(H5N1) in Europe demonstrated that early detection to control HPAIV in poultry has proven pivotal to minimise the risk of zoonotic transmission and prevention of human cases. © 2014 European Centre for Disease Prevention and Control (ECDC). All rights reserved.

Seitz C.,Max Planck Institute for Dynamics of Complex Technical Systems | Frensing T.,Max Planck Institute for Dynamics of Complex Technical Systems | Hoper D.,Friedrich Loeffler Institute FLI | Kochs G.,Albert Ludwigs University of Freiburg | And 2 more authors.
Journal of General Virology | Year: 2010

Because of their high susceptibility to infection with various influenza virus strains, Madin-Darby canine kidney (MDCK) cells have been widely used as a substrate for influenza virus isolation and vaccine production. However, MDCK cells are also interferon (IFN) competent, and the type I IFN response is commonly thought to be a factor strongly inhibiting virus replication. Therefore, the inhibition of influenza virus replication by IFN signalling was analysed for an adherent MDCK cell line used in vaccine manufacturing. Depending on the respective virus strain, different levels of IFN induction and a corresponding upregulation of the IFN-induced myxovirus resistance protein 1 (Mx1) were observed. Suppression of IFN induction by transient expression of the viral non-structural protein 1 protein enhanced replication of an influenza virus lacking NS1, but not wild-type strains. In agreement with this, stimulation of cells with MDCK cell-derived IFN prior to infection resulted only in a decrease in replication rate, and not in a change of final yields for wild-type influenza viruses. This lack of IFN-induced antiviral activity correlated with missing anti-influenza activity of MDCK Mx proteins. No inhibitory effect on viral polymerase activity was found for canine Mx1 (cMx1) and cMx2 in minireplicon assays. In conclusion, in MDCK cells, IFN expression is not a limiting factor for influenza virus replication and this might partially be caused by a lack of anti-influenza activity of canine Mx proteins. © 2010 SGM.

Roedig J.V.,Max Planck Institute for Dynamics of Complex Technical Systems | Rapp E.,Max Planck Institute for Dynamics of Complex Technical Systems | Hoper D.,Friedrich Loeffler Institute FLI | Genzel Y.,Max Planck Institute for Dynamics of Complex Technical Systems | And 2 more authors.
PLoS ONE | Year: 2011

The genome of influenza A viruses is constantly changing (genetic drift) resulting in small, gradual changes in viral proteins. Alterations within antibody recognition sites of the viral membrane glycoproteins hemagglutinin (HA) and neuraminidase (NA) result in an antigenetic drift, which requires the seasonal update of human influenza virus vaccines. Generally, virus adaptation is necessary to obtain sufficiently high virus yields in cell culture-derived vaccine manufacturing. In this study detailed HA N-glycosylation pattern analysis was combined with in-depth pyrosequencing analysis of the virus genomic RNA. Forward and backward adaptation from Madin-Darby Canine Kidney (MDCK) cells to African green monkey kidney (Vero) cells was investigated for two closely related influenza A virus PR/8/34 (H1N1) strains: from the National Institute for Biological Standards and Control (NIBSC) or the Robert Koch Institute (RKI). Furthermore, stability of HA N-glycosylation patterns over ten consecutive passages and different harvest time points is demonstrated. Adaptation to Vero cells finally allowed efficient influenza A virus replication in Vero cells. In contrast, during back-adaptation the virus replicated well from the very beginning. HA N-glycosylation patterns were cell line dependent and stabilized fast within one (NIBSC-derived virus) or two (RKI-derived virus) successive passages during adaptation processes. However, during adaptation new virus variants were detected. These variants carried "rescue" mutations on the genomic level within the HA stem region, which result in amino acid substitutions. These substitutions finally allowed sufficient virus replication in the new host system. According to adaptation pressure the composition of the virus populations varied. In Vero cells a selection for "rescue" variants was characteristic. After back-adaptation to MDCK cells some variants persisted at indifferent frequencies, others slowly diminished and even dropped below the detection limit. © 2011 Roedig et al.

Kronefeld M.,Friedrich Loeffler Institute FLI | Werner D.,Leibniz Center for Agricultural Landscape Research | Kampen H.,Friedrich Loeffler Institute FLI
Parasitology Research | Year: 2014

Based primarily on nucleotide polymorphisms in the internal transcribed spacer 2 (ITS2) of the ribosomal DNA, Anopheles daciae was recently described as an additional member of the Maculipennis Group of species, separate from Anopheles messeae with which it had previously been confused due to morphological and genetic similarity. Species differentiation between A. messeae and A. daciae was possible only by ITS2 polymerase chain reaction (PCR) amplification followed by DNA sequencing or RFLP analysis. In addition to its siblings, Anopheles maculipennis, Anopheles atroparvus and A. messeae, A. daciae has been shown to occur in Germany, although with limited distribution. We here describe additional collection sites for this species in Germany, showing concentrations in East Germany and the northern Upper Rhine Valley in Southwest Germany. A species-specific multiplex PCR assay is presented that is able to differentiate the four Maculipennis Group sibling species occurring in Germany plus Anopheles sacharovi, Anopheles melanoon and Anopheles labranchiae. The correct identification and detailed knowledge of the biology of A. daciae are of relevance since it might be a vector of disease agents, as suggested by the vector potential of its siblings and the recent finding of an A. daciae female infected with Dirofilaria repens in southern Germany. © 2014 Springer-Verlag.

Lehmann K.,Humboldt University of Berlin | Werner D.,Leibniz Center for Agricultural Landscape Research | Hoffmann B.,Friedrich Loeffler Institute FLI | Kampen H.,Friedrich Loeffler Institute FLI
Parasites and Vectors | Year: 2012

Background: Biting midges of the Obsoletus species complex of the ceratopogonid genus Culicoides were assumed to be the major vectors of bluetongue virus (BTV) in northern and central Europe during the 2006 outbreak of bluetongue disease (BT). Most recently, field specimens of the same group of species have also been shown to be infected with the newly emerged Schmallenberg virus (SBV) in Europe. A reliable identification of the cryptic species of this group is fundamental for both understanding the epidemiology of the diseases and for targeted vector control. In the absence of classical morphological characters unambiguously identifying the species, DNA sequence-based tests have been established for the distinction of selected species in some parts of Europe. Since specificity and sensitivity of these tests have been shown to be in need of improvement, an alternative PCR assay targeting the mitochondrial cytochrome oxidase subunit I (COI) gene was developed for the identification of the three Obsoletus complex species endemic to Germany (C. obsoletus, C. scoticus, C. chiopterus) plus the isomorphic species C. dewulfi. Methods. Biting midges of the genus Culicoides caught by UV light traps all over Germany were morphologically pre-identified to species or complex level. The COI region was amplified from their extracted DNA and sequenced. Final species assignment was done by sequence comparison to GenBank entries and to morphologically identified males. Species-specific consensus sequences were aligned and polymorphisms were utilized to design species-specific primers to PCR-identify specimens when combined with a universal primer. Results: The newly developed multiplex PCR assay was successfully tested on genetically defined Obsoletus complex material as well as on morphologically pre-identified field material. The intended major advantage of the assay as compared to other PCR approaches, namely the production of only one single characteristic band for each species, could be realized with high specificity and sensitivity. Conclusion: To elucidate the biological characteristics of potential vectors of disease agents, such as ecology, behaviour and vector competence, and the role of these haematophagous arthropods in the epidemiology of the diseases, simple, cost-effective and, most importantly, reliable identification techniques are necessary. The PCR assay presented will help to identify culicoid vector species and therefore add to bluetongue and Schmallenberg disease research including vector control and monitoring. © 2012 Lehmann et al.; licensee BioMed Central Ltd.

Wahli T.,University of Bern | Bergmann S.M.,Friedrich Loeffler Institute FLI
CAB Reviews: Perspectives in Agriculture, Veterinary Science, Nutrition and Natural Resources | Year: 2011

Viral haemorrhagic septicaemia (VHS) is considered to be one of the most economically important viral fish diseases. This review aims to summarize knowledge on the present distribution of VHS, advances in development of diagnostic methods and implications for resulting interventions against the disease. The disease was originally found as a problem in rainbow trout in European freshwater fish farms. However, the first detection of VHS virus (VHSV) in various marine species in the 1980s and 1990s initiated intensive investigation that have demonstrated the presence of the causative rhabdovirus in a broad range of marine and freshwater species of the Northern Hemisphere including the Atlantic and Pacific Coast of North America and the marine waters around Japan and South Korea. Recent mass mortalities in the Great Lake System of North America have also been attributed to VHSV. The number of known affected and susceptible fish species is steadily growing. At least three serotypes of the virus could be differentiated, which, however, do not allow discrimination between marine and freshwater isolates. Development of molecular methods allowed analysis of sequences of viral genes, particularly of the N, G and NV genes. These analyses revealed four major genotypes, which are divided into different sublineages. The genotypes appear to be more related to geographical than to species origin. The wide distribution of the virus, particularly in wild marine fish populations, requires new methods for the identification of genotypes, especially molecular tools and the investigation of the virulence of the different genotypes to potential aquacultural hosts in marine and freshwater. Further, it raises questions concerning actions to be taken against the disease and success of the initiated actions, particularly in the light of an apparently high adaptability of the virus to new environmental conditions and changes in virulence for certain fish species. These characteristics of the virus mean that taking different actions according to genotype is at least a questionable approach. © CAB International 2011.

Hubner K.,Friedrich Loeffler Institute FLI | Hubner K.,University of Melbourne | Phi-van L.,Friedrich Loeffler Institute FLI
Biochimica et Biophysica Acta - Gene Regulatory Mechanisms | Year: 2010

The 5′-flanking region of the chicken glioma-amplified sequence (GAS) 41 gene is close to the 3′ end of the lysozyme gene and contains no typical TATA box, but several GC boxes. In this study, we have localized the GAS 41 promoter to this narrow region. Electrophoretic mobility shift assays and chromatin immunoprecipitation analyses revealed that Sp1 and Sp3 bind to this promoter. Mapping by a technique of indirect end labeling demonstrated that the Sp1-binding sites contained in this region exactly co-map with two previously identified DNase I hypersensitive (HS) sites, which suggests the important role of Sp1 binding in maintaining an open chromatin structure of the GAS41 promoter. We further found that Sp1 and Sp3 strongly activate CAT expression controlled by the putative GAS41 promoter in Drosophila Schneider S2 cells and that deletion of the Sp1 sites resulted in a loss of promoter activity in chicken HD11 cells. The results indicate that transcription factors of the Sp family play an important role in the transcriptional regulation of the chicken GAS41 gene. © 2010 Elsevier B.V.

Wernike K.,Institute of Diagnostic Virology | Breithaupt A.,Friedrich Loeffler Institute FLI | Keller M.,Institute of Novel and Emerging Infectious Diseases | Hoffmann B.,Institute of Diagnostic Virology | And 2 more authors.
PLoS ONE | Year: 2012

Schmallenberg virus (SBV), a novel orthobunyavirus, was discovered in Europe in late 2011. It causes mild and transient disease in adult ruminants, but fetal infection can lead to abortion or severe malformations. There is considerable demand for SBV research, but in vivo studies in large animals are complicated by their long gestation periods and the cost of high containment housing. The goal of this study was to investigate whether type I interferon receptor knock-out (IFNAR-/-) mice are a suitable small animal model for SBV. Twenty IFNAR-/- mice were inoculated with SBV, four were kept as controls. After inoculation, all were observed and weighed daily; two mice per day were sacrificed and blood, brain, lungs, liver, spleen, and intestine were harvested. All but one inoculated mouse lost weight, and two mice died spontaneously at the end of the first week, while another two had to be euthanized. Real-time RT-PCR detected large amounts of SBV RNA in all dead or sick mice; the controls were healthy and PCR-negative. IFNAR-/- mice are susceptible to SBV infection and can develop fatal disease, making them a handy and versatile tool for SBV vaccine research. © 2012 Wernike et al.

Pietsch C.,University of Basel | Kersten S.,Friedrich Loeffler Institute FLI | Burkhardt-Holm P.,University of Basel | Valenta H.,Friedrich Loeffler Institute FLI | Danicke S.,Friedrich Loeffler Institute FLI
Toxins | Year: 2013

The control of mycotoxins is a global challenge not only in human consumption but also in nutrition of farm animals including aquatic species. Fusarium toxins, such as deoxynivalenol (DON) and zearalenone (ZEN), are common contaminants of animal feed but no study reported the occurrence of both mycotoxins in fish feed so far. Here, we report for the first time the occurrence of DON and ZEN in samples of commercial fish feed designed for nutrition of cyprinids collected from central Europe. A maximal DON concentration of 825 μg kg-1 feed was found in one feed whereas average values of 289 μg kg-1 feed were noted. ZEN was the more prevalent mycotoxin but the concentrations were lower showing an average level of 67.9 μg kg-1 feed. © 2013 by the authors; licensee MDPI, Basel, Switzerland.

Brim S.,University of Munster | Groschup M.H.,Friedrich Loeffler Institute FLI | Kuczius T.,University of Munster
PLoS ONE | Year: 2016

Prion diseases are characterized biochemically by protein aggregation of infectious prion isoforms (PrPSc), which result from the conformational conversion of physiological prion proteins (PrPC ). PrPC are variable post- Translationally modified glycoproteins, which exist as full length and as aminoterminally truncated glycosylated proteins and which exhibit differential detergent solubility. This implicates the presence of heterogeneous phenotypes, which overlap as protein complexes at the same molecular masses. Although the biological function of PrPC is still enigmatic, evidence reveals that PrPC exhibits metal-binding properties, which result in structural changes and decreased solubility. In this study, we analyzed the yield of PrPC metal binding affiliated with low solubility and changes in protein banding patterns. By implementing a high-speed centrifugation step, the interaction of zinc ions with PrPC was shown to generate large quantities of proteins with low solubility, consisting mainly of full-length glycosylated PrPC whereas unglycosylated PrPC remained in the supernatants as well as truncated glycosylated proteins which lack of octarepeat sequence necessary for metal binding. This effect was considerably lower when PrPC interacted with copper ions the presence of other metals tested exhibited no effect under these conditions. The binding of zinc and copper to PrPC demonstrated differentially soluble protein yields within distinct PrPC subtypes. PrPC -Zn2+-interaction may provide a means to differentiate glycosylated and unglycosylated subtypes and offers detailed analysis of metal-bound and metal-free protein conversion assays. © 2016 Brim et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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