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Swine influenza A viruses (swIAVs) can play a crucial role in the generation of new human pandemic viruses. In this study, in-depth passive surveillance comprising nearly 2,500 European swine holdings and more than 18,000 individual samples identified a year-round presence of up to four major swIAV lineages on more than 50% of farms surveilled. Phylogenetic analyses show that intensive reassortment with human pandemic A(H1N1)/2009 (H1pdm) virus produced an expanding and novel repertoire of at least 31 distinct swIAV genotypes and 12 distinct hemagglutinin/neuraminidase combinations with largely unknown consequences for virulence and host tropism. Several viral isolates were resistant to the human antiviral MxA protein, a prerequisite for zoonotic transmission and stable introduction into human populations. A pronounced antigenic variation was noted in swIAV, and several H1pdm lineages antigenically distinct from current seasonal human H1pdm co-circulate in swine. Thus, European swine populations represent reservoirs for emerging IAV strains with zoonotic and, possibly, pre-pandemic potential., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The outbreak of highly pathogenic avian influenza H5Nx viruses in winter 2016/2017 was the most severe HPAI epizootic ever reported in Germany. The H5N8 and H5N5 viruses detected in birds in Germany in 2016/2017 represent a reassortant swarm of at least five distinct genotypes, which carried closely related HA segments derived from clade 2.3.4.4b. The genotypes of these viruses and their spatio-temporal distribution indicated a unique situation with multiple independent entries of HPAIV into Germany.

In November 2016, an influenza A(H5N8) outbreak caused deaths of wild birds and domestic poultry in Germany. Clade 2.3.4.4 virus was closely related to viruses detected at the Russia-Mongolia border in 2016 but had new polymerase acidic and nucleoprotein segments. These new strains may be more efficiently transmitted to and shed by birds.

Highly pathogenic avian influenza (H5N8) virus, like the recently described H5N8 strain from Korea, was detected in November 2014 in farmed turkeys and in a healthy common teal (Anas crecca) in northeastern Germany. Infected wild birds possibly introduced this virus.

Oral emergency vaccination against classical swine fever is a powerful tool to control disease outbreaks among European wild boar and thus to safeguard domestic pigs in affected regions. In the past, when virus detection was mainly done using virus isolation in cell culture or antigen enzyme-linked immunosorbent assays, modified live vaccine strains like C-strain "Riems", were barely detectable after oral vaccination campaigns. Nowadays, the use of highly sensitive molecular techniques has given rise to an increase in vaccine virus detections. This was also the case during the 2009 outbreak among German wild boar and the subsequent vaccination campaigns. To guarantee a rapid differentiation of truly infected from C-strain vaccinated animals, a combination of differentiating multiplex rRT-PCR assays with partial sequencing was implemented. Here, we report on the rational and use of this approach and the lessons learned during execution. It was shown that positive results in the recently developed vaccine strain (genotype) specific rRT-PCR assay can be taken as almost evidentiary whereas negative results should be confirmed by partial sequencing. Thus, combination of multiplex rRT-PCR assays as a first line differentiation with partial sequencing can be recommended for a genetic DIVA strategy in areas with oral vaccination against classical swine fever in wild boars., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Beginning in 2002, a small number of pig farms in western Canada began reporting 4-7-week-old pigs with bilateral hind-end paresis or paralysis. Low numbers of pigs were affected, some died, most had to be euthanized, and those that survived had reduced weight gains and neurological deficits. Necropsies revealed no gross lesions, but microscopic lesions consisted of a nonsuppurative polioencephalomyelitis, most severe in the brain stem and spinal cord. The lesions were most consistent with a viral infection. Tests for circovirus, Porcine reproductive and respiratory syndrome virus, coronavirus, Rabies virus, and Pseudorabies virus were negative. Using immunohistochemistry, virus neutralization, fluorescent antibody test, and nested reverse transcription polymerase chain reaction, Porcine teschovirus was identified in tissues from affected individuals. To the authors' knowledge, this is the first report of teschovirus encephalitis in western Canada and the first reported case of polioencephalomyelitis in pigs in Canada, where teschovirus was confirmed as the cause.

Recurrent outbreaks of H5N1 HPAIV occurred in several Central European countries in 2007. In-depth phylogenetic analyses which included full-length genomic sequences of the viruses involved were performed to elucidate possible origins of incursions and transmission pathways. Tree reconstructions as well as host-shift and ancestral area inferences were conducted in a maximum likelihood framework. All viruses belonged to a separate subgroup (termed "EMA-3") within clade 2.2, and, thus, were distinct from two lineages of HPAIV H5N1 viruses (termed "EMA-1" and "EMA-2") present in the same geographic area in 2006. Analysis of concatenated coding regions of all eight genome segments significantly improved resolution and robustness of the reconstructed phylogenies as compared to single gene analyses. At the same time, the methodological limits to establish retrospectively transmission networks in a comparatively small geographic region and spanning a short period of time became evident when only few corroborating field-epidemiological data are available. Ambiguities remained concerning the origin of the EMA-3 viruses from a region covering Southeast Germany and the Czech Republic as well as routes of spread to other European countries. AIV monitoring programmes in place for wild birds and poultry in these countries did not reveal presence of these viruses in either population. Host switches between domestic poultry and wild bird populations occurred several times. Analysis of outbreaks in Northeast Germany and nearby Northern Poland in December 2007 demonstrated that geographic and even temporal vicinity of outbreaks does not necessarily indicate a common source of incursion., (Copyright 2010 Elsevier B.V. All rights reserved.)

Reemerging bluetongue virus serotype 8 (BTV-8) in Germany was detected first in May 2007 in a sentinel cow and in February 2008 in an export heifer. Reemergence was confirmed by retesting the samples, experimental inoculation, fingerprinting analysis, and virus isolation. Overwintering of BTV-8 and continuous low-level infections are assumed.

The aim of the study was to evaluate a real-time reverse transcription-polymerase chain reaction (RT-PCR) assay for routine diagnosis of classical swine fever (CSF) in wild boar by using 1000 spleen homogenates that were tested previously negative by virus isolation and 26 homogenates from which CSF virus could be isolated. All 26 positive samples in the virus isolation assay also were found to be positive in real-time RT-PCR. Additionally further 10 samples were detected by real-time RT-PCR out of the 1000 negative samples in the virus isolation. With a commercial CSF antigen-ELISA only 14 out of the 36 real-time RT-PCR positive samples could be detected. The sequence analysis of all ten samples that tested positive by real-time RT-PCR and negative by virus isolation revealed CSF virus-specific sequences. Based on the assumption that all samples with a CSF virus-specific sequence or positive in the virus isolation test originated from truly CSF virus-infected wild boar, the following sensitivity values were calculated as antigen-ELISA, 39%; virus isolation, 72% and real-time RT-PCR, 100%. The use of real-time RT-PCR instead of antigen-ELISA and virus isolation as a routine tool for control and eradication of CSF in wild boar populations is recommended.

In Germany, eleven outbreaks of CSF in domestic pig holdings were reported in 2002. They occurred exclusively in regions where CSF virus circulated in the wild boar population. In ten cases the phylogenetic analysis revealed that the isolates from domestic pigs and wild boar had identical sequences in the 5' non-translated region (5'NTR). However, in one case a subtype was isolated which was slightly different from the virus subtype found in the wild boar population of that region. This case is decribed in detail. The epidemiological significance of different diagnostic methods is discussed, in particular the genetic typing of CSF virus isolates.

The genus Pestivirus within the family Flaviviridae currently consists of four different main species: Classical swine fever virus, Bovine viral diarrhea virus types 1 and 2 and Border disease virus. A fifth tentative species is represented by an isolate from a giraffe. In this study, a completely new pestivirus, isolated from a batch of fetal calf serum that was collected in Brazil, is described. It is proposed that the isolate D32/00&#95;'HoBi' may constitute a novel sixth pestivirus species, because it is genetically, as well as antigenically, markedly different from all other pestiviruses. Based on the entire N(pro)- and E2-encoding sequences, identities of <70 % to all other pestivirus species were determined. Similarly, cross-neutralization and binding studies using antisera and mAbs revealed marked antigenic differences between D32/00&#95;'HoBi' and all other pestiviruses.

A pestivirus has been isolated from brain samples of two lambs suffering from clinical signs of border disease. The two identical isolates were allocated to the "true" border disease virus group concerning their reaction pattern with monoclonal antibodies and the 5'UTR sequence data. Nevertheless, alterations of phenotype and genotype in comparison with references of both BDV-subgroups have been shown. The existence of an infection with border disease virus in the flock has been confirmed by serological studies.

The effectiveness of virus isolation, commercial antigen enzyme-linked immunosorbent assay (ELISA), reverse transcriptase-polymerase chain reaction (RT-PCR), and flow cytometry in detection of a low-virulent classical swine fever virus (CSFV) in blood in the early period of infection was evaluated. Domestic pigs at the age of 6-8 weeks and young wild boars were inoculated with a low-virulent field isolate of CSFV originating from a wild boar. This virus induced serious clinical reaction in only one pig which was naturally infected with Pasteurella multocida. Nine of 13 infected domestic pigs showed viremia. All infected weanling pigs were found viraemic by virus isolation on day 6 post infection (p.i.) but virus-free by RT-PCR. The flow cytometry was apparently not as sensitive as the virus isolation. Two young wild boars infected with the virus were viremic only for the first 2 days p.i. Virus isolation and RT-PCR were of similar sensitivity. Three different commercial antigen ELISAs used were not able to detect viral antigen in any animal.

Virus isolation in the PK-15 cell culture, two commercial antigen ELISAs, reverse transcriptional-polymerase chain reaction (RT-PCR), and flow cytometry have been evaluated to detect viremic pigs in the early period of classical swine fever virus (CSFV) infection. Domestic pigs were experimentally inoculated with the virulent CSFV strain 'Alfort 187' and two field isolates. CSFV isolation and RT-PCR were found to be the most sensitive methods for the detection of highly virulent CSFV in the early period of infection which is characterized by the absence of clinical symptoms. Using antigen ELISAs and flow cytometry CSFV could be detected in infected animals after the onset of clinical signs. After infection with a less virulent CSFV field isolate originating from wild boar, viremic pigs could be identified by direct virus isolation. The reasons for the negative results of the RT-PCR still remain unknown. In conclusion we recommend to modify the procedure (antigen ELISA) for the detection of clinically healthy domestic pigs in accordance with the decision 98/413/EC.