Comparison of different in situ hybridization techniques for the detection of various RNA and DNA viruses

In situ hybridization (ISH) is a technique to determine potential correlations between viruses and lesions. The aim of the study was to compare ISH techniques for the detection of various viruses in different tissues. Tested RNA viruses include atypical porcine pestivirus (APPV) in the cerebellum of pigs, equine and bovine hepacivirus (EqHV, BovHepV) in the liver of horses and cattle, respectively, and Schmallenberg virus (SBV) in the cerebrum of goats. Examined DNA viruses comprise canine bocavirus 2 (CBoV-2) in the intestine of dogs, porcine bocavirus (PBoV) in the spinal cord of pigs and porcine circovirus 2 (PCV-2) in cerebrum, lymph node, and lung of pigs. ISH with self-designed digoxigenin-labelled RNA probe

Presence of bovine hepacivirus in Turkish cattle

Hepatitis C virus (HCV), a worldwide distributed human pathogen, causes one of the most important viral infections in human being. HCV is the type species of the genus Hepacivirus (Flaviviridae) in which recently discovered animal viruses i.e. from horses, bats, rodents and cattle are allocated. After preliminary reports in 2015 from German and African cattle, a wide distribution of bovine hepacivirus (BovHepV, Hepacivirus N) was proposed. We investigated the possible presence of BovHepV in serum samples from cattle in different locations of Turkey. Analyzing a total of 120 samples from 98 female (dairy) and 22 male (beef) cattle by real-time RT-PCR resulted in 15 (12.5%) positives. BovHepV infection was detected in 6 out of 10 locations included in the study. There were positive samples both from eastern and western parts of the country indicating possible wide distribution in the Turkish cattle population. Phylogenetic analysis of 9 selected positive samples clearly assigned 8 sequences to a separate cluster on the basis of NS3 gene region, while one of the sequences obtained from an imported animal from north of Italy grouped with sequences obtained from cattle in Germany. The latter finding may indicate possible occurrence of this genetic group of BovHepV not only in Germany but in other European countries. Results of the present study demonstrate the presence of BovHepV infections in Turkey and in The Middle East region.Deutscher Akademischer Austausch Dienst (DAAD)German Center for Infection Research (DZIF) (Germany) - 800280180

Molecular detection of hepatitis E virus in German domestic pigs

In industrial countries, Hepatitis E virus (HEV) transmission to humans is predominantly assumed to be a zoonotic infection. Recently, it has been demonstrated that about 50% of domestic pigs in Germany carry HEV-specific antibodies. However, further investigations concerning the distribution of HEV in different age groups of German domestic pigs, phylogenetic analyses and the viral load in the porcine liver are still pending. Liver samples of all age groups from herds in a pig-dense region in north-western Germany were investigated for the presence and quantity of HEV RNA and subsequently genotyped. Out of 251 liver samples, 34 contained ORF2-specific RNA, whereas 19 samples were positive using ORF1-specific primers, resulting in an overall detection rate of 13.5% and 7.6%, respectively. Especially nursery pigs and growers were tested positive for viral RNA. Furthermore, determination of the HEV copy numbers revealed high replication levels. Up to 10(9) genome copies per g of liver tissue could be detected suggesting a likely high degree of viral spread to the environment. In the HEV-positive liver samples we found no hints for pathohistological changes reflecting the HEV status.The HEV sequences showed marked diversity but could be assigned to HEV genotype 3 without exception. However, by comparing two different genomic fragments, we found indications for infections with two different HEV variants in domestic pigs. Apart from this, the current study confirms the outcome of our recent serological HEV survey and for the first time gives direct proof of HEV infections in the German domestic pig population

Expanded Diversity and Host Range of Bovine Hepacivirus—Genomic and Serological Evidence in Domestic and Wild Ruminant Species

The hepatitis C virus (HCV)-related bovine hepacivirus (BovHepV) can cause acute as well as persistent infections in cattle. The true clinical relevance of the virus is not yet known. As reliable antibody detection methods are lacking and prevalence studies have only been conducted in cattle and few countries to date, the true distribution, genetic diversity, and host range is probably greatly underestimated. In this study, we applied several RT-PCR methods and a nano-luciferase-based immunoprecipitation system (LIPS) assay to analyze bovine serum samples from Bulgaria as well as wild ruminant sera from Germany and the Czech Republic. Using these methods, BovHepV infections were confirmed in Bulgarian cattle, with viral genomes detected in 6.9% and serological reactions against the BovHepV NS3 helicase domain in 10% of bovine serum samples. Genetic analysis demonstrated co-circulation of highly diverse BovHepV strains in Bulgarian cattle, and three novel BovHepV subtypes within the genotype 1 could be defined. Furthermore, application of a nested RT-PCR led to the first description of a BovHepV variant (genotype 2) in a wild ruminant species. The results of this study significantly enhance our knowledge of BovHepV distribution, genetic diversity, and host range

Pathogen Exposure in White Whales (<i>Delphinapterus leucas</i>) in Svalbard, Norway

The Svalbard white whale (Delphinapterus leucas) population is one of the smallest in the world, making it particularly vulnerable to challenges such as climate change and pathogens. In this study, serum samples from live captured (2001–2016) white whales from this region were investigated for influenza A virus (IAV) antibodies (Abs) (n = 27) and RNA (n = 25); morbillivirus (MV) Abs (n = 3) and RNA (n = 25); Brucella spp. Abs; and Toxoplasma gondii Abs (n = 27). IAV Abs were found in a single adult male that was captured in Van Mijenfjorden in 2001, although no IAV RNA was detected. Brucella spp. Abs were found in 59% of the sample group (16/27). All MV and T. gondii results were negative. The results show that Svalbard white whales have been exposed to IAV and Brucella spp., although evidence of disease is lacking. However, dramatic changes in climate and marine ecosystems are taking place in the Arctic, so surveillance of health parameters, including pathogens, is critical for tracking changes in the status of this vulnerable population

Pathological Findings in Eurasian Otters (Lutra lutra) Found Dead between 2015&minus;2020 in Schleswig-Holstein, Germany

In times of massive biodiversity loss and ongoing environmental crises, it is extremely important to ensure long-term conservation efforts of threatened species like Eurasian otters (Lutra lutra). To gain insights into the status of Northern Germany&rsquo;s otter population, 92 otters found dead in Schleswig-Holstein between 2015&minus;2020 were collected and underwent detailed dissection with the aim to establish a monitoring program for this population. Examinations followed a protocol especially designed for otters, including various biological data assessments and extended sampling. The finding sites showed a clear concentration in the Continental region. Seasonal concentration differed among the years, yet peaks were seen from fall to winter. Overall, more males than females were found, although this differed among the years. The majority of otters that could be aged were between 1&ndash;3 years. Placental scars and pregnancy were recorded in only few females. Nutritional status was good in most cases. Infectious diseases found included Vagococcus lutrae, Toxoplasma gondii, and Emmonsia spp. A major cause of death was roadkill. Known sample bias in studies focusing on roadkill was considered in the finding interpretation. Continuation of the population health investigations is mandatory to analyze potential trends and to establish an actual monitoring program for Eurasian otters in Schleswig-Holstein

Hepatitis E virus seroprevalence of domestic pigs in Germany determined by a novel in-house and two reference ELISAs

International audienceAutochthonous hepatitis E virus (HEV) infections by zoonotic transmission of genotype 3 (GT3) have been reported increasingly from industrialized countries. In this paper the development and validation of an IgG ELISA for the detection of HEV-specific antibodies in domestic pigs is described. Comparison of the diagnostic value ofEscherichia coli-expressed HEV-GT3 capsid protein (CP) derivatives revealed a carboxyterminal derivative as most suitable. Validation of the in-house assay using a commercially available IgG ELISA revealed a high diagnostic specificity and sensitivity. The average HEV seroprevalence of domestic pigs from Germany and the federal state Baden-Wuerttemberg determined by the in-house test was 42.7% and 50.3%, respectively. The seroprevalence in different districts of Baden-Wuerttemberg ranged from 34.9% to 60%, but from 0% to 100% between different herds. These data were compared to those achieved by two commercially available ELISA kits and an in-house ratHEV-based ELISA. In conclusion, the CP-based in-house test proved sensitive and specific, indicating that the ORF3-encoded protein might be dispensable for diagnostics. The novel assay also allowed a parallel analysis by a homologous ratHEV-derived antigen. Thus, the novel IgG ELISA represents a useful tool for future standardized seroprevalence studies in domestic pigs from Germany and other regions of Europe. (C) 2013 Elsevier B.V. All rights reserved

Screening for Influenza and Morbillivirus in Seals and Porpoises in the Baltic and North Sea

Bonus BaltHealt

In the Search of Marine Pestiviruses:First Case of Phocoena Pestivirus in a Belt Sea Harbour Porpoise

Pestiviruses are widespread pathogens causing severe acute and chronic diseases among terrestrial mammals. Recently, Phocoena pestivirus (PhoPeV) was described in harbour porpoises (Phocoena phocoena) of the North Sea, expanding the host range to marine mammals. While the role of the virus is unknown, intrauterine infections with the most closely related pestiviruses— Bungowannah pestivirus (BuPV) and Linda virus (LindaV)—can cause increased rates of abortions and deaths in young piglets. Such diseases could severely impact already vulnerable harbour porpoise populations. Here, we investigated the presence of PhoPeV in 77 harbour porpoises, 277 harbour seals (Phoca vitulina), grey seals (Halichoerus grypus) and ringed seals (Pusa hispida) collected in the Baltic Sea region between 2002 and 2019. The full genome sequence of a pestivirus was obtained from a juvenile female porpoise collected along the coast of Zealand in Denmark in 2011. The comparative Bayesian phylogenetic analyses revealed a close relationship between the new PhoPeV sequence and previously published North Sea sequences with a recent divergence from genotype 1 sequences between 2005 and 2009. Our findings provide further insight into the circulation of PhoPeV and expand the distribution from the North Sea to the Baltic Sea region with possible implications for the vulnerable Belt Sea and endangered Baltic Proper harbour porpoise populations