Experimental Infection of Cattle with Highly Pathogenic Avian Influenza Virus (H5N1)

Four calves were experimentally inoculated with highly pathogenic avian influenza virus A/cat/Germany/R606/2006 (H5N1) isolated from a cat in 2006. All calves remained healthy, but several animals shed low amounts of virus, detected by inoculation of nasal swab fluid into embryonated chicken eggs and onto MDCK cells. All calves seroconverted

Exceptional Clinical Resistance and Variable Reservoir Competence

Cowpox virus (CPXV) is a zoonotic virus and endemic in wild rodent populations in Eurasia. Serological surveys in Europe have reported high prevalence in different vole and mouse species. Here, we report on experimental CPXV infections of bank voles (Myodes glareolus) from different evolutionary lineages with a spectrum of CPXV strains. All bank voles, independently of lineage, sex and age, were resistant to clinical signs following CPXV inoculation, and no virus shedding was detected in nasal or buccal swabs. In- contact control animals became only rarely infected. However, depending on the CPXV strain used, inoculated animals seroconverted and viral DNA could be detected preferentially in the upper respiratory tract. The highest antibody titers and virus DNA loads in the lungs were detected after inoculation with two strains from Britain and Finland. We conclude from our experiments that the role of bank voles as an efficient and exclusive CPXV reservoir seems questionable, and that CPXV may be maintained in most regions by other hosts, including other vole species. Further investigations are needed to identify factors that allow and modulate CPXV maintenance in bank voles and other potential reservoirs, which may also influence spill-over infections to accidental hosts

A novel European H5N8 influenza A virus has increased virulence in ducks but low zoonotic potential

We investigated in a unique setup of animal models and a human lung explant culture biological properties, including zoonotic potential, of a representative 2016 highly pathogenic avian influenza virus (HPAIV) H5N8, clade 2.3.4.4 group B (H5N8B), that spread rapidly in a huge and ongoing outbreak series in Europe and caused high mortality in waterfowl and domestic birds. HPAIV H5N8B showed increased virulence with rapid onset of severe disease and mortality in Pekin ducks due to pronounced neuro- and hepatotropism. Cross-species infection was evaluated in mice, ferrets, and in a human lung explant culture model. While the H5N8B isolate was highly virulent for Balb/c mice, virulence and transmissibility were grossly reduced in ferrets, which was mirrored by marginal replication in human lung cultures infected ex vivo. Our data indicate that the 2016 HPAIV H5N8B is avian-adapted with augmented virulence for waterfowl, but has low zoonotic potential. The here tested combination of animal studies with the inoculation of human explants provides a promising future workflow to evaluate zoonotic potential, mammalian replication competence and avian virulence of HPAIV.Peer Reviewe

Assessing the occurrence of the novel zoonotic variegated squirrel bornavirus 1 in captive squirrels in Germany —A prevalence study

The newly described zoonotic variegated squirrel bornavirus 1 (VSBV‐1) in German squirrel holdings has been associated with the death of three private owners and one zoo animal caretaker (confirmed cases). Epidemiological investigations were severely impeded by the general lack of data on holdings of the putative reservoir hosts, the family Sciuridae. To fill this lack of data for detailed epidemiological investigations of the captive squirrel population, a register of private and zoological squirrel holdings was established. The findings show a broad variety of kept species and their frequency distribution. By contacting the different stakeholders via Web‐based social groups and societies, information passed in both directions so that disease awareness could be raised and participants could be recruited for further studies. Cross‐sectional studies revealed a prevalence of VSBV‐1‐positive subpopulations of 0% (95% CI 0%–6.2%) among private squirrel collections and 1.9% (95% CI: 0%–9.9%) among zoos in Germany. The approach presented here can be transferred to other populations of non‐traditional pets, which may be equally difficult to monitor, in the case of an emerging zoonotic infectious disease.Peer Reviewe

Age-Dependent Progression of SARS-CoV-2 Infection in Syrian Hamsters

In late 2019, an outbreak of a severe respiratory disease caused by an emerging coronavirus, SARS-CoV-2, resulted in high morbidity and mortality in infected humans. Complete understanding of COVID-19, the multi-faceted disease caused by SARS-CoV-2, requires suitable small animal models, as does the development and evaluation of vaccines and antivirals. Since age-dependent differences of COVID-19 were identified in humans, we compared the course of SARS-CoV-2 infection in young and aged Syrian hamsters. We show that virus replication in the upper and lower respiratory tract was independent of the age of the animals. However, older hamsters exhibited more pronounced and consistent weight loss. In situ hybridization in the lungs identified viral RNA in bronchial epithelium, alveolar epithelial cells type I and II, and macrophages. Histopathology revealed clear age-dependent differences, with young hamsters launching earlier and stronger immune cell influx than aged hamsters. The latter developed conspicuous alveolar and perivascular edema, indicating vascular leakage. In contrast, we observed rapid lung recovery at day 14 after infection only in young hamsters. We propose that comparative assessment in young versus aged hamsters of SARS-CoV-2 vaccines and treatments may yield valuable information, as this small-animal model appears to mirror age-dependent differences in human patients

Recombinase polymerase amplification assay for rapid detection of lumpy skin disease virus

Background  Lumpy skin disease virus (LSDV) is aCapripoxvirusinfecting cattle and Buffalos. Lumpy skin disease (LSD) leads to significant economic losses due to hide damage, reduction of milk production, mastitis, infertility and mortalities (10 %). Early detection of the virus is crucial to start appropriate outbreak control measures. Veterinarians rely on the presence of the characteristic clinical signs of LSD. Laboratory diagnostics including virus isolation, sequencing and real-time polymerase chain reaction (PCR) are performed at well-equipped laboratories. In this study, a portable, simple, and rapid recombinase polymerase amplification (RPA) assay for the detection of LSDV-genome for the use on farms was developed.  Results  The LSDV RPA assay was performed at 42 °C and detected down to 179 DNA copies/reaction in a maximum of 15 min. Unspecific amplification was observed with neither LSDV-negative samples (n= 12) nor nucleic acid preparations from orf virus, bovine papular stomatitis virus, cowpoxvirus, Peste des petits ruminants and Blue tongue virus (serotypes 1, 6 and 8). The clinical sensitivity of the LSDV RPA assay matched 100 % (n= 22) to real-time PCR results. In addition, the LSDV RPA assay detected sheep and goat poxviruses.  Conclusion  The LSDV RPA assay is a rapid and sensitive test that could be implemented in field or at quarantine stations for the identification of LSDV infected case

Pathogenicity of Highly Pathogenic Avian Influenza Virus (H5N1) in Adult Mute Swans

Adult, healthy mute swans were experimentally infected with highly pathogenic avian influenza virus A/Cygnus cygnus/Germany/R65/2006 subtype H5N1. Immunologically naive birds died, whereas animals with preexisting, naturally acquired avian influenza virus–specific antibodies became infected asymptomatically and shed virus. Adult mute swans are highly susceptible, excrete virus, and can be clinically protected by preexposure immunity

Protection and Virus Shedding of Falcons Vaccinated against Highly Pathogenic Avian Influenza A Virus (H5N1)

Virus shedding by vaccinated birds was markedly reduced

Continuous presence of genetically diverse rustrela virus lineages in yellow-necked field mouse reservoir populations in northeastern Germany.

Rustrela virus (RusV; species Rubivirus strelense, family Matonaviridae) was discovered in different zoo animal species affected by fatal encephalitis. Simultaneous RusV RNA detection in multiple yellow-necked field mice (Apodemus flavicollis) suggested this rodent as a reservoir of RusV. Here, we investigated 1,264 yellow-necked field mice and sympatric other small mammals from different regions in Germany for RusV RNA using an optimized reverse transcription-quantitative polymerase chain reaction (RT-qPCR) protocol and high-throughput sequencing. The investigation resulted in the detection of RusV RNA exclusively in 50 of 396 (12.6 per cent) yellow-necked field mice but absence in other sympatric species. RT-qPCR-determined tissue distribution of RusV RNA revealed the highest viral loads in the central nervous system, with other tissues being only very rarely affected. The histopathological evaluation did not reveal any hints of encephalitis in the brains of infected animals despite the detection of viral RNA in neurons by in situ hybridization (ISH). The positive association between the body mass of yellow-necked field mice and RusV RNA detection suggests a persistent infection. Phylogenetic analysis of partial E1 and full-genome sequences showed a high diversification with at least four RusV lineages (1A-1D) in northeastern Germany. Moreover, phylogenetic and isolation-by-distance analyses indicated evolutionary processes of RusV mostly in local reservoir populations. A comparison of complete genome sequences from all detected RusV lineages demonstrated a high level of amino acid and nucleotide sequence variability within a part of the p150 peptide of the non-structural polyprotein and its coding sequence, respectively. The location of this region within the RusV genome and its genetic properties were comparable to the hypervariable region of the rubella virus. The broad range of detected RusV spillover hosts in combination with its geographical distribution in northeastern Germany requires the assessment of its zoonotic potential and further analysis of encephalitis cases in mammals. Future studies have to prove a putative co-evolution scenario for RusV in the yellow-necked field mouse reservoir

Distribution of zoonotic variegated squirrel bornavirus 1 in naturally infected variegated and Prevost’s squirrels

Recently, the zoonotic capacity of the newly discovered variegated squirrel bornavirus 1 (VSBV-1) was confirmed in humans with a lethal encephalitis. Transmission to humans occurred by variegated and Prevost’s squirrels as presumed reservoir hosts but possible ways of virus shedding and the route of infection still need to be elucidated. Thus, the tissue distribution of VSBV-1 antigen and RNA was investigated in detail via immunohistochemistry (IHC) in six variegated and eight Prevost’s squirrels and by in situ hybridisation (ISH) in one Prevost’s squirrel, respectively. VSBV-1 antigen and RNA positive cells were most numerous in the nervous system and were also found in nearly all tissues and different cell types indicating a broad organ and cell tropism of VSBV-1. Presence of VSBV-1 in several organs might indicate potential virus shedding via various routes and implies the risk of intra- and interspecies transmission, respectively