Differences in neuroinflammation in the olfactory bulb between D614G, Delta and Omicron BA.1 SARS-CoV-2 variants in the hamster model (preprint)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is associated with various neurological complications. SARS-CoV-2 infection induces neuroinflammation in the central nervous system (CNS), whereat the olfactory bulb seems to be involved most frequently. Here we show differences in the neuroinvasiveness and neurovirulence among SARS-CoV-2 variants in the hamster model five days post inoculation. Replication in the olfactory mucosa was observed in all hamsters, but most prominent in D614 inoculated hamsters. We observed neuroinvasion into the CNS via the olfactory nerve in D614G-, but not Delta (B.1.617.2)- or Omicron BA.1 (B.1.1.529) inoculated hamsters. Neuroinvasion was associated with neuroinflammation in the olfactory bulb of hamsters inoculated with D614G but hardly in Delta or Omicron BA.1. Altogether, this indicates that there are differences in the neuroinvasive and neurovirulent potential among SARS-CoV-2 variants in the acute phase of the infection in the hamster model.

Pulmonary lesions following inoculation with the SARS-CoV-2 Omicron BA.1 (B.1.1.529) variant in Syrian golden hamsters (preprint)

The Omicron BA.1 (B.1.1.529) SARS-CoV-2 variant is characterized by a high number of mutations in the viral genome, associated with immune-escape and increased viral spread. It remains unclear whether milder COVID-19 disease progression observed after infection with Omicron BA.1 in humans is due to reduced pathogenicity of the virus or due to pre-existing immunity from vaccination or previous infection. Here, we inoculated hamsters with Omicron BA.1 to evaluate pathogenicity and kinetics of viral shedding, compared to Delta (B.1.617.2) and to animals re-challenged with Omicron BA.1 after previous SARS-CoV-2 614G infection. Omicron BA.1 infected animals showed reduced clinical signs, pathological changes, and viral shedding, compared to Delta-infected animals, but still showed gross- and histopathological evidence of pneumonia. Pre-existing immunity reduced viral shedding and protected against pneumonia. Our data indicate that the observed decrease of disease severity is in part due to intrinsic properties of the Omicron BA.1 variant.

Omicron BA.1 and BA.2 are antigenically distinct SARS-CoV-2 variants (preprint)

The emergence and rapid spread of SARS-CoV-2 variants may impact vaccine efficacy significantly. The Omicron variant termed BA.2, which differs genetically substantially from BA.1, is currently replacing BA.1 in several countries, but its antigenic characteristics have not yet been assessed. Here, we used antigenic cartography to quantify and visualize antigenic differences between SARS-CoV-2 variants using hamster sera obtained after primary infection. Whereas early variants are antigenically similar, clustering relatively close to each other in antigenic space, Omicron BA.1 and BA.2 have evolved as two distinct antigenic outliers. Our data show that BA.1 and BA.2 both escape (vaccine-induced) antibody responses as a result of different antigenic characteristics. Close monitoring of the antigenic changes of SARS-CoV-2 using antigenic cartography can be helpful in the selection of future vaccine strains.

Multi-species ELISA for the detection of antibodies against SARS-CoV-2 in animals (preprint)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a pandemic with millions of infected humans and hundreds of thousands of fatalities. As the novel disease - referred to as COVID-19 - unfolded, occasional anthropozoonotic infections of animals by owners or caretakers were reported in dogs, felid species and farmed mink. Further species were shown to be susceptible under experimental conditions. The extent of natural infections of animals, however, is still largely unknown. Serological methods will be useful tools for tracing SARS-CoV-2 infections in animals once test systems are validated for use in different species. Here, we developed an indirect multi-species ELISA based on the receptor-binding domain (RBD) of SARS-CoV-2. The newly established ELISA was validated using 59 sera of infected or vaccinated animals including ferrets, raccoon dogs, hamsters, rabbits, chickens, cattle and a cat, and a total of 220 antibody-negative sera of the same animal species. Overall, a diagnostic specificity of 100.0% and sensitivity of 98.31% was achieved, and the functionality with every species included in this study could be demonstrated. Hence, a versatile and reliable ELISA protocol was established that enables high-throughput antibody detection in a broad range of animal species, which may be used for outbreak investigations, to assess the seroprevalence in susceptible species or to screen for reservoir or intermediate hosts.

What if we perceive SARS-CoV-2 genomes as documents? Topic modelling using Latent Dirichlet Allocation to identify mutation signatures and classify SARS-CoV-2 genomes (preprint)

Topic modeling is frequently employed for discovering structures (or patterns) in a corpus of documents. Its utility in text-mining and document retrieval tasks in various fields of scientific research is rather well known. An unsupervised machine learning approach, Latent Dirichlet Allocation (LDA) has particularly been utilized for identifying latent (or hidden) topics in document collections and for deciphering the words that define one or more topics using a generative statistical model. Here we describe how SARS-CoV-2 genomic mutation profiles can be structured into a Bag of Words to enable identification of signatures (topics) and their probabilistic distribution across various genomes using LDA. Topic models were generated using ~47000 novel corona virus genomes (considered as documents), leading to identification of 16 amino acid mutation signatures and 18 nucleotide mutation signatures (equivalent to topics) in the corpus of chosen genomes through coherence optimization. The document assumption for genomes also helped in identification of contextual nucleotide mutation signatures in the form of conventional N-grams (e.g. bi-grams and tri-grams). We validated the signatures obtained using LDA driven method against the previously reported recurrent mutations and phylogenetic clades for genomes. Additionally, we report the geographical distribution of the identified mutation signatures in SARS-CoV-2 genomes on the global map. Use of the non-phylogenetic albeit classical approaches like topic modeling and other data centric pattern mining algorithms is therefore proposed for supplementing the efforts towards understanding the genomic diversity of the evolving SARS-CoV-2 genomes (and other pathogens/microbes).

Susceptibility of raccoon dogs for experimental SARS-CoV-2 infection (preprint)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged in China at the end of 2019, and became pandemic. The zoonotic virus most likely originated from bats, but definite intermediate hosts have not yet been identified. Raccoon dogs (Nyctereutes procyonoides) are kept for fur production, in particular in China, and were suspected as potential intermediate host for both SARS-CoV6 and SARS-CoV2. Here we demonstrate susceptibility of raccoon dogs for SARS-CoV-2 infection after intranasal inoculation and transmission to direct contact animals. Rapid, high level virus shedding, in combination with minor clinical signs and pathohistological changes, seroconversion and absence of viral adaptation highlight the role of raccoon dogs as a potential intermediate host. The results are highly relevant for control strategies and emphasize the risk that raccoon dogs may represent a potential SARS-CoV-2 reservoir. Our results support the establishment of adequate surveillance and risk mitigation strategies for kept and wild raccoon dogs. Article Summary LineRaccoon dogs are susceptible to and efficiently transmit SARS-CoV2 and may serve as intermediate host

Experimental Transmission Studies of SARS-CoV-2 in Fruit Bats, Ferrets, Pigs and Chickens (preprint)

Background: A novel zoonotic SARS-related coronavirus emerged in China at the end of 2019. The novel SARS-CoV-2 became pandemic within weeks and the number of human infections and severe cases is increasing. The role of potential animal hosts is still understudied.Methods: We intranasally inoculated fruit bats (Rousettus aegyptiacus; n=9), ferrets (n=9), pigs (n=9) and chickens (n=17) with 105 TCID50 of a SARS-CoV-2 isolate per animal. Animals were monitored clinically and for virus shedding. Direct contact animals (n=3) were included. Animals were humanely sacrificed for virological and immune-pathohistological analysis at different time points.Findings: Under these settings, pigs and chickens were not susceptible to SARS-CoV-2. All swabs as well as organ samples and contact animals remained negative for viral RNA, and none of the animals seroconverted. Rousettus aegyptiacus fruit bats experienced a transient infection, with virus detectable by RT-qPCR, immunohistochemistry (IHC) and in situ hybridization (ISH) in the nasal cavity, associated with rhinitis. Viral RNA was also identified in the trachea, lung and lung associated lymphatic tissue. One of three contact bats became infected. More efficient virus replication but no clinical signs were observed in ferrets with transmission to all direct contact animals. Prominent viral RNA loads of up to 104 viral genome copies/ml were detected in the upper respiratory tract. Mild rhinitis was associated with viral antigen detection in the respiratory and olfactory epithelium. Both fruit bats and ferrets developed SARS-CoV-2 reactive antibodies reaching neutralizing titers of up to 1:1024.Interpretation: Pigs and chickens could not be infected intranasally by SARS-CoV-2, whereas fruit bats showed characteristics of a reservoir host. Virus replication in ferrets resembled a subclinical human infection with efficient spread. These animals might serve as a useful model for further studies e.g. testing vaccines or antivirals.Funding Statement: Intramural funding of the German Federal Ministry of Food and Agriculture provided to the Friedrich-Loeffler-Institut.Declaration of Interests: All authors declare no competing interest.Ethics Approval Statement: The animal experiments were evaluated and approved by the ethics committee of the State Office of Agriculture, Food safety, and Fishery in Mecklenburg – Western Pomerania (LALLF M-V: LVL MV/TSD/7221.3-2-010/18-12). All procedures were carried out in approved biosafety level 3 (BSL3) facilities.