New viruses detected - results from an early warning system - metagenomic screening for unexpected or new viruses. New viruses detected - results from an early warning system

Pro-active approaches in preventing future epidemics include pathogen discovery prior to their emergence in human and/or animal populations. High-throughput sequencing (HTS) enables the characterization of microbial and viral genetic diversity within a given sample. In particular, metagenomic HTS allows the unbiased taxonomic profiling of sequences; hence, it can identify novel and highly divergent pathogens such as viruses. Newly discovered viral sequences must be further investigated using genomic characterization, molecular and serological screening, and/or in-vitro and in-vivo characterization. Several outbreak and surveillance studies applied unbiased generic HTS to characterize whole genome sequences of suspected pathogens. We aimed to screen for novel and unexpected pathogens in previously generated HTS datasets, and use this information as starting point for the early warning system. As proof of concept, we applied this early warning system (EWS) in HTS datasets and archived samples from the 2018-19 West Nile virus (WNV) epidemic in Germany. Metagenomic analyses classified viral sequences belonging to thirteen virus taxonomic families and other unclassified viruses. The EWS investigation focused on viral sequences annotated as families Peribunyaviridae and Reoviridae, which were suspected of causing co-infections in WNV-infected birds. Phylogenetic analyses revealed that the three reovirus genomes clustered with the mosquito-borne Umatilla virus species, whereas the peribunyavirus genome, tentatively named as “Hedwig virus”, belongs to an unclassified genus of the family. In follow up studies, newly developed molecular diagnostic assays detected fifteen Umatilla virus-positive wild birds from different German cities and eight Hedwig virus-positive captive birds from two zoological gardens. We also successfully cultivated Umatilla virus in mosquito C6/36 cells inoculated with a blackbird liver. However, the pathogenicity of the newly detected viruses is still under investigation, and their zoonotic potential is unknown. In conclusion, this study demonstrates the power of the proposed EWS in discovering and characterizing unexpected viruses in repurposed sequence datasets, and virus screening and cultivation using archived sample materials. The EWS enhances the strategies for pathogen recognition before they cause massive sporadic cases and outbreaks and proves to be a reliable tool for modern outbreak preparedness.