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In an attempt to explore the RNA viromes of two German rivers, we searched the virus particle contents of one 50 L water sample each from the Teltow Canal and the Havel River for viruses assumed to infect invertebrates. More than 330 complete and partial virus genomes up to a length of 37 kb were identified, with noda-like and reo-like viruses being most abundant, followed by bunya-like and birna-like viruses. Viruses related to the Permutotetraviridae, Nidovirales, Flaviviridae, Rhabdoviridae and Chuviridae as well as the unclassified Jῑngmén virus and Negev virus groups were also present. The results indicate a broad extent of recombinant virus genomes, supporting the concept of the modularity of eukaryotic viruses. For example, novel combinations of genes encoding replicase and structural proteins with a jellyroll fold have been observed. Less than 35 viruses could be assigned to existing virus genera. These are (i) an avian deltacoronavirus which was represented by only one short contig, albeit with 98% similarity, (ii) a seadornavirus and a rotavirus, and (iii) some 30 nodaviruses. All remaining viruses are novel and too diverse for accommodation in existing genera. Many of the virus genomes exhibit ORFans encoding hypothetical proteins of up to 2000 amino acids without conserved protein domains.

The viromes of freshwater bodies are underexplored. The Picornavirales order, with 371 acknowledged species, is one of the most expansive and diverse groups of eukaryotic RNA viruses. In this study, we add 513 picorna-like viruses to the assemblage of more than 2000 unassigned picorna-like viruses. Our set of the aquatic Picornavirales virome of the Teltow Canal in Berlin, Germany, consists of 239 complete and 274 partial genomes. This urban freshwater body is characterized by the predominance of marna-like viruses (30.8%) and dicistro-like viruses (19.1%), whereas picornaviruses, iflaviruses, solinvi-like viruses, polycipi-like viruses, and nora-like viruses are considerably less prevalent. Caliciviruses and secoviruses were absent in our sample. Although presenting characteristic domains of Picornavirales, more than 100 viruses (20.8%) could not be assigned to any of the 9 Picornavirales families. Thirty-three viruses of the Marnaviridae—mostly locarna-like viruses—exhibit a monocistronic genome layout. Besides a wealth of novel virus sequences, viruses with peculiar features are reported. Among these is a clade of untypeable marna-like viruses with dicistronic genomes, but with the capsid protein-encoding open reading frame located at the 5′ part of their RNA. A virus with a similar genome layout but clustering with dicistroviruses was also observed. We further detected monocistronic viruses with a polymerase gene related to aparaviruses. The detection of Aichi virus and five novel posa-like viruses indicates a slight burden in municipal wastewater.

Plant-infecting RNA viruses from 30 families and floating genera, as well as a great number of uncultured as yet-unclassified plant-associated viruses have been described. Even so, the plant RNA virosphere is still underexplored. RNA extracted from enriched virus particles of 50 L water samples from the Teltow Canal and the Havel River in Berlin, Germany, was sequenced using Illumina next-generation sequencing. Sequences were searched for plant viruses with BLAST and DIAMOND. Phylogenetic analyses were conducted with IQ-TREE 2. Altogether, 647 virus sequences greater than 1 kb were detected and further analyzed. These data revealed the presence of accepted and novel viruses related to Albetovirus, Alphaflexiviridae, Aspiviridae, Bromoviridae, Endornaviridae, Partitiviridae, Potyviridae, Solemoviridae, Tombusviridae and Virgaviridae. The vast majority of the sequences were novel and could not be taxonomically assigned. Several tombus- and endorna-like viruses make use of alternative translation tables that suggest unicellular green algae, ciliates, or diplomonades as their hosts. The identification of 27 albeto-like satellite viruses increases available sequence data five-fold. Sixteen new poty-like viruses align with other poty-like viruses in a link that combines the Astroviridae and Potyviridae families. Further, the identification of viruses with peptidase A6-like and peptidase A21-like capsid proteins suggests horizontal gene transfer in the evolution of these viruses.

To improve the understanding of the virome diversity of riverine ecosystems in metropolitan areas, a metagenome analysis was performed with water collected in June 2018 from the river Havel in Berlin, Germany. After enrichment of virus particles and RNA extraction, paired-end Illumina sequencing was conducted and assignment to virus groups and families was performed. This paper focuses on picorna-like viruses, the most diverse and abundant group of viruses with impact on human, animal, and environmental health. Here, we describe altogether 166 viral sequences ranging in size from 1 to 11.5 kb. The 71 almost complete genomes are comprised of one candidate iflavirus, one picornavirus, two polycipiviruses, 27 marnaviruses, 27 dicistro-like viruses, and 13 untypeable viruses. Many partial picorna-like virus sequences up to 10.2 kb were also investigated. The sequences of the Havel picorna-like viruses represent genomes of seven of eight so far known Picornavirales families. Detection of numerous distantly related dicistroviruses suggests the existence of additional, yet unexplored virus groups with dicistronic genomes, including few viruses with unusual genome layout. Of special interest is a clade of dicistronic viruses with capsid protein-encoding sequences at the 5′-end of the genome. Also, monocistronic viruses with similarity of their polymerase and capsid proteins to those of dicistroviruses are interesting. A second protein with NTP-binding site present in the polyprotein of solinviviruses and related viruses needs further attention. The results underline the importance to study the viromes of fluvial ecosystems. So far acknowledged marnaviruses have been isolated from marine organisms. However, the present study and available sequence data suggest that rivers and limnic habitats are relevant ecosystems with circulation of marnaviruses as well as a plethora of unknown picorna-like viruses.