Expanding diversity of virome associated with ammonium-oxidizing archaea and related lineages

Nitrososphaeria, an archaeal class containing all ammonium-oxidizing archaea (AOA) and their closely related clades (e.g. Ca. Caldarchaeales), is a widely distributed archaeal lineage across diverse ecosystems. Due to the difficulty of isolation and cultivation, the viral diversity and evolution associated with non-marine AOA lineages and Ca. Caldarchaeales remain largely uncharacterized. Here, based on globally sampled genomes and metagenomes, we assembled a genomic catalogue associated with AOA, including 283 species-level viruses and 415 species-level mobile genetic elements (MGEs). Notably, 259 previously unrecognized species-level viruses or MGEs associated with Ca. Caldarchaeales were identified. Across the two major Nitrososphaeria lineages, we uncovered an expansion of archaeal viral families: 11 families associated with AOA, including seven new head-tailed lineages, one of which comprises a mega archaeal virus with a genome size exceeding 220 kb; ten families associated with Ca. Caldarchaeales, including four novel head-tailed groups and two previously unknown rod-shaped family (Demeterviridae and Gaiaviridae). Phylogenetic inference showed that rod-shaped viruses associated with Ca. Caldarchaeales and methanogens are phylogenetically intertwined and likely share a common ancestor with Hoswirudivirus, a Sulfolobales-associated rod-shaped virus characterized by two copies of the coat gene. Through structural modelling, a rare fusion of the coat protein in Demeterviridae was observed, in which a single open reading frame within a gene island adjacent to a glycosyl transferase appears capable of forming a complete capsomer. This provides new insights into the origin and evolution of Adnaviria viral realm. Functional genomics analyses further revealed that these novel rod-shaped viruses encode virus-specific rare quinone biosynthesis genes and arsenic response transcriptional regulator genes, providing potential clues to their mechanisms of adaptation to hyperthermal environments. Collectively, our work uncovers the hidden genomic, evolution and functional diversity of Nitrososphaeria viromes for the first time and closes a long-standing gap in archaeal virome research. HighlightsO_LIFirst genomic catalogue of viruses and MGEs linked to Nitrososphaeria. C_LIO_LIDiscovery of diverse novel archaeal viral families. C_LIO_LINew insights into rod-shaped virus evolution and archaeal virus-host adaptation. C_LI