Spindle-shaped viruses infect marine ammonia-oxidizing thaumarchaea

Abstract

Ammonia-oxidizing archaea (AOA) from the phylum Thaumarchaeota are ubiquitous in marine ecosystems and play a prominent role in carbon and nitrogen cycling. Previous studies have suggested that, like all microbes, thaumarchaea are infected by viruses and that viral predation has a profound impact on thaumarchaeal functioning and mortality, thereby regulating global biogeochemical cycles. However, not a single virus capable of infecting thaumarchaea has been reported thus far. Here we describe the isolation and characterization of three Nitrosopumilus spindle-shaped viruses (NSVs) that infect AOA and are distinct from other known marine viruses. Although NSVs have a narrow host range, they efficiently infect autochthonous Nitrosopumilus strains and display high rates of adsorption to their host cells. The NSVs have linear double-stranded DNA genomes of ∼28 kb that do not display appreciable sequence similarity to genomes of other known archaeal or bacterial viruses and could be considered as representatives of a new virus family, the "Thaspiviridae." Upon infection, NSV replication leads to inhibition of AOA growth, accompanied by severe reduction in the rate of ammonia oxidation and nitrite reduction. Nevertheless, unlike in the case of lytic bacteriophages, NSV propagation is not associated with detectable degradation of the host chromosome or a decrease in cell counts. The broad distribution of NSVs in AOA-dominated marine environments suggests that NSV predation might regulate the diversity and dynamics of AOA communities. Collectively, our results shed light on the diversity, evolution, and potential impact of the virosphere associated with ecologically important mesophilic archaea.

Keywords: ammonia-oxidizing archaea; chronic infection; spindle-shaped virus; viral predation.

Fig. 1.

Transmission electron microscopy images of negatively stained NSV1 virions. (A) NSV1 virions. (Scale bar, 50 nm.) (B) NSV1 particles attached to the surface of an SW cell. (Scale bar, 200 nm.) (C) Elongated NSV1 particles attached to the surface of an SW cell after 12 h of infection. The arrows indicate elongated NSV1 particles. (Scale bar, 200 nm.)

Fig. 2.

Comparative genomics of NSVs. Genomic maps of NSVs and related scaffolds. Shared ORFs are connected by color-coded shaded areas based on sequence identity. Genomes of NSVs are flanked by terminal inverted repeats. %GC represents mol% G+C content of DNA.

Fig. 3.

Phylogeny of pPolB and comparative genome maps of NSV1 and His1. (A) Maximum likelihood phylogenetic analysis of pPolB sequences from bacterial and archaeal viruses. In this tree, archaeal viruses comprise the following taxa: Ampullaviridae, Pleolipoviridae, Ovaliviridae, and Salterprovirus. Sequences originating from marine environments and hyperhalophilic archaeal viruses are highlighted with light blue and green backgrounds, respectively. Sequences from metagenomic datasets are indicated with gray font. (B) Comparison of the NSV1 and His1 genome maps. Functionally equivalent genes are indicated with matching colors. Genes encoding proteins detected in the purified virus particles are shown in cyan. Genes encoding small proteins containing Zn-binding domains are shown in green. Abbreviations: pPolB, protein-primed family B DNA polymerase; MCP, major capsid protein (putative); wHTH, winged helix-turn-helix; GTase, glycosyltransferase; MTase, DNA methyltransferase; PCNA, proliferating cell nuclear antigen; RHH, ribbon-helix-helix. The question mark next to the putative MCP denotes the uncertainty of this prediction.

Fig. 4.

Properties of the NSV1 infection cycle. Strain SW cells were infected with NSV1. Error bars represent SDs for three biological replicates. (A) Comparison of ammonia oxidation by NSV1-infected and noninfected control cells. (B) Virus production by strain SW cells infected with NSV1. AOA growth and virus production were measured by qPCR quantification of 16S rRNA and pPolB genes, respectively. (C) Fraction of nonadsorbed NSV1 virions, estimated by qPCR of viral pPolB gene. Viral genomic DNA of nonadsorbed virions was prepared from the culture supernatant.