| Abstract: | Virioplankton play a crucial role in aquatic ecosystems as top-down regulators of bacterial populations and agents of horizontal gene transfer and nutrient cycling. However, the biology and ecology of virioplankton populations in the environment remain poorly understood. Ribonucleotide reductases (RNRs) are ancient enzymes that reduce ribonucleotides to deoxyribonucleotides and thus prime DNA synthesis. Composed of three classes according to O2 reactivity, RNRs can be predictive of the physiological conditions surrounding DNA synthesis. RNRs are universal among cellular life, common within viral genomes and virioplankton shotgun metagenomes (viromes), and estimated to occur within >90% of the dsDNA virioplankton sampled in this study. RNRs occur across diverse viral groups, including all three morphological families of tailed phages, making these genes attractive for studies of viral diversity. Differing patterns in virioplankton diversity were clear from RNRs sampled across a broad oceanic transect. The most abundant RNRs belonged to novel lineages of podoviruses infecting α-proteobacteria, a bacterial class critical to oceanic carbon cycling. RNR class was predictive of phage morphology among cyanophages and RNR distribution frequencies among cyanophages were largely consistent with the predictions of the “kill the winner–cost of resistance” model. RNRs were also identified for the first time to our knowledge within ssDNA viromes. These data indicate that RNR polymorphism provides a means of connecting the biological and ecological features of virioplankton populations.Viruses are key players in biogeochemical cycling and energy flow and help shape the composition of aquatic microbial communities (1–3). Additionally, viruses influence microbial metabolism through horizontal gene transfer and expression of auxiliary metabolic genes during infection (4). Despite their impact, we understand little about the specific biological features and ecological strategies of viral populations within natural ecosystems. Constraining these second-order issues is critical to building better quantitative models of how viral processes affect ecosystems (5).Methodological limitations have hindered efforts to understand viral ecology. Viruses lack a universally conserved phylogenetic marker, akin to the 16S rRNA gene in cells, which can broadly assay viral distributions and diversity. Marker genes used as proxies of environmental viral diversity are typically limited to specific viral taxa. Furthermore, PCR-based approaches can fail to detect prominent and biologically important viral populations owing to the potential for low nucleotide similarity between homologous genes. Recent work examining the diversity of viral DNA polymerase A genes within virioplankton metagenomic (virome) sequence data revealed that low-efficiency DNA polymerases, undetected by PCR, were predominant within virioplankton (6). That work also highlighted the unique ability of DNA polA sequences to provide insights into the biological features of unknown phages within the virioplankton. In general, the ability to connect biological features with sequence diversity in marker genes—including those widely used in ecological studies, such as the 16S rRNA gene—can be tenuous (7).Ideally, a marker gene of viral diversity should (i) be widely distributed among diverse viral lineages and, therefore, evolutionarily ancient; (ii) be abundant within environmental viral assemblages; (iii) play an important role in viral biology; (iv) have a single evolutionary origin and not be replaceable through nonorthologous gene displacement; (v) be phylogenetically informative; and (vi) be well represented in reference databases. Ribonucleotide reductase (RNR) gene products fulfill these criteria. Nucleotide metabolism pathways, including biosynthesis, are among the most represented within the virioplankton (8, 9). RNRs are the only known enzymes capable of reducing ribonucleotides to deoxyribonucleotides (10), an essential step for DNA synthesis. As such, RNRs are key to nucleotide biosynthesis, under stringent evolutionary selection pressure, and among the most abundant annotated genes in marine virome libraries (11). Importantly, RNR genes are present in all three families of tailed phages in the order Caudovirales and have been identified in viruses infecting hosts within all three domains of life (10). RNRs are strongly tied to lytic marine phages (12), which significantly influence nutrient cycles within the global ocean (5). Therefore, RNRs easily fit the criteria of being functionally nonredundant, abundant, and widely distributed.In addition, RNRs are biologically informative and form three physiological classes according to reactivity with O2. Class I RNRs are O2-dependent. Class II RNRs are O2-independent and rely upon adenosylcobalamin (vitamin B12). Class III RNRs are sensitive to O2. All three classes share a common catalytic center and use similar radical-based chemistry (13). Therefore, all three modern classes of RNR likely evolved from a single common ancestor (14). This study focused on the catalytic (alpha) subunit of the holoenzyme identified in virome libraries spanning a broad oceanic transect. Subsequently these data were used to examine the biological and ecological features of lytic phage populations within the Caudovirales. The outcomes of these analyses were interpreted within the context of known viral diversity and the “kill the winner–cost of resistance” (KTW–COR) model for viral–host interactions (15). Overall, these data show that RNR sequence diversity within the virioplankton connects broadly with phage morphological groups and can be predictive of the ecological strategies within the virioplankton. |
