Abstract
ABSTRACTViruses are ubiquitous microbiome components, shaping ecosystems via strain-specific predation, horizontal gene transfer and redistribution of nutrients through host lysis. Viral impacts are important in groundwater ecosystems, where microbes drive many nutrient fluxes and metabolic processes; however, little is known about the diversity of viruses in these environments. We analyzed four groundwater plasmidomes (the entire plasmid content of an environment) and identified 200 viral sequences, which clustered into 41 genus-level viral clusters (approximately equivalent to viral genera) including 9 known and 32 putative new genera. We used publicly available bacterial whole-genome sequences (WGS) and WGS from 261 bacterial isolates from this groundwater environment to identify potential viral hosts. We linked 76 of the 200 viral sequences to a range of bacterial phyla, the majority associated with Proteobacteria, followed by Firmicutes, Bacteroidetes, and Actinobacteria. The publicly available WGS enabled mapping bacterial hosts to several viral sequences. The WGS of groundwater isolates increased the depth of host prediction by allowing host identification at the strain level. The latter included 4 viruses that were almost entirely (>99% query coverage, >99% identity) identified as integrated in the genomes of Pseudomonas, Acidovorax, and Castellaniella strains, resulting in high-confidence host assignments. Lastly, 21 of these viruses carried putative auxiliary metabolite genes for metal and antibiotic resistance, which might drive their infection cycles and/or provide selective advantage to infected hosts. Exploring the groundwater virome provides a necessary foundation for integration of viruses into ecosystem models where they are key players in microbial adaption to environmental stress.IMPORTANCE To our knowledge, this is the first study to identify the bacteriophage distribution in a groundwater ecosystem shedding light on their prevalence and distribution across metal-contaminated and background sites. Our study is uniquely based on selective sequencing of solely the extrachromosomal elements of a microbiome followed by analysis for viral signatures, thus establishing a more focused approach for phage identifications. Using this method, we detected several novel phage genera along with those previously established. Our approach of using the whole-genome sequences of hundreds of bacterial isolates from the same site enabled us to make host assignments with high confidence, several at strain levels. Certain phage genes suggest that they provide an environment-specific selective advantage to their bacterial hosts. Our study lays the foundation for future research on directed phage isolations using specific bacterial host strains to further characterize groundwater phages, their life cycles, and their effects on groundwater microbiome and biogeochemistry.
Highlights
We leveraged existing data focused on extrachromosomal circular DNA templates by identifying viruses from plasmidome data sets (Fig. 1)
Ecogenomics of Groundwater Viruses that both can be found as extrachromosomal circular DNA molecules, we used VirSorter, a tool designed to predict bacterial and archaeal virus sequences on the plasmidome assemblies [29], and identified 200 sequences as groundwater viral sequences from 13,770 plasmidome contigs (Fig. S1A)
Clustering of the 200 groundwater viral sequences with publicly available bacterial and archaeal viruses revealed that 85 groundwater viral genomes formed 41 viral clusters with at least one representative of groundwater virus (Table S3)
Summary
Viral diversity in groundwater systems has been much less studied, but recent metagenomic studies suggested that groundwater viral communities were clearly distinct from other freshwater environments, that their diversity and structure reflected changes in environmental parameters, including especially pH level and the presence of contaminants, and that viruses may significantly influence groundwater microbe dynamics [19, 20]. The Oak Ridge Field Research Center (ORFRC) [21,22,23] is a well-studied U.S Department of Energy site that includes groundwater areas with and without metal contamination, referred to as the contaminated and background sites, respectively It has been well characterized in terms of the physical parameters, microbiome distribution, and fluctuation in response to different environmental stresses and served as an excellent model groundwater system for studies. We mined the plasmidome data from a well-characterized groundwater system and analyzed the resulting viral sequences complete with genomic and ecological contexts
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