Metagenomic Analysis of Rural Groundwater Viromes Reveals Bacteriophage Contributions to Groundwater Microbial Ecology

Groundwater ecosystems host diverse microbial communities, yet the diversity and ecological roles of their associated viral genomes remain poorly characterized. Here, we investigated viral community composition, diversity, host associations, lifestyles, and auxiliary metabolic potential in groundwater from three hand pumps located in Toukh, Qalyubia, Egypt, representing distinct local surroundings and potential contamination pressures. Using complementary viral detection approaches and a quality assessment workflow, we recovered 9,534 non-redundant viral contigs spanning a wide range of viral genome quality. Taxonomic profiling revealed dominance of tailed dsDNA bacteriophages (Uroviricota/Caudoviricetes) across all pumps, with ~ 99% of contigs not assigned below the class level. Whereas the viral composition of pump 3 was distinct and its diversity was consistently higher, pumps 1 and 2 clustered together, a pattern mirrored across taxonomic scales and diversity metrics. The majority of predicted viral hosts belonged to phylum Pseudomonadota, followed by Actinomycetota, Bacillota and Bacteroidota, with levels that varied between pumps. Correlation and network analyses showed strong concordance between the relative abundance of bacteria and the abundance of viruses that potentially infect them. Lifestyle prediction indicated a descending relative abundance of viruses with lysogenic lifestyle from pumps 1 through 3. Auxiliary metabolic genes (AMGs) related mainly to nucleotide, amino acid, and cofactor metabolism were detected in all pumps, with distinct pump-specific repertoires suggesting localized viral metabolic strategies. Together, these results demonstrate that groundwater viromes are ecologically structured and highly novel, with the potential ability to modulate host metabolism, highlighting their potential role in shaping subsurface microbial communities.