Mojumder, A. | bioRxiv Science

bioRxiv2026

Divergent Fates of Kidney-Resident Polyomaviruses: Stable Shedding Versus Near-Silent Persistence

Polyomaviruses establish long-term infection in the kidney and are intermittently shed in urine. However, the relationship between kidney-resident viral genomes and urinary shedding during persistent infection remains poorly defined. Using a genetically barcoded murine polyomavirus library, we tracked thousands of viral lineages in vivo by pairing longitudinal urine sampling with endpoint barcode sequencing of kidney tissue in four mice. Across all animals, kidney infection consistently resolved into two stable viral populations, with near-silent persistence as the dominant fate. Most kidney-resident barcodes were never detected in late urine at late stages of infection, even though many reached substantial abundance within the kidney, demonstrating that kidney viral genome levels alone do not predict urinary shedding. In contrast, only a small minority of kidney barcodes contributed disproportionately to urine virus output at late timepoints, and these barcodes exhibited stable longitudinal behavior, with repeated detection in urine over time and markedly higher peak urine abundance than late non-shed or random barcode controls. Shedding behavior was not explained by input virus stock abundance, barcode sequence features, predicted miRNA targeting, or ongoing reseeding from blood or other tissues. Instead, barcodes that ultimately dominated late urine already showed elevated urine detection early after infection, indicating that shedding fate is established early and maintained throughout persistent infection. Together, these findings reveal that persistent kidney infection is a structured reservoir composed of a large population of deeply restricted viral genomes and a smaller, stable subset that repeatedly produces urine-detectable viruses, with concurrent smoldering infections and latency-like restriction representing one possible model to explain the sharply different probabilities of shedding among kidney-resident genomes.

microbiology↗

bioRxiv2025

Shedding dynamics of a DNA virus population during acute and long-term persistent infection

Although much is known of the molecular mechanisms of virus infection within cells, substantially less is understood about within-host infection. Such knowledge is key to understanding how viruses take up residence and transmit infectious virus, in some cases throughout the life of the host. Here, using murine polyomavirus (muPyV) as a tractable model, we monitor parallel infections of thousands of differentially barcoded viruses within a single host. In individual mice, we show that numerous viruses (>2600) establish infection and are maintained for long periods post-infection. Strikingly, a low level of many different barcodes is shed in urine at all times post-infection, with a minimum of at least 80 different barcodes present in every sample throughout months of infection. During the early acute phase, bulk shed virus genomes derive from numerous different barcodes. This is followed by long term persistent infection detectable in diverse organs. Consistent with limited productive exchange of virus genomes between organs, each displays a unique pattern of relative barcode abundance. During the persistent phase, constant low-level shedding of typically hundreds of barcodes is maintained but is overlapped with rare, punctuated shedding of high amounts of one or a few individual barcodes. In contrast to the early acute phase, these few infrequent highly shed barcodes comprise the majority of bulk shed genomes observed during late times of persistent infection, contributing to a stark decrease in bulk barcode diversity that is shed over time. These temporally shifting patterns, which are conserved across hosts, suggest that polyomaviruses balance continuous transmission potential with reservoir-driven high-level reactivation. This offers a mechanistic basis for polyomavirus ubiquity and long-term persistence, which are typical of many DNA viruses. Author Summary / ImportancePolyomavirus infections, mostly benign but potentially fatal for immunocompromised individuals, undergo acute and long-term persistent infections. Typically, polyomavirus-associated diseases arise due to virus infection occurring in the context of a persistently infected individual. However, little is understood regarding the mechanisms of how polyomaviruses establish, maintain, and reactivate from persistent infection. We developed a non-invasive virus shedding assay combining barcoded murine polyomavirus, massively parallel sequencing technology, and novel computational approaches to track long-term infections in mice. We expect these methods to be of use not only to the study of DNA viruses but also for understanding persitent infection of diverse microbes. The study revealed organ-specific virus reservoirs and two distinct shedding patterns: constant low-level shedding of numerous barcodes and episodic high-level shedding of few barcodes. Over time, the diversity of shed barcodes decreased substantially. These findings suggest a persistent low-level infection in multiple reservoirs, with occasional bursts of replication in a small subset of infected cells. This combination of broad reservoirs and varied shedding mechanisms may contribute to polyomavirus success in transmission and maintaining long-term infections.

pathology↗