bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2025.07.29.667219

A Genome Sequence Variant Monitoring Program for Seasonal Influenza A H3N2 and Respiratory Syncytial Virus A using Wastewater-Based Surveillance in Ontario, Canada

Nash, D.·Knapp, J. J.·Overton, A. K.·Hungwe, Y. C.·Menon, R.·Nissimov, J. I.·Charles, T. C.
View original source ↗Export metadata ↓

Abstract

Seasonal respiratory viruses, such as the Influenza A virus and the respiratory syncytial virus, are responsible for over a billion infections worldwide each year resulting in a substantial burden on health care systems. Surveillance of these viruses, including their prevalence in communities and their evolution, are essential for informing public health decisions and recommending vaccine formulations and schedules. Typically, these viruses are monitored using clinical samples from patients seeking medical attention. Recently, wastewater-based surveillance (WBS) has been leveraged to understand transmission dynamics and genome evolution of SARS-CoV-2 and seasonal respiratory viruses. To further the utility of WBS we developed and implemented novel tiled-amplicon sequencing assays to identify and track Influenza A virus H3N2 and respiratory syncytial virus A circulating in Southern Ontario, Canada. We also developed virus specific deconvolution tools to estimate the abundance of mixed lineages in wastewater. These assays were able to accurately determine which lineages were circulating in wastewater with high sensitivity and specificity. If implemented in regular surveillance programs, they could be used to inform real-time public health decisions and determine potential disease surge with impact on emergency room visits and hospitalization, as well as track which emerging strains will become predominant in the future and determine which strains should be the focus of seasonal vaccines.

Explore related subjects

genomicsPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Nash, D., Knapp, J. J., Overton, A. K., Hungwe, Y. C., Menon, R., Nissimov, J. I., Charles, T. C.. 2025-07-31. A Genome Sequence Variant Monitoring Program for Seasonal Influenza A H3N2 and Respiratory Syncytial Virus A using Wastewater-Based Surveillance in Ontario, Canada. https://doi.org/10.1101/2025.07.29.667219

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Structural variation in repeat elements is widespread in normal human tissues and in tumorigenesis

Somatic mosaicism contributes to genomic variation, yet postzygotic structural variants remain under-characterized. We performed long- and short-read WGS from multiple individuals (n=47 normal tissues; n=168 samples) and identified mosaic structural variants in all individuals and germ layers, impacting a median 285.2 kb/genome. Nearly half of breakpoints were independently validated, with tissue distributions reflecting both early and late developmental origins. Most mosaic variants were repeat-mediated and 8.3% overlapped functional elements, an enrichment compared to germline variants. To extend these analyses in samples where long-read sequencing is infeasible, we measured repeat alterations from short-read sequencing, recapitulating mosaic tissue-specific differences. We characterized tumor- and tissue- specific variation in repeats across 15 cancer types and found tumor-related repeat variation to be similar in scale to that of normal mosaic variation. Tracking repeat changes in cell-free DNA provided a noninvasive approach for tumor monitoring. Our analyses revealed widespread repeat-driven structural variation in health and disease.

genomics↗
bioRxiv2026

RNA isoform-resolved multiplexed sequencing with bioorthogonal barcoding

RNA isoform dysregulation drives disease pathogenesis and is the target of FDA-approved splice-switching therapeutics. However, multiplexed sequencing methods discard splice junction information because only 3' termini are barcoded and counted. Here, we repurpose acylation and click chemistries to conjugate bioorthogonal barcodes (bobcodes) directly onto multiple internal positions along cellular RNAs. Bobcoded RNAs from multiple samples are pooled for multiplexed cDNA synthesis, during which reverse transcriptase switches from each RNA template onto its tethered bobcode with greater than 99% accuracy in species mixing experiments. Bobcode attachment intervals set cDNA insert sizes without a library fragmentation step, and priming with poly(dT) or random hexamers selects between 3'-end counting and full-length isoform capture. A bioorthogonal barcode-sequencing (BOB-seq v0.1) drug screen identifies transcriptome-wide on- and off-target RNA splicing effects and outperforms existing multiplexing RNA sequencing methods in workflow simplicity, sample-to-sample variability, and barcoding accuracy. Bobcodes add isoform resolution to scalable multiplexed RNA sequencing.

genomics↗
bioRxiv2026

Structural polymorphism and population-variable coding capacity of HERV-K(HML-2) in human pangenomes

Approximately 8% of the human genome is derived from ancient retroviral infections. The most recently integrated of these endogenous retroviruses is the HERV-K(HML-2) clade, whose expression has been associated with cancer, amyotrophic lateral sclerosis, and embryogenesis. Studies of HERV expression, particularly HML-2, have relied predominantly on short-read sequencing. However, the high similarity among HML-2 proviruses prevents many short reads from being assigned uniquely to individual loci. We therefore compared haplotype-resolved long-read genome assemblies from 292 donors to resolve variation in proviral structure and coding capacity. Several loci previously thought to be fixed were structurally polymorphic. Tandem arrays occurred at 13 loci and contained up to six proviral copies in a single array. At 8q11.23, we identified a previously undescribed full-length provirus in one haplotype. All 583 other haplotypes carried a solo-LTR. We found that standard reference genomes failed to represent the coding capacity retained in many individuals, whose proviruses contained intact open reading frames despite disruptive mutations in the reference sequences. Short-read genotypes left 32.5% of the tested donor-variant pairs unresolved at sites associated with viral reading frames. These findings show why HML-2 expression must be interpreted in the context of the structural and coding alleles each individual carries.

genomics↗