Summary
A bioRxiv preprint describes a single-amplicon sequencing workflow that recovered high-quality rhinovirus genomes from clinical specimens and enterovirus coding sequences from isolates. The authors report lower reagent costs and shorter processing time than hybridization capture sequencing.
A bioRxiv preprint posted on October 21, 2026, describes a single-amplicon sequencing workflow for rhinoviruses and enteroviruses. In testing, it recovered 116 high-quality rhinovirus genomes from 140 rhinovirus-positive respiratory specimens, compared with 113 using hybridization capture sequencing. The authors also report recovering expected enterovirus coding sequences from isolates and detecting two coxsackieviruses in specimens initially identified as rhinovirus-positive.
The approach aims to make it easier to obtain viral sequence across the coding region, rather than relying on the VP1 gene alone. The authors say the workflow used fewer reagents and less time than capture sequencing, supporting its potential use in genomic surveillance. The study is a preprint, so its findings have not yet been peer reviewed.
One amplicon to cover the viral coding region
An amplicon is a segment of genetic material copied during a polymerase chain reaction (PCR). The researchers designed a PCR and sequencing workflow intended to generate coding-complete genomes of both rhinoviruses and enteroviruses in a single reaction.
Routine tests often detect these viruses without reporting their species or genotype, according to the preprint. Researchers can use the VP1 gene for genotyping, but the authors note that VP1 alone cannot resolve transmission chains or detect recombination. Sequencing more of the coding region can provide a broader view of viral variation for surveillance.
The team evaluated the method on 140 rhinovirus-positive respiratory specimens and an isolate panel spanning 26 enterovirus A–D types. The preprint reports that expected polyprotein-coding sequences were recovered from all 40 enterovirus isolates tested. The same primer configuration also recovered coxsackievirus A6 and A16 from two specimens that had tested positive for rhinovirus.
Rhinovirus recovery and comparison with capture sequencing
The single-amplicon workflow produced 116 high-quality rhinovirus genomes across 57 genotypes. Hybridization capture sequencing produced 113 genomes from the comparison set. The authors describe the single-amplicon results as having higher fidelity and more uniform sequencing depth than capture sequencing.
Recovery was associated with viral abundance measured by RT-qPCR and droplet digital RT-PCR: specimens with more viral material were more likely to yield sequence. A visible PCR product band also predicted sequencing success. These findings make viral abundance an important factor when interpreting whether the workflow will produce a genome from a clinical specimen.
Cost and time reported by the authors
The preprint estimates reagent costs at about $30 per sample for single-amplicon sequencing, compared with $85 for hybridization capture. It also reports that the workflow required one-third as much time. These comparisons suggest a possible practical advantage for laboratories seeking to sequence many specimens, although the reported recovery results came from the study’s particular clinical specimens and isolate panel.
Because the work is a preprint, the results are preliminary. The evidence supports further evaluation of the method as a surveillance tool, with recovery depending in part on the amount of virus in a specimen.