Performance comparison of rapid and native barcoding methods for Oxford Nanopore sequencing of Poliovirus Viral Protein 1 (VP1) amplicons.

James E Bullows; Christina J Castro; Caeden Meade; Katherine Poston; Jaume Jorba

doi:10.1016/j.jviromet.2026.115436

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Performance comparison of rapid and native barcoding methods for Oxford Nanopore sequencing of Poliovirus Viral Protein 1 (VP1) amplicons.

James E Bullows·Christina J Castro·Caeden Meade·Katherine Poston·Jaume Jorba

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Abstract

Accurate and timely sequencing of poliovirus is critical for global eradication efforts, particularly for molecular epidemiology based on the typing region of the genome, viral protein 1 (VP1). While Oxford Nanopore Technologies (ONT) sequencing has expanded capabilities for poliovirus surveillance, the relative performance of different ONT library preparation methods, including ligation-based (Native Barcoding) and transposase-based (Rapid Barcoding) approaches, has not been systematically evaluated. In this study, we compared rapid barcoding and native barcoding workflows for sequencing VP1 amplicons from 17 type 2 poliovirus-positive samples, each processed in triplicate. Native barcoding generated significantly more sequencing output, producing approximately 2.3-fold greater total read yield than rapid barcoding, and demonstrated higher run-to-run reproducibility (R2 = 0.979-0.998 vs. 0.847-0.929, respectively; p < 0.001). In addition, native barcoding generated 80% of the total yield achieved by rapid barcoding within approximately 7 h, whereas rapid barcoding required approximately 40 h to reach the same output. Despite these differences, both methods produced identical VP1 consensus sequences across all samples, with comparable read quality (median per-base Q-scores of approximately Q17-Q18). Rapid barcoding provided substantial practical advantages, reducing hands-on library preparation time (55 vs. 200 min) and per-sample cost ($12.82 vs. $16.54), while simplifying workflow and reducing technical complexity. These findings indicate that sequencing yield may not be a determinant of downstream analytical outcomes for poliovirus VP1 ONT sequencing. Rapid barcoding therefore represents a cost-effective and efficient approach for routine poliovirus surveillance, whereas native barcoding remains advantageous in applications requiring rapid data generation or maximal sequencing depth.

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Poliovirus Humans DNA Barcoding, Taxonomic Reproducibility of Results Capsid Proteins Nanopore Sequencing Poliomyelitis High-Throughput Nucleotide Sequencing MinION Native barcoding Next generation sequencing Oxford Nanopore Rapid barcoding Journal Article Comparative Study

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James E Bullows, Christina J Castro, Caeden Meade, Katherine Poston, Jaume Jorba. 2026-07-16. Performance comparison of rapid and native barcoding methods for Oxford Nanopore sequencing of Poliovirus Viral Protein 1 (VP1) amplicons.. https://doi.org/10.1016/j.jviromet.2026.115436

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Journal Article; Comparative Study resource Comparative Study subject Humans subject Journal Article subject Next generation sequencing subject Poliomyelitis subject Poliovirus subject

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