A nanopore-based amplicon-sequencing panel to determine the origin and zoonotic host species of Plasmodium knowlesi infections
Abstract
Population genomic studies have revealed strong regional structure in Plasmodium knowlesi across Southeast Asia. To facilitate routine genomic surveillance, we developed a targeted amplicon sequencing panel that preserves the large- and small-scale population genetic structure. We screened published genomes for short, highly polymorphic targets and a 30-SNP barcode. We validated multiplexed amplification and sequencing using a portable Oxford Nanopore Technologies MinION sequencer on 28 clinical infections from four southern Thai provinces collected between 2018-2023. Twenty-seven loci yielded sequence data and 23 infections with data from at least eight loci were retained for analysis. Across the 23 Thai infections and 104 published Malaysian genomes, 586 diverse SNPs were observed. The amplicon panel and 30-SNP barcode retained the major population structure observed in genome-wide data, including a clear distinction between Bornean and Peninsular Malaysian infections and two clusters in Borneo. Across newly sequenced Thai infections, 25 loci were diverse and showed substantial diversity (mean expected heterozygosity = 0.69). All infections were effectively monoclonal. Pairwise identity-by-descent revealed substantial recent relatedness, with 46 of 253 pairs at or above 0.5. Two Thai genetic groups (n = 12 and n = 11) were observed, mirroring previously observed structure in peninsular Malaysia. Groups were not significantly associated with province or collection year. In conclusion, targeted sequencing can reveal geographically informative population structure across scales and thus support surveillance.
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BibTeX
@misc{silva2026nanoporebased,
title = {{A nanopore-based amplicon-sequencing panel to determine the origin and zoonotic host species of Plasmodium knowlesi infections}},
author = {Gustavo da Silva and Parsakorn Tapaopong and Aurel Holzschuh and Jetsumon Sattabongkot and Wang Nguitragool and Cristian Koepfli},
howpublished = {medRxiv},
year = {2026},
month = oct,
doi = {10.64898/2026.10.16.26365752},
url = {https://www.medrxiv.org/content/10.64898/2026.10.16.26365752v1},
note = {CC BY},
}
RIS
TY - UNPB
TI - A nanopore-based amplicon-sequencing panel to determine the origin and zoonotic host species of Plasmodium knowlesi infections
AU - Gustavo da Silva
AU - Parsakorn Tapaopong
AU - Aurel Holzschuh
AU - Jetsumon Sattabongkot
AU - Wang Nguitragool
AU - Cristian Koepfli
PY - 2026
DA - 2026/10/16
PB - medRxiv
DO - 10.64898/2026.10.16.26365752
UR - https://www.medrxiv.org/content/10.64898/2026.10.16.26365752v1
AB - Population genomic studies have revealed strong regional structure in Plasmodium knowlesi across Southeast Asia. To facilitate routine genomic surveillance, we developed a targeted amplicon sequencing panel that preserves the large- and small-scale population genetic structure. We screened published genomes for short, highly polymorphic targets and a 30-SNP barcode. We validated multiplexed amplification and sequencing using a portable Oxford Nanopore Technologies MinION sequencer on 28 clinical infections from four southern Thai provinces collected between 2018-2023. Twenty-seven loci yielded sequence data and 23 infections with data from at least eight loci were retained for analysis. Across the 23 Thai infections and 104 published Malaysian genomes, 586 diverse SNPs were observed. The amplicon panel and 30-SNP barcode retained the major population structure observed in genome-wide data, including a clear distinction between Bornean and Peninsular Malaysian infections and two clusters in Borneo. Across newly sequenced Thai infections, 25 loci were diverse and showed substantial diversity (mean expected heterozygosity = 0.69). All infections were effectively monoclonal. Pairwise identity-by-descent revealed substantial recent relatedness, with 46 of 253 pairs at or above 0.5. Two Thai genetic groups (n = 12 and n = 11) were observed, mirroring previously observed structure in peninsular Malaysia. Groups were not significantly associated with province or collection year. In conclusion, targeted sequencing can reveal geographically informative population structure across scales and thus support surveillance.
ER -