High-Throughput Oxford Nanopore Sequencing Unveils Complex Viral Population in Kansas Wheat: Implications for Sustainable Virus Management.

Wheat viruses are major yield-reducing factors, with mixed infections causing substantial economic losses. Determining field virus populations is crucial for effective management and developing virus-resistant cultivars. This study utilized the high-throughput Oxford Nanopore sequencing technique (ONT) to characterize wheat viral populations in major wheat-growing counties of Kansas from 2019 to 2021. Wheat leaves exhibiting virus-like symptoms were collected, total RNA was extracted, and cDNA libraries were prepared using a PCR-cDNA barcoding kit, then loaded onto ONT MinION flow cells. Sequencing reads aligned with cereal virus references identified eight wheat virus species. (wheat streak mosaic virus, WSMV), (Triticum mosaic virus, ), (brome mosaic virus, ), as well as , , , , and Mixed infections involving two to five viruses in a single sample were common, with the most prevalent being WSMV + TriMV at 16.7% and WSMV + TriMV + BMV at 11.9%. Phylogenetic analysis revealed a wide distribution of WSMV isolates, including European and recombinant variants. A phylogenetic analysis of based on RNA 3A and 3B segments and whole-genome characterization of were also conducted. These findings advance understanding of genetic variability, phylogenetics, and viral co-infections, supporting the development of sustainable management practices through host genetic resistance.