Letter to the Editor: Development of proliferative verrucous leukoplakia (PVL) in oral lichen planus: Is it not a clinical spectrum of PVL?

Thalassemia, hemoglobinopathies, and glucose-6-phosphate dehydrogenase (G6PD) deficiency represent highly prevalent and genetically heterogeneous inherited hematologic disorders or diseases in Thailand. Conventional molecular approaches require multiple targeted assays to detect deletional and non-deletional variants across globin gene clusters and the G6PD gene, resulting in increased complexity, turnaround time, and cost. We developed and analytically validated a comprehensive long-read sequencing workflow using Oxford Nanopore Technologies for simultaneous detection of pathogenic variants in HBA 1, HBA 2, HBB, and G6PD. A total of 63 genomic DNA samples with previously characterized genotypes were analyzed using multiplex long-amplicon PCR followed by nanopore sequencing and bioinformatic variant calling with Clair 3. Variant annotation was performed using Ensembl VEP, and classification followed ACMG guidelines. All samples were successfully genotyped, demonstrating 100% concordance with conventional PCR, and Sanger sequencing. The platform accurately identified large α-globin deletions, β-thalassemia mutations, structural hemoglobin variants, and major Southeast Asian G6PD variants. A total of 39 distinct variants were detected and classified. Long-read sequencing effectively resolved homologous regions between HBA 1 and HBA 2 and enabled accurate characterization of multilocus co-inheritance patterns. This integrated nanopore-based workflow provides a robust, high-resolution, and scalable single-platform solution for comprehensive molecular diagnosis of thalassemia, hemoglobinopathies, and G6PD deficiency in genetically heterogeneous populations.