A new study has developed a sequencing protocol using Oxford Nanopore Technology (ONT), addressing critical limitations in current diagnostic methods for Hepatitis B Virus (HBV). Chronic HBV infection remains a significant public health concern, particularly in Africa, where the burden is substantial. HBV is classified into ten phylogenetically distinct genotypes (A–J), and existing diagnostic tests often fall short in providing comprehensive genetic insights.
Standard methods, such as full-genome sequencing or reverse hybridization, have inherent limitations. Diagnostic sequencing, often using the Sanger method, typically focuses only on the S-gene, offering limited information on intra-patient HBV genetic diversity. Meanwhile, reverse hybridization detects only known genotype-specific mutations. To overcome these challenges, this ONT-based protocol delivers complete genome sequences and detailed intra-patient diversity data.
The protocol involves tiling-based PCR amplification of HBV sequences, library preparation using the ONT Rapid Barcoding Kit, ONT GridION sequencing, and advanced analysis tools such as genome detective software for genotyping, jpHMM and RDP5.61 software for recombination analysis, and Geno2pheno for drug resistance profiling. It was validated on 148 residual diagnostic samples from HBV-infected patients in South Africa’s Western Cape province, generating high-quality near full-length HBV genomes. The findings provide valuable insights into HBV’s genetic diversity, recombination events, and drug resistance mutations.
Clinicians can use this protocol to improve treatment outcomes by identifying mutations associated with drug resistance, enabling personalized therapy decisions. Public health officials can leverage the sequencing data to track HBV’s spread, monitor emerging strains, and refine vaccination strategies tailored to regional challenges.
Beyond its clinical and public health applications, the protocol’s affordability and ease of use make it ideal for laboratories with limited resources. High-throughput sequencing of HBV genomes can become routine, enhancing diagnostic capabilities and improving patient care.
By integrating cutting-edge genomic research with practical applications, this ONT-based sequencing protocol exemplifies how innovation can directly impact healthcare and disease management. Its potential to enhance diagnostics, inform treatment, and support public health strategies marks a significant advancement in combating HBV globally.
Open Access Publication: https://www.mdpi.com/1422-0067/25/21/11702
