In the context of clinical and public health settings, HBV sequencing can provide information that is useful in characterizing virus genotype, potential transmission networks, drug and vaccine resistance, and aspects of the dynamics of infection 5, 7, 8. New sequencing initiatives are required with improved methodologies that are efficient, accurate, sensitive and cost-effective 6. Large-scale virus genome sequencing to provide more complete genetic information at the population and individual level can shed light on the limitations of current interventions 5, and inform new strategies for elimination.
Despite the availability of a robust prophylactic vaccine and safe suppressive antiviral therapy, HBV has remained endemic - and neglected - in many populations 4. An improved understanding of the molecular biology, epidemiology, infection dynamics and pathophysiology of HBV is a crucial step towards reducing the global burden of HBV disease. The status of HBV infection as a globally important public health problem is highlighted by United Nations Sustainable Development Goals, which set a target for HBV elimination by the year 2030 3. The combination of isothermal amplification and Nanopore sequencing also offers appealing potential to develop point-of-care tests for HBV, and for other viruses.Ĭhronic hepatitis B virus (HBV) infection affects an estimated 250–290 million individuals worldwide, resulting in around 800,000 deaths from chronic liver disease and hepatocellular carcinoma each year 1, 2. Thus while Illumina sequencing may still be the most accurate way to capture within-sample diversity, Nanopore data can contribute to an understanding of linkage between polymorphisms within individual virions. With this approach, we can achieve an improved consensus sequencing accuracy of 99.7% and resolve intra-sample sequence variants to form whole-genome haplotypes.
#Illumina adapter sequences plus#
Nanopore reads can be used to implement a straightforward method for error correction that reduces the per-read error rate, by comparing multiple genome copies combined into a single concatemer and by analysing reads generated from plus and minus strands. We show that this product is suitable for Nanopore sequencing as single reads, as well as for generating short-read Illumina sequences. Here we describe advances to a protocol that exploits the circular HBV genome structure, using isothermal rolling-circle amplification to enrich HBV DNA, generating concatemeric amplicons containing multiple successive copies of the same genome. Advancing interventions to tackle the huge global burden of hepatitis B virus (HBV) infection depends on improved insights into virus epidemiology, transmission, within-host diversity, drug resistance and pathogenesis, all of which can be advanced through the large-scale generation of full-length virus genome data.
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