A custom hepatitis A virus assay for whole-genome sequencing

Nora G Cleary; Patrick W Bryant; Daryl M Lamson; Alexandra P Newman; Kirsten St George

doi:10.1016/j.jviromet.2022.114649

A custom hepatitis A virus assay for whole-genome sequencing

J Virol Methods. 2023 Feb:312:114649. doi: 10.1016/j.jviromet.2022.114649. Epub 2022 Nov 12.

Authors

Nora G Cleary¹, Patrick W Bryant², Daryl M Lamson¹, Alexandra P Newman³, Kirsten St George⁴

Affiliations

¹ Laboratory of Viral Diseases, Wadsworth Center, New York State Department of Health, Albany, New York 12208, USA.
² Laboratory of Viral Diseases, Wadsworth Center, New York State Department of Health, Albany, New York 12208, USA. Electronic address: patrick.bryant@health.ny.gov.
³ Bureau of Communicable Disease Control, New York State Department of Health, Albany, New York 12237, USA.
⁴ Laboratory of Viral Diseases, Wadsworth Center, New York State Department of Health, Albany, New York 12208, USA; Department of Biomedical Science, University at Albany, SUNY, Albany, New York 12144, USA.

PMID: 36375537
DOI: 10.1016/j.jviromet.2022.114649

Abstract

Since 2016, the United States has experienced a resurgence in the number of hepatitis A virus (HAV) cases and outbreaks. These outbreaks have been sustained by person-to-person transmission with cases occurring predominantly in high-risk populations including intravenous drug users, individuals experiencing homelessness, and men who have sex with men. To investigate HAV transmission, a molecular-surveillance system consisting of real-time RT-PCR (rRT-PCR) for detection, and a conventional RT-PCR assay for genotyping of HAV, was established in New York State (NYS) in 2019. Since then, a total of 271 HAV-positive serum samples collected from cases across NYS between 2019 and 2021 were identified by rRT-PCR. To rapidly and efficiently generate HAV whole-genome sequences, a custom AmpliSeq™ panel was designed in collaboration with Thermo Fisher. To streamline the process, sample preparation was performed on an Ion Chef and sequencing on an Ion S5XL. Of the 271 HAV-positive samples, the whole-genome sequencing (WGS) assay successfully generated 134 near-complete, high-quality HAV sequences. Phylogenetic analysis of the VP1-2A region identified 216 IB, 48 IA, and 2 IIIA genotypes, while 5 were unable to be typed due to poor sequence in this key region. The HAV whole-genome sequencing approach provided a more efficient and streamlined approach for genotyping HAV compared to previous methods and resulted in phylogenetic trees with enhanced resolution compared to the HAV VP1-2A region alone.

Keywords: AmpliSeq™; GHOST; Hepatitis A; Molecular epidemiology; Whole-genome sequencing.

Publication types

Research Support, U.S. Gov't, P.H.S.

MeSH terms

Genotype
Hepatitis A virus* / genetics
Hepatitis A* / diagnosis
Homosexuality, Male
Humans
Male
Phylogeny
RNA, Viral / genetics
Real-Time Polymerase Chain Reaction
Sexual and Gender Minorities*

Substances

RNA, Viral