Related Experiment Video
Updated: Jan 29, 2026
Amplicon Sequencing using the Long-Read Sequencing Technologies
Published on: August 29, 2025
Tiled-Amplicon Whole-Genome Sequencing Method Reveals Endemic Circulation of Human Adenovirus Type 3 in Japan
Gabriel Gonzalez1, Naganori Nao1, Koshiro Tabata1
1Institute for Vaccine Research and Development (HU-IVReD), Hokkaido University, Sapporo 001-0021, Japan.
Abstract:
Human adenovirus type 3 (HAdV-3) in the species Mastadenovirus blackbeardi is a frequent cause of hundreds of respiratory infections in Japan, with outbreaks varying in clinical severity. Such a high frequency of cases could be due to regular migration of novel variants or persistent circulation of endemic strains. Either scenario would require different measures to ameliorate the burden on public health. We designed a new cost-effective whole-genome sequencing protocol based on tiled amplicons and nanopore sequencing to clarify the circumstances behind the frequent outbreaks. This protocol was used with clinical samples collected between 2011 and 2020 from Japanese patients with acute respiratory illness (n = 110), resulting in near whole-genome sequences (~99% length) for 105 samples with high read coverage (~262.6 ± 192 reads). Phylogenetic analysis indicated sustained circulation of endemic strains in Japan during the analyzed decade and their relation to other strains worldwide with publicly available genome sequences. However, a comparison with other Japanese HAdV-3 strains circulating since 2023 suggested the public health measures introduced during the COVID-19 pandemic (2020-2022) indirectly affected the prevalence of circulating HAdV-3 variants. Additionally, our approach enabled the detection and partial sequencing (~71% completion) of co-infecting strains from the species Mastadenovirus caesari (n = 4) in the examined samples. The detection of adenoviruses belonging to different species in the same sample highlights how our protocol enhances the distinction of circulating viruses. In conclusion, these results and the introduced protocol will enable timely characterization and clinical interventions to mitigate this virus.
Insights
Human adenovirus type 3 (HAdV-3) causes frequent respiratory infections in Japan. Our new nanopore sequencing protocol reveals sustained endemic circulation and potential impacts of COVID-19 measures on variant prevalence.
Area of Science:
- Virology
- Genomics
- Public Health
Background:
- Human adenovirus type 3 (HAdV-3) is a significant cause of respiratory illness in Japan, with frequent outbreaks.
- Understanding the dynamics of HAdV-3 circulation, whether endemic or variant-driven, is crucial for public health interventions.
Purpose of the Study:
- To develop a cost-effective whole-genome sequencing protocol for HAdV-3 using tiled amplicons and nanopore sequencing.
- To investigate the molecular epidemiology of HAdV-3 in Japan between 2011 and 2020.
- To assess the impact of COVID-19 public health measures on HAdV-3 variant prevalence.
Main Methods:
- Whole-genome sequencing protocol based on tiled amplicons and nanopore sequencing.
- Phylogenetic analysis of HAdV-3 strains from clinical samples (n=110) collected between 2011-2020.
- Comparison with publicly available HAdV-3 genome sequences and recent Japanese strains.
Main Results:
- Near whole-genome sequences (~99%) were obtained for 105 samples, revealing sustained circulation of endemic HAdV-3 strains in Japan.
- Phylogenetic analysis placed Japanese strains in a global context.
- A comparison with post-2023 strains suggested indirect effects of COVID-19 pandemic measures on HAdV-3 variant prevalence.
- Co-infecting strains of *Mastadenovirus caesari* (n=4) were detected, showcasing the protocol's ability to distinguish different adenovirus species.
Conclusions:
- The developed nanopore sequencing protocol provides a cost-effective method for near whole-genome sequencing of HAdV-3.
- Endemic HAdV-3 strains have circulated persistently in Japan over the past decade.
- Public health interventions during the COVID-19 pandemic may have influenced HAdV-3 variant dynamics.
- The protocol's capability to detect co-infections aids in distinguishing circulating viruses and informs clinical interventions.
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