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Updated: Aug 29, 2025
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Sequencing of mRNA from Whole Blood using Nanopore Sequencing
Published on: June 3, 2019
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Rapid phylogenetic analysis using open reading frame content patterns acquired by Oxford nanopore sequencing.
Kengo Hayashi1, Yohei Doi1,2,3, Masahiro Suzuki1
1Department of Microbiology, Fujita Health University School of Medicine, Toyoake, Japan.
Journal of Applied Microbiology
|September 8, 2022
Summary
A new phylogenetic method uses open reading frame (ORF) content from Oxford Nanopore Technologies (ONT) sequencing data for bacterial epidemiology. This ORF-based approach bypasses core genome single nucleotide polymorphism (cgSNP) limitations, offering a faster, more accessible alternative for real-time analysis.
Area of Science:
- Microbiology
- Genomics
- Bioinformatics
Background:
- Whole-genome sequencing (WGS) and core genome single nucleotide polymorphism (cgSNP) analysis are vital for bacterial epidemiology.
- Oxford Nanopore Technologies (ONT) sequencing offers real-time data but has a high error rate, limiting its use in cgSNP-based phylogenetics.
- Current methods are resource-intensive and time-consuming.
Purpose of the Study:
- To develop a cgSNP-independent phylogenetic analysis method using ONT sequencing data.
- To leverage open reading frame (ORF) content patterns for bacterial phylogenetic analysis.
- To overcome the limitations of high error rates in ONT sequencing for epidemiological studies.
Main Methods:
- Whole-genome sequencing (WGS) data from Enterobacter hormaechei strains were analyzed.
- Open reading frame (ORF) presence/absence was converted into binary sequences using BLASTn.
- Phylogenetic trees were constructed from binary ORF sequences and compared to cgSNP-based trees.
Main Results:
- ORF-based phylogenetic trees showed comparable clustering and similar topologies to cgSNP-based trees.
- The Fowlkes-Mallows index confirmed the similarity between ORF trees and cgSNP trees.
- The method effectively utilizes ONT sequencing data despite its inherent error rate.
Conclusions:
- ORF-based phylogenetic analysis using ONT sequencing is a viable alternative to cgSNP methods for bacterial epidemiology.
- This approach circumvents ONT's read error issues by focusing on ORF content.
- The method's real-time capability and lower cost offer potential for point-of-care epidemiological analysis.
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