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Comparison Analysis of Different DNA Extraction Methods on Suitability for Long-Read Metagenomic Nanopore Sequencing

Authors :
Lei Zhang
Ting Chen
Ye Wang
Shengwei Zhang
Qingyu Lv
Decong Kong
Hua Jiang
Yuling Zheng
Yuhao Ren
Wenhua Huang
Peng Liu
Yongqiang Jiang
Source :
Frontiers in Cellular and Infection Microbiology, Vol 12 (2022)
Publication Year :
2022
Publisher :
Frontiers Media S.A., 2022.

Abstract

Metagenomic next-generation sequencing (mNGS) is a novel useful strategy that is increasingly used for pathogens detection in clinic. Some emerging mNGS technologies with long-read ability are useful to decrease sequencing time and increase diagnosed accuracy, which is of great significance in rapid pathogen diagnosis. Reliable DNA extraction is considered critical for the success of sequencing; hence, there is thus an urgent need of gentle DNA extraction method to get unbiased and more integrate DNA from all kinds of pathogens. In this study, we systematically compared three DNA extraction methods (enzymatic cell lysis based on MetaPolyzyme, mechanical cell lysis based on bead beating, and the control method without pre–cell lysis, respectively) by assessing DNA yield, integrity, and the microbial diversity based on long-read nanopore sequencing of urine samples with microbial infections. Compared with the control method, the enzymatic-based method increased the average length of microbial reads by a median of 2.1-fold [Inter Quartile Range (IQR), 1.7–2.5; maximum, 4.8) in 18 of the 20 samples and the mapped reads proportion of specific species by a median of 11.8-fold (Inter Quartile Range (IQR), 6.9–32.2; maximum, 79.27]. Moreover, it provided fully (20 of 20) consistent diagnosed results to the clinical culture and more representative microbial profiles (P < 0.05), which all strongly proves the excellent performance of enzymatic-based method in long-read mNGS–based pathogen identification and potential diseases diagnosis of microbiome related.

Details

Language :
English
ISSN :
22352988
Volume :
12
Database :
Directory of Open Access Journals
Journal :
Frontiers in Cellular and Infection Microbiology
Publication Type :
Academic Journal
Accession number :
edsdoj.4c9b0725a0314cc3880678c2f7148a40
Document Type :
article
Full Text :
https://doi.org/10.3389/fcimb.2022.919903