Multilaboratory assessment of metagenomic next-generation sequencing for unbiased microbe detection

Han, Dongsheng; Diao, Zhenli; Lai, Huiying; Han, Yanxi; Xie, Jiehong; Zhang, Rui; Li, Jinming

doi:10.1016/j.jare.2021.09.011

Cited by 16 publications

(12 citation statements)

References 41 publications

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“…As with all new technologies, there are still significant difficulties and challenges in interpreting and reporting the data. Therefore, there is a high variability between laboratories in identifying microorganisms and true pathogens (39). This is related not only to different laboratory environments and sequencing platforms but also to the expertise and experience of interpreters.…”

Section: Discussionmentioning

confidence: 99%

Clinical evaluation of metagenomic next-generation sequencing for detecting pathogens in bronchoalveolar lavage fluid collected from children with community-acquired pneumonia

et al. 2022

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This study is to evaluate the usefulness of pathogen detection using metagenomic next-generation sequencing (mNGS) on bronchoalveolar lavage fluid (BALF) specimens from children with community-acquired pneumonia (CAP). We retrospectively collected BALF specimens from 121 children with CAP at Tianjin Children's Hospital from February 2021 to December 2021. The diagnostic performances of mNGS and conventional tests (CT) (culture and targeted polymerase chain reaction tests) were compared, using composite diagnosis as the reference standard. The results of mNGS and CT were compared based on pathogenic and non-pathogenic organisms. Pathogen profiles and co-infections between the mild CAP and severe CAP groups were also analyzed. The overall positive coincidence rate was 86.78% (105/121) for mNGS and 66.94% (81/121) for CT. The proportion of patients diagnosed using mNGS plus CT increased to 99.18%. Among the patients, 17.36% were confirmed only by mNGS; Streptococcus pneumoniae accounted for 52.38% and 23.8% of the patients were co-infected. Moreover, Bordetella pertussis and Human bocavirus (HBoV) were detected only using mNGS. Mycoplasma pneumoniae, which was identified in 89 (73.55%) of 121 children with CAP, was the most frequent pathogen detected using mNGS. The infection rate of M. pneumoniae in the severe CAP group was significantly higher than that in the mild CAP group (P = 0.007). The symptoms of single bacterial infections (except for mycoplasma) were milder than those of mycoplasma infections. mNGS identified more bacterial infections when compared to the CT methods and was able to identify co-infections which were initially missed on CT. Additionally, it was able to identify pathogens that were beyond the scope of the CT methods. The mNGS method is a powerful supplement to clinical diagnostic tools in respiratory infections, as it can increase the precision of diagnosis and guide the use of antibiotics.

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Section: Discussionmentioning

confidence: 99%

Clinical evaluation of metagenomic next-generation sequencing for detecting pathogens in bronchoalveolar lavage fluid collected from children with community-acquired pneumonia

et al. 2022

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“…Host depletion methods, such as differential lysis method, can filter human DNA ( Ji et al., 2020 ), increasing pathogen DNA ratio ( Thoendel et al., 2018 ; Ji et al., 2020 ; Gu et al., 2021 ) at the expense of some viruses, parasites, and bacteria ( Ji et al., 2020 ). Besides, host depletion methods lose the cfDNA in supernatant, bring in contamination of engineered strains from reagents ( Gu et al., 2021 ), and decrease the detection rates of pathogens ( Han et al., 2021 ). The long-held conflict in location of proliferation and infection between intracellular and extracellular pathogens ( Casadevall and Fang, 2020 ) challenged whether mNGS of cfDNA can accurately detect causative pathogens.…”

Section: Discussionmentioning

confidence: 99%

“…We propose that the better performance of mNGS using cfDNA is attributed to its advantage in DNA extraction and bioinformatics analysis for trace pathogens. Although sample pretreatment for host depletion is not included in this study, wcDNA extraction involves cell wall lysis ( Wilson et al., 2019 ), increasing the risk of DNA degradation ( Thoendel et al., 2018 ; Gu et al., 2021 ) and decreasing DNA recovery rate of pathogens with low loads ( Han et al., 2021 ), rather than high loads. Mild extraction of cfDNA without cell wall lysis should have few effect on the DNA recovery, which has been confirmed by successful detection of trace pathogens from cerebrospinal fluid using mNGS of cfDNA ( Ji et al., 2020 ).…”

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

“…mNGS of cfDNA can be fully competent for detecting fungal and intracellular pathogens, challenging the opinion that process of cell wall lysis, such as bead-beating process, is necessary for DNA extraction (wcDNA) to ensure mNGS detection for those pathogens ( Casadevall and Fang, 2020 ; Han et al., 2021 ). Adding bead-beating process was reported to significantly improve the detection of Aspergillus fumigatus by mNGS of wcDNA ( Han et al., 2021 ). However, with the host immune attack ( Arciola et al., 2018 ) and microbial autolysis ( Wolf et al., 2017 ), DNA of intracellular and fungal pathogens can be released into body fluids in the form of cfDNA ( Casadevall and Fang, 2020 ).…”

Section: Discussionmentioning

confidence: 99%

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Comparison of metagenomic next-generation sequencing using cell-free DNA and whole-cell DNA for the diagnoses of pulmonary infections

Wang²,

et al. 2022

Front. Cell. Infect. Microbiol.

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Although the fast-growing metagenomic next-generation sequencing (mNGS) has been used in diagnosing infectious diseases, low detection rate of mNGS in detecting pathogens with low loads limits its extensive application. In this study, 130 patients with suspected pulmonary infections were enrolled, from whom bronchoalveolar lavage fluid (BALF) samples were collected. The conventional tests and mNGS of cell-free DNA (cfDNA) and whole-cell DNA (wcDNA) using BALF were simultaneously performed. mNGS of cfDNA showed higher detection rate (91.5%) and total coincidence rate (73.8%) than mNGS of wcDNA (83.1% and 63.9%) and conventional methods (26.9% and 30.8%). A total of 70 microbes were detected by mNGS of cfDNA, and most of them (60) were also identified by mNGS of wcDNA. The 31.8% (21/66) of fungi, 38.6% (27/70) of viruses, and 26.7% (8/30) of intracellular microbes can be only detected by mNGS of cfDNA, much higher than those [19.7% (13/66), 14.3% (10/70), and 6.7% (2/30)] only detected by mNGS of wcDNA. After in-depth analysis on these microbes with low loads set by reads per million (RPM), we found that more RPM and fungi/viruses/intracellular microbes were detected by mNGS of cfDNA than by mNGS of wcDNA. Besides, the abilities of mNGS using both cfDNA and wcDNA to detect microbes with high loads were similar. We highlighted the advantage of mNGS using cfDNA in detecting fungi, viruses, and intracellular microbes with low loads, and suggested that mNGS of cfDNA could be considered as the first choice for diagnosing pulmonary infections.

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PHDtools: A platform for pathogen detection and multi-dimensional genetic signatures decoding to realize pathogen genomics data analyses online

Xiong,

Zhang,

et al. 2024

Gene

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Multilaboratory assessment of metagenomic next-generation sequencing for unbiased microbe detection

Cited by 16 publications

References 41 publications

Clinical evaluation of metagenomic next-generation sequencing for detecting pathogens in bronchoalveolar lavage fluid collected from children with community-acquired pneumonia

Clinical evaluation of metagenomic next-generation sequencing for detecting pathogens in bronchoalveolar lavage fluid collected from children with community-acquired pneumonia

Comparison of metagenomic next-generation sequencing using cell-free DNA and whole-cell DNA for the diagnoses of pulmonary infections

PHDtools: A platform for pathogen detection and multi-dimensional genetic signatures decoding to realize pathogen genomics data analyses online

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