Methanogenic Archaea Quantification in the Human Gut Microbiome with F420 Autofluorescence-Based Flow Cytometry
Abstract
:1. Introduction
2. Materials and Methods
2.1. Validating Flow Cytometry to Identify Methanogenic Archaea in the Human Gut Microbiome Based on F420 Autofluorescence with a Methanobrevibacter Smithii Type Strain
2.1.1. Methanobrevibacter smithii Cultivation and Quantification
2.1.2. Preparation of Faecal Slurries
2.1.3. Flow Cytometry F420 Autofluorescence-Based Detection and Quantification of the Methanogenic Archaea M. smithii in a Spiked Faecal Sample
2.2. Benchmarking Flow Cytometry to Quantify Endogenous Methanogenic Archaea Based on F420 Autofluorescence against 16S rRNA Gene Amplicon Sequencing
2.2.1. Applying Flow Cytometry to Quantify Endogenous Methanogenic Archaea in Faecal Samples of 10 Individuals Based on F420 Autofluorescence
2.2.2. Applying Universal and Archaeal-Specific 16S rRNA Gene Amplicon Sequencing to Detect Endogenous Methanogenic Archaea in Faecal Samples of 10 Individuals
2.3. Benchmarking Flow Cytometry to Quantify Methanogenic Archaea in In Vitro Reactor Samples Based on F420 Autofluorescence against Methane Production Measurements and 16S rRNA Gene Amplicon Sequencing
2.3.1. Applying Flow Cytometry to Quantify Endogenous Methanogenic Archaea in SHIME Samples of 5 Individuals Based on F420 Autofluorescence
2.3.2. Applying Universal 16S rRNA Gene Amplicon Sequencing to Quantify Endogenous Methanogenic Archaea in SHIME Samples of 5 Individuals
2.3.3. Measuring Methane in SHIME Headspace Samples of 5 Individuals
2.4. Bioinformatics and Statistical Analysis
2.5. Ethical Approval
3. Results
3.1. Detection and Quantification of Endogenous Methanogens Is Consistent between Flow Cytometry and 16S rRNA Gene Amplicon Sequencing in Faecal Samples and In Vitro Incubated Faecal Samples
3.2. Methanogens Clustered according to In Vitro Methane Production and Methanogenic Archaea Presence in In Vitro Incubated Faecal Samples
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Minnebo, Y.; De Paepe, K.; Props, R.; Lacoere, T.; Boon, N.; Van de Wiele, T. Methanogenic Archaea Quantification in the Human Gut Microbiome with F420 Autofluorescence-Based Flow Cytometry. Appl. Microbiol. 2024, 4, 162-180. https://doi.org/10.3390/applmicrobiol4010012
Minnebo Y, De Paepe K, Props R, Lacoere T, Boon N, Van de Wiele T. Methanogenic Archaea Quantification in the Human Gut Microbiome with F420 Autofluorescence-Based Flow Cytometry. Applied Microbiology. 2024; 4(1):162-180. https://doi.org/10.3390/applmicrobiol4010012
Chicago/Turabian StyleMinnebo, Yorick, Kim De Paepe, Ruben Props, Tim Lacoere, Nico Boon, and Tom Van de Wiele. 2024. "Methanogenic Archaea Quantification in the Human Gut Microbiome with F420 Autofluorescence-Based Flow Cytometry" Applied Microbiology 4, no. 1: 162-180. https://doi.org/10.3390/applmicrobiol4010012