This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Methodology
Evaluation of Methods to Optimise Shotgun Metagenomic Sequencing-Based Analysis of the Microbiome of Milk
Paul Cotter
Teagasc
Corresponding Author
ORCiD: https://orcid.org/0000-0002-2577-9028
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: Shotgun metagenomic sequencing is a valuable tool for the taxonomic and functional profiling of microbial communities. However, this approach is challenging in samples, such as milk, where a low microbial abundance, combined with high levels of host DNA, result in inefficient and uneconomical sequencing.
Results: Here we evaluate approaches to deplete host DNA or enrich microbial DNA prior to sequencing using three commercially available kits. We compared the percentage of microbial reads obtained from each kit after shotgun metagenomic sequencing. Using bovine and human milk samples, we determined that host depletion with the MolYsis complete5 kit significantly improved microbial sequencing depth compared to other approaches. Importantly, no biases were introduced. Additionally, the increased microbial sequencing depth allowed for further characterization of the microbiome through the generation of metagenome-assembled genomes (MAGs).
Conclusions: This evaluation shows that microbiome analysis can be performed on both bovine and human milk samples at a much greater resolution without the need for more expensive deep-sequencing approaches.
Keywords
Shotgun metagenomics, Milk microbiome, Host depletion, Bioinformatics
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
This is a list of supplementary files associated with this preprint. Click to download.
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 14 May, 2020
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
General Microbiology
Learn more about our company.
This is a preprint. It has not completed peer review.
Methodology
Evaluation of Methods to Optimise Shotgun Metagenomic Sequencing-Based Analysis of the Microbiome of Milk
Paul Cotter
Teagasc
Corresponding Author
ORCiD: https://orcid.org/0000-0002-2577-9028
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 14 May, 2020
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
General Microbiology
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: Shotgun metagenomic sequencing is a valuable tool for the taxonomic and functional profiling of microbial communities. However, this approach is challenging in samples, such as milk, where a low microbial abundance, combined with high levels of host DNA, result in inefficient and uneconomical sequencing.
Results: Here we evaluate approaches to deplete host DNA or enrich microbial DNA prior to sequencing using three commercially available kits. We compared the percentage of microbial reads obtained from each kit after shotgun metagenomic sequencing. Using bovine and human milk samples, we determined that host depletion with the MolYsis complete5 kit significantly improved microbial sequencing depth compared to other approaches. Importantly, no biases were introduced. Additionally, the increased microbial sequencing depth allowed for further characterization of the microbiome through the generation of metagenome-assembled genomes (MAGs).
Conclusions: This evaluation shows that microbiome analysis can be performed on both bovine and human milk samples at a much greater resolution without the need for more expensive deep-sequencing approaches.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5