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A Protocol for Extraction of Infective Viromes Suitable for Metagenomics Sequencing from Low Volume Fecal Samples

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Deng, Ling ; Silins, Ronalds ; Castro-Mejía, Josué L ; Kot, Witold ; Jessen, Leon ; Thorsen, Jonathan ; Shah, Shiraz ; Stokholm, Jakob ; Bisgaard, Hans ; Moineau, Sylvain ; Nielsen, Dennis Sandris. / A Protocol for Extraction of Infective Viromes Suitable for Metagenomics Sequencing from Low Volume Fecal Samples. I: Viruses. 2019 ; Bind 11, Nr. 7.

Bibtex

@article{28da106b0756426ea5b54a0239897244,
title = "A Protocol for Extraction of Infective Viromes Suitable for Metagenomics Sequencing from Low Volume Fecal Samples",
abstract = "The human gut microbiome (GM) plays an important role in human health and diseases. However, while substantial progress has been made in understanding the role of bacterial inhabitants of the gut, much less is known regarding the viral component of the GM. Bacteriophages (phages) are viruses attacking specific host bacteria and likely play important roles in shaping the GM. Although metagenomic approaches have led to the discoveries of many new viruses, they remain largely uncultured as their hosts have not been identified, which hampers our understanding of their biological roles. Existing protocols for isolation of viromes generally require relatively high input volumes and are generally more focused on extracting nucleic acids of good quality and purity for down-stream analysis, and less on purifying viruses with infective capacity. In this study, we report the development of an efficient protocol requiring low sample input yielding purified viromes containing phages that are still infective, which also are of sufficient purity for genome sequencing. We validated the method through spiking known phages followed by plaque assays, qPCR, and metagenomic sequencing. The protocol should facilitate the process of culturing novel viruses from the gut as well as large scale studies on gut viromes.",
author = "Ling Deng and Ronalds Silins and Castro-Mej{\'i}a, {Josu{\'e} L} and Witold Kot and Leon Jessen and Jonathan Thorsen and Shiraz Shah and Jakob Stokholm and Hans Bisgaard and Sylvain Moineau and Nielsen, {Dennis Sandris}",
year = "2019",
month = "7",
day = "20",
doi = "10.3390/v11070667",
language = "English",
volume = "11",
journal = "Viruses",
issn = "1999-4915",
publisher = "M D P I AG",
number = "7",

}

RIS

TY - JOUR

T1 - A Protocol for Extraction of Infective Viromes Suitable for Metagenomics Sequencing from Low Volume Fecal Samples

AU - Deng, Ling

AU - Silins, Ronalds

AU - Castro-Mejía, Josué L

AU - Kot, Witold

AU - Jessen, Leon

AU - Thorsen, Jonathan

AU - Shah, Shiraz

AU - Stokholm, Jakob

AU - Bisgaard, Hans

AU - Moineau, Sylvain

AU - Nielsen, Dennis Sandris

PY - 2019/7/20

Y1 - 2019/7/20

N2 - The human gut microbiome (GM) plays an important role in human health and diseases. However, while substantial progress has been made in understanding the role of bacterial inhabitants of the gut, much less is known regarding the viral component of the GM. Bacteriophages (phages) are viruses attacking specific host bacteria and likely play important roles in shaping the GM. Although metagenomic approaches have led to the discoveries of many new viruses, they remain largely uncultured as their hosts have not been identified, which hampers our understanding of their biological roles. Existing protocols for isolation of viromes generally require relatively high input volumes and are generally more focused on extracting nucleic acids of good quality and purity for down-stream analysis, and less on purifying viruses with infective capacity. In this study, we report the development of an efficient protocol requiring low sample input yielding purified viromes containing phages that are still infective, which also are of sufficient purity for genome sequencing. We validated the method through spiking known phages followed by plaque assays, qPCR, and metagenomic sequencing. The protocol should facilitate the process of culturing novel viruses from the gut as well as large scale studies on gut viromes.

AB - The human gut microbiome (GM) plays an important role in human health and diseases. However, while substantial progress has been made in understanding the role of bacterial inhabitants of the gut, much less is known regarding the viral component of the GM. Bacteriophages (phages) are viruses attacking specific host bacteria and likely play important roles in shaping the GM. Although metagenomic approaches have led to the discoveries of many new viruses, they remain largely uncultured as their hosts have not been identified, which hampers our understanding of their biological roles. Existing protocols for isolation of viromes generally require relatively high input volumes and are generally more focused on extracting nucleic acids of good quality and purity for down-stream analysis, and less on purifying viruses with infective capacity. In this study, we report the development of an efficient protocol requiring low sample input yielding purified viromes containing phages that are still infective, which also are of sufficient purity for genome sequencing. We validated the method through spiking known phages followed by plaque assays, qPCR, and metagenomic sequencing. The protocol should facilitate the process of culturing novel viruses from the gut as well as large scale studies on gut viromes.

U2 - 10.3390/v11070667

DO - 10.3390/v11070667

M3 - Journal article

VL - 11

JO - Viruses

JF - Viruses

SN - 1999-4915

IS - 7

ER -

ID: 57877993