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Optimizing protocols for extraction of bacteriophages prior to metagenomic analyses of phage communities in the human gut

Overview of attention for article published in Microbiome, November 2015
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  • In the top 25% of all research outputs scored by Altmetric
  • High Attention Score compared to outputs of the same age (83rd percentile)
  • Average Attention Score compared to outputs of the same age and source

Mentioned by

twitter
14 tweeters
patent
1 patent

Citations

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85 Dimensions

Readers on

mendeley
299 Mendeley
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2 CiteULike
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Title
Optimizing protocols for extraction of bacteriophages prior to metagenomic analyses of phage communities in the human gut
Published in
Microbiome, November 2015
DOI 10.1186/s40168-015-0131-4
Pubmed ID
Authors

Josué L. Castro-Mejía, Musemma K. Muhammed, Witold Kot, Horst Neve, Charles M. A. P. Franz, Lars H. Hansen, Finn K. Vogensen, Dennis S. Nielsen

Abstract

The human gut is densely populated with archaea, eukaryotes, bacteria, and their viruses, such as bacteriophages. Advances in high-throughput sequencing (HTS) as well as bioinformatics have opened new opportunities for characterizing the viral communities harbored in our gut. However, limited attention has been given to the efficiency of protocols dealing with extraction of phages from fecal communities prior to HTS and their impact on the metagenomic dataset. We describe two optimized methods for extraction of phages from fecal samples based on tangential-flow filtration (TFF) and polyethylene glycol precipitation (PEG) approaches using an adapted method from a published protocol as control (literature-adapted protocol (LIT)). To quantify phage recovery, samples were spiked with low numbers of c2, ϕ29, and T4 phages (representatives of the Siphoviridae, Podoviridae, and Myoviridae families, respectively) and their concentration (plaque-forming units) followed at every step during the extraction procedure. Compared with LIT, TFF and PEG had higher recovery of all spiked phages, yielding up to 16 times more phage particles (PPs) and up to 68 times more phage DNA per volume, increasing thus the chances of extracting low abundant phages. TFF- and PEG-derived metaviromes showed 10 % increase in relative abundance of Caudovirales and unclassified phages infecting gut-associated bacteria (>92 % for TFF and PEG, 82.4 % for LIT). Our methods obtained lower relative abundance of the Myoviridae family (<16 %) as compared to the reference protocol (22 %). This decline, however, was not considered a true loss of Myoviridae phages but rather a greater level of extraction of Siphoviridae phages (TFF and PEG >32.5 %, LIT 22.6 %), which was achieved with the enhanced conditions of our procedures (e.g., reduced filter clogging). A high degree of phage diversity in samples extracted using TFF and PEG was documented by transmission electron microscopy. Two procedures (TFF and PEG) for extraction of bacteriophages from fecal samples were optimized using a set of spiked bacteriophages as process control. These protocols are highly efficient tools for extraction and purification of PPs prior to HTS in phage-metavirome studies. Our methods can be easily modified, being thus applicable and adjustable for in principle any solid environmental material in dissolution.

Twitter Demographics

The data shown below were collected from the profiles of 14 tweeters who shared this research output. Click here to find out more about how the information was compiled.

Mendeley readers

The data shown below were compiled from readership statistics for 299 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
Brazil 2 <1%
Denmark 1 <1%
Canada 1 <1%
United States 1 <1%
Unknown 294 98%

Demographic breakdown

Readers by professional status Count As %
Student > Ph. D. Student 73 24%
Student > Master 47 16%
Researcher 42 14%
Student > Bachelor 34 11%
Student > Doctoral Student 14 5%
Other 39 13%
Unknown 50 17%
Readers by discipline Count As %
Agricultural and Biological Sciences 89 30%
Biochemistry, Genetics and Molecular Biology 72 24%
Immunology and Microbiology 33 11%
Engineering 11 4%
Medicine and Dentistry 7 2%
Other 24 8%
Unknown 63 21%

Attention Score in Context

This research output has an Altmetric Attention Score of 9. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 02 September 2021.
All research outputs
#3,063,123
of 19,529,814 outputs
Outputs from Microbiome
#947
of 1,186 outputs
Outputs of similar age
#63,431
of 388,370 outputs
Outputs of similar age from Microbiome
#53
of 79 outputs
Altmetric has tracked 19,529,814 research outputs across all sources so far. Compared to these this one has done well and is in the 84th percentile: it's in the top 25% of all research outputs ever tracked by Altmetric.
So far Altmetric has tracked 1,186 research outputs from this source. They typically receive a lot more attention than average, with a mean Attention Score of 40.0. This one is in the 20th percentile – i.e., 20% of its peers scored the same or lower than it.
Older research outputs will score higher simply because they've had more time to accumulate mentions. To account for age we can compare this Altmetric Attention Score to the 388,370 tracked outputs that were published within six weeks on either side of this one in any source. This one has done well, scoring higher than 83% of its contemporaries.
We're also able to compare this research output to 79 others from the same source and published within six weeks on either side of this one. This one is in the 34th percentile – i.e., 34% of its contemporaries scored the same or lower than it.