Pros and Cons of Ion-Torrent Next Generation Sequencing versus Terminal Restriction Fragment Length Polymorphism T-RFLP for Studying the Rumen Bacterial Community
The development of next generation sequencing has challenged the use of other molecular fingerprinting methods used to study microbial diversity. We analysed the bacterial diversity in the rumen of defaunated sheep following the introduction of different protozoal populations, using both next genera...
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| Vydané v: | PloS one Ročník 9; číslo 7; s. e101435 |
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| Hlavní autori: | , , , , , |
| Médium: | Journal Article |
| Jazyk: | English |
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United States
Public Library of Science
22.07.2014
Public Library of Science (PLoS) |
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| ISSN: | 1932-6203, 1932-6203 |
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| Abstract | The development of next generation sequencing has challenged the use of other molecular fingerprinting methods used to study microbial diversity. We analysed the bacterial diversity in the rumen of defaunated sheep following the introduction of different protozoal populations, using both next generation sequencing (NGS: Ion Torrent PGM) and terminal restriction fragment length polymorphism (T-RFLP). Although absolute number differed, there was a high correlation between NGS and T-RFLP in terms of richness and diversity with R values of 0.836 and 0.781 for richness and Shannon-Wiener index, respectively. Dendrograms for both datasets were also highly correlated (Mantel test = 0.742). Eighteen OTUs and ten genera were significantly impacted by the addition of rumen protozoa, with an increase in the relative abundance of Prevotella, Bacteroides and Ruminobacter, related to an increase in free ammonia levels in the rumen. Our findings suggest that classic fingerprinting methods are still valuable tools to study microbial diversity and structure in complex environments but that NGS techniques now provide cost effect alternatives that provide a far greater level of information on the individual members of the microbial population. |
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| AbstractList | The development of next generation sequencing has challenged the use of other molecular fingerprinting methods used to study microbial diversity. We analysed the bacterial diversity in the rumen of defaunated sheep following the introduction of different protozoal populations, using both next generation sequencing (NGS: Ion Torrent PGM) and terminal restriction fragment length polymorphism (T-RFLP). Although absolute number differed, there was a high correlation between NGS and T-RFLP in terms of richness and diversity with R values of 0.836 and 0.781 for richness and Shannon-Wiener index, respectively. Dendrograms for both datasets were also highly correlated (Mantel test = 0.742). Eighteen OTUs and ten genera were significantly impacted by the addition of rumen protozoa, with an increase in the relative abundance of Prevotella, Bacteroides and Ruminobacter, related to an increase in free ammonia levels in the rumen. Our findings suggest that classic fingerprinting methods are still valuable tools to study microbial diversity and structure in complex environments but that NGS techniques now provide cost effect alternatives that provide a far greater level of information on the individual members of the microbial population. The development of next generation sequencing has challenged the use of other molecular fingerprinting methods used to study microbial diversity. We analysed the bacterial diversity in the rumen of defaunated sheep following the introduction of different protozoal populations, using both next generation sequencing (NGS: Ion Torrent PGM) and terminal restriction fragment length polymorphism (T-RFLP). Although absolute number differed, there was a high correlation between NGS and T-RFLP in terms of richness and diversity with R values of 0.836 and 0.781 for richness and Shannon-Wiener index, respectively. Dendrograms for both datasets were also highly correlated (Mantel test = 0.742). Eighteen OTUs and ten genera were significantly impacted by the addition of rumen protozoa, with an increase in the relative abundance of Prevotella, Bacteroides and Ruminobacter, related to an increase in free ammonia levels in the rumen. Our findings suggest that classic fingerprinting methods are still valuable tools to study microbial diversity and structure in complex environments but that NGS techniques now provide cost effect alternatives that provide a far greater level of information on the individual members of the microbial population.The development of next generation sequencing has challenged the use of other molecular fingerprinting methods used to study microbial diversity. We analysed the bacterial diversity in the rumen of defaunated sheep following the introduction of different protozoal populations, using both next generation sequencing (NGS: Ion Torrent PGM) and terminal restriction fragment length polymorphism (T-RFLP). Although absolute number differed, there was a high correlation between NGS and T-RFLP in terms of richness and diversity with R values of 0.836 and 0.781 for richness and Shannon-Wiener index, respectively. Dendrograms for both datasets were also highly correlated (Mantel test = 0.742). Eighteen OTUs and ten genera were significantly impacted by the addition of rumen protozoa, with an increase in the relative abundance of Prevotella, Bacteroides and Ruminobacter, related to an increase in free ammonia levels in the rumen. Our findings suggest that classic fingerprinting methods are still valuable tools to study microbial diversity and structure in complex environments but that NGS techniques now provide cost effect alternatives that provide a far greater level of information on the individual members of the microbial population. |
| Audience | Academic |
| Author | Belanche, Alejandro Wilkinson, Toby Girwood, Susan E. de la Fuente, Gabriel Pinloche, Eric Newbold, C. Jamie |
| AuthorAffiliation | Graz University of Technology (TU Graz), Austria Institute of Biological Environmental and Rural Sciences, Aberystwyth University, Aberystwyth, Ceredigion, United Kingdom |
| AuthorAffiliation_xml | – name: Graz University of Technology (TU Graz), Austria – name: Institute of Biological Environmental and Rural Sciences, Aberystwyth University, Aberystwyth, Ceredigion, United Kingdom |
| Author_xml | – sequence: 1 givenname: Gabriel surname: de la Fuente fullname: de la Fuente, Gabriel – sequence: 2 givenname: Alejandro surname: Belanche fullname: Belanche, Alejandro – sequence: 3 givenname: Susan E. surname: Girwood fullname: Girwood, Susan E. – sequence: 4 givenname: Eric surname: Pinloche fullname: Pinloche, Eric – sequence: 5 givenname: Toby surname: Wilkinson fullname: Wilkinson, Toby – sequence: 6 givenname: C. Jamie surname: Newbold fullname: Newbold, C. Jamie |
| BackLink | https://www.ncbi.nlm.nih.gov/pubmed/25051490$$D View this record in MEDLINE/PubMed |
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| Copyright | COPYRIGHT 2014 Public Library of Science 2014 de la Fuente et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License: http://creativecommons.org/licenses/by/4.0/ (the “License”), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License. 2014 de la Fuente et al 2014 de la Fuente et al |
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| Notes | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 14 ObjectType-Article-2 ObjectType-Feature-1 content type line 23 Competing Interests: The authors have declared that no competing interests exist. Conceived and designed the experiments: GF AB CJN. Performed the experiments: GF AB. Analyzed the data: GF AB SEG TW EP. Wrote the paper: GF AB EP SEG CJN. |
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22808168 - PLoS One. 2012;7(7):e40467 |
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| SubjectTerms | Ammonia Animals Base Sequence Biology and Life Sciences Cattle Chemical fingerprinting Ciliophora - physiology Diet DNA fingerprinting DNA, Bacterial - genetics Ecosystems Fermentation Gene polymorphism High-Throughput Nucleotide Sequencing Microbiota - genetics Microorganisms Nutrition research Polymorphism Polymorphism, Restriction Fragment Length Protozoa Relative abundance Restriction fragment length polymorphism Rumen Rumen - microbiology Sequence Analysis, DNA Sheep Sheep, Domestic - microbiology |
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| Title | Pros and Cons of Ion-Torrent Next Generation Sequencing versus Terminal Restriction Fragment Length Polymorphism T-RFLP for Studying the Rumen Bacterial Community |
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