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Comparison of targeted metagenomics and IS-Pro methods for analysing the lung microbiome
BACKGROUND: Targeted metagenomics and IS-Pro method are two of the many methods that have been used to study the microbiome. The two methods target different regions of the 16 S rRNA gene. The aim of this study was to compare targeted metagenomics and IS-Pro methods for the ability to discern the mi...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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BioMed Central
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8371770/ https://www.ncbi.nlm.nih.gov/pubmed/34407769 http://dx.doi.org/10.1186/s12866-021-02288-x |
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author | Goolam Mahomed, T Peters, RPH Pretorius, GHJ Goolam Mahomed, A Ueckermann, V Kock, MM Ehlers, MM |
author_facet | Goolam Mahomed, T Peters, RPH Pretorius, GHJ Goolam Mahomed, A Ueckermann, V Kock, MM Ehlers, MM |
author_sort | Goolam Mahomed, T |
collection | PubMed |
description | BACKGROUND: Targeted metagenomics and IS-Pro method are two of the many methods that have been used to study the microbiome. The two methods target different regions of the 16 S rRNA gene. The aim of this study was to compare targeted metagenomics and IS-Pro methods for the ability to discern the microbial composition of the lung microbiome of COPD patients. METHODS: Spontaneously expectorated sputum specimens were collected from COPD patients. Bacterial DNA was extracted and used for targeted metagenomics and IS-Pro method. The analysis was performed using QIIME2 (targeted metagenomics) and IS-Pro software (IS-Pro method). Additionally, a laboratory cost per isolate and time analysis was performed for each method. RESULTS: Statistically significant differences were observed in alpha diversity when targeted metagenomics and IS-Pro methods’ data were compared using the Shannon diversity measure (p-value = 0.0006) but not with the Simpson diversity measure (p-value = 0.84). Distinct clusters with no overlap between the two technologies were observed for beta diversity. Targeted metagenomics had a lower relative abundance of phyla, such as the Proteobacteria, and higher relative abundance of phyla, such as Firmicutes when compared to the IS-Pro method. Haemophilus, Prevotella and Streptococcus were most prevalent genera across both methods. Targeted metagenomics classified 23 % (144/631) of OTUs to a species level, whereas IS-Pro method classified 86 % (55/64) of OTUs to a species level. However, unclassified OTUs accounted for a higher relative abundance when using the IS-Pro method (35 %) compared to targeted metagenomics (5 %). The two methods performed comparably in terms of cost and time; however, the IS-Pro method was more user-friendly. CONCLUSIONS: It is essential to understand the value of different methods for characterisation of the microbiome. Targeted metagenomics and IS-Pro methods showed differences in ability in identifying and characterising OTUs, diversity and microbial composition of the lung microbiome. The IS-Pro method might miss relevant species and could inflate the abundance of Proteobacteria. However, the IS-Pro kit identified most of the important lung pathogens, such as Burkholderia and Pseudomonas and may work in a more diagnostics-orientated setting. Both methods were comparable in terms of cost and time; however, the IS-Pro method was easier to use. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12866-021-02288-x. |
format | Online Article Text |
id | pubmed-8371770 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-83717702021-08-18 Comparison of targeted metagenomics and IS-Pro methods for analysing the lung microbiome Goolam Mahomed, T Peters, RPH Pretorius, GHJ Goolam Mahomed, A Ueckermann, V Kock, MM Ehlers, MM BMC Microbiol Research BACKGROUND: Targeted metagenomics and IS-Pro method are two of the many methods that have been used to study the microbiome. The two methods target different regions of the 16 S rRNA gene. The aim of this study was to compare targeted metagenomics and IS-Pro methods for the ability to discern the microbial composition of the lung microbiome of COPD patients. METHODS: Spontaneously expectorated sputum specimens were collected from COPD patients. Bacterial DNA was extracted and used for targeted metagenomics and IS-Pro method. The analysis was performed using QIIME2 (targeted metagenomics) and IS-Pro software (IS-Pro method). Additionally, a laboratory cost per isolate and time analysis was performed for each method. RESULTS: Statistically significant differences were observed in alpha diversity when targeted metagenomics and IS-Pro methods’ data were compared using the Shannon diversity measure (p-value = 0.0006) but not with the Simpson diversity measure (p-value = 0.84). Distinct clusters with no overlap between the two technologies were observed for beta diversity. Targeted metagenomics had a lower relative abundance of phyla, such as the Proteobacteria, and higher relative abundance of phyla, such as Firmicutes when compared to the IS-Pro method. Haemophilus, Prevotella and Streptococcus were most prevalent genera across both methods. Targeted metagenomics classified 23 % (144/631) of OTUs to a species level, whereas IS-Pro method classified 86 % (55/64) of OTUs to a species level. However, unclassified OTUs accounted for a higher relative abundance when using the IS-Pro method (35 %) compared to targeted metagenomics (5 %). The two methods performed comparably in terms of cost and time; however, the IS-Pro method was more user-friendly. CONCLUSIONS: It is essential to understand the value of different methods for characterisation of the microbiome. Targeted metagenomics and IS-Pro methods showed differences in ability in identifying and characterising OTUs, diversity and microbial composition of the lung microbiome. The IS-Pro method might miss relevant species and could inflate the abundance of Proteobacteria. However, the IS-Pro kit identified most of the important lung pathogens, such as Burkholderia and Pseudomonas and may work in a more diagnostics-orientated setting. Both methods were comparable in terms of cost and time; however, the IS-Pro method was easier to use. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12866-021-02288-x. BioMed Central 2021-08-18 /pmc/articles/PMC8371770/ /pubmed/34407769 http://dx.doi.org/10.1186/s12866-021-02288-x Text en © The Author(s) 2021 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) . The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/ (https://creativecommons.org/publicdomain/zero/1.0/) ) applies to the data made available in this article, unless otherwise stated in a credit line to the data. |
spellingShingle | Research Goolam Mahomed, T Peters, RPH Pretorius, GHJ Goolam Mahomed, A Ueckermann, V Kock, MM Ehlers, MM Comparison of targeted metagenomics and IS-Pro methods for analysing the lung microbiome |
title | Comparison of targeted metagenomics and IS-Pro methods for analysing the lung microbiome |
title_full | Comparison of targeted metagenomics and IS-Pro methods for analysing the lung microbiome |
title_fullStr | Comparison of targeted metagenomics and IS-Pro methods for analysing the lung microbiome |
title_full_unstemmed | Comparison of targeted metagenomics and IS-Pro methods for analysing the lung microbiome |
title_short | Comparison of targeted metagenomics and IS-Pro methods for analysing the lung microbiome |
title_sort | comparison of targeted metagenomics and is-pro methods for analysing the lung microbiome |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8371770/ https://www.ncbi.nlm.nih.gov/pubmed/34407769 http://dx.doi.org/10.1186/s12866-021-02288-x |
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