Estimation of the relative abundance of species in artificial mixtures of insects using low-coverage shotgun metagenomics

Amplicon metabarcoding is an established technique to analyse the taxonomic composition of communities of organisms using high-throughput DNA sequencing, but there are doubts about its ability to quantify the relative proportions of the species, as opposed to the species list. Here,...

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Autores principales:	Lidia Garrido-Sanz, Miquel Àngel Senar, Josep Piñol
Formato:	article
Lenguaje:	EN
Publicado:	Pensoft Publishers 2020
Materias:	Ecology QH540-549.5
Acceso en línea:	https://doaj.org/article/8b813c8cf0da43398e5349d701faffc8
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spelling	oai:doaj.org-article:8b813c8cf0da43398e5349d701faffc82021-12-02T12:18:19ZEstimation of the relative abundance of species in artificial mixtures of insects using low-coverage shotgun metagenomics10.3897/mbmg.4.482812534-9708https://doaj.org/article/8b813c8cf0da43398e5349d701faffc82020-02-01T00:00:00Zhttps://mbmg.pensoft.net/article/48281/download/pdf/https://mbmg.pensoft.net/article/48281/download/xml/https://mbmg.pensoft.net/article/48281/https://doaj.org/toc/2534-9708 Amplicon metabarcoding is an established technique to analyse the taxonomic composition of communities of organisms using high-throughput DNA sequencing, but there are doubts about its ability to quantify the relative proportions of the species, as opposed to the species list. Here, we bypass the enrichment step and avoid the PCR-bias, by directly sequencing the extracted DNA using shotgun metagenomics. This approach is common practice in prokaryotes, but not in eukaryotes, because of the low number of sequenced genomes of eukaryotic species. We tested the metagenomics approach using insect species whose genome is already sequenced and assembled to an advanced degree. We shotgun-sequenced, at low-coverage, 18 species of insects in 22 single-species and 6 mixed-species libraries and mapped the reads against 110 reference genomes of insects. We used the single-species libraries to calibrate the process of assignation of reads to species and the libraries created from species mixtures to evaluate the ability of the method to quantify the relative species abundance. Our results showed that the shotgun metagenomic method is easily able to set apart closely-related insect species, like four species of Drosophila included in the artificial libraries. However, to avoid the counting of rare misclassified reads in samples, it was necessary to use a rather stringent detection limit of 0.001, so species with a lower relative abundance are ignored. We also identified that approximately half the raw reads were informative for taxonomic purposes. Finally, using the mixed-species libraries, we showed that it was feasible to quantify with confidence the relative abundance of individual species in the mixtures. Lidia Garrido-SanzMiquel Àngel SenarJosep PiñolPensoft PublishersarticleEcologyQH540-549.5ENMetabarcoding and Metagenomics, Vol 4, Iss , Pp 1-18 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	Ecology QH540-549.5
spellingShingle	Ecology QH540-549.5 Lidia Garrido-Sanz Miquel Àngel Senar Josep Piñol Estimation of the relative abundance of species in artificial mixtures of insects using low-coverage shotgun metagenomics
description	Amplicon metabarcoding is an established technique to analyse the taxonomic composition of communities of organisms using high-throughput DNA sequencing, but there are doubts about its ability to quantify the relative proportions of the species, as opposed to the species list. Here, we bypass the enrichment step and avoid the PCR-bias, by directly sequencing the extracted DNA using shotgun metagenomics. This approach is common practice in prokaryotes, but not in eukaryotes, because of the low number of sequenced genomes of eukaryotic species. We tested the metagenomics approach using insect species whose genome is already sequenced and assembled to an advanced degree. We shotgun-sequenced, at low-coverage, 18 species of insects in 22 single-species and 6 mixed-species libraries and mapped the reads against 110 reference genomes of insects. We used the single-species libraries to calibrate the process of assignation of reads to species and the libraries created from species mixtures to evaluate the ability of the method to quantify the relative species abundance. Our results showed that the shotgun metagenomic method is easily able to set apart closely-related insect species, like four species of Drosophila included in the artificial libraries. However, to avoid the counting of rare misclassified reads in samples, it was necessary to use a rather stringent detection limit of 0.001, so species with a lower relative abundance are ignored. We also identified that approximately half the raw reads were informative for taxonomic purposes. Finally, using the mixed-species libraries, we showed that it was feasible to quantify with confidence the relative abundance of individual species in the mixtures.
format	article
author	Lidia Garrido-Sanz Miquel Àngel Senar Josep Piñol
author_facet	Lidia Garrido-Sanz Miquel Àngel Senar Josep Piñol
author_sort	Lidia Garrido-Sanz
title	Estimation of the relative abundance of species in artificial mixtures of insects using low-coverage shotgun metagenomics
title_short	Estimation of the relative abundance of species in artificial mixtures of insects using low-coverage shotgun metagenomics
title_full	Estimation of the relative abundance of species in artificial mixtures of insects using low-coverage shotgun metagenomics
title_fullStr	Estimation of the relative abundance of species in artificial mixtures of insects using low-coverage shotgun metagenomics
title_full_unstemmed	Estimation of the relative abundance of species in artificial mixtures of insects using low-coverage shotgun metagenomics
title_sort	estimation of the relative abundance of species in artificial mixtures of insects using low-coverage shotgun metagenomics
publisher	Pensoft Publishers
publishDate	2020
url	https://doaj.org/article/8b813c8cf0da43398e5349d701faffc8
work_keys_str_mv	AT lidiagarridosanz estimationoftherelativeabundanceofspeciesinartificialmixturesofinsectsusinglowcoverageshotgunmetagenomics AT miquelangelsenar estimationoftherelativeabundanceofspeciesinartificialmixturesofinsectsusinglowcoverageshotgunmetagenomics AT joseppinol estimationoftherelativeabundanceofspeciesinartificialmixturesofinsectsusinglowcoverageshotgunmetagenomics
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Estimation of the relative abundance of species in artificial mixtures of insects using low-coverage shotgun metagenomics

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