Dataset Information

Evaluating metabarcoding to analyse diet composition of species foraging in anthropogenic landscapes using Ion Torrent and Illumina sequencing.

ABSTRACT: DNA metabarcoding of faecal samples is being successfully used to study the foraging niche of species. We assessed the ability of two benchtop high-throughput sequencing (HTS) platforms, to identify a large taxonomic array of food items from domestic cats Felis silvestris catus, including prey and human-related food taxa (pet food and leftovers leaving undetectable solid remains in faeces). Scats from a captive feeding trial (n?=?41) and from free-ranging individuals (n?=?326) were collected and analysed using a cytb mini-barcode in independent PCR replicates on the Ion PGM and the MiSeq platforms. Outputs from MiSeq were more sensitive and reproducible than those from Ion PGM due to a higher sequencing depth and sequence quality on MiSeq. DNA from intact prey taxa was detected more often (82% of the expected occurrences) than DNA from pet food (54%) and raw fish and meat (31%). We assumed that this variability was linked to different degree of DNA degradation: The Ion PGM detected significantly less human-linked food, birds, field voles, murids and shrews in the field-collected samples than the MiSeq platform. Pooling the replicates from both platforms and filtering the data allowed identification of at least one food item in 87.4% of the field-collected samples. Our DNA metabarcoding approach identified 29 prey taxa, of which 25 to species level (90% of items) including 9 rodents, 3 insectivores, 12 birds and 1 reptile and 33 human-related food taxa of which 23 were identified to genus level (75% of items). Our results demonstrate that using HTS platforms such as MiSeq, which provide reads of sufficiently high quantity and quality, with sufficient numbers of technical replicates, is a robust and non-invasive approach for further dietary studies on animals foraging on a wide range of food items in anthropogenic landscapes.

SUBMITTER: Forin-Wiart MA

PROVIDER: S-EPMC6244006 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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Evaluating metabarcoding to analyse diet composition of species foraging in anthropogenic landscapes using Ion Torrent and Illumina sequencing.

Forin-Wiart Marie-Amélie MA Poulle Marie-Lazarine ML Piry Sylvain S Cosson Jean-François JF Larose Claire C Galan Maxime M

Scientific reports 20181120 1

DNA metabarcoding of faecal samples is being successfully used to study the foraging niche of species. We assessed the ability of two benchtop high-throughput sequencing (HTS) platforms, to identify a large taxonomic array of food items from domestic cats Felis silvestris catus, including prey and human-related food taxa (pet food and leftovers leaving undetectable solid remains in faeces). Scats from a captive feeding trial (n = 41) and from free-ranging individuals (n = 326) were collected and ...[more]

PMID: 30459313

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Project description:Fast and effective methods are needed for sequencing of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genome to track genetic mutations and to identify new and emerging variants during the ongoing pandemic. The objectives were to assess the performance of the SARS-CoV-2 AmpliSeq research panel and S5 plug-in analysis tools for whole-genome sequencing analysis of SARS-CoV-2 and to compare the results with those obtained with the MiSeq-based ARTIC analysis pipeline, using metrics such as depth, coverage, and concordance of single-nucleotide variant (SNV) calls. A total of 191 clinical specimens and a single cultured isolate were extracted and sequenced with AmpliSeq technology and analysis tools. Of the 191 clinical specimens, 83 (with threshold cycle [CT] values of 15.58 to 32.54) were also sequenced using an Illumina MiSeq-based method with the ARTIC analysis pipeline, for direct comparison. A total of 176 of the 191 clinical specimens sequenced on the S5XL system and prepared using the SARS-CoV-2 research panel had nearly complete coverage (>98%) of the viral genome, with an average depth of 5,031×. Similar coverage levels (>98%) were observed for 81/83 primary specimens that were sequenced with both methods tested. The sample with the lowest viral load (CT value of 32.54) achieved 89% coverage using the MiSeq method and failed to sequence with the AmpliSeq method. Consensus sequences produced by each method were identical for 81/82 samples in areas of equal coverage, with a single difference present in one sample. The AmpliSeq approach is as effective as the Illumina-based method using ARTIC v3 amplification for sequencing SARS-CoV-2 directly from patient specimens across a range of viral loads (CT values of 15.56 to 32.54 [median, 22.18]). The AmpliSeq workflow is very easily automated with the Ion Chef and S5 instruments and requires less training and experience with next-generation sequencing sample preparation than the Illumina workflow.

Dataset Information

Evaluating metabarcoding to analyse diet composition of species foraging in anthropogenic landscapes using Ion Torrent and Illumina sequencing.

Publications

Evaluating metabarcoding to analyse diet composition of species foraging in anthropogenic landscapes using Ion Torrent and Illumina sequencing.

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