Metabarcoding of Fecal Samples to Determine Herbivore Diets: A Case Study of the Endangered Pacific Pocket Mouse

Iwanowicz, Deborah D.; Vandergast, Amy G.; Cornman, Robert S.; Adams, Cynthia R.; Kohn, Joshua R.; Fisher, Robert N.; Brehme, Cheryl S.

doi:10.1371/journal.pone.0165366

Cited by 44 publications

(55 citation statements)

References 54 publications

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“…Faeces are the most popular sample type for dietary analyses, as they can be collected noninvasively (e.g., Bohmann et al, 2011;Kaunisto, Roslin, Sääksjärvi, & Vesterinen, 2017). Faeces can also be collected from captured animals (e.g., Iwanowicz et al, 2016;Kaunisto et al, 2017), which enables complementing the trophic information derived from the samples with predator's metadata, and thus addressing many biological questions that without predator information would be difficult to tackle, for example, differences between sexes (Mata et al, 2016) or developmental stages (Jedlicka, Vo, & Almeida, 2016). In addition, it renders possible to control the confounding factors introduced by the biological features of consumer individuals in the data analysis step ( Figure 2).…”

Section: Faecesmentioning

confidence: 99%

Promises and pitfalls of using high‐throughput sequencing for diet analysis

Alberdi

Aizpurua

Bohmann

et al. 2018

Molecular Ecology Resources

160

183

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The application of high‐throughput sequencing‐based approaches to DNA extracted from environmental samples such as gut contents and faeces has become a popular tool for studying dietary habits of animals. Due to the high resolution and prey detection capacity they provide, both metabarcoding and shotgun sequencing are increasingly used to address ecological questions grounded in dietary relationships. Despite their great promise in this context, recent research has unveiled how a wealth of biological (related to the study system) and technical (related to the methodology) factors can distort the signal of taxonomic composition and diversity. Here, we review these studies in the light of high‐throughput sequencing‐based assessment of trophic interactions. We address how the study design can account for distortion factors, and how acknowledging limitations and biases inherent to sequencing‐based diet analyses are essential for obtaining reliable results, thus drawing appropriate conclusions. Furthermore, we suggest strategies to minimize the effect of distortion factors, measures to increase reproducibility, replicability and comparability of studies, and options to scale up DNA sequencing‐based diet analyses. In doing so, we aim to aid end‐users in designing reliable diet studies by informing them about the complexity and limitations of DNA sequencing‐based diet analyses, and encourage researchers to create and improve tools that will eventually drive this field to its maturity.

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Section: Faecesmentioning

confidence: 99%

Promises and pitfalls of using high‐throughput sequencing for diet analysis

Alberdi

Aizpurua

Bohmann

et al. 2018

Molecular Ecology Resources

160

183

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“…Morphologic examinations of faeces and guano have therefore initially provided important knowledge on bat diets (Hope et al, 2014;Lam et al, 2013 ). These methods have major limits (time consuming, taxonomic expertise required, low resolution and ascertainment biases due to the reject of insect hard parts, see refs in Iwanowicz et al, 2016). In particular, identifying preys at the species level is not possible based on morphological analyses of faecal samples.…”

Section: Introductionmentioning

confidence: 99%

Metabarcoding for the parallel identification of several hundred predators and their preys: application to bat species diet analysis

Galan

Pons

Tournayre

et al. 2017

Preprint

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Assessing diet variability is of main importance to better understand the biology of bats and design conservation strategies. Although the advent of metabarcoding has facilitated such analyses, this approach does not come without challenges. Biases may occur throughout the whole experiment, from fieldwork to biostatistics, resulting in the detection of false negatives, false positives or low taxonomic resolution. We detail a rigorous metabarcoding approach based on a short COI minibarcode and two-step PCR protocol enabling the ‘all at once’ taxonomic identification of bats and their arthropod preys for several hundreds of samples. Our study includes faecal pellets collected in France from 357 bats representing 16 species, as well as insect mock communities that mimic bat meals of known composition, negative and positive controls. All samples were analysed using three replicates. We compare the efficiency of DNA extraction methods and we evaluate the effectiveness of our protocol using identification success, taxonomic resolution, sensitivity, and amplification biases. Our parallel identification strategy of predators and preys reduces the risk of mis-assigning preys to wrong predators and decreases the number of molecular steps. Controls and replicates enable to filter the data and limit the risk of false positives, hence guaranteeing high confidence results for both prey occurrence and bat species identification. We validate 551 COI variants from arthropod including 18 orders, 117 family, 282 genus and 290 species. Our method therefore provides a rapid, resolutive and cost-effective screening tool for addressing evolutionary ecological issues or developing ‘chirosurveillance’ and conservation strategies.

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“…This highlights the value in taking multiple samples from a single individual, as diets are likely to be highly variable. This has been observed previously in another herbivore, the Pacific pocket mouse (Iwanowicz et al 2016). Only a single sample was taken from each faecal pellet-it might also be useful to compare subsamples from a single pellet to determine its degree of consistency.…”

Section: Discussionmentioning

confidence: 73%

Dropping Hints: Estimating the diets of livestock in rangelands using DNA metabarcoding of faeces

Lee¹,

Alemseged²,

Mitchell³

2018

MBMG

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The introduction of domesticated animals into new environments can lead to considerable ecological disruption, and it can be difficult to predict their impact on the new ecosystem. In this study, we use faecal metabarcoding to characterize the diets of three ruminant taxa in the rangelands of south-western New South Wales, Australia. Our study organisms included goats (Capra aegagrus hircus) and two breeds of sheep (Ovis aries): Merinos, which have been present in Australia for over two hundred years, and Dorpers, which were introduced in the 1990s. We used High-Throughput Sequencing methods to sequence the rbcL and ITS2 genes of plants in the faecal samples, and identified the samples using the GenBank and BOLD online databases, as well as a reference collection of sequences from plants collected in the study area. We found that the diets of all three taxa were dominated by the family Malvaceae, and that the Dorper diet was more diverse than the Merino diet at both the family and the species level. We conclude that Dorpers, like Merinos, are potentially a threat to some vulnerable species in the rangelands of New South Wales.

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Metabarcoding of Fecal Samples to Determine Herbivore Diets: A Case Study of the Endangered Pacific Pocket Mouse

Cited by 44 publications

References 54 publications

Promises and pitfalls of using high‐throughput sequencing for diet analysis

Promises and pitfalls of using high‐throughput sequencing for diet analysis

Metabarcoding for the parallel identification of several hundred predators and their preys: application to bat species diet analysis

Dropping Hints: Estimating the diets of livestock in rangelands using DNA metabarcoding of faeces

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