1. Metabarcoding for the parallel identification of several hundred predators and their prey: Application to bat species diet analysis
- Author
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Orianne Tournayre, Dominique Pontier, Eric Pierre, Nathalie Charbonnel, Jean-Baptiste Pons, Maxime Galan, Maxime Leuchtmann, Centre de Biologie pour la Gestion des Populations (UMR CBGP), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut National de la Recherche Agronomique (INRA)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Université de Montpellier (UM)-Institut de Recherche pour le Développement (IRD [France-Sud])-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Laboratoire de Biométrie et Biologie Evolutive - UMR 5558 (LBBE), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de Recherche en Informatique et en Automatique (Inria)-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS), LabEx Ecofect, Eco-Evolutionary Dynamics of Infectious Diseases, Nature environment, Partenaires INRAE, This work was funded by the LabEx ECOFECT and the internal funding from the CBGP laboratory. Orianne Tournayre PhD is funded by the LabEx CeMEB. Part of this work (JBP, DP) was performed within the framework of the LabEx ECOFECT of Université de Lyon and the Poitou-Charentes Nature Feder project 'Grand Rhinolophe et trame verte bocagère : étude des facteurs environnementaux influant sur la dynamique des populations ' (ML, JBP, DP)., ANR-11-LABX-0048,ECOFECT,Dynamiques eco-évolutives des maladies infectieuses(2011), and ANR-10-LABX-0004,CeMEB,Mediterranean Center for Environment and Biodiversity(2010)
- Subjects
0106 biological sciences ,0301 basic medicine ,Conservation of Natural Resources ,Arthropoda ,predator-prey interactions ,[SDV]Life Sciences [q-bio] ,Zoology ,environmental DNA (eDNA) ,Biology ,010603 evolutionary biology ,01 natural sciences ,Predation ,Feces ,03 medical and health sciences ,Chiroptera ,Diet analysis ,false positives ,Genetics ,False positive paradox ,Animals ,DNA Barcoding, Taxonomic ,Species identification ,14. Life underwater ,Arthropods ,Ecology, Evolution, Behavior and Systematics ,030304 developmental biology ,0303 health sciences ,high-throughput sequencing ,Feeding Behavior ,Sequence Analysis, DNA ,DNA extraction ,Diet ,030104 developmental biology ,Evolutionary biology ,[SDE]Environmental Sciences ,Taxonomic resolution ,Identification (biology) ,Species richness ,Biotechnology ,Faecal pellet - Abstract
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.
- Published
- 2018