New tools for diet analysis: nanopore sequencing of metagenomic DNA from rat stomach contents to quantify diet

Abstract

AbstractBackgroundUsing metagenomics to determine animal diet offers a new and promising alternative to current methods. Here we show that rapid and inexpensive diet quantification is possible through metagenomic sequencing with the portable Oxford Nanopore Technologies (ONT) MinION. Using an amplification-free approach, we profiled the stomach contents from wild-caught rats.ResultsWe conservatively identified diet items from over 50 taxonomic orders, ranging across nine phyla that include plants, vertebrates, invertebrates, and fungi. This highlights the wide range of taxa that can be identified using this simple approach. We calibrate the accuracy of this method by comparing the characteristics of reads matching the ground-truth host genome (rat) to those matching diet items, and show that at the family-level, false positive taxon assignments are approximately 97.5% accurate. We also suggest a way to mitigate for database biases in metagenomic approaches. Finally, we implement a constrained ordination analysis and show that we can identify the sampling location of an individual rat within tens of kilometres based on diet content alone.ConclusionsThis work establishes proof-of-principle for long-read metagenomic methods in quantitative diet analysis. We show that diet content can be quantified even with limited expertise, using a simple, amplification free workflow and a relatively inexpensive and accessible next generation sequencing method. Continued increases in the accuracy and throughput of ONT sequencing, along with improved genomic databases, suggests that a metagenomic approach to quantification of animal diets will become an important method in the future.