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.
Highlights
With increasing urbanisation and loss of natural habitat, many species have adapted to human-dominated landscapes by shifting their dietary niche to include human-processed food items[1]
688,530 and 2,509,592 sequences of cytb were assigned to the faecal samples and were identified to the closest taxonomic rank using two out of three sliding similarity thresholds (i.e. 100% or 98% for prey taxa and 98% or 95% for food processed taxa)
In the calibration study (n = 41 faecal samples), we obtained a high proportion of reads from the host: 41,129 reads (87.5%) for Ion Personal Genome Machine (Ion PGM) and 235,673 reads (98.5%) for MiSeq at the 95% similarity threshold
Summary
With increasing urbanisation and loss of natural habitat, many species have adapted to human-dominated landscapes by shifting their dietary niche to include human-processed food items[1] Scavengers such as gulls Larus spp., ravens Corvus spp., Eurasian badgers Meles meles, opossums Didelphis virginiana, racoons Procyon lotor, or wild boar Sus scrofa, as well as opportunistic generalist carnivores such as coyotes Canis latrans[2], or foxes Vulpes spp.[3,4], presently have dietary compositions that differ substantially from their natural diet. By comparing obtained sequences to a standard reference library of taxonomically identified organisms (e.g. Bold or GenBank), the taxa or species present in a sample can be identified with high confidence In this way, metabarcoding of faecal samples allows ingested food items to be identified by characterizing the barcode sequences in the samples[12]. This technique has been successfully used to analyse the diet of species of birds[13], mammals[11,14,15], reptiles[16,17] and fish[18]
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