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 duplicates 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.
Ion PGM raw sequences of the Cytb minibarcode from 400 faecal samples of cats (Felis silvestris catus) from Eastern France
This ZIP file contains the FASTQ file of 4,042,017 reads produced using the Ion PGM platform. Each faecal sample was amplified and sequenced in replicates. The 840 multiplexed PCR products were tagged using both forward and reverse 7-bp tags. The list of the 415 multiplexed samples and the 10 negative controls are provided in the following CSV file titled: Information concerning the samples multiplexed in the Ion PGM run.
IonPGM_Reads_CytB_Cats.fastqsanger.zip
Information concerning the samples multiplexed in the Ion PGM run
This CSV file contains the list of the sample names, PCR names, replicate numbers, the forward and reverse tag sequences, the forward and reverse primer sequences, the gene name and the species name for each of the 415 samples and 5 controls amplified in replicate and multiplexed in the Ion PGM run. Note that 15 samples were also performed twice as replicates for the DNA extraction step.
MiSeq raw sequences of the Cytb minibarcode from 400 faecal samples of cats (Felis silvestris catus) from Eastern France
This ZIP file contains 1694 FASTQ files of the 4.042.606 paired-end reads (R1: reads 1; R2: reads 2) produced for each faecal samples in replicates using the MiSeq platform. The 847 multiplexed PCR products were indexed using both forward and reverse indices. The list of the 415 multiplexed samples and the 13 negative controls are provided in the following CSV file titled: Information concerning the samples multiplexed in the MiSeq run.
MiSeq_Reads_CytB_Cats.fastq.gz.zip
Information concerning the samples multiplexed in the MiSeq run
This CSV file contains the list of the sample names, PCR names, replicate numbers, dual-index ID, species name and the read 1 and read 2 fatsq file names for each of the 415 samples and 13 controls amplified in replicate and multiplexed in the Ion PGM run. Note that 15 samples were also performed twice as replicates for the DNA extraction step. Note that 15 samples were also performed twice as replicates for the DNA extraction step.
Abundance table before data filtering for Ion PGM and MiSeq sequencing
This TXT file contains the number of reads for each distinct taxonomic affiliation obtains with the SESAME barcode software using the variant sequences of the Ion PGM and MiSeq runs, and for each sample replicate before data filtering. The thresholds TFA described in the article are provided in the columns C to H. Each sample is described by 12 columns: 6 concern the Ion PGM results (suffix “_I”) and 6 concern the MiSeq results (suffix “_M”). For each sequencing platform, the taxonomic affiliation results of the two PCR replicates are provided for three similarity thresholds: 95%, 98% and 100%. The column “type” indicates the putative origin of the food using the taxonomic identification: prey (Pr) or human-related food (Hf).
Occurrence table after data filtering for Ion PGM and MiSeq sequencing
This TXT file contains the number of occurrences for each distinct taxonomic affiliation obtains after data filtering using the thresholds TFA describe in the manuscript. The results are provided for both sequencers, Ion PGM (columns with the suffix “Ion”) and MiSeq (columns with the suffix “Mi”), as well as the intra-platform repeatability between PCR replicates (columns with suffix “SumIon” and “SumMi”, “simp” for one positive PCR and “dup” for two positive PCR) and the final occurrences based of the clustering of the four PCR replicates (IonPGM + MiSeq; columns with the suffix “OccDup” = 1 if at least two PCR replicates on four are positives, “OccSimp” = 1 if at least one PCR replicate on four is positive).
