Taxon‐specific PCR for DNA barcoding arthropod prey in bat faeces

Abstract

The application of DNA barcoding to dietary studies allows prey taxa to be identified in the absence of morphological evidence and permits a greater resolution of prey identity than is possible through direct examination of faecal material. For insectivorous bats, which typically eat a great diversity of prey and which chew and digest their prey thoroughly, DNA-based approaches to diet analysis may provide the only means of assessing the range and diversity of prey within faeces. Here, we investigated the effectiveness of DNA barcoding in determining the diets of bat species that specialize in eating different taxa of arthropod prey. We designed and tested a novel taxon-specific primer set and examined the performance of short barcode sequences in resolving prey species. We recovered prey DNA from all faecal samples and subsequent cloning and sequencing of PCR products, followed by a comparison of sequences to a reference database, provided species-level identifications for 149/207 (72%) clones. We detected a phylogenetically broad range of prey while completely avoiding detection of nontarget groups. In total, 37 unique prey taxa were identified from 15 faecal samples. A comparison of DNA data with parallel morphological analyses revealed a close correlation between the two methods. However, the sensitivity and taxonomic resolution of the DNA method were far superior. The methodology developed here provides new opportunities for the study of bat diets and will be of great benefit to the conservation of these ecologically important predators.

Spatiotemporal and demographic variation in the diet of New Zealand lesser short‐tailed bats (Mystacina tuberculata)

“…PCR and sequencing were performed by the Genome Centre (Queen Mary University of London). In brief: Amplification of a 157‐bp fragment of the mitochondrial cytochrome c oxidase subunit 1 was performed using primers ZBJ‐ArtF1c and ZBJ‐ArtR2c (Zeale et al., 2011) adapted to include Fluidigm tags CS1 and CS2. Each 10 μl PCR contained 5 μl of Qiagen multiplex PCR (Qiagen, CA) master mix, 3 μl of water, 0.5 μl of each 10 μM primer, and 1 μl of eluted DNA.…”

Faeces of generalist predators as ‘biodiversity capsules’: A new tool for biodiversity assessment in remote and inaccessible habitats

“…Hajibabaei et al, 2006), is therefore the ideal candidate for the biodiversity capsule approach. Other possible sources of biodiversity overestimation include secondary prey detection where DNA adheres to or is retained within the gut-contents of the predated items (Sheppard et al, 2005) and amplification of nuclear mitochondrial pseudogenes (NUMTs) (Dunshea et al, 2008;Moulton et al, 2010;Zeale et al, 2011). However, NUMTs and secondary prey DNA are probably less likely to be amplified from faecal samples due to lower copy number and higher degradation.…”

“…Although this is not an issue for species with highly specialised diets (Rougerie et al, 2011), this is particularly limiting for generalist predators that feed on a variety of prey species. Until recently, such mixed DNA samples could be analysed only after the development of large panels of species-specific primers (Jarman et al, 2004) used in complex multiplex PCRs (Harper et al, 2005;King et al, 2011) or cloning analyses (Zeale et al, 2011).…”

Female dietary bias towards large migratory moths in the European free-tailed bat ( Tadarida teniotis )

“…As a result, it is possible that segregation may also occur in the use of different prey types [9]. Testing this hypothesis, however, has been hindered by poor taxonomic resolution of most bat dietary studies, though the recent development of molecular tools for dietary analysis provides the opportunity to examine this issue in great detail [10].…”