Long‐term persistence of horse fecalDNAin the environment makes equids particularly good candidates for noninvasive sampling

Abstract

Fecal DNA collected noninvasively can provide valuable information about genetic and ecological characteristics. This approach has rarely been used for equids, despite the need for conservation of endangered species and management of abundant feral populations. We examined factors affecting the efficacy of using equid fecal samples for conservation genetics. First, we evaluated two fecal collection methods (paper bag vs. ethanol). Then, we investigated how time since deposition and month of collection impacted microsatellite amplification success and genotyping errors. Between May and November 2014, we collected feral horse fecal samples of known age each month in a feral horse Herd Management Area in western Colorado and documented deterioration in the field with photographs. Samples collected and dried in paper bags had significantly higher amplification rates than those collected and stored in ethanol. There was little difference in the number of loci that amplified per sample between fresh fecal piles and those that had been exposed to the environment for up to 2 months (in samples collected in paper bags). After 2 months of exposure, amplification success declined. When comparing fresh (0–2 months) and old (3–6 months) fecal piles, samples from fresh piles had more matching genotypes across samples, better amplification success and less allelic dropout. Samples defecated during the summer and collected within 2 months of deposition had highest number of genotypes matching among samples, and lowest rates of amplification failure and allelic dropout. Due to the digestive system and amount of fecal material produced by equids, as well as their occurrence in arid ecosystems, we suggest that they are particularly good candidates for noninvasive sampling using fecal DNA.

Maximizing the acquisition of unique reads in noninvasive capture sequencing experiments

“…NI samples have a complex nature: they are typically composed of low proportions of host or endogenous DNA (eDNA), are highly degraded (Perry et al, 2010; Taberlet et al, 1999), and contain genetic material from the microbiota of the host and from species living in the environment where the sample was collected (i.e., exogenous DNA) (Hicks et al, 2018). The proportion of endogenous vs. exogenous DNA can be highly variable (Hernandez‐Rodriguez et al, 2018) and as previous literature has proposed, may depend on the environmental conditions, with humidity and ambient temperature having the highest influence (Goossens et al, 2000; Harestad & Bunnell, 1987; King et al, 2018; Nsubuga et al, 2004). Because of this, the employment of techniques that generate sequences of the whole genomic content of the samples, such as NGS, has not been economically feasible until recently.…”

A fecal sequel: Testing the limits of a genetic assay for bat species identification

“…Successful sequencing has been found to be higher for fresher feces [9, 10], and for samples collected in conditions such as cold temperatures [11, 12], dry climates with protection from precipitation [13], and low UV exposure [14]. Further, DNA response to these variables has been found to be taxon-specific (e.g., accurate genotyping after 2 months in feral horses [15], errors after 7 days in coyotes [16]) and target-specific (from bat guano: change in bacterial community composition after 1 hour [17], identification of bat species at 3 months [3]). DNA concentration can also vary by collection/preservation method [15, 18] and by the order the feces traveled through the intestines [19].…”

“…Further, DNA response to these variables has been found to be taxon-specific (e.g., accurate genotyping after 2 months in feral horses [15], errors after 7 days in coyotes [16]) and target-specific (from bat guano: change in bacterial community composition after 1 hour [17], identification of bat species at 3 months [3]). DNA concentration can also vary by collection/preservation method [15, 18] and by the order the feces traveled through the intestines [19]. Hence, the pairing of rapidly evolving genetic sequencing methods with fecal-derived DNA means it is important that the constraints inherent to particular fecal types and their associated assays are explored and delineated to allow for predictability and reproducibility [20, 21].…”

Fecal DNA Isolation and Degradation in Clam Cyclina sinensis: Noninvasive DNA Isolation for Conservation and Genetic Assessment

“…Therefore, mitochondrial and nuclear genomic DNA can be isolated from the remaining epithelial cells in the feces [10]. Fecal molecular biotechnology provides a rapid and dependable way of sampling endangered animals [11][12][13][14]. In addition, with the development of molecular biology technology, fecal DNA is extensively used in genetic biology studies for species identification [15][16][17], individual identification [18][19][20], sex identification [21][22][23][24][25], population genetic structure [26][27][28], and genetic diversity evaluation [29].…”