Blood, sweat and tears: a review of non-invasive DNA sampling

Abstract

The use of DNA data is ubiquitous across animal sciences. DNA may be obtained from an organism for a myriad of reasons including identification and distinction between cryptic species, sex identification, comparisons of different morphocryptic genotypes or assessments of relatedness between organisms prior to a behavioural study. DNA should be obtained while minimizing the impact on the fitness, behaviour or welfare of the subject being tested, as this can bias experimental results and cause long-lasting effects on wild animals. Furthermore, minimizing impact on experimental animals is a key Refinement principle within the '3Rs' framework which aims to ensure that animal welfare during experimentation is optimised. The term 'non-invasive DNA sampling' has been defined to indicate collection methods that do not require capture or cause disturbance to the animal, including any effects on behaviour or fitness. In practice this is not always the case, as the term 'non-invasive' is commonly used in the literature to describe studies where animals are restrained or subjected to aversive procedures. We reviewed the non-invasive DNA sampling literature for the past six years (380 papers published in 2013-2018) and uncovered the existence of a significant gap between the current use of this terminology (i.e. 'non-invasive DNA sampling') and its original definition. We show that 58% of the reviewed papers did not comply with the original definition. We discuss the main experimental and ethical issues surrounding the potential confusion or misuse of the phrase 'non-invasive DNA sampling' in the current literature and provide potential solutions. In addition, we introduce the terms 'non-disruptive' and 'minimally disruptive' DNA sampling, to indicate methods that eliminate or minimise impacts not on the physical integrity/structure of the animal, but on its behaviour, fitness and welfare, which in the literature reviewed corresponds to the situation for which an accurate term is clearly missing. Furthermore, we outline when these methods are appropriate to use.

Mitochondrial genomes assembled from non-invasive eDNA metagenomic scat samples in the endangered Amur tiger Panthera tigris altaica

“…Such efforts are currently being tested in other large vertebrates with major conservation problems ( e.g ., in moose– Lyet et al, 2021 ) and this study is a step forward towards to implementation of indirect field surveillance in P. tigris altaica . Considering that the complete removal of fecal samples from the field might increase agonistic interactions among individuals in territorial species such as tigers, the author suggest the partial removal and sampling of scat from the wild (see Lefort et al, 2022 ).…”

How to quantify factors degrading DNA in the environment and predict degradation for effective sampling design

“…Genetic material left behind in the environment with direct information about its donor (e.g., feces, hair, feather, and scales), defined as noninvasive DNA (Taberlet et al, 1999), or contained in environmental samples (e.g., water, soil, and sediment), defined as environmental DNA (eDNA) (Thomsen & Willerslev, 2015), is increasingly used in ecological studies and biomonitoring (Lefort et al, 2022; Thomsen & Willerslev, 2015). Despite differences in concentration and sampling approach, DNA from both noninvasive DNA and eDNA starts degrading upon detachment from the organism.…”

“…Despite differences in concentration and sampling approach, DNA from both noninvasive DNA and eDNA starts degrading upon detachment from the organism. Thus, environmental factors influencing DNA degradation are critical for all kinds of extra-organismal DNA (eoDNA; sensu Rodriguez-Ezpeleta et al, 2021), regardless of the invasiveness (Lefort et al, 2022) of the sampling. This is particularly problematic because eoDNA samples often contain low initial quantities of DNA (Taberlet et al, 1999).…”

Buccal swabs for amphibian genomics

“…As laboratory techniques improved, conservation biologists have thus turned towards alternative DNA sources that reduce the disturbance on animals, such as buccal, cloacal and skin swabs, along with hairs, feathers, exuviae and environmental DNA (reviewed by Carroll et al, 2018). Despite these options, harmful practices like full specimen collection, biopsy, mutilation and blood sampling, are still preferentially chosen (Zemanova, 2019;Lefort et al, 2022). This is partly due to the cultural difficulty to change "old habits" that have proven efficient, even more so now that genomic analyses are at stake.…”

“…Herpetologists now face a paradox. Implementing genomic approaches, such as RADsequencing (RAD-seq) and sequence-capture, is more DNA-demanding, but at the same time, ethical committees are enforcing increasingly strict policies when issuing sampling permits, which can be frustrating for researchers (Mulcahy, 2017;Lefort et al, 2022). Favoring buccal swabs would appear timely, hence whether they are also suitable for genomic analyses becomes a burning question.…”