DNA barcoding identifies cryptic animal tool materials

Funding: Biotechnology and Biological Sciences Research Council (BBSRC) (Grants BB/G023913/1 and BB/G023913/2 to C.R., and studentship to B.C.K.), the School of Biology at the University of St Andrews (studentships to M.P.S. and B.C.K.), and the Leverhulme Trust (Grant RPG-2015-273 to P.M.H.).Some animals fashion tools or constructions out of plant materials to aid foraging, reproduction, self-maintenance, or protection. Their choice of raw materials can affect the structure and properties of the resulting artifacts, with considerable fitness consequences. Documenting animals’ material preferences is challenging, however, as manufacture behavior is often difficult to observe directly, and materials may be processed so heavily that they lack identifying features. Here, we use DNA barcoding to identify, from just a few recovered tool specimens, the plant species New Caledonian crows (Corvus moneduloides) use for crafting elaborate hooked stick tools in one of our long-term study populations. The method succeeded where extensive fieldwork using an array of conventional approaches—including targeted observations, camera traps, radio-tracking, bird-mounted video cameras, and behavioral experiments with wild and temporarily captive subjects—had failed. We believe that DNA barcoding will prove useful for investigating many other tool and construction behaviors, helping to unlock significant research potential across a wide range of study systems.Publisher PDFPeer reviewe

A pragmatic approach for integrating molecular tools into biodiversity conservation

Molecular tools are increasingly applied for assessing and monitoring biodiversity and informing conservation action. While recent developments in genetic and genomic methods provide greater sensitivity in analysis and the capacity to address new questions, they are not equally available to all practitioners: There is considerable bias across institutions and countries in access to technologies, funding, and training. Consequently, in many cases, more accessible traditional genetic data (e.g., microsatellites) are still utilized for making conservation decisions. Conservation approaches need to be pragmatic by tackling clearly defined management questions and using the most appropriate methods available, while maximizing the use of limited resources. Here we present some key questions to consider when applying the molecular toolbox for accessible and actionable conservation management. Finally, we highlight a number of important steps to be addressed in a collaborative way, which can facilitate the broad integration of molecular data into conservation

Genetic and ecological outcomes of Inga vera subsp. affinis (leguminosae) tree plantations in a fragmented tropical landscape

Planting of native trees for habitat restoration is a widespread practice, but the consequences for the retention and transmission of genetic diversity in planted and natural populations are unclear. Using Inga vera subsp. affinis as a model species, we genotyped five natural and five planted populations in the Atlantic forest of northeastern Brazil at polymorphic microsatellite loci. We studied the breeding system and population structure to test how much genetic diversity is retained in planted relative to natural populations. We then genotyped seedlings from these populations to test whether genetic diversity in planted populations is restored by outcrossing to natural populations of I. vera. The breeding system of natural I. vera populations was confirmed to be highly outcrossing (t = 0.92; FIS = -0.061, P = 0.04), with populations showing weak population substructure (FST = 0.028). Genetic diversity in planted populations was 50% less than that of natural populations (planted: AR = 14.9, HO = 0.865 and natural: AR = 30.8, HO = 0.655). However, seedlings from planted populations showed a 30% higher allelic richness relative to their parents (seedlings AR = 10.5, parents AR = 7.6). Understanding the processes and interactions that shape this system are necessary to provide ecologically sensible goals and successfully restore hyper-fragmented habitats. Future restoration plans for I. vera must consider the genetic diversity of planted populations and the potential for gene flow between natural populations in the landscape, in order to preserve ecological interactions (i.e. pollination), and promote opportunities for outcrossing

Allocating sex in road-killed possums using PCR

Road-killed animals are a readily available but often overlooked source of specimens and data. Determining the sex of road-killed possums is often difficult due to the severe damage carcasses sustain on impact with vehicles. Consequently, valuable information is lost. To assess the potential application of genetic techniques to address this problem, tissue biopsies (n=47) taken from road-killed possums were sexed via PCR, through the simultaneous use of W- (G6PD) and Y-linked (SRY) primers. Comparisons with animals of known sex were used to validate the accuracy of this method, with 100% (n=19) of samples correctly identified in a blind test. This methodology can provide researchers with valuable, additional data for road-kill studies where assigning sex in the field is not possible.4 page(s

Understanding Historical Demographic Processes to Inform Contemporary Conservation of an Arid zone Specialist: The Yellow-Footed Rock-Wallaby

Little genetic research has been undertaken on mammals across the vast expanse of the arid biome in Australia, despite continuing species decline and need for conservation management. Here, we evaluate the contemporary and historical genetic connectivity of the yellow-footed rock-wallaby, Petrogale xanthopus xanthopus, a threatened macropodid which inhabits rocky outcrops across the disconnected mountain range systems of the southern arid biome. We use 17 microsatellite loci together with mitochondrial control region data to determine the genetic diversity of populations and the evolutionary processes shaping contemporary population dynamics on which to base conservation recommendations. Our results indicate the highly fragmented populations have reduced diversity and limited contemporary gene flow, with most populations having been through population bottlenecks. Despite limited contemporary gene flow, the phylogeographic relationships of the mitochondrial control region indicate a lack of structure and suggests greater historical connectivity. This is an emerging outcome for mammals across this arid region. On the basis of our results, we recommend augmentation of populations of P. x. xanthopus, mixing populations from disjunct mountain range systems to reduce the chance of continued diversity loss and inbreeding depression, and therefore maximize the potential for populations to adapt and survive into the future.This research was funded by the South Australian Department of Environment and Natural Resources

Paternally inherited genetic markers reveal new insights into genetic structuring within Macropus fuliginosus and hybridisation with sympatric Macropus giganteus

There are several aspects of biology in which the contribution of males and females is unequal. In these instances the examination of Y chromosome markers may be used to elucidate male-specific attributes. Here, male dispersal patterns and genetic structuring were examined using four Y-microsatellite loci in 186 male western grey kangaroos, Macropus fuliginosus, from throughout the species' trans-continental distribution. In addition, 52 male grey kangaroos were examined to investigate hybridisation between M. fuliginosus and the eastern grey kangaroo, Macropus giganteus, in their region of sympatry in eastern Australia. Detected Y chromosome diversity was low, resulting from low effective male population size due to skewed sex ratios and a polygynous mating system. As expected, male dispersal was high across the range. However, the Lake Torrens–Flinders Ranges region appears to have significantly restricted male movement between eastern and central/western Australia. There was little evidence to suggest that other barriers (Nullarbor Plain and Swan River Valley) previously identified by nuclear and mitochondrial DNA marker studies restrict male movement. Hence, the admixture events previously identified may be associated with high male dispersal. Within the region of sympatry between M. fuliginosus and M. giganteus in eastern Australia, four M. giganteus individuals were found to possess M. fuliginosus Y-haplotypes. These results confirm the occurrence of hybridisation between male M. fuliginosus and female M. giganteus. Additionally, the introgression of M. fuliginosus Y-haplotypes into M. giganteus populations indicates that at least some male hybrids are fertile, despite evidence to the contrary from captive studies. This study has provided insights into the male contribution to population history, structure and hybridisation in M. fuliginosus, which were not predicted by comparisons between biparentally and maternally inherited markers. This highlights the importance of direct examination of the Y chromosome to provide novel insights into male-mediated processes, especially where the contribution of the sexes may differ

Extensive genetic differentiation detected within a model marsupial, the tammar wallaby (Notamacropus eugenii)

The tammar wallaby (Notamacropus eugenii) is one of the most intensively studied of all macropodids and was the first Australasian marsupial to have its genome sequenced. However, comparatively little is known about genetic diversity and differentiation amongst the morphologically distinct allopatric populations of tammar wallabies found in Western (WA) and South Australia (SA). Here we compare autosomal and Y-linked microsatellite genotypes, as well as sequence data (~600 bp) from the mitochondrial DNA (mtDNA) control region (CR) in tammar wallabies from across its distribution. Levels of diversity at autosomal microsatellite loci were typically high in the WA mainland and Kangaroo Island (SA) populations (A = 8.9-10.6; He = 0.77-0.78) but significantly reduced in other endemic island populations (A = 3.8-4.1; He = 0.41-0.48). Autosomal and Y-linked microsatellite loci revealed a pattern of significant differentiation amongst populations, especially between SA and WA. The Kangaroo Island and introduced New Zealand population showed limited differentiation. Multiple divergent mtDNA CR haplotypes were identified within both SA and WA populations. The CR haplotypes of tammar wallabies from SA and WA show reciprocal monophyly and are highly divergent (14.5%), with levels of sequence divergence more typical of different species. Within WA tammar wallabies, island populations each have unique clusters of highly related CR haplotypes and each is most closely related to different WA mainland haplotypes. Y-linked microsatellite haplotypes show a similar pattern of divergence although levels of diversity are lower. In light of these differences, we suggest that two subspecies of tammar wallaby be recognized; Notamacropus eugenii eugenii in SA and N. eugenii derbianus in WA. The extensive neutral genetic diversity and inter-population differentiation identified within tammar wallabies should further increase the species value and usefulness as a model organism.23 page(s