Peter B. S. Spencer

Identification of microsatellites from an extinct moa species using high-throughput (454) sequence data.

Genetic variation in microsatellites is rarely examined in the field of ancient DNA (aDNA) due to the low quantity of nuclear DNA in the fossil record together with the lack of characterized nuclear markers in extinct species. 454 sequencing platforms provide a new high-throughput technology capable of generating up to 1 gigabases per run as short (200-400-bp) read lengths. 454 data were generated from the fossil bone of an extinct New Zealand moa (Aves: Dinornithiformes). We identified numerous short tandem repeat (STR) motifs, and here present the successful isolation and characterization of one polymorphic microsatellite (Moa_MS2). Primers designed to flank this locus amplified all three moa species tested here. The presented method proved to be a fast and efficient way of identifying microsatellite markers in ancient DNA templates and, depending on biomolecule preservation, has the potential of enabling high-resolution population genetic studies of extinct taxa. As sequence read lengths of the 454 platforms and its competitors (e.g., the SOLEXA and SOLiD platforms) increase, this approach will become increasingly powerful in identifying microsatellites in extinct (and extant) organisms, and will afford new opportunities to study past biodiversity and extinction processes.

Genetic diversity in remnant mainland and "pristine"’ island populations of three endemic Australian macropodids (Marsupialia): Macropus eugenii, Lagorchestes hirsutus and Petrogale lateralis

Since European settlement, mainland Australia has experienced a wave of mammal extinctions and population declines. However, some species have persisted on off-shore islands, which are now viewed as important wildlife refuges. In this study, we assessed the level of genetic diversity, at 7–11 microsatellite loci, in island and remnant mainland populations of three endemic species of macropodid marsupial; the tammar wallaby Macropus eugenii(n = 92); rufous hare-wallaby Lagorchestes hirsutus(n = 40) and black-footed rock-wallaby Petrogale lateralis(n = 164). There was a consistent pattern of significantly higher levels of microsatellite diversity in the remnant mainland population (A= 4.9–13.9; He= 0.61–0.86) of each species compared to conspecific “pristine” island populations (A= 1.2–3.7; He= 0.05–0.44). These marked differences are even apparent where island populations currently have a substantially larger census size. In addition, island populations were substantially inbred (Fe= 0.49–0.91). Although island populations have been insulated from the relatively recent threatening processes operating on the mainland, they have nevertheless been significantly impacted by increased inbreeding and the substantial erosion of genetic diversity. Despite the difficulties of ensuring the survival of remnant mainland populations, they appear to retain substantially more genetic diversity than their island counterparts and therefore are more likely to contribute to the long-term persistence of their species. These data also demonstrate that small remnant mainland populations (n < 10–20) are often capable of rapid recovery and are not necessarily genetically depauperate.

Illegal translocation and genetic structure of feral pigs in Western Australia

Abstract Unlike many regions in the world where wild pigs (Sus scrofa) are threatened, in Australia they are a significant invasive species. As such, the molecular ecology of feral pigs was investigated to understand their social and population genetic structure. Samples from 269 adult animals were collected over their distribution in southwestern Australia. Using 14 highly polymorphic microsatellite markers, we identified 7 inferred feral pig populations that had moderate heterozygosity (mean = 0.580) and displayed a high level of differentiation (mean RST = 0.180). In revealing the genetic structure of feral pigs, we detected anomalies in the putative origin of some individuals. Samples from these animals were collected from 2 main areas: recently colonized regions that were previously uninfested, and established feral pig populations, where animals from geographically isolated areas had been introduced. In the latter, these corresponded to areas that were in close proximity to public road access and towns. Given the large distances immigrants were found from their population of origin (from 50 to > 400 km), the generally low levels of dispersal of southwest feral pigs, and the grouping and sex of these pigs, we suggest that these individuals have been deliberately and illegally translocated to supplement recreational hunting stocks. Additionally, we could not detect any genetic contribution in these feral pigs from domestic pig herds, suggesting that the deliberate release of domestic pigs to restock feral populations is relatively uncommon. Our molecular data allowed some inferences regarding the success or lack thereof of current management practices, and offered considerable insights into the dynamics of the feral pig populations and identification of “new” approaches that may allow for better control of this highly destructive species.

Enhancement of reproductive success through mate choice in a social rock-wallaby, Petrogale assimilis (Macropodidae) as revealed by microsatellite markers

Abstract The reason why a female who is socially paired to one particular male seeks extra-pair copulations (EPCs) with others has important implications in life history models and to the study of behaviour. The Allied rock-wallaby, Petrogale assimilis, lives in spatially isolated colonies in tropical north Queensland, Australia. Extensive observations of a colony at Black Rock showed that intense behavioural bonding occurs between pairs of adult males and females; about two-thirds of males paired with one female, the remainder paired with two females simultaneously. Single-locus microsatellite profiling determined the paternity of 63 offspring from 21 females for which long-term behavioural data were available. One-third of the young were fathered by males which were not paired socially with the mother. The mating system was heterogeneous: (1) all offspring of 11 females were fathered by the mothers partner, (2) all young of 5 females were fathered by extra-pair males, and (3) only some of the young of 5 females were fathered by their regular consort. Analysis of individual longitudinal demographic records showed that females whose young were always fathered by their consort had higher reproductive success than those whose young were always fathered as a result of (EPCs). However, females with some offspring fathered by their regular consort and others via EPCs had the highest probability of raising young to independence. These females were significantly more likely to have an offspring fathered as a result of an EPC if their previous young had failed to survive to pouch emergence. These results are consistent with the hypothesis that females choose mates for their genetic quality. Comparison of the males with which these females sought EPCs and the regular consorts suggested that arm length rather than body weight or testes size was used as the index of genetic quality. Results from a second colony of rock-wallabies in which the reproductive rate was accelerated were also consistent with the genetic-quality hypothesis. These results imply that by choosing better-quality fathers irrespective of social pairing, females are able to maximise their overall lifetime reproductive success, and presumably, those of their offspring.

Contribution of genetics to ecological restoration

Ecological restoration of degraded ecosystems has emerged as a critical tool in the fight to reverse and ameliorate the current loss of biodiversity and ecosystem services. Approaches derived from different genetic disciplines are extending the theoretical and applied frameworks on which ecological restoration is based. We performed a search of scientific articles and identified 160 articles that employed a genetic approach within a restoration context to shed light on the links between genetics and restoration. These articles were then classified on whether they examined association between genetics and fitness or the application of genetics in demographic studies, and on the way the studies informed restoration practice. Although genetic research in restoration is rapidly growing, we found that studies could make better use of the extensive toolbox developed by applied fields in genetics. Overall, 41% of reviewed studies used genetic information to evaluate or monitor restoration, and 59% provided genetic information to guide prerestoration decision‐making processes. Reviewed studies suggest that restoration practitioners often overlook the importance of including genetic aspects within their restoration goals. Even though there is a genetic basis influencing the provision of ecosystem services, few studies explored this relationship. We provide a view of research gaps, future directions and challenges in the genetics of restoration.

Highly variable microsatellites in isolated colonies of the rock‐wallaby (Petrogale assimilis)

Habitat fragmentation is a major concern of conservation biologists, since reduced gene flow between isolated subpopulations may further decrease the effective population size of a species. Rock wallaby (genus Petrogale) colonies provide a naturally occurring system to study the genetic consequences of habitat fragmentation. Colonies of less than 10 to more than 50 adult rock wallabies are restricted to isolated rock outcrops, and are thus expected to exhibit the genetic and demographic consequences of small population size. In this paper, we describe the characterization of a series of microsatellite loci from the allied rock-wallaby, Petrogale assirnilis, and their use to estimate genetic variation. Despite the small population sizes, a high degree of heterozygosity was observed at all the loci investiga ted. Single-locus microsatellite typing exploits the natural variability of simple repetitive sequences, and has been extensively employed in the medical sciences and in animal husbandry. Despite the potential applications of this technology in conservation genetics, few marsupial microsatellite loci have been characterized to date. In a wide range of eutherian mammals, microsatellites with the general form (TG), are by far the most common class in the genome (see, for example, Moore et nl. 1991). Our preliminary studies (Odorico et aI. 1992) imply that this is also true for marsupials. We therefore characterized a series of (TG), microsatellite loci from I? ussimilis, with the aim of developing microsatellite PCR systems for kinship and population studies. A partial genomic library was constructed for P. ussimilis and screened by standard techniques (Weber L May 1989; Tautz 1989; reviewed in Queller et nl. 1993). In the initial round of screening, seven clones containing (TG), microsatellites with n > 20 were identified; several additional clones contained imperfect or lower numbers of (TG), repeats. A further single clone contained ( n X o and another a complex locus containing two tetranucleotide

Profiling the Dead: Generating Microsatellite Data from Fossil Bones of Extinct Megafauna—Protocols, Problems, and Prospects

We present the first set of microsatellite markers developed exclusively for an extinct taxon. Microsatellite data have been analysed in thousands of genetic studies on extant species but the technology can be problematic when applied to low copy number (LCN) DNA. It is therefore rarely used on substrates more than a few decades old. Now, with the primers and protocols presented here, microsatellite markers are available to study the extinct New Zealand moa (Aves: Dinornithiformes) and, as with single nucleotide polymorphism (SNP) technology, the markers represent a means by which the field of ancient DNA can (preservation allowing) move on from its reliance on mitochondrial DNA. Candidate markers were identified using high throughput sequencing technology (GS-FLX) on DNA extracted from fossil moa bone and eggshell. From the ‘shotgun’ reads, >60 primer pairs were designed and tested on DNA from bones of the South Island giant moa (Dinornis robustus). Six polymorphic loci were characterised and used to assess measures of genetic diversity. Because of low template numbers, typical of ancient DNA, allelic dropout was observed in 36–70% of the PCR reactions at each microsatellite marker. However, a comprehensive survey of allelic dropout, combined with supporting quantitative PCR data, allowed us to establish a set of criteria that maximised data fidelity. Finally, we demonstrated the viability of the primers and the protocols, by compiling a full Dinornis microsatellite dataset representing fossils of c. 600–5000 years of age. A multi-locus genotype was obtained from 74 individuals (84% success rate), and the data showed no signs of being compromised by allelic dropout. The methodology presented here provides a framework by which to generate and evaluate microsatellite data from samples of much greater antiquity than attempted before, and opens new opportunities for ancient DNA research.

Conservation significance of island versus mainland populations: a case study of dibblers (Parantechinus apicalis) in Western Australia

Island populations are an interesting dichotomy in conservation biology. On the one hand, they can be a refuge for species where mainland populations have been decimated by loss of habitat and predation by exotic predators. On the other hand, island populations usually have reduced genetic diversity and are more susceptible to extinction through genetic and demographic processes. Genetic variation and morphological characters were measured for island and mainland populations of Parantechinus apicalis, small dasyurid marsupials, restricted to southwest Australia. Genetic variation at seven microsatellite loci revealed low levels of heterozygosity (He = 0.20 - 0.44) and high levels of inbreeding (Fe = 0.40 - 0.72) in island populations compared with the mainland population (He = 0.73). A nested clade analysis revealed that allopatric fragmentation was probably responsible for the association between geographical location and control region haplotypes, which is consistent with the isolation of populations on islands and indicative of two main populations of P. apicalis representing separate conservation units for management. While these results are typical of many island populations, they have important implications in terms of the conservation of threatened species in Australia and around the world, where island populations are a common source of founders for captive breeding and translocation to mainland sites.

The sociogenetic structure of a controlled feral pig population

In Australia, the feral pig (Sus scrofa) is a significant vertebrate pest that has an impact on agricultural production, public health and ecosystem integrity. Although feral pigs are controlled throughout much of their range, little is known about the impact that these control programs have had on the social biology, structure and the dispersal of pigs. To begin to address this, we collected demographic data and genetic samples from 123 feral pigs culled during a regional aerial shooting program over 33 pastoral properties in the semi-arid rangelands of southern Queensland, Australia. Sampling was carried out after two years of extensive control efforts (aerial 1080-baiting) and the samples therefore represented a controlled, persecuted population with a bias towards young animals. The analysis of 13 microsatellite loci suggested that females will accept multiple matings, females form loose mobs that appear to be highly dynamic social groups, and males will travel large distances between mobs. These data indicate that feral pigs in this population had a high level of social contact and form a single open population with no evidence of genetic (population) structuring. Such information may be important to integrate into management strategies, particularly the development of contingency plans regarding the spread of wildlife diseases.

A preliminary genetic study of the social biology of feral pigs in south-western Australia and the implications for management

A combination of demographic and genetic data was collected from 354 feral pigs (Sus scrofa), caught using standard trapping methods in south-western Australia, to provide preliminary information on their social biology. This included attempts to identify the putative parentage of 172 juvenile and foetal pigs, characterisation of the genetic mating system observed, and examination of the demographics of those individuals captured by current trapping methods. Findings revealed that in south-western Australia (a) feral pigs displayed moderately polygynous, but not polyandrous, mating behaviour, (b) breeding boars were significantly heavier than non-breeding boars, (c) the most reproductively successful boars, large individuals weighing >90 kg, moved the furthest in order to secure paternity, and (d) that a large proportion of breeding adults, particularly boars, were not captured under the standard trapping method employed. Ultimately, these data may be useful for the improvement of existing control programs, and exotic disease preparedness strategies.