African climate and geomorphology drive evolution and ghost introgression in sable antelope.

The evolutionary history of African ungulates has been explained largely in the light of Pleistocene climatic oscillations and the way these influenced the distribution of vegetation types, leading to range expansions and/or isolation in refugia. In contrast, comparatively fewer studies have addressed the continent's environmental heterogeneity and the role played by its geomorphological barriers. In this study, we performed a range-wide analysis of complete mitogenomes of sable antelope (Hippotragus niger) to explore how these different factors may have contributed as drivers of evolution in southcentral Africa. Our results supported two sympatric and deeply divergent mitochondrial lineages in west Tanzanian sables, which can be explained as the result of introgressive hybridization of a mitochondrial ghost lineage from an archaic, as-yet-undefined, congener. Phylogeographical subdivisions into three main lineages suggest that sable diversification may not have been driven solely by climatic events affecting populations differently across a continental scale. Often in interplay with climate, geomorphological features have also clearly shaped the species' patterns of vicariance, where the East Africa Rift System and the Eastern Arc Mountains acted as geological barriers. Subsequent splits among southern populations may be linked to rearrangements in the Zambezi system, possibly framing the most recent time when the river attained its current drainage profile. This work underlines how the use of comprehensive mitogenomic data sets on a model species with a wide geographical distribution can contribute to a much-enhanced understanding of environmental, geomorphological and evolutionary patterns in Africa throughout the Quaternary.

Human activity strongly influences genetic dynamics of the most widespread invasive plant in the sub-Antarctic.

The link between the successful establishment of alien species and propagule pressure is well-documented. Less known is how humans influence the post-introduction dynamics of invasive alien populations. The latter requires studying parallel invasions by the same species in habitats that are differently impacted by humans. We analysed microsatellite and genome size variation, and then compared the genetic diversity and structure of invasive Poa annua L. on two sub-Antarctic islands: human-occupied Marion Island and unoccupied Prince Edward Island. We also carried out niche modelling to map the potential distribution of the species on both islands. We found high levels of genetic diversity and evidence for extensive admixture between genetically distinct lineages of P. annua on Marion Island. By contrast, the Prince Edward Island populations showed low genetic diversity, no apparent admixture, and had smaller genomes. On both islands, high genetic diversity was apparent at human landing sites, and on Marion Island, also around human settlements, suggesting that these areas received multiple introductions and/or acted as initial introduction sites and secondary sources (bridgeheads) for invasive populations. More than 70 years of continuous human activity associated with a meteorological station on Marion Island led to a distribution of this species around human settlements and along footpaths, which facilitates ongoing gene flow among geographically separated populations. By contrast, this was not the case for Prince Edward Island, where P. annua populations showed high genetic structure. The high levels of genetic variation and admixture in P. annua facilitated by human activity, coupled with high habitat suitability on both islands, suggest that P. annua is likely to increase its distribution and abundance in the future.

The influence of landscape, climate and history on spatial genetic patterns in keystone plants (Azorella) on sub-Antarctic islands.

The distribution of genetic variation in species is governed by factors that act differently across spatial scales. To tease apart the contribution of different processes, especially at intermediate spatial scales, it is useful to study simple ecosystems such as those on sub-Antarctic oceanic islands. In this study, we characterize spatial genetic patterns of two keystone plant species, Azorella selago on sub-Antarctic Marion Island and Azorella macquariensis on sub-Antarctic Macquarie Island. Although both islands experience a similar climate and have a similar vegetation structure, they differ significantly in topography and geological history. We genotyped six microsatellites for 1,149 individuals from 123 sites across Marion Island and 372 individuals from 42 sites across Macquarie Island. We tested for spatial patterns in genetic diversity, including correlation with elevation and vegetation type, and clines in different directional bearings. We also examined genetic differentiation within islands, isolation-by-distance with and without accounting for direction, and signals of demographic change. Marion Island was found to have a distinct northwest-southeast divide, with lower genetic diversity and more sites with a signal of population expansion in the northwest. We attribute this to asymmetric seed dispersal by the dominant northwesterly winds, and to population persistence in a southwestern refugium during the Last Glacial Maximum. No apparent spatial pattern, but greater genetic diversity and differentiation between sites, was found on Macquarie Island, which may be due to the narrow length of the island in the direction of the dominant winds and longer population persistence permitted by the lack of extensive glaciation on the island. Together, our results clearly illustrate the implications of island shape and geography, and the importance of direction-dependent drivers, in shaping spatial genetic structure.

Evolutionary and ecological patterns within the South African Bathyergidae: Implications for taxonomy.

The family Bathyergidae (comprising six genera) is a group of subterranean rodents endemic to sub-Saharan Africa. Our understanding of the evolution and species richness of the South African bathyergid genera Georychus, Bathyergus and Cryptomys is limited, with the majority of species listed as Least Concern by the IUCN Red List of Threatened Species. Genetic data suggest that several cryptic species may be present in these genera. To explore genetic and ecological distinctiveness, and evaluate taxonomic richness across the ranges of Georychus, Bathyergus and to a lesser degree, Cryptomys, as well as evaluate possible scenarios which have historically influenced evolutionary patterns, we employed four protein coding markers (one mitochondrial and three nuclear) along with distribution wide sampling schemes and large sample sizes. In addition, possible ecological differences among the different intra-generic clades were explored. Genera appear to have originated in the north-eastern interior of South Africa, following novel habitats created through the Post-African I erosion cycle and dramatic changes in climate and phytogeography. In each genus, multiple geographically discrete genetic lineages (clades) are supported by both the mitochondrial and nuclear data. These lineages bear signature of the fragmentation of wider historical distributions through major environmental changes since the middle Miocene (major uplift events, Post-African II erosion cycle, drainage evolution of major river systems, sea-level fluctuations as well as climatic changes and vegetation shifts), thereby leading to long-term isolation. Along with protracted periods of separation, it appears that ecological differences further delimit the lineages in relation to geology, phytogeographic preference, elevation, rainfall and temperature. As such, two lineages in Georychus (Clades 1 and 2) and one lineages in Cryptomys (Clade I) occur at higher elevations above the Great Escarpment (in older deposits harbouring grassland vegetation, with higher rainfall and lower daily temperatures), with the remaining lineages within these genera (Clades 3, 4 and 5 in Georychus and Clades III and IV in Cryptomys) occupying a low-land distribution with contrasting climatic and geological characteristics. Although significant differences in ecological variables were also observed between Bathyergus clades, these were not consistent, given their largely low-land distributions. Our results corroborate and expand previous suggestions that several cryptic species are present within the South African Bathyergidae.

The effect of terrain on the fine-scale genetic diversity of sub-Antarctic Collembola: A landscape genetics approach.

Biodiversity patterns are shaped by the interplay between geodiversity and organismal characteristics. Superimposing genetic structure onto landscape heterogeneity (i.e., landscape genetics) can help to disentangle their interactions and better understand population dynamics. Previous studies on the sub-Antarctic Prince Edward Islands (located midway between Antarctica and Africa) have highlighted the importance of landscape and climatic barriers in shaping spatial genetic patterns and have drawn attention to the value of these islands as natural laboratories for studying fundamental concepts in biology. Here, we assessed the fine-scale spatial genetic structure of the springtail, Cryptopygus antarcticus travei, which is endemic to Marion Island, in tandem with high-resolution geological data. Using a species-specific suite of microsatellite markers, a fine-scale sampling design incorporating landscape complexity and generalised linear models (GLMs), we examined genetic patterns overlaid onto high-resolution digital surface models and surface geology data across two 1-km sampling transects. The GLMs revealed that genetic patterns across the landscape closely track landscape resistance data in concert with landscape discontinuities and barriers to gene flow identified at a scale of a few metres. These results show that the island's geodiversity plays an important role in shaping biodiversity patterns and intraspecific genetic diversity. This study illustrates that fine-scale genetic patterns in soil arthropods are markedly more structured than anticipated, given that previous studies have reported high levels of genetic diversity and evidence of genetic structing linked to landscape changes for springtail species and considering the homogeneity of the vegetation complexes characteristic of the island at the scale of tens to hundreds of metres. By incorporating fine-scale and high-resolution landscape features into our study, we were able to explain much of the observed spatial genetic patterns. Our study highlights geodiversity as a driver of spatial complexity. More widely, it holds important implications for the conservation and management of the sub-Antarctic islands.

Molecular tracking and prevalence of the red colour morph restricted to a harvested leopard population in South Africa.

The red leopard (Panthera pardus) colour morph is a colour variant that occurs only in South Africa, where it is confined to the Central Bushveld bioregion. Red leopards have been spreading over the past 40 years, which raises the speculation that the prevalence of this phenotype is related to low dispersal of young individuals owing to high off-take in the region. Intensive selective hunting tends to remove large resident male leopards from the breeding population, which gives young male leopards the chance to mate with resident female leopards that are more likely to be their relatives, eventually increasing the frequency of rare genetic variants. To investigate the genetic mechanisms underlying the red coat colour morph in leopards, and whether its prevalence in South Africa relates to an increase in genetic relatedness in the population, we sequenced exons of six coat colour-associated genes and 20 microsatellite loci in twenty Wild-type and four red leopards. The results were combined with demographic data available from our study sites. We found that red leopards own a haplotype in homozygosity identified by two SNPs and a 1 bp deletion that causes a frameshift in the tyrosinase-related protein 1 (TYRP1), a gene known to be involved in the biosynthesis of melanin. Microsatellite analyses indicate clear signs of a population bottleneck and a relatedness of 0.11 among all pairwise relationships, eventually supporting our hypothesis that a rare colour morph in the wild has increased its local frequency due to low natal dispersal, while subject to high human-induced mortality rate.

Conservation priorities in an endangered estuarine seahorse are informed by demographic history.

Historical demographic events shape genetic diversity that remains evident in the genomes of contemporary populations. In the case of species that are of conservation concern, this information helps to unravel evolutionary histories that can be critical in guiding conservation efforts. The Knysna seahorse, Hippocampus capensis, is the world's most endangered seahorse species, and it presently survives in only three estuaries on the South African south coast. Factors that contributed to the species becoming endangered are unclear; additionally, the lack of information on whether the three populations should be managed separately because of potential long-term isolation hampers effective management efforts. In the present study, we reconstructed the seahorses' demographic history using a suite of microsatellite loci. We found that the largest population (Knysna Estuary) has colonised the other estuaries relatively recently (< 450 years ago), and that its population size is comparatively large and stable. Neither of the other two populations shows signs of long-term reductions in population size. The high conservation status of the species is thus a result of its limited range rather than historical population declines. Our findings indicate that the long-term survival of H. capensis depends primarily on the successful management of the Knysna population, although the other estuaries may serve as reservoirs of genetic diversity.

Correction: Evolutionary history of Carnivora (Mammalia, Laurasiatheria) inferred from mitochondrial genomes.

[This corrects the article DOI: 10.1371/journal.pone.0240770.].

The complete mitogenome of Leptestheria brevirostris Barnard, 1924, a rock pool clam shrimp (Branchiopoda: Spinicaudata) from Central District, Botswana.

Spinicaudatan clam shrimp are a widespread and diverse group of branchiopod crustaceans, yet few mitochondrial genomes have been published for this taxonomic group. Here, we present the mitogenome of Leptestheria brevirostris from a rock pool ecosystem in Botswana. Massively parallel sequencing of a single specimen facilitated the reconstruction of the species' 15,579 bp circularized mitogenome. The reconstructed phylogenetic tree confirms that L. brevirostris forms a monophyletic group with other diplostracan branchiopods, and that these are the sister taxon to Notostraca. The mitogenome reconstructed here is the first to be reported from a leptestherid clam shrimp.

De novo whole-genome assembly and resequencing resources for the roan (Hippotragus equinus), an iconic African antelope.

Roan antelope (Hippotragus equinus) is the second-largest member of the Hippotraginae (Bovidae), and is widely distributed across sub-Saharan mesic woodlands. Despite being listed as "Least Concern" across its African range, population numbers are decreasing with many regional Red List statuses varying between Endangered and Locally Extinct. Although the roan antelope has become an economically-important game species in Southern Africa, the vast majority of wild populations are found only in fragmented protected areas, which is of conservation concern. Genomic information is crucial in devising optimal management plans. To this end, we report here the first de novo assembly and annotation of the whole-genome sequence of a male roan antelope from a captive-breeding program. Additionally, we uncover single-nucleotide variants (SNVs) through re-sequencing of five wild individuals representing five of the six described subspecies. We used 10X Genomics Chromium chemistry to produce a draft genome of 2.56 Gb consisting of 16,880 scaffolds with N50 = 8.42 Mb and a BUSCO completeness of 91.2%. The draft roan genome includes 1.1 Gbp (42.2%) repetitive sequences. De novo annotation identified 20,518 protein-coding genes. Genome synteny to the domestic cow showed an average identity of 92.7%. Re-sequencing of five wild individuals to an average sequencing depth of 9.8x resulted in the identification of a filtered set of 3.4x106 bi-allelic SNVs. The proportion of alternative homozygous SNVs for the individuals representing different subspecies, as well as differentiation as measured by PCA, were consistent with expected divergence from the reference genome and among samples. The roan antelope genome is a valuable resource for evolutionary and population genomic questions, as well as management and conservation actions.

Evolutionary history of Carnivora (Mammalia, Laurasiatheria) inferred from mitochondrial genomes.

The order Carnivora, which currently includes 296 species classified into 16 families, is distributed across all continents. The phylogeny and the timing of diversification of members of the order are still a matter of debate. Here, complete mitochondrial genomes were analysed to reconstruct the phylogenetic relationships and to estimate divergence times among species of Carnivora. We assembled 51 new mitogenomes from 13 families, and aligned them with available mitogenomes by selecting only those showing more than 1% of nucleotide divergence and excluding those suspected to be of low-quality or from misidentified taxa. Our final alignment included 220 taxa representing 2,442 mitogenomes. Our analyses led to a robust resolution of suprafamilial and intrafamilial relationships. We identified 21 fossil calibration points to estimate a molecular timescale for carnivorans. According to our divergence time estimates, crown carnivorans appeared during or just after the Early Eocene Climatic Optimum; all major groups of Caniformia (Cynoidea/Arctoidea; Ursidae; Musteloidea/Pinnipedia) diverged from each other during the Eocene, while all major groups of Feliformia (Nandiniidae; Feloidea; Viverroidea) diversified more recently during the Oligocene, with a basal divergence of Nandinia at the Eocene/Oligocene transition; intrafamilial divergences occurred during the Miocene, except for the Procyonidae, as Potos separated from other genera during the Oligocene.

Transcriptomic Diversity in the Livers of South African Sardines Participating in the Annual Sardine Run.

During austral winter, the southern and eastern coastlines of South Africa witness one of the largest animal migrations on the planet, the KwaZulu-Natal sardine run. Hundreds of millions of temperate sardines, Sardinops sagax, form large shoals that migrate north-east towards the subtropical Indian Ocean. Recent studies have highlighted the role that genetic and environmental factors play in sardine run formation. In the present study, we used massively parallel sequencing to assemble and annotate the first reference transcriptome from the liver cells of South African sardines, and to investigate the functional content and transcriptomic diversity. A total of 1,310,530 transcripts with an N50 of 1578 bp were assembled de novo. Several genes and core biochemical pathways that modulate energy production, energy storage, digestion, secretory processes, immune responses, signaling, regulatory processes, and detoxification were identified. The functional content of the liver transcriptome from six individuals that participated in the 2019 sardine run demonstrated heterogeneous levels of variation. Data presented in the current study provide new insights into the complex function of the liver transcriptome in South African sardines.

Environmental DNA Metabarcoding as a Means of Estimating Species Diversity in an Urban Aquatic Ecosystem.

Adaptation to environments that are changing as a result of human activities is critical to species' survival. A large number of species are adapting to, and even thriving in, urban green spaces, but this diversity remains largely undocumented. In the current study, we explored the potential of environmental DNA (eDNA) to document species diversity in one of the largest green spaces in Johannesburg, South Africa. Using a novel metabarcoding approach that assembles short DNA fragments suitable for massively parallel sequencing platforms to the approximate standard ~710 bp COI barcoding fragment, we document the presence of 26 phyla, 52 classes, 134 orders, 289 families, 380 genera and 522 known species from the study site. Our results highlight the critical role that urban areas play in protecting the world's declining biodiversity.

Physiological tolerances account for range limits and abundance structure in an invasive slug.

Despite the importance of understanding the mechanisms underlying range limits and abundance structure, few studies have sought to do so. Here we use a terrestrial slug species, Deroceras panormitanum, that has invaded a remote, largely predator-free, Southern Ocean island as a model system to do so. Across Marion Island, slug density does not conform to an abundant centre distribution. Rather, abundance structure is characterized by patches and gaps. These are associated with this desiccation-sensitive species' preference for biotic and drainage line habitats that share few characteristics except for their high humidity below the vegetation surface. The coastal range margin has a threshold form, rapidly rising from zero to high density. Slugs do not occur where soil-exchangeable Na values are higher than 3000 mg kg(-1), and in laboratory experiments, survival is high below this value but negligible above it. Upper elevation range margins are a function of the inability of this species to survive temperatures below an absolute limit of -6.4 degrees C, which is regularly exceeded at 200 m altitude, above which slug density declines to zero. However, the linear decline in density from the coastal peak is probably also a function of a decline in performance or time available for activity. This is probably associated with an altitudinal decline in mean annual soil temperature. These findings support previous predictions made regarding the form of density change when substrate or climatic factors set range limits.

Phylogeny and biogeography of the African Bathyergidae: a review of patterns and processes.

BACKGROUND: We review genealogical relationships, biogeographic patterns and broad historical drivers of speciation within the Bathyergidae, a group of endemic African rodents, as well as identify key taxa which need further research. METHODS: We sourced comparable cytochrome b sequence data (comparable data available for all members for the Family) and geographic information for all six genera of the African subterranean rodent. This information was combined into the most comprehensive and geographically representative evolutionary study for the Bathyergidae to date. RESULTS: Species richness within the Bathyergidae appears to be underestimated, with undescribed taxa in five of the six genera. Biogeographic patterns suggest large historical distributions, which were repeatedly fragmented by major landscape changes (especially rifting, uplift and drainage evolution) since the Miocene. Aside from vicariant events, other factors (ecological specialization, population-level responses and climatic change) may have been instrumental in driving divergences in the Bathyergidae. As such, adaptive differences may exist among both populations and species across their discrete ranges, driving independent evolutionary trajectories among taxa. In addition, highly fragmented distributions of divergent (and often relict) lineages indicates the possibility of narrow endemics restricted to diminishing suitable habitats. From this, it is clear that a systematic revision of the Bathyergidae is necessary; such a revision should include comprehensive sampling of all putative taxa, the addition of genomic information to assess adaptive differences, as well as ecological information.

Assessing introgressive hybridization in roan antelope (Hippotragus equinus): Lessons from South Africa.

Biological diversity is being lost at unprecedented rates, with genetic admixture and introgression presenting major threats to biodiversity. Our ability to accurately identify introgression is critical to manage species, obtain insights into evolutionary processes, and ultimately contribute to the Aichi Targets developed under the Convention on Biological Diversity. The current study concerns roan antelope, the second largest antelope in Africa. Despite their large size, these antelope are sensitive to habitat disturbance and interspecific competition, leading to the species being listed as Least Concern but with decreasing population trends, and as extinct over parts of its range. Molecular research identified the presence of two evolutionary significant units across their sub-Saharan range, corresponding to a West African lineage and a second larger group which includes animals from East, Central and Southern Africa. Within South Africa, one of the remaining bastions with increasing population sizes, there are a number of West African roan antelope populations on private farms, and concerns are that these animals hybridize with roan that naturally occur in the southern African region. We used a suite of 27 microsatellite markers to conduct admixture analysis. Our results indicate evidence of hybridization, with our developed tests using a simulated dataset being able to accurately identify F1, F2 and non-admixed individuals at threshold values of qi > 0.80 and qi > 0.85. However, further backcrosses were not always detectable with backcrossed-Western roan individuals (46.7-60%), backcrossed-East, Central and Southern African roan individuals (28.3-45%) and double backcrossed (83.3-98.3%) being incorrectly classified as non-admixed. Our study is the first to confirm ongoing hybridization in this within this iconic African antelope, and we provide recommendations for the future conservation and management of this species.

The genetic tale of a recovering lion population (Panthera leo) in the Savé Valley region (Zimbabwe): A better understanding of the history and managing the future.

The rapid decline of the African lion (Panthera leo) has raised conservation concerns. In the Savé Valley Conservancy (SVC), in the Lowveld of Zimbabwe, lions were presumably reduced to approximately 5 to 10 individuals. After ten lions were reintroduced in 2005, the population has recovered to over 200 lions in 2016. Although the increase of lions in the SVC seems promising, a question remains whether the population is genetically viable, considering their small founding population. In this study, we document the genetic diversity in the SVC lion population using both mitochondrial and nuclear genetic markers, and compare our results to literature from other lion populations across Africa. We also tested whether genetic diversity is spatially structured between lion populations residing on several reserves in the Lowveld of Zimbabwe. A total of 42 lions were genotyped successfully for 11 microsatellite loci. We confirmed that the loss of allelic richness (probably resulting from genetic drift and small number of founders) has resulted in low genetic diversity and inbreeding. The SVC lion population was also found to be genetically differentiated from surrounding population, as a result of genetic drift and restricted natural dispersal due to anthropogenic barriers. From a conservation perspective, it is important to avoid further loss of genetic variability in the SVC lion population and maintain evolutionary potential required for future survival. Genetic restoration through the introduction of unrelated individuals is recommended, as this will increase genetic heterozygosity and improve survival and reproductive fitness in populations.

Spatial genetic diversity in the Cape mole-rat, Georychus capensis: Extreme isolation of populations in a subterranean environment.

The subterranean niche harbours animals with extreme adaptations. These adaptations decrease the vagility of taxa and, along with other behavioural adaptations, often result in isolated populations characterized by small effective population sizes, high inbreeding, population bottlenecks, genetic drift and consequently, high spatial genetic structure. Although information is available for some species, estimates of genetic diversity and whether this variation is spatially structured, is lacking for the Cape mole-rat (Georychus capensis). By adopting a range-wide sampling regime and employing two variable mitochondrial markers (cytochrome b and control region), we report on the effects that life-history, population demography and geographic barriers had in shaping genetic variation and population genetic patterns in G. capensis. We also compare our results to information available for the sister taxon of the study species, Bathyergus suillus. Our results show that Georychus capensis exhibits low genetic diversity relative to the concomitantly distributed B. suillus, most likely due to differences in habitat specificity, habitat fragmentation and historical population declines. In addition, the isolated nature of G. capensis populations and low levels of population connectivity has led to small effective population sizes and genetic differentiation, possibly aided by genetic drift. Not surprisingly therefore, G. capensis exhibits pronounced spatial structure across its range in South Africa. Along with geographic distance and demography, other factors shaping the genetic structure of G. capensis include the historical and contemporary impacts of mountains, rivers, sea-level fluctuations and elevation. Given the isolation and differentiation among G. capensis populations, the monotypic genus Georychus may represent a species complex.

Mitochondrial DNA is unsuitable to test for isolation by distance.

Tests for isolation by distance (IBD) are the most commonly used method of assessing spatial genetic structure. Many studies have exclusively used mitochondrial DNA (mtDNA) sequences to test for IBD, but this marker is often in conflict with multilocus markers. Here, we report a review of the literature on IBD, with the aims of determining (a) whether significant IBD is primarily a result of lumping spatially discrete populations, and (b) whether microsatellite datasets are more likely to detect IBD when mtDNA does not. We also provide empirical data from four species in which mtDNA failed to detect IBD by comparing these with microsatellite and SNP data. Our results confirm that IBD is mostly found when distinct regional populations are pooled, and this trend disappears when each is analysed separately. Discrepancies between markers were found in almost half of the studies reviewed, and microsatellites were more likely to detect IBD when mtDNA did not. Our empirical data rejected the lack of IBD in the four species studied, and support for IBD was particularly strong for the SNP data. We conclude that mtDNA sequence data are often not suitable to test for IBD, and can be misleading about species' true dispersal potential. The observed failure of mtDNA to reliably detect IBD, in addition to being a single-locus marker, is likely a result of a selection-driven reduction in genetic diversity obscuring spatial genetic differentiation.

Local and regional scale genetic variation in the Cape dune mole-rat, Bathyergus suillus.

The distribution of genetic variation is determined through the interaction of life history, morphology and habitat specificity of a species in conjunction with landscape structure. While numerous studies have investigated this interplay of factors in species inhabiting aquatic, riverine, terrestrial, arboreal and saxicolous systems, the fossorial system has remained largely unexplored. In this study we attempt to elucidate the impacts of a subterranean lifestyle coupled with a heterogeneous landscape on genetic partitioning by using a subterranean mammal species, the Cape dune mole-rat (Bathyergus suillus), as our model. Bathyergus suillus is one of a few mammal species endemic to the Cape Floristic Region (CFR) of the Western Cape of South Africa. Its distribution is fragmented by rivers and mountains; both geographic phenomena that may act as geographical barriers to gene-flow. Using two mitochondrial fragments (cytochrome b and control region) as well as nine microsatellite loci, we determined the phylogeographic structure and gene-flow patterns at two different spatial scales (local and regional). Furthermore, we investigated genetic differentiation between populations and applied Bayesian clustering and assignment approaches to our data. Nearly every population formed a genetically unique entity with significant genetic structure evident across geographic barriers such as rivers (Berg, Verlorenvlei, Breede and Gourits Rivers), mountains (Piketberg and Hottentots Holland Mountains) and with geographic distance at both spatial scales. Surprisingly, B. suillus was found to be paraphyletic with respect to its sister species, B. janetta-a result largely overlooked by previous studies on these taxa. A systematic revision of the genus Bathyergus is therefore necessary. This study provides a valuable insight into how the biology, life-history and habitat specificity of animals inhabiting a fossorial system may act in concert with the structure of the surrounding landscape to influence genetic distinctiveness and ultimately speciation.