New insights from RADseq data on differentiation in the Hottentot golden mole species complex from South Africa.

Golden moles (Family Chrysochloridae) are small subterranean mammals, endemic to sub-Saharan Africa, and many of the 21 species are listed as threatened on the IUCN Red List. Most species have highly restricted ranges; however two species, the Hottentot golden mole (Amblysomus hottentotus) and the Cape golden mole (Chrysochloris asiatica) have relatively wide ranges. We recently uncovered cryptic diversity within A. hottentotus, through a phylogeographic analysis of this taxon using two mitochondrial gene regions and a nuclear intron. To further investigate this cryptic diversity, we generated nuclear SNP data from across the genome of A. hottentotus, by means of double-digest restriction-site associated DNA sequencing (ddRADSeq), and mapped reads to the Cape golden mole genome. We conducted a phylogenetic analysis and investigated population differentiation. Our results support the distinctiveness of A. h. meesteri. Furthermore, we provide evidence from nuclear SNPs in support of our previous finding that Central coastal samples represent a unique cryptic lineage that is highly divergent from A. h. pondoliae farther south. Although mtDNA suggests that Umtata may represent a unique lineage sister to A. h. longiceps, mito-nuclear discordance from our RADseq data indicate that these samples may instead be closer to A. h. pondoliae, and therefore may not represent a distinct lineage. We stress the importance of recognizing that understudied populations, such as that of Umtata, may represent populations or ESUs under threat and in need of conservation attention. We present a high-quality filtered SNP dataset, comprising thousands of SNPs, which may serve as a useful resource for future golden mole studies. We have thus added to the growing body of research demonstrating the power and utility of RADseq to investigate population differentiation.

Characterization of genome-wide microsatellite markers in rabbitfishes, an important resource for artisanal fisheries in the Indo-West Pacific.

Rabbitfishes are reef-associated fishes that support local fisheries throughout the Indo-West Pacific region. Sound management of the resource requires the development of molecular tools for appropriate stock delimitation of the different species in the family. Microsatellite markers were developed for the cordonnier, Siganus sutor, and their potential for cross-amplification was investigated in 12 congeneric species. A library of 792 repeat-containing sequences was built. Nineteen sets of newly developed primers, and 14 universal finfish microsatellites were tested in S. sutor. Amplification success of the 19 Siganus-specific markers ranged from 32 to 79% in the 12 other Siganus species, slightly decreasing when the genetic distance of the target species to S. sutor increased. Seventeen of these markers were polymorphic in S. sutor and were further assayed in S. luridus, S. rivulatus, and S. spinus, of which respectively 9, 10 and 8 were polymorphic. Statistical power analysis and an analysis of molecular variance showed that subtle genetic differentiation can be detected using these markers, highlighting their utility for the study of genetic diversity and population genetic structure in rabbitfishes.

Life on the rocks: Multilocus phylogeography of rock hyrax (Procavia capensis) from southern Africa.

Understanding the role of geography and climatic cycles in determining patterns of biodiversity is important in comparative and evolutionary biology and conservation. We studied the phylogeographic pattern and historical demography of a rock-dwelling small mammal species from southern Africa, the rock hyrax Procavia capensis capensis. Using a multilocus coalescent approach, we assessed the influence of strong habitat dependence and fluctuating regional climates on genetic diversity. We sequenced a mitochondrial gene (cytochrome b) and two nuclear introns (AP5, PRKC1) supplemented with microsatellite genotyping, in order to assess evolutionary processes over multiple temporal scales. In addition, distribution modelling was used to investigate the current and predicted distribution of the species under different climatic scenarios. Collectively, the data reveal a complex history of isolation followed by secondary contact shaping the current intraspecific diversity. The cyt b sequences confirmed the presence of two previously proposed geographically and genetically distinct lineages distributed across the southern African Great Escarpment and north-western mountain ranges. Molecular dating suggests Miocene divergence of the lineages, yet there are no discernible extrinsic barriers to gene flow. The nuclear markers reveal incomplete lineage sorting or ongoing mixing of the two lineages. Although the microsatellite data lend some support to the presence of two subpopulations, there is weak structuring within and between lineages. These data indicate the presence of gene flow from the northern into the southern parts of the southern African sub-region likely following the secondary contact. The distribution modelling predictably reveal the species' preference for rocky areas, with stable refugia through time in the northern mountain ranges, the Great Escarpment, as well as restricted areas of the Northern Cape Province and the Cape Fold Mountains of South Africa. Different microclimatic variables appear to determine the distributional range of the species. Despite strong habitat preference, the micro-habitat offered by rocky crevices and unique life history traits likely promoted the adaptability of P. capensis, resulting in the widespread distribution and persistence of the species over a long evolutionary period. Spatio-temporal comparison of the evolutionary histories of other co-distributed species across the rocky landscapes of southern Africa will improve our understanding of the regional patterns of biodiversity and local endemism.

Spatio-temporal genetic structure and the effects of long-term fishing in two partially sympatric offshore demersal fishes.

Environmental gradients have been shown to disrupt gene flow in marine species, yet their influence in structuring populations at depth remains poorly understood. The Cape hakes (Merluccius paradoxus and M. capensis) are demersal species co-occurring in the Benguela Current system, where decades of intense fishing resulted in severely depleted stocks in the past. Previous studies identified conflicting mtDNA genetic substructuring patterns and thus contrasting evolutionary trajectories for both species. Using 10 microsatellite loci, the control region of mtDNA and employing a seascape genetics approach, we investigated genetic connectivity and the impact of prolonged exploitation in the two species, which are characterized by different patterns of fishing pressure. Three consecutive years were sampled covering the entire distribution (N = 2100 fishes). Despite large estimated population sizes, both species exhibited low levels of contemporary genetic diversity (0.581 < HE  < 0.692), implying that fishing has had a significant impact on their genetic composition and evolutionary trajectories. Further, for M. paradoxus, significant temporal, but not spatial, divergence points to the presence of genetic chaotic patchiness. In contrast, M. capensis exhibited a clear latitudinal cline in genetic differentiation between Namibia and South Africa (FST  = 0.063, P < 0.05), with low (0.2% per generation) estimates of contemporary gene flow. Seascape analyses reveal an association with bathymetry and upwelling events, suggesting that adaptation to local environmental conditions may drive genetic differentiation in M. capensis. Importantly, our results highlight the need for temporal sampling in disentangling the complex factors that impact population divergence in marine fishes.

Investigating the origin of vagrant dusky groupers, Epinephelus marginatus (Lowe, 1834), in coastal waters of Réunion Island.

Due to their geographic isolation, biotas of oceanic islands are likely influenced by episodic long distance dispersal events, but such observations are scarce. In June 2012, fishermen from Réunion Island caught an unknown specimen of grouper, identified as dusky grouper Epinephelus marginatus (Lowe, 1834). This was highly unexpected considering the large distance of its closest verified occurrence (South Africa, ∼2500km). To identify the origin of this specimen and the mechanisms driving this potential long distance colonization, we combined genetic analyses and hydrodynamic connectivity modeling approaches. Molecular markers and samples from various locations across the distribution range resulted in the identification of three putative source populations. The Réunion specimen clustered genetically with South Africa. The estimated spawning period in relation to the connectivity modeling of larvae showed no possible direct connection between South Africa and Réunion. However, connectivity was predicted through intermediate stepping stone populations likely located around the southern tip of Madagascar, where the occurrence of the species has yet to be verified. The results further highlight the potential role of the cyclone Bingiza (February 2011) in the connection between Madagascar and Réunion. This shows that cyclones may be an important driver in long distance colonization of oceanic islands.

Evaluation of microsatellite markers for populations studies and forensic identification of African lions (Panthera leo).

The South African lion (Panthera leo) population is highly fragmented. One-third of its wild lions occur in small (<1000 km(2)) reserves. These lions were reintroduced from other areas of the species' historical range. Management practices on these reserves have not prioritized genetic provenance or heterozygosity. These trends potentially constrain the conservation value of these lions. To ensure the best management and long-term survival of these subpopulations as a viable collective population, the provenance and current genetic diversity must be described. Concurrently, poaching of lions to supply a growing market for lion bones in Asia may become a serious conservation challenge in the future. Having a standardized, validated method for matching confiscated lion parts with carcasses will be a key tool in investigating these crimes. We evaluated 28 microsatellites in the African lion using samples from 18 small reserves and 1 captive facility in South Africa, two conservancies in Zimbabwe, and Kruger National and Kgalagadi Transfrontier Parks to determine the loci most suited for population management and forensic genetic applications. Twelve microsatellite loci with a match probability of 1.1×10(-5) between siblings were identified for forensics. A further 10 could be added for population genetics studies.

Multilocus phylogeny of the avian family Alaudidae (larks) reveals complex morphological evolution, non-monophyletic genera and hidden species diversity.

The Alaudidae (larks) is a large family of songbirds in the superfamily Sylvioidea. Larks are cosmopolitan, although species-level diversity is by far largest in Africa, followed by Eurasia, whereas Australasia and the New World have only one species each. The present study is the first comprehensive phylogeny of the Alaudidae. It includes 83.5% of all species and representatives from all recognised genera, and was based on two mitochondrial and three nuclear loci (in total 6.4 kbp, although not all loci were available for all species). In addition, a larger sample, comprising several subspecies of some polytypic species was analysed for one of the mitochondrial loci. There was generally good agreement in trees inferred from different loci, although some strongly supported incongruences were noted. The tree based on the concatenated multilocus data was overall well resolved and well supported by the data. We stress the importance of performing single gene as well as combined data analyses, as the latter may obscure significant incongruence behind strong nodal support values. The multilocus tree revealed many unpredicted relationships, including some non-monophyletic genera (Calandrella, Mirafra, Melanocorypha, Spizocorys). The tree based on the extended mitochondrial data set revealed several unexpected deep divergences between taxa presently treated as conspecific (e.g. within Ammomanes cinctura, Ammomanes deserti, Calandrella brachydactyla, Eremophila alpestris), as well as some shallow splits between currently recognised species (e.g. Certhilauda brevirostris-C. semitorquata-C. curvirostris; Calendulauda barlowi-C. erythrochlamys; Mirafra cantillans-M. javanica). Based on our results, we propose a revised generic classification, and comment on some species limits. We also comment on the extraordinary morphological adaptability in larks, which has resulted in numerous examples of parallel evolution (e.g. in Melanocorypha mongolica and Alauda leucoptera [both usually placed in Melanocorypha]; Ammomanopsis grayi and Ammomanes cinctura/deserti [former traditionally placed in Ammomanes]; Chersophilus duponti and Certhilauda spp.; Eremopterix hova [usually placed in Mirafra] and several Mirafra spp.), as well as both highly conserved plumages (e.g. within Mirafra) and strongly divergent lineages (e.g. Eremopterix hova vs. other Eremopterix spp.; Calandrella cinerea complex vs. Eremophila spp.; Eremalauda dunni vs. Chersophilus duponti; Melanocorypha mongolica and male M. yeltoniensis vs. other Melanocorypha spp. and female M. yeltoniensis). Sexual plumage dimorphism has evolved multiple times. Few groups of birds show the same level of disagreement between taxonomy based on morphology and phylogenetic relationships as inferred from DNA sequences.

The Contribution of Digital Sequence Information to Conservation Biology: A Southern African Perspective.

Many recent contributions have made a compelling case that genetic diversity is not adequately reflected in international frameworks and policies, as well as in local governmental processes implementing such frameworks. Using digital sequence information (DSI) and other publicly available data is supported to assess genetic diversity, toward formulation of practical actions for long-term conservation of biodiversity, with the particular goal of maintaining ecological and evolutionary processes. Given the inclusion of specific goals and targets regarding DSI in the latest draft of the Global Biodiversity Framework negotiated at the 15th Conference of the Parties (COP15) in Montreal in December 2022 and the crucial decisions on access and benefit sharing to DSI that will be taken in the coming months and future COP meetings, a southern African perspective on how and why open access to DSI is essential for the conservation of intraspecific biodiversity (genetic diversity and structure) across country borders is provided.

Ancestral polymorphism at the major histocompatibility complex (MHCIIß) in the Nesospiza bunting species complex and its sister species (Rowettia goughensis).

BACKGROUND: The major histocompatibility complex (MHC) is an important component of the vertebrate immune system and is frequently used to characterise adaptive variation in wild populations due to its co-evolution with pathogens. Passerine birds have an exceptionally diverse MHC with multiple gene copies and large numbers of alleles compared to other avian taxa. The Nesospiza bunting species complex (two species on Nightingale Island; one species with three sub-species on Inaccessible Island) represents a rapid adaptive radiation at a small, isolated archipelago, and is thus an excellent model for the study of adaptation and speciation. In this first study of MHC in Nesospiza buntings, we aim to characterize MHCIIß variation, determine the strength of selection acting at this gene region and assess the level of shared polymorphism between the Nesospiza species complex and its putative sister taxon, Rowettia goughensis, from Gough Island. RESULTS: In total, 23 unique alleles were found in 14 Nesospiza and 2 R. goughensis individuals encoding at least four presumably functional loci and two pseudogenes. There was no evidence of ongoing selection on the peptide binding region (PBR). Of the 23 alleles, 15 were found on both the islands inhabited by Nesospiza species, and seven in both Nesospiza and Rowettia; indications of shared, ancestral polymorphism. A gene tree of Nesospiza MHCIIß alleles with several other passerine birds shows three highly supported Nesospiza-specific groups. All R. goughensis alleles were shared with Nesospiza, and these alleles were found in all three Nesospiza sequence groups in the gene tree, suggesting that most of the observed variation predates their phylogenetic split. CONCLUSIONS: Lack of evidence of selection on the PBR, together with shared polymorphism across the gene tree, suggests that population variation of MHCIIß among Nesospiza and Rowettia is due to ancestral polymorphism rather than local selective forces. Weak or no selection pressure could be attributed to low parasite load at these isolated Atlantic islands. The deep divergence between the highly supported Nesospiza-specific sequence Groups 2 and 3, and the clustering of Group 3 close to the distantly related passerines, provide strong support for preserved ancestral polymorphism, and present evidence of one of the rare cases of extensive ancestral polymorphism in birds.

Spatial population genetic structure and colony dynamics in Damaraland mole-rats (Fukomys damarensis) from the southern Kalahari.

BACKGROUND: Non-random associations within and among groups of social animals can provide valuable insight into the function of group living and the evolution of social behaviour. Damaraland mole-rats (Fukomys damarensis) demonstrate extremely high levels of reproductive skew, and dispersal is considered to be male-biased in onset and frequency, although asymmetry in dispersal distance is yet to be investigated. Dispersal may be positively correlated with increasing favourable environmental conditions, such as rainfall, however, the effects of ecological constraints on dispersal and colony fission-fusion dynamics have not previously been demonstrated on a spatial scale. Here we provide the first spatial population genetic study for this species. We investigated genetic structure in a population of Damaraland mole-rats from the southern Kalahari in South Africa over 3 years, combining observational dispersal data from mark-recapture with population genetic data to evaluate (1) sex-bias in frequency and distance of dispersal in this species, and (2) the effect of rainfall on fission-fusion dynamics of colonies. RESULTS: Our results demonstrate (1) that both males and females favour local dispersal but on rare occasions may disperse over distances greater than 400 m, (2) that males may disperse over greater distances than females, and (3) that males more frequently immigrate into established neighbouring colonies than females, who predominantly disperse by colony fission, i.e. multiple individuals "budding" from their native colony into a neighbouring territory, thereby establishing new colonies. Furthermore, our results demonstrate (4) elevated dispersal and colony fission in association with increased rainfall, supporting the hypothesis that rainfall may play a significant role in the maintenance and/or disruption of reproductive skew in Damaraland mole-rat populations. CONCLUSION: This study represents the first fine-scale spatial population genetic study in Damaraland mole-rats, and provides relevant insights into colony fission-fusion dynamics in a social and cooperatively breeding species.

High diversity, inbreeding and a dynamic Pleistocene demographic history revealed by African buffalo genomes.

Genomes retain records of demographic changes and evolutionary forces that shape species and populations. Remnant populations of African buffalo (Syncerus caffer) in South Africa, with varied histories, provide an opportunity to investigate signatures left in their genomes by past events, both recent and ancient. Here, we produce 40 low coverage (7.14×) genome sequences of Cape buffalo (S. c. caffer) from four protected areas in South Africa. Genome-wide heterozygosity was the highest for any mammal for which these data are available, while differences in individual inbreeding coefficients reflected the severity of historical bottlenecks and current census sizes in each population. PSMC analysis revealed multiple changes in Ne between approximately one million and 20 thousand years ago, corresponding to paleoclimatic changes and Cape buffalo colonisation of southern Africa. The results of this study have implications for buffalo management and conservation, particularly in the context of the predicted increase in aridity and temperature in southern Africa over the next century as a result of climate change.

Bioregion heterogeneity correlates with extensive mitochondrial DNA diversity in the Namaqua rock mouse, Micaelamys namaquensis (Rodentia: Muridae) from southern Africa--evidence for a species complex.

BACKGROUND: Intraspecific variation within the diverse southern African murine rodents has not been extensively investigated, yet cryptic diversity is evident in several taxa studied to date. The Namaqua rock mouse, Micaelamys namaquensis Smith, 1834 is a widespread endemic murine rodent from the subregion. Currently, a single species with four subspecies is recognised, but in the past up to 16 subspecies were described. Thus, this species is a good candidate for the investigation of patterns and processes of diversification in a diverse but under-studied mammalian subfamily and geographic region. Here, we report genetic differentiation based on mitochondrial DNA (mtDNA) cytochrome b (cyt b) sequences among samples collected over an extensive coverage of the species' range. RESULTS: Cytochrome b sequences of 360 widely sampled individuals identified 137 unique maternal alleles. Gene tree and phylogeographic analyses of these alleles suggest the presence of at least eight lineages or haplogroups (A-H), with varying degrees of intra-lineage diversity. This differentiation is in contrast with the most recent taxonomic treatment based on cranial morphometrics which only recognised four subspecies. The mtDNA diversity strongly supports earlier views that this taxon may represent a species complex. We further show statistical support for the association of several of these lineages with particular vegetation biomes of southern Africa. The time to the most recent common ancestor (TMRCA) dates to the Pliocene (~5 Mya) whereas coalescent-based divergence time estimates between lineages vary between 813 Kya [0.22-1.36] and 4.06 Mya [1.21-4.47]. The major diversification within lineages occurred during the Pleistocene. The identification of several regions of sympatry of distinct lineages offers future opportunities for the elucidation of the underlying speciation processes in the suggested species complex. CONCLUSIONS: Similar to other African murine rodents, M. namaquensis radiated during the Pliocene and Pleistocene coinciding with major periods of aridification and the expansion of savanna habitats. The suggested species complex is represented by at least eight lineages of which the majority are confined to only one or a few neighbouring biomes/bioregions. Contrasting intra-lineage phylogeographic patterns suggest differences in adaptation and responses to Plio-Pleistocene climatic and vegetation changes. The role of ecological factors in driving speciation in the group needs further investigation.

A phylogenetic estimate for golden moles (Mammalia, Afrotheria, Chrysochloridae).

BACKGROUND: Golden moles (Chrysochloridae) are small, subterranean, afrotherian mammals from South Africa and neighboring regions. Of the 21 species now recognized, some (e.g., Chrysochloris asiatica, Amblysomus hottentotus) are relatively common, whereas others (e.g., species of Chrysospalax, Cryptochloris, Neamblysomus) are rare and endangered. Here, we use a combined analysis of partial sequences of the nuclear GHR gene and morphological characters to derive a phylogeny of species in the family Chrysochloridae. RESULTS: Although not all nodes of the combined analysis have high support values, the overall pattern of relationships obtained from different methods of phylogeny reconstruction allow us to make several recommendations regarding the current taxonomy of golden moles. We elevate Huetia to generic status to include the species leucorhinus and confirm the use of the Linnean binomial Carpitalpa arendsi, which belongs within Amblysominae along with Amblysomus and Neamblysomus. A second group, Chrysochlorinae, includes Chrysochloris, Cryptochloris, Huetia, Eremitalpa, Chrysospalax, and Calcochloris. Bayesian methods make chrysochlorines paraphyletic by placing the root within them, coinciding with root positions favored by a majority of randomly-generated outgroup taxa. Maximum Parsimony (MP) places the root either between chrysochlorines and amblysomines (with Chlorotalpa as sister taxon to amblysomines), or at Chlorotalpa, with the former two groups reconstructed as monophyletic in all optimal MP trees. CONCLUSIONS: The inclusion of additional genetic loci for this clade is important to confirm our taxonomic results and resolve the chrysochlorid root. Nevertheless, our optimal topologies support a division of chrysochlorids into amblysomines and chrysochlorines, with Chlorotalpa intermediate between the two. Furthermore, evolution of the chrysochlorid malleus exhibits homoplasy. The elongate malleus has evolved just once in the Cryptochloris-Chrysochloris group; other changes in shape have occurred at multiple nodes, regardless of how the root is resolved.

Cytochrome oxidase I sequences reveal possible cryptic diversity in the cosmopolitan symbiotic copepod Nesippus orientalis Heller, 1868 (Pandaridae: Siphonostomatoida) on elasmobranch hosts from the KwaZulu-Natal coast of South Africa.

Over the past decade, numerous molecular phylogenetic studies uncovered cryptic diversity within the Copepoda, yet very few investigations focused on symbiotic copepods. Here we report mitochondrial DNA cytochrome oxidase I diversity in the cosmopolitan elasmobranch symbiont Nesippus orientalis off the KwaZulu-Natal coast of South Africa. Analysis of partial COI sequences of copepods sampled from a diversity of shark hosts, revealed the presence of two divergent clades. Diversity within the clades does not appear to be structured based on host species, host individual, geographic locality or time of sampling. However, divergence between the two clades seems to be related to host species. Phylogenetic analyses of representatives from the two clades, along with Nesippus spp., Caligus spp. and Lepeophtheirus spp. outgroups, further supports the distinction between the two clades. Future molecular phylogenetic investigations of widespread copepod symbionts most likely will reveal far greater levels of biodiversity than currently recognized.

Ophiostoma tsotsi sp. nov., a wound-infesting fungus of hardwood trees in Africa.

Polymorphic sequence-characterised marker assays from a recent diversity study on the Ascomycete fungus Ophiostoma quercus reported that some isolates from Africa were genetically distinct from O. quercus. In the present study, these African isolates were compared with authentic O. quercus isolates by evaluating morphological characters, growth in culture, mating compatibility and DNA sequence data. The isolates from Africa were morphologically similar to O. quercus, presenting Pesotum and Sporothrix synanamorphs in culture. Phylogenetic analyses of the ribosomal internal transcribed spacer regions 1 and 2, beta-tubulin and translation elongation factor 1-alpha gene regions confirmed that the African group represents a distinct species within the hardwood lineage of the O. piceae complex, closely related to O. ulmi and O. himal-ulmi. Mating studies between O. quercus and the African isolates showed that isolates mated predominantly with those of their own group, although there were rare cases of fertile crosses between the groups. Isolates residing in the African lineage are described here as a new species, O. tsotsi sp. nov.

Genetic diversity, relatedness and inbreeding of ranched and fragmented Cape buffalo populations in southern Africa.

Wildlife ranching, although not considered a conventional conservation system, provides a sustainable model for wildlife utilization and could be a source of valuable genetic material. However, increased fragmentation and intensive management may threaten the evolutionary potential and conservation value of species. Disease-free Cape buffalo (Syncerus caffer caffer) in southern Africa exist in populations with a variety of histories and management practices. We compared the genetic diversity of buffalo in national parks to private ranches and found that, except for Addo Elephant National Park, genetic diversity was high and statistically equivalent. We found that relatedness and inbreeding levels were not substantially different between ranched populations and those in national parks, indicating that breeding practices likely did not yet influence genetic diversity of buffalo on private ranches in this study. High genetic differentiation between South African protected areas highlighted their fragmented nature. Structure analysis revealed private ranches comprised three gene pools, with origins from Addo Elephant National Park, Kruger National Park and a third, unsampled gene pool. Based on these results, we recommend the Addo population be supplemented with disease-free Graspan and Mokala buffalo (of Kruger origin). We highlight the need for more research to characterize the genetic diversity and composition of ranched wildlife species, in conjunction with wildlife ranchers and conservation authorities, in order to evaluate the implications for management and conservation of these species across different systems.

Phylogeny and biogeography of the genus Pseudobarbus (Cyprinidae): shedding light on the drainage history of rivers associated with the Cape Floristic Region.

Relationships among the historically isolated lineages of Pseudobarbus were reconstructed using molecular and morphological data. Contradictions between the molecular and morphological phylogenies suggest convergent evolution and homoplasy in some morphological characters. The earliest divergence in Pseudobarbus was between P. quathlambae in Lesotho and the rest of the genus associated with the Cape Foristic Region in South Africa. A close relationship between P. phlegethon from the Olifants River system on the west coast of South Africa and a lineage of P. afer from small river systems in Afrotemperate Forests on the south coast, can only be explained through previous occurrence and subsequent extinction of ancestral populations in the Gourits River system. Several river systems had confluences before reaching lower sea levels, most notably during the last glacial maximum about 18,000 years ago, explaining closely related populations across different river systems. Mainly river capture explains shared lineages across river systems that did not share a common confluence during lower sea levels.

Finding stories in noise: Mitochondrial portraits from RAD data.

Mitochondrial DNA (mtDNA) has formed the backbone of phylogeographic research for many years; however, recent trends focus on genome-wide analyses. One method proposed for calibrating inferences from noisy next-generation data, such as RAD sequencing, is to compare these results with analyses of mitochondrial sequences. Most researchers using this approach appear to be unaware that many single nucleotide polymorphisms (SNPs) identified from genome-wide sequence data are themselves mitochondrial, or assume that these are too few to bias analyses. Here, we demonstrate two methods for mining mitochondrial markers using RAD sequence data from three South African species of yellowfish, Labeobarbus. First, we use a rigorous SNP discovery pipeline using the program stacks, to identify variant sites in mtDNA, which we then combine into haplotypes. Second, we directly map sequence reads against a mitochondrial genome reference. This method allowed us to reconstruct up to 98% of the Labeobarbus mitogenome. We validated these mitogenome reconstructions through blast database searches and by comparison with cytochrome b gene sequences obtained through Sanger sequencing. Finally, we investigate the organismal consequences of these data including ancient genetic exchange and a recent translocation among populations of L. natalensis, as well as interspecific hybridization between L. aeneus and L. kimberleyensis.

Exploring the phylogeography of a hexaploid freshwater fish by RAD sequencing.

The KwaZulu-Natal yellowfish (Labeobarbus natalensis) is an abundant cyprinid, endemic to KwaZulu-Natal Province, South Africa. In this study, we developed a single-nucleotide polymorphism (SNP) dataset from double-digest restriction site-associated DNA (ddRAD) sequencing of samples across the distribution. We addressed several hidden challenges, primarily focusing on proper filtering of RAD data and selecting optimal parameters for data processing in polyploid lineages. We used the resulting high-quality SNP dataset to investigate the population genetic structure of L. natalensis. A small number of mitochondrial markers present in these data had disproportionate influence on the recovered genetic structure. The presence of singleton SNPs also confounded genetic structure. We found a well-supported division into northern and southern lineages, with further subdivision into five populations, one of which reflects north-south admixture. Approximate Bayesian Computation scenario testing supported a scenario where an ancestral population diverged into northern and southern lineages, which then diverged to yield the current five populations. All river systems showed similar levels of genetic diversity, which appears unrelated to drainage system size. Nucleotide diversity was highest in the smallest river system, the Mbokodweni, which, together with adjacent small coastal systems, should be considered as a key catchment for conservation.

Friends and family: A software program for identification of unrelated individuals from molecular marker data.

The identification of related and unrelated individuals from molecular marker data is often difficult, particularly when no pedigree information is available and the data set is large. High levels of relatedness or inbreeding can influence genotype frequencies and thus genetic marker evaluation, as well as the accurate inference of hidden genetic structure. Identification of related and unrelated individuals is also important in breeding programmes, to inform decisions about breeding pairs and translocations. We present Friends and Family, a Windows executable program with a graphical user interface that identifies unrelated individuals from a pairwise relatedness matrix or table generated in programs such as coancestry and genalex. Friends and Family outputs a list of samples that are all unrelated to each other, based on a user-defined relatedness cut-off value. This unrelated data set can be used in downstream analyses, such as marker evaluation or inference of genetic structure. The results can be compared to that of the full data set to determine the effect related individuals have on the analyses. We demonstrate one of the applications of the program: how the removal of related individuals altered the Hardy-Weinberg equilibrium test outcome for microsatellite markers in an empirical data set. Friends and Family can be obtained from https://github.com/DeondeJager/Friends-and-Family.