DNA barcoding of southern African mammal species and construction of a reference library for forensic application.

Combating wildlife crimes in South Africa requires accurate identification of traded species and their products. Diagnostic morphological characteristics needed to identify species are often lost when specimens are processed and customs officials lack the expertise to identify species. As a potential solution, DNA barcoding can be used to identify morphologically indistinguishable specimens in forensic cases. However, barcoding is hindered by the reliance on comprehensive, validated DNA barcode reference databases, which are currently limited. To overcome this limitation, we constructed a barcode library of Cytochrome c oxidase subunit 1 (COI) and Cytochrome b (Cyt b) sequences for threatened and protected mammals exploited in southern Africa. Additionally, we included closely related or morphologically similar species and assessed the database's ability to identify species accurately. Published southern African sequences were incorporated to estimate intraspecific and interspecific variation. Neighbor-joining trees successfully discriminated 94-95% of the taxa. However, some widespread species exhibited high intraspecific distances (>2%), suggesting geographic sub-structuring or cryptic speciation. Lack of reliable published data prevented the unambiguous discrimination of certain species. This study highlights the efficacy of DNA barcoding in species identification, particularly for forensic applications. It also highlights the need for a taxonomic re-evaluation of certain widespread species and challenging genera.

PAReTT: A Python Package for the Automated Retrieval and Management of Divergence Time Data from the TimeTree Resource for Downstream Analyses.

Evolutionary processes happen gradually over time and are, thus, considered time dependent. In addition, several evolutionary processes are either adaptations to local habitats or changing habitats, otherwise restricted thereby. Since evolutionary processes driving speciation take place within the landscape of environmental and temporal bounds, several published studies have aimed at providing accurate, fossil-calibrated, estimates of the divergence times of both extant and extinct species. Correct calibration is critical towards attributing evolutionary adaptations and speciation both to the time and paleogeography that contributed to it. Data from more than 4000 studies and nearly 1,50,000 species are available from a central TimeTree resource and provide opportunities of retrieving divergence times, evolutionary timelines, and time trees in various formats for most vertebrates. These data greatly enhance the ability of researchers to investigate evolution. However, there is limited functionality when studying lists of species that require batch retrieval. To overcome this, a PYTHON package termed Python-Automated Retrieval of TimeTree data (PAReTT) was created to facilitate a biologist-friendly interaction with the TimeTree resource. Here, we illustrate the use of the package through three examples that includes the use of timeline data, time tree data, and divergence time data. Furthermore, PAReTT was previously used in a meta-analysis of candidate genes to illustrate the relationship between divergence times and candidate genes of migration. The PAReTT package is available for download from GitHub or as a pre-compiled Windows executable, with extensive documentation on the package available on GitHub wiki pages regarding dependencies, installation, and implementation of the various functions.

Genomic analyses show extremely perilous conservation status of African and Asiatic cheetahs (Acinonyx jubatus).

We live in a world characterized by biodiversity loss and global environmental change. The extinction of large carnivores can have ramifying effects on ecosystems like an uncontrolled increase in wild herbivores, which in turn can have knock-on impacts on vegetation regeneration and communities. Cheetahs (Acinonyx jubatus) serve important ecosystem functions as apex predators; yet, they are quickly heading towards an uncertain future. Threatened by habitat loss, human-wildlife conflict and illegal trafficking, there are only approximately 7100 individuals remaining in nature. We present the most comprehensive genome-wide analysis of cheetah phylogeography and conservation genomics to date, assembling samples from nearly the entire current and past species' range. We show that their phylogeography is more complex than previously thought, and that East African cheetahs (A. j. raineyi) are genetically distinct from Southern African individuals (A. j. jubatus), warranting their recognition as a distinct subspecies. We found strong genetic differentiation between all classically recognized subspecies, thus refuting earlier findings that cheetahs show only little differentiation. The strongest differentiation was observed between the Asiatic and all the African subspecies. We detected high inbreeding in the Critically Endangered Iranian (A. j. venaticus) and North-western (A. j. hecki) subspecies, and show that overall cheetahs, along with snow leopards, have the lowest genome-wide heterozygosity of all the big cats. This further emphasizes the cheetah's perilous conservation status. Our results provide novel and important information on cheetah phylogeography that can support evidence-based conservation policy decisions to help protect this species. This is especially relevant in light of ongoing and proposed translocations across subspecies boundaries, and the increasing threats of illegal trafficking.

Genome Scan for Variable Genes Involved in Environmental Adaptations of Nubian Ibex.

The Nubian ibex (Capra nubiana) is a wild goat species that inhabits the Sahara and Arabian deserts and is adapted to extreme ambient temperatures, intense solar radiation, and scarcity of food and water resources. To investigate desert adaptation, we explored the possible role of copy number variations (CNVs) in the evolution of Capra species with a specific focus on the environment of Capra nubiana. CNVs are structural genomic variations that have been implicated in phenotypic differences between species and could play a role in species adaptation. CNVs were inferred from Capra nubiana sequence data relative to the domestic goat reference genome using read-depth approach. We identified 191 CNVs overlapping with protein-coding genes mainly involved in biological processes such as innate immune response, xenobiotic metabolisms, and energy metabolisms. We found copy number variable genes involved in defense response to viral infections (Cluster of Differentiation 48, UL16 binding protein 3, Natural Killer Group 2D ligand 1-like, and Interferon-induced transmembrane protein 3), possibly suggesting their roles in Nubian ibex adaptations to viral infections. Additionally, we found copy number variable xenobiotic metabolism genes (carboxylesterase 1, Cytochrome P450 2D6, Glutathione S-transferase Mu 4, and UDP Glucuronosyltransferase-2B7), which are probably an adaptation of Nubian ibex to desert diets that are rich in plant secondary metabolites. Collectively, this study's results advance our understanding of CNVs and their possible roles in the adaptation of Nubian ibex to its environment. The copy number variable genes identified in Nubian ibex could be considered as subjects for further functional characterizations.

Unlocking the potential of a validated single nucleotide polymorphism array for genomic monitoring of trade in cheetahs (Acinonyx jubatus).

Cheetahs (Acinonyx jubatus) are listed as vulnerable on the International Union for Conservation of Nature Red List of Threatened Species. Threats include loss of habitat, human-wildlife conflict and illegal wildlife trade. In South Africa, the export of wild cheetah is a restricted activity under the Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES), however, limited legal trade is permitted of animals born to captive parents. To effectively monitor the legal and illegal trade in South Africa, it was thus essential to develop a validated molecular test. Here, we designed a single nucleotide polymorphism (SNP) array for cheetah from Double Digest Restriction Associated DNA sequencing data for individual identification and parentage testing. In order to validate the array, unrelated individuals and 16 family groups consisting of both parents and one to three offspring were genotyped using the Applied Biosystems™ QuantStudio™ 12K Flex Real-Time PCR System. In addition, parentage assignments were compared to microsatellite data. Cross-species amplification was tested in various felids and cheetah sub-species in order to determine the utility of the SNP array in other species. We obtained successful genotyping results for 218 SNPs in cheetah (A. j. jubatus) with an optimal DNA input concentration ranging from 10 to 30 ng/µl. The combination of SNPs had a higher resolving power for individual identification compared to microsatellites and provided high assignment accuracy in known pedigrees. Cross-species amplification in other felids was determined to be limited. However, the SNP array demonstrated a clear genetic discrimination of two cheetah subspecies tested here. We conclude that the described SNP array is suitable for accurate parentage assignment and provides an important traceability tool for forensic investigations of cheetah trade.

Sperm motility, kinematics, morphometry and morphology over two seasons in free-ranging African elephants (Loxodonta africana).

This study aimed to address the lack of information on quantitative semen and sperm characteristics of free-ranging African elephants. Nineteen ejaculates were collected from 12 elephant bulls by means of electroejaculation in spring (Season 1, end of dry season, n=7) and in autumn (Season 2, end of rainy season, n=12). While most elephant cows are in oestrus in the rainy season, it is not evident whether sperm quality also improves during this period. Semen samples were assessed using computer-aided sperm analysis (CASA), brightfield microscopy and transmission electron microscopy. Seasonal differences and individual variation in sperm quality of bulls were apparent, with ejaculates collected during Season 2 revealing higher percentages for total motility, progressive motility, rapid-swimming spermatozoa and kinematic parameters compared with Season 1 (P<0.05). Although normal sperm morphology percentage was similar over the two seasons, more sperm tail defects were found in Season 2 (P<0.05). The baseline reference data and multivariate sperm parameter associations reported in this study can be used to predict elephant bull sperm quality and potential to fertilise. It is clear that CASA can detect subtle differences in sperm quality of African elephant ejaculates and should be the approach for future investigations.

Pangolin genomes and the evolution of mammalian scales and immunity.

Pangolins, unique mammals with scales over most of their body, no teeth, poor vision, and an acute olfactory system, comprise the only placental order (Pholidota) without a whole-genome map. To investigate pangolin biology and evolution, we developed genome assemblies of the Malayan (Manis javanica) and Chinese (M. pentadactyla) pangolins. Strikingly, we found that interferon epsilon (IFNE), exclusively expressed in epithelial cells and important in skin and mucosal immunity, is pseudogenized in all African and Asian pangolin species that we examined, perhaps impacting resistance to infection. We propose that scale development was an innovation that provided protection against injuries or stress and reduced pangolin vulnerability to infection. Further evidence of specialized adaptations was evident from positively selected genes involving immunity-related pathways, inflammation, energy storage and metabolism, muscular and nervous systems, and scale/hair development. Olfactory receptor gene families are significantly expanded in pangolins, reflecting their well-developed olfaction system. This study provides insights into mammalian adaptation and functional diversification, new research tools and questions, and perhaps a new natural IFNE-deficient animal model for studying mammalian immunity.

Contrasting evolutionary history, anthropogenic declines and genetic contact in the northern and southern white rhinoceros (Ceratotherium simum).

The white rhinoceros (Ceratotherium simum) has a discontinuous African distribution, which is limited by the extent of sub-Saharan grasslands. The southern population (SWR) declined to its lowest number around the turn of the nineteenth century, but recovered to become the world's most numerous rhinoceros. In contrast, the northern population (NWR) was common during much of the twentieth century, declining rapidly since the 1970s, and now only two post-reproductive individuals remain. Despite this species's conservation status, it lacks a genetic assessment of its demographic history. We therefore sampled 232 individuals from extant and museum sources and analysed ten microsatellite loci and the mtDNA control region. Both marker types reliably partitioned the species into SWR and NWR, with moderate nuclear genetic diversity and only three mtDNA haplotypes for the species, including historical samples. We detected ancient interglacial demographic declines in both populations. Both populations may also have been affected by recent declines associated with the colonial expansion for the SWR, and with the much earlier Bantu migrations for the NWR. Finally, we detected post-divergence secondary contact between NWR and SWR, possibly occurring as recently as the last glacial maximum. These results suggest the species was subjected to regular periods of fragmentation and low genetic diversity, which may have been replenished upon secondary contact during glacial periods. The species's current situation thus reflects prehistoric declines that were exacerbated by anthropogenic pressure associated with the rise of late Holocene technological advancement in Africa. Importantly, secondary contact suggests a potentially positive outcome for a hybrid rescue conservation strategy, although further genome-wide data are desirable to corroborate these results.

Mitochondrial genomes of African pangolins and insights into evolutionary patterns and phylogeny of the family Manidae.

BACKGROUND: This study used next generation sequencing to generate the mitogenomes of four African pangolin species; Temminck's ground pangolin (Smutsia temminckii), giant ground pangolin (S. gigantea), white-bellied pangolin (Phataginus tricuspis) and black-bellied pangolin (P. tetradactyla). RESULTS: The results indicate that the mitogenomes of the African pangolins are 16,558 bp for S. temminckii, 16,540 bp for S. gigantea, 16,649 bp for P. tetradactyla and 16,565 bp for P. tricuspis. Phylogenetic comparisons of the African pangolins indicated two lineages with high posterior probabilities providing evidence to support the classification of two genera; Smutsia and Phataginus. The total GC content between African pangolins was observed to be similar between species (36.5% - 37.3%). The most frequent codon was found to be A or C at the 3rd codon position. Significant variations in GC-content and codon usage were observed for several regions between African and Asian pangolin species which may be attributed to mutation pressure and/or natural selection. Lastly, a total of two insertions of 80 bp and 28 bp in size respectively was observed in the control region of the black-bellied pangolin which were absent in the other African pangolin species. CONCLUSIONS: The current study presents reference mitogenomes of all four African pangolin species and thus expands on the current set of reference genomes available for six of the eight extant pangolin species globally and represents the first phylogenetic analysis with six pangolin species using full mitochondrial genomes. Knowledge of full mitochondrial DNA genomes will assist in providing a better understanding on the evolution of pangolins which will be essential for conservation genetic studies.

Advances in canine distemper virus pathogenesis research: a wildlife perspective.

Canine distemper virus (CDV) has emerged as a significant disease of wildlife, which is highly contagious and readily transmitted between susceptible hosts. Initially described as an infectious disease of domestic dogs, it is now recognized as a global multi-host pathogen, infecting and causing mass mortalities in a wide range of carnivore species. The last decade has seen the effect of numerous CDV outbreaks in various wildlife populations. Prevention of CDV requires a clear understanding of the potential hosts in danger of infection as well as the dynamic pathways CDV uses to gain entry to its host cells and its ability to initiate viral shedding and disease transmission. We review recent research conducted on CDV infections in wildlife, including the latest findings on the causes of host specificity and cellular receptors involved in distemper pathogenesis.

Forensic application of DNA barcoding for identification of illegally traded African pangolin scales.

The escalating growth in illegal wildlife trade and anthropogenic habitat changes threaten the survival of pangolin species worldwide. All eight extant species have experienced drastic population size reductions globally with a high extinction risk in Asia. Consequently, forensic services have become critical for law enforcement, with a need for standardised and validated genetic methods for reliable identifications. The seizure of three tonnes of pangolin scales, believed to have originated from Africa, by Hong Kong Customs Authorities provided an opportunity for the application of DNA barcoding in identifying scales. Three mitochondrial DNA gene regions (COI, Cyt b, and D-loop) were amplified for a subsample of the confiscated material and compared with taxonomically verified references. All four African species were recovered as monophyletic with high interspecific uncorrected p-distance estimates (0.048-0.188) among genes. However, only three of four African species (Phataginus tricuspis, Phataginus tetradactyla, and Smutsia gigantea, originating from West and Central Africa) and one of four Asian species (Manis javanica from Southeast Asia) were identified among scales. Although the assignment of unknown scales to specific species was reliable, additional genetic tools and representative reference material are required to determine geographic origins of confiscated pangolin specimens.

SNP discovery and characterisation in White Rhino (Ceratotherium simum) with application to parentage assignment.

The white rhino is one of the great success stories of modern wildlife conservation, growing from as few as 50-100 animals in the 1880s, to approximately 20,000 white rhinoceros remaining today. However, illegal trade in conservational rhinoceros horns is adding constant pressure on remaining populations. Captive management of ex situ populations of endangered species using molecular methods can contribute to improving the management of the species. Here we compare for the first time the utility of 33 Single Nucleotide Polymorphisms (SNPs) and nine microsatellites (MS) in isolation and in combination for assigning parentage in captive White Rhinoceros. We found that a combined dataset of SNPs and microsatellites was most informative with the highest confidence level. This study thus provided us with a useful set of SNP and MS markers for parentage and relatedness testing. Further assessment of the utility of these markers over multiple (> three) generations and the incorporation of a larger variety of relationships among individuals (e.g. half-siblings or cousins) is strongly suggested.

Intrinsic factors, adrenal gland morphology, and disease burden in captive cheetahs (Acinonyx jubatus) in South Africa.

Adrenal gland weight (AW) and corticomedullary ratio (ACMR) are used as indicators of stress in animals. Captive cheetahs (Acinonyx jubatus) have higher ACMRs than free-ranging ones and stress has been linked to gastritis, amyloidosis, glomerulosclerosis, and myocardial fibrosis. We reviewed age, sex, body weight (BW), kidney weight (KW), and left AW and ACMR with necropsy findings in 51 South African captive cheetahs. Eleven common histopathologic lesions were counted for each animal as measure of its disease burden. Adrenal corticomedullary hyperplasia was significantly correlated with left AW and ACMR. Males had significantly higher AWs than females; other parameters showed no difference between the sexes. Disease burden, gastritis, and myocardial fibrosis were moderately correlated with adrenal morphology supporting prior evidence that gastritis and myocardial fibrosis are linked to stress. Glomerulosclerosis was not correlated with adrenal morphology and neither kidney nor liver amyloidosis contributed significantly to variation in AW or ACMR on multivariate analyses. Interstitial nephritis showed much stronger correlations with kidney and liver amyloidosis than gastritis. All three adrenal parameters were correlated with age; age was the only significant variable affecting ACMR on the multivariate analyses; and disease burden as well as systemic amyloidosis and kidney disease (except for fibrosis) showed moderate correlations with age. Age may, therefore, be important in the pathogenesis of disease in captive cheetahs, particularly of amyloidosis and kidney disease. None of the intrinsic measurements or adrenal parameters were sufficiently closely linked to disease to be used as ante-mortem proxies for disease burden or specific diseases. Zoo Biol. 36:40-49, 2017. © 2016 Wiley Periodicals, Inc.

Classical and Molecular Cytogenetics of the Panther Genet Genetta maculata (Mammalia, Carnivora, Viverridae).

Genets (Genetta) are a genus of African mammalian carnivorans with 14 currently recognized species, although taxonomic uncertainties remain, particularly regarding the number of species within the large-spotted genet complex. This study presents the first banded karyotype and molecular cytogenetic analysis of a genetically identified panther genet, Genetta maculata, the most common and widespread taxon of the large-spotted genet complex, with a wide distribution in sub-Saharan Africa. Sampled in Gauteng Province, South Africa, it could be assigned to the subspecies G. m. letabae on geographic grounds and had a similar karyotype (2n = 52, FNa = 96) to those published for a panther genet from Ethiopia and for the West African large-spotted genet G. pardina. Notably, the specimen had a different autosomal morphology (2 acrocentric chromosomes) from that previously attributed to letabae (a single acrocentric chromosome), but the latter assignment was uncertain because the studied individuals were captive born and assigned based solely on a presumed origin in the former Transvaal Province of South Africa. Fluorescence in situ hybridization with a telomere repeat probe revealed the presence of telomeric sequences in the centromeres of most chromosomes, the so-called interstitial telomeric sites (ITSs). Since genets seem to have a unique, highly rearranged karyotype among feliforms and relatively low interspecific karyotypic variation, and considering the known instability of ITSs, we suggest that the large amount of ITSs found here might be due to evolutionarily recent extensive genomic rearrangements. This study provides cytogenetic information that contributes to our understanding of chromosomal variation and genomic rearrangements in genets, and valuable baseline data for future studies of karyotype evolution in carnivores in general and viverrids in particular.

Mercury concentrations in eggshells of the Southern Ground-Hornbill (Bucorvus leadbeateri) and Wattled Crane (Bugeranus carunculatus) in South Africa.

In this study, wild hatched eggshells were collected from the nests of threatened Wattled Crane and South Ground-Hornbill in an attempt to determine their total Hg concentrations. A total of fourteen eggshell samples from both bird species were collected from different study areas in the Mpumlanga and KwaZulu-Natal Provinces of South Africa. The eggshells were acid digested under reflux and their total Hg concentrations were determined using cold-vapour atomic absorption spectrophotometry (CV-AAS). The observed total Hg levels for the South Ground-Hornbill samples ranged from 1.31 to 8.88 µg g(-1) dry weight (dw), except for one outlier which had an elevated 75.0 µg g(-1) dw. The levels obtained for the Wattled Crane samples were relatively high and these ranged from 14.84 to 36.37 µg g(-1) dw. Generally, all the measured total Hg concentrations for the Wattled Crane samples exceeded the estimated total Hg levels derived for eggshell which were known to cause adverse reproductive effects in avian species from previous studies. Based on these findings, it is, therefore, possible that the exposure of these birds to elevated Hg may have contributed to their present population decline.

Interspecific hybridization between greater kudu and nyala.

Hybridization of wildlife species, even in the absence of introgression, is of concern due to wasted reproductive effort and a reduction in productivity. In this study we detail an accidental mating between a female nyala (Tragelaphus angasii) and a male greater kudu (T. strepsiceros). The hybrid was phenotypically nyala and was identified as such based on mitochondrial DNA. Further genetic analysis based on nine microsatellite markers, chromosome number and chromosome morphology however, confirmed its status as an F1 hybrid. Results obtained from a reproductive potential assessment indicated that this animal does not have the potential to breed successfully and can be considered as sterile.

Diet and diet-related disorders in captive ruminants at the national zoological gardens of South Africa.

Although diet-related disorders have received much attention in the zoo literature, evidence-based results on relationships between diet and disease are still rare, often due to a lack of quantitative dietary information that can be linked to clinical or necropsy reports. We investigated 24 species of captive ruminants from one facility for which quantitative feeding instructions and necropsy reports between 1991 and 2012 were available. Species were classified as grazer (GR), intermediate feeder (IM), or browser (BR). Feeding type and body mass were significantly correlated to the diet fed, with smaller and BR species receiving higher proportions of non-roughage diet items. There were no significant differences between feeding types in the occurrence of parakeratosis/ruminitis/acidosis (PRA) at necropsy, but in body condition score, with BR more often in poor and less often in excellent body condition at necropsy. While there was no direct correlation between the proportion of non-roughage diet items and PRA across species, there was a significant effect of the proportion of non-roughage diet items on PRA when body mass was also taken into account: larger species, and those that received more non-roughage diet items, had higher prevalence of PRA. The results underline that diet and lack of structured feed items can be associated with the disease complex of acidosis in ruminants, but also suggest that this is modified by factors related to animal size. These latter may include susceptibility to acidosis, or husbandry-related opportunities to monopolize non-roughage feeds and ingest higher proportions than intended by feeding instructions.

Development and validation of a novel forensic STR multiplex assay for blue (Anthropoides paradiseus), wattled (Bugeranus carunculatus), and grey-crowned crane (Balearica regulorum).

The blue crane (Anthropoides paradiseus), wattled crane (Bugeranus carunculatus), and grey-crowned crane (Balearica regulorum) are species of concern as their populations are declining and they face several threats including habitat loss, disturbance and illegal trade. In South Africa, these species are bred in captivity for trade purposes which is permitted and regulated globally under the Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES). Legal sustainable trade through captive breeding of endangered wildlife species such as cranes has been promoted to counteract the illegal trade of individuals from the wild. Captive breeding independent of wild populations may reduce the harvest pressures on wild bird populations which in turn benefit the recovery of exploited species. This approach is considered to be controversial by some individuals. Although captive breeding of endangered species, for both population sustainability and commercial purposes, is promoted to aid in conserving species, concerns have been raised with regards to breeding facilities being used for laundering of animals. To monitor the legal trade of cranes in South Africa a short tandem repeat (STR) assay following recommendations of the International Society for Forensic Genetics (ISFG) was developed and validated. An STR assay comprising of four multiplexes that include 16 STR markers and two gender determination markers was proven to be highly informative with average polymorphic information content (PIC) values of 0.806, 0.646 and 0.725 for A. paradiseus, B. regulorum and B. carunculatus respectively. In addition, the assay showed sufficient discriminatory power for parentage assignment of closely related individuals in all three species (A. paradiseus: PI = 1.7×10-24, PIsibs = 4.7×10-08, and B. carunculatus: PI = 1.4×10-19, PIsibs = 2.9×10-07 and B. regulorum: PI = 1.7×10-12, PIsibs = 5.0×10-05). Analysis of 251 samples suggested that the validated multiplex assay ensures reliability, reproducibility, and repeatability for applications in forensic case work where illegal trade of offspring is suspected through verifying parentage of captive birds in breeding facilities.

Methylation-based markers for the estimation of age in African cheetah, Acinonyx jubatus.

Age is a key demographic in conservation where age classes show differences in important population metrics such as morbidity and mortality. Several traits, including reproductive potential, also show senescence with ageing. Thus, the ability to estimate age of individuals in a population is critical in understanding the current structure as well as their future fitness. Many methods exist to determine age in wildlife, with most using morphological features that show inherent variability with age. These methods require significant expertise and become less accurate in adult age classes, often the most critical groups to model. Molecular methods have been applied to measuring key population attributes, and more recently epigenetic attributes such as methylation have been explored as biomarkers for age. There are, however, several factors such as permits, sample sovereignty, and costs that may preclude the use of extant methods in a conservation context. This study explored the utility of measuring age-related changes in methylation in candidate genes using mass array technology. Novel methods are described for using gene orthologues to identify and assay regions for differential methylation. To illustrate the potential application, African cheetah was used as a case study. Correlation analyses identified six methylation sites with an age relationship, used to develop a model with sufficient predictive power for most conservation contexts. This model was more accurate than previous attempts using PCR and performed similarly to candidate gene studies in other mammal species. Mass array presents an accurate and cost-effective method for age estimation in wildlife of conservation concern.

Hybridization in an isolated population of blesbok and red hartebeest.

Hybridization in antelope species has been widely reported in South African national parks and provincial reserves as well as on private land due to anthropogenic effects. In a closed management setting, hybridization may occur due to the crossbreeding of closely related species with unequal sex ratios, resulting in either sterile or fertile offspring. In this study, we used molecular techniques to evaluate the risk of anthropogenic hybridization between blesbok (Damaliscus pygargus phillipsi) and red hartebeest (Alcelaphus buselaphus caama) in an isolated group that purposely included the two species with unequal sex ratios (one red hartebeest male and 19 male and female blesbok). Genetic analysis based on microsatellites confirmed the presence of seven hybrid individuals. Mitochondrial analysis verified that hybridization occurred between blesbok females and the red hartebeest male. STRUCTURE and NEWHYBRIDS classified the hybrids as F1. It is suspected that the hybrid individuals were sterile as the males had undeveloped testes and only F1 hybrids were detected. Thus, the risk of hybridization between these two species may be limited in the wild. In captive settings, genetic monitoring should be included in management plans for blesbok and red hartebeest to ensure that the long-term consequences of wasted reproductive effort are limited.