Small populations of endangered species risk losing already eroded genetic diversity, important for adaptive potential, through the effects of genetic drift. The magnitude of drift can be mitigated by maximising the effective population size, as is the goal of genetic management strategies. Different mating systems, specifically those leading to reproductive skew, exacerbate genetic drift by distorting contributions. In the absence of an active management strategy, reproductive skew will have long-term effects on the genetic composition of a population, particularly where admixture is present. Here we examine the contrasting effects of conservation management strategies in two ex situ populations of the Critically Endangered eastern black rhino (Diceros bicornis michaeli), one managed as a semi-wild population in South Africa (SAx), and one managed under a mean-kinship breeding strategy in European zoos. We use molecular data to reconstruct pedigrees for both populations and validate the method using the zoo studbook. Using the reconstructed pedigree and studbook we show there is male sex-specific skew in both populations. However, the zoo’s mean-kinship breeding strategy effectively reduces reproductive skew in comparison to a semi-wild population with little genetic management. We also show that strong male reproductive skew in SAx has resulted in extensive admixture, which may require a re-evaluation of the population’s original intended role in the black rhino meta-population. With a high potential for admixture in many ex situ populations of endangered species, molecular and pedigree data remain vital tools for populations needing to balance drift and selection.

Samples were collected from zoo animals, between 2017 and 2021, using non-invasive nasal swabs (N=75), or whole blood (N=10) where this could be obtained opportunistically. One muscle tissue sample from a deceased animal, preserved in 2ml absolute ethanol at -20°C, was also included. Nasal swabs were preserved using silica gel at room temperature and blood samples collected in EDTA tubes were frozen at -20°C. From the SAx population we extracted DNA from pinna offcuts preserved in sodium chloride. These were obtained during routine ear-notching operations performed to facilitate individual animal identification on the reserve. Samples used in this study dated up to 2018, but samples from historic assignments and some adults were not available. We sampled approximately 80% of the SAx and 85% of the Zoo population.

DNA was extracted using a QIAamp DNA mini kit (QIAGEN GMBH, Hilden, Germany). Mitochondrial DNA was amplified using D-loop primers mt15996L and mt16502H following Moodley et al., (2017). Individuals were Sanger sequenced in the forward and reverse direction (Eurofins Genomics Europe) and consensus sequences generated using Geneious software (Geneious Prime 2023.0.1, https://www.geneious.com). These sequences were aligned with the corresponding homologous sequenced region from the Moodley et al., (2017) metapopulation database and a haplotype network was constructed using PopArt (Version 1.7, Leigh & Bryant, 2015).

We genotyped each sample with 15 polymorphic autosomal microsatellite loci isolated from the black rhino (Diceros bicornis) and white rhino (Ceratotherium simum). This included 10 of the 11 loci applied in Moodley et al., (2017). We included additional markers DB49, DB66, and BlRh1B (Brown & Houlden, 1999), and BlRh1C and DB52 (Harper et al., 2013). We followed the protocol outlined in Moodley et al., (2017). The additional primer sets were amplified as multiplexes using annealing temperatures of 57°C (DB49, DB66, BlRh1B) and 52°C (BlRh1C and DB52). Samples were analysed on a 48-well capillary ABI3730 genetic analyser (Applied Biosystems) with Genescan ROX500 size standards (Applied Biosystems). Allele sizes were determined using using the Geneious software microsatellite plugin (Geneious Prime 2023.0.1, https://www.geneious.com).