Aardvark microsatellite data for southern and east Africa

Aim: As climate change accelerates, assessing how climate shapes gene flow and neutral and adaptive genetic differentiation on landscapes is increasingly important. Aardvarks (Orycteropus afer) are ecologically important in sub-Saharan Africa but are sensitive to human pressures and increasing aridity. We used individual, population, and landscape genetic approaches to infer the influence of landscape, climate, and potential adaptive differences on gene flow. Location: We surveyed 8 protected and 4 privately owned areas in South Africa, 2 protected areas in Eswatini, and one location in Kenya during 2016–2018. Methods: We developed microsatellite markers and methods for DNA extraction from feces, collected and genotyped fecal samples from focal areas, and estimated genetic structure. We inferred space use from individual redetections, tested for close relatives, and estimated genetic neighborhood distance. We applied individual-based landscape genetic analyses at multiple scales across South Africa to test hypotheses about genetic differentiation by landscape resistance and potential adaptive differences. Results: We developed 19 variable microsatellite loci and collected 253 fecal samples from 13 focal areas. We genotyped 104 samples successfully at ≥8 loci as needed for individual identification. Genetic structure suggested 3 regional divisions in South Africa. We detected individuals at locations ≤7.3 km distant and closely related individuals at ≤44 km; genetic neighborhood distance was <55 km. Lower precipitation increased landscape resistance and strongly predicted genetic differentiation at most spatial scales. Temperature differences at sampling sites also influenced structure, suggesting climate-associated adaptive differences. Main Conclusions: Genetic structure of aardvarks in South Africa and Eswatini is strongly shaped by climate, with arid areas limiting gene flow, and reflects apparent isolation by adaptation associated with temperature. Dispersal distances likely are <45 km. The markers we developed will facilitate studies of space use, dispersal, population density, or survival. Aridification will increase fragmentation and we recommend monitoring aardvark presence as an indicator of ecosystem change associated with aridification. This dataset was generated by designing new microsatellite primers, collecting fecal samples from aardvarks in southern Africa and one location in East Africa, extracting aardvark DNA from those fecal samples, and genotyping those DNA extracts at up to 19 loci. Duplicate individuals were removed and samples successfully genotyped at fewer than 8 loci were not included. Laboratory details are provided here: We scraped the exterior surface of the dried fecal samples using a razor blade to obtain 0.03–0.045g of feces, attempting to avoid sand and dietary matter (i.e., ants or termites). We extracted DNA from fecal material using a version of the Aquagenomic and Aquaprecipi fecal extraction protocol (Multitarget Pharmaceuticals, Colorado Springs, CO) modified as follows. We increased the amount of Aquagenomic solution to 450 µL per sample, performed a 15-minute bead-beating step with 1.0 mm silica/zirconium beads (BioSpec Products Inc., Bartlesville, OK) to facilitate cell lysis, and added 12 mAU proteinase K (Qiagen Inc., Valencia, CA). Lastly, we added 150 µL of Aquaprecipi solution to cell lysate to counteract PCR inhibitors present in fecal samples and rehydrated the final DNA pellet in 100 µL 1x TE buffer.  We attempted to amplify 19 microsatellite loci in four multiplex PCR panels, after screening samples for DNA quality by initially running all samples in three separate PCR reactions for Panel A (Table 1). We conducted PCRs in 10 μL reactions consisting of 5x Qiagen Multiplex PCR Master Mix, 10 μg of bovine serum albumin, 0.2 μM of each panel-specific primer and 1 μL of genomic DNA. Reactions were brought to volume with nuclease-free water. For each locus, one primer was fluorescently tagged on the 5’ end with NED, PET, VIC (Applied Biosystems, Carlsbad, CA) or 6-FAM (Sigma-Aldrich, St. Louis, MO). We included negative and positive controls in each PCR to monitor for reagent contamination and align calls across runs. Thermalcycling conditions for the multiplexed loci were as follows: initial denaturation of 15 minutes at 95 °C, followed by 35 cycles of [95 °C for 30 seconds, 60 °C for 90 seconds, 72 °C for 60 seconds], and a final elongation of 30 minutes at 60 °C.  We ran PCRs on BioRad C1000, T100, and MyCycler thermalcycler machines (Bio-Rad Laboratories Inc., Hercules, CA). We verified amplifications by visualizing 4 µL of PCR product from one replicate of each sample on a 2% agarose gel prestained with GelRed (Biotium, Fremont, CA); products were then diluted accordingly, ethanol-precipitated to remove salts, and submitted for genotyping on the ABI 3730 DNA analyzer (Applied Biosystems) at the Oregon State University Center for Quantitative Life Sciences (Corvallis, OR). We used GeneScan500 LIZ dye size standard and called allele sizes in GeneMapper v.4.1 (Applied Biosystems). Samples that produced data at fewer than 50% of the loci in the first three replicates for Panel A were not analyzed further. Samples that produced partial genotypes at ≥ 50% of microsatellite loci were rerun 3–6 more times depending on the completeness of initial replicates, while samples that produced complete and consistent genotypes in the first three replicates were considered finalized and a consensus genotype was accepted. For a genotype to be accepted for a particular locus, each allele in a heterozygote genotype had to be observed twice, while the single allele in a homozygote genotype had to be observed three times. We ran Panels B-D for samples that had generated sufficient data at Panel A. Any sample that consistently showed more than two alleles at a single locus was considered contaminated and removed. R, or Microsoft Excel or similar spreadsheet programs that can read a text (.txt) file. # Aardvark microsatellite data for southern and east Africa [doi.org](doi.org) Data for 19 microsatellite loci for aardvarks (Orycteropus afer), genotyped from feces. Missing data are scored as ?, otherwise, fragment sizes are reported. Each locus is reported in two columns, with those columns named according to the convention "LocusName\_1" for the first allele and "LocusName\_2" for the second allele. Sampling location (descriptive, as well as GPS coordinates in WGS84, Decimal Degrees) is included for each sample. Samples included were genotyped at a minimum of 8 loci according to the genotyping criteria; duplicate samples determined to be derived from the same individuals have been removed.