Recovering species are often managed in small numbers, requiring management strategies that maintain genetic variation for long-term viability. Here, we evaluate the genetic outcomes of two restored American bison (Bison bison) populations 15 generations after their reintroduction as Colorado’s first wildlife reintroduction. After initial reintroduction in 1914 to Genesee park, the herd was split into two separate populations in 1938. To determine the genetic viability of the restored populations, we genotyped 36 individuals from both herds, analyzing 52 microsatellite markers to asses heterozygosity, allelic richness, inbreeding, and population structure. Both herds exhibit relatively high observed heterozygosity (Genesee: 0.775 sd = 0.183; Daniels: 0.781 sd = 0.178), high allelic richness (Genesee: 5.17 sd = 1.45; Daniels: 4.96 sd = 1.46), and negative FIS values (Genesee: -0.112 bootstraps = -0.158, -0.065; Daniels: -0.15, bootstraps = -0.191, -0.108), indicating a lack of inbreeding. Despite ongoing gene flow, the herds remain genetically distinct, as supported by pairwise FST (0.0354, bootstraps = 0.024, 0.046), Nei’s D (0.136), and AMOVA results (FST = 0.078, p = 0.001). STRUCTURE analysis further confirmed that the herds maintain genetic clustering despite some admixture. These results suggest that Denver Mountain Parks’ long-term management strategies – promoting controlled gene flow while preventing inbreeding – have been effective in maintaining genetic variation. Intentional individual movement between herds and introductions from external metapopulations have contributed to the long-term viability of these herds. This study highlights the success of small, intensively managed bison populations in maintaining genetic health over many generations and underscores the importance of gene flow strategies in wildlife restoration.

Denver Mountain Parks staff collected tail hairs from all available offspring (n = 33) born in 2021, along with a subset of reproductively active cows (n = 3), for a total of 36 individuals. This included 18 yearlings and one adult cow from Daniels Park as well as 15 yearlings and two adult cows from Genesee Mountain Park (n = 36). Our dataset encompassed 47% of all individuals in both herds at the time of sampling, including an entire cohort of offspring, which incorporates a substantial proportion of the total genetic variation within these herds. Tail hairs were stored in cool, dry conditions before being sent to the University of California Davis Veterinary Genetics Lab (Davis, CA) for DNA extraction and sequencing.

A unique Veterinary Genetics Lab case number was assigned to each sample, and DNA extractions were completed using a lysis buffer that dissolved cell membranes and released nuclear and mitochondrial DNA. Aliquots with two-to-three microliters of DNA solution were used for PCR. PCR was implemented using multiplexed panels (five total) with 52 previously-developed microsatellite markers to identify genetic diversity in bison using primer mixes where one of a pair is fluorescence-labeled. To separate fluorescence-labeled PCR product for each marker panel, an aliquot of each sample was subjected to electrophoresis. Laser detection was used to collect fluorescence signals from PCR products, and electropherograms were developed for each sample based on this data. Both the PCR and electrophoresis were performed with a reference sample as a positive control.

The STRand software was implemented for genotype analysis by the UCDavis Veterinary Genetics lab. This software processed the electropherogram output by the PCR gel electrophoreses and generated the genotypes.