Estimates of heterozygosity from single nucleotide polymorphism markers are context‐dependent and often wrong

Abstract

Genetic diversity is frequently described using heterozygosity, particularly in a conservation context. Often, it is estimated using single nucleotide polymorphisms (SNPs); however, it has been shown that heterozygosity values calculated from SNPs can be biased by both study design and filtering parameters. Though solutions have been proposed to address these issues, our own work has found them to be inadequate in some circumstances. Here, we aimed to improve the reliability and comparability of heterozygosity estimates, specifically by investigating how sample size and missing data thresholds influenced the calculation of autosomal heterozygosity (heterozygosity calculated from across the genome, i.e. fixed and variable sites). We also explored how the standard practice of tri‐ and tetra‐allelic site exclusion could bias heterozygosity estimates and influence eventual conclusions relating to genetic diversity. Across three distinct taxa (a frog, Litoria rubella; a tree, Eucalyptus microcarpa; and a grasshopper, Keyacris scurra), we found heterozygosity estimates to be meaningfully affected by sample size and missing data thresholds, partly due to the exclusion of tri‐ and tetra‐allelic sites. These biases were inconsistent both between species and populations, with more diverse populations tending to have their estimates more severely affected, thus having potential to dramatically alter interpretations of genetic diversity. We propose a modified framework for calculating heterozygosity that reduces bias and improves the utility of heterozygosity as a measure of genetic diversity, whilst also highlighting the need for existing population genetic pipelines to be adjusted such that tri‐ and tetra‐allelic sites be included in calculations.

Previously unmeasured genetic diversity explains part of Lewontin’s paradox in a k-mer-based meta-analysis of 112 plant species

“…4), suggesting that k-mer-based diversity may scale with population size better by capturing additional neutral genome size variation. Overall, these results suggests that diversity missed by SNPs explains part of Lewontin ’ s paradox in plants, consistent with literature suggesting that SNPs provide an incomplete picture of genome-wide polymorphism [Schmidt et al, 2021, VanWallendael and Alvarez, 2022, Jaegle et al, 2023, Sopniewski and Catullo, 2024].…”

“…Finally, previous meta-analyses of population size and diversity data rely on scraping diversity estimates from previously published studies (Frankham [2012], Buffalo [2021], except see Corbett-Detig et al [2015]). However, many studies report inaccurate SNP calls and diversity estimates due to errors in the handling of missing data [Korunes and Samuk, 2021, Schmidt et al, 2021, Sopniewski and Catullo, 2024 and may filter genotype calls differently, making comparisons across species difficult. Re-analyzing whole genome sequencing data with a common pipeline would make diversity estimates across species more comparable [Buffalo, 2021, Mirchandani et al, 2024.…”

Range size variably predicts genetic diversity in Gehyra geckos

“…Unexpected patterns in genetic diversity may be the result of recently recognised biases in SNP-only measures of heterozygosity, that do not account for invariant (Schmidt et al, 2021) or multi-allelic (Sopniewski & Catullo, 2024) sites, which may have affected results in a previous study of genetic diversity in northern Australian lizards (Fenker et al, 2021). To ensure methodological factors did not impact our conclusions, we leveraged two high-quality Gehyra reference genomes for bioinformatic processing, that allowed us to estimate individual heterozygosity using both variant and invariant sites.…”