Genome-wide runs of homozygosity revealed sources of
inconsistencies in the AxiomTM Equine Genotyping
array
Annik Imogen Gmel1,2 and Markus
Neuditschko1
1Animal GenoPhenomics, Agroscope, Posieux,
Switzerland, 2Equine Department, Vetsuisse Faculty,
University of Zurich, Switzerland
Corresponding author:
markus.neuditschko@agroscope.admin.ch
Runs of homozygosity (ROH), haplotypes identical-by-descent (IBD), are
key tools for deriving genomic inbreeding1. The
genomic inbreeding coefficient (FROH) for an animal is
derived by dividing the sum of all homozygous segments
(SROH) by the total length of the genome. In livestock,
previously reported studies have shown high concordance rates between
FROH and pedigree derived inbreeding
(FPED). In the framework of a global genetic diversity
study of modern horse breeds, we recently observed notably low
FROH values compared to the corresponding
FPED. In this study, a total of 4’520 horses from 21
different breeds were genotyped on the 670K AxiomTMEquine Genotyping array (602,131 autosomal SNPs mapped to
EquCab3.0)2, 3. Quality of genotyping was considered
acceptable with a dish QC (DQC) ≥ 0.82 and QC call rate (CR) ≥ 97
according to Affymetrix Axiom best practices4. Runs of
homozygosity segments (NROH) for each horse were
determined with an overlapping window approach in PLINK
v1.95 with the following parameters: a minimum SNP
density of one SNP per 50 kb, a maximum gap length of 100 kb, a minimum
length of homozygous segments of 500 kb (including more than 80
homozygous SNPs), and allowing for one heterozygous SNP per
segment6. In total there were 18 horses with
NROH equal to 0 and another 62 horses with
NROH less than 30. Within the Franches-Montagnes (FM)
breed it was particularly notable, that five purebred horses had fewer
ROH segments (NROH < 88) compared to F1
outcrosses. Of particular concern were one horse with
NROH equal to 0 and two others with NROHless than 60. We re-genotyped these horses, along with two additional
control horses that had NROH between 123 and 180, using
the same DNA sample and genotype platform. Based on this data, we
computed genotype concordance rates between the two SNP batches. We
found that FM horses with none or few NROH exhibited low
genotype concordances rates (Figure 1a; green dots). The same result was
observed for eight re-genotyped Lusitano (LUS) horses, with
NROH ranging from 1 to 99 (Figure 1a; brown dots). A
comparison of discordant SNPs between the three FM outlier horses
suggested that individual genotype errors occurred randomly, as there
was only an 8% overlap in erroneous markers among the horses (Figure
1b). Notably, we observed over 50,000 discordant SNPs for the FM horse
showing none NROH. Based on the updated genotype
information, all horses (FM and LUS) exhibited significantly higher
NROH, ranging from 148 to 175. Our findings demonstrate
that NROH is a reliable indicator for assessing the
genotype quality of individual horses. Consequently, we recommend
excluding horses exhibiting extremely low NROH (e.g.,
NROH < 30) from downstream analyses. However,
further research is necessary to enhance the reproducibility of
AxiomTM Equine Genotyping array.
Figure 1: Analysis of runs of homozygosity segments
(NROH), genotype concordance rate, and discordant SNPs
in re-genotyped horses. (a) Correlation between NROHand genotype concordance between two independent genotyping efforts
using the same DNA extract and SNP platform: This panel illustrates the
association between NROH and genotype concordance in
Franches-Montagnes (FM) and Lusitano horses. (b) Overlap of discordant
SNPs: The Venn diagram shows the common and unique discordant SNPs among
the three re-genotyped FM horses.