Mutation rate and spectrum of germline de novo mutations in a closed population of Thoroughbred horses.

Overview

• This study analyzed new genetic mutations occurring for the first time (de novo mutations) in a closed breed of Thoroughbred horses.
• Using whole-genome sequencing of a horse family (sire, dam, foal), the researchers measured how often these mutations arise and characterized their origin and mutation types.

Background

• Thoroughbred horses have been bred as a closed population for over 300 years, originating from crosses between Arabian stallions and English mares.
• Understanding germline de novo mutations, which are changes in DNA that appear newly in offspring, is important as they provide a source of genetic variation and may impact traits or diseases.
• While de novo mutation rates and patterns have been studied in humans and some other species, this information was lacking for Thoroughbred horses.

Study Objectives

• Identify de novo mutations in Thoroughbred horses at the whole-genome level.
• Estimate the frequency (mutation rate) of these mutations within a genetically closed Thoroughbred population.
• Determine the parental origin of the mutations (paternal vs maternal).

Methods

• The researchers performed deep whole-genome sequencing on a trio consisting of a Thoroughbred sire (father), dam (mother), and foal (offspring).
• Sequencing depth was very high (≥230×) with 150 base pair paired-end reads, increasing confidence in mutation detection.
• Sequencing reads were aligned to the EquCab3.0 horse reference genome.
• Analysis focused on well-covered regions (≥20× depth), which included 99.6% of the genome.
• Mendelian inconsistency analysis was used to detect candidate de novo mutations, defined as variants present in the foal but absent in both parents.
• Sanger sequencing was used to validate candidate mutations, confirming their presence in the foal but absence in parents.
• Parental origin of mutations was assigned through statistical and sequence context analyses.

Results

• A total of 48 de novo mutations were discovered:
• 46 were single nucleotide substitutions (point mutations)
• 2 were deletions
• None of these mutations were found in existing public variant databases, indicating they are novel.
• Among the mutations with assigned origin, 18 were paternal (from the father) versus 6 maternal (from the mother), a statistically significant bias (P < 0.05) toward paternal origin.
• The estimated germline mutation rate was approximately 9.59 × 10⁻⁹ mutations per base per generation.
• This rate is comparable to mutation rates reported in humans and other mammals.
• Notably, one mutation caused a nonsense variant (introducing a premature stop codon) in the zinc finger protein 77 gene on chromosome 7, which is believed to be a transcriptional regulator.

Conclusions

• This study successfully identified de novo mutations in Thoroughbred horses despite the breed’s genetic closedness.
• The measured mutation rate aligns with rates found in other mammalian species, reinforcing evolutionary conservation of mutation frequency.
• A significant paternal bias in mutation origin is consistent with findings in humans, likely reflecting the higher number of cell divisions in the male germline.
• De novo mutations contribute new genetic variation even in closed Thoroughbred populations, potentially impacting traits and breed evolution.
• The mutation causing a nonsense variant in a gene related to transcription regulation provides an example of potential functional impact from these novel mutations.

Significance and Implications

• These findings provide foundational data for genetic and evolutionary studies within horse breeds.
• Understanding mutation rates helps in managing genetic health and diversity in Thoroughbreds.
• Future work could explore the phenotypic consequences of identified mutations, including their effects on horse performance or disease susceptibility.
• Knowledge of parental origin biases can inform breeding strategies and genetic counseling in equine populations.