Genome-Wide Analysis of Genetic Predispositions Linked to Damaged Membranes and Impaired Fertility as Indicators of Compromised Sperm-Egg Interaction Mechanisms in Frozen-Thawed Rooster Semen.

Cryopreservation cannot be widely used for rooster sperm due to high incidences of cryoinjury, including damage to sperm membranes. Thus, cryopreserved rooster sperm has limited use due to low sperm motility and reduced fertilizing ability, which disrupts the mechanisms involved in sperm-egg interactions. Previously, we used an Illumina 60K single-nucleotide polymorphism (SNP) array to search for genes associated with rooster sperm quality, before and after freeze-thawing. As a continuation of these genome-wide association studies (GWAS), the present investigation used a denser 600K SNP chip. Consequently, the screen depth was expanded by many markers for cryo-resistance in rooster sperm while more candidate genes were identified. Thus, our study aimed to identify genome-wide associations with ejaculate quality indicators, including those concerning sperm membrane damage. We selected sperm quality indicators after freezing-thawing using samples from a proprietary cryobank collection created to preserve generative and germ cells of rare and endangered breeds of chickens and other animal species. A total of 258 ejaculates from 96 roosters of 16 different breeds were analyzed. Moreover, 96 respective DNA samples were isolated for genotyping using a 600K Affymetrix® Axiom® high-density genotyping array. In total, 31 SNPs and 26 candidate genes were associated with characteristics of sperm membrane damage, progressive motility, and sperm cell respiration induction using 2,4-dinitrophenol. In particular, we identified the ENSGALG00000029931 gene as a candidate for progressive motility, PHF14 and ARID1B for damaged sperm membranes, and KDELR3, DDX17, DMD, CDKL5, DGAT2, ST18, FAM150A, DIAPH2, MTMR7, NAV2, RAG2, PDE11A, IFT70A, AGPS, WDFY1, DEPDC5, TSC1, CASZ1, and PLEKHM2 for sperm cell respiration induction. Our findings provide important information for understanding the genetic basis of sperm membrane integrity and other traits that can potentially compromise the mechanisms involved in sperm-egg interactions. These findings are relevant to the persistence of fertility after thawing previously frozen rooster semen.