Semen Quality Of Bulls Research Articles

Sperm acrosome-associated 1 (SPACA1) mRNA and protein molecules deficiency indicate low fertility and semen quality of Bali bulls (Bos sondaicus)

Sperm acrosome-associated 1 (SPACA1) mRNA and protein molecules deficiency indicate low fertility and semen quality of Bali bulls (Bos sondaicus)

EFFECT OF SEASON ON METABOLITES AND SEMEN TRAITS OF BULLS

This study was conducted to investigate the effect of the winter and spring seasons on some biomarkers and semen characteristics of Holstein bulls (n = 20) during the Coronavirus pandemic period. Semen was collected for 16 weeks and evaluated weekly, along with the determination of some biomarkers. Sperm concentration, live sperm percentage, the concentrations of the most amino acids, essential, non-essential, and total amino acids, all carboxylic acids, omega 3, 6, 9, total fatty acids, and total saturated and unsaturated fatty acids, increased (P<0.05) in semen of spring compared to the winter season. Moreover, the percentage of plasma membrane integrity was higher in the winter than the spring season. The concentration of malondialdehyde in semen decreased (P<0.05) in the semen during the winter compared to the spring season. These biomarkers may have played a role in the fertility and semen quality or were affected by the season and repercussions of the Corona pandemic (feed shortage stress). In conclusion, the level of some amino and carboxylic acids can be adopted as an indicator of climate change (season) and semen quality of bulls or energy level in the diet.

Pyrroroquinoline Quinone (PQQ) Improves the Quality of Holstein Bull Semen during Cryopreservation.

Cryopreserved semen is extensively utilized in the artificial insemination (AI) of domestic animals; however, suboptimal conception rates due to oxidative damage following AI continue to pose a challenge. The present study investigated the effects of Pyrroroquinoline Quinone (PQQ), a novel antioxidant, on the semen quality of Holstein bulls during cryopreservation, as well as its potential molecular mechanisms. Semen samples were diluted with varying concentrations of PQQ (0, 50 μmol/L, 100 μmol/L, 150 μmol/L) prior to cryopreservation. Following the freeze-thaw process, a comprehensive evaluation was conducted to assess sperm motility, plasma membrane integrity, acrosome integrity, and the levels of reactive oxygen species (ROS), malondialdehyde (MDA), and adenosine triphosphate (ATP). Western blot analysis was employed to examine the levels of proteins including PGAM2, CAPZB, CAT, SOD1, and GPX1. Notably, the inclusion of 100 μmol/L PQQ significantly enhanced sperm motility, membrane integrity, and acrosome integrity post freeze-thawing (p < 0.05). Furthermore, the group treated with 100 μmol/L PQQ exhibited reduced levels of MDA and ROS (p < 0.05), while ATP levels were significantly elevated (p < 0.05). Interestingly, treatment with 100 μmol/L PQQ resulted in decreased consumption of PGAM2, CAPZB, CAT, SOD1, and GPX1 proteins in sperm after freeze-thawing, compared to the control group (p < 0.05). These findings indicate that PQQ treatment enhances the quality of bull semen, mitigates oxidative stress damage, and ultimately improves the efficacy of sperm cryopreservation.

Polymorphism detection and characterization of sperm cells chromatin remodeling associated genes in Murrah buffalo.

Seasonal variations significantly impact buffalo bull semen production and quality, particularly during the summer months. Understanding the genetic basis of these changes is important for managing bull fertility and improving sperm quality. The present study focused on characterizing and identifying polymorphisms in chromatin remodeling genes, protamines (PRMs) and Transition Nuclear Proteins (TNPs) in Murrah buffalo bulls with varying semen quality due to seasonal effects. Our findings revealed none of the coding region variation in PRM1, PRM2, TNP1, and TNP2, these genes are highly conserved in buffalo. Two intronic variants were identified, including G16C in PRM1 intron 1 and intronic SNP in PRM2 intron 1(G96A). The complete CDS of consensus sequence of bubaline PRM1 was 86.3% identical and 94.1% similar to the bovine PRM1. Whereas the complete CDS of consensus sequence of bubaline TNP2 was 78.2% identical and 91.0% similar to bovine TNP2. Further, no statistically significant differences in the fold change of TNP1, TNP2, PRM1, and PRM2 levels between the hot summer SNA and SA groups and the winter SNA and SA groups This study represents the first comprehensive report on the characterization of bubaline PRM1 (complete CDS), PRM2 (partial CDS), TNP1 (partial CDS), and TNP2 (complete CDS) genes in buffalo sperm cells. Results of the study, clearly indicate that the genes associated with protamine (PRM1 and TNP2) are highly conserved in Bubalus bubalis. Understanding these genetic underpinnings can have implications for improving buffalo bull fertility and semen quality.

Comparison of computer-assisted sperm analysis and smartphone-applied sperm analysis for evaluation of frozen-thawed bull semen.

This study aims to compare the efficacy of computer-assisted sperm analysis (CASA) and smartphone-applied sperm analysis (SASA) in assessing the quality of frozen-thawed bull semen. A total of 75 straws (n = 75) semen samples were used from different production batches of five Holstein bulls. The semen analyses were conducted in three groups: Group I (CASA-37°C), semen samples were evaluated using the CASA system at 37°C (n = 25); Group II (SASA-25°C), semen samples were assessed using the SASA system at a temperature of room heat (25°C) (n = 25); and Group III (SASA-37°C), semen samples were evaluated using the SASA system at 37°C (n = 25). The frozen-thawed bull semen samples were analysed in terms of total motility (TM), progressive motility (PM), immotile, velocity average path (VAP), velocity curve linear (VCL), velocity straight line (VSL) and sperm concentration. There was no significant difference between the groups in terms of spermatozoa concentration (p > .05). However, significant differences among the groups were observed for total motile spermatozoa values (p < .001). Values of progressive motile spermatozoa were lower in Group I and Group II compared to Group III (p < .001). The immotile spermatozoa values were significant between the groups (p < .001) and were found to be proportional to total motile spermatozoa values. Additionally, the VAP, VCL and VSL values were comparable between Group II and Group III, but lower when compared to Group I. In conclusion, the results of the study demonstrate that the Sperm Cell™ system can accurately analyse the concentration of frozen-thawed bull semen. The analyses performed at room temperature indicate a parallelism between the PM value and CASA results. However, it is thought that SASA devices require a series of standardization studies in different semen extenders, different sample concentrations and different animal species, analogous to the standardization evolution process of CASA devices in semen analysis.

The Effects of Different Thawing Procedures on Spermatological Parameters of Frozen Bull Sperm

One of the most crucial elements influencing the efficacy of artificial insemination applications is semen quality. It is known that post-thaw semen quality was affected by thawing method. Errors made during the thawing process can compromise the motility, viability, morphology and DNA integrity of spermatozoa after thawing. This study was conducted to investigate the effects of four distinct thawing procedures on post-thaw semen quality in bulls. The first group was designated as 24°C (n=8); the straws in this group were thawed in a water bath at 24°C for 60 seconds. Second group was designated as 32°C (n=8); the straws in this group were thawed in a water bath at 32°C for 60 seconds. Third group was designated as 38°C (n=8); the straws in this group were thawed in a water bath at 38°C for 30 seconds. And the fourth group was designated as 70°C (n=8); the straws in this group were thawed in a water bath at 70°C for 7 seconds. Following the thawing process, motility, sperm morphology, viability, and DNA damage rate in spermatozoa were evaluated. Results showed that in 32°C group and 24°C group sperm motility significantly decreased when compared to 70°C and 38°C groups. Additionally, the 24°C group exhibited a significantly elevated DNA damage rate compared to the 70°C group, while no differences were observed in the other groups. In conclusion, the most ideal thawing procedures are at 70°C for 7 seconds and at 38°C for 30 seconds are recommended to get optimum sperm quality from frozen bull semen after thawing.

Impact of Zinc Supplementation on Tharparkar Bull Semen Quality: Comparative Study of Commercial and Lab Extenders

Impact of Zinc Supplementation on Tharparkar Bull Semen Quality: Comparative Study of Commercial and Lab Extenders

Heat stress-induced dysbiosis of the gut microbiota impairs spermatogenesis by regulating secondary bile acid metabolism in the gut

Heat stress (HS) poses a substantial challenge to livestock. Studies have demonstrated that HS reduces fertility and leads to gut microbiota dysbiosis in bulls. However, the impact of the gut microbiota on fertility in bulls during HS is still unclear. Our research revealed that HS exposure decreased semen quality in bulls, and fecal microbiota transplantation (FMT) from heat-stressed bulls to recipient mice resulted in a significant decrease in number of testicular germ cells and epididymal sperm. Untargeted metabolomics methodology and 16S rDNA sequencing conjoint analysis revealed that Akkermansia muciniphila (A. muciniphila) seemed to be a key bacterial regulator of spermatogenesis after HS exposure. Moreover, the research indicated that A. muciniphila regulated secondary bile acid metabolism by promoting the colonization of bile salt hydrolase (BSH)-metabolizing bacteria, leading to increase of retinol absorption in the host gut and subsequently elevation of testicular retinoic acid level, thereby improving spermatogenesis. This study sheds light on the relationship between HS-induced microbiota dysbiosis and spermatogenesis, offering a potential therapeutic approach for addressing bull spermatogenic dysfunction triggered by HS exposure.

Role of trace minerals in bull reproductive physiology and semen quality

Testicular dysfunction is often associated with an imbalance in antioxidant/oxidant homeostasis, leading to negative effects of oxidative stress on germ cell proliferation, steroidogenesis, and sperm function. Trace minerals (TM) are involved in basic homeostatic and enzymatic processes like free radical detoxification, cellular respiration, carbohydrates, lipids, nucleic acids, synthesis and metabolism of proteins, and stabilization of membranes and DNA. Dietary source of TM is necessary to support these processes. Supplementing cattle with TM is nowadays a common practice to help support growth, reproduction, and immunity. This review provides information on roles of TM in bulls testicular and sperm functions, and effects of TM mineral supplementation on bull reproductive parameters.

Supplementation with MitoTEMPO before cryopreservation improves sperm quality and fertility potential of Piedmontese beef bull semen.

The purpose of this study was to improve the quality of frozen-thawed Piedmontese bull semen by incorporating MitoTEMPO (MT) in extended semen before cryopreservation. Semen was collected from 4 fertile bulls, using an artificial vagina, once weekly for 6 consecutive weeks. Semen samples were pooled, diluted with Bullxcell® extender, and supplemented with different concentrations of MT (0 as control, 5, 10, 20, 40, and 80 μM) before cooling, equilibration, and freezing procedures. The frozen-thawed semen was assessed for motility, vitality, acrosome intactness, plasma membrane integrity, DNA integrity, apoptosis, mitochondrial membrane potential, intracellular ROS level and in vitro fertilizing capability. The results showed that MT at concentrations of 10, 20, and 40 μM improved the total, progressive, and rapid motility directly after thawing while, at the highest tested concentration (80 μM), it decreased the progressive and rapid motility after 1, 2, and 3 h of incubation. The sperm kinetics including STR and LIN were noticeably increased at concentrations of 10, 20, and 40 μM directly after thawing (0 h), whereas the MT effect was variable on the other sperm kinetics during the different incubation periods. MitoTEMPO improved the sperm vitality at all tested concentrations, while the acrosomal and DNA integrity were improved at 20 μM and the mitochondrial membrane potentials was increased at 80 μM. The cleavage and blastocyst formation rates were significantly increased by using semen treated with 20 μM MT compared with controls. These findings suggest a potential use of MT mainly at a concentration of 20 μM as an additive in the cryopreservation media of bull semen to improve sperm quality.

Effect of the addition of soursop juice (Annona muricata L., 1753) to egg yolk citrate extender on the quality of bull semen

The present study was carried out to investigate the effect of the addition of soursop (Annona muricata L., 1753) juice to the citrate-egg yolk extender (EYC) on the quality of cryopreserved bull semen. After pre-evaluation, the semen was distributed in five extenders T1, T2, T3, T4 and T5 containing respectively 0% soursop juice (SJ) (T1), 25% SJ (T2), 50% SJ (T3), 75% SJ (T4), 100% SJ (T5) and microscopic evaluation was performed daily to assess plasma membrane integrity (PMI), progressive motility of spermatozoa (MP) and sperm pH until all sperm were dead. The results indicate a significant increase in spermatozoa PMI and MP (p&lt;0.05) in the case of treatment with 50% SJ (T3). Spermatozoa showed a motility of 50.78% 9 days later in the citrate-egg yolk extender containing 50% SJ (T3), whereas at day 9, spermatozoa in the other extenders had a progressive motility of less than 50%. Similar results were obtained for PMI with 50.78% of spermatozoa having an intact plasma membrane 9 days later in the citrate-egg yolk extender containing 50% SJ whereas the PMI of cells in the other stents was less than 50%. However, the pH of the extender containing 50% SJ was closest to 7 during the experimental period. The results of this experiment suggest that the addition of 50% soursop juice (T3) to the citrate-egg yolk extender could be adopted as an acceptable alternative to traditional egg yolk-based extenders for the cryopreservation of bull semen.

Selection of reference genes for normalizing gene expression data across seasons in spermatozoa of water buffalo (Bubalus bubalis).

Selection of the most stably expressed reference genes is key to monitoring accurate target gene expression across any tissue or cell type. The mRNA in spermatozoa stores valuable information related to changes in spermatogenesis due to variations in environmental conditions, especially during heat stress, which affects various sperm functions. Semen quality in buffalo bulls is significantly influenced by the seasons. In the study, a panel of nine genes was evaluated to identify the most stably expressed internal control gene (ICG) for the normalization of real-time gene expression data generated across various seasons for Murrah buffalo bulls' spermatozoa. Sperm cells were purified from the semen samples collected during different seasons, with temperature-humidity index (THI) ranging from 80.80 ± 1.47 (hot summer) to 55.88 ± 1.98 (winter), using the BoviPure™ gradient purification method. The RNA isolated from the purified spermatozoa fraction was quality checked prior to reverse transcription and subjected to qPCR (quantitative real-time PCR) based expression analysis. An automated 'endoGene' pipeline was employed to apply the geNorm, NormFinder, and BestKeeper algorithms for data analysis. The result indicated that GAPDH and PP1A were the most stably expressed among the gene panel, whereas ATPSF1 and ACTB were the two least stable expressed reference genes. Further, the most suitable ICGs identified were validated by normalization of real time expression data of heat stress and sperm quality genes, HSFY2 and AKAP4, respectively. The genes identified would help in generating the most reliable results for the expression profiling of the genes dictating sperm quality and heat stress cope-up mechanism in buffalo spermatozoa, collected during different seasons.

Effect of exogenous melatonin implant on post-thaw semen quality of buffalo bulls.

Melatonin is an intracellular antioxidant of sperm membrane that protects the cells from lipid peroxidation. Yet, its role as an antioxidant on semen quality of buffalo bulls is still obscure. The present study was undertaken to assess the effect of exogenous melatonin implant (18 mg/50 kg bodyweight) on post-thaw sperm characteristics, oxidative stress, endocrinological profiles and fertility of buffalo bulls. Six apparently healthy breeding Murrah buffalo bulls were randomly selected at bull farm, Guru Angad Dev Veterinary and Animal Sciences University for the present study and divided into two groups viz. control (n = 3) and melatonin implanted group (n = 3). A total of 120 ejaculates were collected from bulls of both groups (n = 60 each) throughout the study period. Most beneficial effects of melatonin implants were observed during post-implantation period. The percentages of post-thaw sperm total and progressive motility, viability and mitochondrial membrane potential were higher (p < .05) in melatonin implanted buffalo bulls compared to controls during post-implantation period. Following melatonin implantation, MDA production in post-thaw semen was lower (p < .05) in melatonin implanted group than in control group. Plasma melatonin and testosterone concentrations were higher (p < .05) in buffalo bulls implanted with melatonin as compared to their control counterparts. No differences (p > .05) in plasma LH concentrations were observed in both groups. First service pregnancy rate was 43.3% using semen of melatonin implanted bulls and 30.0% with semen of controls (p > .05). Thus, melatonin was able to protect sperm membrane against oxidative damage and improve post-thaw semen quality, thereby resulting in higher fertilizing potential of spermatozoa.

CFAP58 is involved in the sperm head shaping and flagellogenesis of cattle and mice.

CFAP58 is a testis-enriched gene that plays an important role in the sperm flagellogenesis of humans and mice. However, the effect of CFAP58 on bull semen quality and the underlying molecular mechanisms involved in spermatogenesis remain unknown. Here, we identified two single-nucleotide polymorphisms (rs110610797, A>G and rs133760846, G>T) and one indel (g.-1811_ g.-1810 ins147bp) in the promoter of CFAP58 that were significantly associated with semen quality of bulls, including sperm deformity rate and ejaculate volume. Moreover, by generating gene knockout mice, we found for the first time that the loss of Cfap58 not only causes severe defects in the sperm tail, but also affects the manchette structure, resulting in abnormal sperm head shaping. Cfap58 deficiency causes an increase in spermatozoa apoptosis. Further experiments confirmed that CFAP58 interacts with IFT88 and CCDC42. Moreover, it may be a transported cargo protein that plays a role in stabilizing other cargo proteins, such as CCDC42, in the intra-manchette transport/intra-flagellar transport pathway. Collectively, our findings reveal that CFAP58 is required for spermatogenesis and provide genetic markers for evaluating semen quality in cattle.

Effects of apigenin and trans-ferulic acid on microscopic and oxidative stress parameters in the semen of water buffalo bulls during cryopreservation

Cryopreservation involves exposing sperm to stressful conditions that affect cell viability. The high quality of the Azerbaijani water buffalo's by-products, such as buffalo milk, makes it a species of significant importance. Our focus is on protecting its genetic resources by preserving its sperm, as their numbers will decrease in the coming years and they are at risk of extinction. This study's goal was to ascertain how apigenin (A) and trans-ferulic acid (t-FA) affected the semen quality of Azari water buffalo bulls under cryopreservation. Pooled buffalo sperm (n = 35 ejaculations) were diluted in a Tris-based diluent also containing varying amounts of apigenin (0.2, 0.4, 0.6, and 0.8 mM) and trans-ferulic acid (2.5, 5, 10 and 20 mM). Following a freeze-thaw procedure, samples were assayed for total antioxidant capacity (TAC), catalase (CAT), superoxide dismutase (SOD), malondialdehyde (MDA), glutathione activity (GSH), glutathione peroxidase (GPx), progressive motility and total motility, motility properties, plasma membrane functionality, and viability. Sixty days after insemination, the rectal examination was performed on 38 buffaloes that had undergone sexual breeding to confirm pregnancy. The results of the study show that the addition of A-0.2, A-0.4, and t-FA-10 to buffalo semen increases the percentage of intact plasma membrane, motility, and sperm viability, as well as the levels of GSH, GPx, CAT. and TAC. In addition, there is a decrease in MDA and DNA damage after cryopreservation. Furthermore, the results show that 0.4 mM apigenin significantly increases conception rates compared to the control group. The base extender of Tris supplemented with A (0.4 and 0.2 mM) and t-FA (10 mM) improves the antioxidant indices of both frozen and thawed buffalo sperm, which in turn improves post-thawing sperm quality and in vivo fertility improves buffalo sperm.

Genome-wide 5′-C-phosphate-G-3′ methylation patterns reveal the effect of heat stress on the altered semen quality in Bubalus bubalis

Semen production and quality are closely correlated with different environmental factors in bovines, particularly for the buffalo (Bubalus bubalis) bulls reared under tropical and sub-tropical conditions. Factors including DNA methylation patterns, an intricate process in sperm cells, have an impact on the production of quality semen in buffalo bulls under abiotic stress conditions. The present study was conducted to identify DNA methylome signatures for semen quality in Murrah buffalo bulls, acclaimed as a major dairy breed globally, under summer heat stress. Based on semen quality parameters that significantly varied between the two groups over the seasons, the breeding bulls were classified into seasonally affected (SA = 6) and seasonally non-affected (SNA = 6) categories. DNA was isolated from purified sperm cells and sequenced using the RRBS (Reduced Representation Bisulfite Sequencing) technique for genome-wide methylome data generation. During the hot summer months, the physiological parameters such as scrotal surface temperature, rectal temperature, and respiration rate for both the SA and SNA bulls were significantly higher in the afternoon than in the morning. Whereas, the global CpG% of SA bulls was positively correlated with the afternoon's scrotal surface and rectal temperature. The RRBS results conveyed differentially methylated cytosines in the promoter region of the genes encoding the channels responsible for Ca2+ exchange, NPTN, Ca2+ activated chloride channels, ANO1, and a few structure-related units such as septins (SEPT4 and SEPT6), SPATA, etc. Additionally, the hypermethylated set of genes in SA was significantly enriched for pathways such as the FOXO signaling pathway and oocyte meiosis. The methylation patterns suggest promoter methylation in the genes regulating the sperm structure as well as surface transporters, which could contribute to the reduced semen quality in the Murrah buffalo bulls during the season-related heat stress.

Analysis of Correlation between PRM1, STK35, and IFT27 Gene Expression Levels and Holstein Bull Semen Quality Parameters

Analysis of Correlation between PRM1, STK35, and IFT27 Gene Expression Levels and Holstein Bull Semen Quality Parameters

Regulatory functions of lecithin nanoparticles on post-thaw quality of buffalo bull semen: a morphological analysis of spermatozoa

Regulatory functions of lecithin nanoparticles on post-thaw quality of buffalo bull semen: a morphological analysis of spermatozoa

The Libido and Semen Quality of Bali Bulls Supplemented with Micro-Nutrients of Zinc, Selenium, Vitamin A and Vitamin E

Low quality and quantity of food intake of bulls in the tropics might adversely affect libido and semen quality through their effects on spermatogenesis process. The purpose of this study was to evaluate the effect of micro-minerals (Zn and Se) and vitamins (A and E) on the libido and semen quality of Bali bulls. 2 (two) Bali bulls, aged 4-5 years old with a body weight 300-350 kg. The study was divided into 2 periods and each period was conducted for 8 weeks. At the first period (control), the experimental bulls were fed with natural grasses and concentrates (1% of body weight). Furthermore, at the second period (treatment period), they were fed diet similar to that used in the first period with an additional of a mineral mix (Zn and Se) and vitamin (A and E). Evaluation of libido and semen quality were carried out simultaneously twice a week. Libido was measured by calculating the time required for the bulls from approaching the teaser until they mounted for the first time and from approaching until they ejaculated, respectively. Semen was collected by using an artificial vagina. The volume, colour, pH, and, viscosity, as were recorded. Evaluation of semen quality in the form of the sperm concentration, motility, viability and abnormality were done by using Computer Assisted Semen Analyser (CASA). Fresh semen was diluted by Andromed®. The diluted semen was preserved at the temperature 50C for 144 hrs. Motility, viability and abnormality were observed every 12 hrs. Data were reported as mean and standard error of mean analysed by using the paired student’s t-test. Mean interval between the bulls approaching and mounting the teaser for the first time and between approaching and ejaculation after the supplementation of micro-nutrients were significantly shorter (P&lt;0.05) in compared to those before the supplementation (2.68 vs. 6.52 and 5.09 vs. 10.06 minutes, respectively). The sperm viability after the supplementation of micro-nutrients was significantly higher in compared to that before supplementation. (82.25 vs. 87.50 %). Preservation of diluted bull semen at the temperature of 50C for 144 hrs showed that the sperm viability was significantly higher (P&lt;0.05) and sperm abnormality was significantly lower (P&lt;0.05) after the supplementation of micro-nutrients in compared to those before the supplementation. It can be concluded that the supplementation micro-nutrients in Bali bulls could increase libido, sperm viability and reduce the sperm abnormality.

Risk of Sperm Disorders and Impaired Fertility in Frozen-Thawed Bull Semen: A Genome-Wide Association Study.

Cryopreservation is a widely used method of semen conservation in animal breeding programs. This process, however, can have a detrimental effect on sperm quality, especially in terms of its morphology. The resultant sperm disorders raise the risk of reduced sperm fertilizing ability, which poses a serious threat to the long-term efficacy of livestock reproduction and breeding. Understanding the genetic factors underlying these effects is critical for maintaining sperm quality during cryopreservation, and for animal fertility in general. In this regard, we performed a genome-wide association study to identify genomic regions associated with various cryopreservation sperm abnormalities in Holstein cattle, using single nucleotide polymorphism (SNP) markers via a high-density genotyping assay. Our analysis revealed a significant association of specific SNPs and candidate genes with absence of acrosomes, damaged cell necks and tails, as well as wrinkled acrosomes and decreased motility of cryopreserved sperm. As a result, we identified candidate genes such as POU6F2, LPCAT4, DPYD, SLC39A12 and CACNB2, as well as microRNAs (bta-mir-137 and bta-mir-2420) that may play a critical role in sperm morphology and disorders. These findings provide crucial information on the molecular mechanisms underlying acrosome integrity, motility, head abnormalities and damaged cell necks and tails of sperm after cryopreservation. Further studies with larger sample sizes, genome-wide coverage and functional validation are needed to explore causal variants in more detail, thereby elucidating the mechanisms mediating these effects. Overall, our results contribute to the understanding of genetic architecture in cryopreserved semen quality and disorders in bulls, laying the foundation for improved animal reproduction and breeding.