Boar Semen Quality Research Articles

Supplementation of glutamine in a short-term boar semen extender during 17°C holding time enhances post-thaw sperm quality for cryopreservation.

Glutamine is a nonessential amino acid and the most abundant amino acid found in the seminal plasma and sperm-rich fraction of boar semen. Glutamine plays an important role in enhancing glutathione (GSH) synthesis. It acts as an effective antioxidant in semen and provides intracellular defense to sperm against oxidative stress. This study aimed to improve the quality of frozen-thawed boar semen by using glutamine supplementation in a short-term semen extender during the holding time at 17°C before cryopreservation. The results indicate that the total motility, progressive motility, LIN, STR, and WOB were the highest in the 20 mM supplementation group at the 2h timepoint after thawing. Thus, the optimal concentration for glutamine supplementation in short-term boar semen extender during the holding time at 17°C was 20 mM. Interestingly, at all of the time points after thawing, 20 mM glutamine supplementation exhibited the highest level of sperm viability and membrane integrity when compared to the CONTROL (0 mM) and other experimental dilution groups. Moreover, the acrosome integrity, mitochondrial activity, and capacitation status (F pattern) were significantly greater in the 20 mM supplementation group than the other groups at the 2h timepoint after thawing. Supplementation of glutamine at a concentration of 20 mM in a short-term semen extender (Bio Pig®) during the 17°C holding time before cryopreservation, which had a standard freezing extender (9.0% glycerol and 1.9% Equex paste), could enhance the post-thaw sperm motility and quality parameters of cryopreservation.

GPX5-Enriched Exosomes Improve Sperm Quality and Fertilization Ability

Semen preservation quality affects the artificial insemination success rate, and seminal exosomes are rich in various proteins that are transferable to sperm and conducive to sperm-function preservation during storage. However, the specific effects of these proteins remain unclear. In this study, the specific effects of these proteins on semen preservation quality and fertilization capacity were investigated through a proteomic analysis of seminal exosomes from boars with high conception rates (HCRs) and low conception rates (LCRs). The results revealed significant differences in the expression of 161 proteins between the two groups, with the GPX5 level being significantly higher in the HCR group (p < 0.05). The role of GPX5 was further investigated by constructing engineered exosomes enriched with GPX5 (Exo-GPX5), which could successfully transfer GPX5 to sperm. Compared to the control group, Exo-GPX5 could significantly improve sperm motility on storage days 4 and 5 and enhance the acrosome integrity on day 5 (p < 0.05). Additionally, Exo-GPX5 increased the total antioxidant capacity (T-AOC) of sperm, reduced the malondialdehyde (MDA) level, and decreased the expression of antioxidant proteins SOD1 and CAT (p < 0.05). In simulated fertilization experiments, Exo-GPX5-treated sperm exhibited higher capacitation ability and a significant increase in the acrosome reaction rate (p < 0.05). Overall, Exo-GPX5 can improve boar semen quality under 17 °C storage conditions and enhance sperm fertilization capacity.

Temporal effect of flaxseed oil in boar’s diet on semen quality, antioxidant status and in-vivo fertility under hot humid sub-tropical condition

The present study investigates the temporal effects of flaxseed supplementation on boar semen quality, antioxidant status, and in-vivo fertility under high-temperature humidity index (THI) conditions in a sub-tropical climate. Twelve Hampshire crossbreed boars were randomly assigned to control and treatment groups, with the treatment group receiving flaxseed oil supplementation. Semen samples were collected and analyzed for semen quality parameters, sperm kinematics, and antioxidant status. Fertility outcomes were assessed through in-vivo mating trials. Flaxseed supplementation resulted in time dependent significant improvements in semen volume, sperm concentration, total and progressive sperm motility, sperm quality parameters, and antioxidant status. Fertility outcomes, including farrowing rates and litter sizes, were also enhanced in the flaxseed-supplemented group. These findings highlight the potential of flaxseed supplementation to improve boar fertility under high ambient stress conditions, with implications for optimizing reproductive performance in swine production systems.

Deep learning classification method for boar sperm morphology analysis.

Boar semen quality emphasizes three major criteria: sperm concentration, motility, and morphology. Methods to analyze concentration and motility quickly and objectively readily exist, but few exist for analyzing morphology outside of subjective manual counting. Other vital factors for fertilization, like acrosome health, lack efficient detection methods due to limitations in detection by the human eye and costly biomarker analysis, which is rarely used in semen diagnostics. To overcome these challenges, we propose a novel approach integrating deep-learning technology with high-throughput image-based flow cytometry (IBFC) for objective and accurate analysis of both morphology and label-free acrosome health of thousands of individual spermatozoa at once, as opposed to manually counting on a microscope slide. Images of 10,000 spermatozoa were captured using an IBFC and manually annotated based on the primary morphological defect or acrosome health status for the training of the convolutional neural network (CNN). The CNN used these images to train and then applied that training to unannotated images to predict the model accuracy. Using the CNNs, high F1 scores of 96.73%, 98.55%, and 99.31% for 20x, 40x, and 60x magnifications, respectively, for morphological classification were attained. Additionally, the model demonstrates an F1 score of 99.8% in detecting subtle acrosome health variations at the 60x magnification. We have established an integrated approach to rapidly collect and classify morphological defects and acrosome health status, without the use of manual counting or biomarker labeling. Our study underscores the potential of artificial intelligence in semen diagnostics, reducing technician variability, streamlining assays, and facilitating the development of additional label-free detection methods. This innovative approach addresses the barriers hindering biomarker adoption in semen analysis, offering a promising avenue for enhancing reproductive health assessments.

PSVII-28 Unraveling the mechanisms governing boar semen quality during chilled storage

Abstract Artificial insemination (AI) appears as a core of reproductive management in the hog industry. AI involves the use of chilled semen doses meeting routine thresholds for motility (≥ 70%) and morphology (≥ 80%) at collection. However, semen doses meeting these standards exhibit a gradual decline in sperm quality, with likely divergent profiles during preservation that impact fertility outcomes. Hence, this study aims to unravel the sperm quality dynamics of boar semen doses during 7-d chilled storage at 17ºC. Semen doses (n = 25) from proven-fertile Duroc boars were provided by a commercial hog farm (Prestage Farm, MS) and transported to the laboratory within 30 min of collection. Each semen sample was aliquoted and analyzed on the day of collection (d 0) and after 7 d of storage (d 7) at 17°C. Sperm total motility (TM), progressive motility (PM), and normal morphology (NM) were assessed using a computer-assisted sperm analyzer (CASA; CEROS II, IMV Biotechnologies). Apoptosis (Annexin V/7-AAD), viability (Propidium iodide – PI), and intracellular reactive oxygen species (ROS; DCFH-DA) were evaluated using flow cytometry. Total antioxidant activity (TAOC) and lipid peroxidation (malondialdehyde; MDA) were assessed using commercial kits (Cayman Chemical). Data analysis was conducted using parametric statistics (SPSS), with P < 0.05 considered significant. Results were presented as mean ± SEM. Data analysis confirmed the significant decrease of CASA parameters, particularly TM from d 0 to d 7 (78.87 ± 1.37 vs. 73.19 ± 1.98%), with extreme variance on d 7 allowing to discriminate high motility (HM, n = 6) vs low motility (LM, n = 6) semen doses. HM doses evidenced significantly greater (P < 0.05) TM (83.73 ± 1.17%), PM (36.07 ± 1.11%), and NM (89.45 ± 2.30%) compared with LM doses (TM: 59.40 ± 3.35%, PM: 16.57 ± 3.25%, NM: 79.93 ± 2.61%) on d 7. Unlike HM samples, which were comparable between d 0 and d 7, LM doses displayed significantly greater proportions of apoptotic and viable cells, and increased intracellular ROS levels on d 0. Likewise, LM samples revealed significantly greater (P < 0.05) proportions of apoptotic cells (3.69 ± 1.66%), dead cells (38.41 ± 3.71%), and cells with intracellular ROS (36.21 ± 3.62%) than HM samples (apoptotic cells: 0.03 ± 0.02%, dead cells: 4.73 ± 4.65%, ROS: 5.34 ± 4.91%) on d 0; however, only the proportions of dead cells displayed significant increase in LM on d 7. MDA concentrations were comparable between HM and LM samples on d 0. However, on d 7, LM samples exhibited significantly greater MDA concentrations than HM samples. In contrast, TAOC was significantly greater in HM samples (2.53 ± 0.14 mM) compared with LM samples (2.17 ± 0.07 mM) on d 0. In conclusion, semen samples with similar gross characteristics at collection exhibited differential responses during storage. Their initial redox potential and apoptosis-like changes may contribute to their varying ability to withstand prolonged storage. Work supported by USDA-ARS grant# 6066-31000-015-00D.

Alginate oligosaccharide supplementation improves boar semen quality under heat stress

Heat stress is a serious problem that affects animal husbandry by reducing growth and reproductive performance of animals. Adding plant extracts to the diet is an effective way to help overcome this problem. Alginate oligosaccharide (AOS) is a natural non-toxic antioxidant with multiple biological activities. This study analyzed the potential mechanism of AOS in alleviating heat stress and improving semen quality in boars through a combination of multiple omics tools. The results indicated that AOS could significantly increase sperm motility (P < 0.001) and sperm concentration (P < 0.05). At the same time, AOS improved the antioxidant capacity of blood and semen, and increased blood testosterone (P < 0.05) level. AOS could improve the metabolites in sperm, change the composition of gut microbiota, increase the relative abundance of beneficial bacteria such as Pseudomonas (P < 0.01), Escherichia-Shigella (P < 0.05), Bifidobacterium (P < 0.01), reduce the relative abundance of harmful bacteria such as Prevotella_9 (P < 0.05), Prevotellaceae_UCG-001 (P < 0.01), and increase the content of short chain fatty acids. Proteomic results showed that AOS increased proteins related to spermatogenesis, while decreasing heat shock protein 70 (P < 0.05) and heat shock protein 90 (P < 0.01). These results were verified using immunofluorescence staining technology. There was a good correlation among sperm quality, sperm metabolome, sperm proteome, and gut microbiota. In conclusion, AOS can be used as a feed additive to increase the semen quality of boars to enhance reproductive performance under heat stress.

A preliminary study of Lasia spinosa Thw. feed supplementation improving semen quality of subfertile purebred boars

A preliminary study of Lasia spinosa Thw. feed supplementation improving semen quality of subfertile purebred boars

Supplementation of Moringa oleifera leaf meal in feed on sexual behavior and semen quality of Landrace-cross boars

Supplementation of Moringa oleifera leaf meal in feed on sexual behavior and semen quality of Landrace-cross boars

Source and level of dietary iron influence semen quality by affecting inflammation, oxidative stress and iron utilization levels in boars

BackgroundBoars fed a mixed form of inorganic and organic iron in excess of the NRC recommended levels still develop anemia, which suggested that the current level and form of iron supplementation in boar diets may be inappropriate. Therefore, 56 healthy Topeka E line boars aged 15–21 months were randomly divided into 5 groups: basal diet supplemented with 96 mg/kg ferrous sulfate (FeSO4) and 54 mg/kg glycine chelated iron (Gly-Fe, control); 80 mg/kg or 115 mg/kg Gly-Fe; 80 mg/kg or 115 mg/kg methionine hydroxyl analogue chelated iron (MHA-Fe, from Calimet-Fe) for 16 weeks. The effects of dietary iron supplementation with different sources and levels on semen quality in boars were investigated.Results1) Serum Fe and hemoglobin concentrations were not affected by reduced dietary iron levels in the 80 mg/kg or 115 mg/kg Gly-Fe and MHA-Fe groups compared with the control group (P > 0.05). 2) Serum interleukin-6 (IL-6) and sperm malondialdehyde (MDA) levels in the 80 mg/kg or 115 mg/kg MHA-Fe groups were lower than those in the control group (P < 0.05), and higher serum superoxide dismutase levels and lower MDA levels in the 115 mg/kg MHA-Fe group (P < 0.05). 3) Boars in the 80 mg/kg or 115 mg/kg Gly-Fe and MHA-Fe groups had lower serum hepcidin (P < 0.01), ferritin (P < 0.05), and transferrin receptor (P < 0.01) concentrations, and boars in the 115 mg/kg MHA-Fe group had higher seminal plasma Fe concentrations compared with the control group. 4) Boars in the 80 mg/kg and 115 mg/kg MHA-Fe groups had lower abnormal sperm rate and in situ oscillating sperm ratio compared to the control group at weeks 12 and/or 16 of the trial. However, the effect of Gly-Fe on improving semen quality in boars was not evident. 5) Serum IL-6 level was positively correlated with hepcidin concentration (P < 0.05), which in turn was significantly positively correlated with abnormal sperm rate (P < 0.05). Furthermore, significant correlations were also found between indicators of iron status and oxidative stress and semen quality parameters.ConclusionsDietary supplementation with 80 mg/kg or 115 mg/kg MHA-Fe did not induce iron deficiency, but rather reduced serum inflammatory levels and hepcidin concentration, alleviated oxidative stress, increased body iron utilization, and improved semen quality in adult boars.

Proteomic Analysis of Differences in the Freezability of Porcine Sperm Identifies α-Amylase As a Key Protein.

To investigate the mechanisms underlying the differences in the freezability of boar semen, Yorkshire boars with freezing-tolerant semen (YT, n = 3), Yorkshire boars with freezing-sensitive semen (YS, n = 3), Landrace boars with freezing-tolerant semen (LT, n = 3), and Landrace boars with freezing-sensitive semen (LS, n = 3) were selected for this study. Their sperm was subjected to protein extraction, followed by data-independent acquisition proteomics and functional bioinformatics analysis. A total of 3042 proteins were identified, of which 2810 were quantified. Some key KEGG pathways were enriched, such as starch and sucrose metabolism, carbohydrate digestion and absorption, mineral absorption, the HIF-1 signaling pathway, and the necroptosis pathways. Through PRM verification, we found that several proteins, such as α-amylase and epididymal sperm-binding protein 1, can be used as molecular markers of the freezing resistance of boar semen. Furthermore, we found that the addition of α-amylase to cryoprotective extender could significantly improve the post-thaw motility and quality of boar semen. In summary, this study revealed some molecular markers and potential molecular pathways contributing to the high or low freezability of boar sperm, identifying α-amylase as a key protein. This study is valuable for optimizing boar semen cryopreservation technology.

The Effects of Different Antimicrobial Peptides (A-11 and AP19) on Isolated Bacteria from Fresh Boar Semen and Semen Quality during Storage at 18 °C.

Antibiotic resistance (AMR) is a major public health concern. Antimicrobial peptides (AMPs) could be an alternative to conventional antibiotics. The purpose of this research was to investigate the antimicrobial ability of the synthetic AMPs (i.e., A-11 and AP19) on the most frequently isolated bacteria in boar semen and their effect on extended boar semen quality during storage. We tested the antimicrobial effect of A-11 and AP19 at different concentrations and compared them with gentamicin for inhibiting the growth of E. coli, Pseudomonas aeruginosa and Proteus mirabilis that were isolated from fresh boar semen. In order to evaluate the effect of AMP on semen qualities on days 0, 1, 3, and 5 after storage at 18 °C, seven fresh boar semen samples were collected, diluted with semen extender with antibiotic (i.e., gentamicin at 200 µg/mL, positive control) or without (negative control), and semen extender contained only A-11 or AP19 at different concentrations (i.e., 62.50, 31.25, and 15.625 µg/mL). The total bacterial count was also measured at 0, 24, 36, 48, and 72 h after storage. Comparable to gentamicin, both A-11 and AP19 inhibited the growth of E. coli, Pseudomonas aeruginosa, and Proteus mirabilis at 62.50, 31.25, and 15.625 µg/mL, respectively. Comparing the total bacterial count at 0, 24, 36, 48 and 72 h after storage, the lowest total bacterial concentration was found in the positive control group (p < 0.05), and an inferior total bacterial concentration was found in the treatment groups than in the negative control. On day 1, there is a lower percentage of all sperm parameters in the AP19 group at a concentration of 62.50 µg/mL compared with the other groups. On day 3, the highest percentage of all sperm parameters was found in the positive control and A-11 at a concentration of 31.25 µg/mL compared with the other groups. The AP19 group at 62.5 µg/mL constantly yielded inferior sperm parameters. On day 5, only A-11 at a concentration of 15.625 µg/mL showed a total motility higher than 70%, which is comparable to the positive control. A-11 and AP19 showed antimicrobial activity against E. coli, Pseudomonas aeruginosa and Proteus mirabilis isolated from boar semen. Considering their effect on semen quality during storage, these antimicrobial peptides are an alternative to conventional antibiotics used in boar semen extenders. Nevertheless, the utilization of these particular antimicrobial peptides relied on the concentration and duration of storage.

Patterns of alteration in boar semen quality from 9 to 37 months old and improvement by protocatechuic acid

BackgroundComprehending the patterns of alteration in boar semen quality and identifying effective nutritional interventions are crucial for enhancing the productivity of commercial pig systems. This study aimed to examine the alteration in semen quality in boars, and assess the impact of protocatechuic acid (PCA) on semen quality during the phase of declining semen quality.MethodsIn Exp. 1, a total of 38 Pig Improvement Company (PIC) boars were selected and their semen quality data were recorded from the age of 9 to 37 months. In Exp. 2, 18 PIC boars (28 months old) were randomly assigned into three groups (n = 6) and fed a basal diet, a basal diet containing 500 or 1,000 mg/kg PCA, respectively. The experiment lasted for 12 weeks.ResultsThe semen volume, concentration, and total number of spermatozoa in boars exhibited an increase from 9 to 19 months old and showed a significant linear decreased trend in 28, 24, and 22 months old. Sperm motility displayed an upward trajectory, reaching its peak at 20 months of age, and showed a significant linear decreased trend at 20 months old. Dietary supplementation of PCA demonstrated an effect to mitigate the decrease in semen volume, concentration of spermatozoa, total number of spermatozoa (P > 0.05), and significantly increased the sperm motility (P < 0.05). Moreover, supplementation of 1,000 mg/kg PCA significantly increased the sperm viability (P < 0.05). Analysis on cellular signaling pathways revealed that PCA restored serum testosterone levels and alleviated oxidative damage by upregulating the expression of HO-1, SOD2, and NQO1 in testicular stromal cells. Notably, PCA can enhance phosphorylation by selectively binding to AMP-activated protein kinase (AMPK) protein, thereby improving sperm mitochondrial function and augmenting sperm motility via PGC-1/Nrf1.ConclusionsThese data elucidated the pattern of semen quality variation in boars within the age range of 9 to 37 months old, and PCA has the potential to be a natural antioxidant to enhance sperm quality through modulation of the AMPK/PGC-1/Nrf1 signaling pathway.

20 Effects of a stabilized fish oil on boar semen quality when fed during summer months

Abstract The objective of this study was to investigate the effects of feeding a stabilized fish oil source to different genetic lines on boar semen quality. A total of 127 boars representing three genetic lines (PIC 337, PIC 800, and PIC L03; Hendersonville, TN) at a commercial boar stud were balanced by age, body condition, and previous semen quality and randomly allotted to one of two dietary treatments within genetic line. Boars were assigned to either receive Control; a daily top dress of 32g of corn starch; or Fish oil; a daily top dress of 32g of Salmate DHA+ (The Ballard Group, 2407 Ashland Ave., Cincinnati, OH 45206 USA) to provide 2.38 g of docosahexaenoic acid (DHA). The top-dress supplementation was provided by placing it on top of the daily ration of feed for each individual boar on the floor once each day immediately after their feed has been delivered to them. Supplementation lasted for 16 wk from June-September, 2023 during which weekly semen collection was performed. Semen was evaluated at the stud for concentration and motility parameters using computer assisted semen analysis. Data was analyzed using generalized linear mixed models with the lmer and glmer functions of R. Boar was the experimental unit, with dietary treatment, genetic line, and their interaction tested as fixed effects while age at collection served as a covariate. Boar nested within treatment combinations was used as a random effect to account for repeated measurements within the same boar. There was no evidence (P &amp;gt; 0.10) for dietary treatment by genetic line interaction for any response criteria (Table 1). There was no evidence of fish oil supplementation effects (P &amp;gt; 0.10) on semen volume, concentration, total sperm cells per ejaculate, percentage of accepted ejaculates, or number of doses produced per accepted ejaculate. There was no evidence of fish oil supplementation effects (P &amp;gt; 0.10) on percentage motile sperm cells, or percentage of proximal and distal droplets and abnormal tails. Line 800 boars had the least (P &amp;lt; 0.05) semen volume, but greatest (P &amp;lt; 0.05) sperm concentration, resulting in no evidence for differences (P &amp;gt; 0.10) between the three genetic lines in total sperm cells per ejaculate. Line 03 boars had increased (P &amp;lt; 0.05) percentage of rejected ejaculates as well as decreased number of doses produced per accepted ejaculate (P &amp;lt; 0.05) compared with lines 337 and 800 boars. There was no evidence for differences (P &amp;gt; 0.10) in sperm motility, proximal and distal droplets, or abnormal tails between the three genetic lines. In summary, maternal line boars had an increased proportion of rejected ejaculates per boar and a decreased number of doses produced compared with terminal line boars. Feeding a stabilized fish oil to provide 2.38 g of DHA did not improve semen during summer months.

Effect of selenium nanoparticles on the quality and fertility of short-term preserved boar semen.

This study was carried out to investigate the effect of different concentrations of selenium nanoparticles (Se-NPs) in the Beltsville Thawing Solution (BTS) extender on the semen quality and fertility of Hampshire crossbred pigs. For the study, semen was collected from four boars (10 ejaculates/boar) by the gloved hand method. Each ejaculate was extended @ 1:2 with the BTS extender and split into four aliquots. The control (C) samples were without the supplementation of Se-NPs, whereas the other three were supplemented with 0.5 (T1), 1 (T2), and 2 μg ml-1 of Se-NPs (T3) and stored at 15°C in a BOD incubator. Extended semen was evaluated at 0 (immediately after dilution), 24, 48, 72, and 96 h of storage for sperm motility, live sperm, plasma membrane integrity, acrosome integrity, DNA integrity, and mitochondrial membrane potential (MMP). The mean percentage of sperm motility, live sperm, and sperm with intact plasma membrane and acrosome, and MMPs were significantly (p < 0.01) higher in all treated groups in comparison to control at 24, 48, 72, and 96 h of storage. Sperm with intact DNA in all treated groups increased significantly at 48 (p < 0.05), and 72 and 96 (p < 0.01) h of storage in comparison to the control group. The concentration of 1 μg ml-1 of Se-NPs was found to be the best among other concentrations. In each group, 10 sows were artificially inseminated with the liquid semen preserved for 72 h at 15°C. Supplementation of 1 μg ml-1 of Se-NPs yielded the highest conception rate in comparison to other groups. In conclusion, supplementation of 1 μg ml-1 of Se-NPs in the BTS extender resulted in the best semen quality and conception rate during the short-time liquid preservation of boar semen.

Effects of ergothioneine supplementation on the quality of liquid-preserved and frozen-thawed boar semen.

This study examined the effects of ergothioneine (EGT) supplementation as an antioxidant on the quality of boar spermatozoa when using liquid and frozen preservation methods. In the first experiment, boar semen was preserved in an extender supplemented with 0, 50, 100 and 200 µM EGT, at15 °C, part of the samples for one and another part for three weeks. In comparison with the control (without EGT), EGT supplementation at 100 µM significantly increased the percentage of total motility of spermatozoa that were preserved as a liquid both for one and three weeks (P < 0.05). EGT supplementation did not affect the quality of preserved spermatozoa, irrespective of the EGT concentration. In the second experiment, semen was frozen and thawed in the freezing extender supplemented with 0,50, 100 and 200 µM EGT. In comparison with the control, the 100 µM EGT supplementation significantly increased the percentages of total and progressive motility of frozen-thawed spermatozoa (P < 0.05). EGT (100 µM) supplementation did not affect the viability, the plasma membrane integrity, or the acrosomal integrity of frozen-thawed spermatozoa. These findings indicate that supplementing extenders with 100 µM EGT may improve the motility of boar sperm in both liquid and freezing preservation methods.

Alginate oligosaccharide extends the service lifespan by improving the sperm metabolome and gut microbiota in an aging Duroc boars model.

Alginate oligosaccharide (AOS), as a natural non-toxic plant extract, has been paid more attention in recent years due to its strong antioxidant, anti-inflammatory, and even anti-cancer properties. However, the mechanism by which AOS affects animal reproductive performance is still unclear. The purpose of this study is to use multi-omics technology to analyze the effects of AOS in extending the service lifespan of aging boars. The results showed that AOS can significantly improve the sperm motility (p < 0.05) and sperm validity rate (p < 0.001) of aging boars and significantly reduce the abnormal sperm rate (p < 0.01) by increasing the protein levels such as CatSper 8 and protein kinase A (PKA) for semen quality. At the same time, AOS significantly improved the testosterone content in the blood of boars (p < 0.01). AOS significantly improved fatty acids such as adrenic acid (p < 0.05) and antioxidants such as succinic acid (p < 0.05) in sperm metabolites, significantly reducing harmful substances such as dibutyl phthalate (p < 0.05), which has a negative effect on spermatogenesis. AOS can improve the composition of intestinal microbes, mainly increasing beneficial bacteria Enterobacter (p = 0.1262) and reducing harmful bacteria such as Streptococcus (p < 0.05), Prevotellaceae_UCG-001 (p < 0.05), and Prevotellaceae_NK3B31_group (p < 0.05). Meanwhile, short-chain fatty acids in feces such as acetic acid (p < 0.05) and butyric acid (p < 0.05) were significantly increased. Spearman correlation analysis showed that there was a close correlation among microorganisms, sperm metabolites, and sperm parameters. Therefore, the data indicated that AOS improved the semen quality of older boars by improving the intestinal microbiota and sperm metabolome. AOS can be used as a feed additive to solve the problem of high elimination rate in large-scale boar studs.

Effect of Zinc Supplementation to Diluent Boar Semen on Sperm Characteristics and Activity of Antioxidant Enzymes

Background. Zinc is essential for male reproductive function as it is necessary for spermatozoa maturation, capacitation, acrosome reaction, and fertilization. It has been established that the use of chelates compounds of metals with amino acids, nucleotides, peptides, and carbohydrates is not only more effective but also economically justified. Objective. The aim of this study was to evaluate and compare the effect of different concentrations of Zn glutamic amino acid chelate (ZnGlu) on boar spermatozoa motility, viability parameters, and prooxidant-antioxidant homeostasis during in vitro incubation. Methods. Freshly ejaculated boar semen, after the addition of "Ecosperm" diluent, was divided into groups: a control group and three experimental groups. Zinc glutamate was supplemented to the experimental samples at concentrations of 1.0, 2.0, and 5.0 µg/ml, respectively. Semen samples were stored at 18 °C for 4 days. Every 24 hours, sperm motility and viability and the antioxidant status were assessed by the level of diene con­jugates (DC), concentration of malondialdehydere (MDA), and the activities of catalase and superoxide dismutase. Results. It was observed that boar sperm motility decreased with the extension of storage time, while the addition of 2.0 and 5.0 µg ZnGlu significantly improved sperm total motility and the percentage of vitality spermatozoa during 48–96 hours of incubation. Our studies also demonstrate that ZnGlu possess a protective effect in alleviating oxidative stress in boar sperm in vitro. The addition of ZnGlu significantly reduced the content of MDA and DC in ejaculate samples in all experimental groups during incubation, compared to the control group. Moreover, the activity of superoxide dismutase and catalase increased after adding ZnGlu to the boar semen, especially at concentrations of 2.0 and 5.0 mg/ml (P &lt; 0.05). Conclusions. These data demonstrate that the supplementation of zinc glutamate enhances the antioxidant defence system of sperm and improves quality of boar semen in vitro.

Effect of L-proline on sperm quality during cryopreservation of boar semen

L-proline has been reported to be useful in semen cryopreservation. However, its use has rarely been reported in the freezing of boar semen. The objective of this study was to evaluate the effects of different concentrations of L-proline (0, 10, 30, 50, and 90 mM) on the quality of boar semen after freezing and thawing. Semen samples from boars (n = 6) were frozen using freezing extenders with added concentrations of L-proline. Total sperm motility, progressive motility, survival time at 37 °C, acrosome integrity, mitochondrial activity, DNA integrity, the content of the lipid peroxidation product malondialdehyde (MDA), total antioxidant capacity (T-AOC) and, expression levels of apoptosis protein (cleaved caspase 3 and Bax) were evaluated after thawing. The results showed that total sperm viability (73.96% vs. 63.58%) and progressive motility (56.88% vs. 47.26%) after thawing were significantly higher in the 10 mM L-proline treatment group than in the control group. The survival time at 37 °C and the total motility of sperm in the 10 mM group within one hour after thawing were significantly higher than in the control group. Acrosome integrity and mitochondrial activity of sperm in the 10 mM group were significantly higher than those in the control, 50 mM, and 90 mM groups. The DNA integrity rate in the 10 mM group was significantly higher than in the control group. The L-proline treatment did not affect sperm MDA content or T-AOC. The expression levels of apoptosis protein (cleaved caspase 3 and Bax) in the 10 mM L-proline supplemented group were lower than those in the control group. In conclusion, the freezing extender containing 10 mM L-proline improved semen quality after freezing and thawing and thus would be a useful reagent for boar semen cryopreservation.

Effect of certain extenders on semen quality of boars during preservation at 17°C

The present study was designed to determine the effect of three selected extenders on liquid preservation of boar semen at 17°C. Ejaculates (30) were collected from three boars (one ejaculate/boar/week) housed at University pig farm, Ludhiana using dummy sow. Only ejaculates having &gt;70% initial progressive motility were extended in Beltsville Thawing Solution (BTS), Safe Cell (SFC) and Tris-Egg Yolk (TEY) extenders in the ratio 1:4 and preserved in a BOD incubator at 17°C for 120 h. The semen was evaluated for different sperm attributes and lipid peroxidation (MDA) at 0, 24, 48, 72, 96 and 120 h of preservation. The mean percentage of sperm motility, viability,plasma membrane integrity and acrosome integrity were significantly higher in BTS as compared to SFC and TEY at different hours of storage period. At 120 h of preservation, highest percentage of in vitro capacitation/acrosome reaction was seen in semen stored in BTS followed by SFC and lowest in TEY after 6 h of incubation. Eventually, significantly lower levels of MDA were noticed in semen extended in BTS than in their contemporary extenders (SFC and TEY) at 96 and 120 h of storage period. The mean percentage of most sperm parameters decreased gradually from day of collection (0 h) up till 120 h in all the extenders. In conclusion, Beltsville Thawing Solution was better than Safe Cell and Tris-Egg Yolk extenders in liquid preservation of boar semen at 17oC owing to improved spermcharacteristics and reduced oxidative stress.

Effects of gelatin and oxytocin supplementation in a long-term semen extender on boar semen quality and fertility potential.

This study investigated the efficacy of different concentrations of gelatin supplementation in long-term semen extender on boar semen quality during storage for 10 days at 17°C. Additionally, oxytocin was added to stored semen to enhance fertility. In Experiment 1, boar semen was collected, diluted with gelatin at concentrations between 0% and 2.5% (w/v) and mixed with a semen extender. Then, it was kept in a refrigerator at 17°C and stored for 10 days. In Experiment 2, the sperm quality was examined after adding 0, 5, and 10 IU oxytocin per artificial insemination dose to the most effective semen extender from Experiment 1 and placing it in a refrigerator at 17°C for 10 days. In Experiment 3, the fertility potential in terms of non-return rate and litter size was determined using the most effective solid-stored semen supplemented with oxytocin. The results indicated that sperm quality decreased with increasing storage time (p<0.05). The sperm quality in terms of total motility, progressive motility, and viable sperm with intact acrosomes and high mitochondrial potential was the highest with 1.5% gelatin supplementation (p<0.001) on all days of storage. Treatment with oxytocin did not affect sperm quality (p>0.05). The non-return rate and litter size after insemination with semen supplemented with 1.5% gelatin and 10 IU of oxytocin after 8 to 10 days of storage were comparable to those of the control group (p>0.05). A semen extender as a solid medium supplemented with 1.5% gelatin successfully preserved boar semen for a long storage duration. Treatment with oxytocin did not affect sperm quality. In addition, the fertility capacity using 1.5% gelatin with 10 IU oxytocin and stored for 8 to 10 days was acceptable and comparable to that of short-term storage.