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This study aimed to examine how varying concentrations (0.5%, 1.0%, 2.0%, 3.0%, and 4.0%) of nano-L-a phosphatidylcholine (nano-L-a-PC) affected the quality of post-thawed buffalo bull semen compared to the same concentrations of L-a-phosphatidylcholine (L-a-PC), while 20% egg yolk (EY) was used as a control. Ejaculates (n = 72) were collected from fertile buffalo bulls (n = 12) for 6 weeks. The ejaculates were pooled and divided into 11 groups (3 aliquots per group) extended with 20% EY (Group 1 without supplement), 0.5%, 1.0%, 2.0%, 3.0%, and 4.0% nano-L-a-PC (Groups 2-6), and 0.5%, 1.0%, 2.0%, 3.0%, and 4.0% L-a-PC (Groups 7-11) in tris buffer. After cryopreservation procedures, semen samples were thawed at 37 °C and then evaluated for sperm kinematics, acrosomes, plasma membranes, and DNA integrities. The seminal plasma was analyzed for malondialdehyde (MDA), superoxide dismutase (SOD), glutathione reduced (GSH), and catalase (CAT). Results demonstrated that extended semen samples containing 1.0-3.0% nano-L-a-PC and 1.0-4.0% L-a-PC showed high total motility compared to control, whereas at 0.5-4.0% of nano-L-a-PC and L-a-PC showed high progressive motility. Significant (P< 0.05) wobble levels were seen at 0.5, 1.0, and 3.0% nano-L-a-PC, and distance average path and velocity average path at 2.0% L-a-PC. Acrosome and plasma membrane integrities were markedly elevated (P< 0.0001) at 2.0% nano-L-a-PC and 4.0% L-a-PC. High DNA integrity metrics were noticed at 0.5-1.0% nano-L-a-PC, as well as 3.0% and 4% L-a-PC. Low MDA and high (SOD, GSH, and CAT) levels were observed at 2.0% nano-L-a-PC and 4.0% L-a-PC. In conclusion, the optimum concentration of nano-L-a-PC that improved semen quality was 2.0%, which is roughly equivalent to the effect of 4.0% L-a-PC. [ABSTRACT FROM AUTHOR]

This study aimed to examine how varying concentrations (0.5%, 1.0%, 2.0%, 3.0%, and 4.0%) of nano-L-α phosphatidylcholine (nano-L-α-PC) affected the quality of post-thawed buffalo bull semen compared to the same concentrations of L-α-phosphatidylcholine (L-α-PC), while 20% egg yolk (EY) was used as a control. Ejaculates (n = 72) were collected from fertile buffalo bulls (n = 12) for 6 weeks. The ejaculates were pooled and divided into 11 groups (3 aliquots per group) extended with 20% EY (Group 1 without supplement), 0.5%, 1.0%, 2.0%, 3.0%, and 4.0% nano-L-α-PC (Groups 2–6), and 0.5%, 1.0%, 2.0%, 3.0%, and 4.0% L-α-PC (Groups 7–11) in tris buffer. After cryopreservation procedures, semen samples were thawed at 37 °C and then evaluated for sperm kinematics, acrosomes, plasma membranes, and DNA integrities. The seminal plasma was analyzed for malondialdehyde (MDA), superoxide dismutase (SOD), glutathione reduced (GSH), and catalase (CAT). Results demonstrated that extended semen samples containing 1.0-3.0% nano-L-α-PC and 1.0-4.0% L-α-PC showed high total motility compared to control, whereas at 0.5–4.0% of nano-L-α-PC and L-α-PC showed high progressive motility. Significant (P< 0.05) wobble levels were seen at 0.5, 1.0, and 3.0% nano-L-α-PC, and distance average path and velocity average path at 2.0% L-α-PC. Acrosome and plasma membrane integrities were markedly elevated (P< 0.0001) at 2.0% nano-L-α-PC and 4.0% L-α-PC. High DNA integrity metrics were noticed at 0.5–1.0% nano-L-α-PC, as well as 3.0% and 4% L-α-PC. Low MDA and high (SOD, GSH, and CAT) levels were observed at 2.0% nano-L-α-PC and 4.0% L-α-PC. In conclusion, the optimum concentration of nano-L-α-PC that improved semen quality was 2.0%, which is roughly equivalent to the effect of 4.0% L-α-PC.

Simple Summary: Cryopreserved semen, widely utilized in the artificial insemination of domestic animals, encounters challenges due to oxidative damage. This study investigated the effects of the antioxidant PQQ on the semen quality of Holstein bulls during cryopreservation and elucidated the associated mechanisms. Our findings revealed that PQQ significantly enhanced sperm motility, membrane integrity, acrosome integrity, and ATP levels, while simultaneously reducing MDA and ROS levels. Additionally, increased PGAM2, CAPZB, CAT, SOD1, and GPX1 protein expressions were also observed in the PQQ-treated group. Based on these results, PQQ not only improved semen quality but also mitigated oxidative stress, thereby enhancing the efficacy of sperm cryopreservation in bulls. 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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Regular verification of the quality of cryopreserved semen derived from native cattle is one of the tasks performed at the bank as recommended by the Food and Agriculture Organization. The purpose of the present study was to evaluate the quality of semen from PR bulls stored for 40, 50 and 60 years in the BMB using standard evaluation parameters such as sperm motility as well as structural-functional parameters such as plasma membrane integrity, transmembrane mitochondrial potential and sperm chromatin damage. Semen pellets from 27 PR bulls (3 ejaculates/bull) were tested. The data were analysed by one-way and two-way ANOVA, and the significance of the difference (P≤ 0.01) between the means was determined using Duncan's test. Our study results revealed that the long-term storage of semen had no effect on sperm characteristics after thawing. However, statistically significant differences (P≤0.01) in sperm plasma membrane integrity and transmembrane mitochondrial potential, following storage in liquid nitrogen were noted between bulls at all time points. However, there were no significant differences (P>0.01) in sperm chromatin damage between breeds or between different storage times, and the degree of DNA fragmentation ranged from 0.4 to 0.8%. [ABSTRACT FROM AUTHOR]

The present study was designed to determine the effect of different concentrations of quail egg yolk (5%, 10%, 15% and 20%) in tris based semen extender on progressive motility of Sahiwal bull spermatozoa at pre-freeze as well as post-thaw stage. The progressive motility significantly (P<0.01) improved in 10% and 15% quail egg yolk concentration at prefreeze stage whereas it increases significantly in 10% quail egg yolk concentration at post-thaw stage. The findings of the present study suggested that quail egg yolk improves post-thaw sperm progressive motility at 10% concentration. [ABSTRACT FROM AUTHOR]

The present investigation was undertaken on cattle bull semen reared at Frozen Semen Bull Station, Karimnagar, Telangana. The study was carried out on four cattle breeds (3 to 5 years of age) viz., Gir, Ongole, Jersey and Crossbred Jersey (CBJY). Semen was collected twice weekly from each bull using artificial vagina technique with a total of 32 ejaculates evaluated under study. The semen was diluted with Egg Yolk Tris Citrate (EYTC) extender and was evaluated for physical characteristics of neat semen, motility parameters and sperm viability percentage. The investigation revealed that Gir bull semen had significantly higher (p<0.05) ejaculate volume, mass motility, individual motility, sperm concentration compared to other breeds in present study. Whereas, Jersey bull semen had significantly higher (p<0.05) sperm viability percentage. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

The goal of current investigation was to ascertain the impact of carnitine as an additive in tris-egg yolk-based extender on Hariana bull semen (32 ejaculates) chosen for cryopreservation. Progressive motility, viability, HOS response, and acrosomal integrity were estimated after equilibration (pre-freeze) and after thawing. The activity of enzymes malondialdehyde (MDA), glutathione-S-transferase (GST), and superoxide dismutase (SOD) were quantified in post-thaw seminal plasma. Semen ejaculates from Hariana bulls (n=4) were diluted in tris egg yolk glycerol (TEYG) and were divided into four equal groups. Aliquot/Group I was processed without carnitine and served as control, while Groups II, III and IV received carnitine at concentration of 2.5 mM, 5 mM and 10 mM per 80x106 spermatozoa, respectively. Carnitine at 5 mM concentration significantly (p<0.05) enhanced sperm viability, acrosomal integrity, and intact plasma membranes both at pre-freeze and post-thaw stage. The addition of 5 mM and 2.5 mM carnitine reduced oxidative stress considerably (p<0.05). The results of this investigation revealed that carnitine has cryoprotective properties on bull spermatozoa via reducing oxidative stress and free radicals. Hence to improve post-thaw sperm functional dynamics, it was suggested that 5 mM of carnitine be supplemented as an additive to semen extender. [ABSTRACT FROM AUTHOR]

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.

It is postulated that alterations in membranous lipids construction, surface characteristics, fluidity, Ca2+ permeability, and cholesterol levels collectively contribute to the process of capacitation induction. The change in sperm membrane calcium permeability following capacitation is the main signal for the acrosome reaction. The acrosome response is commonly induced in vitro using calcium ionophore. The study's objectives are to determine how the calcium ionophore (ionomycin) and the solvent dimethyl sulfoxide (DMSO) affect the motility, hyperactivity, and velocity of bovine frozen–thawed spermatozoa at varying concentrations and incubation times. The capacitation medium consisted of Bracket–Oliphant medium. Four capacitation media were prepared; two treated groups were supplemented with 25 nM and 50 nM of ionomycin, one group was without ionomycin, and the control group was supplemented with DMSO. Sperm motility decreased substantially with increasing time period in all four groups, according to the findings. The result found that there was some variance in the sperm parameters between the groups and treatment times. Overall, the group exposed to 25 nM had better semen and sperm parameters than the group exposed to 50 nM. In conclusion, the role of calcium ionophore had better semen and sperm parameters in the group exposed with a lower concentration than a higher concentration on sperm.

Semen samples contain bacteria originating from the animal urogenital tract, environment, and/or contamination during semen processing, negatively affecting sperm quality by producing toxins and/or competing for nutrients in extenders. The aims of this study were to evaluate two methods of Single-layer centrifuges (SLC), high and low density colloid, as a method for bacterial removal from bull semen, and to evaluate sperm quality after treatment. In total, semen samples from 20 bulls (3 ejaculates per bull) were used in this study. Bacterial reduction was evaluated by bacterial quantification (colony forming unit - CFU/mL) while bacterial identification was performed by matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) after culturing bacteria on blood agar. Sperm motility parameters were evaluated by Computer Assisted Sperm Analyses (CASA), and sperm chromatin structure assay (SCSA) by Flow cytometry. Both, High and Low density SLC reduced number of bacteria significantly (p < 0.001) compared with control. The difference in bacterial count between High and Low SLC was also significant (p < 0.001). Furthermore, High density SLC was successful in removing almost all Bacillus and Proteus spp. Most CASA parameters were significantly improved after both treatments (p < 0.001, p < 0.01, p < 0.05). The Deoxyribonucleic acid (DNA) fragmentation index evaluated by SCSA in High (p < 0.01) and Low (p < 0.05) SLC group differed significantly compared with control. Single-layer centrifugation (SLC) with either a high or a low density colloid is a suitable method for bacterial removal in bull semen. [ABSTRACT FROM AUTHOR]

The aim of the present study was to evaluate the effects of insulin supplemented extender on bull semen at a post-thaw stage. Semen samples were collected four times from a Zavot bull which is endangered species in Türkiye. Semen samples were diluted with Tris based extender supplemented with 10 IU or 15 IU of insulin and control. Motility and plasma membrane integrity were evaluated by a phase-contrast microscope. Acrosome integrity (Fitc-Peanut Agglutinin), mitochondrial membrane potential (Rhodamine 123), nitric oxide level (4,5-Diaminofluorescein-2/diacetate) were evaluated with dual staining propidium iodide using flow cytometry. Motility and plasma membrane functional integrity were better preserved compared to control group (P0.05), but increased insulin negatively affected acrosome integrity (p0.05). There were not any significant differences among groups in terms of nitric oxide level (P>0.05). In conclusion, it was thought that insulin involved in energy metabolism and improved preservation of bull semen.

Simple Summary: As the basis of artificial insemination technology, semen cryopreservation technology is a very valuable technology in the cattle industry and directly affects the success rate of fertilization. The effectiveness of semen cryopreservation is closely associated with quality alterations of fresh semen and semen diluent during semen cryopreservation. Therefore, in this study, in order to improve the effect of semen cryopreservation and fresh semen quality of breeding bulls, enough antioxidant-efficient grape seed proanthocyanidins were added to semen diluent and breeder bull diets. It was found that grape seed proanthocyanidins as a semen diluent extender could improve frozen–thawed sperm morphology and quality (including frozen–thawed sperm motility, membrane integrity, acrosome integrity, and mitochondrial activity). As a dietary additive, it could improve sperm motility, reduce sperm deformity rate, and improve the quality of fresh semen. This study revealed that grape seed proanthocyanidins could improve effect of semen cryopreservation and the reproductive performance of breeding bulls, which was significantly related with the antioxidant properties of grape seed proanthocyanidins. This research promotes the preservation and breeding of bovine semen and provides a basis for the fundamental science of mammalian gametes and reproductive biotechnology. Oxidative stress leads to a decrease in semen quality during semen cryopreservation and fresh semen production. Grape seed proanthocyanidins (GSPs) are endowed with well-recognized antioxidant, anti-inflammatory, anti-cancer, and anti-aging activities. Therefore, the objective of this experiment was to explore the effects of GSPs on the quality of fresh and cryopreserved semen to provide a basis for GSPs as a new dietary additive and semen diluent additive for males' reproduction. Fresh semen from three healthy bulls aged 3 to 5 years old were gathered and mixed with semen diluents dissolved with 0 µg/mL, 30 µg/mL, 40 µg/mL, 50 µg/mL, and 60 µg/mL GSPs respectively. The motility, physiological structures (acrosome integrity, membrane integrity, mitochondrial activity), and antioxidant capacity of frozen–thawed sperm were measured after storage in liquid nitrogen for 7 days (d). Bulls were fed with 20 mg/kg body weight (BW) GSPs in their diet for 60 days; the weight of the bull is about 600 kg. Then, the reproductive performance and antioxidant indexes of bulls were measured before and after feeding. The results demonstrated that GSPs supplementation significantly increased sperm motility, physiological structures, GSH-Px, and CAT enzyme activities and significantly decreased MDA content in sperm during semen cryopreservation. The optimal concentration of GSPs was 40 µg/mL (p < 0.05). After 20 mg/kg (body weight) GSP supplementation, sperm motility was significantly heightened (p < 0.05), the sperm deformity rate was significantly reduced (p < 0.05), and antioxidant enzyme activities (such as SOD, CAT, and GSH-Px) were significantly enhanced (p < 0.05), and the production of MDA was significantly suppressed (p < 0.05) in serum compared with that before feeding. In conclusion, these results reveal that a certain concentration of GSPs has a good protective effect on sperm damage caused by semen cryopreservation and the reproductive performance reduction caused by stress in bulls, which may be attributed to the antioxidant function of GSPs. In summary, GSPs are a useful cryoprotective adjuvant and dietary additive for bull sperm quality. [ABSTRACT FROM AUTHOR]

The aim of the present study was to evaluate the effects of insulin supplemented extender on bull semen at a post-thaw stage. Semen samples were collected four times from a Zavot bull which is endangered species in Türkiye. Semen samples were diluted with Tris based extender supplemented with 10 IU or 15 IU of insulin and control. Motility and plasma membrane integrity were evaluated by a phase-contrast microscope. Acrosome integrity (Fitc-Peanut Agglutinin), mitochondrial membrane potential (Rhodamine 123), nitric oxide level (4,5-Diaminofluorescein-2/diacetate) were evaluated with dual staining propidium iodide using flow cytometry. Motility and plasma membrane functional integrity were better preserved compared to control group (P<0.05). Acrosome integrity results were statistically similar between control and 10 IU insulin groups (P>0.05), but increased insulin negatively affected acrosome integrity (p<0.05). Mitochondrial membrane potential was found to be higher when compared all cells in insulin containing groups compared to control (P<0.05). However, results were found similar in all groups with PI negative/mitochondrial membrane potential positive (P>0.05). There were not any significant differences among groups in terms of nitric oxide level (P>0.05). In conclusion, it was thought that insulin involved in energy metabolism and improved preservation of bull semen. [ABSTRACT FROM AUTHOR]

Sperm survives for a very short time in fresh semen, and slow cooling to 5.00 °C kills a large number of sperms. This study was aimed to compare the semen quality parameters and anti-oxidant levels in four extenders (manual, Triladyl, Steridyl and AndroMed). Semen samples were obtained from a total number of 12 dual-purpose Simmental bulls kept in the Simmental Cattle Breeding Center for a period of 3 months using an artificial vagina. Sperm viability, motility, abnormal morphology, plasma membrane integrity, DNA damage, chromatin quality, total antioxidant capacity (TAC) and lipid peroxidation were evaluated. The highest progressive motility, viability, plasma membrane integrity, and TAC and the lowest levels of malondi-aldehyde in the frozen-thawed semen belonged to the semen group frozen with Triladyl. Parameters of motility were higher in the frozen-thawed semen with Triladyl than in other groups, indicating a significant difference from the manual extender. Among the extenders studied, Triladyl was the most suitable for semen freezing in Simmental bulls. [ABSTRACT FROM AUTHOR]

Introduction: Flaxseed oil (FO) and vitamin E (VE) both have antioxidant effects on sperm. The present study investigated the effects of dietary supplementation with FO and/or VE on semen quality. Methods: 16 fertile Simmental bulls were selected and randomly divided into 4 groups (n = 4): the control group (control diet), FO group (control diet containing 24 g/kg FO), VE group (control diet containing 150 mg/kg VE) and FOVE group (control diet containing 150 mg/kg VE and 24 g/kg FO), and the trial lasted 10 weeks. Results: The results showed that the addition of FO independently can increase sperm motion parameters, the levels of catalase (CAT), glutathione peroxidase (GSH-Px), testosterone (T) and estradiol (E2), while reduce oxidative stress in seminal plasma (P < 0.05). Supplement of VE independently can increased the motility, motility parameters, CAT and superoxide dismutase (SOD) levels, and reduce oxidative stress in seminal plasma (P < 0.05). There was an interaction effect of FO × VE on motility and reactive oxygen species (ROS), while GSH-Px and ROS were affected by week × VE 2-way interaction, levels of T and E2 were also affected by the dietary FO × week interaction (P < 0.05). The triple interaction effects of FO, VE and week were significant for malondialdehyde (MDA) (P < 0.05). Compared with the control group, sperm from the FOVE group had a significantly higher in vitro fertilization (IVF) rate, and subsequent embryos had increased developmental ability with reduced ROS levels at the eight-cell stage, then increased adenosine triphosphate (ATP) content and gene expression levels of CAT, CDX2, Nanog, and SOD at the blastocyst stage (P < 0.05). Metabolomic and transcriptomic results indicated that dietary supplementation of FO and VE increased the expression of the metabolite aconitic acid, as well as the expression of ABAT and AHDHA genes. Conclusion: With in-silico analysis, it can be concluded that the effects of dietary FO and VE on improving semen quality and embryo development may be related to increased aconitic acid via the ABAT and AHDHA genes involved in the propionic acid metabolism pathway. [ABSTRACT FROM AUTHOR]

Technologies of artificial insemination of cattle are actively used in livestock farms in Russia. At the same time, quality control of semen production does not involve testing the bull semen for microorganisms of the genus Mycoplasma. The active use of molecular genetic methods allows for the rapid determination and species identification of pathogenic mycoplasmas; however, it does not indicate the viability of the identified pathogen. The paper aims to study mycoplasma contamination of semen products presented on the Russian market and determine the conformity of the results of the Polymerase Chain Reaction [PCR] method and the traditionally used microbiological method. The paper presents the results of the first Russian study of the occurrence and viability of pathogenic species Mycoplasma bovis, M. bovigenitalium, M. californicum, and Ureaplasma diversum in semen production. We investigated 447 samples of cryopreserved semen of bulls-producers of various breeds from Russian breeding centers and breeding farms of the USA, Great Britain, and the Netherlands, supplying these products to the Russian market. The analysis shows a high frequency of occurrence in the bull semen of DNA of microorganisms of the genus Mycoplasma (up to 70.9%). Besides, M. bovigenitalium appeared to be the most frequently detected type of mycoplasma from those tested. Cases of identifying several types of mycoplasmas in one sample were revealed: in 58.8% of the studied samples of semen products from Russian farms and 15% of semen samples from international breeding centers. The viability of mycoplasmas that were positive according to PCR analysis when testing 65 samples was demonstrated using the microbiological method for 94% of samples. The correspondence of the results of microbiological and molecular genetic detection of mycoplasmas and ureaplasmas was 88.3%. The results obtained indicate the prevalence of mycoplasmas in semen production and the need to improve the control system of breeding material in order to prevent the spread of mycoplasma infections of cattle using artificial insemination technologies.

Low fertility rates in lactating dairy cows as well as restricted availability of semen doses of young bulls with high genetic merit are two major problems in the reproduction of dairy cows. By using liquid semen (LS), the number of doses per ejaculate can be increased. One of the challenges of optimizing the reproductive performance of dairy cows is the phenomenon of variable estrus lengths. The objective of this study was to determine whether the use of LS affects pregnancy outcome of dairy cows with delayed ovulation, when compared with frozen semen (FS). A randomized controlled clinical trial was implemented. In a split-sample procedure, 131 ejaculates were processed into LS (Caprogen, LIC, New Zealand) and FS (BioXcell, IMV, France). Both semen types of each ejaculate were inseminated under the same field conditions to cows showing natural or induced heat. Cows and semen type were allocated according to the last digit of the cows identification number (even = frozen semen, odd = liquid semen). Inseminations (n = 667) were conducted after localization of the pre-ovulatory follicle. Determination of ovulation was performed 24 h post AI per transrectal ultrasonographic examination. Ovulations were classified as delayed when the pre-ovulatory follicle was still present at ovulation control. The prevalence of delayed ovulations was 25.2%. Data of 667 inseminations were analyzed with a generalized linear mixed model including semen type (P = 0.016), parity (P = 0.014), backfat thickness (P = 0.006), estrus induction (P = 0.010), ovulation (P = 0.265) and the interaction term 'semen type by ovulation' (P = 0.094). Overall, a higher pregnancy per AI (P/AI) of LS (45.4%) than P/AI of FS (33.7%) was found. In cases of delayed ovulations, use of LS resulted in higher P/AI (46.8%) compared with FS (27.7%; P = 0.017). We concluded that the fertilizing capacity of LS in prolonged intervals from AI to ovulation might be greater when compared with FS and could be an efficient tool to improve fertility of lactating dairy cows with delayed ovulations. • Delayed ovulations occurred in a quarter of all cases. • Liquid semen showed significant increase of pregnancy per insemination. • a backfat thickness of 10–13 mm at time of insemination yielded highest pregnancy outcome. [ABSTRACT FROM AUTHOR]

The rapid emergence of antibacterial resistance requires alternatives to antibiotics to be found, including for semen preservation. One of the possible alternatives would be to use plant-based substances with known antimicrobial effects. The objective of this study was to test the antimicrobial effect of pomegranate powder, ginger, and curcumin extract in two concentrations on bull semen microbiota after exposure for <2 h and 24 h. An additional aim was to evaluate the effect of these substances on sperm quality parameters. The bacterial count in semen was low from the beginning; however, a reduction was present for all tested substances compared with control. A reduction in bacterial count in control samples was also observed with time. Curcumin at a concentration of 5%, reduced bacterial count by 32% and was the only substance that had a slight positive effect on sperm kinematics. The other substances were associated with a decline in sperm kinematics and viability. Neither concentration of curcumin had a deleterious effect on sperm viability parameters measured by flow cytometry. The results of this study indicate that curcumin extract at a concentration of 5% can reduce the bacterial count and does not have a negative influence on bull sperm quality. [ABSTRACT FROM AUTHOR]

Simple Summary: We evaluated the impact of imported semen of Holstein bulls from different countries on their daughters' economic traits using the Lifetime Net Income (LNI) index in different climates of Iran. The first lactation records were collected from 10 big dairy farms in various regions of Iran. Studied traits included milk, fat and protein yields, calving age and calving interval. There were significant differences between sire groups for studied traits in each climate (p < 0.01). In semi-cold, moderate and warm climates, daughters of French sires had the highest least-square means of LNI index. In cold climate, daughters of German sires had the highest least-square means. The aim of the present study was to evaluate the effects of imported semen of Holstein bulls from different countries on the economic traits of their daughters using the Lifetime Net Income (LNI) index in various climates of Iran. The data included the first lactation records of 274,057 Holstein cows collected during 1993 to 2017 by the Animal Breeding Center of Iran from 10 large dairy farms located in various provinces of Iran. The investigated traits included milk, fat and protein yields, calving age and calving interval. Breeding values of progenies were predicted by the Best Linear Unbiased Prediction (BLUP) method under the multi-trait animal model using DMU software. The genetic-economic merit of the progenies was estimated by the LNI index. There were significant differences between the estimated breeding values (EBVs) of sire groups (based on bull semen origin) for milk, fat and protein yields, calving age and calving interval in each climate (p < 0.01). The obtained results showed that the highest least-square means of LNI index in semi-cold, moderate and warm climates belonged to the daughters of French sires; however, daughters of German sires were estimated to have the highest least-square means in the cold climate. [ABSTRACT FROM AUTHOR]

This study aimed to determine the probable protective eff ect of gallic acid (GA) on the spermatological parameters of frozen‐thawed bull semen. Ejaculates were collected from four Holstein bulls. The mixed ejaculate was divided into four equal groups as control (0 μg/mL GA), 50, 100 and 200 μg/mL GA with a Tris-based extender. All extended groups were cooled, equilibrated into a +4°C cold cabinet and loaded into straws. The straws were then frozen and stored in a liquid nitrogen container (-196°C). Subsequently, samples were thawed in a water bath for analyzing motility and kinematic parameters, morphological integrity, DNA damage and biochemical alterations. GA50 (28.76±0.51%) and GA100 (29.32±0.31%) had improved progressive motility in comparison to the other groups (P

ABSTRACT Background Wide variation in fertility rates is observed when using frozen bull semen, even when the bulls have met quality standards for semen production. Therefore, a simple and reliable test to assess the freezing potential of bull semen based on the analysis of fresh semen or blood would be of great value. Attention is now turning to assessment of seminal plasma components such as proteins and elements. In the present study, the concentrations of macro- and microelements in fresh bull semen plasma and in serum and their correlation with quality characteristics of fresh semen and with semen quality after freezing and thawing were determined. Ejaculates were collected from 30 mature bulls, and semen volume, concentration, sperm motility, morphology, tail membrane integrity, plasma membrane permeability and DNA fragmentation were determined on the day of collection and after freezing and thawing. The concentrations of macroelements (Na, Mg, K and Ca) and microelements (Cu, Fe, Zn and Se) were determined in the seminal plasma and serum. The semen samples were classified into satisfactory and unsatisfactory groups according to the fresh semen quality. Results Zinc and Se levels measured in serum were associated with almost all fresh and frozen-thawed semen quality characteristics, while Fe levels were associated only with acrosomal defects in fresh semen. Zinc and Fe levels in fresh seminal plasma were associated with various quality characteristics of fresh and frozen-thawed semen, while Se level in fresh seminal plasma was not associated with any of the semen quality characteristics. Conclusions Microelements were shown to be useful as biomarkers involved in the analysis of bull sperm quality and could be used as an additional tool to predict bull semen quality after freezing and thawing. Our results confirm that the analysis of Zn and Se levels in serum and Zn, Cu and Fe levels in fresh seminal plasma can provide information to discriminate between bull semen samples with spermatozoa with high or low cryotolerance.

Schmallenberg orthobunyavirus (SBV) was initially detected in 2011 in Germany from dairy cattle with fever and decreased milk yield. The virus infection is now established in many parts of the world with recurrent epidemics. SBV is transmitted through midges and transplacental. No direct virus transmission including via breeding has ever been demonstrated. In some bulls, however, the virus is detectable transiently, in low to minute quantities, in semen post‐infection. While the infection is considered of low impact for the dairy industry, some SBV‐free countries have adopted a zero‐risk approach requiring bull semen batches to be tested for SBV RNA residues prior to import. This, in turn, obligates a protocol to enable sensitive detection of SBV RNA in semen samples for export purposes. Here, we describe how we established a now ISO/IEC 17025 accredited protocol that can effectively detect minute quantities of SBV RNA in semen and also its application to monitor bull semen during two outbreaks in the United Kingdom in 2012 and 2016. The data demonstrate that only a small number of bulls temporarily shed low amounts of SBV. [ABSTRACT FROM AUTHOR]

Distinguishing between bull Y- and X-bearing sperm populations is advantageous for techniques using sexed bull semen. The aim of this study was to produce a single-chain fragment variable (scFv) antibody against plasma membrane epitopes on bull Y-bearing sperm. Variable heavy (VH)- and variable light (VL)-region genes generated from a hybridoma cell secreting a specific Y-bearing sperm monoclonal antibody (mAb-1F9) were cloned and expressed. The expected sizes of the DNA bands were ∼350 bp for the VH gene and ∼318 bp for the VL gene. The VH and VL genes were generated and used to construct an scFv gene (∼650 bp), which was expressed in E.coli TG1 cells and produced the corresponding soluble scFv antibody. Compared with the parent mAb-1F9, the scFv antibodies presented a high affinity for Y-bearing sperm and low cross-reactivity with X-bearing sperm. An immunofluorescence analysis confirmed that the scFv antibodies and mAb-1F9 recognize epitopes on the Y-bearing sperm surface. The fluorescence signal was strong on the plasma membrane of Y-bearing sperm but very weak for X-bearing sperm. This study aids the application and production of engineered scFv antibodies specific to Y-bearing sperm to distinguish between Y- and X-bearing sperm populations for techniques involving sexed bull semen. [ABSTRACT FROM AUTHOR]

Simple Summary: Female calves are favored for milk production and genetic advancement in the dairy industry, and sex selection by using sexed semen has been long considered. A potential alternative sperm sexing technique is magnetic-activated cell sorting combined with an immunological method that uses scFv antibodies against male-specific sites on Y-chromosome-bearing sperm; however, the technique should be evaluated for validity and accuracy. This study focuses on how well bovine sperm are separated by the use of magnetic microbeads coupled with scFv antibodies against Y-chromosome-bearing sperm (PY-microbeads). The results showed that sexed bovine sperm using PY-microbeads was a highly effective technique for distinguishing X- and Y-chromosome-bearing sperm. It had no negative impact on the quality of X-chromosome-bearing sperm. The technique produced 82.65% of X-chromosome sperm in the X-enriched fraction semen and 81.43% of Y-chromosome sperm in the Y-enriched fraction semen, which was utilized to generate target sexed bovine semen. Sperm sexing technique is favored in the dairy industry. This research focuses on the efficiency of bovine sperm sexing using magnetic-activated cell sorting (MACS) by scFv antibody against Y-chromosome-bearing sperm (Y-scFv) coupled to magnetic microbeads and its effects on kinematic variables, sperm quality, and X/Y-sperm ratio. In this study, the optimal concentration of Y-scFv antibody coupling to the surface of magnetic microbeads was 2–4 mg/mL. PY-microbeads revealed significantly enriched Y-chromosome-bearing sperm (Y-sperm) in the eluted fraction (78.01–81.43%) and X-chromosome-bearing sperm (X-sperm) in the supernatant fraction (79.04–82.65%). The quality of frozen–thawed sexed sperm was analyzed by CASA and imaging flow cytometer, which showed that PY-microbeads did not have a negative effect on X-sperm motility, viability, or acrosome integrity. However, sexed Y-sperm had significantly decreased motility and viability. The X/Y-sperm ratio was determined using an imaging flow cytometer and real-time PCR. PY-microbeads produced sperm with up to 82.65% X-sperm in the X-enriched fraction and up to 81.43% Y-sperm in the Y-enriched fraction. Bovine sperm sexing by PY-microbeads showed high efficiency in separating Y-sperm from X-sperm and acceptable sperm quality. This initial technique is feasible for bovine sperm sexing, which increases the number of heifers in dairy herds while lowering production expenses. [ABSTRACT FROM AUTHOR]

To determine the effects of epigallocatechin‐3‐gallate (EGCG) on the cryopreservation of bovine semen, epigallocatechin‐3‐gallate dissolved with double distilled water to 0.2, 0.4, 0.6 and 0.8 mg/ml were added to the cryopreservation diluent of the bull semen. Then, we used computer‐assisted analysis of semen kinematic parameters, staining method to detect membrane function, acrosome integrity, enzyme‐linked immunosorbent assay to detect catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GSH‐Px), malondialdehydes (MDA) and reactive oxygen levels. The results showed that adding 0.6 mg/L of epigallocatechin‐3‐gallate could improve the cryopreserved sperm quality, which significantly increased the total motility, distance average path, distance straight line, distance curved line, average path velocity, curvilinear‐velocity, straight‐line velocity, amplitude of lateral head displacement and beat/cross frequency, as well as sperm CAT, GSH‐Px and SOD levels (p < 0.05), whilst reducing the reactive oxygen species and MDA levels (p < 0.05). Hence, these results indicate that the addition of 0.6 mg/ml of EGCG has a protective effect on the cryopreservation of the bovine semen. [ABSTRACT FROM AUTHOR]

In this study, the cryoprotective potential of natural antioxidant curcumin in Hariana bull semen was evaluated as an additive in a tris‐based extender with the assessment of motility and motion parameters of spermatozoa, membrane intactness, progesterone‐receptor binding, protein carbonyl content, cervical mucus penetration, cryocapacitation‐associated and apoptotic‐like changes. The collected ejaculates were divided into five groups in the tris‐based extender (control without curcumin‐I, 10 µM‐II, 25 µM‐III, 50 µM‐IV and 75µM‐V) and were cryopreserved. Groups II and III containing 10 and 25 µM curcumin substantially (p <.05) improved the post‐thaw sperm parameters like viability, motility, and velocity parameters; intact acrosome and membrane; lowered protein carbonyl content; DNA fragmentation and cryocapacitation‐associated changes in comparison to control. It was interesting to note that early apoptotic‐like changes in sperm cells were significantly (p <.05) decreased in Group II along with an increase in a higher population of sperm cells having high mitochondrial transmembrane potential. Higher progesterone‐receptor binding, Vanguard distance and in vitro capacitation response were observed only in Group II (10µM) compared to other groups. In conclusion, curcumin in a semen extender manifests cryoprotective effects and may be incorporated at 10 µM concentration in a Hariana bull semen extender for better post‐thaw sperm quality. [ABSTRACT FROM AUTHOR]

The aim of this study was to investigate whether vaccenic acid (VA) added to TRIS semen extender has an effect on freezing bull semen. The semen was collected at intervals of three days through an artificial vagina then pooled. The pooled ejaculates were divided into five components as the control (C; no supplement) and VA (25, 50, 100, and 200 µg / mL) groups then extended with the TRIS-based medium. The samples were left in a cold air cabinet for cooling process (4 °C), then they were equilibrated for 4 h. Afterward, the samples in the groups were frozen by using a programmable freezer. The greatest progressive and total motility results were obtained in the VA25 group (p < 0.001). Besides, the total abnormality and chromatin damage was determined to be on the lowest level in the VA25 group (p < 0.05). Except for VA200, the glutathione level was greater in the treatment groups in comparison to the control group (p < 0.05), while the lowest malondialdehyde was determined in VA25. In conclusion, VA supplemented with TRIS-based extender at the concentration of 25 μg / mL improves the progressive and total motility properties of spermatozoa, reduces the abnormal spermatozoa rate, and has a positive effect on chromatin integrity. [ABSTRACT FROM AUTHOR]

Reports on the use of 16S sequencing for the identification of bacteria in healthy animals are lacking. Bacterial contamination of bull semen can have a negative effect on the sperm quality. The aims of this study were threefold: to identify bacteria in the semen of healthy bulls using 16S sequencing; to investigate the differences in the bacterial community between individual bulls; and to establish if there was a relationship between the bacteria isolated and bull fertility. Semen from 18 bulls of known fertility was used for the DNA extraction and 16S sequencing; 107 bacterial genera were identified. The differences in the amplicon sequence variants (ASVs) and the numbers of genera between bulls were noted. Negative correlations (p < 0.05) between several bacterial genera with Curvibacter, Rikenellaceae RC9-gut-group and Dyella spp. were seen. Other negatively correlated bacteria were Cutibacterium, Ruminococcaceae UCG-005, Ruminococcaceae UCG-010 and Staphylococcus, all within the top 20 genera. Two genera, W5053 and Lawsonella, were enriched in bulls of low fertility; this is the first time that these bacteria have been reported in bull semen samples. The majority of the bacteria were environmental organisms or were species originating from the mucous membranes of animals and humans. The results of this study indicate that differences in the seminal microbiota of healthy bulls occur and might be correlated with fertility. [ABSTRACT FROM AUTHOR]

A seasonal effect on sperm quality parameters was observed previously. Although identification of the bull semen microbiota by 16S rRNA sequencing was performed previously, it has not been carried out in commercial semen samples from different seasons, and its connection with sperm quality parameters has not been evaluated yet. The objectives in this study were; (i) to evaluate diversity of bull semen microbiota and sperm quality parameters in different seasons, and (ii) to find if specific bacteria were associated with seasonal differences in specific sperm quality parameters. Bull semen microbiota was identified in 54 commercial bull semen samples from 3 seasons (winter, spring, summer). Sperm quality was analysed by Computer Assisted Sperm Analyses (CASA) and Flow Cytometry (FC). From 28 phyla in all samples, six phyla were identified in samples from all seasons, with observed seasonal differences in their distribution. At genus level, 388 genera were identified, of which 22 genera had a relative abundance over 1 % and showed seasonal differences in bacterial diversity, and 9 bacteria genera were present in all seasons. Differences between spring and summer (P < 0.05) were observed for live hydrogen peroxide positive sperm cells. A trend towards significance (0.10 > P > 0.05) was observed for some CASA kinematics (VCL and LIN) and FC parameters (High respiratory activity, and live hydrogen peroxide positive sperm cells) between seasons. Nevertheless, associations between sperm quality parameters and specific bacteria were observed in spring. • Seasonal difference in the relative abundance of the top six phyla were present. • The number of genera and their amount in different seasons vary. • Only small differences in sperm quality between seasons were detected. • Genera were correlated with hydrogen peroxide (H 2 0 2) positive sperm cells in spring. • Genus Cutibacterium was negatively correlated with H 2 0 2 positive sperm cells. [ABSTRACT FROM AUTHOR]

Copyright of Kafkas Universitesi Veteriner Fakultesi Dergisi is the property of University of Kafkas, Faculty of Veterinary Medicine and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Eurasian Journal of Veterinary Sciences is the property of Eurasian Journal of Veterinary Sciences and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Background: Wide variation in fertility rates is observed when using frozen bull semen, even when the bulls have met quality standards for semen production. Therefore, a simple and reliable test to assess the freezing potential of bull semen based on the analysis of fresh semen or blood would be of great value. Attention is now turning to assessment of seminal plasma components such as proteins and elements. In the present study, the concentrations of macro- and microelements in fresh bull semen plasma and in serum and their correlation with quality characteristics of fresh semen and with semen quality after freezing and thawing were determined. Ejaculates were collected from 30 mature bulls, and semen volume, concentration, sperm motility, morphology, tail membrane integrity, plasma membrane permeability and DNA fragmentation were determined on the day of collection and after freezing and thawing. The concentrations of macroelements (Na, Mg, K and Ca) and microelements (Cu, Fe, Zn and Se) were determined in the seminal plasma and serum. The semen samples were classified into satisfactory and unsatisfactory groups according to the fresh semen quality. Results: Zinc and Se levels measured in serum were associated with almost all fresh and frozen-thawed semen quality characteristics, while Fe levels were associated only with acrosomal defects in fresh semen. Zinc and Fe levels in fresh seminal plasma were associated with various quality characteristics of fresh and frozen-thawed semen, while Se level in fresh seminal plasma was not associated with any of the semen quality characteristics. Conclusions: Microelements were shown to be useful as biomarkers involved in the analysis of bull sperm quality and could be used as an additional tool to predict bull semen quality after freezing and thawing. Our results confirm that the analysis of Zn and Se levels in serum and Zn, Cu and Fe levels in fresh seminal plasma can provide information to discriminate between bull semen samples with spermatozoa with high or low cryotolerance. [ABSTRACT FROM AUTHOR]

The aim of this study was to evaluate the effect of 7-dehydrocholesterol loaded cyclodextrin (7-DHCLC) and cholesterol loaded cyclodextrin (CLC) with the addition of different glycerol (G) rates on bull semen cryopreservation. Semen samples of three bulls were pooled and divided into seven groups; control group (C) and the other six were extended with standard Tris extender including different G rates (3, 6 and 9%) with CLC or 7-DHCLC to reach 1.5 mg/120x106. Extended samples were frozen in 0.25 mL straws. After thawing, motion characteristics and motility, viability, acrosome integrity, mitochondrial activation, functional membrane integrity and abnormal spermatozoa rates were assessed. The highest membrane integrity, viability and total motility were detected in 7-DHCLC with 9% of G group (P0.05). According to these results, 7-DHCLC with 9% G addition maintained some spermatological parameters better than other groups after thawing in bull semen thus, it was concluded that 7-DHCLC could be used instead of CLC in bull semen cryopreservation.

444 samples of bull frozen semen used for artificial insemination from local and foreign breeding centers were investigated by PCR for the presence of DNA of Coxiella burnetii. C. burnetii DNA was detected in 32 semen samples from US and was not found in semen straws from local breeding centers and breeding centers of Netherlands and Great Britain. According to the results, it is evident that despite the restrictions and controls on the international movement of semen, bull semen is still a potential source of pathogen transmission.

Several domestic and wildlife species have been shown to possess antibacterial defenses in their ejaculate most probably in order to increase the fertilization success and protect against sexually transmitted pathogens. However, very little is known about the consequences and factors influencing the differences within and between species as far as ejaculate-associated immunity. In the present study, we have analyzed bacterial killing activity (BKA) against Escherichia (E.) coli and Staphylococcus (S.) aureus as well as lysozyme concentrations (LC) in seminal plasma from 60 Fleckvieh bulls. Further, sperm quality and its association with BKA and LC were determined. Twenty percent of the individuals displayed BKA against both bacteria, 78.3% against S. aureus only and 1.7% of the bulls did not indicate any BKA in seminal plasma. No bulls with seminal plasma BKA only against E. coli were identified; implying that 80.0% of the tested bulls had no ejaculate associated defense mechanisms against this gram-negative bacterial species in place. This is in striking contrast to results of Pietrain boars within our previous study, in which 42.8% of the 119 boars expressed an antibacterial activity against E. coli in seminal plasma, 10.9% amongst them with BKA against E. coli only. LC was higher in the bull group with BKA against both bacteria (1.2 ± 0.6 μg/mL) compared to the group with BKA against S. aureus only (0.7 ± 0.3 μg/mL), but – if calculated over all individuals – LC in bulls (0.8 ± 0.4 μg/mL) was lower compared to boars (2.4 ± 1.2 μg/mL). LC showed positive correlations to the age of the bulls and sperm quality as well as a negative relation to bacterial load in raw semen although the highest bacterial contamination was found in animals with seminal plasma BKA against both strains. We discuss the obtained results with regards to possible differences within the microbiome of female and male genital tracts and the reproductive strategies (vaginal vs. uterine depositors) in these two livestock species. Besides identifying the responsible molecules, future phylogenetically controlled comparative studies are needed for a better understanding of the evolution of species differences in ejaculate-associated antibacterial defenses. • Sixty bull ejaculates were analyzed for bacterial killing activity and lysozyme. • Four-fifths of the samples had no ejaculate-associated defense against E. coli. • Lysozyme concentration in seminal plasma correlated with age of bull. • Lysozyme concentration in seminal plasma correlated with bull sperm quality. • Reproductive strategies & microbiome explain BKA differences between bull & boar ejaculate. [ABSTRACT FROM AUTHOR]

This study aimed to evaluate the effect of sodium caseinate added into freezing extender on the sperm parameters of cryopreserved bull semen and in vitro and in vivo fertility. One ejaculate of 30 bulls was used and processed using Botu-Bov (Botupharma, Botucatu, Brazil) with the addition of 20% egg yolk (EY) or 15% egg yolk with 2% sodium caseinate (EY + SC), subsequently submitted to freezing. Semen from both groups were evaluated immediately after thawing (T0) and after thermic stress at 37 °C for 90 min (T90), for sperm kinetics, by CASA method, and plasma membrane integrity (PMI), superoxide (O 2 −) concentration and high mitochondrial potential (HMP) by flow cytometry. In vitro fertilization (IVF) was performed to assess embryo cleavage rate on day 3, and blastocyst rate on day 8. The in vivo fertility test was performed using fixed-time artificial insemination (FTAI). In sperm evaluation, trajectory velocity, linear velocity, curvilinear velocity, and lateral head movement were higher (P < 0.05) in EY + SC at T0. At T90, while rectilinearity and linearity did not differ between EY and EY + SC (P > 0.05), the other parameters evaluated were higher in EY + SC. Similarly, the integrity of the plasma and acrosomal membranes (iPAM) was higher (P < 0.05) at T90 in EY + SC, but did not differ (P > 0.05) between the groups at T0. For O 2 − and HMP, the values were lower (P < 0.05) in EY + SC group in both moments; furthermore, EY + SC showed higher cleavage and blastocyst rates in IVF. Likewise, pregnancy rates by FTAI were higher (P < 0.05) in the EY + SC group. In conclusion, the addition of sodium caseinate into freezing extender improves sperm parameters of frozen-thawed bull semen and fertility rates on during in vitro and in vivo tests. • Sperm parameters of bull frozen-thawed semen after were improved by the addition of sodium caseinate into the freezing semen extender. • The diluent containing 2% sodium caseinate increased embryos producing rate and pregnancy by IATF. • Semen diluted with sodium caseinate presented better quality after heat stress. [ABSTRACT FROM AUTHOR]

Contents: Breed and sire differences in sperm cryosurvival have been noted, with negative implications for sperm cryobanking and assisted reproduction programmes. This study hypothesized that these differences could be modified by using lower molecular weight cryoprotectants. Therefore, the effect of replacing glycerol (GLY) with ethylene glycol (EG) on differential cryosurvival of semen from two Sanga cattle breeds (Mashona vs. Tuli) was determined. Three to five ejaculates were collected from each of ten bulls (3‐8 years) by electro‐ejaculation, diluted in three Tris‐egg yolk extenders (Triladyl®, 7% GLY‐based and 7% EG‐based) and evaluated for sperm motility, viability and morphology at three time periods (fresh – 0 hr, pre‐freeze – 4 hr and post‐thaw). Tuli bulls produced larger (11.8 ± 0.31 ml vs. 8.5 ± 0.38 ml) and more concentrated ejaculates of lower fresh semen quality. Breeds differed across time for motility and morphology, but not viability. Mashona bull semen had significantly higher motility and normal morphology values at each sampling time. Bulls classified as poor freezers had lower concentration (0.70 ± 0.09 × 109 sperm/ml vs. 1.37 ± 0.10 × 109 sperm/ml), sperm motility index (SMI, 35.0 ± 3.4 % vs. 67.8 ± 2.1 %) and viability (69.7 ± 1.1 % vs. 75.7 ± 1.0 %) compared to good freezers. Maintenance of semen quality by GLY and EG did not differ between breeds, poor and good freezers, or age groups. The interaction breed by extender across time did not reach statistical significance for all variables. The study revealed that bull and breed variation in sperm quality and cryosurvival is not modified by replacing GLY with EG, suggesting that cryostress tolerance of sperm may be under control of mechanisms other than differential response to GLY cytotoxicity. [ABSTRACT FROM AUTHOR]

Improving livestock reproductive performances is a high priority of worldwide breeders. The knowledge on cows accurate estrous detection associated with a good semen bull quality and artificial insemination appropriate timing, contribute in improving animal production yield. According to some studies, it is possible to achieve good quality semen from bulls by applying methods such the selection of bull presenting good conformation and health, followed by the collection and analysis of their semen, to be insured that this contains enough well fitted spermatozoids. For better success of artificial insemination, it is important to have a good mastery of the technique. The semen should also be protected against any agent of destruction and degeneration of spermatozoa, such as oxidizing processes. The present review gathers some technics of collecting and analyzing semen bull as well as some types of semen extender supplementation with antioxidant substances. [ABSTRACT FROM AUTHOR]

This study was designed to evaluate the effects of encapsulated reduced glutathione added to extender on the quality of bull sperm after freezing and thawing process. Extender contains lecithin nanoliposomes supplemented with different concenfrations of glutathione (0, 1, 2.5 and 5 Mm concenfration). The supplemented nanoliposomes with glutathione were prepared by thin-film preparation method. The particle size decreased to nanometer by sonication. 40 ejaculations were collected from 6 Holstein bulls and frozen for 6 weeks. Evaluated characteristics after freezing and thawing process were kinetic parameters (CASA), membrane activity (HOST), membrane integrity (eosinnigrosin) and morphology of sperm (Hancock's solution). Results showed that level 1 mM significantly increased the total and progressive motility (47.5±1.7, لح 32.72%). Although, there was no significant difference with level 2.5 mM glutathione (1.7 لح 45.02, لح 26.2%). Also, the concenfration of 2.5 mM showed the highest value of cell membrane integrity and cell membrane fimctionality but did not differ with the concenfration of ImM. However, supplementation of nanoliposomes with 5 mM glutathione reduced the quality of post-thaw sperm. Thus, level 1 M encapsulated glutathione can provide better protection compare to the other level of that for bull sperm. [ABSTRACT FROM AUTHOR]

It is postulated that alterations in membranous lipids construction, surface characteristics, fluidity, Ca2+ permeability, and cholesterol levels collectively contribute to the process of capacitation induction. The change in sperm membrane calcium permeability following capacitation is the main signal for the acrosome reaction. The acrosome response is commonly induced in vitro using calcium ionophore. The study's objectives are to determine how the calcium ionophore (ionomycin) and the solvent dimethyl sulfoxide (DMSO) affect the motility, hyperactivity, and velocity of bovine frozen–thawed spermatozoa at varying concentrations and incubation times. The capacitation medium consisted of Bracket–Oliphant medium. Four capacitation media were prepared; two treated groups were supplemented with 25 nM and 50 nM of ionomycin, one group was without ionomycin, and the control group was supplemented with DMSO. Sperm motility decreased substantially with increasing time period in all four groups, according to the findings. The result found that there was some variance in the sperm parameters between the groups and treatment times. Overall, the group exposed to 25 nM had better semen and sperm parameters than the group exposed to 50 nM. In conclusion, the role of calcium ionophore had better semen and sperm parameters in the group exposed with a lower concentration than a higher concentration on sperm. [ABSTRACT FROM AUTHOR]

Reports on the use of 16S sequencing for the identification of bacteria in healthy animals are lacking. Bacterial contamination of bull semen can have a negative effect on the sperm quality. The aims of this study were threefold: to identify bacteria in the semen of healthy bulls using 16S sequencing; to investigate the differences in the bacterial community between individual bulls; and to establish if there was a relationship between the bacteria isolated and bull fertility. Semen from 18 bulls of known fertility was used for the DNA extraction and 16S sequencing; 107 bacterial genera were identified. The differences in the amplicon sequence variants (ASVs) and the numbers of genera between bulls were noted. Negative correlations (p < 0.05) between several bacterial genera with Curvibacter, Rikenellaceae RC9-gut-group and Dyella spp. were seen. Other negatively correlated bacteria were Cutibacterium, Ruminococcaceae UCG-005, Ruminococcaceae UCG-010 and Staphylococcus, all within the top 20 genera. Two genera, W5053 and Lawsonella, were enriched in bulls of low fertility; this is the first time that these bacteria have been reported in bull semen samples. The majority of the bacteria were environmental organisms or were species originating from the mucous membranes of animals and humans. The results of this study indicate that differences in the seminal microbiota of healthy bulls occur and might be correlated with fertility.

Bu çalışma sperma sulandırıcısına katılan balık yağının Bali boğa spermatozoonlarının çözdürme sonrası kalitesi üzerine etkisini araştırmak amacıyla yapıldı. Süt tozu-yumurta sarısı sulandırıcısına 0 (kontrol), 50, 100, 150 ve 200 mg/100 mL dozunda balık yağı ilave edildi. 100 mg/100 mL balık yağı ilavesi kontrol grubu ile karşılaştırıldığında hem canlı sperm oranını hem de motiliteyi önemli derecede (P

Aim: The objective of this study was to evaluate the effects of different concentrations of soybean lecithin (SL) and virgin coconut oil (VCO) in Tris-based extender on chilled and frozen-thawed bull semen quality parameters. Materials and Methods: A total of 24 ejaculates were collected from four bulls via an electroejaculator. Semen samples were diluted with 2% VCO in Tris-based extender which consists of various concentrations of SL (1, 1.25, 1.5, and 1.75%). A 20% egg yolk in Tris used as a positive control (C+). The diluted semen samples were divided into two fractions; one for chilling which were stored at 4°C for 24, 72, and 144 h before evaluated for semen quality parameters. The second fraction used for freezing was chilled for 3 h at 4°C, packed into 0.25 mL straws and then cryopreserved in liquid nitrogen. The samples were then evaluated after 7 and 14 days. Chilled and frozen semen samples were thawed at 37°C and assessed for general motility using computer-assisted semen analysis, viability, acrosome integrity and morphology (eosin-nigrosin stain), membrane integrity, and lipid peroxidation using thiobarbituric acid reaction test. Results: The results showed that all the quality parameters assessed were significantly (p

Many microorganisms from various sources may be present in ejaculates of bulls. This study identified and isolated bacteria from bull sperm samples in a commercial stud and evaluated their resistance to antibiotics. The number of colony‐forming units was determined in semen samples collected at distinct steps during freezing and thawing. The minimum inhibitory concentration and the minimum bactericidal concentration were determined for four antibiotics commonly used in commercial studs. A total of 135 microorganisms from 25 genera were isolated. After a sensitivity test, all evaluated microorganisms (n = 55) were resistant to penicillin and most of them were resistant to tylosin and lincomycin (n = 54). Resistance to all tested antibiotics was observed in 22% of all isolates, whereas only 3.9% of the isolates were inhibited by the tested antibiotics at the concentrations recommended by the international legislation. As the isolated microorganisms presented high resistance to frequently used antibiotics, sensitivity tests should be periodically conducted in commercial bull semen studs to prevent the use of contaminated semen in artificial insemination. [ABSTRACT FROM AUTHOR]

This study was designed to evaluate the effects of Treduced glutathione added to extender on the quality of bull sperm after freezing and thawing process. Extender contains lecithin nanoliposomes supplemented with different concentrations of glutathione (0, 1, 2.5 and 5 Mm concentration). The supplemented nanoliposomes with glutathione were prepared by thin-film preparation method. The particle size decreased to nanometer by sonication. 40 ejaculations were collected from 6 Holstein bulls and frozen for 6 weeks. Evaluated characteristics after freezing and thawing process were kinetic parameters (CASA), membrane activity (HOST), membrane integrity (eosinnigrosin) and morphology of sperm (Hancock's solution). Results showed that level 1 mM significantly increased the total and progressive motility (47.5±1.7, 32.7±2 %). Although, there was no significant difference with level 2.5 mM glutathione (45.0±1.7, 26.2±2 %). Also, the concentration of 2.5 mM showed the highest value of cell membrane integrity and cell membrane functionality but did not differ with the concentration of 1mM. However, supplementation of nanoliposomes with 5 mM glutathione reduced the quality of post-thaw sperm. Thus, level 1 M encapsulated glutathione can provide better protection compare to the other level of that for bull sperm. [ABSTRACT FROM AUTHOR]

Aim: The present study was designed to evaluate enrofloxacin (EN), a fluoroquinolone broad-spectrum antibacterial drug for use in cryopreservation of Zebu bull semen. Materials and Methods: Semen was collected from three Zebu bulls of the Sahiwal breed (Bos indicus) for 3 weeks (replicates) for each experiment. Experiment I: semen samples were cultured for isolation of bacteria and in vitro sensitivity tests against streptomycin, penicillin, and EN. Proteus spp., Pseudomonas aeruginosa, Staphylococcus aureus, and Escherichia coli were isolated. P. aeruginosa was found to be resistant to streptomycin and penicillin (SP), while Proteus spp. was resistant to streptomycin. EN effectively inhibited the growth of all four bacteria in an in vitro sensitivity test. Experiment II: in vitro dose toxicity of EN (0–2000 μg/mL) for bull sperm was assessed in 2.9% sodium citrate buffer. EN up to a concentration of 1200 μg/mL was found to be safe for motility and viability of bull spermatozoa. Experiment III: semen was cryopreserved in a tris-citric acid extender containing either SP or EN (400 μg/mL) or without antibiotics as the control. Results: Post-thaw sperm motility was found to be higher in the extender containing EN and SP compared to control. Nevertheless, sperm viability was recorded higher (p < 0.05) with EN compared to SP and control. Sperm chromatin integrity did not differ (p > 0.05) in EN, SP, and controls. The total aerobic bacterial count was recorded as zero in the extender containing EN compared to SP and control. The fertility rate was higher (p < 0.05) with 400 μg/mL EN (59%) compared to SP (44%). Conclusions: It is concluded that EN is capable of controlling the bacterial load in frozen Zebu bull semen, resulting in a higher fertility rate under field conditions. [ABSTRACT FROM AUTHOR]