Wael Khalil | Mansoura University (original) (raw)

Papers by Wael Khalil

Journal of Animal and Poultry Production

The aim of this study was to evaluate the effect of adding zeolite mineral in semen extender on r... more The aim of this study was to evaluate the effect of adding zeolite mineral in semen extender on rabbit sperm cryopreservation. Ten healthy, fertile rabbit bucks were used, and the ejaculates were obtained using an artificial vagina. Semen of all bucks were pooled and diluted in a tris-yolk fructose (TYF) extender supplemented with zeolite at concentration of 0 (control) and 1% (w/v) for a final sperm concentration of 25 x 106 sperm cells/ ml. Diluted semen was packed in straws (0.25 ml) and stored in liquid nitrogen (-196 °C) for one month. After thawing, semen of each treatment was evaluated for sperm quality parameters, including sperm progressive motility, livability, morphological abnormalities and plasma membrane integrity. Apoptosis and sperm ultrastructure were also examined. Total antioxidant capacity and lipid peroxidation markers were determined in extender after thawing. Results showed that zeolite had a positive effect (P < 0.05) on sperm characteristics (progressive motility, livability and membrane integrity) after equilibration period and post-thawing as compared with the control. Percentage of viable sperm increased (P < 0.05), while percentages of early apoptotic, apoptotic and necrotic sperm cells decreased (P < 0.05) in treatment of zeolite as compared to control. In contrary, total antioxidants capacity in extender decreased (P < 0.05) and malondialdhyde and H 2 O 2 concentration increased (P < 0.05) in treatment of zeolite compared to control. In conclusion, addition of semen extender with zeolite improved post-thaw sperm quality of rabbit by enhancing sperm characteristics, reducing apoptosis and sperm damage occurring by cryopreservation.

Animals, 2021

This study evaluated the efficacy of Turraea fischeri leaf extract for maintaining the viability ... more This study evaluated the efficacy of Turraea fischeri leaf extract for maintaining the viability of cryopreserved goat sperm. Ejaculated semen was collected from 5 mature Baladi bucks (50–60 kg, 2–4 years of age) and those samples with mass motility ≥ 70% and sperm concentration ≥ 2.5 × 109/mL were selected, pooled, and divided into 4 aliquots. Each aliquot was diluted in Tris-citric-soybean lecithin extender containing a different concentration of T. fischeri leaf extract (0, 125, 250, or 375 µg/mL). Treated semen samples were cooled to 5 °C, transferred to 0.25-mL French straws, and stored in liquid nitrogen (LN2) at −196 °C. After thawing, membrane integrity was examined by transmission electron microscopy, apoptotic activity by Annexin/propidium iodide staining and flow cytometry, and both enzyme activities and antioxidant capacity by spectroscopic assays. The leaf extract at 375 µg/mL significantly improved semen quality as indicated by enhanced total antioxidant capacity, reduced H2O2 concentration, a greater proportion of structurally intact motile sperm, and concomitant reductions in apoptosis and necrosis. The extract also significantly increased the proportion of sperm with a contiguous plasma membrane and intact acrosome (p < 0.05). Furthermore, LC-MS revealed numerous secondary metabolites in the extract that may contribute to sperm cryopreservation

Cryobiology, 2021

The aim of the present study was to investigate the effect of supplementing rabbit semen extender... more The aim of the present study was to investigate the effect of supplementing rabbit semen extender with zeolite loaded with different charges (Z + or Z − , Z ±) on sperm cryopreservation. Semen was collected from six healthy, fertile New Zealand rabbit bucks using an artificial vagina. The collected ejaculates were pooled and diluted with a tris-yolk fructose (TYF) extender supplemented with Z ± (+16, +12, +8, − 16, − 12, and − 8) at a concentration of 1% for a final sperm concentration of 25 × 10 6 sperm cells/mL. The diluted semen samples were then cryopreserved in 0.25 mL straws and stored in liquid nitrogen for 1 month. To evaluate sperm quality, we examined sperm progressive motility, vitality, morphological abnormalities, and plasma membrane integrity. In addition, apoptotic rates were determined using flow cytometry and by examining sperm ultrastructure under a transmission electron microscope (TEM). Moreover, total antioxidant capacity and markers of lipid peroxidation were measured in the extender after thawing. Addition of Z ± had a positive effect on progressive motility, vitality, and membrane integrity after an equilibration period and post-thawing as compared with the controls (P < 0.05). Z ± supplementation, particularly with a strong negative charge, also decreased the percentages of apoptotic and necrotic sperm cells compared to controls (P < 0.05), as shown both by flow cytometry and TEM. This was not associated with any marked effects on the oxidative biomarkers in the extender. In conclusion, addition of Z ± to semen extender improved post-thawing sperm quality by improving sperm characteristics, decreasing apoptosis, and minimizing sperm damage during cryopreservation.

Tropical Animal Health and Production, 2021

This study determined the effects of antioxidant supplementation and storage time at cool tempera... more This study determined the effects of antioxidant supplementation and storage time at cool temperatures on the characteristics of epididymal camel spermatozoa. Camel testes were collected at the abattoir after animal slaughtering and kept at 4°C during transportation and until processing (max 6 h). Spermatozoa were retrieved and diluted with SHOTOR extender, split in aliquots, supplemented with the following antioxidants: 200 μm/mL vitamin E, 1.0 g/L vitamin C, 1 μg/mL selenium nanoparticles, 50 μg/mL zinc nanoparticles, 2 μg/mL sodium selenite, and 100 μg/mL zinc sulfate, and stored at 4°C for 2, 48, 96, and 144 h. The storage time significantly affected (P < 0.05) the sperms' motility and livability, the sperms' membrane integrity, and the percentages of cytoplasmic droplets as well as the percentage of morphologically normal spermatozoa. Epididymal sperm characteristics (progressive motility, livability, membrane integrity, and abnormalities) were significantly improved (P < 0.05) when the spermatozoa were diluted with antioxidants as compared with the control group, and the best additives were identified as nano-selenium, sodium selenite, nano-zinc, and zinc sulfate. In conclusion, adding nano-sized minerals or inorganic trace elements and vitamins maintained the progressive motility, livability, and membrane integrity, and decreased abnormalities and cytoplasmic droplet percentages of epididymal camel spermatozoa stored at 4°C up to 144 h.

The Splinter bean, Entada abyssinica, is widely used in folk medicine. In the current work, we pr... more The Splinter bean, Entada abyssinica, is widely used in folk medicine. In the current work, we profiled the secondary metabolites from E. abyssinica bark extract using LC-MS and investigated its effect on cryopreserved ram semen. Twenty-eight compounds, including tannins and gallic acid derivatives that prevailed in the extract, were tentatively identified. Results showed that the quality of the post-thawed semen showed a significant improvement when the extract was added to the extender at a concentration of 375 µg/mL. The progressive motility and plasma membrane integrity of sperm cells were significantly increased in the post-thawed semen; however, the total antioxidant capacity (TAC) was insignificantly increased. A significant decrease in the concentration of hydrogen peroxide was detected as well. No significant changes were observed in activities of lactate dehydrogenase (LDH), alanine aminotransaminase (ALT), and aspartate transaminase (AST) within the treated samples. Intact sperm percentage was significantly increased, while apoptotic and necrotic sperm percentages were reduced significantly. Molecular docking of some individual components from the extract revealed their potential to interfere with the apoptosis cascade in which Bcl-2 is involved. In conclusion, Entada abyssinica appears to be useful for cryopreservation presumably owing to its polyphenol content that has potent antioxidant capacity scavenging reactive oxygen species (ROS), enhancing the endogenous antioxidant system and inhibiting lipid peroxidation.

Reprod Dom Anim. , 2020

Increased reactive oxygen species (ROS) generation may disrupt the oocytes function and their com... more Increased reactive oxygen species (ROS) generation may disrupt the oocytes function and their competence. In this study, we introduced BTZO-1, a new supplement that can regulate the oxidative stress. Addition of BTZO-1 during IVM of bovine oocytes
improved their developmental competence in the term of improvement of blastocyst
rates. In addition, the quality of the produced embryos was improved by decreasing
the apoptosis level by showing a decreased number of TUNNEL positive cells.

his study was conducted to evaluate whether adding Moringa oleifera dry leaves powder (MODLP) as ... more his study was conducted to evaluate whether adding Moringa oleifera dry leaves powder (MODLP) as a natural anti-oxidants in the diet, has the potential to attenuate the negative effects of heat stress on growth performance, healthy status, anti-oxidant defense system and immunity of growing rabbits. Weaned NZW rabbits (n=120) were divided into four groups fed basal diet with 0, 100, 200 and 300 mg MODLP/kg. Growth performance parameters were recorded at age intervals from 5-13 wk. Caecal activity, carcass traits, hematological, serum biochemicals, antioxidant and immunity were determined at 13 wk of age. Results show that dietary addition of 200 mg MODLP/kg increased (P<0.05) pulse rate, growth performance parameters, net carcass weight, dressing rate and spleen percentages, beneficial bacteria (lactobacillus) count, hematological parameters, serum high-density lipoproteins, total antioxidant capacity, antibody titer and lysozyme. Abdominal fat percentage, E. coli count, and total cholesterol, triglycerides and low-density lipoproteins and thiobarbituric acid-reactive substances concentration reduced (P<0.05) 200 mg/kg group. The current study indicated that alleviation of adverse impacts due to heat stress on productivity, blood constituents, oxidative stress and immunity status, can be achieved in growing rabbits through dietary adding Moringa Oleifera dried leaves at a level of 200 mg/kg diet.

The current study aimed to evaluate the effect of dietary supplementation with sunflower oil (SFO... more The current study aimed to evaluate the effect of dietary supplementation with sunflower oil (SFO), flaxseed oil (FSO) or 1% SFO+1% FSO on growth performance, and lipid, testosterone and fatty acid profiles of California rabbit bucks. Number of 24 California bucks having 5.3-5.5 months of age and weighing 2316-2429 g were divided into four groups (n=6). Bucks in the control group were fed free commercial diet, while those in the 2 nd , 3 rd and 4 th groups were fed the same diet with 2% SFO, 2% FSO, or 1% SFO+1% FSO, respectively. Growth parameters were determined during the experiment (16 wk). Serum lipid, testosterone and fatty acid profiles were determined. Results showed insignificant treatment effect on body weight, feed intake and weight gain during the experiment (0-16 wk). The differences in serum triglycerides and total lipids between treatment groups and control were not significant, but triglycerides increased (P<0.05) in FSO-than SFO-bucks. Phospholipids decreased (P<0.05) only by SFO+FSO, while total cholesterol and LDL increased (P<0.05) by SFO or SFO+FSO, but HDL was not affected. All oil types increased (P<0.05) Non-HDL, being higher (P<0.05) in SFO+FSO than in SFO or FSO. Serum Castelli I and II indices, AST and ALT activity, serum total fatty acids and fatty acid profile were not affected by treatment. In conclusion, balancing the ratio between n-6: n-3 fatty acids is important to be the easiest approach to indicate this would be intake of oils rich in n-3 or n-6 fatty acids, such as sunflower and flaxseed oils, respectively.

There are various challenges during semen cryopreservation used for artificial insemination, oxid... more There are various challenges during semen cryopreservation used for artificial insemination, oxidative stress is considered as the major factor. Usage of nano-particle materials is an efficient tool to overcome these challenges. In the present article, effects of adding green synthesized gold nanoparticles (GSGNPs, 5; 10ppm/mL) to Tris-extender on sperm characteristics in post-equilibrated and post-thawed semen. Apoptosis and sperm ultrastructure post-thawed were also examined as well as enzyme activity and antioxidant status in post-thawed extender of goat semen were evaluated. Supplementation of GSGNPs (10 ppm) increased (P<0.05) motility, livability, and membrane integrity post-equilibration, and also in addition to acrosome integrity post-thawing. Also, viable sperm increased (P<0.05), early apoptotic, apoptotic, and necrotic sperm decreased (P<0.05) by 10 ppm GSGPNs. In the extender of thawed semen, AST and ALT activities were not affected, total antioxidant capacity and catalase activity were higher (P<0.05), while malondialdehyde decreased (P<0.05) by 10 ppm GSGNPs. Frequency distribution of sperm cells with intact plasma or acrosomal membranes increased (P<0.05), while of those with swollen plasma membranes or typical acrosome reaction decreased (P<0.05). However, frequency distribution of sperm cells with slightly swollen or lost plasma membranes as well as atypical acrosome reaction and lost acrosomal membrane was not affected by GSGNPs. Transmission electron microscopy examination revealed no entry of GSGNPs with sperm cells.The present article may suggest considerable addition of GSGNPs at a level of 10 ppm/mL to Tris-extender of cryopreserved goat semen during cryopreservation.

Simple Summary: This is a comprehensive study to compare between the effects of different supplem... more Simple Summary: This is a comprehensive study to compare between the effects of different supplements (vitamins C and E, trace elements Na 2 SeO 3 and ZnSO 4 , and nanoparticles of zinc oxide and selenium) to the semen extender of camel epididymal spermatozoa during cooling and freezing/thawing cryopreservation. Supplementation of the semen SHOTOR extender with zinc oxide and selenium nanoparticles lead to improved progressive motility, vitality, and anti-oxidative defense, and reduced the ultrastructural abnormalities in camel epididymal spermatozoa. Abstract: There are several obstacles in camel semen cryopreservation; such as increasing semen viscosity and the reduction in motile spermatozoa after ejaculation. Epididymal spermatozoa offer an efficient alternative to overcome these problems and are well-suited for artificial insemination in camels. In the current study, we compared the effects of supplementation with vitamin C, E, inorganic trace elements of selenium (Na 2 SeO 3) and zinc (ZnSO 4), and zinc and selenium nanoparticles (ZnONPs and SeNPs, respectively) on the cryopreservation of dromedary camel epididymal spermatozoa. When the SHOTOR extender was supplemented with ZnONPs and SeNPs; the sperm showed increased progressive motility; vitality; and membrane integrity after cooling at 5 • C for 2 h; when compared to the control and vitamin-supplemented groups. Moreover, the ZnONPs and SeNPs supplementation improved the progressive motility, vitality, sperm membrane integrity, ultrastructural morphology, and decreased apoptosis when frozen and thawed. SeNPs significantly increased reduced glutathione (GSH), superoxide dismutase (SOD), and decreased lipid peroxide malondialdehyde (MDA) levels. The advantageous effects of the trace elements were potentiated by reduction into a nano-sized particle, which could increase bioavailability and reduce the undesired liberation of toxic concentrations. We recommend the inclusion of SeNPs or ZnONPs to SHOTOR extenders to improve the cryotolerance of camel epididymal spermatozoa.

Objective: To test the effects of extenders and packaging methods on morphological and functional... more Objective: To test the effects of extenders and packaging methods on morphological and functional characteristics of frozen thawed Ossimi ram semen. Methods: Ram semen was pooled, diluted in 3 different extenders: Tris-egg yolk (TEY), Tris-soybean lecithin (TSBL), and Tris-butylatedhydroxytoluene (TBHT), equilibrated at 5 C for 4 h, and packaged in straws or pellets for freezing. Semen was evaluated for sperm progressive motility, viability, abnormality, and membrane integrity after dilution, equilibration and thawing. The percentages of viable, early apoptotic, apoptotic, and necrotic spermatozoa as well as comet assay parameters were determined in post-thawed semen. Total antioxidants capacity, malondialdehyde and lactic dehydrogenase were assayed in thawed seminal plasma. Results: After equilibration, only sperm membrane integrity was significantly higher (P<0.05) in TEY and TSBL than in TBHT. After thawing, TEY or TSBL in straws significantly improved sperm progressive motility and vitality (P<0.05). In thawed seminal plasma, TBHT or TSBL in straws and TBHT in pellets significantly reduced malondialdehyde, and TBHT in pellets significantly increased lactic dehydrogenase (P<0.05). TEY in straws increased viable sperm, while significantly decreased early apoptotic and apoptotic sperm (P<0.05). DNA damage was significantly decreased (P<0.05) in straws with TEY and TSBL, and tail moment decreased in straws with all extenders (P<0.05). Conclusions: Despite the disadvantages of dilution of cryopreserved semen with egg yolk, ram semen cryopreserved with TEY gives the best physical, morphological and functional characteristics in straws compared with pellets, followed by semen diluted with TSBL. However, semen diluted with TBHT or TSBL, regardless of packaging method, showed the highest impact on antioxidant status of cyopreserved ram semen.

A B S T R A C T Recent advances in nanotechnology have tremendously expanded its possible applica... more A B S T R A C T Recent advances in nanotechnology have tremendously expanded its possible applications in biomedicine. Although, the effects of nanoparticles (NPs) at cellular and tissue levels have not been fully understood, some of these biological effects might be employed in assisted reproduction to improve male fertility particularly by enhancing sperm cell quality either in vivo or in vitro. This review summarises the available literature regarding the potential applications of nanomaterials in farm animal reproduction, with a specific focus on the male gamete and on different strategies to improve breeding performances, transgenesis and targeted delivery of substances to a sperm cell. Antioxidant, antimicrobial properties and special surface binding ligand functiona-lization and their applications for sperm processing and cryopreservation have been reviewed. In addition, nanotoxicity and detrimental effects of NPs on sperm cells are also discussed due to the increasing concerns regarding the environmental impact of the expanding use of nanotechnologies on reproduction.

Twenty four adult Barki eweswere used to investigate the effect of addition of rumen protected am... more Twenty four adult Barki eweswere used to investigate the effect of addition of rumen protected amino acids, lysine or/and methionineon some wool characteristics. This study was carried out from September 2016 to February 2017. Animals were divided into four groups (6ewes/group). The first group (Control): fed only the control diet, second group (LYS): fed the control diet and rumen protected lysine (6g Lysi pearl /kg concentrate, 3g /animal/day), third group (MET): fed the control diet and rumen protected methionine (14g Smartamin/kg concentrate, 7g /animal/day) and fourth group (LYS+MET): fed the control diet and mixture of (3g LYS and 7g MET/animal/day) for six months. Some wool measurements were investigated such as fiber length, fiber diameter, staple strength and wool growth per 10 cm 2 and clean wool yield. Results showed that supplementing methionine caused significant (P<0.05) increase in fiber length, fiber diameter, staple strength and clean wool yield. Lysine increased fiber length significantly (P<0.05), but did not affect clean wool yield and caused small increase in both staple strength and fiber diameter. Supplementing of (mixture of methionine and lysine) increased significantly fiber length, wool yield and staple strength, but caused insignificant increase in clean wool production per 10cm 2 , but had no significant effect on fiber diameter.From these results, it can be concluded that addition of rumen protected methionine (7g Smartamin/animal/day) forsix monthsenhanced significantly wool characteristics of Barki ewes (fiber length, fiber diameter, staple strength and clean wool yield).

Aim of this study was to investigate performance and puberty of ram lambs produced from pregnant ... more Aim of this study was to investigate performance and puberty of ram lambs produced from pregnant ewes treated with different levels of arginine. This study was carried out at Sakha Animal Production Research Station, belonging to the Animal Production Research Institute in a cooperation with Animal Production Department, Faculty of Agriculture, Mansoura University. During November 2016, total of 45 Ossimi ewes (September mating season), 3-4 years of age and 39.59±0.15 kg live body weight (LBW) were divided into three groups (15 head/each). Ewes in the 1 st group (G1) served as a control, ewes in the 2 nd (G2) and 3 rd (G3) groups were weekly treated with oral dose of 20 and 30 mg arginine (AR), respectively, for the duration of the treatment from the second month of pregnancy up to lambing. After lambing, total of 8 ram lambs produced from each group were taken and allotted in three groups similar to treatment of their dams. LBW of ram lambs was recorded at birth and monthly up to 12 month of age. In blood plasma of ram lambs, concentration of IgG and IgM at 2 days and 1, 3 and 4 wk of age, while concentration of total proteins, albumin, creatinine and glucose, and activity of aspartate (AST) and alanine (ALT) aminotransferase at 2, 4, 6 and 8 month of age were determined. Age and plasma testosterone concentration (PTC) of each ram lambs were determined at three pubertal stages, while semen of 1 st ejaculates (at puberty) was evaluated. Results showed that ram lamb LBW was higher (P<0.05) only in G3 than G1 at birth and at 5 to 12 month of age. Final ram lamb LBW at 12 month of age was higher by 10.2 and 15.9% in G2 and G3 than in G1, respectively. Plasma concentration of Immunoglobulin G (IgG) and Immunoglobulin M (IgM) after 2 days, 1, 3, and 4 wk of lambing was highest (P<0.05) in G3, followed by G2, while, G1 showed the lowest values. Plasma concentration of total proteins at 4 month of age and of albumin at 4 and 6 month of age increased (P<0.05), while glucose decreased (P<0.05) at 2 and 8 month of age in G2 and G3 as compared to those of G1. Creatinine increased (P<0.05) only in G3 as compared to G2 and G1 at 6 and 8 month of age, while activity of AST and ALT was not affected at all ages. At puberty, age of ram lambs was earlier (P<0.05) by 51.4 and 33.0 days and PTC was higher (P<0.05) by 22.5 and 18.8% in G2 and G3 than in G1. Ram lambs in G3 showed the best (P<0.05) semen characteristics of the 1 st ejaculation (volume, sperm motility and live sperm output / ejaculate), followed by G2. PTC was higher (P<0.05) in G2 and G3 than in G1 at 6 and 8 month of age, showing significantly (P<0.05) an opposite trend at 7 month of age, while insignificantly different at 5, 9 and 10 month of age. In conclusion, ram lambs of ewes treated with weekly oral dose of L-arginine-HCL (30 mg / ewe) from the second month of pregnancy up to lambing achieved early age and higher weight at puberty.

This study aimed to evaluate the effect of alternative components of egg yolk in Tris extender an... more This study aimed to evaluate the effect of alternative components of egg yolk in Tris extender and semen packaging methods during cryopreservation on sperm characteristics and antioxidant system in seminal plasma of frozen-thawed semen of Finnish Landrace rams. Semen was collected from 5 sexually matured Finnish rams (50-70 kg LBW and 2-4 years old) by artificial vagina once weekly for 7 weeks. Only semen with mass motility of ≥70% was pooled and diluted with Tris-citric extender containing 15% egg yolk (Tris-EY) or 1% soybean lecithin (Tris-SBL) or 2 mM butylated hydroxytoluene (Tris-BHT). Semen was extended at a rate of 1:5 (semen/extender) with three extender types. After semen extension, semen was placed for cooling in the refrigerator (5 o C) for 4 hours as equilibration period and packaged in 0.25 ml French straws or 0.25 ml pellets in liquid nitrogen. Semen was thawed at 37°C for 30 s. Semen was evaluated after dilution, equilibration and thawing, for progressive motility, livability and abnormality of spermatozoa and curled tail spermatozoa responded to a solution of osmolarity of 75 mOsm for 30 min. The concentration of total antioxidants (TAC), malondialdehyde (MDA) and lactic dehydrogenase (LDH) activity in post-thawed seminal plasma were determined. Results showed that sperm characteristics, including percentages of progressive motility, livability, abnormality and curled tail in post-diluted, post-equilibrated or post-thawed semen were not affected significantly by the type of extender. Progressive motility and curled tail percentages in post-thawed semen were higher (P<0.001; P<0.05) in straws than in pellets. Livability and abnormality percentages were insignificantly better at straws than in pellets. The recovery rate of motility and livability was higher at straws than in pellets. All sperm characteristics indicated insignificant effect of interaction between type of extender and semen packaging method. TAC was higher (P<0.05), while MDA concentration was lower (P<0.05) in Tris-SBL and Tris-BHT than in Tris-EY. The activity of LDH was insignificantly the highest in Tris-BHT than in other extenders. The TAC, MDA concentration and LDH activity in post-thawed semen were not affected significantly by semen packaging method. Effects of interaction between type of extender and semen packaging method on each of TAC, MDA concentration and LDH activity were not significant. The current study concluded the successful usage of Tris-SBL or Tris-BHT in comparing with the possible disadvantages of using egg yolk in Tris-based extender of ram semen.

This study designed to examine effects of adding different concentrations of propolis ethanolic e... more This study designed to examine effects of adding different concentrations of propolis ethanolic extract (PEE) as antioxidant to ram semen extenders; tris egg yolk (TEY) or tris-soybean lecithin (TSBL) on sperm characteristics, lipid peroxidation and enzymatic activities in seminal plasma of ram semen preserved at 5°C for 48 hours. Semen was collected from five adult Rahmani rams using an artificial vagina twice/week for 5 weeks. Pooled semen was extended with TEY or TSBL supplemented with different concentrations from PEE (0, 0.1, 0.5 and 1.0 mg/ml). Polyphenols and flavonoid compounds in propolis ethanolic extract were identified. In each treatment, percentage of progressive motility, livability, and abnormality were determined. Also, percentages of sperm membrane integrity or chromatin damage were determined. Concentrations of total antioxidant (TAO) and malondialdehyde (MDA), and activity of acid phosphatase (ACP), lactate dehydrogenase (LDH), and asprtate (AST) and alanine (ALT) transaminase, were determined. Results showed that the most effective polyphenolic compounds in the prolopolis ethanolic extract was the chlorogenic acids, catechein, protocatchuic and pyrogallol (120.623, 53.006, 23.907 and 11.048 mg/100g extract, respectively). Luteo.6-arbinose 8-glucose was the predominant identified flavonoid component in propolis extract (5268.78ppm), followed by acacetin, hisperidin, apig.6-rhamnose8-glucose, apig.6-glucose8-rhamnose, kamp.3-(2-p-comaroyl) glucose and luteo.6-glucose 8-arbinose with concentrations (647.53, 561.09, 366.47, 150.46, 144.51 and 109.42 ppm, respectively. The effect of propolis ethanolic extract supplementation to ram semen extenders was significant (P<0.05) only on the overall percentages of sperm progressive motility and sperm chromatin damage while, sperm livability, abnormality and membrane integrity were not affected significantly (P<0.05). Concentration of TAC was numerically (P= 0.82) higher in TEY extender with different concentrations of PEE than TSBL. However, MDA was lower (P<0.05) in free TEY, followed by TSBL with PEE. Activity of LDH, AST and ALT tended to be lower in seminal plasma of semen extended with TSBL than TEY. These results showed beneficial effects of soybean lecithin and propolis ethanolic extract in tris based extender on progressive motility and chromatin integrity of spermatozoa up to 48 h of preservation at 5°C. Also, lipid peroxidation and enzyme activity were lower in TSBL than TEY. In addition, soybean lecithin can effectively replace egg yolk as a protective additive with propolis ethanolic extract for preservation extender without any detrimental effects on post-chilling semen quality in rams.

Semen cryopreservation is a well-established procedure used in veterinary assisted reproduction t... more Semen cryopreservation is a well-established procedure used in veterinary assisted reproduction technology applications. We investigated damaging effects of cryopreservation on the structural and ultra-structural characteristics of bull sperm induced at different temperatures and steps during standard cryopreservation procedure using transmission (TEM) and scanning electron microscopy. We also examined the effect of cryopreservation on sperm DNA and chromatin integrity. Five healthy, fertile Friesian bulls were used, and the ejaculates were obtained using an artificial vagina method. The semen samples were pooled and diluted in a tris-yolk fructose (TYF) for a final concentration of 80 Â 10 6 spermatozoa/ ml. The semen samples were packed in straws (0.25 ml), and stored in liquid nitrogen (À196°C). Samples were evaluated before dilution, just after dilution (at 37°C), at 2 h and 4 h during equilibration, and after thawing (37°C for 30 s in water bath). In association with step-wise decline in motility and viability , our results showed that the plasma membrane surrounding the sperm head was the most vulnerable structure to cryo-damage with various degrees of swelling, undulation, or loss affecting about 50% of the total sperm population after equilibration and freezing. Typical acrosome reaction was limited to 10% of the spermatozoa after freezing. We also observed increased number of mitochondria with distorted cristae (15%). Chromatin damage was significantly increased by cryopreservation as evident by TEM (9%). This was mainly due to DNA breaks as confirmed by Sperm Chromatin Structure Assay (SCSA) (8.4%) whereas the chromatin structure was less affected as evaluated microscopically by toluidine blue staining. We concluded that, using standard cryopreservation protocol, the most pronounced damage induced by cryopreservation is observed in the plasma membrane. Further improvement of cryopreser-vation protocols should thus be targeted at reducing plasma membrane damage. Acrosomal, mitochon-drial and chromatin damage are also evident but appear to be within acceptable limits as discussed.

A B S T R A C T Dietary polyunsaturated fatty acids (PUFAs) can influence fertility in farm anima... more A B S T R A C T Dietary polyunsaturated fatty acids (PUFAs) can influence fertility in farm animals. Some evidence in mice and sheep have suggested that PUFAs may influence offspring sex ratio, which may have significant value for cattle production. To test this hypothesis, three groups of Holstein cows were supplemented with either 0%, 3% or 5% protected fat (PF) in the form of calcium salt of fatty acids (rich in omega-6) from 14–21 days pre-partum until conception. Proven-fertile frozen semen from the same ejaculate was used for insemination. Calf sex recorded at birth was 8/19 (42.1%) male offspring in the control group, increasing to 14/20 (70%, P > 0.05) and 17/20 (85%, P < 0.05) in 3% and 5% PF, respectively. To test if this effect was caused by a direct influence on the oocyte, we supplemented bovine cumulus oocyte complexes during in vitro maturation with either omega-3 alpha-linolenic acid (ALA), omega-6 linoleic acid (LA) or trans-10, cis-12 conjugated linoleic acid (CLA). Sex ratio of the produced transferable embryos was determined using PCR of SRY gene. Similar to the in vivo results, sex ratio was skewed to the male side in the embryos derived from LA-and CLA-treated oocytes (79% and 71%) compared to control and ALA-treated oocytes (44% and 54%, respectively). These results indicate that both dietary and in vitro supplementation of omega-6 PUFAs can skew the sex ratio towards the male side in cattle. Further experiments are required to confirm this effect on a larger scale and to study the mechanisms of action that might be involved.

Albizia harveyi is a tropical deciduous tree, found across South and Eastern Africa and widely us... more Albizia harveyi is a tropical deciduous tree, found across South and Eastern Africa and widely used in traditional medicine. The leaf extract ameliorated the damaging effects of the frozen-thawing process in cryopreserved bull semen. In a dose-dependent pattern, sperm motility, viability, and membrane integrity were improved compared to the untreated control. Furthermore, the extract increased the percentage of viable sperm cells and reduced the percentages of early apoptotic and apoptotic sperm cells as well as the damage in sperm ultra-structure. These activities are in agreement with the robust antioxidant properties in vitro and in the seminal fluid as observed in the total antioxidant capacity and the lipid peroxidation parameter malondialdehyde. LC-MS yielded 35 compounds. The extract was dominated by quercetin-O-galloyl-hexoside and quercetin-O-pentoside, along with other flavonoid glycosides. The polyphenols are probably responsible for the observed activities. In conclusion, the current findings show that A. harveyi leaves are rich in bioactive polyphenols with functional properties, validating its traditional use.

Journal of Animal and Poultry Production

Animals, 2021

Cryobiology, 2021

Tropical Animal Health and Production, 2021

Reprod Dom Anim. , 2020