Type-1 cannabinoid receptors reduce membrane fluidity of capacitated boar sperm by impairing their activation by bicarbonate.

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Barbara Barboni

Full Text Available BACKGROUND: Mammalian spermatozoa acquire their full fertilizing ability (so called capacitation within the female genital tract, where they are progressively exposed to inverse gradients of inhibiting and stimulating molecules. METHODOLOGY/PRINCIPAL FINDINGS: In the present research, the effect on this process of anandamide, an endocannabinoid that can either activate or inhibit cannabinoid receptors depending on its concentration, and bicarbonate, an oviductal activatory molecule, was assessed, in order to study the role exerted by the type 1 cannabinoid receptor (CB1R in the process of lipid membrane remodeling crucial to complete capacitation. To this aim, boar sperm were incubated in vitro under capacitating conditions (stimulated by bicarbonate in the presence or in the absence of methanandamide (Met-AEA, a non-hydrolysable analogue of anandamide. The CB1R involvement was studied by using the specific inhibitor (SR141716 or mimicking its activation by adding a permeable cAMP analogue (8Br-cAMP. By an immunocytochemistry approach it was shown that the Met-AEA inhibits the bicarbonate-dependent translocation of CB1R from the post-equatorial to equatorial region of sperm head. In addition it was found that Met-AEA is able to prevent the bicarbonate-induced increase in membrane disorder and the cholesterol extraction, both preliminary to capacitation, acting through a CB1R-cAMP mediated pathway, as indicated by MC540 and filipin staining, EPR spectroscopy and biochemical analysis on whole membranes (CB1R activity and on membrane enriched fraction (C/P content and anisotropy. CONCLUSIONS/SIGNIFICANCE: Altogether, these data demonstrate that the endocannabinoid system strongly inhibits the process of sperm capacitation, acting as membrane stabilizing agent, thus increasing the basic knowledge on capacitation-related signaling and potentially opening new perspectives in diagnostics and therapeutics of male infertility.

Bovine serum albumin and skim-milk improve boar sperm motility by enhancing energy metabolism and protein modifications during liquid storage at 17 °C.

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Fu, Jieli; Li, Yuhua; Wang, Lirui; Zhen, Linqing; Yang, Qiangzhen; Li, Peifei; Li, Xinhong

2017-10-15

Both bovine serum albumin (BSA) and skim-milk have been reported to improve sperm quality, primarily by enhancing sperm motility, but the underlying molecular mechanism remains unknown. In this study, boar semen samples were collected and diluted with Androstar ® Plus extender containing different concentrations (0, 2, 4 g/l) of BSA and skim-milk. On days 0, 3, 5 and 7, the sperm motility parameters were determined using computer-assisted sperm analysis (CASA), and the ATP concentrations, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) activity and mitochondrial membrane potential were evaluated using commercial kits. The levels of protein phosphorylation, acylation and ubiquitination were analyzed by western blot. The results showed that supplementation with BSA and skim-milk provided higher sperm motility parameters, ATP levels, GAPDH activity and mitochondrial membrane potential than the control group (P levels of protein phosphorylation, acetylation and succinylation of the spermatozoa in the treated groups were dramatically higher than those in the control group (P level had a decreasing trend, the change in ubiquitination modification was not significantly different between the control group and treated groups. Moreover, the changes in protein modifications between the BSA treated group and skim-milk treated group were not distinctly dissimilar. Taken together, these results suggest that BSA and skim-milk had a positive role in the regulation of boar sperm motility by influencing sperm protein modifications changes as well as increasing the GAPDH activity, mitochondrial membrane potential, and intracellular ATP content. This research provides novel insights into the molecular mechanisms underlying BSA and skim-milk protective effects on boar sperm in the male reproductive system and suggests the feasibility of using skim-milk instead of BSA as a boar semen extender supplement. Copyright © 2017 Elsevier Inc. All rights reserved.

The cryoprotective effect of trehalose supplementation on boar spermatozoa quality.

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Hu, Jian-Hong; Li, Qing-Wang; Li, Gang; Jiang, Zhong-Liang; Bu, Shu-hai; Yang, Hai; Wang, Li-Qiang

2009-05-01

In order to improve boar sperm quality during frozen-thawed process, the influence of the presence of trehalose on success of cryopreservation of boar sperm were investigated. We evaluated freeze-thawing tolerance of boar spermatozoa in a base cooling extender with the addition of different trehalose concentrations (0, 25, 50, 100 and 200mmol/l), and tried to determine the optimum concentration of trehalose. We chose sperm motility, acrosome integrity, membrane integrity and cryocapacitation as parameters to evaluate cryopreservation capacity of boar spermatozoa. We obtained the best results for 100mmol/l trehalose-supplemented extenders, with values of 49.89% for motility, 66.52% for acrosome integrity and 44.61% for membrane integrity, while freeze-thawing tolerance was diminished significantly for 200mmol/l of trehalose. Before and after capacitation, the CTC score for semen diluted by extender containing 100mmol/l trehalose was 3.68% and 43.82%, respectively. In conclusion, trehalose could confer a greater cryoprotective capacity to boar spermatozoa. Trehalose-supplementation with 100mmol/l concentration in basic extender could significantly improve sperm motility, membrane integrity and acrosome integrity parameters, and reduce boar spermatozoa cryocapacitation during the cryopreservation process.

The improving effect of reduced glutathione on boar sperm cryotolerance is related with the intrinsic ejaculate freezability.

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Yeste, Marc; Estrada, Efrén; Pinart, Elisabeth; Bonet, Sergi; Miró, Jordi; Rodríguez-Gil, Joan E

2014-04-01

Reduced glutathione (GSH) improves boar sperm cryosurvival and fertilising ability when added to freezing extenders. Poor freezability ejaculates (PFE) are known to present lower resistance than good freezability ejaculates (GFE) to cryopreservation procedures. So far, no study has evaluated whether the ability of GSH to counteract the cryopreservation-induced injuries depends on ejaculate freezability (i.e. GFE vs. PFE). For this reason, thirty boar ejaculates were divided into three equal volume fractions and cryopreserved with or without GSH at a final concentration of either 2 or 5mM in freezing media. Before and after freeze-thawing, sperm quality was evaluated through analysis of viability, motility, integrity of outer acrosome membrane, ROS levels, integrity of nucleoprotein structure, and DNA fragmentation. Ejaculates were classified into two groups (GFE or PFE) according to their post-thaw sperm motility and viability assessments in negative control (GSH 0mM), after running cluster analyses. Values of each sperm parameter were then compared between treatments (GSH 0mM, GSH 2mM, GSH 5mM) and freezability groups (GFE, PFE). In the case of GFE, GSH significantly improved boar sperm cryotolerance, without differences between 2 and 5mM. In contrast, PFE freezability was significantly increased when supplemented with 5mM GSH, but not when supplemented with 2mM GSH. In conclusion, PFE need a higher concentration of GSH than GFE to improve their cryotolerance. Copyright © 2014 Elsevier Inc. All rights reserved.

Antioxidative effects of magnetized extender containing bovine serum albumin on sperm oxidative stress during long-term liquid preservation of boar semen.

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Lee, Sang-Hee; Park, Choon-Keun

2015-08-21

Magnetized water is defined as water that has passed through a magnet and shows increased permeability into cells and electron-donating characteristics. These attributes can protect against membrane damage and remove reactive oxygen species (ROS) in mammalian cells. We explored the effects of improved magnetized semen extenders containing bovine serum albumin (BSA) as antioxidants on apoptosis in boar sperm. Ejaculated semen was diluted in magnetized extender (0G and 6000G) with or without BSA (0G + BSA and 6000G + BSA), and sperm were analyzed based on viability, acrosome reaction, and H2O2 level of live sperm using flow cytometry. Sperm were then preserved for 11 days at 18 °C. We found that viability was significantly higher in 6000G + BSA than under the other treatments (P extenders have antioxidative effects on the liquid preservation of boar sperm. Copyright © 2015 Elsevier Inc. All rights reserved.

Influence of cryoprotectants glycerol and amides, combined with antioxidants on quality of frozen-thawed boar sperm.

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Buranaamnuay, K; Grossfeld, R; Struckmann, C; Rath, D

2011-08-01

The present study was undertaken to examine whether the cooling and freezing extenders containing a mixture of antioxidants (AOs) catalase, Na-pyruvate and mercaptoethanol and one of three types of cryoprotectants (CPs) would be able to improve the quality of frozen-thawed boar sperm. The collected semen, only the sperm-rich fraction, was diluted 1:1 with Androhep plus™ extender, stored at 15°C for 2 h and centrifuged. The centrifuged sperm pellet was re-suspended in lactose-egg yolk extender and divided into four groups for mixing with freezing extenders containing different kinds of CPs at 5°C: (I) glycerol (GLY) as control; (II) GLY with AOs; (III) dimethyl formamide (DMF) with AOs and (IV) dimethyl acetamide (DMA) with AOs. Processed sperm were packaged in 0.25-mL straws and frozen using a controlled rate freezer. After thawing, the diluted thawed sperm were incubated at 38°C for 10 min and was assessed for motility by CASA, membrane/acrosome integrity by FITC-PNA/PI and DNA integrity (DFI) by SCSA. All sperm parameters evaluated, except DFI, were negatively affected (P0.05). There was no difference in DFI among the studied groups (P>0.05). In conclusion, based on the present results, addition of AOs to cooling and freezing extenders and/or replacement of GLY with DMF or DMA could not improve quality of frozen-thawed boar sperm. Copyright © 2011 Elsevier B.V. All rights reserved.

Antioxidative effects of magnetized extender containing bovine serum albumin on sperm oxidative stress during long-term liquid preservation of boar semen

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Lee, Sang-Hee; Park, Choon-Keun

2015-01-01

Magnetized water is defined as water that has passed through a magnet and shows increased permeability into cells and electron-donating characteristics. These attributes can protect against membrane damage and remove reactive oxygen species (ROS) in mammalian cells. We explored the effects of improved magnetized semen extenders containing bovine serum albumin (BSA) as antioxidants on apoptosis in boar sperm. Ejaculated semen was diluted in magnetized extender (0G and 6000G) with or without BSA (0G + BSA and 6000G + BSA), and sperm were analyzed based on viability, acrosome reaction, and H 2 O 2 level of live sperm using flow cytometry. Sperm were then preserved for 11 days at 18 °C. We found that viability was significantly higher in 6000G + BSA than under the other treatments (P < 0.05). The acrosome reaction was significantly lower in the 6000G + BSA group compared with the other treatments (P < 0.05). Live sperm with high intracellular H 2 O 2 level were significantly lower in the 6000G + BSA group than under other treatments (P < 0.05). Based on our results, magnetized extenders have antioxidative effects on the liquid preservation of boar sperm. - Highlights: • Magnetized water is water that has been passed through a magnetic field. • Magnetized extender improve viability and decrease oxidative stress of boar sperm for preservation. • Ejaculated semen diluted with magnetized extender can improve liquid preservation period

Effect of storage in short--and long-term commercial semen extenders on the motility, plasma membrane and chromatin integrity of boar spermatozoa.

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De Ambrogi, Marco; Ballester, Juan; Saravia, Fernando; Caballero, Ignacio; Johannisson, Anders; Wallgren, Margareta; Andersson, Magnus; Rodriguez-Martinez, Heriberto

2006-10-01

For artificial insemination (AI) in pigs, preservation of liquid boar semen at 16-20 degrees C is still common practice as sperm cryopreservation remains suboptimal in this species. To meet the different needs of the swine industry, several extenders have been developed to preserve semen in liquid form for short--and long-term storage. In the present study, three different commercial extenders devised for short-term (BTS+) or long-term preservation (MR-A and X-Cell), were used to test whether storage of semen from four mature, fertile boars at 17 degrees C for 96 h would affect sperm characteristics relevant for fertility, such as motility, membrane integrity and chromatin stability. Computer-assisted sperm analysis, and stainings with the acylated membrane dye SYBR-14/propidium iodide, and acridine orange in connection with flow cytometry were used to evaluate these variables. Percentages of total motile spermatozoa decreased slightly, but significantly, after 72-96 h. While membrane integrity values varied during the period of study, no significant changes in either membrane integrity or chromatin stability were, however, registered. This suggests a customary 96-day storage at 17 degrees C in these extenders was too short an interval to cause losses of integrity in nuclear DNA in the boar population studied.

Effect of dietary selenium and vitamin E on the ultrastructure and ATP concentration of boar spermatozoa, and the efficacy of added sodium selenite in extended semen on sperm motility.

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Marin-Guzman, J; Mahan, D C; Whitmoyer, R

2000-06-01

Three experiments evaluated the effects of dietary Se and vitamin E on the ultrastructure of spermatozoa, ATP concentration of spermatozoa, and the effects of adding sodium selenite to semen extenders on subsequent sperm motility. The experiment was a 2 x 2 arrangement of treatments in a randomized complete block design. A total of 10 mature boars were fed from weaning to 18 mo of age diets fortified with two levels of supplemental Se (0 or .5 ppm) or vitamin E (0 or 220 IU/kg diet). The nonfortified diets contained .06 ppm Se and 4.4 IU vitamin E/kg. In Exp. 1, the spermatozoa from all boars were examined by electron microscopy. Vitamin E had no effect on structural abnormalities in the spermatozoa. When the low-Se diet was fed the acrosome or nuclei of the spermatozoa was unaffected, but the mitochondria in the tail midpiece were more oval with wider gaps between organelles. The plasma membrane connection to the tail midpiece was not tightly bound as when boars were fed Se. Immature spermatozoa with cytoplasmic droplets were more numerous when boars were fed the low-Se diet, but the occurrence of midpiece abnormalities occurred in boars fed diets with or without Se or vitamin E. Our results suggest that Se may enhance spermatozoa maturation in the epididymis and may reduce the number of sperm with cytoplasmic droplets. In Exp. 2, the concentration of ATP in the spermatozoa was evaluated in the semen of all treatment boars. When the low-Se diet was fed, ATP concentration was lower (P boar semen with a semen extender with sodium selenite added at 0, .3, .6, or .9 ppm Se. Three ejaculates from each boar were used to evaluate these effects on sperm motility to 48 h after dilution. Sperm motility declined (P extender, and this decline was exacerbated as the concentration of added Se increased (P boars resulted in abnormal spermatozoal mitochondria, a lower ATP concentration in the spermatozoa, and a loose apposition of the plasma membrane to the helical coil of the

In search of epigenetic marks in testes and sperm cells of differentially fed boars.

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Rémy Bruggmann

Full Text Available In search of transmittable epigenetic marks we investigated gene expression in testes and sperm cells of differentially fed F0 boars from a three generation pig feeding experiment that showed phenotypic differences in the F2 generation. RNA samples from 8 testes of boars that received either a diet enriched in methylating micronutrients or a control diet were analyzed by microarray analysis. We found moderate differential expression between testes of differentially fed boars with a high FDR of 0.82 indicating that most of the differentially expressed genes were false positives. Nevertheless, we performed a pathway analysis and found disparate pathway maps of development_A2B receptor: action via G-protein alpha s, cell adhesion_Tight junctions and cell adhesion_Endothelial cell contacts by junctional mechanisms which show inconclusive relation to epigenetic inheritance. Four RNA samples from sperm cells of these differentially fed boars were analyzed by RNA-Seq methodology. We found no differential gene expression in sperm cells of the two groups (adjusted P-value>0.05. Nevertheless, we also explored gene expression in sperm by a pathway analysis showing that genes were enriched for the pathway maps of bacterial infections in cystic fibrosis (CF airways, glycolysis and gluconeogenesis p.3 and cell cycle_Initiation of mitosis. Again, these pathway maps are miscellaneous without an obvious relationship to epigenetic inheritance. It is concluded that the methylating micronutrients moderately if at all affects RNA expression in testes of differentially fed boars. Furthermore, gene expression in sperm cells is not significantly affected by extensive supplementation of methylating micronutrients and thus RNA molecules could not be established as the epigenetic mark in this feeding experiment.

Antioxidative effects of magnetized extender containing bovine serum albumin on sperm oxidative stress during long-term liquid preservation of boar semen

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Lee, Sang-Hee; Park, Choon-Keun, E-mail: parkck@kangwon.ac.kr

2015-08-21

Magnetized water is defined as water that has passed through a magnet and shows increased permeability into cells and electron-donating characteristics. These attributes can protect against membrane damage and remove reactive oxygen species (ROS) in mammalian cells. We explored the effects of improved magnetized semen extenders containing bovine serum albumin (BSA) as antioxidants on apoptosis in boar sperm. Ejaculated semen was diluted in magnetized extender (0G and 6000G) with or without BSA (0G + BSA and 6000G + BSA), and sperm were analyzed based on viability, acrosome reaction, and H{sub 2}O{sub 2} level of live sperm using flow cytometry. Sperm were then preserved for 11 days at 18 °C. We found that viability was significantly higher in 6000G + BSA than under the other treatments (P < 0.05). The acrosome reaction was significantly lower in the 6000G + BSA group compared with the other treatments (P < 0.05). Live sperm with high intracellular H{sub 2}O{sub 2} level were significantly lower in the 6000G + BSA group than under other treatments (P < 0.05). Based on our results, magnetized extenders have antioxidative effects on the liquid preservation of boar sperm. - Highlights: • Magnetized water is water that has been passed through a magnetic field. • Magnetized extender improve viability and decrease oxidative stress of boar sperm for preservation. • Ejaculated semen diluted with magnetized extender can improve liquid preservation period.

Effect of addition of coconut water (Cocos nucifera) to the freezing media on post-thaw viability of boar sperm.

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Bottini-Luzardo, María; Centurión-Castro, Fernando; Alfaro-Gamboa, Militza; Aké-López, Ricardo; Herrera-Camacho, José

2013-01-01

The aims of this experiment were to evaluate the addition of coconut water in natura to the freezing media, compare the effect of deionized water vs filtered water of coconut over the post-thaw seminal characteristics, and evaluate the effect of the deionized water and in natura coconut water on the seminal characteristics of boar sperm at different post-thaw times. Thirty-four ejaculates were used divided in three aliquots which received one of the following treatments (T): T1, LEY (bidistilled water, lactose, and egg yolk) and LEYGO (LEY + glycerol and Orvus ET paste); T2, LEY(A) (coconut deionized water, lactose, and egg yolk)-LEYGO(A); and T3, LEY(B) (in natura coconut water, lactose, and egg yolk)-LEYGO(B). Samples of boar semen were frozen according to the Westendorf method, thawed at 38°C, and evaluated at three incubation times (0, 30, and 60 min). Seminal characteristics assessed were motility (Mot), acrosomal integrity (AInt), membrane integrity (MInt), and mitochondrial activity (MAct). T1 showed a higher percentage of viable sperm than T3 (Mot 36.5 vs 5.4 %, AInt 61.8 vs 41.2 %, MInt 50.4 vs 41.3 %, and MAct 56.9 vs 50.5 %). T2 kept a higher percentage of viable sperm at all incubation times. In natura coconut water showed a detrimental effect over the viability of the frozen-thawed boar semen. Deionized coconut water improved the boar semen viability post-thaw, outperforming results of in natura coconut water.

Bacterial Contamination of Boar Semen and its Relationship to Sperm Quality Preserved in Commercial Extender Containing Gentamicin Sulfate.

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Gączarzewicz, D; Udała, J; Piasecka, M; Błaszczyk, B; Stankiewicz, T

2016-09-01

This study was designed to determine the degree and type of bacterial contamination in boar semen (79 ejaculates from Large White and Landrace boars) and its consequences for sperm quality during storage (27 extended semen samples, 16°C for five days) under practical conditions of artificial insemination (AI). The results revealed the presence of aerobic bacteria in 99% of the ejaculates (from 80 to 370 ×106 colony-forming units/mL). Most of the ejaculates contained two or three bacterial contaminants, while the Staphylococcus, Streptococcus, and Pseudomonas bacterial genera were most frequently isolated. Also detected were Enterobacter spp., Bacillus spp., Proteus spp., Escherichia coli, P. fluorescens, and P. aeruginosa. In general, the growth of certain bacterial types isolated prior to semen processing (Enterobacter spp., E. coli, P. fluorescens, and P. aeruginosa) was not discovered on different days of storage, but fluctuations (with a tendency towards increases) were found in the frequencies of Bacillus spp., Pseudomonas spp., and Staphylococcus spp. isolates up to the end of storage. Semen preserved for five days exhibited decreases in sperm motility and increases in the average number of total aerobic bacteria; this was associated with sperm agglutination, plasma membrane disruption, and acrosome damage. We inferred that, due to the different degrees and types of bacterial contaminants in the boar ejaculates, the inhibitory activity of some antimicrobial agents used in swine extenders (such as gentamicin sulfate) may be limited. Because such agents can contribute to the overgrowth of certain aerobic bacteria and a reduction in the quality of stored semen, procedures with high standards of hygiene and microbiological control should be used when processing boar semen.

Egg Yolk Protective Effect in Boar Spermatozoa Cooled at 5ºC

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Alexandru-Vasile Rusu

2011-05-01

Full Text Available Nowadays, many boar reproduction researches are directed to improve extenders and to increase cold shock protection of semen. Little research is focused on the influence of egg yolk combined with alternative cold shock protective media. Egg yolk could interfere with other compounds present in the extender composition. The influence of egg yolk addition was assessed in boar sperm cells, cooled at 5ºC, to elucidate its effect on motility and membrane integrity. Flow Cytometry and Computer Assisted Semen Analysis (CASA were used to determine the rate of sperm with intact plasma and acrosomal membrane, respectively the sperm cells motility. Statistical analyses (T-Test were performed using GraphPad Prism version 5.00. Androhep Plus supplemented with 20% egg yolk (AhPlus+20%EY indicated a higher cold shock protection in progressive motility (93.9±2.64% and membrane integrity (79.78±4.14%, rather than the extender without egg yolk (p0.05. The combination egg yolk-AhPlus seems to be an alternative to standard extenders, conferring stability in boar sperm cells against cold shock.

Fluorescent multiple staining and CASA system to assess boar sperm viability and membranes integrity in short and long-term extenders.

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Lange-Consiglio, A; Meucci, A; Cremonesi, F

2013-01-01

The aim of this study was to assess the effect on boar spermatozoa quality of in vitro storage in short and long-term extenders by fluorescent multiple staining (FMS) and computer assisted semen analyzer (CASA). Fresh ejaculates from three healthy, sexually mature boars were diluted with equal volumes of six short-term or three long-term commercial extenders and stored at 19°C for 6 days (short-term) or 12 days (long-term). The integrity of spermatozoa membranes was analyzed by FMS using propidium iodide, 5,5',6,6'-tetrachloro-1,1',3,3' tetraethylbenzimidazolyl-carbocyanine iodide (JC-1) and fluorescein isothiocyanate-conjugated peanut agglutinin (PNA). The results obtained from this staining were compared with spermatozoa motility assessed by CASA. Our study showed that the number of viable spermatozoa with non-reacted acrosomes and intact mitochondria was positively correlated with the rate of motile spermatozoa (r(2)>0.9) irrespective of the extender used. In all extenders the number of motile spermatozoa significantly decreased as preservation period increased (P<0.05). FMS test is a potent indicator of sperm motility because it analyses mitochondrial integrity independently from observable alterations in motility. The best performing extenders were BTS for short-term storage and TRI-x-Cell for long-term storage.

Characteristics of sperm motility in boar semen diluted in different extenders and stored for seven days at 18 degrees C.

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Estienne, Mark J; Harper, Allen F; Day, Jennifer L

2007-11-01

Although numerous extenders exist for diluting boar semen, little research has been conducted comparing commercial extenders with regard to maintaining sperm motility during storage. The objective was to use a computer- assisted sperm analysis system to assess motility of boar spermatozoa diluted in Beltsville Thawing Solution, Merck-III, Androhep-lite, Sperm Aid, MR-A, Modena, X-Cell, VSP, and Vital. Ejaculates from boars (n=10) were collected and sub-samples were diluted (35x10(6) spermatozoa/ml) in the different extenders and stored for seven days at 18 degrees. Extender by day interactions were detected (pextenders. For example, on day 7, the percentages of motile and progressively motile spermatozoa were highest (pextender utilized, but with the exception of Sperm Aid, all extenders maintained a high degree of sperm motility through 7 days of storage.

Effect of commercial long-term extenders on metabolic activity and membrane integrity of boar spermatozoa stored at 17 degrees C.

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Dziekońska, A; Fraser, L; Majewska, A; Lecewicz, M; Zasiadczyk, Ł; Kordan, W

2013-01-01

This study was aimed to analyze the metabolic activity and membrane integrity of boar spermatozoa following storage in long-term semen extenders. Boar semen was diluted with Androhep EnduraGuard (AeG), DILU-Cell (DC), SafeCell Plus (SCP) and Vitasem LD (VLD) extenders and stored for 10 days at 17 degrees C. Parameters of the analyzed sperm metabolic activity included total motility (TMOT), progressive motility (PMOT), high mitochondrial membrane potential (MMP) and ATP content, whereas those of the membrane integrity included plasma membrane integrity (PMI) and normal apical ridge (NAR) acrosome. Extender type was a significant (P semen storage. In all extenders the metabolic activity and membrane integrity of the stored spermatozoa decreased continuously over time. Among the four analyzed extenders, AeG and SCP showed the best performance in terms of TMOT and PMI on Days 5, 7 and 10 of storage. Marked differences in the proportions of spermatozoa with high MMP were observed between the extenders, particularly on Day 10 of storage. There were not any marked differences in sperm ATP content between the extenders, regardless of the storage time. Furthermore, the percentage of spermatozoa with NAR acrosomes decreased during prolonged storage, being markedly lower in DC-diluted semen compared with semen diluted with either AeG or SCP extender. The results of this study indicated that components of the long-term extenders have different effects on the sperm functionality and prolonged semen longevity by delaying the processes associated with sperm ageing during liquid storage.

Adverse effects of members of the Enterobacteriaceae family on boar sperm quality.

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Ubeda, Juan Luis; Ausejo, Raquel; Dahmani, Yahya; Falceto, Maria V; Usan, Adan; Malo, Clara; Perez-Martinez, Francisco C

2013-10-01

Semen samples collected in 2012 from 1785 boars belonging to five different breeds were recruited from the quality control laboratory of Magapor SL, Spain. These samples came from 43 boar studs and resulted from diluting the ejaculates in commercial semen extenders. Evaluation of the semen sample characteristics (color, smell, pH, osmolality, concentration, motility of sperm cells, agglutination, acrosome integrity, short hypoosmotic swelling test, and abnormal forms) revealed that they met the international standards. The samples were also tested for the presence of aerobic bacterial contamination. In the present study, 14.73% (n = 263) of the semen samples were contaminated above 3 × 10(2) colony-forming units/mL with at least one type of bacteria. The Enterobacteriaceae family was by far the major contaminant, being present in 40.68% of the contaminated samples (n = 107). Bacterial strains of the Enterobacteriaceae family isolated from boar semen samples were in order of incidence (percentage of the contaminated samples): Serratia marcescens (12.55%), Klebsiella oxytoca (11.79%), Providencia stuartii (9.12%), Morganella morganii (3.80%), Proteus mirabilis (1.90%), and Escherichia coli (1.52%). We have seen that the presence in semen samples of S. marcescens, K. oxytoca, M. morganii, or P. mirabilis, but not P. stuartii or E. coli, was negatively associated with sperm motility (P semen samples infected with M. morganii compared with uninfected ones. Moreover, P. mirabilis was negatively associated with the presence of abnormal forms. Thus, on the basis of the pathological effects that some of these strains may have on boar sperm quality, bacterial contamination should always be examined in semen samples prepared for artificial insemination. Copyright © 2013 Elsevier Inc. All rights reserved.

Effects of different extenders on DNA integrity of boar spermatozoa following freezing-thawing.

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Hu, Jian-hong; Li, Qing-wang; Jiang, Zhong-liang; Li, Wen-ye

2008-12-01

The sperm-rich fraction, collected from eight mature Yorkshire boars, was frozen in an extender containing 9% LDL (w/v), 100mM trehalose, or 20% yolk (v/v), respectively. Sperm DNA integrity was assessed using the single-cell gel electrophoresis (SCGE). Other sperm quality characteristics such as motility, acrosome and membrane integrity were also monitored. The results showed that freezing-thawing caused an increase in sperm DNA fragmentation, and extender containing 9% LDL could significantly protect sperm DNA integrity (Pextender containing 100mM trehalose and 20% yolk (v/v). No significant difference in damaged DNA was detected between frozen and unfrozen semen samples for extender of 9% LDL and 100mM trehalose, but cryopreservation could increase the degree of DNA damage (Pboar sperm DNA damage and protecting DNA integrity. It can be suggested that evaluation of sperm DNA integrity, coupled with correlative and basic characteristics such as motility, acrosome integrity and membrane integrity, may aid in determining the quality of frozen boar semen.

Fluorescent multiple staining and CASA system to assess boar sperm viability and membranes integrity in short and long-term extenders

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F. Cremonesi

2013-03-01

Full Text Available The aim of this study was to assess the effect on boar spermatozoa quality of in vitro storage in short and long-term extenders by fluorescent multiple staining (FMS and computer assisted semen analyzer (CASA. Fresh ejaculates from three healthy, sexually mature boars were diluted with equal volumes of six short-term or three long-term commercial extenders and stored at 19°C for 6 days (short-term or 12 days (long-term. The integrity of spermatozoa membranes was analyzed by FMS using propidium iodide, 5,5’,6,6’-tetrachloro-1,1’,3,3’ tetraethylbenzimidazolyl-carbocyanine iodide (JC-1 and fluorescein isothiocyanate-conjugated peanut agglutinin (PNA. The results obtained from this staining were compared with spermatozoa motility assessed by CASA. Our study showed that the number of viable spermatozoa with non-reacted acrosomes and intact mitochondria was positively correlated with the rate of motile spermatozoa (r2>0.9 irrespective of the extender used. In all extenders the number of motile spermatozoa significantly decreased as preservation period increased (P<0.05. FMS test is a potent indicator of sperm motility because it analyses mitochondrial integrity independently from observable alterations in motility. The best performing extenders were BTS for short-term storage and TRI-x-Cell for long-term storage.

Decrease in glutathione content in boar sperm after cryopreservation. Effect of the addition of reduced glutathione to the freezing and thawing extenders.

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Gadea, Joaquín; Sellés, Elena; Marco, Marco Antonio; Coy, Pilar; Matás, Carmen; Romar, Raquel; Ruiz, Salvador

2004-08-01

Although glutathione content in boar spermatozoa has been previously reported, the effect of reduced glutathione (GSH) on semen parameters and the fertilizing ability of boar spermatozoa after cryopreservation has never been evaluated. In this study, GSH content was determined in ejaculated boar spermatozoa before and after cryopreservation. Semen samples were centrifuged and GSH content in the resulting pellet monitored spectrophotometrically. The fertilizing ability of frozen-thawed boar sperm was also tested in vitro by incubating sperm with in vitro matured oocytes obtained from gilts. GSH content in fresh semen was 3.84 +/- 0.21 nM GSH/10(8) sperm. Following semen cryopreservation, there was a 32% decrease in GSH content (P boars and after various preservation protocols (P = 0.0102 ). The effect of addition of GSH to the freezing and thawing extenders was also evaluated. Addition of 5 mM GSH to the freezing extender did not have a significant effect on standard semen parameters or sperm fertilizing ability after thawing. In contrast, when GSH was added to the thawing extender, a dose-dependent tendency to increase in sperm fertilizing ability was observed, although no differences were observed in standard semen parameters. In summary, (i) there was a loss in GSH content after cryopreservation of boar semen; (ii) addition of GSH to the freezing extender did not result in any improvement in either standard semen parameters or sperm fertilizing ability; and (iii) addition of GSH to the thawing extender resulted in a significant increase in sperm fertilizing ability. Nevertheless, future studies must conclude if this is the case for all boars. Furthermore, since addition of GSH to the thawing extender did not result in an improvement in standard semen parameters, this suggests that during the thawing process, GSH prevents damage of a sperm property that is critical in the fertilization process but that is not measured in the routine semen analysis.

Impact of Storage and Purification on Mitochondrial Membrane Potential of Boar Spermatozoa

OpenAIRE

Aristotelis G. Lymberopoulos; TAREK KHALIFA

2013-01-01

This study aimed to evaluate the effect of semen purification and storage on sperm mitochondrial membrane potential (ΔΨm). Gel-free whole ejaculates were collected from five proven fertile Large White boars aged two to three years. Aliquots of fresh semen were split, diluted in one step with commercial extenders and incubated at 37oC for 5-10 minutes. Semen was cooled to 18oC and packaged in 15-ml sterile propylene tubes. After 4-10 hours post-semen collection, stored semen was purified by co...

Boar sperm storage capacity of BTS and Androhep Plus: viability, motility, capacitation, and tyrosine phosphorylation.

Science.gov (United States)

Dubé, Charlotte; Beaulieu, Martin; Reyes-Moreno, Carlos; Guillemette, Christine; Bailey, Janice L

2004-09-01

Androhep Plus, a long-term extender (up to 7 days) and Beltsville Thawing Solution (BTS), a short-term extender (up to 3 days), are commonly used for liquid storage of porcine semen. To test the hypothesis that modifications in sperm viability, motility, chlortetracycline (CTC) fluorescence patterns, and protein tyrosine phosphorylation occur during semen storage in extenders, we compared these end points at different periods of storage in either Androhep Plus or BTS. Sperm from five boars were assessed daily over 12 days of storage (n = 5 ejaculates from different boars). Viability was not different (P extenders, except on Day 2, when Androhep Plus maintained better viability. Differences in the percentage of motile (total) sperm due to extender were evident on Days 2, 4, 5, and 6, when Androhep Plus was superior to BTS (P extender as early as Day 2; storage in Androhep Plus induced higher levels of pattern B sperm (P extenders; these may affect the fertilizing capacity of the semen.

Sperm quality and oxidative status as affected by homogenization of liquid-stored boar semen diluted in short- and long-term extenders.

Science.gov (United States)

Menegat, Mariana B; Mellagi, Ana Paula G; Bortolin, Rafael C; Menezes, Tila A; Vargas, Amanda R; Bernardi, Mari Lourdes; Wentz, Ivo; Gelain, Daniel P; Moreira, José Cláudio F; Bortolozzo, Fernando P

2017-04-01

Homogenization of diluted boar semen during storage has for a long time been regarded as beneficial. Recent studies indicated an adverse effect of homogenization on sperm quality for yet unknown reasons. This study aimed to verify the effect of homogenization on sperm parameters and to elucidate the impact of oxidative stress. Twenty-one normospermic ejaculates (21 boars) were diluted with Androstar ® Plus (AND) and Beltsville Thawing Solution (BTS). Semen doses were submitted to no-homogenization (NoHom) or twice-a-day manual homogenization (2xHom) during storage at 17°C for 168h. NoHom and 2xHom were similar (P>0.05) for both short- and long-term extenders with respect to motility and kinematics parameters (CASA system), membrane viability (SYBR-14/PI), acrosome integrity, lipid peroxidation, protein oxidation, intracellular reactive oxygen species, sulfhydryl content, and total radical-trapping antioxidant potential. 2xHom reduced sperm motility and motion kinematics (VCL, VSL, VAP, BCF, and ALH) following the thermoresistance test and presented with a slight increase in pH along the storage (P=0.05) as compared to NoHom. Furthermore, 2xHom semen doses presented with a constant SOD and GSH-Px activity during storage whereas enzymatic activity increased for NoHom at the end of the storage. These findings confirm that homogenization of semen doses is detrimental to sperm quality. Moreover, it is shown that the effect of homogenization is unlikely to be primarily related to oxidative stress. Homogenization is not recommended for storage of liquid boar semen for up to 168h in both short- and long-term extenders. Copyright © 2017 Elsevier B.V. All rights reserved.

Cryopreservation of boar semen by egg yolk-based extenders containing lactose or fructose is better than sorbitol.

Science.gov (United States)

Chanapiwat, Panida; Kaeoket, Kampon; Tummaruk, Padet

2012-03-01

The present study determined the effect of different types of sugars (lactose, fructose, glucose and sorbitol) used in egg yolk-based extender on the post-thawed boar semen quality. Twenty-two ejaculates from 6 fertility-proven Yorkshire boars were cryopreserved by liquid nitrogen vapor method. Sperm motility, viability, acrosome integrity and intact functional plasma membrane were determined at 0, 2 and 4 hr after thawing. It was found that the lactose-based extender resulted in a higher percentage of post-thawed sperm motility, viability, intact acrosome and functional plasma membrane than sorbitol-based extender (Pextender yielded a higher post-thawed sperm motility and viability than sorbitol-based extender (Pboar semen.

Method agreement between measuring of boar sperm concentration using Makler chamber and photometer

OpenAIRE

Mrkun Janko; Kosec M.; Zakošek Maja; Zrimšek Petra

2007-01-01

Determination of boar sperm concentration using a photometer is used routinely by many artificial insemination (AI) laboratories. The agreement between determining sperm concentration using Makler chamber and a photometer has been assessed. Method agreement was evaluated on the basis of scatter plots with Deming regression line, absolute bias plots with limits of agreement, and relative bias plots. Coefficients of variance for the Makler chamber and a photometer were calculated as 6.575+3.461...

Ophiobolin A from Bipolaris oryzae Perturbs Motility and Membrane Integrities of Porcine Sperm and Induces Cell Death on Mammalian Somatic Cell Lines

Directory of Open Access Journals (Sweden)

Ottó Bencsik

2014-09-01

Full Text Available Bipolaris oryzae is a phytopathogenic fungus causing a brown spot disease in rice, and produces substance that strongly perturbs motility and membrane integrities of boar spermatozoa. The substance was isolated from the liquid culture of the fungal strain using extraction and a multi-step semi-preparative HPLC procedures. Based on the results of mass spectrometric and 2D NMR techniques, the bioactive molecule was identified as ophiobolin A, a previously described sesterterpene-type compound. The purified ophiobolin A exhibited strong motility inhibition and viability reduction on boar spermatozoa. Furthermore, it damaged the sperm mitochondria significantly at sublethal concentration by the dissipation of transmembrane potential in the mitochondrial inner membrane, while the plasma membrane permeability barrier remained intact. The study demonstrated that the cytotoxicity of ophiobolin A toward somatic cell lines is higher by 1–2 orders of magnitude compared to other mitochondriotoxic mycotoxins, and towards sperm cells unique by replacing the progressive motility by shivering tail beating at low exposure concentration.

Liquid storage of miniature boar semen.

Science.gov (United States)

Shimatsu, Yoshiki; Uchida, Masaki; Niki, Rikio; Imai, Hiroshi

2002-04-01

The effects of liquid storage at 15 degrees C on the fertilizing ability of miniature pig semen were investigated. Characterization of ejaculated semen from 3 miniature boars was carried out. Semen volume and pH were similar among these boars. In one of the boars, sperm motility was slightly low, and sperm concentration and total number of sperm were significantly lower than in the others (P semen was substituted with various extenders (Kiev, Androhep, BTS and Modena) by centrifugation and semen was stored for 7 days at 15 degrees C. Sperm motility was estimated daily at 37 degrees C. For complete substitution of seminal plasma, Modena was significantly more efficient than the other extenders (P Semen from each of the 3 miniature boars that had been stored for 5 to 7 days at 15 degrees C in Modena was used for artificial insemination of 15 miniature sows. The farrowing rates were 100, 100 and 60%, and litter sizes were 6.4 +/- 1.5, 5.8 +/- 0.8 and 5.0 +/- 1.0 for each boar semen, respectively. The boar that sired the smallest farrowing rate was the same one that showed lower seminal quality with respect to sperm motility, sperm concentration and total number of sperm. These results suggest that miniature boar semen can be stored for at least 5 days at 15 degrees C by the substitution of seminal plasma with Modena extender.

Evaluation of amides and centrifugation temperature in boar semen cryopreservation.

Science.gov (United States)

Bianchi, I; Calderam, K; Maschio, E F; Madeira, E M; da Rosa Ulguim, R; Corcini, C D; Bongalhardo, D C; Corrêa, E K; Lucia, T; Deschamps, J C; Corrêa, M N

2008-03-15

Two experiments were conducted to evaluate the use of amides as cryoprotectants and two centrifugation temperatures (15 or 24 degrees C) in boar semen cryopreservation protocols. Semen was diluted in BTS, cooled centrifuged, added to cooling extenders, followed by the addition of various cryoprotectants. In experiment 1, mean (+/-S.E.M.) sperm motility for 5% dimethylformamide (DMF; 50.6+/-1.9%) and 5% dimethylacetamide (DMA; 53.8+/-1.7%) were superior (P0.05). In experiment 2, we tested MF, DMF, and DMA at 3, 5, and 7%. Sperm motility and membrane integrity were higher for 5% DMA (53.8+/-1.7 and 50.9+/-1.9%) and 5% DMF (50.6+/-1.9 and 47.9+/-2.1%), in comparison with 7% DMF and all MF concentrations (P0.05). In conclusion, boar semen was successfully cryopreserved by replacement of glycerol with amides (especially 5% DMA) and centrifugation at 15 degrees C, with benefits for post-thaw sperm motility and membrane integrity.

Response to capacitating stimuli indicates extender-related differences in boar sperm function.

Science.gov (United States)

Schmid, S; Henning, H; Petrunkina, A M; Weitze, K F; Waberski, D

2013-10-01

Spermatozoa, especially those of the porcine species, are highly susceptible to in vitro chilling and ageing. Extenders are continuously developed to protect boar spermatozoa from chilling injury. New semen extenders and other modified preservation strategies require sensitive testing for essential sperm functions. The key process on the pathway of fertilization is capacitation. The aim of the present study was to examine whether the specific response to capacitating stimuli is sensitive enough to indicate different preservation capacities of extenders during hypothermic storage of boar spermatozoa. Semen was diluted in Beltsville Thawing Solution (BTS) and Androstar Plus and kept for 3 h at 22°C or stored at 17°C, 10°C, and 5°C. Semen was analyzed at 24 and 96 h of storage. Motility and membrane integrity remained at high levels, except for lower values when stored in BTS at 5°C. Washed subsamples were incubated in capacitating medium (Tyrode) and control medium and were assessed for intracellular calcium concentration and integrity of plasma membranes using a flow cytometer. On the basis of the loss of low-calcium live cells in a kinetic approach, the specific response to capacitation stimuli was determined. There was a higher loss of response in semen stored hypothermically in the standard extender BTS compared to Androstar Plus. Assessment of the extent of phospholipid disorder under capacitating and control conditions by use of merocyanine staining did not reveal any significant extender-related differences. A field insemination trial with 778 sows was performed to relate in vitro results to fertility. Fertility parameters did not differ in semen stored up to 48 h at 10°C in Androstar Plus compared to controls stored at 17°C in BTS. In conclusion, assessment of specific reactivity to capacitating stimuli appears to be a sensitive tool for detection of extender-dependent alterations in functionality of chilled boar spermatozoa.

Assessment of sperm viability, mitochondrial activity, capacitation and acrosome intactness in extended boar semen during long-term storage.

Science.gov (United States)

Huo, Li-Jun; Ma, Xing-Hong; Yang, Zeng-Ming

2002-10-15

The purpose of this study was to assess sperm quality in extended boar semen during in vitro storage in order to determine which extender should be used and how long boar semen can be stored. Freshly ejaculated boar semen was diluted with equal volumes of Beltsville thaw solution (BTS), Androhep, KIEV or Zorlesco extenders and stored at 17 degrees C for up to 15 days. Sperm quality was evaluated by examining viability using SYBR-14/PI and Hoechst 33258 staining, mitochondrial activity using 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolyl-carbocyanine iodide (JC-1) staining, acrosome intactness by Coomassie blue staining, and capacitation status by chlortetracycline (CTC) staining. There were over 50% viable spermatozoa in boar semen extended with Zorlesco and Androhep extenders on Day 13 of storage. The percentage of JC-1-stained spermatozoa was 53.8 +/- 2.1% for Zorlesco and 57.7 +/- 1.60% for Androhep extenders on Day 13 of storage. The percentage of acrosome-intact spermatozoa detected by Coomassie blue staining was higher than that in the SYBR-14PI-, Hoechst 33258-, and JC-1-stained samples in our study. The results from SYBR-14/PI, Hoechst 33258, JC-1, and Coomassie blue staining were highly correlated (r > or = 0.9461). There were less than 15% capacitated spermatozoa in the semen extended with BTS, Androhep and Zorlesco extenders during 9 days of storage. However, most viable boar spermatozoa became capacitated by Day 13 of storage. The rank order of four extenders for maintaining sperm viability and mitochondrial activity was as follows: Androhep, Zorlesco, BTS, KIEV.

Oxidoreductive capability of boar sperm mitochondria in fresh semen and during their preservation in BTS extender.

Science.gov (United States)

Gaczarzewicz, Dariusz; Piasecka, Małgorzata; Udała, Jan; Błaszczyk, Barbara; Laszczyńska, Maria; Kram, Andrzej

2003-07-01

The purpose of our study was to determine the effect of dilution and liquid-preservation of boar sperm on oxidoreductive capability of their mitochondria. The semen was diluted with BTS extender produced from water purified by destillation or by reverse osmosis. The spermatozoa were stored over a four-day period at 16-18 degrees C. The function of sperm mitochondria was assessed using the screening cytochemical test for NADH-dependent oxidoreductases (diaphorase/NADH, related to flavoprotein). Morphological assessment of cytochemical reaction was carried out using a light microscope. The intensity of the reaction was evaluated by means of a computer image analysing system (Quantimet 600S), measuring the integrated optical density (IOD) and mean optical density (MOD) of the reaction product (formazans) occurring in the sperm midpieces. In the non-diluted semen, intensive cytochemical reaction throughout the length of the sperm midpiece was observed. Furthermore, spermatozoa with the intensive reaction displayed the high optical density values. After dilution the semen with two variants of experimental extender, and as the conservation time expired, the cytochemical reaction was less intensive. Moreover, the absence of formazan deposits in various parts of the sperm midpiece was also noted. These morphological features corresponded to low values of optical density. These findings suggest that the dilution of semen and the time of sperm preservation may be critical factors that handicap energy metabolism of sperm mitochondria. The type of water used in preparing BTS extender does not have any significant effect on the oxidoreductive capability of sperm boar mitochondria.

Isolation of total ribonucleic acid from fresh and frozen-thawed boar ...

African Journals Online (AJOL)

RNA) from raw fresh semen and frozen-thawed boar semen, using a protocol comprising the conventional TRIzol assay and a membrane-based technique, the PureLink RNA mini kit. Bioanalyzer profile revealed that the sperm RNA size ...

Validation of the FACSCount AF system for determination of sperm concentration in boar semen

DEFF Research Database (Denmark)

Hansen, C.; Christensen, P.; Stryhn, H.

2002-01-01

microspheres ( beads). Satisfactory staining can be achieved within 2-3 min and the following flow cytometric analysis on the FACSCount AF System rapidly provides the user with a precise and accurate assessment of the sperm concentration. In this study, the FACSCount AF System and Sperm Counting Reagent ( BD...... Biosciences) was compared with microscopic counting using a Burker-Turk haemocytometer. In addition, sperm concentration was determined using the Corning 254 spectrophotometer which is used routinely by Danish artificial insemination stations for boars. The results show that the agreement between flow...... cytometry and microscopic counting is very high. The slope for the regression line was 1.12 (SE = 0.03) with an estimated intercept with the Y-axis of 22 x 10(6) sperm/ml (SE = 10 x 10(6) sperm/ml) and an estimated error of the model of 10 x 10(6) sperm/ml. For the spectrophotometer, the slope...

Effect of different procedures of ejaculate collection, extenders and packages on DNA integrity of boar spermatozoa following freezing-thawing.

Science.gov (United States)

Fraser, L; Strzezek, J

2007-06-01

Whole ejaculate or sperm-rich fraction, collected from four sexually mature boars, was frozen in an extender containing lactose-hen egg yolk with glycerol (lactose-HEY-G) or extender containing lactose, lyophilized lipoprotein fractions isolated from ostrich egg yolk and glycerol (lactose-LPFo-G), and Orvus Es Paste, respectively. The sperm samples were also frozen in a standard boar semen extender (Kortowo-3), without the addition of cryoprotective substances. Sperm DNA integrity was assessed using a modified neutral comet assay. Sperm characteristics such as motility, plasma membrane integrity (SYBR-14/PI), mitochondrial function (rhodamine 123) and acrosome integrity were monitored. Freezing-thawing caused a significant increase (Pextender type. Sperm DNA fragmentation was significantly lower (Pextender exhibited lower (Pboar semen.

Fluorescent multiple staining and CASA system to assess boar sperm viability and membranes integrity in short and long-term extenders

OpenAIRE

F. Cremonesi; A. Meucci; A. Lange-Consiglio

2013-01-01

The aim of this study was to assess the effect on boar spermatozoa quality of in vitro storage in short and long-term extenders by fluorescent multiple staining (FMS) and computer assisted semen analyzer (CASA). Fresh ejaculates from three healthy, sexually mature boars were diluted with equal volumes of six short-term or three long-term commercial extenders and stored at 19?C for 6 days (short-term) or 12 days (long-term). The integrity of spermatozoa membranes was analyzed by FMS using prop...

Evaluation of porcine beta defensins-1 and -2 as antimicrobial peptides for liquid-stored boar semen: Effects on bacterial growth and sperm quality.

Science.gov (United States)

Puig-Timonet, Adrià; Castillo-Martín, Miriam; Pereira, Barbara A; Pinart, Elisabeth; Bonet, Sergi; Yeste, Marc

2018-04-15

The present study evaluated whether two different antimicrobial peptides (AMP): porcine beta defensins-1 (PBD1) and -2 (PBD2) at three concentrations (1.5 μM, 3 μM and 5 μM) could be a suitable alternative to antibiotics in liquid-stored boar semen. Two separate experiments were conducted with liquid-stored boar semen preserved at 17 °C for 9-10 days. In the first one, we evaluated the impact of adding three concentrations of each AMP on the bacterial growth and sperm quality of boar semen stored for 10 days. In the second experiment, the ability of these AMPs to control bacterial growth was determined over a 9-day period, following artificial inoculation with Escherichia coli at 10 7 and 10 8  CFU mL -1 . In both experiments, sperm viability was assessed through flow cytometry, sperm motility was determined with Computer Assisted Sperm Analysis (CASA) and the inhibitory effect on microbial growth was evaluated by bacteria culture on Luria Bertani agar. PBD1 and PBD2 were found to significantly (P extenders for boar semen at a concentration of 3 μM, but do not completely control all bacterial growth. Copyright © 2018 Elsevier Inc. All rights reserved.

Membrane status and in vitro capacitation of porcine sperm preserved in long-term extender at 16 degrees C.

Science.gov (United States)

Conejo-Nava, J; Fierro, R; Gutierrez, C G; Betancourt, M

2003-01-01

Preservation of porcine semen in long-term extenders at 15-18 degrees C for more than 5 days results in decreased farrowing rates and reduced litter size after artificial insemination, despite the high progressive motility rates of sperm. To improve this preservation system it is necessary to understand sperm physiology under storage conditions. The purpose of this study was to determine the effect of storing diluted porcine semen (during 0, 2, 4, 6, and 8 days) on the sperm membranes status and the ability of sperm to respond to in vitro capacitation treatment. Ten semen samples from 5 adult boars were analyzed. Two aliquots were obtained from the sperm-rich fraction: one was used to assess fresh semen and the other was diluted in Reading extender and stored at 16 degrees C. Both semen samples were stained with chlortetracycline to assess the status of sperm membranes and with Hoechst 33258 to determine viability. Semen storage for 4-8 days increased the proportion of prematurely capacitated sperm. After 4 days of storage, in vitro capacitation treatment did not increase the percentage of capacitated sperm, but increased the percentage of acrosome reacted sperm. This phenomenon could explain the reduced fertilizing ability of porcine semen stored at 16 degrees C for over 4 days, in spite of the acceptable sperm viability and progressive motility.

Quality Control of Boar Sperm Processing : Implications from European AI Centres and Two Spermatology Reference Laboratories

NARCIS (Netherlands)

Riesenbeck, A; Schulze, M; Rüdiger, K; Henning, H; Waberski, D

In recent years, increased automatization has resulted in a higher efficiency of boar semen processing in AI laboratories. Sophisticated laboratory management and efficient quality control programmes are needed for current tendencies in major pork-producing countries to reduce the sperm number per

Deep-freezing of boar semen in plastic film 'cochettes'.

Science.gov (United States)

Eriksson, B M; Rodriguez-Martinez, H

2000-03-01

The motility and membrane integrity of spermatozoa from nine boars frozen with a programmable freezing machine in plastic bags, 'cochettes', and in 'maxi-straws', in total doses of 5 x 10(9) spermatozoa/5 ml with glycerol (3%) used as cryoprotectant, were assessed after thawing. A computer-based cell motion analyser was used to evaluate sperm motility, while the integrity of the plasmalemma was assessed with fluorescent supravital dyes (C-FDA/PI). The fertilizing capacity of the semen frozen in the two containers was investigated by inseminating (AI) gilts. Pregnancy was monitored by Doppler-ultrasound, and the numbers of corpora lutea and viable embryos counted at slaughter, between days 30 and 38 after AI. The cochettes sustained the overall procedure of freezing/thawing (FT), with 30 min post-thaw (PT) sperm motility being significantly higher than for straws, 46.9 vs. 39.5%. The only significant difference in motility patterns detected when comparing the packages was a higher sperm velocity (VCL) in cochettes at 30 min PT. However, percentages of FT-spermatozoa with intact membranes, detected with the supravital probes, were higher in maxi-straws than in cochettes, 46.8 vs. 43.0% (P straws and those frozen in cochettes. The results indicate that although the deep-freezing of AI-doses of boar semen in large plastic bags is feasible, problems such as their inconvenient size for storage and inconsistent thawing must be solved before this type of container can be used for the commercial cryopreservation of boar semen.

The effect of extender, method of thawing, and duration of storage on in vitro fertility measures of frozen-thawed boar sperm.

Science.gov (United States)

Knox, R V; Ringwelski, J M; McNamara, K A; Aardsma, M; Bojko, M

2015-08-01

Frozen-thawed boar sperm (FTS) has reduced in vitro and in vivo life span compared to liquid semen. Experiments tested whether extenders, thawing procedures, and storage temperatures could extend the fertile life span of FTS. Experiment 1 tested the effect of six extenders on postthaw motility (MOT) and viability (VIA). Straws from boars (n = 6) were thawed, diluted into each extender, and evaluated at 20, 60, and 120 minutes. There was a trend (P = 0.08) for an extender-by-time interaction for MOT and effect of extender and time for MOT (P extender (P = 0.10) and time (P boar ejaculates (n = 15) were thawed at 50 °C for 10, 20, or 30 seconds or at 70 °C for 5, 10, or 20 seconds and evaluated at 5, 30, and 60 minutes. There was an effect of thawing treatment on MOT, VIA, and ACR (viable sperm with intact acrosomes, P extenders, thawing, and storage. Copyright © 2015 Elsevier Inc. All rights reserved.

Supplementation of different concentrations of Orvus Es Paste (OEP) to ostrich egg yolk lipoprotein extender improves post-thaw boar semen quality.

Science.gov (United States)

Fraser, L; Jasiewicz, E; Kordan, W

2014-01-01

This study aimed to compare post-thaw quality of boar semen following freezing in an ostrich egg yolk lipoprotein (LPFo) extender supplemented with 0%, 0.25% and 0.50% Orvus Es Paste (OEP). Sperm assessments included total motility (TMOT), mitochondrial function (MF), plasma membrane integrity (PMI) and acrosome integrity (normal apical ridge, NAR). Considerable variations among boars and OEP treatments had a significant effect (P semen samples frozen in the absence of OEP. By contrast, lactose-LPFo-glycerol extender supplemented with either 0.25% OEP or 0.50% OEP markedly enhanced post-thaw sperm characteristics. In all boars, there were no marked differences in post-thaw sperm TMOT between the freezing extenders supplemented with 0.25% and 0.50% OEP. However, a decline in the percentage of post-thaw motile spermatozoa was more pronounced in the extender supplemented with 0.50% OEP following a 120-min incubation period. Furthermore, the proportions of frozen-thawed spermatozoa with MF, PMI and NAR acrosomes varied significantly among the boars in the OEP-supplemented extenders. The findings of this study indicate that different OEP concentrations, in the presence of ostrich egg yolk lipoproteins, could have varying effects on post-thaw sperm survival.

Effects of the antimicrobial peptide protegrine 1 on sperm viability and bacterial load of boar seminal doses.

Science.gov (United States)

Sancho, S; Briz, M; Yeste, M; Bonet, S; Bussalleu, E

2017-10-01

The presence of bacteria adversely affects boar sperm quality of seminal doses intended for artificial insemination. Currently, the most common measure to prevent bacteriospermia is the addition of antibiotics in semen extenders; however, mounting evidence shows that microbial resistance exists. A promising alternative to replace antibiotics are antimicrobial peptides. In this study, the effects of the antimicrobial peptide protegrine 1 (PG1) on the sperm viability and bacterial load of boar seminal doses were evaluated. Three different concentrations of PG1 (2.5, 25 and 100 μg/ml) were tested over a storing period of 10 days at 17°C. Sperm viability was analysed by fluorescence microscopy (SYBR14/propidium iodide), and bacterial load was assessed by plating 100 μl of each sample in Luria-Bertani medium and incubated at 37°C for 72 hr under aerobic conditions. Protegrine 1 was effective in controlling the bacterial load in all the assessed concentrations (p < .05), reaching the lowest values at the highest concentrations of the antimicrobial peptide. Nevertheless, sperm viability was significantly (p < .05) reduced by all tested concentrations of this peptide, the most cytotoxic effects being observed at the highest PG1 concentrations. Despite these results, the use of PG1 as an alternative to antibiotics cannot be totally discarded, as further studies using the truncated form of this peptide are needed. © 2017 Blackwell Verlag GmbH.

Supplemental effect of varying L-cysteine concentrations on the quality of cryopreserved boar semen

Science.gov (United States)

Kaeoket, Kampon; Chanapiwat, Panida; Tummaruk, Padet; Techakumphu, Mongkol

2010-01-01

Cryopreservation is associated with the production of reactive oxygen species, which leads to lipid peroxidation of the sperm membrane and consequently a reduction in sperm motility and decreased fertility potential. The aim of this study was to determine the optimal concentration of L-cysteine needed for cryopreservation of boar semen. Twelve boars provided semen of proven motility and morphology for this study. The semen was divided into four portions in which the lactose-egg yolk (LEY) extender used to resuspend the centrifuged sperm pellet was supplemented with various concentrations of L-cysteine to reach 0 mmol L−1 (group I, control), 5 mmol L−1 (group II), 10 mmol L−1 (group III) and 15 mmol L−1 (group IV). Semen suspensions were loaded in straws (0.5 mL) and placed in a controlled-rate freezer. After cryopreservation, frozen semen samples were thawed and investigated for progressive motility, viability using SYBR-14/EthD-1 staining and acrosome integrity using FITC-PNA/EthD-1 staining. There was a significantly higher (P extender for improving the quality of frozen–thawed boar semen. PMID:20601963

EFFECT OF THE ADDITION OF SEMINAL PLASMA, VITAMIN E AND INCUBATION TIME ON POST-THAWED SPERM VIABILITY IN BOAR SEMEN

Directory of Open Access Journals (Sweden)

Alba G C Pech-Sansores

2011-09-01

Full Text Available The objective of the study was to evaluate the effect of seminal plasma (SP, vitamin E (VE, and incubation time on sperm viability of post-thawed boar semen. Thirty six ejaculates were used and allocated to four treatments: T1, semen + BTS (Belstville Thawing Solution + 10% SP; T2, semen + BTS + 200μg/ml VE; T3, semen + BTS + 10% SP + 200μg/ml VE; T4, semen + BTS (control. Motility (MOT, intact acrosomes (IA, membrane integrity (MI and mitochondrial activity (MA were evaluated, at 0 and 30 min after thawing. A split plot design was used and the data analyzed using a mixed model analysis of variance. There was a significant effect of SP and VE on IA and MI (P0.05. There was significant effect of incubation time on MOT (21.3 and 27.9% and IA (46.0 and 36.0%, at 0 and 30 min post-thawing (P <0.05. It is concluded that the vitamin E, seminal plasma or the combination of both, affected the thawed sperm traits here studied. Incubation time increased motility and intact acrosome percentage but did not affect the membrane integrity and mitochondrial activity.

Survivability of boar sperm stored under room temperature in extenders containing some natural products

OpenAIRE

Machebe,; Ugwu,Simeon; Akandi,Alafuro

2015-01-01

Alafuro Akandi,1 Simeon Ogochukwu Ugwu,1 Ndubuisi Samuel Machebe,1,2 1Department of Animal Science, University of Nigeria, Nsukka, Nigeria; 2Laboratory of Animal Reproduction, College of Agriculture, Kinki University, Nara, Japan Background: Semen extenders are used to create multiple insemination doses from a single ejaculate and contain buffers and nutrients suitable for storage of spermatozoa. The purpose of this study was to evaluate the survivability of boar sperm stored under room temp...

INFLUENCE OF CERTAIN BIOACTIVE PREPARATIONS ON THE DURATION OF BOAR SEMEN PRESERVATION

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V. HAREA

2009-05-01

Full Text Available The experiences were held on the boar sperm. There were studied the bioactive substances with the role of antioxidizer made at the Institute of Genetic of Science Academy of Republic of Moldova. The bioactive substances (GL-2 were used as a structure dilution GHTS what is used for boars sperm dilution with the concentration of 0,1 – 1%. The experimental researches showed that the studied substances were not toxic for sperm used in the structure of GHTS dilution with the concentration of 0,1-1 whit gave the possibility to increase the period of boar sperm stoking till 168 hours, keeping the sperms mobility at the level of standard of artificial insemination.

Influence of Boar and Semen Parameters on Motility and Acrosome Integrity in Liquid Boar Semen Stored for Five Days

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2002-01-01

Ninety ejaculates from a total of 76 AI boars were extended in Beltsville Thawing Solution (BTS). Boar identity, breed, weight of the ejaculate and sperm concentration were registered. Motility and acrosome integrity were assessed after storage at 16–18°C for 6, 30, 54, 78, and 102 h. Storage time had a significant influence on both motility (p boar (p boar (p boars and weight of the ejaculate did not influence the dependent variables. PMID:12071116

Effects of cationic antimicrobial peptides on liquid-preserved boar spermatozoa.

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Martin Schulze

Full Text Available Antibiotics are mandatory additives in semen extenders to control bacterial contamination. The worldwide increase in resistance to conventional antibiotics requires the search for alternatives not only for animal artificial insemination industries, but also for veterinary and human medicine. Cationic antimicrobial peptides are of interest as a novel class of antimicrobial additives for boar semen preservation. The present study investigated effects of two synthetic cyclic hexapeptides (c-WFW, c-WWW and a synthetic helical magainin II amide derivative (MK5E on boar sperm during semen storage at 16 °C for 4 days. The standard extender, Beltsville Thawing Solution (BTS containing 250 µg/mL gentamicin (standard, was compared to combinations of BTS with each of the peptides in a split-sample procedure. Examination revealed peptide- and concentration-dependent effects on sperm integrity and motility. Negative effects were more pronounced for MK5E than in hexapeptide-supplemented samples. The cyclic hexapeptides were partly able to stimulate a linear progressive sperm movement. When using low concentrations of cyclic hexapeptides (4 µM c-WFW, 2 µM c-WWW sperm quality was comparable to the standard extender over the course of preservation. C-WFW-supplemented boar semen resulted in normal fertility rates after AI. In order to investigate the interaction of peptides with the membrane, electron spin resonance spectroscopic measurements were performed using spin-labeled lipids. C-WWW and c-WFW reversibly immobilized an analog of phosphatidylcholine (PC, whereas MK5E caused an irreversible increase of PC mobility. These results suggest testing the antimicrobial efficiency of non-toxic concentrations of selected cyclic hexapeptides as potential candidates to supplement/replace common antibiotics in semen preservation.

Effects of cationic antimicrobial peptides on liquid-preserved boar spermatozoa.

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Schulze, Martin; Junkes, Christof; Mueller, Peter; Speck, Stephanie; Ruediger, Karin; Dathe, Margitta; Mueller, Karin

2014-01-01

Antibiotics are mandatory additives in semen extenders to control bacterial contamination. The worldwide increase in resistance to conventional antibiotics requires the search for alternatives not only for animal artificial insemination industries, but also for veterinary and human medicine. Cationic antimicrobial peptides are of interest as a novel class of antimicrobial additives for boar semen preservation. The present study investigated effects of two synthetic cyclic hexapeptides (c-WFW, c-WWW) and a synthetic helical magainin II amide derivative (MK5E) on boar sperm during semen storage at 16 °C for 4 days. The standard extender, Beltsville Thawing Solution (BTS) containing 250 µg/mL gentamicin (standard), was compared to combinations of BTS with each of the peptides in a split-sample procedure. Examination revealed peptide- and concentration-dependent effects on sperm integrity and motility. Negative effects were more pronounced for MK5E than in hexapeptide-supplemented samples. The cyclic hexapeptides were partly able to stimulate a linear progressive sperm movement. When using low concentrations of cyclic hexapeptides (4 µM c-WFW, 2 µM c-WWW) sperm quality was comparable to the standard extender over the course of preservation. C-WFW-supplemented boar semen resulted in normal fertility rates after AI. In order to investigate the interaction of peptides with the membrane, electron spin resonance spectroscopic measurements were performed using spin-labeled lipids. C-WWW and c-WFW reversibly immobilized an analog of phosphatidylcholine (PC), whereas MK5E caused an irreversible increase of PC mobility. These results suggest testing the antimicrobial efficiency of non-toxic concentrations of selected cyclic hexapeptides as potential candidates to supplement/replace common antibiotics in semen preservation.

Supplemental effect of different levels of taurine in Modena on boar semen quality during liquid preservation at 17°C.

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Li, Hao; Zhang, Xiao-Gang; Fang, Qian; Liu, Qi; Du, Ren-Rang; Yang, Gong-She; Wang, Li-Qiang; Hu, Jian-Hong

2017-11-01

Peroxidation damage induces sublethal injury to boar sperm during the storage process. Taurine has already been demonstrated to protect cells effectively from oxidant-induced injury. This study was aimed to evaluate the effect of different concentrations of taurine (0.5, 1, 5 and 10 mmol/L) in Modena diluent on boar sperm quality during liquid storage at 17°C. Ejaculates from sexually mature Duroc pigs were collected, pooled and preserved in the Modena containing different concentrations of taurine. Sperm motility, plasma membrane integrity, acrosome integrity, total antioxidative capacity (T-AOC) activity and malondialdehyde content (MDA) were examined every 24 h. Modena diluent containing taurine suppressed the reduction in sperm qualities during the process of liquid preservation compared with those of the control group. After 5 days of liquid preservation, the addition of taurine at 5 mmol/L had the optimal effect on survival time as well as maintenance of motility, plasma membrane integrity, acrosomal integrity, T-AOC activity and MDA content. These results may suggest the possibility that the proper addition of taurine to the semen extender improves the swine production system using artificial insemination by the suppressing of sperm damage and subsequent dysfunction during liquid preservation. © 2017 Japanese Society of Animal Science.

Is there a relationship between the chromatin status and DNA fragmentation of boar spermatozoa following freezing-thawing?

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Fraser, L; Strzezek, J

2007-07-15

In this study a radioisotope method, which is based on the quantitative measurements of tritiated-labeled actinomycin D ((3)H-AMD) incorporation into the sperm nuclei ((3)H-AMD incorporation assay), was used to assess the chromatin status of frozen-thawed boar spermatozoa. This study also tested the hypothesis that frozen-thawed spermatozoa with altered chromatin were susceptible to DNA fragmentation measured with the neutral comet assay (NCA). Boar semen was diluted in lactose-hen egg yolk-glycerol extender (L-HEY) or lactose ostrich egg yolk lipoprotein fractions-glycerol extender (L-LPFo), packaged into aluminum tubes or plastic straws and frozen in a controlled programmable freezer. In Experiment 1, the chromatin status and DNA fragmentation were measured in fresh and frozen-thawed spermatozoa from the same ejaculates. There was a significant increase in sperm chromatin destabilization and DNA fragmentation in frozen-thawed semen as compared with fresh semen. The proportions of spermatozoa labeled with (3)H-AMD were concurrent with elevated levels of sperm DNA fragmentation in K-3 extender, without cryoprotective substances, compared with L-HEY or L-LPFo extender. Regression analysis revealed that the results of the (3)H-AMD incorporation assay and NCA for frozen-thawed spermatozoa were correlated. Boars differed significantly in terms of post-thaw sperm DNA damage. In Experiment 2, the susceptibility of sperm chromatin to decondensation was assessed using a low concentration of heparin. Treatment of frozen-thawed spermatozoa with heparin revealed enhanced (3)H-AMD binding, suggesting nuclear chromatin decondensation. The deterioration in post-thaw sperm viability, such as motility, mitochondrial function and plasma membrane integrity, was concurrent with increased chromatin instability and DNA fragmentation. This is the first report to show that freezing-thawing procedure facilitated destabilization in the chromatin structure of boar spermatozoa, resulting in

Effect of Salvia miltiorrhiza polysaccharides on boar spermatozoa during freezing-thawing.

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Shen, Tao; Jiang, Zhong-Liang; Liu, Hong; Li, Qing-Wang

2015-08-01

Salvia miltiorrhiza polysaccharides (SMPs) were extracted from S. miltiorrhiza in this study. The aim of the present study was to evaluate the effect of SMP on the motility of boar sperm, including the antioxidant effect of SMP on boar sperm and the effect of SMP on the in vivo fertilizing ability of frozen-thawed boar sperm. Fifty ejaculates from 5 Swagger boars were collected and diluted with an extender, which contained 3% glycerol (v/v) with five concentrations of SMP (0.2, 0.4, 0.6, 0.8, and 1.0mg/mL). The semen was frozen in 0.25mL straws at 1.0×10(9) cells/mL. Sixty gilts were inseminated using fresh semen, frozen semen with 0.4mg/mL of SMP and frozen semen without SMP. The results indicate that the addition of SMP to the extender results in a higher percentage of motile sperm post-thaw (Pextender (Pboar sperm from peroxidative damage and increase sperm motility and litter size during the process of freezing-thawing. The optimal concentration of SMP for the frozen extenders in this study was determined to be 0.4mg/mL. Copyright © 2015. Published by Elsevier B.V.

Validation of the polysemen admixture on viability and acrosomal morphology of boar spermatozoa

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Ogbuewu IP

2007-07-01

Full Text Available Semen were collected using artificial vagina (AV, from 5 large white boars aged 2-2.5 years twice a week for 16 weeks in each of the two seasons, early rainy (ER and late rainy (LR seasons, to determine the effects of multiple semen pool admixture on the viability and acrosomal morphology. The semen qualities studied were sperm motility, live sperm and sperm concentration, while the acrosomal parameters includes normal apical ridge (NAR, damaged apical ridge (DAR, missing apical ridge (MAR and loose apical ridge (LAC. There were no significant (P>0.05 seasonal effects. Three-boar semen admixture gave the highest percentage NAR, motility, live sperm concentration and least DAR and LAC, although these were not significantly (P>0.05 different from the 2-boar semen admixture. The result of this study suggests that 3-boar semen admixture is most suitable for use in artificial insemination program.

Trehalose in glycerol-free freezing extender enhances post-thaw survival of boar spermatozoa

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ATHURUPANA, Rukmali; TAKAHASHI, Daisen; IOKI, Sumire; FUNAHASHI, Hiroaki

2015-01-01

Cryopreservation of boar semen is still considered suboptimal due to lower fertility as compared with fresh samples when glycerol, a permeating cryoprotectant, is used. Trehalose is a non-permeable cryoprotectant and nonreducing disaccharide known to stabilize proteins and biologic membranes. The aim of this study was to evaluate the cryosurvival and in vitro penetrability of boar spermatozoa when glycerol was replaced with trehalose in a freezing extender. Ejaculated Berkshire semen samples were diluted in egg yolk-based freezing extender containing glycerol (100 mM) or trehalose (0, 50, 100, 150, 200 and 250 mM) and cryopreserved using a straw freezing procedure. Thawed samples were analyzed for motility, viability, mitochondrial membrane potential (MMP), and acrosome integrity. In experiment 2, penetrability of spermatozoa cryopreserved with 100 mM glycerol or trehalose was examined. Replacement of cryoprotectant glycerol (100 mM) with trehalose had no effect on sperm viability, but replacing it with 100 mM trehalose improved motility, MMP and acrosome integrity significantly. Sperm motility and MMP were considerably higher in 100 mM trehalose, whereas the acrosome integrity was substantially higher in 100–250 mM trehalose. The in vitro penetration rate was also significantly higher in spermatozoa cryopreserved with trehalose (61.3%) than in those cryopreserved with glycerol (43.6%). In conclusion, 100 mM non-permeable trehalose can be used to replace glycerol, a permeating cryoprotectant, for maintenance of better post-thaw quality of boar spermatozoa. PMID:25754239

Inter- and intra-breed comparative study of sperm motility and viability in Iberian and Duroc boar semen during long-term storage in MR-A and XCell extenders.

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Martín-Hidalgo, D; Barón, F J; Robina, A; Bragado, M J; Llera, A Hurtado de; García-Marín, L J; Gil, M C

2013-06-01

During boar semen liquid preservation, extender is one of the factors that influence storage tolerance of spermatozoa. However, there are few studies about intra-breed variation in the preservation of semen quality during storage in different extenders. Similarly, boar breed is generally not considered a possible factor influencing variation in the semen storage tolerance in a particular extender. The aim of this study was to compare boar semen storage potential, in terms of the ability to maintain sperm viability and motility, of two currently used long-term extenders, MR-A and XCell. Extended semen from two breeds, Iberian and Duroc that had been stored at 17°C for up to 7 days was used. Intra- and inter-breed effect was studied. On Days 1, 4 and 7 (Day 0=day of semen collection), motility parameters and the percentage of total motile sperm and progressively motile sperm using a CASA system was evaluated. Viability (SYBR-14/PI) was evaluated by flow cytometry. Within each breed and for each storage day, there were differences between extenders, although semen tolerance to preservation was more influenced by the extender in the Iberian than in the Duroc breed. Neither breed nor extender influenced the percentage of viable spermatozoa during the storage time. Moreover, differences in motility parameters were observed between breeds, although the differences were greater when the XCell extender was used. In conclusion, both extender and breed influence motility characteristics of liquid-stored boar semen, so both aspects have to be considered in the design of comparative studies about stored boar semen quality from different breeds or with different extenders. Further studies are needed to corroborate these findings. Copyright © 2013 Elsevier B.V. All rights reserved.

Crossreactivity of boar sperm monoclonal antibodies with human ...

African Journals Online (AJOL)

Monoclonal antibodies against the head (H mabs) and tail (Tmabs) of boar spermatozoa were produced. Spermatozoa from boar, stallion, bull, human, ram, goat and rabbit were independently incubated with the monoclonal antibodies and later stained by immunofluorescence method. There were positive reactions of the ...

Comparative study on five different commercial extenders for boar semen.

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Vyt, P; Maes, D; Dejonckheere, E; Castryck, F; Van Soom, A

2004-02-01

Increasing interest in a longer preservation of diluted boar sperm raises questions in the field concerning the choice of the extender. The aim of this study was to evaluate the longevity of boar sperm extended in currently used commercial semen extenders. Three long-term extenders and two short-term extenders were compared for different semen quality parameters that can be assessed under routine laboratory conditions. Sperm morphology, motility, pH and bacteriological contamination were investigated during a 7-day period. The number of dead spermatozoa did not differ significantly among the extenders (p > 0.05). Sperm motility was not only related with storage period but most of all with pH, especially in long-term extenders. Differences between the different extenders were prominent (p extender showed good motility during the whole test period. In all cases, the pH of the extended semen increased by 0.3-0.5 in the first days of storage and was significantly correlated with a decrease in motility. Bacteriological quality had no significant influence on motility or pH of the semen. In conclusion, we can state that in both short-term extenders and in only one long-term extender, sperm longevity, as evaluated by the parameters used in this study, was sufficient during the preservation period. To preserve the quality of diluted boar semen during long-term storage, the choice of the long-term extender is important. In addition, the monitoring of the pH of extended boar semen in our study emphasizes the importance of the buffering capacity of semen extenders.

Cryopreservation of Iberian pig spermatozoa. Comparison of different freezing extenders based on post-thaw sperm quality.

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De Mercado, Eduardo; Rodríguez, Ana; Gómez, Emilio; Sanz, Elena

2010-03-01

The aim of this study was to evaluate the cryoprotective effect of different freezing extenders against cryopreservation injuries on Iberian boar sperm. The sperm-rich fraction was collected and pooled from six sexually mature Iberian boars, and was frozen in different extenders containing glucose, lactose or fructose as sugar source and including Orvus ES Paste only in the freezing extender-2 (Glucose; Lactose and Fructose) or in both freezing extenders (Glucose2; Lactose2 and Fructose2). During the cryopreservation process, the supernatant was removed after the centrifugation step, then was extended with freezing extender-1 for the equilibration period and with freezing extender-2 immediately before freezing. Post-thaw sperm characteristics, such as plasma membrane integrity (SYBR-14/PI), mitochondrial function (Rhodamine 123) and acrosome integrity (NAR), were monitored. Overall sperm motility and the individual kinematic parameters of motile spermatozoa (assessed by the computer-aided sperm analysis system Sperm Class Analyzer [SCA]) were recorded in the different experimental treatments. Measurements were taken at 30 and 150 min post-thaw. The state of the acrosome after thawing did not show significant differences between the freezing extenders studied. Freezing-thawing caused a significant decrease (Pextenders. Furthermore, spermatozoa frozen with Orvus ES Paste in both freezing extenders exhibited lower (Pextender. The spermatozoa frozen with the Lactose extender and with Orvus ES Paste only in the second freezing extender showed a better evolution of the motility and kinematic characteristics (Pextenders studied in the present experiment affected the quality of frozen-thawed semen in Iberian boar.

A comparison of mycotoxin adsorbents and their effects on some selected parameters of boar semen

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Zdeněk Tvrdoň

2006-01-01

Full Text Available Effect of two mycotoxin adsorbents with different mechanisms of action were followed in 180 purebred and terminal combination boars reared in artificial insemination stations. The following parameters of boar sperm quality were investigated: volume in millilitres, concentration and motility, and numbers of pathological spermatozoa. When processing the boar semen, the dilution ration and numbers of AI doses were recorded. Compared were two preparations: in case of Preparation 1 the number of evaluated ejaculates was 1,037 while in case of Preparation 2 altogether 1,109 ejaculates were evaluated. Boars receiving diets with Preparation 1 produced more voluminous ejaculates (by 4.1 ml and their concentration of spermatozoa was also higher (by 39 thous./ml; P ≤ 0.001. Dilution parameters were better as well and numbers produced of AI doses were also higher. In case of Preparation 2 the motility of spermatozoa was a higher while the numbers of sperms were lower. The obtained results demonstrated that a suitable adsorbent can show a positive effect on both quantitative and qualitative parameters of boar sperm.

Characterization of the functions and proteomes associated with membrane rafts in chicken sperm.

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Ai Ushiyama

Full Text Available Cellular membranes are heterogeneous, and this has a great impact on cellular function. Despite the central role of membrane functions in multiple cellular processes in sperm, their molecular mechanisms are poorly understood. Membrane rafts are specific membrane domains enriched in cholesterol, ganglioside GM1, and functional proteins, and they are involved in the regulation of a variety of cellular functions. Studies of the functional characterization of membrane rafts in mammalian sperm have demonstrated roles in sperm-egg binding and the acrosomal reaction. Recently, our biochemical and cell biological studies showed that membrane rafts are present and might play functional roles in chicken sperm. In this study, we isolated membrane rafts from chicken sperm as a detergent-resistant membranes (DRM floating on a density gradient in the presence of 1% Triton X-100, and characterized the function and proteomes associated with these domains. Biochemical comparison of the DRM between fresh and cryopreserved sperm demonstrated that cryopreservation induces cholesterol loss specifically from membrane rafts, indicating the functional connection with reduced post-thaw fertility in chicken sperm. Furthermore, using an avidin-biotin system, we found that sperm DRM is highly enriched in a 60 KDa single protein able to bind to the inner perivitelline layer. To identify possible roles of membrane rafts, quantitative proteomics, combined with a stable isotope dimethyl labeling approach, identified 82 proteins exclusively or relatively more associated with membrane rafts. Our results demonstrate the functional distinctions between membrane domains and provide compelling evidence that membrane rafts are involved in various cellular pathways inherent to chicken sperm.

Head (H) And Tail (T) Monoclonal Antibodies of Boar Spermatozoa ...

African Journals Online (AJOL)

Semen collected from three boars were pooled and the sperm cells separated from the semen, washed, sonicated and mixed with equal volume of Freund's adjuvant. The mixture of sperm with complete Freund's adjuvant was used to immunize five mice of the BALB/C strain intraperitoneally on Day O. The sonicated sperm ...

Comparison of semen variables, sperm DNA damage and sperm membrane proteins in two male layer breeder lines.

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M, Shanmugam; T R, Kannaki; A, Vinoth

2016-09-01

Semen variables are affected by the breed and strain of chicken. The present study was undertaken to compare the semen quality in two lines of adult chickens with particular reference to sperm chromatin condensation, sperm DNA damage and sperm membrane proteins. Semen from a PD3 and White Leghorn control line was collected at 46 and 47 weeks and 55 weeks of age. The semen was evaluated for gross variables and sperm chromatin condensation by aniline blue staining. Sperm DNA damage was assessed by using the comet assay at 47 weeks of age and sperm membrane proteins were assessed at 55 weeks of age. The duration of fertility was studied by inseminating 100 million sperm once into the hens of the same line as well as another line. The eggs were collected after insemination for 15days and incubated. The eggs were candled on 18th day of incubation for observing embryonic development. The White Leghorn control line had a greater sperm concentration and lesser percentage of morphologically abnormal sperm at the different ages where assessments occurred. There was no difference in sperm chromatin condensation, DNA damage and membrane proteins between the lines. Only low molecular weight protein bands of less than 95kDa were observed in samples of both lines. The line from which semen was used had no effect on the duration over which fertility was sustained after insemination either when used in the same line or another line. Thus, from the results of the present study it may be concluded that there was a difference in gross semen variables between the lines that were studied, however, the sperm chromatin condensation, DNA damage, membrane proteins and duration over which fertility was sustained after insemination did not differ between the lines. Copyright © 2016 Elsevier B.V. All rights reserved.

Changing rooster sperm membranes to facilitate cryopreservation

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Cryopreservation damages rooster sperm membranes. Part of this damage is due to membrane transitioning from the fluid to the gel state as temperature is reduced. This damage may be prevented by increasing membrane fluidity at low temperatures by incorporating cholesterol or unsaturated lipids into t...

Effects of dietary supplementation with an organic source of selenium on characteristics of semen quality and in vitro fertility in boars.

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Speight, S M; Estienne, M J; Harper, A F; Crawford, R J; Knight, J W; Whitaker, B D

2012-03-01

Semen characteristics in boars fed organic or inorganic sources of Se were assessed in 3 experiments. Crossbred boars were randomly assigned at weaning to 1 of 3 dietary treatments: I) basal diets with no supplemental Se (control), II) basal diets with 0.3 mg/kg of supplemental Se from an organic source (Sel-Plex, Alltech Inc., Nicholasville, KY), and III) basal diets supplemented with 0.3 mg/kg of supplemental Se from sodium selenite (Premium Selenium 270, North American Nutrition Co. Inc., Lewisburg, OH). For Exp. 1, semen was collected from boars (n = 10/dietary treatment) on 5 consecutive days at 15 mo of age. Effects of treatment × day were detected for the proportions of progressively motile (P = 0.02) and rapidly moving (P = 0.03) spermatozoa, and measures of sperm velocity, including path velocity of the smoothed cell path (P = 0.05) and average velocity measured in a straight line from the beginning to the end of the track (P = 0.05). Negative effects of day of semen collection on sperm motility were least pronounced in boars fed Sel-Plex. Experiment 2 was conducted when boars were 17 mo of age, and semen was collected (n = 10 boars/dietary treatment), diluted in commercially available extenders, and stored at 18°C for 9 d. Effects of treatment × day were detected for percentages of motile (P = 0.01) and static (P = 0.01) spermatozoa, amplitude of lateral head displacement (P = 0.02), frequency with which the sperm track crossed the sperm path (P = 0.04), straightness (P = 0.01), and average size of all sperm heads (P = 0.03). In general, sperm cells from boars fed Sel-Plex were better able to maintain motility during liquid storage compared with boars fed sodium selenite. For Exp. 3, semen was collected from boars (n = 6/dietary treatment) at 23 mo of age, and spermatozoa were evaluated at d 1 and 8 after semen collection using in vitro fertilization procedures. There was a tendency for an effect (P = 0.11) of dietary treatment on fertilization rate

Long-term liquid storage and reproductive evaluation of an innovative boar semen extender (Formula12®) containing a non-reducing disaccharide and an enzymatic agent.

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Bresciani, Carla; Bianchera, Annalisa; Bettini, Ruggero; Buschini, Annamaria; Marchi, Laura; Cabassi, Clotilde Silvia; Sabbioni, Alberto; Righi, Federico; Mazzoni, Claudio; Parmigiani, Enrico

2017-05-01

There are no reports of saccharolytic enzymes being used in the preparation of formulations for animal semen extenders. In the present study, the use of an innovative semen extender (Formula12 ® ) in the long-term liquid storage of boar semen at 17°C was evaluated. The formulation included use of a disaccharide (sucrose) as the energy source precursor coupled to an enzymatic agent (invertase). The innovative extender was evaluated and compared in vitro to a commercial extender (Vitasem LD ® ) for the following variables: Total Motility (TM), Forward Progressive Motility (FPM), sperm morphology, membrane integrity, acrosome integrity, and chromatin instability. Boar sperm diluted in Formula12 ® and stored for 12 days at 17°C maintained a commercially acceptable FPM (>70%). Using the results from the in vitro study, an AI field trial was performed. A total of 170 females were inseminated (135 with Formula12 ® and 35 with Vitasem LD ® ). The pregnancy rates were 97.8% compared with 91.4%, and the farrowing rates were 96.3% compared with 88.6% when Formula12 ® and Vitasem LD ® were used, respectively. The mean number of piglets born/sow were 14.92±0.46 compared with 13.83±0.70, and the number of piglets born alive/sow were 14.07±0.46 compared with 12.12±0.70 (Pextender allowed for meeting the metabolic requirements of boar sperm during storage at 17°C. It is suggested that there was a beneficial effect on fertilizing capacity of boar sperm in the female reproductive tract with use of these technologies. Copyright © 2017 Elsevier B.V. All rights reserved.

Effect of freezing and thawing rates on the post-thaw viability of boar spermatozoa frozen in FlatPacks and Maxi-straws.

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Eriksson, B M; Rodriguez-Martinez, H

2000-11-01

The effects of different freezing and thawing rates on the post-thaw motility and membrane integrity of boar spermatozoa, processed as split samples in Maxi-straws or flat PET-plastic packages (FlatPack) were studied. A programmable freezing device was used to obtain freezing rates of either 20, 50 or 80 degrees C/min. Thawing of the samples was performed in a bath of circulating water; for 40s at 50 degrees C or 27s at 70 degrees C for Maxi-straws and 23s at 35 degrees C, 13s at 50 degrees C or 8s at 70 degrees C for the FlatPacks. Sperm motility was assessed both visually and with a computer assisted semen analysis (CASA) apparatus, while plasma membrane integrity was assessed using the fluorescent probes Calcein AM and ethidium homodimer-1. Temperature changes during freezing and thawing were monitored in both forms of packaging. Values for motile spermatozoa, sperm velocity and lateral head displacement variables were significantly (pstraws, with superior results at higher thawing rates. Freezing at 50 degrees C/min yielded better motility than 20 or 80 degrees C/min, although the effect was rather small. Neither freezing rate nor thawing rate had any effect on membrane integrity (p>0.05). A significant boar effect was seen for several parameters. The most striking difference in temperature courses between containers was a 4-5-fold lowering of the thawing rate, between -20 and 0 degrees C, in the center of the Maxi-straw, compared with the FlatPack. This is apparently due to the insulating effect of the thawed water in the periphery of the Maxi-straw. The improvement in sperm motility seen when using the FlatPack appears to be related to the rapid thawing throughout the sample, which decreases the risk of cell damage due to recrystallization during thawing. Since sperm motility patterns have been reported to be correlated with fertility both in vitro and in vivo it is speculated that the use of the FlatPack might improve the results when using frozen

Effect of Gynostemma Pentaphyllum polysaccharide on boar spermatozoa quality following freezing-thawing.

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Hu, Jian-Hong; Li, Qing-Wang; Zhang, Ting; Jiang, Zhong-Liang

2009-12-01

Gynostemma Pentaphyllum Polysaccharide (GPP) was added at concentrations of 0.25, 0.5, 1.0, 1.5 and 2.0 mg/ml to the extenders used to freeze boar semen and its effects on the quality of frozen-thawed sperm were assessed. The sperm motility was significantly higher in the extenders containing 0.25 and 0.5 mg/ml GPP, as compared to other groups (Pextender supplemented with 0.5 mg/ml GPP favored the highest intact membrane and intact acrosome percentages in comparison with other groups (Pextender supplemented with 0.25 and 0.5 mg/ml GPP significantly improved SOD levels, compared to other groups (P>0.05). However, the extenders supplemented with GPP did not cause significant differences in levels of CAT and GSH-Px, compared to the control (P>0.05). In summary, GPP exhibited a dose-related response and the lower concentration produced greater protective effect. According to the standard semen quality parameters and antioxidant activities measured in this study, the concentration of 0.5 mg/ml GPP caused a beneficial cryoprotective effects on the quality of frozen-thawed boar semen. It is proposed that an extender containing 0.5 mg/ml GPP could be used as cryoprotective medium of better efficiency.

Antioxidant effects of cultured wild ginseng root extracts on the male reproductive function of boars and guinea pigs.

Science.gov (United States)

Yun, Suk Jun; Bae, Gui-Seck; Park, Jae Hawn; Song, Tae Ho; Choi, Ahreum; Ryu, Buom-Yong; Pang, Myung-Geol; Kim, Eun Joong; Yoon, Minjung; Chang, Moon Baek

2016-07-01

The main objective of this study was to investigate the effects of cultured wild ginseng root extracts (cWGRE) on the sperm of boars and the reproductive system of guinea pigs. Firstly, semen collected from boars (n=10) were incubated in 38°C for 1h with xanthine and xanthine oxidase to generate ROS. The cWGRE was added to the sperm culture system to test its antioxidant effect on the boar sperm. The amount of Reactive Oxygen Species (ROS) was measured by a chemiluminescence assay using luminol. The results indicated that the addition of cWGRE to boar sperm culture inhibited xanthine and xanthine oxidase-induced ROS concentrations. Treatment with cWGRE also had a positive effect on maintaining sperm motility. Effects of cWGRE administration on vitamin C-deficient guinea pigs were further investigated. Hartley guinea pigs (n=25) at 8 weeks of age were randomly divided into five groups. With the exception of the positive control group, each group was fed vitamin C-deficient feed for 21days (d). Respective groups were also orally administered cWGRE, ginseng extract, or mixed ginsenosides for 21 days. In comparison to the control group, oral administration of cWGRE reduced (Pproduction. Copyright © 2016 Elsevier B.V. All rights reserved.

Liquid Storage of Boar Semen Using Commercial Extenders

OpenAIRE

Liviu BOGDAN; Mihai CENARIU; Mihai BORZAN; Simona CIUPE; Lehel SZABO; Emoke PALL

2018-01-01

The research was focused on the modern evaluation of boar semen parameters, using computer assisted sperm analysis (CASA), before and after liquid storage at 15ºC. Semen was collected from 15 sexually mature boars by manual stimulation. Macroscopical and microscopical evaluation of semen was performed, followed by a detailed CASA analysis of all ejaculates. Subsequently, semen was diluted using 4 different extenders (Semtest, Androstar, MIII and Cronos) and stored at 15ºC for 24 hours. Next, ...

Effect of natural betaine on estimates of semen quality in mature AI boars during summer heat stress.

Science.gov (United States)

Cabezón, F A; Stewart, K R; Schinckel, A P; Barnes, W; Boyd, R D; Wilcock, P; Woodliff, J

2016-07-01

This study evaluated the effect of supplemental dietary betaine at three concentrations (0.0%, 0.63% and 1.26%) on semen characteristics, quality and quality after storage on boars. The trial was conducted between 22 July and 1 October 2014 in a boar stud located in Oklahoma. Boars were blocked by age within genetic line and randomly allotted to receive 0% (CON, n (line T)=22, n (line L)=10), 0.63% (BET-0.63%, n (line T)=21, n (line L)=6) or 1.26% (BET-1.26%, n (line T)=23, n (line L)=7). The diets containing betaine were fed over 10 weeks, to ensure supplemental betaine product (96% betaine) daily intakes of 16.34 and 32.68g, for the BET-0.63% and BET-1.26% diets, respectively. Serum homocysteine concentrations were less for animals with betaine treatments (P=0.016). Rectal temperatures of the boars were unaffected by betaine diets. Betaine tended to increase total sperm in the ejaculates when collectively compared with data of the control animals (P=0.093). Sperm morphology analysis indicated there was a greater percent of sperm with distal midpiece reflex (P=0.009) and tail (P=0.035) abnormalities in boars fed the BET-1.26% than boars fed the BET-0.63% diet. Betaine concentration in the seminal plasma was greater in boars with betaine treatments, with animals being fed the 0.63% and 1.26% diets having 59.2% and 54.5% greater betaine concentrations in seminal plasma as compared with boars of the control group (P=0.046). In conclusion, betaine supplementation at 0.63% and 1.26% tended to increase sperm concentration in the ejaculates by 6% and 13%, respectively, with no negative impacts on semen quality when 0.63% of betaine was included in the diet. Copyright © 2016 Elsevier B.V. All rights reserved.

Effects of DHA-enriched hen egg yolk and L-cysteine supplementation on quality of cryopreserved boar semen.

Science.gov (United States)

Chanapiwat, Panida; Kaeoket, Kampon; Tummaruk, Padet

2009-09-01

The objective of the present study was to determine the effects of docosahexaenoic acid (DHA)-enriched hen egg yolks and L-cysteine supplementation on the qualities of the cryopreserved boar semen. A total of 15 ejaculates from 5 Pietrain boars were divided into 4 groups according to the compositions of the freezing extenders used, that is, normal hen egg yolk (group I), DHA-enriched hen egg yolk (group II), normal hen egg yolk with 5 mmol L(-1) of cysteine supplementation (group III) and DHA-enriched hen egg yolk with 5 mmol L(-1) of cysteine supplementation (group IV). The semen was cryopreserved using controlled rate freezer and was thawed at 50 degrees C for 12 s. Progressive motility, sperm viability, acrosome integrity and functional integrity of sperm plasma membrane of the post-thawed semen were evaluated. The supplementation of L-cysteine in the freezing extender alone (group III) improved progressive motility (P semen qualities (P > 0.05). In conclusion, the supplementation of antioxidant L-cysteine alone or in combination with DHA-enriched hen egg yolk significantly improved the post-thawed semen qualities, especially progressive motility and acrosome integrity.

Effects of commercial selenium products on glutathione peroxidase activity and semen quality in stud boars

Science.gov (United States)

The aim of this study was to determine how dietary supplementation of inorganic and organic selenium affects selenium concentration and glutathione peroxidase activity in blood and sperm of sexually mature stud boars. Twenty-four boars of the Large White, Landrace, Pietrain, and Duroc breeds of opt...

Dimethylformamide is not better than glycerol for cryopreservation of boar semen.

Science.gov (United States)

Malo, C; Gil, L; Cano, R; Martínez, F; García, A; Jerez, R A

2012-05-01

To improve the boar sperm cryopreservation process, the influence of the sugar (lactose, trehalose) source and the cryoprotectant [glycerol, dimethylformamide (DMF)] on the success of freezing was investigated. Sperm samples were frozen in one of six extenders: lactose plus 3% glycerol (LG); lactose plus 1.5% glycerol and 1.5% DMF (LGD); lactose plus 3% DMF (LD); trehalose plus 3% glycerol (TG); trehalose plus 1.5% glycerol and 1.5% DMF (TGD); trehalose plus 3% DMF (TD). Effects on motility, viability, acrosome integrity and hypoosmotic test (HOST) were measured. The results showed that extender containing 3% glycerol retained the highest motility percentages. In regard to viability and acrosome integrity, all extenders yielded similar rates except for the decreasing values of TD. Endosmosis was diminished in TD and LD at 2 h (P = 0.0018), as compared with the others. The results of the study demonstrated that the use of DMF as a cryoprotectant adversely affected boar sperm quality after cryopreservation. © 2011 Blackwell Verlag GmbH.

Defined Combinations of Cryomedia and Thawing Extenders Influence the Viable X-Y Boar Sperm Ratio in Vitro.

Science.gov (United States)

Korchunjit, W; Kaeoket, K; Kitiyanant, Y; Taylor, J; Wongtawan, T

It is believed that plasma membrane X- and Y-chromosome bearing sperm are different; therefore the freezing and thawing process may affect X- and Y-sperm differently. The objective of this study was to investigate the effect of cryomedia and thawing extenders on the survival of X and Y-sperm. Three different cryomedia and thawing extenders were compared. Viable motile sperm were separated using a swim-up technique. Real-time PCR was used to identify the sperm type. Using CryoA for freezing and Beltsville-Thawing-Solution (BTS) as the thawing extender yielded significantly higher numbers of viable motile Y sperm (64 percent) than control (48 percent) (P semen freezing with CryoC and thawing with Androstar Plus gave a significantly lower number of viable motile Y sperm (32 percent) than control (51 percent). Our results revealed that defined combinations of cryomedia and thawing extenders significantly altered the survival ratio of frozen-thawed X-Y sperm in vitro, which has potential implications for artificial insemination.

The effects of three extenders on refrigerated boar semen

African Journals Online (AJOL)

sofiateixeira

2015-03-19

Mar 19, 2015 ... South African Journal of Animal Science 2015, 45 (No. .... influence of various extenders on boar sperm survival (Vyt et al., 2004), especially quality (Huo ..... Boe-Hansen, G.B., Ersboll, A.K., Greve, T. & Christensen, P., 2005.

Use of refractometry as a new management tool in AI boar centers for quality assurance of extender preparations.

Science.gov (United States)

Schulze, M; Rüdiger, K; Jung, M; Grossfeld, R

2015-01-01

A study was performed to see if refractometry can be used as a new quality control tool for boar semen extenders. For this the refractive index and osmolality of BTS extender concentrations (EC) were recorded in 10%-steps from 50% to 150% and 200% of the correct amount. Twelve boar ejaculates were evaluated for semen quality. The refractive index for the correctly prepared extender was 4.6±0.0°Bx, corresponded to 316±16mOsmkg(-1), and correlated highly with osmolality (r=0.99; P<0.001). Total sperm motility with 100% EC differed significantly from ≤70% EC (P<0.001) and 200% EC (P<0.001) on day 1 (d1) and d4, respectively. The percentage of motile spermatozoa in a thermoresistance test on d2 showed a significant drop using ≤70% EC (P=0.047) and ≥140% EC (P=0.004). Secondary apical ridge defects were significantly higher using 50% EC (P<0.001) and ≥150% EC (P=0.032) compared to 100% EC, respectively. An increased number of coiled tails were observed using ≤60% EC (P<0.001). Percentages of spermatozoa with intact membranes on d2 resulted in a significant decrease using 50% EC (P<0.001) and ≥150% EC (P=0.005), respectively. The mean percentage of PI negative spermatozoa with active mitochondria on d2 showed a significant difference using ≤60% EC (P=0.016) and ≥140% EC (P<0.001) compared to 100% EC, respectively. Boar sperm quality is affected by inexact extender preparation. The refractive-index is an indicator of osmolality and may be used to verify semen extender preparation. The sensitivity is sufficient to detect deviations from correct extender preparation before negative effects on sperm quality occur. Copyright © 2014 Elsevier B.V. All rights reserved.

Sperm DNA damage in relation to lipid peroxidation following ...

African Journals Online (AJOL)

This study investigated the relationships between lipid peroxidation (LPO) and sperm DNA damage following freezing-thawing of boar semen in different extenders. The comet assay was used to measure the extent of sperm DNA damage in a cryoprotectant-free extender or in cryoprotectant-based extenders after single ...

The Effect of Curcumin on Intracellular pH (pHi), Membrane Hyperpolarization and Sperm Motility.

Science.gov (United States)

Naz, Rajesh K

2014-04-01

Curcumin has shown to affect sperm motility and function in vitro and fertility in vivo. The molecular mechanism(s) by which curcumin affects sperm motility has not been delineated. Since modulation of intracellular pH (pHi) and plasma membrane polarization is involved in sperm motility, the present study was conducted to investigate the effect of curcumin on these sperm (human and murine) parameters. The effect of curcumin on sperm forward motility was examined by counting percentages of forward moving sperm. The effect of curcumin on intracellular pH (pHi) was measured by the fluorescent pH indicator 2,7-bicarboxyethyl-5,6-carboxyfluorescein-acetoxymethyl ester (BCECF-AM). The effect of curcumin on plasma membrane polarization was examined using the fluorescence sensitive dye bis (1,3-dibarbituric acid)-trimethine oxanol [DiBAC4(3)]. Curcumin caused a concentration-dependent (ppHi) in both human and mouse sperm. Curcumin induced significant (ppHi and membrane polarization that affect sperm forward motility. These exciting findings will have application in deciphering the signal transduction pathway involved in sperm motility and function and in development of a novel non-steroidal contraceptive for infertility.

Comparing sugar type supplementation for cryopreservation of boar semen in egg yolk based extender.

Science.gov (United States)

Malo, C; Gil, L; Gonzalez, N; Cano, R; de Blas, I; Espinosa, E

2010-08-01

Cryopreservation of boar semen is still considered suboptimal due to lower fertility when compared to fresh semen. The aim of this study was to evaluate the effects of the addition of different sugars (lactose, trehalose and glucose) on boar spermatozoa cryopreserved in an egg yolk based extender. Ejaculates were collected from a boar previously selected and semen samples were processed using the straw freezing procedure. In experiment 1, subsamples of semen were frozen in three different extenders: recommended lactose egg yolk extender (LEY); trehalose egg yolk extender (TEY) and glucose egg yolk extender (GEY). Sperm quality was assessed for motility, viability, acrosome integrity and hypoosmotic swelling test response upon collection, after freezing and thawing and then every hour for 3h. Results showed that total motility at 1 and 3h, progressive motility at 3h, positive hypoosmotic response at 2 and 3h and acrosome integrity at all times were significantly improved when trehalose was added to the extender. In experiment 2, sugar influence was also demonstrated in vitro fertilization. A total of 1691 oocytes were in vitro matured and inseminated with frozen-thawed sperm at 2000:1 sperm:oocyte ratio and coincubated for 6h. Presumptive zygotes were cultured in NCSU-23 medium to assess fertilization parameters and embryo development. Both penetration and monospermy rates were significantly higher for trehalose frozen semen. A significant increase was observed in efficiency and blastocyst formation rates from TEY to the other groups. Our results demonstrated that trehalose extender enhances spermatozoa viability and its in vitro fertilization parameters in boar ejaculates with good sperm freezability. Further studies are necessary to assess the impact of sugars on the entire population. (c) 2010 Elsevier Inc. All rights reserved.

Egg yolk and glycerol requirements for freezing boar spermatozoa treated with methyl β-cyclodextrin or cholesterol-loaded cyclodextrin.

Science.gov (United States)

Blanch, Eva; Tomás, Cristina; Hernández, Marta; Roca, Jordi; Martínez, Emilio A; Vázquez, Juan M; Mocé, Eva

2014-04-24

Egg yolk (EY) and glycerol are common constituents of extenders used for sperm cryopreservation. It has been demonstrated that using cholesterol-loaded cyclodextrins (CLC) improves sperm cryosurvival in several species. However, standard freezing extenders might not be the most appropriate for CLC-treated sperm. This study evaluated the EY and glycerol requirements for freezing CLC-treated boar spermatozoa. Semen samples from 34 ejaculates coming from 4 boars were used. Each ejaculate was split into three aliquots: one was used untreated (control), and the other two were treated with 1 mg of CLC or methyl-β-cyclodextrin/120 × 10(6) sperm for 15 min at 22 C prior to cryopreservation. Our results indicated that reducing the concentration of EY was detrimental for sperm viability after thawing (31.57 ± 2 vs. 19.89% ± 2 for 20 and 10% EY, respectively; P semen treated with CLC. On the other hand, it was observed that the traditional concentration of glycerol (3%) was not the appropriate for freezing CLC-treated sperm (61.10 ± 3 vs. 47.87% ± 3 viable sperm for control and CLC-treated sperm, respectively; P extenders for CLC-treated sperm. Nevertheless, additional studies will be needed to evaluate alternative cryoprotectants and to determine the effect of high glycerol concentrations on sperm functionality.

Effects of different cryoprotectants and freezing methods on post-thaw boar semen quality.

Science.gov (United States)

Yang, Chung-Hsun; Wu, Ting-Wen; Cheng, Feng-Pang; Wang, Jiann-Hsiung; Wu, Jui-Te

2016-03-01

The current study aimed to investigate the effects of different concentrations of glycerol (0%, 1%, 2%, 3%, and 5%) and dimethylacetamide (DMA: 0%, 1%, 3%, and 5%) on post-sperm quality characteristics following semen freezing in dry ice (D) or liquid nitrogen (N). Semen was collected from Duroc boars and was allocated to 32 treatment groups for cryopreservation. Analysis of post-thaw semen quality and fertility after artificial insemination (AI) was used to examine the combinatorial effects of different treatments. The best scores for post-thaw sperm motility, sperm viability, and sperm acrosomal integrity were observed in semen frozen in: (a) dry ice in the presence of 5% glycerol and no DMA (16D-treatment); (b) dry ice in the presence of 3% glycerol and no DMA (9D-treatment); and (c) liquid nitrogen in the presence of 3% glycerol and 1% DMA (10N-treatment), with no significant difference observed among these three treatments. The farrowing rates after AI with post-thawed semen after 9D- and 10N-treatments were 33% and 50%, respectively. To summarize, the results of the present study indicated that the freezing extender containing 3% glycerol in combination with the straw-freezing method using dry ice produced the best post-thaw quality parameters of boar semen. Combinations of glycerol and DMA did not enhance the cryosurvival of boar spermatozoa. Copyright © 2016 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

COMPARISON OF CD46 EXPRESSION ON THE INNER ACROSOMAL MEMBRANE OF SPERMS FROM NORMOSPERMIC AND ASTHENOSPERMIC INDIVIDUALS

Directory of Open Access Journals (Sweden)

M NASR ESFAHANI

2003-03-01

Full Text Available Introduction: CD46 is a membrane cofactor protein (MCP of complement system wich is present on the membrane of all somatic cells except RBC. It is also present on the inner acrosmal membrane of human sperm. Thus, the aim of this study was to compare the expression of this prote, in on the inner acrosmal membrane of sperms from normospermic and asthenospermic individuals. Method: Semen from 6 normospermic and 17 asthenospermic individuals were examined for CD46 expression. After solublization of sperms in solublizing detergent, the solublized sperm membrane was separated from the rest of cell organelles by centrifugation. Solublized sperm membrane were divide to equal parts and SOS-PAGE gel was canied out in paired on the same gel for each sample. Western blot was carried out on half of the gel and then the nitrocellose papers were stained by a monocolonal Ab and HRP conjugate Ab. The other half were stained by silver stain for identification of MW. Results: After scoring the stained nitrocellose papen in each groups, no statistical significant difference was observed for C046 expression between the two groups. However, a significant Spearmen correlation was observed between CD46 expression and sperm motility (r=0.597, P=0.003. The MW of C046 was between 36 to 45 KD. with a mean of 42 KD. Discussion: This is the first report of a positive Spearmen correlation between sperm CD46 expression and sperm motility which suggest that there might be relation between CD46 expression and sperm motility.

Dietary lipids differentially affect membranes from different areas of rooster sperm.

Science.gov (United States)

Bongalhardo, D C; Leeson, S; Buhr, M M

2009-05-01

The present work aimed to compare the effect of dietary flax with other oil sources on rooster sperm membranes and on semen characteristics. White Leghorn roosters (16 per diet) were fed 1 of 4 treatments: control diet (CON), or a diet containing corn oil (CORN), fish oil (FISH), or flax seed (FLAX) as the lipid source. Semen from 4 birds (30 wk old) of each treatment was pooled, the sperm head (HM) and body membranes (BM) were isolated, and lipids were extracted and analyzed. Aspects of lipid composition tested were as follows: percentage of individual fatty acids (C14:0 to C24:1) in total fatty acids, percentage of fatty acid categories [saturated, monounsaturated, polyunsaturated (PUFA), n-3 and n-6 PUFA, and n-6:n-3 ratio] within total fatty acids, and percentage of phospholipids [phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylinositol, phosphatidylserine, and sphingomyelin] in total phospholipids. Sperm characteristics evaluated were as follows: volume, concentration, viability, percentage of motile cells, average path velocity, track speed, progressive velocity, lateral head displacement, straightness, and linearity. Diet did not affect membrane phospholipid ratios in either membrane but modified major fatty acids within certain phospholipids. Birds fed FISH and CORN showed, respectively, the highest and the lowest n-3 in sperm, causing reciprocal significant changes in n-6:n-3 ratio. Feeding FLAX caused intermediate effects in n-3, with values significantly lower than FISH but higher than CORN in HM (PC, PE, and phosphatidylinositol) and PC in BM (P < 0.05). In the PE phospholipids, FISH, followed by FLAX, increased n-3 in BM and decreased n-6 PUFA in HM. Sperm concentration was specifically correlated with the amount of 20:4n-6 in FLAX and 22:4n-6 in CON. In FLAX diets, straightness correlated with C18:0, n-3, and n-6:n-3 ratio. Diets containing distinct lipid sources differentially modify the lipid contents of HM and BM, with minor

Cationic synthetic peptides: assessment of their antimicrobial potency in liquid preserved boar semen.

Directory of Open Access Journals (Sweden)

Stephanie Speck

Full Text Available Various semen extender formulas are in use to maintain sperm longevity and quality whilst acting against bacterial contamination in liquid sperm preservation. Aminoglycosides are commonly supplemented to aid in the control of bacteria. As bacterial resistance is increasing worldwide, antimicrobial peptides (AMPs received lively interest as alternatives to overcome multi-drug resistant bacteria. We investigated, whether synthetic cationic AMPs might be a suitable alternative for conventional antibiotics in liquid boar sperm preservation. The antibacterial activity of two cyclic AMPs (c-WWW, c-WFW and a helical magainin II amide analog (MK5E was studied in vitro against two Gram-positive and eleven Gram-negative bacteria. Isolates included ATCC reference strains, multi-resistant E. coli and bacteria cultured from boar semen. Using broth microdilution, minimum inhibitory concentrations were determined for all AMPs. All AMPs revealed activity towards the majority of bacteria but not against Proteus spp. (all AMPs and Staphylococcus aureus ATCC 29213 (MK5E. We could also demonstrate that c-WWW and c-WFW were effective against bacterial growth in liquid preserved boar semen in situ, especially when combined with a small amount of gentamicin. Our results suggest that albeit not offering a complete alternative to traditional antibiotics, the use of AMPs offers a promising solution to decrease the use of conventional antibiotics and thereby limit the selection of multi-resistant strains.

Cationic Synthetic Peptides: Assessment of Their Antimicrobial Potency in Liquid Preserved Boar Semen

Science.gov (United States)

Speck, Stephanie; Courtiol, Alexandre; Junkes, Christof; Dathe, Margitta; Müller, Karin; Schulze, Martin

2014-01-01

Various semen extender formulas are in use to maintain sperm longevity and quality whilst acting against bacterial contamination in liquid sperm preservation. Aminoglycosides are commonly supplemented to aid in the control of bacteria. As bacterial resistance is increasing worldwide, antimicrobial peptides (AMPs) received lively interest as alternatives to overcome multi-drug resistant bacteria. We investigated, whether synthetic cationic AMPs might be a suitable alternative for conventional antibiotics in liquid boar sperm preservation. The antibacterial activity of two cyclic AMPs (c-WWW, c-WFW) and a helical magainin II amide analog (MK5E) was studied in vitro against two Gram-positive and eleven Gram-negative bacteria. Isolates included ATCC reference strains, multi-resistant E. coli and bacteria cultured from boar semen. Using broth microdilution, minimum inhibitory concentrations were determined for all AMPs. All AMPs revealed activity towards the majority of bacteria but not against Proteus spp. (all AMPs) and Staphylococcus aureus ATCC 29213 (MK5E). We could also demonstrate that c-WWW and c-WFW were effective against bacterial growth in liquid preserved boar semen in situ, especially when combined with a small amount of gentamicin. Our results suggest that albeit not offering a complete alternative to traditional antibiotics, the use of AMPs offers a promising solution to decrease the use of conventional antibiotics and thereby limit the selection of multi-resistant strains. PMID:25148109

TRIXcell+, a new long-term boar semen extender containing whey protein with higher preservation capacity and litter size

Directory of Open Access Journals (Sweden)

B.M. van den Berg

2014-02-01

Full Text Available It was the aim of the present study to test whey as protective protein for the sperm cell in the long-term boar semen preservation medium TRIXcell. Analyses of sperm cell motility using computer-assisted semen analysis (CASA indicated that the whey protein Porex has a similar protective effect as bovine serum albumin (BSA in maintaining viability of stored boar sperm. Boar sperm diluted in TRIXcell+ maintains commercially acceptable motility (>60% for 10 days, while swine sperm diluted in the semen preservation medium Beltsville Thawing Solution (BTS maintains commercially acceptable motility (>60% for 3-5 days for most boars. To test the on-farm fertility performance of TRIXcell+ compared to BTS, inseminations were started on 35 commercial pig production farms in the summer of 2006. During the period of July 2006 until July 2012 for each farm and each calendar year the mean farrowing rate and litter size for semen diluted in TRIXcell+ and stored for 3-5 days was found higher than that of semen stored for 1-2 days in BTS. Based on data gained from a total of 583.749 sows inseminated through the years 2006-2012, the mean farrowing rate for semen diluted in TRIXcell+ and BTS was 90.4 ± 4.0 and 87.9 ± 3.6, respectively, which is not significantly different. Based on the same data, the mean total number of piglets born alive for semen diluted in TRIXcell+ and BTS was 14.2 ± 0.7 and 13.6 ± 0.6, respectively, which is significantly different. We conclude that whey protein can effectively be used in the long-term preservation medium TRIXcell resulting in a higher litter size.

TRIXcell+, a new long-term boar semen extender containing whey protein with higher preservation capacity and litter size

Science.gov (United States)

van den Berg, B.M.; Reesink, J.; Reesink, W.

2014-01-01

It was the aim of the present study to test whey as protective protein for the sperm cell in the long-term boar semen preservation medium TRIXcell. Analyses of sperm cell motility using computer-assisted semen analysis (CASA) indicated that the whey protein Porex has a similar protective effect as bovine serum albumin (BSA) in maintaining viability of stored boar sperm. Boar sperm diluted in TRIXcell+ maintains commercially acceptable motility (>60%) for 10 days, while swine sperm diluted in the semen preservation medium Beltsville Thawing Solution (BTS) maintains commercially acceptable motility (>60%) for 3-5 days for most boars. To test the on-farm fertility performance of TRIXcell+ compared to BTS, inseminations were started on 35 commercial pig production farms in the summer of 2006. During the period of July 2006 until July 2012 for each farm and each calendar year the mean farrowing rate and litter size for semen diluted in TRIXcell+ and stored for 3-5 days was found higher than that of semen stored for 1-2 days in BTS. Based on data gained from a total of 583.749 sows inseminated through the years 2006-2012, the mean farrowing rate for semen diluted in TRIXcell+ and BTS was 90.4 ± 4.0 and 87.9 ± 3.6, respectively, which is not significantly different. Based on the same data, the mean total number of piglets born alive for semen diluted in TRIXcell+ and BTS was 14.2 ± 0.7 and 13.6 ± 0.6, respectively, which is significantly different. We conclude that whey protein can effectively be used in the long-term preservation medium TRIXcell resulting in a higher litter size. PMID:26623335

Effect of Selenium, Zinc, Vitamin C and E on Boar Ejaculate Quality at Heat Stress

Directory of Open Access Journals (Sweden)

Pavel Horký

2016-01-01

Full Text Available The aim of experiment was to test effect of selected antioxidants (selenium, zinc, vitamin C and E to reduce the impact of heat stress at boars. In the experiment, boars of Duroc breed were tested. The first control group (n = 10 was not supplemented with antioxidants. The second experimental group (n = 10 was supplemented with antioxidants in the following quantities of 0.5 mg of selenium (seleno-methionine, 100 mg of zinc (zinc-methionine, 70 mg of vitamin E (alpha‑tocopherol and 350 mg of vitamin C (ascorbic acid per kilogram of their feed. The experiment was carried out for 120 days and took place in summer (June to September. During the experiment, average and maximum daily temperatures, where boars were stabled, were monitored. Average daily temperature ranged from 12 to 28 °C. Maximum temperature during the day was from 13 to 32 °C. The evaluation of the semen quality has revealed increased number of abnormal spermatozoa in the control group of boars by 39 % (P < 0.05. There were observed no significant changes at other monitored parameters (ejaculate volume, total count of produced sperm, motility and sperm concentration. The results show that the addition of selenium, zinc, vitamin C and E may reduce the effect of heat stress to some extent at breeding boars.

Influence of chamber type integrated with computer-assisted semen analysis (CASA) system on the results of boar semen evaluation.

Science.gov (United States)

Gączarzewicz, D

2015-01-01

The objective of the study was to evaluate the effect of different types of chambers used in computer-assisted semen analysis (CASA) on boar sperm concentration and motility parameters. CASA measurements were performed on 45 ejaculates by comparing three commonly used chambers: Leja chamber (LJ), Makler chamber (MK) and microscopic slide-coverslip (SL). Concentration results obtained with CASA were verified by manual counting on a Bürker hemocytometer (BH). No significant differences were found between the concentrations determined with BH vs. LJ and SL, whereas higher (p0.05). The results obtained show that CASA assessment of boar semen should account for the effect of counting chamber on the results of sperm motility and concentration, which confirms the need for further study on standardizing the automatic analysis of boar semen.

Liquid Storage of Boar Semen Using Commercial Extenders

Directory of Open Access Journals (Sweden)

Liviu BOGDAN

2018-05-01

Full Text Available The research was focused on the modern evaluation of boar semen parameters, using computer assisted sperm analysis (CASA, before and after liquid storage at 15ºC. Semen was collected from 15 sexually mature boars by manual stimulation. Macroscopical and microscopical evaluation of semen was performed, followed by a detailed CASA analysis of all ejaculates. Subsequently, semen was diluted using 4 different extenders (Semtest, Androstar, MIII and Cronos and stored at 15ºC for 24 hours. Next, evaluation of progressive motility, total motility and viability was performed, using the same CASA system. All experiments were performed in triplicates and results were statistically analyzed. The average progressive motility after 24 hours was as follows: 69.56 ± 6.38 for MIII, 65.92% ± 2.63 for Semtest, 67.07% ± 5.58 for Androstar Plus and 68.93% ± 3.40 for Cronos. The viability results after 24 hours were: 86.34% ± 1.38 for Semtest extender, 93.55% ± 3.38% for Androstrar Plus, 89.19% ± 3.42 for MIII and 91.35% ± 2.37 for Cronos. The findings of this study suggest that the use of commercial extenders for short-term storage of swine semen is important in order to increase sperm longevity with minimal sperm function deterioration.

Genetic parameters for male fertility and its relationship to skatole and androstenone in Danish Landrace boars

DEFF Research Database (Denmark)

Strathe, Anders Bjerring; Velander, I.H.; Mark, Thomas

2013-01-01

Concerns have been raised regarding selection against the boar taint compounds, androstenone and skatole, due to potential unfavorable genetic correlations with important male fertility traits (i.e., selection of boars with low levels of these boar taint compounds might also reduce male fertility......). Hence, the objective of this investigation was to study the genetic association between direct measures of male fertility and the boar taint compounds in Danish Landrace pigs. Concentrations of skatole and androstenone in the back fat were available for approximately 6,000 and 1,000 Landrace boars......, and total number of sperm were available from 95,267 ejaculates. These ejaculates were collected between 2005 and 2012 and originated from 3,145 Landrace boars from 12 AI stations in Denmark. The traits were analyzed using single and multitrait animal models including univariate random regression models...

Effect of production management on semen quality during long-term storage in different European boar studs.

Science.gov (United States)

Schulze, M; Kuster, C; Schäfer, J; Jung, M; Grossfeld, R

2018-03-01

The processing of ejaculates is a fundamental step for the fertilizing capacity of boar spermatozoa. The aim of the present study was to identify factors that affect quality of boar semen doses. The production process during 1 day of semen processing in 26 European boar studs was monitored. In each boar stud, nine to 19 randomly selected ejaculates from 372 Pietrain boars were analyzed for sperm motility, acrosome and plasma membrane integrity, mitochondrial activity and thermo-resistance (TRT). Each ejaculate was monitored for production time and temperature for each step in semen processing using the special programmed software SEQU (version 1.7, Minitüb, Tiefenbach, Germany). The dilution of ejaculates with a short-term extender was completed in one step in 10 AI centers (n = 135 ejaculates), in two steps in 11 AI centers (n = 158 ejaculates) and in three steps in five AI centers (n = 79 ejaculates). Results indicated there was a greater semen quality with one-step isothermal dilution compared with the multi-step dilution of AI semen doses (total motility TRT d7: 71.1 ± 19.2%, 64.6 ± 20.0%, 47.1 ± 27.1%; one-step compared with two-step compared with the three-step dilution; P < .05). There was a marked advantage when using the one-step isothermal dilution regarding time management, preservation suitability, stability and stress resistance. One-step dilution caused significant lower holding times of raw ejaculates and reduced the possible risk of making mistakes due to a lower number of processing steps. These results lead to refined recommendations for boar semen processing. Copyright © 2018 Elsevier B.V. All rights reserved.

Ocean acidification impacts on sperm mitochondrial membrane potential bring sperm swimming behaviour near its tipping point.

Science.gov (United States)

Schlegel, Peter; Binet, Monique T; Havenhand, Jonathan N; Doyle, Christopher J; Williamson, Jane E

2015-04-01

Broadcast spawning marine invertebrates are susceptible to environmental stressors such as climate change, as their reproduction depends on the successful meeting and fertilization of gametes in the water column. Under near-future scenarios of ocean acidification, the swimming behaviour of marine invertebrate sperm is altered. We tested whether this was due to changes in sperm mitochondrial activity by investigating the effects of ocean acidification on sperm metabolism and swimming behaviour in the sea urchin Centrostephanus rodgersii. We used a fluorescent molecular probe (JC-1) and flow cytometry to visualize mitochondrial activity (measured as change in mitochondrial membrane potential, MMP). Sperm MMP was significantly reduced in ΔpH -0.3 (35% reduction) and ΔpH -0.5 (48% reduction) treatments, whereas sperm swimming behaviour was less sensitive with only slight changes (up to 11% decrease) observed overall. There was significant inter-individual variability in responses of sperm swimming behaviour and MMP to acidified seawater. We suggest it is likely that sperm exposed to these changes in pH are close to their tipping point in terms of physiological tolerance to acidity. Importantly, substantial inter-individual variation in responses of sperm swimming to ocean acidification may increase the scope for selection of resilient phenotypes, which, if heritable, could provide a basis for adaptation to future ocean acidification. © 2015. Published by The Company of Biologists Ltd.

Strategic use of anti-GnRH vaccine allowing selection of breeding boars without adverse effects on reproductive or production performances.

Science.gov (United States)

Oliviero, Claudio; Ollila, Anna; Andersson, Magnus; Heinonen, Mari; Voutila, Liisa; Serenius, Timo; Peltoniemi, Olli

2016-02-01

Boar stations raise only entire male pigs for selection as reproductive boars, but the majority of them will fail the selection process, ending at slaughter with a high risk of boar tainted meat. The aim of this study was to investigate whether a single dose of Improvac given to 16-week old boars had a negative effect on their subsequent sperm numbers and motility in 16 artificial insemination boars. We also aimed to generate more knowledge on incidence of boar taint at slaughter among Finnish pigs, compare production performances as average daily gain, feed conversion ratio, and carcass and meat quality (lean meat percentage, back fat, pH, color, androstenone, and skatole) of immunocastrated boars (n = 248) with those of entire boars (n = 268). Moreover, we aimed also to explore whether a fat biopsy taken at 16 weeks of age could already reveal the presence of boar taint compounds and be predictive of boar taint development at slaughter age. We found that 32% of entire boars (Figen Landrace, Figen Large White, and their crossbreed) slaughtered at an age of 25 weeks presented levels of androstenone and/or skatole above the threshold for boar taint in their meat. These boars (control) had higher androstenone and skatole levels in the back fat samples at slaughter (0.77 ± 0.55 and 0.09 ± 0.06 μg/g, respectively, mean ± standard deviation) than those in the immuno group (0.20 ± 0.25 and 0.06 ± 0.03 μg/g, respectively, P artificial insemination. We found no difference in the levels of testosterone, anti-GnRH antibodies titers, testicle morphology, and sperm numbers and motility between the boars vaccinated once, at 16 weeks of age, with anti-GnRH vaccine and the control boars (no vaccination). There were no differences in average daily gain, feed conversion ratio, lean meat percentage, and back fat between the immunocastrated boars and entire boars. Meat from immunocastrated boars had a higher pH and better color than meat from entire boars (P meat quality

Assessing in vivo fertilizing capacity of liquid-preserved boar semen according to the 'Hanover gilt model'.

Science.gov (United States)

Ardón, F; Döhring, A; Le Thi, X; Weitze, K F; Waberski, D

2003-04-01

The goal of this study was to determine the ability of the Hanover gilt model to assess in vivo fertilizing capacity of preserved sperm and to consider whether any modifications to this model were needed. This model evaluates the fertilizing capacity of semen based on the fertilization rate, the rate of normal embryos and the accessory sperm count of 3-5-day embryos. Its distinguishing characteristics are the use of one-time insemination of sperm in reduced numbers, of spontaneously ovulating gilts and of ovulation detection through ultrasound examination of ovaries. Reduced sperm numbers allow for an accurate evaluation of the fertilizing potential of different semen treatments, thereby avoiding the compensatory effect of doses calibrated to maximize fertility. The model's usefulness was assessed in a trial run designed to compare the fertilizing capacity of liquid boar semen diluted into two different extenders. The diluent, the boar and the backflow, had no significant effect on any of the parameters studied. Gilts inseminated less than 24 h before ovulation had a significantly higher (p semen), while using relatively few gilts and little time.

Boar genotype as a factor shaping age-related changes in semen parameters and reproduction longevity simulations.

Science.gov (United States)

Knecht, Damian; Jankowska-Mąkosa, Anna; Duziński, Kamil

2017-08-01

The aim of this study was a detailed analysis of the boar genotypes used in AI stations with an indication of their production capacity, including age and a precise analysis of their culling time and reason. The study included 334 boars: 81 Polish Large White (PLW), 108 Polish Landrace (PL), 49 Pietrain (P), 56 Duroc × Pietrain (D × P) and 40 Hampshire × Pietrain (H × P). Semen volume, spermatozoa concentration, total number of spermatozoa, number of motile spermatozoa, and number of insemination doses were analyzed. Quadratic regression was used to illustrate the selected sperm parameters at specific ages. Among all the studied boars the lowest motilities of spermatozoa were identified in white breeds PLW and PL, and the difference between motility extremes was 3.53% (P ≤ 0.01). The highest number of insemination doses were produced from D × P crossbreed boars: about 0.7 portions more compared to PL, 1.13 to PLW, 1.18 to H × P and 1.8 to P (all differences P ≤ 0.01). It has been shown in the case of ejaculate volume that for PLW and H × P boars the culling moment was far too early in terms of production capacity and differences were, respectively, 16.35 ml for PLW and 12.61 ml for H × P. Based on the developed regression equations, the earliest maximum number of motile sperm (73.82 × 10 9 ) was obtained by H × P crossbreed boars as early as at age 24 months. The highest values for this parameter were achieved, however, by other D × P crossbreed boars: 74.30 × 10 9  at the later age of 32 months. A consequence of the high number of motile sperm in young H × P boars was that the theoretical maximum value of the number of AI doses was produced as early as the 14th month (25.59 portions). Curves of similar shape were obtained for PL and D × P boars; the difference in maximal values was 0.54 portions in favor of crossbreeds, at a later age of 7 months. It was noted that for PLW and D × P boars the highest number

A technical assessment of the porcine ejaculated spermatozoa for a sperm-specific RNA-seq analysis.

Science.gov (United States)

Gòdia, Marta; Mayer, Fabiana Quoos; Nafissi, Julieta; Castelló, Anna; Rodríguez-Gil, Joan Enric; Sánchez, Armand; Clop, Alex

2018-04-26

The study of the boar sperm transcriptome by RNA-seq can provide relevant information on sperm quality and fertility and might contribute to animal breeding strategies. However, the analysis of the spermatozoa RNA is challenging as these cells harbor very low amounts of highly fragmented RNA, and the ejaculates also contain other cell types with larger amounts of non-fragmented RNA. Here, we describe a strategy for a successful boar sperm purification, RNA extraction and RNA-seq library preparation. Using these approaches our objectives were: (i) to evaluate the sperm recovery rate (SRR) after boar spermatozoa purification by density centrifugation using the non-porcine-specific commercial reagent BoviPure TM ; (ii) to assess the correlation between SRR and sperm quality characteristics; (iii) to evaluate the relationship between sperm cell RNA load and sperm quality traits and (iv) to compare different library preparation kits for both total RNA-seq (SMARTer Universal Low Input RNA and TruSeq RNA Library Prep kit) and small RNA-seq (NEBNext Small RNA and TailorMix miRNA Sample Prep v2) for high-throughput sequencing. Our results show that pig SRR (~22%) is lower than in other mammalian species and that it is not significantly dependent of the sperm quality parameters analyzed in our study. Moreover, no relationship between the RNA yield per sperm cell and sperm phenotypes was found. We compared a RNA-seq library preparation kit optimized for low amounts of fragmented RNA with a standard kit designed for high amount and quality of input RNA and found that for sperm, a protocol designed to work on low-quality RNA is essential. We also compared two small RNA-seq kits and did not find substantial differences in their performance. We propose the methodological workflow described for the RNA-seq screening of the boar spermatozoa transcriptome. FPKM: fragments per kilobase of transcript per million mapped reads; KRT1: keratin 1; miRNA: micro-RNA; miscRNA: miscellaneous

High resolution DNA flow cytometry of boar sperm cells in identification of boars carrying cytogenetic aberrations

DEFF Research Database (Denmark)

Larsen, Jacob; Christensen, Knud; Larsen, Jørgen K

2004-01-01

The cytogenetic quality of boars used for breeding determines the litter outcome and thus has large economical consequences. Traditionally, quality controls based on the examination of simple karyograms are time consuming and sometimes give uncertain results. As an alternative, the use of high...

Single-cell-based evaluation of sperm progressive motility via fluorescent assessment of mitochondria membrane potential.

Science.gov (United States)

Moscatelli, Natalina; Spagnolo, Barbara; Pisanello, Marco; Lemma, Enrico Domenico; De Vittorio, Massimo; Zara, Vincenzo; Pisanello, Ferruccio; Ferramosca, Alessandra

2017-12-20

Sperm cells progressive motility is the most important parameter involved in the fertilization process. Sperm middle piece contains mitochondria, which play a critical role in energy production and whose proper operation ensures the reproductive success. Notably, sperm progressive motility is strictly related to mitochondrial membrane potential (MMP) and consequently to mitochondrial functionality. Although previous studies presented an evaluation of mitochondrial function through MMP assessment in entire sperm cells samples, a quantitative approach at single-cell level could provide more insights in the analysis of semen quality. Here we combine laser scanning confocal microscopy and functional fluorescent staining of mitochondrial membrane to assess MMP distribution among isolated spermatozoa. We found that the sperm fluorescence value increases as a function of growing progressive motility and that such fluorescence is influenced by MMP disruptors, potentially allowing for the discrimination of different quality classes of sperm cells in heterogeneous populations.

Supramolecular organization of the sperm plasma membrane during maturation and capacitation.

Science.gov (United States)

Jones, Roy; James, Peter S; Howes, Liz; Bruckbauer, Andreas; Klenerman, David

2007-07-01

In the present study, a variety of high resolution microscopy techniques were used to visualize the organization and motion of lipids and proteins in the sperm's plasma membrane. We have addressed questions such as the presence of diffusion barriers, confinement of molecules to specific surface domains, polarized diffusion and the role of cholesterol in regulating lipid rafts and signal transduction during capacitation. Atomic force microscopy identified a novel region (EqSS) within the equatorial segment of bovine, porcine and ovine spermatozoa that was enriched in constitutively phosphorylated proteins. The EqSS was assembled during epididymal maturation. Fluorescence imaging techniques were then used to follow molecular diffusion on the sperm head. Single lipid molecules were freely exchangeable throughout the plasma membrane and showed no evidence for confinement within domains. Large lipid aggregates, however, did not cross over the boundary between the post-acrosome and equatorial segment suggesting the presence of a molecular filter between these two domains. A small reduction in membrane cholesterol enlarges or increases lipid rafts concomitant with phosphorylation of intracellular proteins. Excessive removal of cholesterol, however, disorganizes rafts with a cessation of phosphorylation. These techniques are forcing a revision of long-held views on how lipids and proteins in sperm membranes are assembled into larger complexes that mediate recognition and fusion with the egg.

A genome-wide association study reveals a novel candidate gene for sperm motility in pigs

NARCIS (Netherlands)

Diniz, D.B.; Lopes, M.S.; Broekhuijse, M.L.W.J.; Lopes, P.S.; Harlizius, B.; Guimaraes, S.E.F.; Duijvesteijn, N.; Knol, E.F.; Silva, F.F.

2014-01-01

Sperm motility is one of the most widely used parameters in order to evaluate boar semen quality. However, this trait can only be measured after puberty. Thus, the use of genomic information appears as an appealing alternative to evaluate and improve selection for boar fertility traits earlier in

Effect of a pre-freezing treatment with cholesterol-loaded cyclodextrins on boar sperm longevity, capacitation dynamics, ability to adhere to porcine oviductal epithelial cells in vitro and DNA fragmentation dynamics.

Science.gov (United States)

Tomás, C; Blanch, E; Fazeli, A; Mocé, E

2013-01-01

The aim of this work was to examine how a pre-freezing treatment with cholesterol-loaded cyclodextrins (CLC) affects boar sperm longevity, capacitation dynamics, ability to bind to a porcine telomerase-immortalised oviductal epithelial cell line (TERT-OPEC) in vitro and DNA integrity dynamics after freeze-thawing. Although the samples treated with CLC exhibited lower sperm quality than the control samples (P0.05) after long-term incubation (26h at 37 or 16°C). Additionally, the CLC-treated spermatozoa underwent similar capacitation and DNA fragmentation dynamics as the control spermatozoa (P>0.05). However, CLC-treated spermatozoa were better able to bind to TERT-OPEC in vitro (POPEC in vitro, which could have an effect on the establishment of the sperm reservoir in the ampullary--isthmic junction in vivo. Additionally, frozen-thawed spermatozoa can be stored at 16°C for at least 6h without a significant observable decline in sperm quality, which could be beneficial for the transport of thawed diluted doses of spermatozoa from the laboratory to the farm.

High resolution DNA flow cytometry of boar sperm cells in identification of boars carrying cytogenetic aberrations

DEFF Research Database (Denmark)

Larsen, Jacob; Christensen, Knud; Larsen, Jørgen K

2004-01-01

The cytogenetic quality of boars used for breeding determines the litter outcome and thus has large economical consequences. Traditionally, quality controls based on the examination of simple karyograms are time consuming and sometimes give uncertain results. As an alternative, the use of high...... necessitate essential improvements in standardization and measurement precision....

Effects of storage in different semen extenders on the pre-freezing and post-thawing quality of boar spermatozoa.

Science.gov (United States)

Dziekońska, A; Zasiadczyk, Ł; Lecewicz, M; Strzeżek, R; Koziorowska-Gilun, M; Fraser, L; Mogielnicka-Brzozowska, M; Kordan, W

2015-01-01

The aim of this study was to investigate the effects of storage of semen in different commercial extenders on the pre-freezing and post-thawing quality of boar spermatozoa. Semen was diluted in BTS, Androhep (AH) and Gedil (GD), stored for 24 h at 17°C, and then frozen in accordance with the cryopreservation protocol. Analyses of the quality of spermatozoa included: motility, normal apical ridge (NAR) acrosome, plasma membrane integrity (PMI), mitochondrial membrane potential (MMP), measurements of ATP content and activity of superoxidase dismutase (SOD) and glutathione peroxidase (GPx). Prior to the freezing process, no significant effect of the extender on the sperm quality parameters was noted. After thawing the spermatozoa it was demonstrated that the type of extender used influenced PMI, MMP, ATP content and activity of GPx. In the AH extender the percentage of spermatozoa with PMI and ATP content in spermatozoa was significantly higher (Pextenders. In addition, semen stored in the AH was characterised by a statistically higher (Pboar spermatozoa stored for 24 hours in liquid state can be used. However, the type of extender used prior to freezing may have a significant effect on the post-thawing quality of the spermatozoa. The AH extender better secured the quality of thawed boar spermatozoa as compared with the BTS or GD.

Flow cytometric sex sorting affects CD4 membrane distribution and binding of exogenous DNA on bovine sperm cells.

Science.gov (United States)

Domingues, William Borges; da Silveira, Tony Leandro Rezende; Komninou, Eliza Rossi; Monte, Leonardo Garcia; Remião, Mariana Härter; Dellagostin, Odir Antônio; Corcini, Carine Dahl; Varela Junior, Antônio Sergio; Seixas, Fabiana Kömmling; Collares, Tiago; Campos, Vinicius Farias

2017-08-01

Bovine sex-sorted sperm have been commercialized and successfully used for the production of transgenic embryos of the desired sex through the sperm-mediated gene transfer (SMGT) technique. However, sex-sorted sperm show a reduced ability to internalize exogenous DNA. The interaction between sperm cells and the exogenous DNA has been reported in other species to be a CD4-like molecule-dependent process. The flow cytometry-based sex-sorting process subjects the spermatozoa to different stresses causing changes in the cell membrane. The aim of this study was to elucidate the relationship between the redistribution of CD4-like molecules and binding of exogenous DNA to sex-sorted bovine sperm. In the first set of experiments, the membrane phospholipid disorder and the redistribution of the CD4 were evaluated. The second set of experiments was conducted to investigate the effect of CD4 redistribution on the mechanism of binding of exogenous DNA to sperm cells and the efficiency of lipofection in sex-sorted bovine sperm. Sex-sorting procedure increased the membrane phospholipid disorder and induced the redistribution of CD4-like molecules. Both X-sorted and Y-sorted sperm had decreased DNA bound to membrane in comparison with the unsorted sperm; however, the binding of the exogenous DNA was significantly increased with the addition of liposomes. Moreover, we demonstrated that the number of sperm-bound exogenous DNA was decreased when these cells were preincubated with anti-bovine CD4 monoclonal antibody, supporting our hypothesis that CD4-like molecules indeed play a crucial role in the process of exogenous DNA/bovine sperm cells interaction.

Modification of trout sperm membranes associated with activation and cryopreservation. Implications for fertilizing potential

Science.gov (United States)

Abstract We investigated the effects of two trout sperm activation solutions on sperm physiology and membrane organization prior to and following cryopreservation using flow cytometry and investigated their impact on in vitro fertility. Cryopreservation caused greater phospholipid disorder (high pl...

Ion channel activity of membrane vesicles released from sea urchin sperm during the acrosome reaction

International Nuclear Information System (INIS)

Schulz, Joseph R.; Vega-Beltran, Jose L. de la; Beltran, Carmen; Vacquier, Victor D.; Darszon, Alberto

2004-01-01

The sperm acrosome reaction (AR) involves ion channel activation. In sea urchin sperm, the AR requires Ca 2+ and Na + influx and K + and H + efflux. During the AR, the plasma membrane fuses with the acrosomal vesicle membrane forming hybrid membrane vesicles that are released from sperm into the medium. This paper reports the isolation and preliminary characterization of these acrosome reaction vesicles (ARVs), using synaptosome-associated protein of 25 kDa (SNAP-25) as a marker. Isolated ARVs have a unique protein composition. The exocytosis regulatory proteins vesicle-associated membrane protein and SNAP-25 are inside ARVs, as judged by protease protection experiments, and membrane associated based on Triton X-114 partitioning. ARVs fused with planar bilayers display three main types of single channel activity. The most frequently recorded channel is cationic, weakly voltage dependent and has a low open probability that increases with negative potentials. This channel is activated by cAMP, blocked by Ba 2+ , and has a PK + /PNa + selectivity of 4.5. ARVs represent a novel membrane preparation suitable to deepen our understanding of ion channel activity in the AR and during fertilization

Preliminary characterization of multiple hyaluronidase forms in boar reproductive tract

Czech Academy of Sciences Publication Activity Database

Cibulková, Eva; Maňásková, Pavla; Jonáková, Věra; Tichá, M.

2007-01-01

Roč. 68, č. 7 (2007), s. 1047-1054 ISSN 0093-691X R&D Projects: GA ČR GA303/06/0895; GA MŠk 1M06011 Institutional research plan: CEZ:AV0Z50520514; CEZ:AV0Z50520701 Keywords : hyaluronidase * boar sperm * seminal plasma * epididymis * seminal vesicle Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.911, year: 2007

Anti-oxidant supplementation improves boar sperm characteristics and fertility after cryopreservation: comparison between cysteine and rosemary (Rosmarinus officinalis).

Science.gov (United States)

Malo, C; Gil, L; Gonzalez, N; Martínez, F; Cano, R; de Blas, I; Espinosa, E

2010-08-01

Anti-oxidants partially ameliorated the detrimental effects of reactive oxidative substances produced during cryopreservation. The objective of the study was to determine the effect of anti-oxidant addition to the freezing extender on boar semen qualities and fertility capacity. Ejaculates were collected from a previously selected boar and semen samples were processed using the straw freezing procedure. In experiment 1, semen samples were cryopreserved in lactose-egg yolk solution supplemented with various concentrations of cysteine (0, 5 and 10mM) to determinate a cysteine concentration capable of producing a protective effect during cryopreservation. Semen quality (total motility, progressive motility, viability, acrosome integrity and hypoosmotic swelling test) was evaluated after freezing and thawing and then every hour for 3h. In experiment 2, ejaculates were cryopreserved with lactose-egg yolk extender with or without the following anti-oxidants: cysteine, rosemary (Rosmarinus officinalis) and cysteine plus rosemary. Semen quality was evaluated. In the experiment 3, fertility capacity of semen frozen in anti-oxidant supplementation extenders was examined in vitro. A total of 2232 oocytes were in vitro matured and inseminated with frozen-thawed sperm. In summary: (i) the effective concentration of cysteine in freezing extender was 10mM; (ii) the addition of exogenous rosemary or cysteine to the freezing extender positively affected post-thawed viability and acrosome integrity. Only rosemary supplementation improved total motility at 3h and progressive motility at any time; (iii) the inclusion of rosemary into the extender was effective in penetration and cleavage rate and also in the efficiency of the fertilization system. (c) 2010 Elsevier Inc. All rights reserved.

Evaluation of glucose as a cryoprotectant for boar semen.

Science.gov (United States)

De los Reyes, M; Saenz, L; Lapierre, L; Crosby, J; Barros, C

2002-10-19

Fertility parameters of boar spermatozoa were evaluated in vitro, after freeze-thawing the semen in three different extenders containing permeable and non-permeable cryoprotectants: A (111.0 mM Tris, 31.4 mM citric acid, 185.0 mM glucose, 20 per cent egg yolk, 3 per cent glycerol and 100 iu/ml penicillin G); B (200 mM Tris; 70.8 mM citric acid, 55.5 mM glucose, 20 per cent egg yolk, three per cent glycerol and 100 iu/ml penicillin G); C (200 mM Tris, 70.8 mM citric acid, 55.5 mM fructose, 20 per cent egg yolk, 3 per cent glycerol and 100 iu/ml penicillin G). The freeze-thawing techniques were the same for each extender. Eight ejaculates from four boars were obtained; the sperm-rich fraction of each ejaculate was extended in each of the three media at a final concentration of 400 x 106 sperm/ml, loaded into 0.5 ml straws and frozen at a rate of 30 degrees C/minute to -196 degrees C. The straws were thawed at 60 degrees C for eight seconds. Sperm motility, acrosomal integrity and in vitro sperm penetration through the zona pellucida of gilt oocytes matured in vitro were evaluated. The motility of unfrozen spermatozoa was 93.1 per cent compared with 60.7 per cent, 48.2 per cent and 35 per cent for sperm frozen in extenders A, B and C respectively; these values were all significantly different (Pextender A, but there were significant decreases in sperm motility after 30 minutes of incubation in B and C. The percentage acrosomal integrities were 97.2 per cent for the control and 45.5 per cent, 30.3 per cent and 16.8 per cent for the frozen-thawed spermatozoa in extenders A, B and C respectively. The results of the in vitro penetration assay were 80.7 per cent when using control spermatozoa, and 42.2 per cent, 18.4 per cent and 3.3 per cent when using frozen-thawed spermatozoa in extenders A, B and C respectively

Mitochondrial outer membrane permeabilization increases reactive oxygen species production and decreases mean sperm velocity but is not associated with DNA fragmentation in human sperm.

Science.gov (United States)

Treulen, F; Uribe, P; Boguen, R; Villegas, J V

2016-02-01

Does induction of mitochondrial outer membrane permeabilization (MOMP) in vitro affect specific functional parameters of human spermatozoa? Our findings show that MOMP induction increases intracellular reactive oxygen species (ROS) and decreases mean sperm velocity but does not alter DNA integrity. MOMP in somatic cells is related to a variety of apoptotic traits, such as alteration of mitochondrial membrane potential (ΔΨm), and increase in ROS production and DNA fragmentation. Although the presence of these apoptotic features has been reported in spermatozoa, to date the effects of MOMP on sperm function and DNA integrity have not been analysed. The study included spermatozoa from fertile donors. Motile sperm were obtained using the swim-up method. The highly motile sperm were collected and diluted with human tubal fluid to a final cell concentration of 5 × 10(6) ml(-1). To induce MOMP, selected sperm were treated at 37°C for 4 h with a mimetic of a Bcl-2 pro-apoptotic protein, ABT-737. MOMP was evaluated by relocating of cytochrome c. In addition, the effect of ABT-737 on mitochondrial inner membrane permeabilization was assessed using the calcein-AM/cobalt chloride method. In turn, ΔΨm was evaluated with JC-1 staining, intracellular ROS production with dihydroethidium, sperm motility was analysed by computer-assisted sperm analysis and DNA fragmentation by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) assay. Measurements were performed by flow cytometry. MOMP was associated with ΔΨm dissipation (P < 0.05), increased ROS production (P < 0.05) and decreased mean sperm velocity (P < 0.05), but it was not associated with DNA fragmentation. MOMP did not induce a large increase in ROS, which could explain the negligible effect of MOMP on sperm DNA fragmentation under our experimental conditions. The study was carried out in vitro using highly motile sperm, selected by swim-up, from healthy donors. The results obtained in this

Quality and fertilizing capacity of boar spermatozoa during liquid storage in extender supplemented with different antibiotics.

Science.gov (United States)

Bryła, Magdalena; Trzcińska, Monika

2015-12-01

The aim of the study was to determine the effect of antibiotics on quality parameters and fertilizing capacity of boar sperm during liquid preservation. In the first experiment, semen was diluted in an extender containing 200 μg/mL of gentamicin as a control and diluted in a modified extenders: Ext I (contained 200 μg/mL florfenicol), Ext II (contained 200 μg/mL polymyxin B), Ext III (contained 100 μg/mL gentamicin and 100 μg/mL florfenicol) and Ext IV (contained 100 μg/mL gentamicin and 100 μg/mL polymyxin B). The semen was stored for ten days. Sperm quality was evaluated based on the motility (CASA; TM: total motility; PM: progressive motility), membrane integrity (YO-PRO-1/PI assay), mitochondrial activity (JC-1) and DNA integrity (TUNEL). The highest PM% (62.5 ± 9.6) was observed in Ext III at Day 6 of storage. The highest sperm viability and mitochondrial transmembrane potential was noticed at the end of the storage period in Ext III. Long-term storage did not induce DNA fragmentation in the extenders analyzed. In the second experiment, semen diluted in the control extender and in the extender providing the highest quality spermatozoa on Day 10 (Ext III) was used for artificial insemination (AI) of synchronized gilts. Our studies showed that the highest reproductive performance of inseminated gilts (pregnant gilts: 97.0%, litter size: 11.4 ± 1.2) occurred with Ext III semen dilution. The combination of 100 μg/mL gentamicin and 100 μg/mL florfenicol in the extender maintained sperm motility, membrane integrity and mitochondrial activity and enhanced the higher reproduction success. Copyright © 2015 Elsevier B.V. All rights reserved.

The combinatorial effect of different Equex STM paste concentrations, cryoprotectants and the straw-freezing methods on the post-thaw boar semen quality.

Science.gov (United States)

Wu, T-W; Cheng, F-P; Chen, I-H; Yang, C-H; Tsai, M-Y; Chang, M-H; Wang, J-H; Wu, J-T

2013-02-01

This study was to evaluate the combinatorial effect (14 treatments, A-N) of different Equex STM paste concentrations, cryoprotectants and the straw-freezing method on the post-thaw boar semen quality. Two ejaculates were collected from each of nine boars (three boars from each of three breeds). Semen was diluted in extenders with different concentrations of Equex STM paste and different cryoprotectants [glycerol or dimethylacetamide (DMA)] before cryopreserving via liquid nitrogen or dry ice. Motility, viability, percentage of spermatozoa with intense acrosomal staining and with normal morphology of post-thaw sperm were evaluated. The qualities of thawed semen were best preserved in treatment H (extender with 0.5% Equex STM paste and 5% glycerol and freezing by dry ice) and were worst in treatment B (extender with 0% Equex STM paste and 5% DMA and freezing by dry ice). Significant difference (p 0.05). Moreover, statistical analysis suggests that no significant difference was present in semen quality among breed or individual donors (p > 0.05). These findings suggest that Equex STM paste improved the cryosurvival efficiency of boar sperm, and the favourable straw-freezing method changes between glycerol and DMA. © 2012 Blackwell Verlag GmbH.

Cryo-scanning electron microscopy discloses differences in dehydration of frozen boar semen stored in large containers.

Science.gov (United States)

Ekwall, H

2009-02-01

In general, freezing in flat plastic polyethylene terephthalate (PET) bags (FlatPacks) at 50 degrees C/min gives better post-thaw viability, in terms of sperm motility and membrane integrity, than does freezing in plastic maxi-straws, probably owing to differences in cryobiology. To test the hypothesis that this better survival post-thaw relates to the degree of sperm dehydration during freezing, the present study investigated the structure of boar semen in a frozen state using cryo-scanning electron microscopy (cryo-SEM) to compare two different packages (FlatPacks and maxi-straws) for single artificial insemination (AI) doses, and three different freezing rates. The semen was split-sample frozen in maxi-straws or FlatPacks (both holding 5 ml) using 3% glycerol as cryoprotectant. Three freezing rates were applied from -5 degrees C to -100 degrees C, namely 2 degrees C/min, 50 degrees C/min and 1200 degrees C/min, the lattermost by plunging the samples into liquid nitrogen (LN(2)). The samples were thereafter fractured into LN(2) and larger areas of extra-cellular, unbound frozen water ('ice lakes') were measured to determine the degree of dehydration of the spermatozoa. These areas decreased in size with an increase in cooling rate, the differences in size being more dramatic for maxi-straws than for FlatPacks. Size of ice lakes was also influenced by location within package in relation to cooling rate, the central values being always smaller in maxi-straws than in Flatpacks (p < 0.05 at 2 degrees C/min and 50 degrees C/min) but not at 1200 degrees C/min, which suggested the FlatPack allows for more homogenous freezing of boar semen.

Effect of semen extenders on frozen-thawed boar sperm characteristics and distribution in the female genital tract after deep intrauterine insemination in sows.

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Noguchi, Michiko; Yoshioka, Koji; Hikono, Hirokazu; Suzuki, Chie; Kikuchi, Kazuhiro

2015-12-01

We compared the effects of extenders of frozen-thawed semen on post-thaw sperm characteristics and the distribution of frozen-thawed spermatozoa in the female genital tract after fixed-timed deep intrauterine insemination (DIUI) in sows. Frozen semen samples were thawed and diluted in either modified Modena solution (mMS) or porcine fertilization medium (PFM) containing theophylline, adenosine and cysteine. Sperm quality, assessed in vitro based on motility using a computer-assisted sperm analyzer and the integrity of the plasma and acrosomal membranes using flow cytometry, was evaluated at 0.5, 1.5, 3 and 6h after thawing. Progressive motility and the percentage of spermatozoa with damaged acrosomal membranes in PFM were significantly better than in mMS throughout the 6h. Sows with estrus synchronized using prostaglandin F2 alpha, equine chorionic gonadotropin and human chorionic gonadotropin (hCG) were inseminated once with mMS- or PFM-diluted 5 × 10(8) frozen-thawed spermatozoa by DIUI at 34 h after the hCG injection. At 4h after DIUI, reproductive tracts were recovered from 30 sows. There were significantly fewer polymorphonuclear leukocytes (PMNs) and more spermatozoa outside PMNs in the uterine horn after PFM treatment than with mMS. When 22 sows were administered DIUI with 10 × 10(8) frozen-thawed spermatozoa at 36 h after hCG, the pregnancy rates did not differ significantly between the mMS- (36%) and PFM- (64%) treated groups. Thus, PFM enhanced progressive sperm motility but increased sperm membrane damage compared with mMS; it also suppressed the migration of PMNs into the uterine lumen. Copyright © 2015 Elsevier B.V. All rights reserved.

Effects of hepatitis B virus S protein exposure on sperm membrane integrity and functions.

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XiangJin Kang

Full Text Available BACKGROUND: Hepatitis B is a public health problem worldwide. Viral infection can affect a man's fertility, but only scant information about the influence of hepatitis B virus (HBV infection on sperm quality is available. The purpose of this study was to investigate the effect of hepatitis B virus S protein (HBs on human sperm membrane integrity and functions. METHODS/PRINCIPAL FINDINGS: Reactive oxygen species (ROS, lipid peroxidation (LP, total antioxidant capacity (TAC and phosphatidylserine (PS externalization were determined. The terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL assays and flow cytometric analyses were performed. (1 After 3 h incubation with 25 µg/ml of HBs, the average rates of ROS positive cells, annexin V-positive/propidium iodide (PI-negative cells, Caspases-3,-8,-9 positive cells and TUNEL-positive cells were significantly increased in the test groups as compared to those in the control groups, while TAC level was decreased when compared with the control. The level of malondialdehyde (MDA in the sperm cells exposed to 50 µg/ml of HBs for 3 h was significantly higher than that in the control (P<0.05-0.01. (2 HBs increased the MDA levels and the numbers of ROS positive cells, annexin V-positive/PI-negative cells, caspases-3, -8, -9 positive cells and TUNEL-positive cells in a dose-dependent manner. (3 HBs monoclonal antibody (MAb and N-Acetylcysteine (NAC reduced the number of ROS-positive sperm cells. (4 HBs decreased the TAC levels in sperm cells in a dose-dependent manner. CONCLUSION: HBs exposure could lead to ROS generation, lipid peroxidation, TAC reduction, PS externalization, activation of caspases, and DNA fragmentation, resulting in increased apoptosis of sperm cells and loss of sperm membrane integrity and causing sperm dysfunctions.

The use of comet assay to assess DNA integrity of boar spermatozoa following liquid preservation at 5 degrees C and 16 degrees C.

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J Strzezek

2004-03-01

Full Text Available The comet assay, under neutral conditions, allows the assessment of DNA integrity influenced by sperm ageing, which is manifested in DNA double-strand breaks. Here, we attempted to use a modified neutral comet assay test (single-cell gel electrophoresis, to our knowledge for the first time, to assess DNA integrity of boar spermatozoa during liquid storage for 96 h at 5 degrees C and 16 degrees C. In this comet assay protocol we used 2% beta-mercaptoethanol prior to the lysis procedure, to aid in removing nuclear proteins. Ejaculates from 3 boars (designated A, C and G were diluted with a standard semen extender, Kortowo-3 (K-3, which was supplemented with lipoprotein fractions extracted from hen egg yolk (LPFh or ostrich egg yolk (LPFo. Irrespective of the extender type, the percentage of comet-detected spermatozoa with damaged DNA increased gradually during prolonged storage at 5 degrees C and 16 degrees C. Spermatozoa stored in K-3 extender exhibited elevated levels of DNA damage at both storage temperatures. Significant differences in DNA damage among the boars were more pronounced during storage in LPF-based extenders at 5 degrees C: spermatozoa of boars A and G were less susceptible to DNA damage. The percent of tail DNA in comets was lower in LPF-based extenders, and there were individual variations among the boars. We observed that changes in DNA integrity were dependent on the extender type and storage temperature. A higher level of DNA instability was observed in K-3 extended semen compared with K-3/LPFh or K-3/LPFo extended semen during storage at 5 degrees C. No significant difference in the level of DNA damage between K-3/LPFh and K-3/LPFo was observed. It seems that a long-term storage can affect genomic integrity of boar spermatozoa. The modified neutral comet assay can be used to detect low levels of DNA damage in boar spermatozoa during liquid preservation. Therefore, screening for sperm DNA damage may be used as an additional

The use of comet assay to assess DNA integrity of boar spermatozoa following liquid preservation at 5 degrees C and 16 degrees C.

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Fraser, L; Strzezek, J

2004-01-01

The comet assay, under neutral conditions, allows the assessment of DNA integrity influenced by sperm ageing, which is manifested in DNA double-strand breaks. Here, we attempted to use a modified neutral comet assay test (single-cell gel electrophoresis), to our knowledge for the first time, to assess DNA integrity of boar spermatozoa during liquid storage for 96 h at 5 degrees C and 16 degrees C. In this comet assay protocol we used 2% beta-mercaptoethanol prior to the lysis procedure, to aid in removing nuclear proteins. Ejaculates from 3 boars (designated A, C and G) were diluted with a standard semen extender, Kortowo-3 (K-3), which was supplemented with lipoprotein fractions extracted from hen egg yolk (LPFh) or ostrich egg yolk (LPFo). Irrespective of the extender type, the percentage of comet-detected spermatozoa with damaged DNA increased gradually during prolonged storage at 5 degrees C and 16 degrees C. Spermatozoa stored in K-3 extender exhibited elevated levels of DNA damage at both storage temperatures. Significant differences in DNA damage among the boars were more pronounced during storage in LPF-based extenders at 5 degrees C: spermatozoa of boars A and G were less susceptible to DNA damage. The percent of tail DNA in comets was lower in LPF-based extenders, and there were individual variations among the boars. We observed that changes in DNA integrity were dependent on the extender type and storage temperature. A higher level of DNA instability was observed in K-3 extended semen compared with K-3/LPFh or K-3/LPFo extended semen during storage at 5 degrees C. No significant difference in the level of DNA damage between K-3/LPFh and K-3/LPFo was observed. It seems that a long-term storage can affect genomic integrity of boar spermatozoa. The modified neutral comet assay can be used to detect low levels of DNA damage in boar spermatozoa during liquid preservation. Therefore, screening for sperm DNA damage may be used as an additional test of sperm

Boar spermatozoa cryopreservation in low glycerol/trehalose enriched freezing media improves cellular integrity.

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Gutiérrez-Pérez, Oscar; Juárez-Mosqueda, María de Lourdes; Carvajal, Salvador Uribe; Ortega, María Elena Trujillo

2009-06-01

The use of glycerol for boar semen cryopreservation results in low fertility, possibly due to toxicity. This has led to recommend the use of solutions with less than 4% glycerol. Trehalose is a disaccharide known to stabilize proteins and biologic membranes during processes such as cryopreservation. Thus, it was decided to evaluate the cryoprotective effect of glycerol/trehalose mixtures. Effects on motility (M), viability (Vb) and acrosomal integrity (nA) were evaluated. Sperm samples were frozen in three different extenders: G4 contained 4% glycerol; T1 contained 1% glycerol plus 250 mM trehalose and T0.5 was constituted by 0.5% glycerol plus 250 mM trehalose. All extenders yielded similar post-freezing/thawing motility rates. Viability was diminished in T0.5 as compared to the others. In regard to acrosome integrity, it was twice as high (Pextender. Thus, T1 twice as many spermatozoa were alive, motile and intact, than in either T0.5 or G4, i.e. during freeze/thawing the use of T1 resulted in twice as many fertile cells as when using the other extenders. During our study, we noted that there were wide individual variations both in sperm viability and in motility.

Effect of cooling to different sub-zero temperatures on boar sperm cryosurvival

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Angelica Garcia-Olivares

2016-03-01

Conclusions: Cooling of pig sperm to −7 °C (no freezing damaged sperm function and structure; in contrast, cooling to either −3 °C or −5 °C did not change pig sperm survival after freeze-thawing.

Characterization and possible function of glyceraldehyde-3-phosphate dehydrogenase-spermatogenic protein GAPDHS in mammalian sperm.

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Margaryan, Hasmik; Dorosh, Andriy; Capkova, Jana; Manaskova-Postlerova, Pavla; Philimonenko, Anatoly; Hozak, Pavel; Peknicova, Jana

2015-03-08

Sperm proteins are important for the sperm cell function in fertilization. Some of them are involved in the binding of sperm to the egg. We characterized the acrosomal sperm protein detected by a monoclonal antibody (MoAb) (Hs-8) that was prepared in our laboratory by immunization of BALB/c mice with human ejaculated sperms and we tested the possible role of this protein in the binding assay. Indirect immunofluorescence and immunogold labelling, gel electrophoresis, Western blotting and protein sequencing were used for Hs-8 antigen characterization. Functional analysis of GAPDHS from the sperm acrosome was performed in the boar model using sperm/zona pellucida binding assay. Monoclonal antibody Hs-8 is an anti-human sperm antibody that cross-reacts with the Hs-8-related protein in spermatozoa of other mammalian species (boar, mouse). In the immunofluorescence test, Hs-8 antibody recognized the protein localized in the acrosomal part of the sperm head and in the principal piece of the sperm flagellum. In immunoblotting test, MoAb Hs-8 labelled a protein of 45 kDa in the extract of human sperm. Sequence analysis identified protein Hs-8 as GAPDHS (glyceraldehyde 3-phosphate dehydrohenase-spermatogenic). For this reason, commercial mouse anti-GAPDHS MoAb was applied in control tests. Both antibodies showed similar staining patterns in immunofluorescence tests, in electron microscopy and in immunoblot analysis. Moreover, both Hs-8 and anti-GAPDHS antibodies blocked sperm/zona pellucida binding. GAPDHS is a sperm-specific glycolytic enzyme involved in energy production during spermatogenesis and sperm motility; its role in the sperm head is unknown. In this study, we identified the antigen with Hs8 antibody and confirmed its localization in the apical part of the sperm head in addition to the principal piece of the flagellum. In an indirect binding assay, we confirmed the potential role of GAPDHS as a binding protein that is involved in the secondary sperm

Osmotic tolerance of avian spermatozoa: Influence of time, temperature, cryoprotectant and membrane ion pump function on sperm viability

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Blanco, J.M.; Long, J.A.; Gee, G.; Donoghue, A.M.; Wildt, D.E.

2008-01-01

Potential factors influencing sperm survival under hypertonic conditions were evaluated in the Sandhill crane (Grus canadensis) and turkey (Meleagridis gallopavo). Sperm osmotolerance (300-3000 mOsm/kg) was evaluated after: (1) equilibration times of 2, 10, 45 and 60 min at 4 ?C versus 21 ?C; (2) pre-equilibrating with dimethylacetamide (DMA) or dimethylsulfoxide (Me2SO) at either 4 ?C or 21 ?C; and (3) inhibition of the Na+/K+ and the Na+/H+ antiporter membrane ionic pumps. Sperm viability was assessed using the eosin-nigrosin live/dead stain. Species-specific differences occurred in response to hypertonic conditions with crane sperm remaining viable under extreme hypertonicity (3000 mOsm/kg), whereas turkey sperm viability was compromised with only slightly hypertonic (500 mOsm/kg) conditions. The timing of spermolysis under hypertonic conditions was also species-specific, with a shorter interval for turkey (2 min) than crane (10 min) sperm. Turkey sperm osmotolerance was slightly improved by lowering the incubation temperature from 21 to 4 ?C. Pre-equilibrating sperm with DMA reduced the incidence of hypertonic spermolysis only in the crane, at both room and refrigeration temperature. Inhibiting the Na+/K+ and the Na+/H+ antiporter membrane ion pumps did not impair resistance of crane and turkey spermatozoa to hypertonic stress; pump inhibition actually increased turkey sperm survival compared to control sperm. Results demonstrate marked species specificity in osmotolerance between crane and turkey sperm, as well as in the way temperature and time of exposure affect sperm survival under hypertonic conditions. Differences are independent of the role of osmotic pumps in these species.

Effects of reduced glutathione on acrosin activity in frozen-thawed boar spermatozoa.

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Estrada, Efrén; Rodríguez-Gil, Joan E; Rivera Del Álamo, Maria M; Peña, Alejandro; Yeste, Marc

2017-02-01

In pigs, acrosin activity in extended semen is correlated with reproductive performance and has recently been identified as a freezability marker. Reduced glutathione (GSH) is known to decrease sperm cryodamage and increase the reproductive performance of frozen-thawed boar spermatozoa. However, the effects of GSH on the acrosin activity of good and poor freezability ejaculates (GFE and PFE, respectively) is yet to be examined. The present study investigated how supplementing cryopreservation media with GSH affected acrosin activity in GFE and PFE, as well as the relationship between acrosin activity and reproductive performance in frozen-thawed boar spermatozoa. In addition, we examined whether the increase in fertility rates and litter sizes observed after the addition of 2mM GSH to cryopreservation extenders was related to acrosin activity. Supplementing freezing media with 2mM GSH partially counteracted the cryopreservation-related decrease in acrosin activity in GFE but not PFE. Acrosin activity was found to be significantly correlated with in vivo reproductive performance of frozen-thawed boar semen. In conclusion, the effects of adding GSH to freezing extenders on the acrosin activity of frozen-thawed boar spermatozoa rely on the intrinsic freezability of the ejaculate. Furthermore, the maintenance of proper acrosin activity could contribute to the increase in reproductive performance mediated by GSH.

Hyaluronic acid binding ability of human sperm reflects cellular maturity and fertilizing potential: selection of sperm for intracytoplasmic sperm injection.

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Huszar, Gabor; Ozkavukcu, Sinan; Jakab, Attila; Celik-Ozenci, Ciler; Sati, G Leyla; Cayli, Sevil

2006-06-01

The current concepts of sperm biochemical markers and the central role of the HspA2 chaperone protein, a measure of sperm cellular maturity and fertilizing potential, are reviewed. Because HspA2 is a component of the synaptonemal complex, low HspA2 levels and increased frequency of chromosomal aneuploidies are related in diminished maturity sperm. We also suggest a relationship between HspA2 expression in elongating spermatids and events of late spermiogenesis, such as cytoplasmic extrusion and plasma membrane remodeling that aid the formation of the zona pellucida binding and hyaluronic acid binding sites. The presence of hyaluronic acid receptor on the plasma membrane of mature sperm, coupled with hyaluronic acid coated glass or plastic surfaces, facilitates testing of sperm function and selection of single mature sperm for intracytoplasmic sperm injection. The frequencies of sperm with chromosomal disomy are reduced approximately fourfold to fivefold in hyaluronic acid selected sperm compared with semen sperm, comparable to the increase in such abnormalities in intracytoplasmic sperm injection offspring. Hyaluronic acid binding also excludes immature sperm with cytoplasmic extrusion, persistent histones, and DNA chain breaks. Hyaluronic acid mediated sperm selection is a novel technique that is comparable to sperm zona pellucida binding. Hyaluronic acid selected sperm will also alleviate the risks related to intracytoplasmic sperm injection fertilization with sperm of diminished maturity that currently cause worldwide concern.

Dose rates of antimicrobial substances in boar semen preservation-time to establish new protocols.

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Schulze, M; Grobbel, M; Riesenbeck, A; Brüning, S; Schaefer, J; Jung, M; Grossfeld, R

2017-06-01

To achieve a standardized number of spermatozoa in the final AI dose, varying amounts of extender fluid with a fixed concentration of antimicrobial substances are currently added to boar ejaculates. This practice ignores the different degrees of dilution of the antimicrobials in the end product. In calculating the final concentration of gentamicin in AI doses from 27,538 processed boar ejaculates, we demonstrated varying gentamicin concentrations in the resultant extended boar semen samples. The median concentration was 220.37 mg/L. In 25 of the samples (0.09%), the gentamicin concentration fell below 5 mg/L, which is close to or below the epidemiological cut-off value for many bacteria. We calculated the minimum inhibitory concentration of gentamicin for bacteria isolated from raw and extended ejaculates. Five of the isolates from extended ejaculates exceeded the maximum test concentration of 512 mg/L. As a result, we are presenting an alternative method of boar semen preservation whereby a particular combination of gentamicin concentrate and antibiotic-free extender is incorporated that standardizes the antibiotic concentration in the diluted semen. The addition of standardized antibiotic concentrations did not negatively affect sperm quality when compared to the use of ready-to-use extenders. In conclusion, an end volume-based and standardized addition of gentamicin to boar ejaculates can be a helpful alternative to prevent insufficient dosage of antibiotics in liquid preserved boar semen without affecting semen quality. © 2017 Blackwell Verlag GmbH.

Presence and function of dopamine transporter (DAT in stallion sperm: dopamine modulates sperm motility and acrosomal integrity.

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Javier A Urra

Full Text Available Dopamine is a catecholamine with multiple physiological functions, playing a key role in nervous system; however its participation in reproductive processes and sperm physiology is controversial. High dopamine concentrations have been reported in different portions of the feminine and masculine reproductive tract, although the role fulfilled by this catecholamine in reproductive physiology is as yet unknown. We have previously shown that dopamine type 2 receptor is functional in boar sperm, suggesting that dopamine acts as a physiological modulator of sperm viability, capacitation and motility. In the present study, using immunodetection methods, we revealed the presence of several proteins important for the dopamine uptake and signalling in mammalian sperm, specifically monoamine transporters as dopamine (DAT, serotonin (SERT and norepinephrine (NET transporters in equine sperm. We also demonstrated for the first time in equine sperm a functional dopamine transporter using 4-[4-(Dimethylaminostyryl]-N-methylpyridinium iodide (ASP(+, as substrate. In addition, we also showed that dopamine (1 mM treatment in vitro, does not affect sperm viability but decreases total and progressive sperm motility. This effect is reversed by blocking the dopamine transporter with the selective inhibitor vanoxerine (GBR12909 and non-selective inhibitors of dopamine reuptake such as nomifensine and bupropion. The effect of dopamine in sperm physiology was evaluated and we demonstrated that acrosome integrity and thyrosine phosphorylation in equine sperm is significantly reduced at high concentrations of this catecholamine. In summary, our results revealed the presence of monoamine transporter DAT, NET and SERT in equine sperm, and that the dopamine uptake by DAT can regulate sperm function, specifically acrosomal integrity and sperm motility.

Soluble products of Escherichia coli induce mitochondrial dysfunction-related sperm membrane lipid peroxidation which is prevented by lactobacilli.

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Arcangelo Barbonetti

Full Text Available Unidentified soluble factors secreted by E. coli, a frequently isolated microorganism in genitourinary infections, have been reported to inhibit mitochondrial membrane potential (ΔΨm, motility and vitality of human spermatozoa. Here we explore the mechanisms involved in the adverse impact of E. coli on sperm motility, focusing mainly on sperm mitochondrial function and possible membrane damage induced by mitochondrial-generated reactive oxygen species (ROS. Furthermore, as lactobacilli, which dominate the vaginal ecosystem of healthy women, have been shown to exert anti-oxidant protective effects on spermatozoa, we also evaluated whether soluble products from these microorganisms could protect spermatozoa against the effects of E. coli. We assessed motility (by computer-aided semen analysis, ΔΨm (with JC-1 dye by flow cytometry, mitochondrial ROS generation (with MitoSOX red dye by flow cytometry and membrane lipid-peroxidation (with the fluorophore BODIPY C11 by flow cytometry of sperm suspensions exposed to E. coli in the presence and in the absence of a combination of 3 selected strains of lactobacilli (L. brevis, L. salivarius, L. plantarum. A Transwell system was used to avoid direct contact between spermatozoa and microorganisms. Soluble products of E. coli induced ΔΨm loss, mitochondrial generation of ROS and membrane lipid-peroxidation, resulting in motility loss. Soluble factors of lactobacilli prevented membrane lipid-peroxidation of E. coli-exposed spermatozoa, thus preserving their motility. In conclusion, sperm motility loss by soluble products of E. coli reflects a mitochondrial dysfunction-related membrane lipid-peroxidation. Lactobacilli could protect spermatozoa in the presence of vaginal disorders, by preventing ROS-induced membrane damage.

Localized accumulation of cytosolic calcium near the fused sperm is associated with the calcium- and voltage-dependent block of sperm entry in the sea urchin egg.

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Ivonnet, Pedro I; Mohri, Tatsuma; McCulloh, David H

2017-10-01

Interaction of the sperm and egg depolarizes the egg membrane, allowing the sperm to enter; however, if the egg membrane is not allowed to depolarize from its resting potential (e.g., by voltage-clamp), the sperm will not enter. Previous studies demonstrated that sperm entry into sea urchin eggs that are voltage-clamped at negative membrane potentials is regulated both by the egg's membrane potential and a voltage-dependent influx of calcium into the egg. In these cases, electrical or cytoplasmic continuity (sperm-egg membrane fusion) occurs at negative membrane potentials, but subsequent loss of cytoplasmic continuity results in failure of sperm entry (unfusion). The work presented herein examined where, in relation to the sperm, and when, in relation to the sperm-induced electrophysiological events, the egg's calcium influx occurs, and how these events relate to successful or failed sperm entry. When sperm entered the egg, elevation of intracellular calcium concentration ([Ca 2+ ] i ) began near the fused sperm on average 5.9 s after sperm-egg membrane fusion. Conversely, when sperm failed to enter the egg, [Ca 2+ ] i elevated near the site of sperm-egg fusion on average 0.7 s after sperm-egg membrane fusion, which is significantly earlier than in eggs for which sperm entered. Therefore, the accumulation of calcium near the site of sperm-egg fusion is spatially and temporally consistent with the mechanism that may be responsible for loss of cytoplasmic continuity and failure of sperm entry. © 2017 Wiley Periodicals, Inc.

Evaluation of Lasting Effects of Heat Stress on Sperm Profile and Oxidative Status of Ram Semen and Epididymal Sperm

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Thais Rose dos Santos Hamilton

2016-01-01

Full Text Available Higher temperatures lead to an increase of testicular metabolism that results in spermatic damage. Oxidative stress is the main factor responsible for testicular damage caused by heat stress. The aim of this study was to evaluate lasting effects of heat stress on ejaculated sperm and immediate or long-term effects of heat stress on epididymal sperm. We observed decrease in motility and mass motility of ejaculated sperm, as well as an increase in the percentages of sperm showing major and minor defects, damaged plasma and acrosome membranes, and a decrease in the percentage of sperm with high mitochondrial membrane potential in the treated group until one spermatic cycle. An increased enzymatic activity of glutathione peroxidase and an increase of stressed cells were observed in ejaculated sperm of the treated group. A decrease in the percentage of epididymal sperm with high mitochondrial membrane potential was observed in the treated group. However, when comparing immediate and long-term effects, we observed an increase in the percentage of sperm with low mitochondrial membrane potential. In conclusion, testicular heat stress induced oxidative stress that led to rescuable alterations after one spermatic cycle in ejaculated sperm and also after 30 days in epididymal sperm.

A NEW MODEL OF BOAR SEMEN EVALUATION AND THE IMPACT OF CRYOGENIC FACTOR ON SPERMATIC CELLS

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A. V. RUSU

2009-05-01

Full Text Available Nowadays, sperm evaluation is mostly used to predict fertility and freezability. The aim of this study is to evaluate the possibility of investigating the effects of the cryogenic agent on boar spermatozoa, by identifying a set of laboratory tests for a rapid and efficient evaluation of semen quality. Usual sperm analysis such as sperm concentration, motility and spermatozoa morphology are not able to show subtle abnormalities, which are having a basic role in the fertilizing ability. Moreover, it seems that other sperm characteristics, involved in the fertilizing ability, can interfere with the freezing-thawing processes, being not evaluated or maybe not known. Morphological (microscopic analysis of stained spermatozoa, functional (motility analysis and hypo-osmotic swelling test and chromatin integrity (Acridine Orange Test and Comet Assay analysis were performed aiming to show the differences in spermatozoon integrity and functionality, caused by the cryogenic factor.

A comparative study of the morphometry of sperm head components in cattle, sheep, and pigs with a computer-assisted fluorescence method

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Jesús L Yániz

2016-01-01

Full Text Available The aim of this study was to compare the sperm nuclear and acrosomal morphometry of three species of domestic artiodactyls; cattle (Bos taurus, sheep (Ovis aries, and pigs (Sus scrofa. Semen smears of twenty ejaculates from each species were fixed and labeled with a propidium iodide-Pisum sativum agglutinin (PI/PSA combination. Digital images of the sperm nucleus, acrosome, and whole sperm head were captured and analyzed. The use of the PI/PSA combination and CASA-Morph fluorescence-based method allowed the capture, morphometric analysis, and differentiation of most sperm nuclei, acrosomes and whole heads, and the assessment of acrosomal integrity with a high precision in the three species studied. For the size of the head and nuclear area, the relationship between the three species may be summarized as bull > ram > boar. However, for the other morphometric parameters (length, width, and perimeter, there were differences in the relationships between species for sperm nuclei and whole sperm heads. Bull sperm acrosomes were clearly smaller than those in the other species studied and covered a smaller proportion of the sperm head. The acrosomal morphology, small in the bull, large and broad in the sheep, and large, long, and with a pronounced equatorial segment curve in the boar, was species-characteristic. It was concluded that there are clear variations in the size and shape of the sperm head components between the three species studied, the acrosome being the structure showing the most variability, allowing a clear distinction of the spermatozoa of each species.

Monitoring sperm mitochondrial respiration response in a laser trap using ratiometric fluorescence

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Mei, Adrian; Botvinick, Elliot; Berns, Michael

2005-08-01

Sperm motility is an important area in understanding male infertility. Various techniques, such as the Computer Assisted Sperm Analysis (CASA), have been used to understand sperm motility. Sperm motility is related to the energy (ATP) production of sperm. ATP is produced by the depolarization of the membrane potential of the inner membrane of the mitochondria. In this study, a mitochondrial dye, JC-1, has been used to monitor the energetics of the mitochondria. This fluorescent dye can emit at two different wavelengths, depending on the membrane potential of the mitochondria. It can fluoresce green at low membrane potential and red at high membrane potential. The ratio of the two colors (red/green) allows for an accurate measurement of the change of membrane potential. Various experiments were conducted to quantify the behavior of the dye within the sperm and the reaction of the sperm to trap. Sperm were trapped using laser tweezers. Results have shown that the ratio drops dramatically when sperm are trapped, indicating a depolarization of the membrane. The physiological response to this depolarization is yet to be determined, but the studies indicate that the sperm could have been slightly damaged by the laser. However, knowing that sperm depolarizes their membrane when trapped can help understand how sperm react to their environment and consequently help treat male infertility.

Metabolic incorporation of unsaturated fatty acids into boar spermatozoa lipids and de novo formation of diacylglycerols

DEFF Research Database (Denmark)

Svetlichnyy, V.; Müller, P.; Günther-Pomorski, Thomas

2014-01-01

Lipids play an important role in the maturation, viability and function of sperm cells. In this study, we examined the neutral and polar lipid composition of boar spermatozoa by thin-layer chromatography/mass spectrometry. Main representatives of the neutral lipid classes were diacylglycerols...... containing saturated (myristoyl, palmitoyl and stearoyl) fatty acyl residues. Glycerophosphatidylcholine and glycerophosphatidylethanolamine with alk(en)yl ether residues in the sn-1 position and unsaturated long chained fatty acyl residues in sn-2 position were identified as the most prominent polar lipids....... The only glycoglycerolipid was sulfogalactosylglycerolipid carrying 16:0-alkyl- and 16:0-acyl chains. Using stable isotope-labelling, the metabolic incorporation of exogenously supplied fatty acids was analysed. Boar spermatozoa incorporated hexadecenoic (16:1), octadecenoic (18:1), octadecadienoic (18...

Milk proteins interact with goat Binder of SPerm (BSP) proteins and decrease their binding to sperm.

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de Menezes, Erika Bezerra; van Tilburg, Mauricio; Plante, Geneviève; de Oliveira, Rodrigo V; Moura, Arlindo A; Manjunath, Puttaswamy

2016-11-01

Seminal plasma Binder of SPerm (BSP) proteins bind to sperm at ejaculation and promote capacitation. When in excess, however, BSP proteins damage the sperm membrane. It has been suggested that milk components of semen extenders associate with BSP proteins, potentially protecting sperm. Thus, this study was conducted to investigate if milk proteins interact with BSP proteins and reduce BSP binding to goat sperm. Using gel filtration chromatography, milk was incubated with goat seminal plasma proteins and loaded onto columns with and without calcium. Milk was also fractionated into parts containing mostly whey proteins or mostly caseins, incubated with seminal plasma proteins and subjected to gel filtration. Eluted fractions were evaluated by immunoblot using anti-goat BSP antibodies, confirming milk protein-BSP protein interactions. As determined by ELISA, milk proteins coated on polystyrene wells bound to increasing of goat BSP proteins. Far-western dot blots confirmed that BSP proteins bound to caseins and β-lactoglobulin in a concentration-dependent manner. Then, cauda epididymal sperm from five goats was incubated with seminal plasma; seminal plasma followed by milk; and milk followed by seminal plasma. Sperm membrane proteins were extracted and evaluated by immunoblotting. The pattern of BSP binding to sperm membrane proteins was reduced by 59.3 % when epididymal sperm were incubated with seminal plasma and then with skimmed milk (p  0.05). In conclusion, goat BSP proteins have an affinity for caseins and whey proteins. Milk reduces BSP binding to goat sperm, depending whether or not sperm had been previously exposed to seminal plasma. Such events may explain the protective effect of milk during goat sperm preservation.

A NEW MODEL OF BOAR SEMEN EVALUATION AND THE IMPACT OF CRYOGENIC FACTOR ON SPERMATIC CELLS

Directory of Open Access Journals (Sweden)

M. ZĂHAN

2009-05-01

Full Text Available Nowadays, sperm evaluation is mostly used to predict fertility and freezability. Theaim of this study is to evaluate the possibility of investigating the effects of thecryogenic agent on boar spermatozoa, by identifying a set of laboratory tests for arapid and efficient evaluation of semen quality. Usual sperm analysis such as spermconcentration, motility and spermatozoa morphology are not able to show subtleabnormalities, which are having a basic role in the fertilizing ability. Moreover, itseems that other sperm characteristics, involved in the fertilizing ability, can interferewith the freezing-thawing processes, being not evaluated or maybe not known.Morphological (microscopic analysis of stained spermatozoa, functional (motilityanalysis and hypo-osmotic swelling test and chromatin integrity (Acridine OrangeTest and Comet Assay analysis were performed aiming to show the differences inspermatozoon integrity and functionality, caused by the cryogenic factor

[Eosin Y-water test for sperm function examination].

Science.gov (United States)

Zha, Shu-wei; Lü, Nian-qing; Xu, Hao-qin

2015-06-01

Based on the principles of the in vitro staining technique, hypotonic swelling test, and water test, the Eosin Y-water test method was developed to simultaneously detect the integrity of the sperm head and tail and sperm membrane structure and function. As a widely used method in clinical laboratories in China, the Eosin Y-water test is methodologically characterized by three advantages. Firstly, both the sperm head and tail can be detected at the same time, which allows easy and comprehensive assessment of membrane damage in different parts of sperm. Secondly, distilled water is used instead of the usual formula solution to simplify and standardize the test by eliminating any potential effects on the water molecules through the sperm membrane due to different osmotic pressure or different sugar proportions and electrolyte solutions. Thirdly, the test takes less time and thus can be repeated before and after treatment. This article focuses on the fundamental principles and modification of the Eosin Y-water test and its application in sperm function examination and routine semen analysis for male infertility, assessment of the quality of sperm retrieved by testicular fine needle aspiration, semen cryopreservation program development, and evaluation of sperm membrane integrity after microwave radiation.

Ionic regulation of the plasma membrane potential of rainbow trout (Salmo gairdneri) spermatozoa: Role in the initiation of sperm motility

International Nuclear Information System (INIS)

Gatti, J.L.; Billard, R.; Christen, R.

1990-01-01

The ionic dependence of the trout sperm plasma membrane potential was analysed by measuring the accumulation of the lipophilic ions 3 H-tetraphenylphosphonium (TPP) and 14 C-thiocyanate (SCN) following dilution in artificial media isotonic to the seminal fluid. Our data showed that the trout sperm plasma membrane has a mixed conductance: the plasma membrane potential is sensitive upon the transmembrane gradients of K+, Na+, and H+. This potential is negative (less than -40 mV) in a 125 mM choline chloride media (ChM) at pH 8.5. Replacement of choline by sodium has a small depolarizing effect. The membrane potential is about -15 mV in a 125 mM potassium chloride and falls near zero mV only if valinomycin is added. In ChM changing the external pH (pHe) greatly affects the membrane potential: its value rises from less than -40 mV at pHe 9.0 to -17 mV at pHe 5.0. This pH effect is observed also in presence of sodium or potassium. A decrease in the transmembrane proton gradient produced by increasing internal pH without changing pHe induces also a depolarisation of the plasma membrane. In the different media in which trout sperm remain immotile after dilution (media with [K+] greater than 20-40 mM or a pH less than 7.5) the plasma membrane is more depolarized than in media allowing motility, suggesting a relationship between the state of membrane polarization and the intracellular effectors of the axonemal movement

Dilution of boar ejaculates with BTS containing HEPES in place of bicarbonate immediately after ejaculation can reduce the increased inducibility of the acrosome reaction by treatment with calcium and calcium ionophore A23187, which is potentially associated with boar subfertility.

Science.gov (United States)

Murase, Tetsuma; Imaeda, Noriaki; Yamada, Hiroto; Takasu, Masaki; Taguchi, Kazuo; Katoh, Tsutomu

2010-06-01

The present study investigated whether substitution of HEPES for bicarbonate in BTS (BTS-H) used to dilute boar ejaculates immediately after ejaculation could reduce the increased inducibility of the acrosome reaction by calcium and calcium ionophore A23187. When an ejaculate was split, diluted 5-fold with regular BTS (BTS-B) and BTS-H and stored at 17 C for 12 h or 60 h, the extender or storage time had no significant influence on sperm motility or viability measured by the eosin-nigrosin method. When spermatozoa diluted serially with BTS-B and stored (36 h) were stimulated with Ca2+ (3 mM) and A23187 (0.3 microM), the proportion of spermatozoa that underwent the acrosome reaction (% acrosome reactions) significantly increased as the magnifications of dilution increased (bicarbonate content almost unchanged by dilution). By contrast, the % acrosome reactions in spermatozoa similarly diluted and stored with BTS-H decreased with the increasing magnifications of dilution (bicarbonate decreased). Sperm motility immediately after the end of incubation without A23178 tended to be lower for BTS-H than BTS-B, and the ejaculates for BTS-H had a tendency to have a lower total protein in seminal plasma than those for BTS-B. These results implied that the samples for BTS-H could be used as a model for ejaculates possibly collected during summer and showing subfertility. When an ejaculate was split, diluted serially with BTS-B and BTS-H and stored, viability measured by staining with propidium iodide was extremely similar between the 2 extenders and among the different dilution magnifications, regardless of whether spermatozoa were washed (stored for 36-66 h) or not (stored for 66-72 h). These results suggest that boar ejaculate can be stored with BTS-H at least according to the results for sperm motility and viability and that hypersensitivity of spermatozoa to Ca2+ and A23187 potentially associated with boar subfertility could be lessened by diluting ejaculates with BTS-H.

Do antimicrobial peptides PR-39, PMAP-36 and PMAP-37 have any effect on bacterial growth and quality of liquid-stored boar semen?

Science.gov (United States)

Bussalleu, Eva; Sancho, Sílvia; Briz, Maria D; Yeste, Marc; Bonet, Sergi

2017-02-01

The use of antimicrobial peptides (AMP) has become one of the most promising alternatives to the use of antibiotics (Abs) in semen extender's formulation to overcome the increasing bacterial resistance to antibiotics. However, AMP may impair boar sperm quality, so that their deleterious effects might be higher than their effectiveness against bacteria. Thus, the aim of this study was to determine whether three different AMP, the proline-arginine-rich antimicrobial peptide PR-39 (PR-39), and the porcine myeloid antimicrobial peptides 36 (PMAP-36) and 37 (PMAP-37) had any effect upon boar sperm quality and bacterial growth. For this purpose, three different concentrations of each peptide (1 μM, 10 μM and 20 μM for PR-39 and 0.5 μM, 1 μM and 3 μM for PMAP-36 and PMAP-37) were added to 2 mL of a pool of extended semen with BTS without Abs; two controls, one without AMPs and Abs, and the other with Abs only were used for each peptide (n = 3). Total (TMOT) and progressive (PMOT) sperm motility, sperm viability and bacterial concentration were assessed before the addition of each AMP or Abs and at 1, 3, 6, 8 and 10 days post-addition. For each AMP, results revealed a drop in the TMOT and PMOT in all treatments and controls. In regard to sperm viability, while PR-39 at 10 μM maintained it in values similar to those of the control with Abs and PMAP-36 kept also the sperm viability in a similar fashion to the treatment with Abs, PMAP-37 was more effective in keeping sperm viability than controls (P semen, as it hardly impairs sperm viability and controls the bacterial load. Nevertheless, further studies are still required to improve its effectiveness. Copyright © 2016. Published by Elsevier Inc.

Regulatory properties of 6-phosphofructokinase and control of glycolysis in boar spermatozoa.

Science.gov (United States)

Kamp, G; Schmidt, H; Stypa, H; Feiden, S; Mahling, C; Wegener, G

2007-01-01

Glycolysis is crucial for sperm functions (motility and fertilization), but how this pathway is regulated in spermatozoa is not clear. This prompted to study the location and the regulatory properties of 6-phosphofructokinase (PFK, EC 2.7.1.11), the most important element for control of glycolytic flux. Unlike some other glycolytic enzymes, PFK showed no tight binding to sperm structures. It could readily be extracted from ejaculated boar spermatozoa by sonication and was then chromatographically purified. At physiological pH, the enzyme was allosterically inhibited by near-physiological concentrations of its co-substrate ATP, which induced co-operativity, i.e. reduced the affinity for the substrate fructose 6-phosphate. Inhibition by ATP was reinforced by citrate and H+. Above pH 8, PFK lost all its regulatory properties and showed maximum activity. However, in the physiological pH range, PFK activity was very sensitive to small changes in effectors. At near-physiological substrate concentrations, PFK activity requires activators (de-inhibitors) of which the combination of AMP and fructose 2,6-bisphosphate (F2,6P2) was most efficient as a result of synergistic effects. The kinetics of PFK suggest AMP, F2,6P2, H+, and citrate as allosteric effectors controlling PFK activity in boar spermatozoa. Using immunogold labeling, PFK was localized in the mid-piece and principal piece of the flagellum as well as in the acrosomal area at the top of the head and in the cytoplasmic droplets released from the mid-piece after ejaculation.

Effect of cooling to different sub-zero temperatures on boar sperm cryosurvival

OpenAIRE

Angelica Garcia-Olivares; Cesar Garzon-Perez; Oscar Gutierrez-Perez; Alfredo Medrano

2016-01-01

Objective: To compare different cooling temperatures before ice formation on pig sperm quality, before and after cryopreservation. Methods: Semen diluted in BF5 was cooled from 23 °C to 5 °C (1% glycerol, 200 × 106 cells/mL). Sperm were packaged in plastic straws, and maintained at +5 °C per 16 h. 1. Freezing point of diluted spermatozoa was determined by exposing straws to nitrogen vapors. 2. Straws (at +5 °C) were further cooled to −3 °C, −5 °C, and −7 °C, and rewarmed. 3. Straws (at +5 ...

Short communication Relationship between sperm plasma ...

African Journals Online (AJOL)

Matshidiso MB. Masenya

2017-01-04

Jan 4, 2017 ... observed between sperm plasma membrane integrity and fertility. There was a weak positive correlation between normal sperm morphology and conception rate (r = 0.11). Additionally ..... been approved by all of us. Authors' ...

Plasma Membrane Ca2+-ATPase 4 in Murine Epididymis: Secretion of Splice Variants in the Luminal Fluid and a Role in Sperm Maturation1

OpenAIRE

Patel, Ramkrishna; Al-Dossary, Amal A.; Stabley, Deborah L.; Barone, Carol; Galileo, Deni S.; Strehler, Emanuel E.; Martin-DeLeon, Patricia A.

2013-01-01

Plasma membrane Ca2+-ATPase isoform 4 (PMCA4) is the primary Ca2+ efflux pump in murine sperm, where it regulates motility. In Pmca4 null sperm, motility loss results in infertility. We have shown that murine sperm PMCA4b interacts with Ca2+/CaM-dependent serine kinase (CASK) in regulating Ca2+ homeostasis and motility. However, recent work indicated that the bovine PMCA4a splice variant (missing in testis) is epididymally expressed, along with 4b, and may be transferred to sperm. Here we sho...

Characterization of Na+K+-ATPase in bovine sperm.

Science.gov (United States)

Hickey, Katie D; Buhr, Mary M

2012-04-15

Existing as a ubiquitous transmembrane protein, Na(+)K(+)-ATPase affects sperm fertility and capacitation through ion transport and a recently identified signaling function. Functional Na(+)K(+)-ATPase is a dimer of α and β subunits, each with isoforms (four and three, respectively). Since specific isoform pairings and locations may influence or indicate function, the objective of this study was to identify and localize subunits of Na(+)K(+)-ATPase in fresh bull sperm by immunoblotting and immunocytochemistry using antibodies against α1 and 3, and all β isoforms. Relative quantity of Na(+)K(+)-ATPase in head plasma membranes (HPM's) from sperm of different bulls was determined by densitometry of immunoblot bands, and compared to bovine kidney. Sperm and kidney specifically bound all antibodies at kDa equivalent to commercial controls, and to additional lower kDa bands in HPM. Immunofluorescence of intact sperm confirmed that all isoforms were present in the head region of sperm and that α3 was also uniformly distributed post-equatorially. Permeabilization exposing internal membranes typically resulted in an increase in fluorescence, indicating that some antibody binding sites were present on the inner surface of the HPM or the acrosomal membrane. Deglycosylation of β1 reduced the kDa of bands in sperm, rat brain and kidney, with the kDa of the deglycosylated bands differing among tissues. Two-dimensional blots of β1 revealed three distinct spots. Based on the unique quantity, location and structure Na(+)K(+)-ATPase subunits in sperm, we inferred that this protein has unique functions in sperm. Copyright © 2012 Elsevier Inc. All rights reserved.

The Sperm-surface glycoprotein, SGP, is necessary for fertilization in the frog, Xenopus laevis.

Science.gov (United States)

Nagai, Keita; Ishida, Takuya; Hashimoto, Takafumi; Harada, Yuichirou; Ueno, Shuichi; Ueda, Yasushi; Kubo, Hideo; Iwao, Yasuhiro

2009-06-01

To identify a molecule involved in sperm-egg plasma membrane binding at fertilization, a monoclonal antibody against a sperm-surface glycoprotein (SGP) was obtained by immunizing mice with a sperm membrane fraction of the frog, Xenopus laevis, followed by screening of the culture supernatants based on their inhibitory activity against fertilization. The fertilization of both jellied and denuded eggs was effectively inhibited by pretreatment of sperm with intact anti-SGP antibody as well as its Fab fragment, indicating that the antibody recognizes a molecule on the sperm's surface that is necessary for fertilization. On Western blots, the anti-SGP antibody recognized large molecules, with molecular masses of 65-150 kDa and minor smaller molecules with masses of 20-28 kDa in the sperm membrane vesicles. SGP was distributed over nearly the entire surface of the sperm, probably as an integral membrane protein in close association with microfilaments. More membrane vesicles containing SGP bound to the surface were found in the animal hemisphere compared with the vegetal hemisphere in unfertilized eggs, but the vesicle-binding was not observed in fertilized eggs. These results indicate that SGP mediates sperm-egg membrane binding and is responsible for the establishment of fertilization in Xenopus.

Reproductive tissue expression and sperm localization of porcine beta-microseminoprotein

Czech Academy of Sciences Publication Activity Database

Maňásková-Postlerová, Pavla; Davidová, Nina; Šulc, Miroslav; Philimonenko, Anatoly; Hozák, Pavel; Jonáková, Věra

2011-01-01

Roč. 344, č. 2 (2011), s. 341-353 ISSN 0302-766X R&D Projects: GA ČR(CZ) GA303/09/1285; GA ČR GD523/08/H064; GA MZd(CZ) NS10009; GA MŠk(CZ) 1M06011; GA MŠk(CZ) LC07017; GA MŠk(CZ) LC545 Institutional research plan: CEZ:AV0Z50520701; CEZ:AV0Z50520514; CEZ:AV0Z50200510 Keywords : porcine beta-microseminoprotein * reproductive organs * spermatozoa * immunofluorescence * sperm acrosome * boar Subject RIV: CE - Biochemistry Impact factor: 3.114, year: 2011

An automatic system to study sperm motility and energetics.

Science.gov (United States)

Shi, Linda Z; Nascimento, Jaclyn M; Chandsawangbhuwana, Charlie; Botvinick, Elliot L; Berns, Michael W

2008-08-01

An integrated robotic laser and microscope system has been developed to automatically analyze individual sperm motility and energetics. The custom-designed optical system directs near-infrared laser light into an inverted microscope to create a single-point 3-D gradient laser trap at the focal spot of the microscope objective. A two-level computer structure is described that quantifies the sperm motility (in terms of swimming speed and swimming force) and energetics (measuring mid-piece membrane potential) using real-time tracking (done by the upper-level system) and fluorescent ratio imaging (done by the lower-level system). The communication between these two systems is achieved by a gigabit network. The custom-built image processing algorithm identifies the sperm swimming trajectory in real-time using phase contrast images, and then subsequently traps the sperm by automatically moving the microscope stage to relocate the sperm to the laser trap focal plane. Once the sperm is stably trapped (determined by the algorithm), the algorithm can also gradually reduce the laser power by rotating the polarizer in the laser path to measure the trapping power at which the sperm is capable of escaping the trap. To monitor the membrane potential of the mitochondria located in a sperm's mid-piece, the sperm is treated with a ratiometrically-encoded fluorescent probe. The proposed algorithm can relocate the sperm to the center of the ratio imaging camera and the average ratio value can be measured in real-time. The three parameters, sperm escape power, sperm swimming speed and ratio values of the mid-piece membrane potential of individual sperm can be compared with respect to time. This two-level automatic system to study individual sperm motility and energetics has not only increased experimental throughput by an order of magnitude but also has allowed us to monitor sperm energetics prior to and after exposure to the laser trap. This system should have application in both the

Methods of sperm vitality assessment.

Science.gov (United States)

Moskovtsev, Sergey I; Librach, Clifford L

2013-01-01

Sperm vitality is a reflection of the proportion of live, membrane-intact spermatozoa determined by either dye exclusion or osmoregulatory capacity under hypo-osmotic conditions. In this chapter we address the two most common methods of sperm vitality assessment: eosin-nigrosin staining and the hypo-osmotic swelling test, both utilized in clinical Andrology laboratories.

USE OF ALTERNATIVE EXTENDERS AND TEMPERATURES IN LONG TERM STORAGE OF BOAR SEMEN

Directory of Open Access Journals (Sweden)

Lina Raquel Santos Araújo

2016-01-01

Full Text Available The use of appropriate extenders is important for the success of an artificial insemination program. The objective of this study was to evaluate the efficiency of alternative extenders for swine semen at different temperatures (17 to 10 °C. The following extenders were used: Beltsville Thawing Solution (BTS, powdered coconut water (ACP-103®, and skimmed milk powder (LPD. The 50 ejaculates were analyzed daily, in natura and after dilution, during the 5-day period of semen preservation  (D0 to D4, regarding spermatic vigor and motility. Acrosome integrity and sperm viability were evaluated on D0 and D4. Data were analyzed using Mann-Whitney, Students, Tukey and chi-square tests (p<0.05. The LPD extender at 10 °C presented higher motility and sperm vigor compared to BTS and ACP until D2, and to treatments stored at 17 °C. Acrosome vitality and integrity remained higher (p<0.001 with LPD at 10 °C on D0 and D4. LPD showed to be a good extender for the swine semen at lower temperature (10 °C. Furthermore, it provided better protection to sperm cells, by allowing greater integrity and vitality of the acrosome. Keywords: coconut water; conservation; skimmed milk; semen boar.

Ion permeabilities in mouse sperm reveal an external trigger for SLO3-dependent hyperpolarization.

Directory of Open Access Journals (Sweden)

Julio C Chávez

Full Text Available Unlike most cells of the body which function in an ionic environment controlled within narrow limits, spermatozoa must function in a less controlled external environment. In order to better understand how sperm control their membrane potential in different ionic conditions, we measured mouse sperm membrane potentials under a variety of conditions and at different external K(+ concentrations, both before and after capacitation. Experiments were undertaken using both wild-type, and mutant mouse sperm from the knock-out strain of the sperm-specific, pH-sensitive, SLO3 K(+ channel. Membrane voltage data were fit to the Goldman-Hodgkin-Katz equation. Our study revealed a significant membrane permeability to both K(+ and Cl(- before capacitation, as well as Na(+. The permeability to both K(+ and Cl(- has the effect of preventing large changes in membrane potential when the extracellular concentration of either ion is changed. Such a mechanism may protect against undesired shifts in membrane potential in changing ionic environments. We found that a significant portion of resting membrane potassium permeability in wild-type sperm was contributed by SLO3 K(+ channels. We also found that further activation of SLO3 channels was the essential mechanism producing membrane hyperpolarization under two separate conditions, 1 elevation of external pH prior to capacitation and 2 capacitating conditions. Both conditions produced a significant membrane hyperpolarization in wild-type which was absent in SLO3 mutant sperm. Hyperpolarization in both conditions may result from activation of SLO3 channels by raising intracellular pH; however, demonstrating that SLO3-dependent hyperpolarization is achieved by an alkaline environment alone shows that SLO3 channel activation might occur independently of other events associated with capacitation. For example sperm may undergo stages of membrane hyperpolarization when reaching alkaline regions of the female genital tract

Cryopreservation of donkey sperm using non-permeable cryoprotectants.

Science.gov (United States)

Diaz-Jimenez, M; Dorado, J; Ortiz, I; Consuegra, C; Pereira, B; Gonzalez-De Cara, C A; Aguilera, R; Mari, G; Mislei, B; Love, C C; Hidalgo, M

2018-02-01

The aim of this study was to evaluate the effect of different concentrations of sucrose combined with bovine serum albumin (BSA), as non-permeable cryoprotectants, on donkey sperm parameters after cryopreservation, in comparison to a control extender containing glycerol. Semen from five Andalusian donkeys (n = 12) were centrifuged and resuspended with a commercial extender for equine sperm (Gent A, Minitube) adding 1% BSA and different concentrations (M, mol/l) of water-diluted sucrose: 0.05, 0.1, 0.25, 0.35 and 0.45. Thereafter, semen (n = 24) were diluted in the same base extender containing 0.25 M sucrose (S25) or glycerol (GLY, Gent B). Sperm were slowly cooled, filled in 0.5 ml straws and frozen in nitrogen vapours. Post-thaw samples were assessed for sperm motility, plasma membrane and DNA integrity and results were compared by ANOVA. In Experiment 1, sperm motility was significantly higher (P < 0.001) for S25 than the remaining treatments, and no differences were found for plasma membrane or DNA integrity. In Experiment 2, no differences were found between S25 or GLY for sperm motility and DNA integrity but plasma membrane integrity was significantly higher (P < 0.05) for S25. In conclusion, the extender with sucrose 0.25 M combined with BSA can be considered as an alternative to conventional extenders with glycerol for donkey sperm cryopreservation. Copyright © 2017 Elsevier B.V. All rights reserved.

Epididymosomes: transfer of fertility-modulating proteins to the sperm surface.

Science.gov (United States)

Martin-DeLeon, Patricia A

2015-01-01

A variety of glycosylphosphatidylinositol (GPI)-linked proteins are acquired on spermatozoa from epididymal luminal fluids (ELF) during sperm maturation. These proteins serve roles in immunoprotection and in key steps of fertilization such as capacitation, acrosomal exocytosis and sperm-egg interactions. Their acquisition on sperm cells is mediated both by membrane vesicles (epididymosomes, EP) which were first reported to dock on the sperm surface, and by lipid carriers which facilitate the transfer of proteins associated with the membrane-free fraction of ELF. While the nonvesicular fraction is more efficient, both pathways are dependent on hydrophobic interactions between the GPI-anchor and the external lipid layer of the sperm surface. More recently proteomic and hypothesis-driven studies have shown that EP from several mammals carry transmembrane (TM) proteins, including plasma membrane Ca 2 + -ATPase 4 (PMCA4). Synthesized in the testis, PMCA4 is an essential protein and the major Ca 2 + efflux pump in murine spermatozoa. Delivery of PMCA4 to spermatozoa from bovine and mouse EP during epididymal maturation and in vitro suggests that the docking of EP on the sperm surface precedes fusion, and experimental evidence supports a fusogenic mechanism for TM proteins. Fusion is facilitated by CD9, which generates fusion-competent sites on membranes. On the basis of knowledge of PMCA4's interacting partners a number of TM and membrane-associated proteins have been identified or are predicted to be present, in the epididymosomal cargo deliverable to spermatozoa. These Ca 2 + -dependent proteins, undetected in proteomic studies, play essential roles in sperm motility and fertility, and their detection highlights the usefulness of the hypothesis-driven approach.

Epididymosomes: transfer of fertility-modulating proteins to the sperm surface

Directory of Open Access Journals (Sweden)

Patricia A Martin-DeLeon

2015-01-01

Full Text Available A variety of glycosylphosphatidylinositol (GPI-linked proteins are acquired on spermatozoa from epididymal luminal fluids (ELF during sperm maturation. These proteins serve roles in immunoprotection and in key steps of fertilization such as capacitation, acrosomal exocytosis and sperm-egg interactions. Their acquisition on sperm cells is mediated both by membrane vesicles (epididymosomes, EP which were first reported to dock on the sperm surface, and by lipid carriers which facilitate the transfer of proteins associated with the membrane-free fraction of ELF. While the nonvesicular fraction is more efficient, both pathways are dependent on hydrophobic interactions between the GPI-anchor and the external lipid layer of the sperm surface. More recently proteomic and hypothesis-driven studies have shown that EP from several mammals carry transmembrane (TM proteins, including plasma membrane Ca 2 + -ATPase 4 (PMCA4. Synthesized in the testis, PMCA4 is an essential protein and the major Ca 2 + efflux pump in murine spermatozoa. Delivery of PMCA4 to spermatozoa from bovine and mouse EP during epididymal maturation and in vitro suggests that the docking of EP on the sperm surface precedes fusion, and experimental evidence supports a fusogenic mechanism for TM proteins. Fusion is facilitated by CD9, which generates fusion-competent sites on membranes. On the basis of knowledge of PMCA4′s interacting partners a number of TM and membrane-associated proteins have been identified or are predicted to be present, in the epididymosomal cargo deliverable to spermatozoa. These Ca 2 + -dependent proteins, undetected in proteomic studies, play essential roles in sperm motility and fertility, and their detection highlights the usefulness of the hypothesis-driven approach.

Effect of an isotonic lubricant on sperm collection and sperm quality.

Science.gov (United States)

Agarwal, Ashok; Malvezzi, Helena; Sharma, Rakesh

2013-05-01

To assess the influence of an isotonic lubricant used during sperm sample collection on [1] ease of collection and [2] resultant sperm quality. Paired randomized cross-over design. Tertiary hospital. Healthy men over 18 years old with normal semen analysis as per World Health Organization 2010 guidelines. Collection of semen sample from 22 subjects by masturbation with or without the use of Pre-Seed personal lubricant. Qualitative survey results and quantitative sperm function outcomes were measured to determine resultant sperm quality and collection experience with and without Pre-Seed lubricant. The qualitative questionnaire results showed that 73% of donors prefer the semen collection process with the isotonic lubricant and 55% recommended the use of lubricant in their everyday collection. The motility, viability, membrane integrity, levels of reactive oxygen species, total antioxidant capacity, and percentage of DNA damage in collected semen samples were not affected by the use of the lubricant. More donors prefer, and find it easier, to collect semen samples with the use of the lubricant. The isotonic lubricant Pre-Seed did not compromise sperm quality as evaluated in an array of sperm assays, suggesting its safe use in fertility patients as required during sperm collection. Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Sperm storage and duration of fertility in female ostriches ( Struthio ...

African Journals Online (AJOL)

, the duration of sperm storage and the fertile period following separation of sexes were investigated by egg break-out and by counting the sperm in the perivitelline membrane (spermOPVL) above the germinal disc (GD) region. Fertilisation ...

Plasma membrane Ca2+-ATPase 4 in murine epididymis: secretion of splice variants in the luminal fluid and a role in sperm maturation.

Science.gov (United States)

Patel, Ramkrishna; Al-Dossary, Amal A; Stabley, Deborah L; Barone, Carol; Galileo, Deni S; Strehler, Emanuel E; Martin-DeLeon, Patricia A

2013-07-01

Plasma membrane Ca(2+)-ATPase isoform 4 (PMCA4) is the primary Ca(2+) efflux pump in murine sperm, where it regulates motility. In Pmca4 null sperm, motility loss results in infertility. We have shown that murine sperm PMCA4b interacts with Ca(2+)/CaM-dependent serine kinase (CASK) in regulating Ca(2+) homeostasis and motility. However, recent work indicated that the bovine PMCA4a splice variant (missing in testis) is epididymally expressed, along with 4b, and may be transferred to sperm. Here we show, via conventional and in situ RT-PCR, that both the splice variants of Pmca4 mRNA are expressed in murine testis and throughout the epididymis. Immunofluorescence localized PMCA4a to the apical membrane of the epididymal epithelium, and Western analysis not only confirmed its presence but showed for the first time that PMCA4a and PMCA4b are secreted in the epididymal luminal fluid (ELF), from which epididymosomes containing PMCA4a were isolated. Flow cytometry indicated the presence of PMCA4a on mature caudal sperm where it was increased ~5-fold compared to caput sperm (detected by Western blotting) and ~2-fold after incubation in ELF, revealing in vitro uptake and implicating PMCA4a in epididymal sperm maturation. Coimmunoprecipitation using pan-PMCA4 antibodies, revealed that both variants associate with CASK, suggesting their presence in a complex. Because they have different kinetic properties for Ca(2+) transport and different abilities to bind to CASK, our study suggests a mechanism for combining the functional attributes of both PMCA4 variants, leading to heightened efficiency of the pump in the maintenance of Ca(2+) homeostasis, which is crucial for normal motility and male fertility.

Conception rate and litter size in multiparous sows after intrauterine insemination using frozen-thawed boar semen in a commercial swine herd in Thailand.

Science.gov (United States)

Chanapiwat, Panida; Olanratmanee, Em-On; Kaeoket, Kampon; Tummaruk, Padet

2014-10-01

The aim of the present study was to determine the conception rate and litter size in sows after fixed time intra-uterine insemination using frozen-thawed boar semen in a commercial swine herd in Thailand. Sixty-nine Landrace multiparous sows were randomly allocated into two groups, including control (n=36) and treatment (n=33). The control sows were inseminated with extended fresh semen (3 × 10(9) motile sperm/dose, 100 ml) at 24, 36 and 48 hr after the onset of estrus. The treatment sows were inseminated with frozen-thawed semen (2 × 10(9) motile sperm/dose, 20 ml) at 24 and 36 hr after induction of ovulation by human chorionic gonadotropin. All inseminations were carried out by using an intra-uterine insemination technique. The time of ovulation was determined by using transrectal real-time B-mode ultrasonography. The conception rate, farrowing rate, total number of piglets born/litter (TB) and number of piglets born alive/litter (BA) were evaluated. The sows inseminated with extended fresh semen yield a higher TB (10.8 versus 9.0 piglets/l, P=0.015) and tended to have a higher conception rate (88.9% versus 75.8%, P=0.150) than sows inseminated with frozen-thawed semen. In conclusion, insemination using frozen-thawed boar semen can be practiced with convinced fertility under field conditions by fixed-time intrauterine insemination with 2 × 10(9) sperm/ dose of 20 ml at 24 and 36 hr after the onset of estrus.

Conception Rate and Litter Size in Multiparous Sows after Intrauterine Insemination Using Frozen-Thawed Boar Semen in a Commercial Swine Herd in Thailand

Science.gov (United States)

CHANAPIWAT, Panida; OLANRATMANEE, Em-On; KAEOKET, Kampon; TUMMARUK, Padet

2014-01-01

ABSTRACT The aim of the present study was to determine the conception rate and litter size in sows after fixed time intra-uterine insemination using frozen-thawed boar semen in a commercial swine herd in Thailand. Sixty-nine Landrace multiparous sows were randomly allocated into two groups, including control (n=36) and treatment (n=33). The control sows were inseminated with extended fresh semen (3 × 109 motile sperm/dose, 100 ml) at 24, 36 and 48 hr after the onset of estrus. The treatment sows were inseminated with frozen-thawed semen (2 × 109 motile sperm/dose, 20 ml) at 24 and 36 hr after induction of ovulation by human chorionic gonadotropin. All inseminations were carried out by using an intra-uterine insemination technique. The time of ovulation was determined by using transrectal real-time B-mode ultrasonography. The conception rate, farrowing rate, total number of piglets born/litter (TB) and number of piglets born alive/litter (BA) were evaluated. The sows inseminated with extended fresh semen yield a higher TB (10.8 versus 9.0 piglets/l, P=0.015) and tended to have a higher conception rate (88.9% versus 75.8%, P=0.150) than sows inseminated with frozen-thawed semen. In conclusion, insemination using frozen-thawed boar semen can be practiced with convinced fertility under field conditions by fixed-time intrauterine insemination with 2 × 109 sperm/ dose of 20 ml at 24 and 36 hr after the onset of estrus. PMID:24954517

Boar Semen Studies

Science.gov (United States)

King, G. J.; Macpherson, J. W.

1966-01-01

A successful method for low temperature preservation of bull semen was modified for use with boar semen. Observations were made on the effects of varying cooling rate, equilibration time, freezing rate, glycerol concentration, method of glycerol addition, packaging containers, extender pH and tonicity. Observations indicate that boar semen should be cooled and frozen at a slower rate than bull semen. Within the ranges or methods examined, the other factors had little effect on recovery of motility after freezing. PMID:4226548

A role for carbohydrate recognition in mammalian sperm-egg binding

International Nuclear Information System (INIS)

Clark, Gary F.

2014-01-01

Highlights: • Mammalian sperm-egg binding as a carbohydrate dependent species recognition event. • The role of carbohydrate recognition in human, mouse and pig sperm-egg binding. • Historical perspective and future directions for research focused on gamete binding. - Abstract: Mammalian fertilization usually requires three sequential cell–cell interactions: (i) initial binding of sperm to the specialized extracellular matrix coating the egg known as the zona pellucida (ZP); (ii) binding of sperm to the ZP via the inner acrosomal membrane that is exposed following the induction of acrosomal exocytosis; and (iii) adhesion of acrosome-reacted sperm to the plasma membrane of the egg cell, enabling subsequent fusion of these gametes. The focus of this review is on the initial binding of intact sperm to the mammalian ZP. Evidence collected over the past fifty years has confirmed that this interaction relies primarily on the recognition of carbohydrate sequences presented on the ZP by lectin-like egg binding proteins located on the plasma membrane of sperm. There is also evidence that the same carbohydrate sequences that mediate binding also function as ligands for lectins on lymphocytes that can inactivate immune responses, likely protecting the egg and the developing embryo up to the stage of blastocyst hatching. The literature related to initial sperm-ZP binding in the three major mammalian models (human, mouse and pig) is discussed. Historical perspectives and future directions for research related to this aspect of gamete adhesion are also presented

A role for carbohydrate recognition in mammalian sperm-egg binding

Energy Technology Data Exchange (ETDEWEB)

Clark, Gary F., E-mail: clarkgf@health.missouri.edu

2014-08-01

Highlights: • Mammalian sperm-egg binding as a carbohydrate dependent species recognition event. • The role of carbohydrate recognition in human, mouse and pig sperm-egg binding. • Historical perspective and future directions for research focused on gamete binding. - Abstract: Mammalian fertilization usually requires three sequential cell–cell interactions: (i) initial binding of sperm to the specialized extracellular matrix coating the egg known as the zona pellucida (ZP); (ii) binding of sperm to the ZP via the inner acrosomal membrane that is exposed following the induction of acrosomal exocytosis; and (iii) adhesion of acrosome-reacted sperm to the plasma membrane of the egg cell, enabling subsequent fusion of these gametes. The focus of this review is on the initial binding of intact sperm to the mammalian ZP. Evidence collected over the past fifty years has confirmed that this interaction relies primarily on the recognition of carbohydrate sequences presented on the ZP by lectin-like egg binding proteins located on the plasma membrane of sperm. There is also evidence that the same carbohydrate sequences that mediate binding also function as ligands for lectins on lymphocytes that can inactivate immune responses, likely protecting the egg and the developing embryo up to the stage of blastocyst hatching. The literature related to initial sperm-ZP binding in the three major mammalian models (human, mouse and pig) is discussed. Historical perspectives and future directions for research related to this aspect of gamete adhesion are also presented.

Plasma membrane calcium ATPase 4 (PMCA4) co-ordinates calcium and nitric oxide signaling in regulating murine sperm functional activity.

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Olli, Kristine E; Li, Kun; Galileo, Deni S; Martin-DeLeon, Patricia A

2018-01-01

Reduced sperm motility (asthenospermia) and resulting infertility arise from deletion of the Plasma Membrane Ca 2+ -ATPase 4 (Pmca4) gene which encodes the highly conserved Ca 2+ efflux pump, PMCA4. This is the major Ca 2+ clearance protein in murine sperm. Since the mechanism underlying asthenospermia in PMCA4's absence or reduced activity is unknown, we investigated if sperm PMCA4 negatively regulates nitric oxide synthases (NOSs) and when absent NO, peroxynitrite, and oxidative stress levels are increased. Using co-immunoprecipitation (Co-IP) and Fluorescence Resonance Energy Transfer (FRET), we show an association of PMCA4 with the NOSs in elevated cytosolic [Ca 2+ ] in capacitated and Ca 2+ ionophore-treated sperm and with neuronal (nNOS) at basal [Ca 2+ ] (ucapacitated sperm). FRET efficiencies for PMCA4-eNOS were 35% and 23% in capacitated and uncapacitated sperm, significantly (p < 0.01) different, with the molecules being <10 nm apart. For PMCA4-nNOS, this interaction was seen only for capacitated sperm where FRET efficiency was 24%, significantly (p < 0.05) higher than in uncapacitated sperm (6%). PMCA4 and the NOSs were identified as interacting partners in a quaternary complex that includes Caveolin1, which co-immunoprecipitated with eNOS in a Ca 2+ -dependent manner. In Pmca4 -/- sperm NOS activity was elevated twofold in capacitated/uncapacitated sperm (vs. wild-type), accompanied by a twofold increase in peroxynitrite levels and significantly (p < 0.001) increased numbers of apoptotic germ cells. The data support a quaternary complex model in which PMCA4 co-ordinates Ca 2+ and NO signaling to maintain motility, with increased NO levels resulting in asthenospermia in Pmca4 -/- males. They suggest the involvement of PMCA4 mutations in human asthenospermia, with diagnostic relevance. © 2017 Wiley Periodicals, Inc.

Plasma membrane Ca2+-ATPase 4: interaction with constitutive nitric oxide synthases in human sperm and prostasomes which carry Ca2+/CaM-dependent serine kinase

OpenAIRE

Andrews, Rachel E.; Galileo, Deni S.; Martin-DeLeon, Patricia A.

2015-01-01

Deletion of the gene encoding the widely conserved plasma membrane calcium ATPase 4 (PMCA4), a major Ca2+ efflux pump, leads to loss of sperm motility and male infertility in mice. PMCA4's partners in sperm and how its absence exerts its effect on fertility are unknown. We hypothesize that in sperm PMCA4 interacts with endothelial nitric oxide synthase (eNOS) and neuronal nitric oxide synthase (nNOS) which are rapidly activated by Ca2+, and that these fertility-modulating proteins are present...

Percoll gradient-centrifuged capacitated mouse sperm have increased fertilizing ability and higher contents of sulfogalactosylglycerolipid and docosahexaenoic acid-containing phosphatidylcholine compared to washed capacitated mouse sperm.

Science.gov (United States)

Furimsky, Anna; Vuong, Ngoc; Xu, Hongbin; Kumarathasan, Premkumari; Xu, Min; Weerachatyanukul, Wattana; Bou Khalil, Maroun; Kates, Morris; Tanphaichitr, Nongnuj

2005-03-01

Although Percoll gradient centrifugation has been used routinely to prepare motile human sperm, its use in preparing motile mouse sperm has been limited. Here, we showed that Percoll gradient-centrifuged (PGC) capacitated mouse sperm had markedly higher fertilizing ability (sperm-zona pellucida [ZP] binding and in vitro fertilization) than washed capacitated mouse sperm. We also showed that the lipid profiles of PGC capacitated sperm and washed capacitated sperm differed significantly. The PGC sperm had much lower contents of cholesterol and phospholipids. This resulted in relative enrichment of male germ cell-specific sulfogalactosylglycerolipid (SGG), a ZP-binding ligand, in PGC capacitated sperm, and this would explain, in part, their increased ZP-binding ability compared with that of washed capacitated sperm. Analyses of phospholipid fatty acyl chains revealed that PGC capacitated sperm were enriched in phosphatidylcholine (PC) molecular species containing highly unsaturated fatty acids (HUFAs), with docosahexaenoic acid (DHA; C22: 6n-3) being the predominant HUFA (42% of total hydrocarbon chains of PC). In contrast, the level of PC-HUFAs comprising arachidonic acid (20:4n-6), docosapentaenoic acid (C22:5n-6), and DHA in washed capacitated sperm was only 27%. Having the highest unsaturation degree among all HUFAs in PC, DHA would enhance membrane fluidity to the uppermost. Therefore, membranes of PGC capacitated sperm would undergo fertilization-related fusion events at higher rates than washed capacitated sperm. These results suggested that PGC mouse sperm should be used in fertilization experiments and that SGG and DHA should be considered to be important biomarkers for sperm fertilizing ability.

Perfringolysin O as a useful tool to study human sperm physiology.

Science.gov (United States)

Pocognoni, Cristián A; De Blas, Gerardo A; Heuck, Alejandro P; Belmonte, Silvia A; Mayorga, Luis S

2013-01-01

To evaluate perfringolysin O, a cholesterol-dependent pore-forming cytolysin, as a tool to study several aspects of human sperm physiology. Prospective study. Basic research laboratory. Human semen samples with normal parameters obtained from healthy donors. Interaction of recombinant perfringolysin O with human spermatozoa. Assessment of perfringolysin O binding to spermatozoa, tests for acrosome and plasma membrane integrity, and acrosomal exocytosis assays. Perfringolysin O associated with human spermatozoa at 4°C. The binding was sensitive to changes in cholesterol concentrations and distribution occurring in the plasma membrane of these cells during capacitation. When perfringolysin O-treated sperm were incubated at 37°C, the plasma membrane became permeable, whereas the acrosome membrane remained intact. Permeabilized spermatozoa were able to respond to exocytic stimuli. The process was inhibited by proteins that interfere with membrane fusion, indicating that large molecules, including antibodies, were able to permeate into the spermatozoa. PFO is a useful probe to assess changes in the amount and distribution of the active sterol fraction present in the sperm plasma membrane. The toxin can be used for the efficient and selective permeabilization of this membrane, rendering a flexible experimental model suitable for studying molecular processes occurring in the sperm cytoplasm. Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Boar semen bacterial contamination in Italy and antibiotic efficacy in a modified extender

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Carla Bresciani

2014-02-01

Full Text Available The aims of the study were to identify microbial flora in boar semen under field conditions in northern Italy, to investigate antibiotic resistance and sensitivity of isolated bacteria, and to evaluate elimination of bacteria after storage in two types of extenders added with different antibiotics (amikacin vs gentamicin. A total of 60 boars were collected in 13 pig farms. Bacteriological and mycological investigations were performed immediately on raw semen samples, then at 48 and 120 h of storage on semen diluted randomly in a new short-term modified extender (ME-S or in a commercial one (CRONOSTM. Bacterial contamination was found in 63% of raw semen samples and different bacterial species were isolated: E.coli, Serratia marcescens, Staphylococcus epidermidis and aureus, Proteus spp., Streptococcus spp. and Pseudomonas aeruginosa. E. coli was the most isolated contaminant (53%; Pseudomonas aeruginosa was found only in one semen sample. The analysis of variance of factors affecting contamination levels was significant for the farm of origin (P<0.05 and not significant for the breed. Antibiotic resistance of these bacteria was assessed using different antibiotics. Significant differences (P<0.05 between observed and expected frequencies of bacterial isolates resistant or not to the antibiotics contained in the extenders were found. At 48 h of storage a reduction of aerobic contamination was found after ME-S dilution by 85.3% and after CRONOSTM by 63.8%. This paper proved the presence of pathogenic bacteria in semen. We thus believe it is highly advisable to perform periodic microbiological screening of boar semen in the swine industry to avoid the use of low sperm quality.

Nutrition and Reproductive Health: Sperm versus Erythrocyte Lipidomic Profile and ω-3 Intake

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Gabriela Ruth Mendeluk

2015-01-01

Full Text Available Fatty acid analyses of sperm and erythrocyte cell membrane phospholipids in idiopathic infertile patients evidenced that erythrocyte contents of EPA, DHA, omega-6–omega-3 ratio and arachidonic acid provide a mathematical correspondence for the prediction of EPA level in sperm cells. The erythrocyte lipidomic profile of patients was significantly altered, with signatures of typical Western pattern dietary habits and no fish intake. A supplementation with nutritional levels of EPA and DHA and antioxidants was then performed for 3 months, with the follow-up of both erythrocyte and sperm cell membranes composition as well as conventional sperm parameters. Some significant changes were found in the lipidomic membrane profile of erythrocyte but not in sperm cells, which correspondently did not show significant parameter ameliorations. This is the first report indicating that membrane lipids of different tissues do not equally metabolize the fatty acid elements upon supplementation. Molecular diagnostic tools are necessary to understand the cell metabolic turnover and monitor the success of nutraceuticals for personalized treatments.

The effect of cooling to different subzero temperatures on dog sperm cryosurvival.

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Alcantar-Rodriguez, A; Medrano, A

2017-06-01

The objective was to assess the effect of cooling to different subzero temperatures around ice formation (-5°C) on dog sperm cryosurvival and plasma membrane fluidity. Semen was centrifuged, and sperm were resuspended in a Tris-egg yolk medium (3% glycerol). Diluted sperm were cooled from 22 to 5°C, and then, a Tris-egg yolk medium containing 7% glycerol was added (final concentration of 5% glycerol and 200 × 10 6  cells/ml). Sperm were packaged in 0.5-ml plastic straws, and equilibration was done 16 hr at 5°C before freezing. I. Straws (n = 47) at 5°C were exposed to nitrogen vapours to determine the freezing point. II. Other straws (from different ejaculates) processed as mentioned, were further cooled to -3, -5 or -7°C and immediately rewarmed in a water bath at 37°C. Motility, plasma membrane functionality and acrosome integrity were assessed. III. Other straws (from different ejaculates) processed as mentioned were further cooled to -3 or -5°C, frozen over nitrogen vapours and stored in liquid nitrogen for one month. Straws were thawed in a water bath at 38°C for 30 s. Motility, plasma membrane functionality, plasma membrane integrity, acrosome integrity, capacitation status and plasma membrane fluidity were assessed. Ice nucleation temperature was -14.3 ± 2.05°C (mean ± SD); cooling to +5, -3, -5 and -7°C, without freezing, produces no differences on sperm quality between target temperatures; cooling to +5, -3, and -5°C produced no differences on sperm survival and plasma membrane fluidity after freeze-thawing. In conclusion, cooling of dog spermatozoa to different subzero temperatures did not improve sperm cryosurvival and had no effect on plasma membrane fluidity after thawing. © 2017 Blackwell Verlag GmbH.

Comparison of the external physical damages between laser-assisted and mechanical immobilized human sperm using scanning electronic microscopy.

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David Y L Chan

Full Text Available We aim to visualize the external physical damages and distinct external phenotypic effects between mechanical and laser-assisted immobilized human spermatozoa using scanning electronic microscopy (SEM. Human spermatozoa were immobilized mechanically or with laser assistance for SEM examination and the membrane integrities were checked on both types of immobilized spermatozoa. We found evidence of external damages at SEM level on mechanically kinked sperm, but not on laser-assisted immobilized sperm. Although no external damage was found on laser-assist immobilized sperm, there were two distinct types of morphological changes when spermatozoa were stricken by infra-red laser. Coiled tails were immediately formed when Laser pulse was applied to the sperm end piece area, whereas laser applied to the sperm principal piece area resulted in a sharp bend of sperm tails. Sperm immobilized by laser did not exhibit any morphological change if the laser did not hit within the on-screen central target zone or if the laser hit the sperm mid piece or head. Our modified membrane integrity assay revealed that the external membrane of more than half of the laser-assisted immobilized sperm remained intact. In conclusion, mechanical immobilization produced membrane damages whilst laser-assisted immobilization did not result in any external membrane damages besides morphological changes at SEM level.

Mass spectrometry profiling of oxysterols in human sperm identifies 25-hydroxycholesterol as a marker of sperm function

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Chiara Zerbinati

2017-04-01

Full Text Available Cholesterol is a main lipid component of sperm cell that is essential for sperm membrane fluidity, capacitation, and acrosomal reaction. Recent data obtained in bovine sperm showed that sperm capacitation is associated to the formation of oxysterols, oxidized products of cholesterol. The aim of this study was to profile oxysterol content in human semen, and to investigate their potential role in sperm pathophysiology. Among the 12 oxysterols analyzed, 25-hydroxycholesterol (25-HC resulted the most represented in normozoospermic samples, and its concentration positively correlated with spermatozoa number. We detected Cholesterol 25-hydroxylase, the enzyme responsible for 25-HC production, in human spermatozoa at the level of the neck and the post acrosomal area. Upon incubation with spermatozoa, 25-HC induced calcium and cholesterol transients in connection with the acrosomal reaction. Our results support a role for 25-HC in sperm function.

Characterization of sperm surface protein patterns of ejaculated and capacitated boar sperm, with the detection of ZP binding candidates

Czech Academy of Sciences Publication Activity Database

Zigo, Michal; Jonáková, Věra; Šulc, Miroslav; Maňásková-Postlerová, Pavla

2013-01-01

Roč. 61, oct (2013), s. 322-328 ISSN 0141-8130 R&D Projects: GA ČR GAP503/12/1834 Institutional research plan: CEZ:AV0Z50520701 Institutional support: RVO:61388971 Keywords : Sperm surface protein * Zona pellucida-binding receptors * PKDREJ protein Subject RIV: CE - Biochemistry Impact factor: 3.096, year: 2013

Egg CD9 protein tides correlated with sperm oscillations tune the gamete fusion ability in mammal.

Science.gov (United States)

Ravaux, Benjamin; Favier, Sophie; Perez, Eric; Gourier, Christine

2018-01-23

Mammalian fertilization involves membrane events -adhesion, fusion, sperm engulfment, membrane block to polyspermy- whose causes remain largely unknown. Recently, specific oscillations of the sperm in contact with the egg were shown to be necessary for fusion. Using a microfluidic chip to impose the venue for the encounter of two gametes allowed real-time observation of the membrane remodelling occurring at the sperm/egg interface. The spatiotemporal mapping of egg CD9 revealed that this protein concentrates at the egg/sperm interface as a result of sperm oscillations, until a CD9-rich platform is nucleated on which fusion immediately takes place. Within 2 to 5 minutes after fusion, most of the CD9 leaves the egg for the external aqueous medium. Then an egg membrane wave engulfs the sperm head in approximately 25 minutes. These results show that sperm oscillations initiate the CD9 recruitment that causes gamete fusion after which CD9 and associated proteins leave the membrane in a process likely to contribute to block polyspermy. They highlight that the gamete fusion story in mammals is an unexpected interplay between mechanical constraints and proteins. © The Author(s) (2018). Published by Oxford University Press on behalf of Journal of Molecular Cell Biology, IBCB, SIBS, CAS. All rights reserved.

Epididymosomes: transfer of fertility-modulating proteins to the sperm surface

OpenAIRE

Patricia A Martin-DeLeon

2015-01-01

A variety of glycosylphosphatidylinositol (GPI)-linked proteins are acquired on spermatozoa from epididymal luminal fluids (ELF) during sperm maturation. These proteins serve roles in immunoprotection and in key steps of fertilization such as capacitation, acrosomal exocytosis and sperm-egg interactions. Their acquisition on sperm cells is mediated both by membrane vesicles (epididymosomes, EP) which were first reported to dock on the sperm surface, and by lipid carriers which facilitate the ...

Computer-assisted sperm analysis (CASA): capabilities and potential developments.

Science.gov (United States)

Amann, Rupert P; Waberski, Dagmar

2014-01-01

Computer-assisted sperm analysis (CASA) systems have evolved over approximately 40 years, through advances in devices to capture the image from a microscope, huge increases in computational power concurrent with amazing reduction in size of computers, new computer languages, and updated/expanded software algorithms. Remarkably, basic concepts for identifying sperm and their motion patterns are little changed. Older and slower systems remain in use. Most major spermatology laboratories and semen processing facilities have a CASA system, but the extent of reliance thereon ranges widely. This review describes capabilities and limitations of present CASA technology used with boar, bull, and stallion sperm, followed by possible future developments. Each marketed system is different. Modern CASA systems can automatically view multiple fields in a shallow specimen chamber to capture strobe-like images of 500 to >2000 sperm, at 50 or 60 frames per second, in clear or complex extenders, and in information for ≥ 30 frames and provide summary data for each spermatozoon and the population. A few systems evaluate sperm morphology concurrent with motion. CASA cannot accurately predict 'fertility' that will be obtained with a semen sample or subject. However, when carefully validated, current CASA systems provide information important for quality assurance of semen planned for marketing, and for the understanding of the diversity of sperm responses to changes in the microenvironment in research. The four take-home messages from this review are: (1) animal species, extender or medium, specimen chamber, intensity of illumination, imaging hardware and software, instrument settings, technician, etc., all affect accuracy and precision of output values; (2) semen production facilities probably do not need a substantially different CASA system whereas biology laboratories would benefit from systems capable of imaging and tracking sperm in deep chambers for a flexible period of time

Epididymal sperm from Spix's yellow-toothed cavies sperm successfully cryopreserved in Tris extender with 6% glycerol and 20% egg yolk.

Science.gov (United States)

Silva, Andréia M; Praxedes, Erica C G; Campos, Lívia B; Bezerra, Luana G P; Moreira, Samara S J; Maia, Keilla M; Souza, Ana L P; Silva, Alexandre R

2018-04-01

As a non-threatened hystricognath rodent species, Spix's yellow-toothed cavies can be used as a model for the development of assisted reproductive techniques for the conservation of closely related species. The objective was to establish a functional protocol for cryopreservation of epididymal sperm from these cavies. Twelve sexually mature males, ∼2 y old and weighing ∼300 g, were euthanized. Sperm were recovered by retrograde flushing of the vas deferens and cauda epididymis with Tris extender. Thereafter, sperm were extended in Tris plus 20% egg yolk, with 3%, 6% or 9% glycerol or dimethyl sulfoxide (DMSO), placed in 0.25 mL straws and cryopreserved in liquid nitrogen. Sperm concentration, motility (using computer-assisted sperm analysis; CASA), plasma membrane integrity, osmotic response, morphology and sperm binding-ability were determined in fresh and frozen-thawed sperm. For most sperm endpoints, glycerol was a more desirable cryoprotectant than DMSO. Data (mean ± SEM) were similar with use of 3%, 6%, and 9% glycerol (P > 0.05) in osmotic response (40.66 ± 6.3%, 42.5 ± 7.1%, and 39.5 ± 5.0% respectably), and membrane integrity (55.17 ± 5.5%, 68.4 ± 4.1%, and 59.1 ± 4.9% respectably). Among concentrations assessed, the use of 6% glycerol resulted in the greatest (P < 0.05) post-thaw values for total motility (60.9 ± 4.4%), rapid subpopulation motility (27.7 ± 3.1%) and sperm-binding capability (227.0 ± 20.2). In conclusion, epididymal sperm from the Spix's yellow-toothed cavies (G. spixii) are optimally cryopreserved in Tris extender with 6% glycerol and 20% egg yolk. Copyright © 2018 Elsevier B.V. All rights reserved.

Characterizing the glycocalyx of poultry spermatozoa: III. Semen cryopreservation methods alter the carbohydrate component of rooster sperm membrane glycoconjugates.

Science.gov (United States)

Peláez, J; Bongalhardo, D C; Long, J A

2011-02-01

The carbohydrate-rich zone on the sperm surface is essential for inmunoprotection in the female tract and early gamete interactions. We recently have shown the glycocalyx of chicken sperm to be extensively sialylated and to contain residues of mannose, glucose, galactose, fucose, N-acetyl-galactosamine, N-acetyl-glucosamine, and N-acetyl-lactosamine. Our objective here was to evaluate the effects of 3 different cryopreservation methods on the sperm glycocalyx. Semen from roosters was pooled, diluted, cooled to 5°C, and aliquoted for cryopreservation using 6% dimethylacetamide (DMA), 11% dimethylsulfoxide (DMSO), or 11% glycerol (GOH). For the DMA method, semen was equilibrated for 1 min with cryoprotectant and rapidly frozen by dropping 25-µL aliquots into liquid nitrogen. For the other methods, semen was equilibrated for either 1 min (DMSO) or 20 min (GOH), loaded into straws, and frozen with a programmable freezer. Thawing rates mimicked the freezing rates (e.g., rapid for DMA; moderate for DMSO and GOH). Aliquots of thawed and fresh, unfrozen semen were incubated with 1 of 12 fluorescein isothiocyanate-conjugated lectins and counterstained with propidium iodide, and mean fluorescence intensity (MFI) was assessed by flow cytometry. For each lectin, the MFI of propidium iodide-negative (viable sperm) was compared among the fresh and frozen-thawed treatments (n = 5). For sperm frozen with GOH and DMA, the MFI of most lectins was similar (P > 0.05) to that of fresh sperm, whereas only 5 of 12 lectins were similar between fresh and DMSO-frozen sperm. Sperm from all 3 methods had higher (P < 0.05) MFI for lectins specific for N-acetyl-glucosamine and β-galactose than did fresh sperm. Fewer sperm were damaged (P < 0.001) with GOH than with DMA or DMSO, and membrane integrity was correlated with MFI for 9 of 12 lectins (P < 0.05). These data indicate that surface carbohydrates are altered during cryopreservation, and that cryoprotectant type and freezing

Heat shock proteins on the human sperm surface.

Science.gov (United States)

Naaby-Hansen, Soren; Herr, John C

2010-01-01

The sperm plasma membrane is known to be critical to fertilization and to be highly regionalized into domains of head, mid- and principal pieces. However, the molecular composition of the sperm plasma membrane and its alterations during genital tract passage, capacitation and the acrosome reaction remains to be fully dissected. A two-dimensional gel-based proteomic study previously identified 98 human sperm proteins which were accessible for surface labelling with both biotin and radioiodine. In this report twelve dually labelled protein spots were excised from stained gels or PDVF membranes and analysed by mass spectrometry (MS) and Edman degradation. Seven members from four different heat shock protein (HSP) families were identified including HYOU1 (ORP150), HSPC1 (HSP86), HSPA5 (Bip), HSPD1 (HSP60), and several isoforms of the two testis-specific HSP70 chaperones HSPA2 and HSPA1L. An antiserum raised against the testis-specific HSPA2 chaperone reacted with three 65kDa HSPA2 isoforms and three high molecular weight surface proteins (78-79kDa, 84kDa and 90-93kDa). These proteins, together with seven 65kDa HSP70 forms, reacted with human anti-sperm IgG antibodies that blocked in vitro fertilization in humans. Three of these surface biotinylated human sperm antigens were immunoprecipitated with a rabbit antiserum raised against a linear peptide epitope in Chlamydia trachomatis HSP70. The results indicate diverse HSP chaperones are accessible for surface labelling on human sperm. Some of these share epitopes with C. trachomatis HSP70, suggesting an association between genital tract infection, immunity to HSP70 and reproductive failure. 2009 Elsevier Ireland Ltd. All rights reserved.

Sperm variables as predictors of fertility in Black Castellana roosters; use in the selection of sperm donors for genome resource banking purposes

Energy Technology Data Exchange (ETDEWEB)

Santiago Moreno, J.; Lopez Sebastian, A.; Castano, C.; Coloma, M. A.; Gomez Brunet, A.; Toledano Diaz, A.; Prieto, M. T.; Campo, J. L.

2009-07-01

Semen was collected from 10 Black Castellana roosters and the classic sperm variables (ejaculate volume, sperm concentration and sperm motility) examined. In addition, the hypo-osmotic swelling test was used to investigate sperm cell membrane integrity, and acidic aniline blue staining used to screen for morphological abnormalities (including acrosome integrity) and to examine the condensation status of the chromatin. The latter was also examined by Gram staining. Large and small semen volumes were associated high and low sperm concentrations respectively (R2=0.04, P<0.05). The percentage of motile spermatozoa correlated strongly with the percentage of sperm cells showing an intact acrosome (R2=0.13, P<0.001) and with the percentage of morphologically normal spermatozoa (R2=0.04, P<0.05). The percentage of Gram positive spermatozoa was positively correlated with semen appearance (R2=0.12, P<0.05), sperm cell concentration (R2=0.13, P<0.05), and with the sperm motility variables studied (R2=0.14, P<0.05 for percentage mobility, and R2=0.12, P<0.05 for quality of movement). Only three of the 10 roosters, all with fertilisation potentials of 80-90%, were considered potential sperm donors for genome resource banking purposes. The remaining birds were all of low fertility (. 50%); in fact, some produced semen volumes too small to perform fertility tests. Semen volume and membrane integrity were found to be the best variables for predicting the fertilisation potential of rooster ejaculates. (Author) 37 refs.

The biopsy of the boar testes using ultrasonographic examination

Directory of Open Access Journals (Sweden)

Laima Liepa

2014-03-01

Full Text Available The biopsy of live animal testes is an important clinical manipulation to control spermatogenesis and reproductive system pathologies. The aim was to develop a method of boar testes biopsy using a biopsy gun with ultrasound guidance and to investigate the influence of this procedure on the boar testes parenchyma and quality of ejaculate. The biopsy was carried out in six 8-month-old boars. Fourteen days prior to and 21 days after biopsy, the quality of ejaculate was examined (weight of ejaculate; concentration and motility of spermatozoa with a seven-day intervals. Ultrasound images of the testes parenchyma were recorded three times: directly before and 15 minutes after the biopsy, then 21 days after the procedure. The testes biopsies of generally anesthetized boars were performed with the biopsy gun for needle biopsy with a 12cm long, disposable 16-gauge needle 1.8mm in diameter (Vitesse through 1cm skin incision in the depth of 1.2-1.6cm of parenchyma. Fifteen minutes after the biopsy, macroscopic injures of the parenchyma of all the boar testes were not detected in the ultrasound image. Twenty one days after biopsy, the hyperechogenic line 0.1-0.2cm in diameter was seen in the testes parenchyma of six boars in the depth of 1.2-1.6cm. The biopsy of boar testes did not influence the quality of boars ejaculate. The ultrasonographic examination of boar testicles before the biopsy reduced possibilities to traumatize large blood vessels of the testes. A perfect boar testicular biopsy was easy to perform using ultrasonographic examination in the pigsty conditions.

Differences in the fatty-acid composition of rodent spermatozoa are associated to levels of sperm competition

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Javier delBarco-Trillo

2015-03-01

Full Text Available Sperm competition is a prevalent phenomenon that drives the evolution of sperm function. High levels of sperm competition lead to increased metabolism to fuel higher sperm velocities. This enhanced metabolism can result in oxidative damage (including lipid peroxidation and damage to the membrane. We hypothesized that in those species experiencing high levels of sperm competition there are changes in the fatty-acid composition of the sperm membrane that makes the membrane more resistant to oxidative damage. Given that polyunsaturated fatty acids (PUFAs are the most prone to lipid peroxidation, we predicted that higher sperm competition leads to a reduction in the proportion of sperm PUFAs. In contrast, we predicted that levels of sperm competition should not affect the proportion of PUFAs in somatic cells. To test these predictions, we quantified the fatty-acid composition of sperm, testis and liver cells in four mouse species (genus Mus that differ in their levels of sperm competition. Fatty-acid composition in testis and liver cells was not associated to sperm competition levels. However, in sperm cells, as predicted, an increase in sperm competition levels was associated with an increase in the proportion of saturated fatty-acids (the most resistant to lipid peroxidation and by a concomitant decrease in the proportion of PUFAs. Two particular fatty acids were most responsible for this pattern (arachidonic acid and palmitic acid. Our findings thus indicate that sperm competition has a pervasive influence in the composition of sperm cells that ultimately may have important effects in sperm function.

Differences in the fatty-acid composition of rodent spermatozoa are associated to levels of sperm competition

Science.gov (United States)

delBarco-Trillo, Javier; Mateo, Rafael; Roldan, Eduardo R. S.

2015-01-01

Sperm competition is a prevalent phenomenon that drives the evolution of sperm function. High levels of sperm competition lead to increased metabolism to fuel higher sperm velocities. This enhanced metabolism can result in oxidative damage (including lipid peroxidation) and damage to the membrane. We hypothesized that in those species experiencing high levels of sperm competition there are changes in the fatty-acid composition of the sperm membrane that makes the membrane more resistant to oxidative damage. Given that polyunsaturated fatty acids (PUFAs) are the most prone to lipid peroxidation, we predicted that higher sperm competition leads to a reduction in the proportion of sperm PUFAs. In contrast, we predicted that levels of sperm competition should not affect the proportion of PUFAs in somatic cells. To test these predictions, we quantified the fatty-acid composition of sperm, testis and liver cells in four mouse species (genus Mus) that differ in their levels of sperm competition. Fatty-acid composition in testis and liver cells was not associated to sperm competition levels. However, in sperm cells, as predicted, an increase in sperm competition levels was associated with an increase in the proportion of saturated fatty-acids (the most resistant to lipid peroxidation) and by a concomitant decrease in the proportion of PUFAs. Two particular fatty acids were most responsible for this pattern (arachidonic acid and palmitic acid). Our findings thus indicate that sperm competition has a pervasive influence in the composition of sperm cells that ultimately may have important effects in sperm function. PMID:25795911

Effect of transfection and co-incubation of bovine sperm with exogenous DNA on sperm quality and functional parameters for its use in sperm-mediated gene transfer.

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Arias, María Elena; Sánchez-Villalba, Esther; Delgado, Andrea; Felmer, Ricardo

2017-02-01

Sperm-mediated gene transfer (SMGT) is based on the capacity of sperm to bind exogenous DNA and transfer it into the oocyte during fertilization. In bovines, the progress of this technology has been slow due to the poor reproducibility and efficiency of the production of transgenic embryos. The aim of the present study was to evaluate the effects of different sperm transfection systems on the quality and functional parameters of sperm. Additionally, the ability of sperm to bind and incorporate exogenous DNA was assessed. These analyses were carried out by flow cytometry and confocal fluorescence microscopy, and motility parameters were also evaluated by computer-assisted sperm analysis (CASA). Transfection was carried out using complexes of plasmid DNA with Lipofectamine, SuperFect and TurboFect for 0.5, 1, 2 or 4 h. The results showed that all of the transfection treatments promoted sperm binding and incorporation of exogenous DNA, similar to sperm incorporation of DNA alone, without affecting the viability. Nevertheless, the treatments and incubation times significantly affected the motility parameters, although no effect on the integrity of DNA or the levels of reactive oxygen species (ROS) was observed. Additionally, we observed that transfection using SuperFect and TurboFect negatively affected the acrosome integrity, and TurboFect affected the mitochondrial membrane potential of sperm. In conclusion, we demonstrated binding and incorporation of exogenous DNA by sperm after transfection and confirmed the capacity of sperm to spontaneously incorporate exogenous DNA. These findings will allow the establishment of the most appropriate method [intracytoplasmic sperm injection (ICSI) or in vitro fertilization (IVF)] of generating transgenic embryos via SMGT based on the fertilization capacity of transfected sperm.

Migration of the guinea pig sperm membrane protein PH-20 from one localized surface domain to another does not occur by a simple diffusion-trapping mechanism.

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Cowan, A E; Myles, D G; Koppel, D E

1991-03-01

The redistribution of membrane proteins on the surface of cells is a prevalent feature of differentiation in a variety of cells. In most cases the mechanism responsible for such redistribution is poorly understood. Two potential mechanisms for the redistribution of surface proteins are: (1) passive diffusion coupled with trapping, and (2) active translocation. We have studied the process of membrane protein redistribution for the PH-20 protein of guinea pig sperm, a surface protein required for sperm binding to the egg zona pellucida (P. Primakoff, H. Hyatt, and D. G. Myles (1985). J. Cell Biol. 101, 2239-2244). PH-20 protein is localized to the posterior head plasma menbrane of the mature sperm cell. Following the exocytotic acrosome reaction, PH-20 protein moves into the newly incorporated inner acrosomal membrane (IAM), placing it in a position favorable for a role in binding sperm to the egg zona pellucida (D. G. Myles, and P. Primakoff (1984), J. Cell Biol. 99, 1634-1641). To analyze the mechanistic basis for this protein migration, we have used fluorescence microscopy and digital image processing to characterize PH-20 protein migration in individual cells. PH-20 protein was observed to move against a concentration gradient in the posterior head plasma membrane. This result argues strongly against a model of passive diffusion followed by trapping in the IAM, and instead suggests that an active process serves to concentrate PH-20 protein toward the boundary separating the posterior head and IAM regions. A transient gradient of PH-20 concentration observed in the IAM suggests that once PH-20 protein reaches the IAM, it is freely diffusing. Additionally, we observed that migration of PH-20 protein was calcium dependent.

Plasma membrane Ca2+-ATPase 4: interaction with constitutive nitric oxide synthases in human sperm and prostasomes which carry Ca2+/CaM-dependent serine kinase.

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Andrews, Rachel E; Galileo, Deni S; Martin-DeLeon, Patricia A

2015-11-01

Deletion of the gene encoding the widely conserved plasma membrane calcium ATPase 4 (PMCA4), a major Ca(2+) efflux pump, leads to loss of sperm motility and male infertility in mice. PMCA4's partners in sperm and how its absence exerts its effect on fertility are unknown. We hypothesize that in sperm PMCA4 interacts with endothelial nitric oxide synthase (eNOS) and neuronal nitric oxide synthase (nNOS) which are rapidly activated by Ca(2+), and that these fertility-modulating proteins are present in prostasomes, which deliver them to sperm. We show that in human sperm PMCA4 is present on the acrosome, inner acrosomal membrane, posterior head, neck, midpiece and the proximal principal piece. PMCA4 localization showed inter- and intra-individual variation and was most abundant at the posterior head/neck junction, co-localizing with NOSs. Co-immunoprecipitations (Co-IP) revealed a close association of PMCA4 and the NOSs in Ca(2+) ionophore-treated sperm but much less so in uncapacitated untreated sperm. Fluorescence resonance energy transfer (FRET) showed a similar Ca(2+)-related association: PMCA4 and the NOSs are within 10 nm apart, and preferentially so in capacitated, compared with uncapacitated, sperm. FRET efficiencies varied, being significantly (P < 0.001) higher at high cytosolic Ca(2+) concentration ([Ca(2+)]c) in capacitated sperm than at low [Ca(2+)]c in uncapacitated sperm for the PMCA4-eNOS complex. These dynamic interactions were not seen for PMCA4-nNOS complexes, which had the highest FRET efficiencies. Further, along with Ca(2+)/CaM-dependent serine kinase (CASK), PMCA4 and the NOSs are present in the seminal plasma, specifically in prostasomes where Co-IP showed complexes similar to those in sperm. Finally, flow cytometry demonstrated that following co-incubation of sperm and seminal plasma, PMCA4 and the NOSs can be delivered in vitro to sperm via prostasomes. Our findings indicate that PMCA4 interacts simultaneously with the NOSs preferentially at

Effects of cryoprotectant treatments on bovine sperm function and osmolyte content

OpenAIRE

Setyawan, Erif E. M.; Cooper, Trevor G.; Widiasih, Dyah A.; Junaidi, Aris; Yeung, Ching-Hei

2009-01-01

The hypothesis that addition and removal of cryoprotectants to and from spermatozoa would initiate regulatory volume decrease, and lead to osmolyte loss and reduced sperm function, was tested. Common cryoprotectants, in the absence of freezing and thawing, affected bovine ejaculated spermatozoa by lowering their total and progressive motility in medium, reducing their migration through surrogate cervical mucus, damaging sperm head membranes and inducing sperm tail coiling. Sperm function was ...

Sperm-Hybrid Micromotor for Targeted Drug Delivery.

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Xu, Haifeng; Medina-Sánchez, Mariana; Magdanz, Veronika; Schwarz, Lukas; Hebenstreit, Franziska; Schmidt, Oliver G

2018-01-23

A sperm-driven micromotor is presented as a targeted drug delivery system, which is appealing to potentially treat diseases in the female reproductive tract. This system is demonstrated to be an efficient drug delivery vehicle by first loading a motile sperm cell with an anticancer drug (doxorubicin hydrochloride), guiding it magnetically, to an in vitro cultured tumor spheroid, and finally freeing the sperm cell to deliver the drug locally. The sperm release mechanism is designed to liberate the sperm when the biohybrid micromotor hits the tumor walls, allowing it to swim into the tumor and deliver the drug through the sperm-cancer cell membrane fusion. In our experiments, the sperm cells exhibited a high drug encapsulation capability and drug carrying stability, conveniently minimizing  toxic side effects and unwanted drug accumulation in healthy tissues. Overall, sperm cells are excellent candidates to operate in physiological environments, as they neither express pathogenic proteins nor proliferate to form undesirable colonies, unlike other cells or microorganisms. This sperm-hybrid micromotor is a biocompatible platform with potential application in gynecological healthcare, treating or detecting cancer or other diseases in the female reproductive system.

Na/K-ATPase regulates bovine sperm capacitation through raft- and non-raft-mediated signaling mechanisms.

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Rajamanickam, Gayathri D; Kastelic, John P; Thundathil, Jacob C

2017-11-01

Highly dynamic lipid microdomains (rafts) in the sperm plasma membrane contain several signaling proteins that regulate sperm capacitation. Na/K-ATPase isoforms (testis-specific isoform ATP1A4 and ubiquitous isoform ATP1A1) are abundant in bovine sperm plasma membrane. We previously reported that incubation of bovine sperm with ouabain, a specific Na/K-ATPase ligand, induced tyrosine phosphorylation of several sperm proteins during capacitation. The objective of this study was to investigate the roles of lipid rafts and non-rafts in Na/K-ATPase enzyme activity and signaling during bovine sperm capacitation. Content of ATP1A4 and, to a lesser extent, ATP1A1 was increased in raft and non-raft fractions of capacitated sperm, although non-raft enzyme activities of both isoforms were higher than the corresponding activities in rafts from capacitated sperm. Yet, ATP1A4 was the predominant isoform responsible for total Na/K-ATPase activity in both rafts and non-rafts. A comparative increase in phosphorylation of signaling molecules was observed in both raft (CAV1) and non-raft (EGFR and ERK1/2) membrane fractions during capacitation. Although SRC was phosphorylated in both membrane fractions, the non-raft fraction possessed more of this activated form. We also inferred, by immunoprecipitation, that ATP1A4 interacted with CAV1 and EGFR in the raft fraction, whereas interactions of ATP1A4 with SRC, EGFR, and ERK1/2 occurred in the non-raft fraction of ouabain-capacitated sperm; conversely, ATP1A1 interacted only with CAV1 in both fractions of uncapacitated and capacitated sperm. In conclusion, both raft and non-raft cohorts of Na/K-ATPase isoforms contributed to phosphorylation of signaling molecules during bovine sperm capacitation. © 2017 Wiley Periodicals, Inc.

Conception Rate and Litter Size in Multiparous Sows after Intrauterine Insemination Using Frozen-Thawed Boar Semen in a Commercial Swine Herd in Thailand

OpenAIRE

CHANAPIWAT, Panida; OLANRATMANEE, Em-On; KAEOKET, Kampon; TUMMARUK, Padet

2014-01-01

ABSTRACT The aim of the present study was to determine the conception rate and litter size in sows after fixed time intra-uterine insemination using frozen-thawed boar semen in a commercial swine herd in Thailand. Sixty-nine Landrace multiparous sows were randomly allocated into two groups, including control (n=36) and treatment (n=33). The control sows were inseminated with extended fresh semen (3 × 109 motile sperm/dose, 100 ml) at 24, 36 and 48 hr after the onset of estrus. The treatment s...

A new approach to sperm preservation based on bioenergetic theory.

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Froman, D P; Feltmann, A J

2010-04-01

To date, attempts to preserve chicken sperm have been based on a trial-and-error experimental approach. The present work outlines the development of an alternative approach based on empiricism and bioenergetic theory. In previous work, we found fowl sperm motility to be dependent on mitochondrial calcium cycling, phospholipase A(2), and long-chain fatty acids as an endogenous energy source. It is noteworthy that fowl sperm reside within the sperm storage tubules (SST) of the oviduct over an interval of days to weeks after insemination. In this regard, a model for in vivo sperm storage was developed and tested in additional previous research. Sperm penetration of the SST, sperm residence within the SST, and sperm egress from the SST can be explained in terms mitochondrial function. Understanding sperm function and longevity in terms of bioenergetics presented the possibility that sperm could be inactivated by disrupting mitochondrial calcium cycling and could thereby be preserved. However, this possibility also posed a problem: maintenance of the inner membrane potential of the mitochondrion within inactivated sperm. This report describes a series of experiments in which fowl sperm were inactivated by treatment with the calcium chelator tetrasodium 1,2-bis-(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid, and then reactivated by treatment with calcium ions. The effect of tetrasodium 1,2-bis-(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid on mitochondrial calcium cycling was confirmed by flow cytometry and confocal microscopy. When treated sperm were cooled to 10 degrees C, inactivated sperm could be reactivated throughout a 5-h storage interval. When stored sperm were held for 3 h before reactivation and insemination, fertility was 88% of the control. Storage did not affect hatchability. In summary, short-term storage was realized by manipulating mitochondrial function. We propose that 1) complex V consumes ATP within inactivated sperm and, by doing so, maintains

Capacitation dependent changes in the sperm plasma membrane influence porcine gamete interaction

NARCIS (Netherlands)

Flesch, F.M.

2000-01-01

Although the sperm cell was first seen through Van Leeuwenhoek’s microscope in the late seventieth century and despite much effort in the last 40 years in particular, we still do not know a great deal of the sperm cell and its interaction with the oocyte. Mammalian sperm-oocyte interaction is a

[Study on sperm damage caused by trichloroethylene in male rats].

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Wu, De-sheng; Yang, Lin-qing; Huang, Sui; Liu, Jian-jun; Xu, Xin-yun; Huang, Hai-yan; Gong, Chun-mei; Hu, Gong-hua; Liu, Qing-cheng; Yang, Xi-fei; Hong, Wen-xu; Zhou, Li; Huang, Xin-feng; Yuan, Jian-hui; Zhuang, Zhi-xiong

2013-11-01

To study in vitro sperm damage caused by trichloroethylene in male rats. Sperms of Sprague-Dawley (SD) rats were collected 4 hours after being contaminated by trichloroethylene of 0, 2, 4, 6, 8, and 10 mmol/L in vitro. Giemsa staining was performed to observe the morphological changes of sperms, and flow cytometer was used to detect the changes in mitochondrial membrane potential. The sperm motilities in 6, 8, and 10 mmol/L trichloroethylene groups decreased significantly compared with that in control group (P trichloroethylene groups were significantly higher than that in control group (Ptrichloroethylene groups and control group (Ptrichloroethylene can reduce sperm motility and increase the aberration rate and apoptosis rate of sperms in male SD rats.

Superoxide dismutase (SOD) in boar spermatozoa: purification, biochemical properties and changes in activity during semen storage (16°C) in different extenders.

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Orzołek, Aleksandra; Wysocki, Paweł; Strzeżek, Jerzy; Kordan, Władysław

2013-03-01

The antioxidant system in semen is composed of enzymes, low-molecular weight antioxidants and seminal plasma proteins. Loss of enzymatic activity of superoxide dismutase (SOD) during semen preservation may cause insufficient antioxidant defense of boar spermatozoa. The aim of this study was to isolate and characterize SOD molecular forms from spermatozoa and to describe changes in SOD activity in boar sperm during preservation at 16°C. Sperm extracts were prepared from fresh or diluted semen and used for SOD purification or activity measurement. Ion-exchange chromatography and gel filtration was used to purify SOD molecular forms. BTS, Dilu Cell, M III and Vitasem were used as diluents for 5-day storage of semen at +16°C. The molecular form of SOD released from spermatozoa after cold shock and homogenization had a molecular weight of approximately 67kDa. The activity of the SOD form was the highest at pH 10 within the temperature range between 20 and 45°C. The enzymatic activity of form released after cold shock was inhibited by H2O2 and diethyldithiocarbamate (DDC; by 65 and 40%, respectively). The SOD form released by homogenization was inhibited by H2O2 and DDC (40%). The molecular form released after urea treatment was a 30kDa protein with maximum activity at 20°C and pH 10. Enzymatic activity of this form was inhibited by H2O2 by 35%, DDC by 80% and 2-mercaptoethanol by 15%. The antigenic determinants of SOD isolated from boar seminal plasma and spermatozoa were similar to each other. Susceptibility of spermatozoa to cold shock increased during storage, but the differences between extenders were statistically non-significant. Copyright © 2013 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

Effects of alternative cryoprotectants, diluents, straw size and cholesterol addition on cryopreserved rooster sperm

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Rooster sperm do not cryopreserve well. This is due in part to osmotic changes that sperm undergo during addition and removal of cryoprotectant (CPA), as well as membrane damage during the membrane phase transition from fluid at 37 degrees C to gel at low temperatures. When a CPA is removed from spe...

Effects of alginate on frozen-thawed boar spermatozoa quality, lipid peroxidation and antioxidant enzymes activities.

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Hu, Jinghua; Geng, Guoxia; Li, Qingwang; Sun, Xiuzhu; Cao, Hualin; Liu, Yawei

2014-06-30

Although alginate was reported to play an important role as free radical scavengers in vitro and could be used as sources of natural antioxidants, there was no study about the cryoprotective effects of alginate on boar spermatozoa freezing. The objective of this research was to evaluate the effects of different concentrations of alginate added to the freezing extenders on boar spermatozoa motility, plasma membrane integrity, acrosomal integrity, mitochondrial activities, lipid peroxidation and antioxidative enzymes activities (SOD and GSH-Px) after thawing. Alginate was added to the TCG extender to yield six different final concentrations: 0, 0.2, 0.4, 0.6, 0.8, and 1.0mg/mL. The semen extender supplemented with various doses of alginate increased (Pboar spermatozoa acrosomal integrity at concentrations of 0.6, 0.8, 1.0mg/mL, compared with that of the control (Pextenders with the presence of alginate led to higher SOD and GSH-Px activities and lower MDA levels, in comparison to the control (Pboar spermatozoa motility, functional integrity and antioxidative capacity at appropriate concentrations. Therefore alginate could be employed as an effective cryoprotectant in boar spermatozoa cryopreservation. Copyright © 2014 Elsevier B.V. All rights reserved.

Boars for breeding: A study of methods of evaluation used at testing stations

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Elsi Ettala

1973-01-01

Full Text Available The phenotype testing of highly selected boars was studied. Two groups, each of 30 boars, were tested centrally at stations for growth rate and ultrasonically measured fat thickness. According to test points, made up of a combination of these two traits, the 5 best boars, 5 average boars and the 5 poorest boars where selected for progeny evaluation. In all 26 boars and 441 progeny were tested. The progeny evaluation showed that ultrasonic measurement of the fat thickness of the boars gave a very reliable estimate of the meatiness of their progeny. Those boars, as a group, giving the poorest carcass quality could be distinguished with statistical reliability from the other groups. The correlations between sires and progeny were significant for both daily gain (period 20—88 kg and feed efficiency. The importance of rate of growth and feed efficiency has been neglected in selection for breeding as the test points used for the selection of boars depended almost entirely on fat thickness. More than 30 % of both boars and progeny boars suffered from some form of difficulty in walking. 13 % of the boars were eliminated because of leg faults. Leg faults in progeny were mainly inherited or caused through injury. A phenotype evaluation of progeny boars accounts much more effectively for the variation in their carcass value than does a full barrow sib evaluation alone. For best results, a progeny boar phenotype evaluation should be combined with a full barrow sib evaluation.

Hypercholesterolemia Impaired Sperm Functionality in Rabbits

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Monclus, Maria A.; Cabrillana, Maria E.; Clementi, Marisa A.; Espínola, Leandro S.; Cid Barría, Jose L.; Vincenti, Amanda E.; Santi, Analia G.; Fornés, Miguel W.

2010-01-01

Hypercholesterolemia represents a high risk factor for frequent diseases and it has also been associated with poor semen quality that may lead to male infertility. The aim of this study was to analyze semen and sperm function in diet-induced hypercholesterolemic rabbits. Twelve adult White New Zealand male rabbits were fed ad libitum a control diet or a diet supplemented with 0.05% cholesterol. Rabbits under cholesterol-enriched diet significantly increased total cholesterol level in the serum. Semen examination revealed a significant reduction in semen volume and sperm motility in hypercholesterolemic rabbits (HCR). Sperm cell morphology was seriously affected, displaying primarily a “folded head”-head fold along the major axe-, and the presence of cytoplasmic droplet on sperm flagellum. Cholesterol was particularly increased in acrosomal region when detected by filipin probe. The rise in cholesterol concentration in sperm cells was determined quantitatively by Gas chromatographic-mass spectrometric analyses. We also found a reduction of protein tyrosine phosphorylation in sperm incubated under capacitating conditions from HCR. Interestingly, the addition of Protein Kinase A pathway activators -dibutyryl-cyclic AMP and iso-butylmethylxanthine- to the medium restored sperm capacitation. Finally, it was also reported a significant decrease in the percentage of reacted sperm in the presence of progesterone. In conclusion, our data showed that diet-induced hypercholesterolemia adversely affects semen quality and sperm motility, capacitation and acrosomal reaction in rabbits; probably due to an increase in cellular cholesterol content that alters membrane related events. PMID:20976152

Impaired protamination and sperm DNA damage in a Nellore bull with high percentages of morphological sperm defects in comparison to normospermic bulls

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J.T. Carreira

2015-04-01

Full Text Available The routine semen evaluation assessing sperm concentration, motility and morphology, does not identify subtle defects in sperm chromatin architecture. Bulls appear to have stable chromatin, with low levels of DNA fragmentation. However, the nature of fragmentation and its impact on fertility remain unclear and there are no detailed reports characterizing the DNA organization and damage in this species. The intensive genetic selection, the use of artificial insemination and in vitro embryo production associated to the cryopreservation process can contribute to the chromatin damage and highlights the importance of sperm DNA integrity for the success of these technologies. Frozen-thawed semen samples from three ejaculates from a Nellore bull showed high levels of morphological sperm abnormalities (55.8±5.1%, and were selected for complementary tests. Damage of acrosomal (76.9±8.9% and plasma membranes (75.7±9.3% as well as sperm DNA strand breaks (13.8±9.5% and protamination deficiency (3.7±0.6% were significantly higher compared to the values measured in the semen of five Nellore bulls with normospermia (24.3±3.3%; 24.5±6.1%; 0.6±0.5%; 0.4±0.6% for acrosome, plasma membrane, DNA breaks and protamine deficiency, respectively (P<0.05. Motility and percentage of spermatozoa with low mitochondrial potential showed no differences between groups. This study shows how routine semen analyses (in this case morphology may point to the length and complexity of sperm cell damage emphasizing the importance of sperm function testing.

AMP-activated kinase in human spermatozoa: identification, intracellular localization, and key function in the regulation of sperm motility

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Calle-Guisado, Violeta; de Llera, Ana Hurtado; Martin-Hidalgo, David; Mijares, Jose; Gil, Maria C; Alvarez, Ignacio S; Bragado, Maria J; Garcia-Marin, Luis J

2017-01-01

AMP-activated kinase (AMPK), a protein that regulates energy balance and metabolism, has recently been identified in boar spermatozoa where regulates key functional sperm processes essential for fertilization. This work's aims are AMPK identification, intracellular localization, and their role in human spermatozoa function. Semen was obtained from healthy human donors. Sperm AMPK and phospho-Thr172-AMPK were analyzed by Western blotting and indirect immunofluorescence. High- and low-quality sperm populations were separated by a 40%–80% density gradient. Human spermatozoa motility was evaluated by an Integrated Semen Analysis System (ISAS) in the presence or absence of the AMPK inhibitor compound C (CC). AMPK is localized along the human spermatozoa, at the entire acrosome, midpiece and tail with variable intensity, whereas its active form, phospho-Thr172-AMPK, shows a prominent staining at the acrosome and sperm tail with a weaker staining in the midpiece and the postacrosomal region. Interestingly, spermatozoa bearing an excess residual cytoplasm show strong AMPK staining in this subcellular compartment. Both AMPK and phospho-Thr172-AMPK human spermatozoa contents exhibit important individual variations. Moreover, active AMPK is predominant in the high motility sperm population, where shows a stronger intensity compared with the low motility sperm population. Inhibition of AMPK activity in human spermatozoa by CC treatment leads to a significant reduction in any sperm motility parameter analyzed: percent of motile sperm, sperm velocities, progressivity, and other motility coefficients. This work identifies and points out AMPK as a new molecular mechanism involved in human spermatozoa motility. Further AMPK implications in the clinical efficiency of assisted reproduction and in other reproductive areas need to be studied. PMID:27678462

AMP-activated kinase in human spermatozoa: identification, intracellular localization, and key function in the regulation of sperm motility

Directory of Open Access Journals (Sweden)

Violeta Calle-Guisado

2017-01-01

Full Text Available AMP-activated kinase (AMPK, a protein that regulates energy balance and metabolism, has recently been identified in boar spermatozoa where regulates key functional sperm processes essential for fertilization. This work′s aims are AMPK identification, intracellular localization, and their role in human spermatozoa function. Semen was obtained from healthy human donors. Sperm AMPK and phospho-Thr172-AMPK were analyzed by Western blotting and indirect immunofluorescence. High- and low-quality sperm populations were separated by a 40%-80% density gradient. Human spermatozoa motility was evaluated by an Integrated Semen Analysis System (ISAS in the presence or absence of the AMPK inhibitor compound C (CC. AMPK is localized along the human spermatozoa, at the entire acrosome, midpiece and tail with variable intensity, whereas its active form, phospho-Thr172-AMPK, shows a prominent staining at the acrosome and sperm tail with a weaker staining in the midpiece and the postacrosomal region. Interestingly, spermatozoa bearing an excess residual cytoplasm show strong AMPK staining in this subcellular compartment. Both AMPK and phospho-Thr172-AMPK human spermatozoa contents exhibit important individual variations. Moreover, active AMPK is predominant in the high motility sperm population, where shows a stronger intensity compared with the low motility sperm population. Inhibition of AMPK activity in human spermatozoa by CC treatment leads to a significant reduction in any sperm motility parameter analyzed: percent of motile sperm, sperm velocities, progressivity, and other motility coefficients. This work identifies and points out AMPK as a new molecular mechanism involved in human spermatozoa motility. Further AMPK implications in the clinical efficiency of assisted reproduction and in other reproductive areas need to be studied.

Cellular biophysics during freezing of rat and mouse sperm predicts post-thaw motility.

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Hagiwara, Mie; Choi, Jeung Hwan; Devireddy, Ramachandra V; Roberts, Kenneth P; Wolkers, Willem F; Makhlouf, Antoine; Bischof, John C

2009-10-01

Though cryopreservation of mouse sperm yields good survival and motility after thawing, cryopreservation of rat sperm remains a challenge. This study was designed to evaluate the biophysics (membrane permeability) of rat in comparison to mouse to better understand the cooling rate response that contributes to cryopreservation success or failure in these two sperm types. In order to extract subzero membrane hydraulic permeability in the presence of ice, a differential scanning calorimeter (DSC) method was used. By analyzing rat and mouse sperm frozen at 5 degrees C/min and 20 degrees C/min, heat release signatures characteristic of each sperm type were obtained and correlated to cellular dehydration. The dehydration response was then fit to a model of cellular water transport (dehydration) by adjusting cell-specific biophysical (membrane hydraulic permeability) parameters L(pg) and E(Lp). A "combined fit" (to 5 degrees C/min and 20 degrees C/min data) for rat sperm in Biggers-Whitten-Whittingham media yielded L(pg) = 0.007 microm min(-1) atm(-1) and E(Lp) = 17.8 kcal/mol, and in egg yolk cryopreservation media yielded L(pg) = 0.005 microm min(-1) atm(-1) and E(Lp) = 14.3 kcal/mol. These parameters, especially the activation energy, were found to be lower than previously published parameters for mouse sperm. In addition, the biophysical responses in mouse and rat sperm were shown to depend on the constituents of the cryopreservation media, in particular egg yolk and glycerol. Using these parameters, optimal cooling rates for cryopreservation were predicted for each sperm based on a criteria of 5%-15% normalized cell water at -30 degrees C during freezing in cryopreservation media. These predicted rates range from 53 degrees C/min to 70 degrees C/min and from 28 degrees C/min to 36 degrees C/min in rat and mouse, respectively. These predictions were validated by comparison to experimentally determined cryopreservation outcomes, in this case based on motility. Maximum

Sperm Na+, K+-ATPase and Ca2+-ATPase activity: A preliminary study of comparison of swim up and density gradient centrifugation methods for sperm preparation

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Lestari, Silvia W.; Larasati, Manggiasih D.; Asmarinah, Mansur, Indra G.

2018-02-01

As one of the treatment for infertility, the success rate of Intrauterine Insemination (IUI) is still relatively low. Several sperm preparation methods, swim-up (SU) and the density-gradient centrifugation (DGC) are frequently used to select for better sperm quality which also contribute to IUI failure. Sperm selection methods mainly separate the motile from the immotile sperm, eliminating the seminal plasma. The sperm motility involves the structure and function of sperm membrane in maintaining the balance of ion transport system which is regulated by the Na+, K+-ATPase, and Ca2+-ATPase enzymes. This study aims to re-evaluate the efficiency of these methods in selecting for sperm before being used for IUI and based the evaluation on sperm Na+,K+-ATPase and Ca2+-ATPase activities. Fourteen infertile men from couples who underwent IUI were involved in this study. The SU and DGC methods were used for the sperm preparation. Semen analysis was performed based on the reference value of World Health Organization (WHO) 2010. After isolating the membrane fraction of sperms, the Na+, K+-ATPase activity was defined as the difference in the released inorganic phosphate (Pi) with and without the existence of 10 mM ouabain in the reaction, while the Ca2+-ATPase was determined as the difference in Pi contents with and without the existence of 55 µm CaCl2. The prepared sperm demonstrated a higher percentage of motile sperm compared to sperm from the whole semen. Additionally, the percentage of motile sperm of post-DGC showed higher result than the sperm from post-SU. The velocity of sperm showed similar pattern with the percentage of motile sperm, in which the velocity of prepared sperm was higher than the sperm from whole semen. Furthermore, the sperm velocity of post-DGC was higher compared to the sperm from post-SU. The Na+, K+-ATPase activity of prepared sperm was higher compared to whole semen, whereas Na+, K+-ATPase activity in the post DGC was higher than post SU. The Ca2

The zinc transporter ZIPT-7.1 regulates sperm activation in nematodes.

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Yanmei Zhao

2018-06-01

Full Text Available Sperm activation is a fascinating example of cell differentiation, in which immotile spermatids undergo a rapid and dramatic transition to become mature, motile sperm. Because the sperm nucleus is transcriptionally silent, this transition does not involve transcriptional changes. Although Caenorhabditis elegans is a leading model for studies of sperm activation, the mechanisms by which signaling pathways induce this transformation remain poorly characterized. Here we show that a conserved transmembrane zinc transporter, ZIPT-7.1, regulates the induction of sperm activation in Caenorhabditis nematodes. The zipt-7.1 mutant hermaphrodites cannot self-fertilize, and males reproduce poorly, because mutant spermatids are defective in responding to activating signals. The zipt-7.1 gene is expressed in the germ line and functions in germ cells to promote sperm activation. When expressed in mammalian cells, ZIPT-7.1 mediates zinc transport with high specificity and is predominantly located on internal membranes. Finally, genetic epistasis places zipt-7.1 at the end of the spe-8 sperm activation pathway, and ZIPT-7.1 binds SPE-4, a presenilin that regulates sperm activation. Based on these results, we propose a new model for sperm activation. In spermatids, inactive ZIPT-7.1 is localized to the membranous organelles, which contain higher levels of zinc than the cytoplasm. When sperm activation is triggered, ZIPT-7.1 activity increases, releasing zinc from internal stores. The resulting increase in cytoplasmic zinc promotes the phenotypic changes characteristic of activation. Thus, zinc signaling is a key step in the signal transduction process that mediates sperm activation, and we have identified a zinc transporter that is central to this activation process.

Bacteriological and genetic assessment of game meat from Japanese wild boars.

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Naya, Yuka; Horiuchi, Motohiro; Ishiguro, Naotaka; Shinagawa, Morikazu

2003-01-15

Bacterial tests were used to assess bacterial contamination of game meat from Japanese wild boars. The bacterial contamination of wild boar meat was less than that of domestic pork, as determined by aerobic plate counts (APC) and coliform counts. None of the meat examined in this study was contaminated by Salmonella or E. coli O-157. To detect adulteration by domestic pig meat or European wild boar meat, 46 samples of game meat sold as Japanese wild boar were examined genetically. A total of 17 samples showed genetic haplotypes of European and Asian domestic pigs in the D-loop of mitochondrial DNA (mtDNA), and 16 samples showed nuclear glucosephosphate isomerase-processed pseudogene (GPIP) genotypes of European domestic pigs. The European GPIP genotypes of these samples were confirmed by PCR-RFLP analysis. These results indicate that some game meat sold as Japanese wild boar is adulterated by cross-breeding between pigs and wild boars or by contamination with meat from domestic pigs or European wild boars.

In vivo fertilizing capacity of deep frozen boar semen packaged in plastic bags and maxi-straws.

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Bwanga, C O; Hofmo, P O; Grevle, I S; Einarsson, S; Rodriguez-Martinez

1991-05-01

Pooled ejaculates from six fertile boars were frozen under controlled conditions in Teflon FEP-film plastic bags (5 ml) and maxi-straws (2.5 ml) using 3% glycerol as cryoprotectant. The percentages of both post-thaw motility and normal apical ridges were significantly higher (P less than 0.001) for the bags (54.5 and 75%) than for the maxi-straws (40.1 and 59.4%) respectively. For evaluation of the in vivo fertilizing capacity of the frozen-thawed spermatozoa, 26 gilts were inseminated once 24 h after the first observation of standing reflex in their second oestrus, with 5 ml of semen (containing 5 billion spermatozoa) reconstituted in 80 ml of BTS from either bags or maxi-straws. Ova were recovered from the oviducts/uteri 2-4 days following insemination and examined for cleavage and sperm binding to the zona pellucida (ZP). Significantly higher rates (P less than 0.02) of fertilized ova were found in the bag-inseminated (75%) than in maxi-straw inseminated gilts (63%); and similarly their ova had significantly more spermatozoa in the ZP, irrespective of whether they were fertilized or nonfertilized. This study confirmed that the plastic bags are suitable and may be used for packaging single insemination doses of deep frozen boar semen for routine A.I. work.

The Effects of Different Doses of Ketamine on Quality of Normal Ejaculated Sperm

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Forouzan Absalan

2014-07-01

Full Text Available Background: Ketamine, an injectable anesthetic in human and animal medicine, is also a recreational drug used by young adults. The aim of this study is to evaluate the effects of ketamine on membrane integrity, DNA fragmentation and sperm parameters in humans. Materials and Methods: This prospective study was conducted on 40 males with normal semen samples over one month (August 2012. Subjects were randomly allocated to four groups (Control and case I, II and III whose semen samples were adjusted to different concentrations of ketamine (1, 3, 5 μL for one hour. Sperm analysis was performed for routine parameters, motility and morphology. Evaluation of membrane integrity and DNA fragmentation was done by eosin-Y staining and the sperm chromatin dispersion (SCD test, respectively. The results were analyzed by ANOVA and Tukey’s tests. P≤0.05 was considered statistically significant. Results: Total sperm motility in all case groups were significantly lower compared with the control group. In case group III, progressive motility showed significant difference with case group II. After addition of ketamine, sperm had evidence of coiled tails in all case groups compared to the control group however this observation was not significant. Evaluation of membrane integrity showed the rate of necrospermia increased in all case groups. However, ketamine only significantly affected membrane integrity in case group III. SCD staining showed that in the control group nucleoids with medium halos (63.44 ± 1.2 were significantly different compared to the case groups I (15.44 ± 0.45, II (9.05±1.16 and III (10.55 ± 1.14, respectively. Between case groups, nucleoids with large and medium halos showed significant differences in case groups II and III compared with case group I. Nucleoids with medium halos were significantly different between case groups II and III. Conclusion: Ketamine, through its effect on membrane integrity and DNA fragmentation, decreased

Cryopreservation of boar semen. II: Effect of cooling rate and duration of freezing point plateau on boar semen frozen in mini- and maxi-straws and plastic bags.

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Bwanga, C O; Einarsson, S; Rodriguez-Martinez, H

1991-01-01

The post-thaw motility and the acrosome integrity of semen from 4 boars frozen with a programmable freezing machine, in mini (0.25 ml) and maxi (5 ml) plastic straws and in 10 x 5 cm Teflon FEP-plastic bags (0.12 mm thick, 5 ml), were compared. The freezing of the semen was monitored by way of thermo-couples placed in the straws and the bags. Three freezing programmes were used, namely A: from +5 degrees C, at a rate of 3 degrees C/min, to -6 degrees C, held for 1 min at -6 degrees C, and followed by a cooling rate of 20 degrees C/min to -100 degrees C; B: a similar curve except that there was no holding time at -6 degrees C and that the cooling rate was 30 degrees C/min, and C: from +5 degrees C to -100 degrees C, with a cooling rate of 35 degrees C/min, followed by storage in liquid N2. Despite the freezing curve assayed, both the mini-straws and the bags depicted much shorter freezing point plateaus as compared to the maxi-straws. Post-thaw sperm motility as well as the amount of normal apical ridges were equally significantly higher when semen was frozen in mini-straws or in bags than in maxi-straws. Significant differences in these post-thawing parameters were obtained between the freezing curves used. The stepwise freezing procedure A appeared as the best alternative for boar semen, considering this in vitro evaluation.

Sperm competition, sperm numbers and sperm quality in muroid rodents.

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Gómez Montoto, Laura; Magaña, Concepción; Tourmente, Maximiliano; Martín-Coello, Juan; Crespo, Cristina; Luque-Larena, Juan José; Gomendio, Montserrat; Roldan, Eduardo R S

2011-03-25

Sperm competition favors increases in relative testes mass and production efficiency, and changes in sperm phenotype that result in faster swimming speeds. However, little is known about its effects on traits that contribute to determine the quality of a whole ejaculate (i.e., proportion of motile, viable, morphologically normal and acrosome intact sperm) and that are key determinants of fertilization success. Two competing hypotheses lead to alternative predictions: (a) sperm quantity and quality traits co-evolve under sperm competition because they play complementary roles in determining ejaculate's competitive ability, or (b) energetic constraints force trade-offs between traits depending on their relevance in providing a competitive advantage. We examined relationships between sperm competition levels, sperm quantity, and traits that determine ejaculate quality, in a comparative study of 18 rodent species using phylogenetically controlled analyses. Total sperm numbers were positively correlated to proportions of normal sperm, acrosome integrity and motile sperm; the latter three were also significantly related among themselves, suggesting no trade-offs between traits. In addition, testes mass corrected for body mass (i.e., relative testes mass), showed a strong association with sperm numbers, and positive significant associations with all sperm traits that determine ejaculate quality with the exception of live sperm. An "overall sperm quality" parameter obtained by principal component analysis (which explained 85% of the variance) was more strongly associated with relative testes mass than any individual quality trait. Overall sperm quality was as strongly associated with relative testes mass as sperm numbers. Thus, sperm quality traits improve under sperm competition in an integrated manner suggesting that a combination of all traits is what makes ejaculates more competitive. In evolutionary terms this implies that a complex network of genetic and

Improved cryopreservability of stallion sperm using a sorbitol-based freezing extender.

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Pojprasath, T; Lohachit, C; Techakumphu, M; Stout, T; Tharasanit, T

2011-06-01

Cryopreservation of stallion semen is often associated with poor post-thaw sperm quality. Sugars are among the important components of a freezing extender and act as non-permeating cryoprotectants. This study aimed to compare the quality of stallion sperm frozen with glucose, fructose or sorbitol-containing freezing extenders. Semen was collected from six stallions of proven fertility and cryopreserved using a freezing extender containing different types of monosaccharide sugars (glucose, fructose or sorbitol). After thawing, the semen was examined for sperm motility, viability, acrosome integrity, plasma membrane functionality and sperm longevity. The fertility of semen frozen in the presence of sorbitol was also tested by artificial insemination. Sperm quality was significantly decreased following freezing and thawing (P sorbitol and glucose (P sorbitol-based extender when examined at 2 and 4 h post-thaw, all of these parameters plus plasma membrane functionality were improved for sperm frozen in the sorbitol extender than in the glucose extender when examined 10 min post-thaw. Two of four mares (50%) inseminated with semen frozen with a sorbitol-containing freezing extender became pregnant. It is concluded that different sugars have different abilities to protect against cryoinjury during freezing and thawing of stallion sperm. This study demonstrated that an extender containing sorbitol as primary sugar can be used to successfully cryopreserve equine sperm; moreover, the quality of frozen-thawed sperm appeared to be better than when glucose or fructose was the principle sugar in the freezing extender. Copyright © 2011 Elsevier Inc. All rights reserved.

The control of classical swine fever in wild boar

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Volker eMoennig

2015-11-01

Full Text Available Classical swine fever (CSF is a viral disease with severe economic consequences for domestic pigs. Natural hosts for the CSF virus (CSFV are members of the family Suidae, i.e. Eurasian wild boar (sus scrofa are also susceptible. CSF in wild boar poses a serious threat to domestic pigs. CSFV is an enveloped RNA virus belonging to the pestivirus genus of the Flaviviridae family. Transmission of the infection is usually by direct contact or by feeding of contaminated meat products. In recent decades CSF has been successfully eradicated from Australia, North America, and the European Union. In areas with dense wild boar populations CSF tends to become endemic whereas it is often self-limiting in small, less dense populations. In recent decades eradication strategies of CSF in wild boar have been improved considerably. The reduction of the number of susceptible animals to a threshold level where the basic reproductive number is R0<1 is the major goal of all control efforts. Depending on the epidemiological situation, hunting measures combined with strict hygiene may be effective in areas with a relatively low density of wild boar. Oral immunization was shown to be highly effective in endemic situations in areas with a high density of wild boar.

Cellular Biophysics During Freezing of Rat and Mouse Sperm Predicts Post-thaw Motility1

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Hagiwara, Mie; Choi, Jeung Hwan; Devireddy, Ramachandra V.; Roberts, Kenneth P.; Wolkers, Willem F.; Makhlouf, Antoine; Bischof, John C.

2009-01-01

Though cryopreservation of mouse sperm yields good survival and motility after thawing, cryopreservation of rat sperm remains a challenge. This study was designed to evaluate the biophysics (membrane permeability) of rat in comparison to mouse to better understand the cooling rate response that contributes to cryopreservation success or failure in these two sperm types. In order to extract subzero membrane hydraulic permeability in the presence of ice, a differential scanning calorimeter (DSC) method was used. By analyzing rat and mouse sperm frozen at 5°C/min and 20°C/min, heat release signatures characteristic of each sperm type were obtained and correlated to cellular dehydration. The dehydration response was then fit to a model of cellular water transport (dehydration) by adjusting cell-specific biophysical (membrane hydraulic permeability) parameters Lpg and ELp. A “combined fit” (to 5°C/min and 20°C/min data) for rat sperm in Biggers-Whitten-Whittingham media yielded Lpg = 0.007 μm min−1 atm−1 and ELp = 17.8 kcal/mol, and in egg yolk cryopreservation media yielded Lpg = 0.005 μm min−1 atm−1 and ELp = 14.3 kcal/mol. These parameters, especially the activation energy, were found to be lower than previously published parameters for mouse sperm. In addition, the biophysical responses in mouse and rat sperm were shown to depend on the constituents of the cryopreservation media, in particular egg yolk and glycerol. Using these parameters, optimal cooling rates for cryopreservation were predicted for each sperm based on a criteria of 5%–15% normalized cell water at −30°C during freezing in cryopreservation media. These predicted rates range from 53°C/min to 70°C/min and from 28°C/min to 36°C/min in rat and mouse, respectively. These predictions were validated by comparison to experimentally determined cryopreservation outcomes, in this case based on motility. Maximum motility was obtained with freezing rates between 50°C/min and 80�

Bovine binder-of-sperm protein BSP1 promotes protrusion and nanotube formation from liposomes

International Nuclear Information System (INIS)

Lafleur, Michel; Courtemanche, Lesley; Karlsson, Goeran; Edwards, Katarina; Schwartz, Jean-Louis; Manjunath, Puttaswamy

2010-01-01

Research highlights: → Binder-of-sperm protein 1 (BSP1) modifies the morphology of lipidic vesicles inducing bead necklace-like and thread-like structures. → In the presence of multilamellar liposomes, BSP1 leads to the formation of long nanotubes. → The insertion of BSP1 in the external lipid leaflet of membranes induces local changes in bilayer curvature. -- Abstract: Binder-of-sperm (BSP) proteins interact with sperm membranes and are proposed to extract selectively phosphatidylcholine and cholesterol from these. This change in lipid composition is a key step in sperm capacitation. The present work demonstrates that the interactions between the protein BSP1 and model membranes composed with phosphatidylcholine lead to drastic changes in the morphology of the lipidic self-assemblies. Using cryo-electron microscopy and fluorescence microscopy, we show that, in the presence of the protein, the lipid vesicles elongate, and form bead necklace-like structures that evolve toward small vesicles or thread-like structures. In the presence of multilamellar vesicles, where a large reservoir of lipid is available, the presence of BSP proteins lead to the formation of long nanotubes. Long spiral-like threads, associated with lipid/protein complexes, are also observed. The local curvature of lipid membranes induced by the BSP proteins may be involved in lipid domain formation and the extraction of some lipids during the sperm maturation process.

Antibody Prevalence to Influenza Type A in Wild Boar of Northern Ukraine.

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Kovalenko, Ganna; Molozhanova, Alona; Halka, Ihor; Nychyk, Serhiy

2017-12-01

A preliminary serological survey was carried out to assess the likelihood of influenza A (IA) infection in wild boar and begin to characterize the role of wild boar in the epidemiology of the IA virus (IAV). Sera collected from 120 wild boar that were hunted in 2014 were tested. To detect antibodies to IA, a blocking the enzyme-linked immunosorbent assay (ELISA) was used. Thirty boar were collected from each of four oblasts in the north central and northwestern regions of Ukraine. Antibodies against IAV were detected in 27 samples (22.5%; 95% confidence interval 16.0-30.8) and in at least some of the wild boar from all of the four oblasts. This preliminary survey of IA antibodies in wild boar populations of northern Ukraine indicates a substantial frequency of exposure to IAV throughout the region. Infection of wild boar populations could provide an alternative or additional route for spillover from wild populations to domestic animals and humans.

Molecular basis of sperm capacitation acrosome reaction and interaction with eggs

International Nuclear Information System (INIS)

Sheikhnejad, G.

1985-01-01

A phospholipase C (PLC) which can hydrolyze 14 C-phosphatidylcholine was purified from bull seminal plasma. This PLC has an optimum at pH 7.2 and its PI was about 5.0. The enzyme was inhibited by EDTA, Cd 2+ , Pb 2+ , Ni 2+ , Fe 2+ , and Zn 2+ . PLC consists of two subunits one 69,000 and the other 55,000 daltons. The purified PLC was examined for induction of capacitation and acrosome reaction of guinea pig spermatozoa. Sperm were examined for the acrosome reaction 10 min after addition of 3.4 mM Ca 2+ . Fifty percent of the sperm underwent the acrosome reaction while the control had less than 5% acrosome reacted sperm. The antiserum to the inneracrosomal membrane isolated from sperm was labeled with FITC conjugated goat anti-guinea pig IgG. The conjugated antibody was used to localize sperm antigens. The antigens located on the IAM were only fluoresced when rabbit sperm were treated with methanol and/or MgCl 2 . Therefore anti-IAM antibody did not bind to the sperm plasma membrane. In vivo capacitated rabbit sperm were incubated with anti-IAM antibody (intact IgG and F(ab') 2 fragments) for 30 min prior to addition of rabbit eggs. After 24 h the eggs were examined for cleavage. The control eggs were fertilized (90%) while the antibody completely inhibited the fertilization of ova in vitro. The eggs incubated with antibody prior to the addition of sperm were still fertilizable. Thus, anti-IAM did not have any noticeable effect on the eggs. It was also shown that antibody inhibited fertilization of zona-free rabbit eggs in vitro as well

Activation of free sperm and dissociation of sperm bundles (spermatozeugmata) of an endangered viviparous fish, Xenotoca eiseni.

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Liu, Yue; Yang, Huiping; Torres, Leticia; Tiersch, Terrence R

2018-04-01

Knowledge of sperm motility activation for viviparous fishes has been limited to study of several species in Poeciliidae, and the dissociation of sperm bundles is even less understood. The goal of this study was to use the endangered Redtail Splitfin (Xenotoca eiseni) as a model to investigate the activation of sperm from viviparous fishes by study of free sperm and spermatozeugmata (unencapsulated sperm bundles). The specific objectives were to evaluate the effects of: (1) osmotic pressure and refrigerated storage (4 °C) on activation of free sperm, (2) osmotic pressure, ions, and pH on dissociation of spermatozeugmata, and (3) CaCl 2 concentration and pH on sperm membrane integrity. Free sperm were activated in Ca 2+ -free Hanks' balanced salt solution at 81-516 mOsmol/kg. The highest motility (19 ± 6%) was at 305 mOsmol/kg and swim remained for 84 h. Glucose (300-700 mOsmol/kg), NaCl (50-600 mOsmol/kg), and KCl, MgCl 2 , and MnCl 2 at 5-160 mM activated sperm within spermatozeugmata, but did not dissociate spermatozeugmata. CaCl 2 at 5-160 mM dissociated spermatozeugmata within 10 min. Solutions of NaCl-NaOH at pH 11.6 to 12.4 dissociated spermatozeugmata within 1 min. The percentage of viable cells had no significant differences (P = 0.2033) among different concentrations of CaCl 2 , but it was lower (P fishes, and for development of germplasm repositories for imperiled goodeids. Copyright © 2018 Elsevier Inc. All rights reserved.

Optical tweezers and non-ratiometric fluorescent-dye-based studies of respiration in sperm mitochondria

International Nuclear Information System (INIS)

Chen, Timothy; Shi, Linda Z; Zhu, Qingyuan; Chandsawangbhuwana, Charlie; Berns, Michael W

2011-01-01

The purpose of this study is to investigate how the mitochondrial membrane potential affects sperm motility using laser tweezers and a non-ratiometric fluorescent probe, DiOC 6 (3). A 1064 nm Nd:YVO4 continuous wave laser was used to trap motile sperm at a power of 450 mW in the trap spot. Using customized tracking software, the curvilinear velocity (VCL) and the escape force from the laser tweezers were measured. Human (Homo sapiens), dog (Canis lupis familiaris) and drill (Mandrillus leucophaeus) sperm were treated with DiOC 6 (3) to measure the membrane potential in the mitochondria-rich sperm midpieces. Sperm from all three species exhibited an increase in fluorescence when treated with the DiOC 6 (3). When a cyanide inhibitor (CCCP) of aerobic respiration was applied, sperm of all three species exhibited a reduction in fluorescence to pre-dye levels. With respect to VCL and escape force, the CCCP had no effect on dog or human sperm, suggesting a major reliance upon anaerobic respiration (glycolysis) for ATP in these two species. Based on the preliminary study on drill sperm, CCCP caused a drop in the VCL, suggesting potential reliance on both glycolysis and aerobic respiration for motility. The results demonstrate that optical trapping in combination with DiOC 6 (3) is an effective way to study sperm motility and energetics

Optical tweezers and non-ratiometric fluorescent-dye-based studies of respiration in sperm mitochondria

Science.gov (United States)

Chen, Timothy; Shi, Linda Z.; Zhu, Qingyuan; Chandsawangbhuwana, Charlie; Berns, Michael W.

2011-04-01

The purpose of this study is to investigate how the mitochondrial membrane potential affects sperm motility using laser tweezers and a non-ratiometric fluorescent probe, DiOC6(3). A 1064 nm Nd:YVO4 continuous wave laser was used to trap motile sperm at a power of 450 mW in the trap spot. Using customized tracking software, the curvilinear velocity (VCL) and the escape force from the laser tweezers were measured. Human (Homo sapiens), dog (Canis lupis familiaris) and drill (Mandrillus leucophaeus) sperm were treated with DiOC6(3) to measure the membrane potential in the mitochondria-rich sperm midpieces. Sperm from all three species exhibited an increase in fluorescence when treated with the DiOC6(3). When a cyanide inhibitor (CCCP) of aerobic respiration was applied, sperm of all three species exhibited a reduction in fluorescence to pre-dye levels. With respect to VCL and escape force, the CCCP had no effect on dog or human sperm, suggesting a major reliance upon anaerobic respiration (glycolysis) for ATP in these two species. Based on the preliminary study on drill sperm, CCCP caused a drop in the VCL, suggesting potential reliance on both glycolysis and aerobic respiration for motility. The results demonstrate that optical trapping in combination with DiOC6(3) is an effective way to study sperm motility and energetics.

Implementing an open-access CASA software for the assessment of stallion sperm motility: Relationship with other sperm quality parameters.

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Giaretta, Elisa; Munerato, Mauro; Yeste, Marc; Galeati, Giovanna; Spinaci, Marcella; Tamanini, Carlo; Mari, Gaetano; Bucci, Diego

2017-01-01

Setting an open-access computer assisted sperm analysis (CASA) may benefit the evaluation of motility in mammalian sperm, especially when economic constraints do not allow the use of a commercial system. There have been successful attempts to develop such a device in Zebra fish sperm and the system has been used in very few studies on mammalian spermatozoa. Against this background, the present study aimed at developing an open-access CASA system for mammalian sperm using the horse as a model and based upon the Image J software previously established for Zebra fish sperm. Along with determining the sperm progressive motility and other kinetic parameters (such as amplitude of lateral head displacement), the "results" window was adjusted to simplify subsequent statistical analyses. The path window was enriched with colored sperm trajectories on the basis of the subpopulation they belong to and a number that allowed the sperm track to be associated to the sperm motility data shown in the "results" window. Data obtained from the novel plugin (named as CASA_bgm) were compared with those of the commercial CASA Hamilton-Thorn IVOS Vers.12, through Bland Altman's plots. While the percentage of total and progressive motile sperm, VCL, VAP, VSL, LIN and STR and ALH were in agreement with those obtained with the commercial system, BCF significantly differed between the two systems probably due to their settings. Interestingly, a positive and significant correlation between the percentages of total motile sperm evaluated through CASA_bgm and those showing high mitochondrial membrane potential evaluated by JC-1 staining was found. In conclusion, CASA_bgm ImageJ plugin could be useful and reliable for stallion sperm motility analysis and it is our aim to apply this system to other mammalian species. Copyright © 2016 Elsevier B.V. All rights reserved.

Ulceroglandular Tularemia Following Contact with a Boar.

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Passiouk, Nikola; Heininger, Ulrich

2016-04-01

We report on a 13-year-old patient who developed ulceroglandular tularemia after having assisted in slaughtering a hunted boar. He presented with a digital skin ulcer and enlarged lymph nodes. Clinically suspected tularemia was proven by real-time polymerase chain reaction performed on a skin ulcer biopsy and swab and by positive serology. This is the first reported case of tularemia after contact with a boar.

Effects of supplemental seminal plasma on cryopreserved boar ...

African Journals Online (AJOL)

To analyse the effects of supplemental autologous seminal plasma on boar semen quality before freezing and after thawing, thirty ejaculates were collected from six Pietrain boars. The main factors of a 2 x 2 x 2 factorial arrangement of treatments were Beltsville thawing solution (BTS), seminal plasma before freezing, and ...

Mitochondrial DNA control region variability in wild boars from west Balkans

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Đan Mihajla

2013-01-01

Full Text Available The wild boar (Sus scrofa is one of most abundant game species in hunting areas of Balkan region. The large fraction of pre-glacial genetic diversity in wild boar populations from the Balkans was addressed due to high proportion of unique mtDNA haplotypes found in Greece, indicating Balkan as main refugial area for wild boars. The aim of the present study is to characterize mitochondrial DNA control region variability in wild boars from different areas in the West Balkan region, in order to evaluate level of genetic variability, to detect unique haplotypes and to infer possible structuring. The total number of 163 individuals from different sampling localities were included in the study. A fragment of the mtDNA control region was amplified and sequenced by standard procedures. Population genetic analyses were performed using several computer packages: BioEdit, ARLEQUIN 3.5.1.2., Network 4.6.0.0 and MEGA5. Eleven different haplotypes were identified and haplotype diversity was 0.676, nucleotide diversity 0.0026, and the average number of nucleotide differences (k 1.169. The mismatch distribution and neutrality tests indicated the expansion of the all populations. It is shown that high level of genetic diversity is present in the wild boars from the West Balkan region and we have managed to detect regional unique haplotypes in high frequency. Genetic diversity differences have been found in regional wild boar groups, clustering them in two main clusters, but further speculations on the reasons for the observed clustering are prevented due to restricted informativness of the single locus marker. Obtained knowledge of genetic variation in the wild boar may be relevant for improving knowledge of the phylogeny and phylogeography of the wild boars, but as well as for hunting societies and responsible authorities for the effective control of wild boar populations.

Depolarization of sperm membrane potential is a common feature of men with subfertility and is associated with low fertilization rate at IVF.

Science.gov (United States)

Brown, Sean G; Publicover, Stephen J; Mansell, Steven A; Lishko, Polina V; Williams, Hannah L; Ramalingam, Mythili; Wilson, Stuart M; Barratt, Christopher L R; Sutton, Keith A; Da Silva, Sarah Martins

2016-06-01

Are significant abnormalities in outward (K(+)) conductance and resting membrane potential (Vm) present in the spermatozoa of patients undertaking IVF and ICSI and if so, what is their functional effect on fertilization success? Negligible outward conductance (≈5% of patients) or an enhanced inward conductance (≈4% of patients), both of which caused depolarization of Vm, were associated with a low rate of fertilization following IVF. Sperm-specific potassium channel knockout mice are infertile with defects in sperm function, suggesting that these channels are essential for fertility. These observations suggest that malfunction of K(+) channels in human spermatozoa might contribute significantly to the occurrence of subfertility in men. However, remarkably little is known of the nature of K(+) channels in human spermatozoa or the incidence and functional consequences of K(+) channel defects. Spermatozoa were obtained from healthy volunteer research donors and subfertile IVF and ICSI patients attending a hospital assisted reproductive techniques clinic between May 2013 and December 2015. In total, 40 IVF patients, 41 ICSI patients and 26 normozoospermic donors took part in the study. Samples were examined using electrophysiology (whole-cell patch clamping). Where abnormal electrophysiological characteristics were identified, spermatozoa were further examined for Ca(2+) influx induced by progesterone and penetration into viscous media if sufficient sample was available. Full exome sequencing was performed to specifically evaluate potassium calcium-activated channel subfamily M α 1 (KCNMA1), potassium calcium-activated channel subfamily U member 1 (KCNU1) and leucine-rich repeat containing 52 (LRRC52) genes and others associated with K(+) signalling. In IVF patients, comparison with fertilization rates was done to assess the functional significance of the electrophysiological abnormalities. Patch clamp electrophysiology was used to assess outward (K

Age effect on post freezing sperm viability of Bali cattle (Bos javanicus)

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Hapsari, R. D.; Khalifah, Y.; Widyas, N.; Pramono, A.; Prastowo, S.

2018-03-01

Post freezing sperm viability is one of factors which determine artificial insemination success. In the other side, bull’s or sire age influences the semen quality through sperm membrane constituent. It is known that freezing process change the sperm membrane during the processing stage. This research aims to know the effect of sire age on post freezing sperm viability of Bali cattle. The samples were collected in Singosari Artificial Insemination Centre, Malang, East Java, Indonesia on September - November 2016. Eight Bali cattle (4 and 7 y.o, 4 heads in each group) were used as semen source. Semen was collected using artificial vagina, 10 times spanning for 5 weeks (2 times per week, interval 3 and 4 days) in a row. The samples were then evaluated at fresh, chill and frozen stage. Fresh semen was diluted in Tris-citrate-egg yolk 20% (v/v) followed with chilling and freezing. Semen qualities were observed as sperm % motility (MOT), % live sperm using eosin-nigrosine staining (EOS) and % sperm membrane integrity using hypoosmotic swelling test (HOS). Variable comparisons between age groups were done using t-test. On the average, 4 y.o bulls showed higher semen quality at fresh, chill and frozen compared to 7 y.o in MOT (68.00±6.39 vs 65.9±7.62 56.40±3.71 vs 54.33±5.83 44.25±3.52 vs 40.40±7.06), EOS (72.08±6.63 vs 71.82±7.38 57.81±3.83 vs 57.41±6.32 53.16 ±8.41 vs 46.49±9.13) and HOS (60.85±13.91 vs 54.84±13.43 53.16 ±8.41 vs 46.49±9.13 44.6±9.39 vs 33.8±10.70) respectively. Statistical analysis results showed that age was significantly (PBali cattle (4 y.o) have more viable post freezing sperm compared to the older ones (7 y.o).

Hormonal and behavioral correlates of emotional states in sexually trained boars.

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Bishop, J D; Malven, P V; Singleton, W L; Weesner, G D

1999-12-01

Physiological and behavioral traits of sexually mature boars were compared between episodes of copulation and sexual frustration in order to determine reliable indicators of the differences in emotional states. Ten boars, approximately 6 mo of age, were trained to mount a stationary artificial sow (ArtSow) and to ejaculate when digital pressure was applied to the extended penis. This method of semen collection is the typical procedure of the industry. All 10 boars used in this study were fully trained to this procedure before the onset of the study. Each boar was subjected to trials in which one of the following two treatments was applied. In the control (CTRL) treatment, boars were treated the same as during their training (i.e., allowed to complete ejaculation). In the frustration (FRUS) treatment, boars were allowed to mount the ArtSow, but because no manual pressure was applied to the extended penis, ejaculation never occurred. Blood was collected via indwelling catheters before onset of the trial, during exposure to the ArtSow, and after returning to their home pen. Concentrations of testosterone, cortisol, and beta-endorphin were quantified. Behavior of the boars was recorded during exposure to the ArtSow and for 30 min after return to their home pen. Relative to preexposure levels, serum cortisol increased (Pcortisol did not allow us to distinguish between the excitement of copulation and the negative affect associated with sexual frustration, whereas increases in serum beta-endorphin and motor activity seemed to be indicators of the negative emotional state of sexual frustration in trained boars.

Rooster semen cryopreservation: effect of pedigree line and male age on postthaw sperm function.

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Long, J A; Bongalhardo, D C; Pelaéz, J; Saxena, S; Settar, P; O'Sullivan, N P; Fulton, J E

2010-05-01

The fertility rates of cryopreserved poultry semen are highly variable and not reliable for use in preservation of commercial genetic stocks. Our objective was to evaluate the cryosurvival of semen from 8 pedigreed layer lines at 2 different ages: the onset and end of commercial production. Semen from 160 roosters (20/line) was frozen individually with 11% glycerol at 6 and 12 mo of age. Glycerol was removed from thawed semen by Accudenz gradient centrifugation. The viability of thawed sperm from each male was determined using fluorescent live-dead staining and flow cytometry; sperm velocity parameters were measured using computerized motion analysis. The fertilizing ability of thawed sperm was evaluated in vitro by assessing hydrolysis of the inner perivitelline membrane. The postthaw function of sperm from the elite lines varied widely, despite the fact that fresh semen from all of these lines typically yielded high fertility rates. The percentage of thawed sperm with intact plasma membranes ranged from 27.8 + or - 2.1 to 49.6 + or - 1.9 and varied among lines and between age groups. Thawed sperm from 2 lines consistently demonstrated the highest and lowest motility parameters, whereas the velocity parameters of the remaining 6 lines varied widely. The mean number of hydrolysis points per square millimeter of inner perivitelline membrane ranged from 12.5 + or - 4.1 (line 2) to 103.3 + or - 30.2 (line 6). Age effects were observed for 4 out of 8 lines; however, improved postthaw sperm function at 12 mo of age was not consistent for all 3 assays. These results demonstrate variability among pedigreed lines in withstanding glycerol-based semen cryopreservation and provide a model for delineating genotypic and phenotypic factors affecting sperm cryosurvival.

Concentration-dependent Sildenafil citrate (Viagra) effects on ROS production, energy status, and human sperm function.

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Sousa, Maria Inês; Amaral, Sandra; Tavares, Renata Santos; Paiva, Carla; Ramalho-Santos, João

2014-04-01

Literature regarding the effects of sildenafil citrate on sperm function remains controversial. In the present study, we specifically wanted to determine if mitochondrial dysfunction, namely membrane potential, reactive oxygen species production, and changes in energy content, are involved in in vitro sildenafil-induced alterations of human sperm function. Sperm samples of healthy men were incubated in the presence of 0.03, 0.3, and 3 μM sildenafil citrate in a phosphate buffered saline (PBS)-based medium for 2, 3, 12, and 24 hours. Sperm motility and viability were evaluated and mitochondrial function, i.e., mitochondrial membrane potential and mitochondrial superoxide production were assessed using flow-cytometry. Additionally, adenosine triphosphate (ATP) levels were determined by high performance liquid chromatography (HPLC) analysis. Results show a decrease in sperm motility correlated with the level of mitochondria-generated superoxide, without a visible effect on mitochondrial membrane potential or viability upon exposure to sildenafil. The effect on both motility and superoxide production was higher for the intermediate concentration of sildenafil (0.3 µM) indicating that the in vitro effects of sildenafil on human sperm do not vary linearly with drug concentration. Adenosine triphosphate levels also decreased following sildenafil exposure, but this decrease was only detected after a decrease in motility was already evident. These results suggest that along with the level of ATP and mitochondrial function other factors are involved in the early sildenafil-mediated decline in sperm motility. However, the further decrease in ATP levels and increase in mitochondria-generated reactive oxygen species after 24 hours of exposure might further contribute towards declining sperm motility.

The effect of breeding facility and sexual stimulation on plasma cortisol in boars.

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Levis, D G; Barnett, J L; Hemsworth, P H; Jongman, E

1995-12-01

Nine boars were used to evaluate effects of breeding facility design and sexual activity on plasma cortisol concentrations . In one breeding facility (conventional), boars were housed individually in small pens, and female pigs were mated in those boar pens. In another breeding facility (Detection-Mating Area [DMA] system), boars were housed individually in stalls, and female pigs were mated in a specific mating pen adjacent to the front of stalls where boars were housed. After 51 d of housing treatment, a catheter was surgically implanted in the cephalic vein for collection of blood samples. Daytime profiles (hourly collections from 0900 to 1700) of cortisol did not differ among boars in the two treatment groups. Cortisol was greater (P cortisol in boars, but breeding facility did not affect the ACTH-induced changes in cortisol concentrations. There was a treatment x time interaction (P cortisol after sexual stimulation, and the magnitude and duration of increase in cortisol were greater (P Cortisol was greater (P cortisol concentration in boars seems to be a normal biological response to sexual activity. However, magnitude and duration of the increase in cortisol may be influenced by breeding facility design and mating procedure. There is no evidence, based on physiological data, that housing boars in stalls in the DMA system has any adverse effects on their welfare.

First TBEV serological screening in Flemish wild boar

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Sophie Roelandt

2016-04-01

Full Text Available In the frame of a Flemish wildlife surveillance in 2013, a serological screening was performed on sera from wild boar (Sus scrofa; n=238 in order to detect tick-borne encephalitis virus (TBEV-specific antibodies. Neutralising antibodies were titrated with a seroneutralisation test (SNT, using two cut-off titres (1/10–1/15. Seven wild boars were found TBEV-seropositive and showed moderate (>1/15 to high (>1/125 SNT-titres; three individuals had borderline results (1/10–1/15. This study demonstrated the presence of TBEV-specific antibodies in wild boar and highlighted potential TBEV-foci in Flanders. Additional surveillance including direct virus testing is now recommended.

Mitochondrial PKA mediates sperm motility.

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Mizrahi, Rashel; Breitbart, Haim

2014-12-01

Mitochondria are the major source of ATP to power sperm motility. Phosphorylation of mitochondrial proteins has been proposed as a major regulatory mechanism for mitochondrial bioenergetics. Sperm motility was measured by a computer-assisted analyzer, protein detection by western blotting, membrane potential by tetramethylrhodamine, cellular ATP by luciferase assay and localization of PKA by immuno-electron microscopy. Bicarbonate is essential for the creation of mitochondrial electro-chemical gradient, ATP synthesis and sperm motility. Bicarbonate stimulates PKA-dependent phosphorylation of two 60kDa proteins identified as Tektin and glucose-6-phosphate isomerase. This phosphorylation was inhibited by respiration inhibition and phosphorylation could be restored by glucose in the presence of bicarbonate. However, this effect of glucose cannot be seen when the mitochondrial ATP/ADP exchanger was inhibited indicating that glycolytic-produced ATP is transported into the mitochondria and allows PKA-dependent protein phosphorylation inside the mitochondria. Bicarbonate activates mitochondrial soluble adenylyl cyclase (sAC) which catalyzes cAMP production leading to the activation of mitochondrial PKA. Glucose can overcome the lack of ATP in the absence of bicarbonate but it cannot affect the mitochondrial sAC/PKA system, therefore the PKA-dependent phosphorylation of the 60kDa proteins does not occur in the absence of bicarbonate. Production of CO2 in Krebs cycle, which is converted to bicarbonate is essential for sAC/PKA activation leading to mitochondrial membrane potential creation and ATP synthesis. Copyright © 2014 Elsevier B.V. All rights reserved.

Sperm from sneaker male squids exhibit chemotactic swarming to CO₂.

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Hirohashi, Noritaka; Alvarez, Luis; Shiba, Kogiku; Fujiwara, Eiji; Iwata, Yoko; Mohri, Tatsuma; Inaba, Kazuo; Chiba, Kazuyoshi; Ochi, Hiroe; Supuran, Claudiu T; Kotzur, Nico; Kakiuchi, Yasutaka; Kaupp, U Benjamin; Baba, Shoji A

2013-05-06

Behavioral traits of sperm are adapted to the reproductive strategy that each species employs. In polyandrous species, spermatozoa often form motile clusters, which might be advantageous for competing with sperm from other males. Despite this presumed advantage for reproductive success, little is known about how sperm form such functional assemblies. Previously, we reported that males of the coastal squid Loligo bleekeri produce two morphologically different euspermatozoa that are linked to distinctly different mating behaviors. Consort and sneaker males use two distinct insemination sites, one inside and one outside the female's body, respectively. Here, we show that sperm release a self-attracting molecule that causes only sneaker sperm to swarm. We identified CO2 as the sperm chemoattractant and membrane-bound flagellar carbonic anhydrase as its sensor. Downstream signaling results from the generation of extracellular H(+), intracellular acidosis, and recovery from acidosis. These signaling events elicit Ca(2+)-dependent turning behavior, resulting in chemotactic swarming. These results illuminate the bifurcating evolution of sperm underlying the distinct fertilization strategies of this species. Copyright © 2013 Elsevier Ltd. All rights reserved.

First report of Mycoplasma hyopneumoniae seroprevalence in farmed wild boars in China.

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Liang, Qin-Li; Zou, Yang; Gao, Yun-Hang; Nie, Lan-Bi; Zhang, Xiao-Xuan; Hu, Gui-Xue; Du, Rui; Zhu, Xing-Quan

2018-06-01

Porcine enzootic pneumonia caused by Mycoplasma hyopneumoniae affects the global pig industry with significant economic losses. It is yet to know whether wild boars in China were infected with M. hyopneumoniae. The present study was conducted to examine the seroprevalence and to evaluate risk factors of M. hyopneumoniae infection in farmed wild boars in China. A total of 882 serum samples were collected from farmed wild boars in Jilin City, Siping City and Baishan City in Jilin Province, northeastern China from April 2015 to February 2016, and were examined by the double sandwich enzyme-linked immunosorbent assay (ELISA). Seventy-eight out of 882 (8.8%) serum samples were M. hyopneumoniae-seropositive. Among region groups, wild boars from Jilin city (11.7%, 33/281) had the highest seropositivity, followed by Siping city (11%, 29/263) and Baishan city (4.7%, 16/338), and the difference was statistically significant (P = 0.0031). The M. hyopneumoniae seroprevalence in the female wild boars (9.0%, 75/831) was higher than that in the male wild boars (5.9%, 3/51) (P = 0.4429). The results of this investigation showed that farmed wild boars were susceptible to M. hyopneumoniae. Logistic regression analysis showed that there is a significant correlation between the geographical area and M. hyopneumoniae infection, which may be related to the regional environment. This is the first report of M. hyopneumoniae seroprevalence in farmed wild boars in China, which provided baseline information for further studies and control of M. hyopneumoniae infection in wild boars in China. Copyright © 2018 Elsevier B.V. All rights reserved.

Effect of sperm concentration on characteristics and fertilization capacity of rooster sperm frozen in the presence of the antioxidants catalase and vitamin E.

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Moghbeli, Morteza; Kohram, Hamid; Zare-Shahaneh, Ahmad; Zhandi, Mahdi; Sharideh, Hossein; Sharafi, Mohsen

2016-10-01

The objective of this study conducted was to determine the influence of different levels of sperm concentration, including catalase (CAT) and vitamin E (VitE) in rooster semen extender on postthawed quality and fertility of rooster semen. Semen was collected twice a week from six roosters (Arian) and diluted according to experimental treatments consisting of sperm suspensions containing different sperm concentrations (200, 400, and 600 × 106 sperm/mL) without antioxidant supplementation as control (Con) groups (Con200, Con400, and Con600, respectively), sperm suspensions containing different sperm concentrations (200, 400, and 600 × 106 sperm/mL) supplemented with 5-μg/mL VitE (VitE200, VitE400, and VitE600, respectively) and different sperm concentrations (200, 400, and 600 × 106 sperm/mL) supplementation with 100 IU/mL CAT (CAT200, CAT400, and CAT600, respectively). After thawing; sperm motility, membrane integrity, and mitochondrial function were assessed. Fertility and hatchability rates were determined by using 100 artificially inseminated hens. The percentage of total motility (TM) and activity of mitochondria decreased (P rooster extender with different level sperm concentrations had no effect (P > 0.05) on fertility and hatchability rates. In conclusion, although adding VitE and CAT in extender with different levels of sperm concentration improved postthawed quality of rooster semen, but adding VitE and CAT in the extender have no effect on fertility rate. Copyright © 2016 Elsevier Inc. All rights reserved.

Infection with Paragonimus westermani of boar-hunting dogs in Western Japan maintained via artificial feeding with wild boar meat by hunters.

Science.gov (United States)

Irie, Takao; Yamaguchi, Yohei; Doanh, Pham Ngoc; Guo, Zhi Hong; Habe, Shigehisa; Horii, Yoichiro; Nonaka, Nariaki

2017-08-18

Infection of boar-hunting dogs with Paragonimus westermani was investigated in Western Japan. Blood and rectal feces were collected from 441 dogs in the three districts (205 in Kinki, 131 in Chugoku and 105 in Shikoku District). In a screening ELISA for serum antibody against P. westermani antigen, 195 dogs (44.2%) showed positive reaction. In the 195 dogs, 8 dogs were found excreting P. westermani eggs after molecular analysis of fecal eggs, and additional 7 were identified serologically for the parasite infection because of their stronger reactivity against P. westermani antigen than against antigens of other species of Paragonimus. A spatial analysis showed that all of the P. westermani infections were found in Kinki and Chugoku Districts. In this area, dogs' experience of being fed with raw boar meat showed high odds ratio (3.35) to the sero-positivity in the screening ELISA, and the frequency of such experiences was significantly higher in sero-positive dogs. While clear relationship was not obtained between predation of boars by dogs during hunting and their sero-positivity. Therefore, it is suggested that human activity of feeding with wild boar meat is the risk factor for P. westermani infection in boar-hunting dogs. Considering that hunting dogs could play as a major definitive host and maintain the present distribution of P. westermani in Western Japan, control measures for the infection in hunting dogs, such as prohibition of raw meat feeding and regular deworming, should be undertaken.

NÍVEIS DE TESTOSTERONA NA SALIVA E NO PLASMA SEMINAL DE REPRODUTORES SUÍNOS

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Thais Schwarz Gaggini

2016-01-01

Full Text Available The aims of this study were evaluated testosterone levels in saliva and seminal plasma and correlate these informations with libido and sperm production of two lines of boars. The hormonal analysis was done using ELISA (Enzyme Linked Immuno Sorbent Assay and test F and Sperman correlation of SAS program was used for statistical analysis. There was no difference (P>0.05 between boar lines and testosterone levels in saliva and seminal plasma, collection length, semen volume and concentration and sperm motility and viability. Boars used in this study had libido, semen production and sperm cells considered normal and there was no difference between the lines. The results obtained can be classified as normal parameters expected in this situation.

Etiologies of sperm oxidative stress

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Parvin Sabeti

2016-04-01

Full Text Available Sperm is particularly susceptible to reactive oxygen species (ROS during critical phases of spermiogenesis. However, the level of seminal ROS is restricted by seminal antioxidants which have beneficial effects on sperm parameters and developmental potentials. Mitochondria and sperm plasma membrane are two major sites of ROS generation in sperm cells. Besides, leukocytes including polymer phonuclear (PMN leukocytes and macrophages produce broad category of molecules including oxygen free radicals, non-radical species and reactive nitrogen species. Physiological role of ROS increase the intracellular cAMP which then activate protein kinase in male reproductive system. This indicates that spermatozoa need small amounts of ROS to acquire the ability of nuclear maturation regulation and condensation to fertilize the oocyte. There is a long list of intrinsic and extrinsic factors which can induce oxidative stress to interact with lipids, proteins and DNA molecules. As a result, we have lipid peroxidation, DNA fragmentation, axonemal damage, denaturation of the enzymes, over generation of superoxide in the mitochondria, lower antioxidant activity and finally abnormal spermatogenesis. If oxidative stress is considered as one of the main cause of DNA damage in the germ cells, then there should be good reason for antioxidant therapy in these conditions

Seasonal and cryopreservation impacts on semen quality in boars

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Seasonal boar infertility occurs worldwide and contributes to economic loss to the pork industry. The current study evaluated cooled vs cryopreserved semen quality of 11 Duroc boars collected in June (cool season) and August 2014 (warm season). Semen was cooled to 16°C (cooled) or frozen over liquid...

Zoonotic tick-borne bacteria among wild boars (Sus scrofa in Central Italy

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Valentina Virginia Ebani

2017-03-01

Full Text Available The objective of this investigation was to estimate the occurrence of infections by the three zoonotic bacteria Anaplasma phagocytophilum (A. phagocytophilum, Borrelia burgdorferi sensu lato (B. burgdorferi s.l. and Coxiella burnetii in wild boars (Sus scrofa in Central Italy. The spleen samples from 100 hunted wild boars were submitted to DNA extraction and PCR assays were carried out to detect the three agents. One (1% animal was positive for A. phagocytophilum, and three (3% for B. burgdorferi s.l. No positive reactions were observed for Coxiella burnetii. Wild boars did not seem to play an important role in the epidemiology of the three investigated agents. However, the detection of A. phagocytophilum and B. burgdorferi s.l. in the spleen of the tested animals showed that wild boars can harbor these pathogens, thus ticked that feeding on infected wild boars are likely to become infected, too, which represents a source of infection for other animals and humans. This is the first detection of A. phagocytophilum in wild boars in Italy.

Fat tissue is not a reservoir for radiocesium in wild boars.

Science.gov (United States)

Steinhauser, Georg; Knecht, Christian; Sipos, Wolfgang

2017-01-01

Meat of wild boars is not only known for high 137 Cs activity concentrations but also for the remarkable constancy of these levels. Even decades after the Chernobyl accident, the 137 Cs levels in wild boar meat in Central Europe have not declined but even partly increased. In the present study, we investigated an unusual hypothesis for this very unusual phenomenon: may the boars' fat tissue act as a reservoir for radiocesium? We investigated fat and muscle tissues of four wild boars in Western Germany and found that the 137 Cs concentrations in fat were in the range of 10-30% of the respective activities in muscle tissue. Hence, the hypothesis was refuted.

Challenges in Development of Sperm Repositories for Biomedical Fishes: Quality Control in Small-Bodied Species.

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Torres, Leticia; Liu, Yue; Guitreau, Amy; Yang, Huiping; Tiersch, Terrence R

2017-12-01

Quality control (QC) is essential for reproducible and efficient functioning of germplasm repositories. However, many biomedical fish models present significant QC challenges due to small body sizes (<5 cm) and miniscule sperm volumes (<5 μL). Using minimal volumes of sperm, we used Zebrafish to evaluate common QC endpoints as surrogates for fertilization success along sequential steps of cryopreservation. First, concentrations of calibration bead suspensions were evaluated with a Makler ® counting chamber by using different sample volumes and mixing methods. For sperm analysis, samples were initially diluted at a 1:30 ratio with Hanks' balanced salt solution (HBSS). Motility was evaluated by using different ratios of sperm and activation medium, and membrane integrity was analyzed with flow cytometry at different concentrations. Concentration and sperm motility could be confidently estimated by using volumes as small as 1 μL, whereas membrane integrity required a minimum of 2 μL (at 1 × 10 6 cells/mL). Thus, <5 μL of sperm suspension (after dilution to 30-150 μL with HBSS) was required to evaluate sperm quality by using three endpoints. Sperm quality assessment using a combination of complementary endpoints enhances QC efforts during cryopreservation, increasing reliability and reproducibility, and reducing waste of time and resources.

Detection of serum anti-sperm antibody in infertile couples with dot-immunogold filtration assay (DIGFA)

International Nuclear Information System (INIS)

Xie Xiaoxian

2005-01-01

Objective: To develop a new method for rapid detection of serum anti-sperm antibody in infertile couples. Methods: Human sperm antigen was prepared from pooled semen specimens of fertile males. Nitro-cellulose membrane was used as solid-phase carrier of the antigen. Colloidal gold pellet combined goat anti-human IgG was taken as labelled antibody. A dot-immunogold filtration assay system was established for test of serum anti-human sperm antibody. Serum specimens from 137 infertile couples were tested and the result compared with flat from ELISA. Results: The human sperm antigen would react with the anti-sperm antibody in the tested serum over the cellulose membrane through filtration and the result could be read with naked eye within 6 minutes. In this study of 137 infertile coupled, the anti-sperm antibody was positive in 21.9% of the female serum specimens and 13.19% of the males. Compared with the result from ELISA, the consistency rate was 96.1%. The sensitivity of the assay was 90.2% and specificity was 95.4%. The p reparation was stable after 6 months refrigerator storage. Conclusion: This newly developed DIGFA is very adequate for rap id detection of anti-sperm antibody and deserves popularization. (authors)

Egg cell-secreted EC1 triggers sperm cell activation during double fertilization.

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Sprunck, Stefanie; Rademacher, Svenja; Vogler, Frank; Gheyselinck, Jacqueline; Grossniklaus, Ueli; Dresselhaus, Thomas

2012-11-23

Double fertilization is the defining characteristic of flowering plants. However, the molecular mechanisms regulating the fusion of one sperm with the egg and the second sperm with the central cell are largely unknown. We show that gamete interactions in Arabidopsis depend on small cysteine-rich EC1 (EGG CELL 1) proteins accumulating in storage vesicles of the egg cell. Upon sperm arrival, EC1-containing vesicles are exocytosed. The sperm endomembrane system responds to exogenously applied EC1 peptides by redistributing the potential gamete fusogen HAP2/GCS1 (HAPLESS 2/GENERATIVE CELL SPECIFIC 1) to the cell surface. Furthermore, fertilization studies with ec1 quintuple mutants show that successful male-female gamete interactions are necessary to prevent multiple-sperm cell delivery. Our findings provide evidence that mutual gamete activation, regulated exocytosis, and sperm plasma membrane modifications govern flowering plant gamete interactions.

Role of WNT signaling in epididymal sperm maturation.

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Cheng, Jin-Mei; Tang, Ji-Xin; Li, Jian; Wang, Yu-Qian; Wang, Xiu-Xia; Zhang, Yan; Chen, Su-Ren; Liu, Yi-Xun

2018-02-01

Spermatozoa maturation, a process required for spermatozoa to acquire progressive motility and the ability to fertilize ova, primarily occurs in the caput and corpus of the epididymis. Despite considerable efforts, the factor(s) promoting epididymal sperm maturation remains unclear. Recently, WNT signaling has been implicated in epididymal sperm maturation. To further investigate WNT signaling function in epididymal sperm maturation, we generated Wntless conditional knockout mice (Wls cKO), Wls flox/flox ; Lcn5-Cre. In these mice, WNTLESS (WLS), a conserved membrane protein required for all WNT protein secretion, was specifically disrupted in the principal cells of the caput epididymidis. Immunoblot analysis showed that WLS was significantly reduced in the caput epididymidis of Wls cKO mice. In the caput epididymidis of Wls cKO mice, WNT 10A and WNT 2b, which are typically secreted by the principal cells of the caput epididymis, were not secreted. Interestingly, sperm motility analysis showed that the WLS deficiency in the caput epididymidis had no effect on sperm motility. Moreover, fertility tests showed that Wls cKO male mice had normal fertility. These results indicate that the disruption of WLS in principal cells of the caput epididymidis inhibits WNT protein secretion but has no effect on sperm motility and male fertility, suggesting that WNT signaling in the caput epididymidis may be dispensable for epididymal sperm maturation in mice.

Linear model analysis of the influencing factors of boar longevity in Southern China.

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Wang, Chao; Li, Jia-Lian; Wei, Hong-Kui; Zhou, Yuan-Fei; Jiang, Si-Wen; Peng, Jian

2017-04-15

This study aimed to investigate the factors influencing the boar herd life month (BHLM) in Southern China. A total of 1630 records of culling boars from nine artificial insemination centers were collected from January 2013 to May 2016. A logistic regression model and two linear models were used to analyze the effects of breed, housing type, age at herd entry, and seed stock herd on boar removal reason and BHLM, respectively. Boar breed and the age at herd entry had significant effects on the removal reasons (P linear models (with or without removal reason including) showed boars raised individually in stalls exhibited shorter BHLM than those raised in pens (P introduction. Copyright © 2017. Published by Elsevier Inc.

HORMONAL PROFILE AND NONSPECIFIC RESISTANCE IN BOAR UNDER PRE-SLAUGHTER STRESS

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S.S. Grabovskyi

2016-03-01

Full Text Available The article is devoted to determination of hormonal profile and nonspecific resistance in boars blood before slaughter after using of biologically active substances — animal origin antistressors andimmunostimulators. The purpose of research — determination of changes of insulin, adrenocorticotropic hormone (ACTH, cortisol content in boars blood before slaughter and their correction of natural origin biologically active substances of spleen extract. Object and research methods. The spleen extract has been additionally entered to the boars feed at 5 days before slaughter as an antistressors and immunostimulators in pre-slaughter period. The experiment was conducted on 15 boars with standard diet. Three groups of boars six months of age (5 boars each were formed for research. The pig’s spleen extract was obtained with ultrasound application (Iresearch group were using as a biologically active substances to the feed boars in pre-slaughter period. The extracts were applied to dry feed by aerosol method (70 % alcohol solution of spleen extract volume of 1.4 ml per kg body weight. The boars of II research group in the same way received to the feed of 70 % ethanol solution in the same volume. The boars of control group received only dry feed economy. Theboars slaughter was held on day 13 hours a.m. Mathematical treatment of the research results worked statistically using the software package Statistica 6.0. Results and discussion. The ACTH and cortisol level in the boars’ blood plasma of experimental and control groups significantly increased after transportation (before the slaughter compared with the indexes before transportation to meat plant. The ACTH concentration in the boars’ blood plasma of І experimental group, which was added to the basic diet spleen extract, was 10 % lower than in the control group boars compared with indicators before and after transportation

Spatiotemporal trends in Canadian domestic wild boar production and habitat predict wild pig distribution

DEFF Research Database (Denmark)

Michel, Nicole; Laforge, Michel; van Beest, Floris

2017-01-01

eradication of wild pigs is rarely feasible after establishment over large areas, effective management will depend on strengthening regulations and enforcement of containment practices for Canadian domestic wild boar farms. Initiation of coordinated provincial and federal efforts to implement population...... wild boar and test the propagule pressure hypothesis to improve predictive ability of an existing habitat-based model of wild pigs. We reviewed spatiotemporal patterns in domestic wild boar production across ten Canadian provinces during 1991–2011 and evaluated the ability of wild boar farm...... distribution to improve predictive models of wild pig occurrence using a resource selection probability function for wild pigs in Saskatchewan. Domestic wild boar production in Canada increased from 1991 to 2001 followed by sharp declines in all provinces. The distribution of domestic wild boar farms in 2006...

Fat tissue is not a reservoir for radiocesium in wild boars

International Nuclear Information System (INIS)

Steinhauser, Georg; Knecht, Christian; Sipos, Wolfgang

2017-01-01

Meat of wild boars is not only known for high "1"3"7Cs activity concentrations but also for the remarkable constancy of these levels. Even decades after the Chernobyl accident, the "1"3"7Cs levels in wild boar meat in Central Europe have not declined but even partly increased. In the present study, we investigated an unusual hypothesis for this very unusual phenomenon: may the boars' fat tissue act as a reservoir for radiocesium? We investigated fat and muscle tissues of four wild boars in Western Germany and found that the "1"3"7Cs concentrations in fat were in the range of 10-30% of the respective activities in muscle tissue. Hence, the hypothesis was refuted. (author)

The effect of two packaging systems on the post-thaw characteristics of canine sperm.

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Strzeżek, R; Polakiewicz, P; Kordan, W

2015-01-01

The aim of this study was to compare the effect of different packaging systems on some parameters of cryopreserved canine spermatozoa. The experimental material consisted of the sperm-rich fractions of ejaculates collected from four Beagle dogs. Semen samples for cryopreservation were stored in 0.25 ml plastic straws and two aluminum tubes with a total volume of 5.0 ml. Semen was frozen in static nitrogen vapor for 10 minutes (0.25 ml straws) or 15 and 20 minutes (aluminum tubes). Post-thaw assessments involved the determination of sperm motility parameters using a computer assisted sperm analyzer (CASA), sperm plasma membrane integrity (SPMI), mitochondrial membrane potential (MMP) and acrosome integrity (normal apical ridge, NAR). Regardless of the packaging system applied, no significant differences in total sperm motility (TMOT) or selected kinematic parameters were observed after freezing-thawing. However, spermatozoa frozen in 0.25 mL straws were characterized by improved functionality, in particular mitochondrial function, after thawing. The results indicate that large quantities of canine semen can be frozen in aluminum tubes. Further studies are required, however, to evaluate different freezing and thawing rates of aluminum tubes.

Sperm motility in fishes. (II) Effects of ions and osmolality: a review.

Science.gov (United States)

Alavi, Sayyed Mohammad Hadi; Cosson, Jacky

2006-01-01

The spermatozoa of most fish species are immotile in the testis and seminal plasma. Therefore, motility is induced after the spermatozoa are released into the aqueous environment during natural reproduction or into the diluent during artificial reproduction. There are clear relationships between seminal plasma composition and osmolality and the duration of fish sperm motility. Various parameters such as ion concentrations (K+, Na+, and Ca2+), osmotic pressure, pH, temperature and dilution rate affect motility. In the present paper, we review the roles of these ions on sperm motility in Salmonidae, Cyprinidae, Acipenseridae and marine fishes, and their relationship with seminal plasma composition. Results in the literature show that: 1. K+ is a key ion controlling sperm motility in Salmonidae and Acipenseridae in combination with osmotic pressure; this control is more simple in other fish species: sperm motility is prevented when the osmotic pressure is high (Cyprinidae) or low (marine fishes) compared to that of the seminal fluid. 2. Cations (mostly divalent, such as Ca2+) are antagonistic with the inhibitory effect of K+ on sperm motility. 3. In many species, Ca2+ influx and K+ or Na+ efflux through specific ionic channels change the membrane potential and eventually lead to an increase in cAMP concentration in the cell, which constitutes the initiation signal for sperm motility in Salmonidae. 4. Media that are hyper- and hypo-osmotic relative to seminal fluid trigger sperm motility in marine and freshwater fishes, respectively. 5. The motility of fish spermatozoa is controlled through their sensitivity to osmolality and ion concentrations. This phenomenon is related to ionic channel activities in the membrane and governs the motility mechanisms of axonemes.

Role of C-type natriuretic peptide in the function of normal human sperm

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Hui Xia

2016-01-01

Full Text Available C-type natriuretic peptide (CNP is a newly discovered type of local regulatory factor that mediates its biological effects through the specific, membrane-bound natriuretic peptide receptor-B (NPR-B. Recent studies have established that CNP is closely related to male reproductive function. The aims of this study were to determine the distribution of CNP/NPR-B in human ejaculated spermatozoa through different methods (such as immunolocalization, real time polymerase chain reaction and Western Blot, and then to evaluate the influence of CNP on sperm function i n vitro, such as motility and acrosome reaction. Human semen samples were collected from consenting donors who met the criteria of the World Health Organization for normozoospermia. Our results show that the specific receptor NPR-B of CNP is localized in the acrosomal region of the head and the membrane of the front-end tail of the sperm, and there is no signal of CNP in human sperm. Compared with the control, CNP can induce a significant dose-dependent increase in spermatozoa motility and acrosome reaction. In summary, CNP/NPR-B can affect sperm motility and acrosome reaction, thus regulating the reproductive function of males. CNP may be a new key factor in regulating sperm function.

Effect of calcium, bicarbonate, and albumin on capacitation-related events in equine sperm.

Science.gov (United States)

Macías-García, B; González-Fernández, L; Loux, S C; Rocha, A M; Guimarães, T; Peña, F J; Varner, D D; Hinrichs, K

2015-01-01

Repeatable methods for IVF have not been established in the horse, reflecting the failure of standard capacitating media to induce changes required for fertilization capacity in equine sperm. One important step in capacitation is membrane cholesterol efflux, which in other species is triggered by cholesterol oxidation and is typically enhanced using albumin as a sterol acceptor. We incubated equine sperm in the presence of calcium, BSA, and bicarbonate, alone or in combination. Bicarbonate induced an increase in reactive oxygen species (ROS) that was abolished by the addition of calcium or BSA. Bicarbonate induced protein tyrosine phosphorylation (PY), even in the presence of calcium or BSA. Incubation at high pH enhanced PY but did not increase ROS production. Notably, no combination of these factors was associated with significant cholesterol efflux, as assessed by fluorescent quantitative cholesterol assay and confirmed by filipin staining. By contrast, sperm treated with methyl-β-cyclodextrin showed a significant reduction in cholesterol levels, but no significant increase in PY or ROS. Presence of BSA increased sperm binding to bovine zonae pellucidae in all three stallions. These results show that presence of serum albumin is not associated with a reduction in membrane cholesterol levels in equine sperm, highlighting the failure of equine sperm to exhibit core capacitation-related changes in a standard capacitating medium. These data indicate an atypical relationship among cholesterol efflux, ROS production, and PY in equine sperm. Our findings may help to elucidate factors affecting failure of equine IVF under standard conditions. © 2015 Society for Reproduction and Fertility.

Assessing the Risk of African Swine Fever Introduction into the European Union by Wild Boar.

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De la Torre, A; Bosch, J; Iglesias, I; Muñoz, M J; Mur, L; Martínez-López, B; Martínez, M; Sánchez-Vizcaíno, J M

2015-06-01

The presence of African swine fever (ASF) in the Caucasus region and Russian Federation has increased concerns that wild boars may introduce the ASF virus into the European Union (EU). This study describes a semi-quantitative approach for evaluating the risk of ASF introduction into the EU by wild boar movements based on the following risk estimators: the susceptible population of (1) wild boars and (2) domestic pigs in the country of origin; the outbreak density in (3) wild boars and (4) domestic pigs in the countries of origin, the (5) suitable habitat for wild boars along the EU border; and the distance between the EU border and the nearest ASF outbreak in (6) wild boars or (7) domestic pigs. Sensitivity analysis was performed to identify the most influential risk estimators. The highest risk was found to be concentrated in Finland, Romania, Latvia and Poland, and wild boar habitat and outbreak density were the two most important risk estimators. Animal health authorities in at-risk countries should be aware of these risk estimators and should communicate closely with wild boar hunters and pig farmers to rapidly detect and control ASF. © 2013 Blackwell Verlag GmbH.

Enhancement of mouse sperm motility by trophinin-binding peptide

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Park Seong

2012-11-01

Full Text Available Abstract Background Trophinin is an intrinsic membrane protein that forms a complex in the cytoplasm with bystin and tastin, linking it microtubule-associated motor dynein (ATPase in some cell types. Previously, we found that human sperm tails contain trophinin, bystin and tastin proteins, and that trophinin-binding GWRQ (glycine, tryptophan, arginine, glutamine peptide enhanced motility of human sperm. Methods Immunohistochemistry was employed to determine trophinin protein in mouse spermatozoa from wild type mouse, by using spermatozoa from trophinin null mutant mice as a negative control. Multivalent 8-branched GWRQ (glycine, tryptophan, arginine, glutamine peptide or GWRQ-MAPS, was chemically synthesized, purified by HPLC and its structure was confirmed by MALDI-TOF mass spectrometry. Effect of GWRQ-MAPS on mouse spermatozoa from wild type and trophinin null mutant was assessed by a computer-assisted semen analyzer (CASA. Results Anti-trophinin antibody stained the principal (central piece of the tail of wild type mouse sperm, whereas the antibody showed no staining on trophinin null sperm. Phage particles displaying GWRQ bound to the principal piece of sperm tail from wild type but not trophinin null mice. GWRQ-MAPS enhanced motility of spermatozoa from wild type but not trophinin null mice. CASA showed that GWRQ-MAPS enhanced both progressive motility and rapid motility in wild type mouse sperm. Conclusions Present study established the expression of trophinin in the mouse sperm tail and trophinin-dependent effect of GWRQ-MAPS on sperm motility. GWRQ causes a significant increase in sperm motility.

Sperm Production Rate, Gonadal and Extragonadal Sperm ...

African Journals Online (AJOL)

Five healthy West African Dwarf (WAD) rams, 1.5 to 2.5 years of age and weighing between 15 kg to 20 kg were used to determine daily sperm production, gonadal and exragonadal sperm reserves. Gonadal and extragonadal sperm reserves were estimated by the haemocytometric method, while the daily sperm production ...

How does supplementary feeding affect endoparasite infection in wild boar?

DEFF Research Database (Denmark)

Oja, Ragne; Velstrom, Kaisa; Moks, Epp

2017-01-01

was associated with both wild boar and feeding site density, whereas the presence of Eimeria sp. oocysts in faecal samples was only associated with wild boar density. Helminth eggs were found more often from the soil of active and abandoned feeding sites than from control areas. This could reflect parasitic...

Functional integrity of Colossoma macropomum (Cuvier, 1816 sperm cryopreserved with enriched extender solutions

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Raycon Roberto Freitas Garcia

Full Text Available Cryoprotectant solutions are used to protect the sperm from alterations caused by the low temperature in the cryopreservation process. We evaluated the quality of Colossoma macropomum semen after freezing, using dimethyl sulfoxide (DMSO as a cryoprotectant, combined with two extender solutions (T1 - Solution 1: Glucose 90.0 g/L, Sodium Citrate 6.0 g/L, EDTA 1.5 g/L, Sodium Bicarbonate 1.5 g/L, Potassium Chloride 0.8 g/L, Gentamycin Sulphate 0.2 g/L, and T2 - Solution 2: Glucose 90.0 g/L, ACP(r-104 10.0 g/L. Motility rate and motility time did not differ between T1 and T2 and were lower than fresh semen. The number of normal sperm was significantly different in treatments T1 (15.1% and T2 (21.9%, and both showed a reduction in the percentage of normal sperm compared to fresh semen (57.4%. The values found for the rates of fertilization and hatching, mitochondrial functionality and sperm DNA, did not differ between the treatments (T1 and T2. Regarding membrane integrity, there was a higher percentage of spermatozoa with intact membranes in T1 (53.4% than T2 (43.7%. The extender solutions, combined with 10% DMSO, maintained the sperm DNA intact in almost all the C. macropomum sperm cells, however there was a loss in their functionality.

Avaliação da suplementação de vitamina A nas características seminais em reprodutores suínos Assessment of vitamin A supplementation on boar semen characteristics

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Simone Maria Massami Kitamura Martins

2009-08-01

Full Text Available Avaliou-se o efeito da suplementação de vitamina A na dieta sobre as características seminais (volume, motilidade, pH, vigor e concentração espermática, número total de espermatozoides, percentual de espermatozoides vivos e anormalidades morfológicas de reprodutores suínos no período de um ano (junho de 2004 a junho de 2005. Utilizaram-se dez reprodutores híbridos com 328,5 ± 2,12 dias de idade e 191,0 ± 12,0 kg distribuídos em duas rações, uma controle, com 10.000 UI de vit. A por kg de ração por dia, e outra com 16.000 UI de vit. A, fornecidas na quantidade de 2,5 kg/dia. O delineamento foi inteiramente casualizado com medidas repetidas no tempo. Não houve efeito significativo da suplementação de vitamina A na dieta nem de interação com o tempo nas características seminais estudadas. O tempo teve efeito significativo no pH, na concentração espermática, no número total de espermatozoides, no percentual de espermatozoides vivos, nas anormalidades morfológicas, na motilidade e no vigor espermático. Apesar da não-significância da suplementação de vitamina A, foi possível detectar diferenças numéricas no aumento da motilidade e de espermatozoides vivos, bem como na diminuição das anormalidades morfológicas, resultados que indicam ação da vitamina A.The objective of this study was to evaluate the effect of vitamin A feed supplementation on the following boar semen characteristics: volume, motility, vigor, seminal pH, spermatic concentration, total number of spermatozoa, percentage of living sperm cells and morphologic abnormalities. The experiment was carried out with 10 hybrid boars (328.5 ± 2.12 days of age and 191.0 ± 12.0 kg live weight. The boars were separated into two treatments and fed with two vitamin A levels: control, 10.000 UI/kg feed daily; and vitamin A, 16.000 UI. The boars received 2.5 kg feed daily. A randomized complete design was used with replication measurements in time. There was no

Hyaluronic acid improves frozen-thawed sperm quality and fertility potential in rooster.

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Lotfi, Saied; Mehri, Morteza; Sharafi, Mohsen; Masoudi, Reza

2017-09-01

Beneficial effects of Hyaluronic acid (HA) has not been yet assessed for cryopreservation of rooster sperm. This study was conducted to evaluate the effects of different concentrations of HA (0, 1, 2, 4 and 8mM) in Beltsville extender on the cryopreservation of rooster sperm. Semen samples were collected from six Ross broiler breeders (24-week) using abdominal massage, then divided into five equal aliquots and cryopreserved in Beltsville extender that contained different concentrations of HA. Motion characteristics, morphology, membrane functionality, viability, acrosome integrity, lipid peroxidation and fertility potential of sperm were assessed after thawing. HA at concentration of 2mM (HA2) resulted in the highest (Prooster sperm after freeze thawing. Copyright © 2017 Elsevier B.V. All rights reserved.

Tamoxifen is a potent antioxidant modulator for sperm quality in patients with idiopathic oligoasthenospermia.

Science.gov (United States)

Guo, Li; Jing, Jun; Feng, Yu-Ming; Yao, Bing

2015-09-01

To explore the new mechanisms of tamoxifen (TAM) in the treatment for patients with idiopathic oligoasthenospermia-antioxidation. In a prospective, randomized, controlled clinical trial, 120 cases of idiopathic oligoasthenospermia were enrolled and randomly assigned to the indomethacin group (n = 60) treated with indomethacin (25 mg, bid) and TAM group (n = 60) treated with TAM (10 mg, bid) for 3 months. Before and after treatment, we evaluated semen parameters, serum malondialdehyde (MDA) and total antioxidant capacity (TAC), seminal plasma MDA and TAC, spermatozoa intracellular reactive oxygen species (ROS), sperm succinate dehydrogenase (SDH) activity, sperm mitochondrial membrane potential (MMP), and sperm adenosine triphosphate (ATP) content. The independent t test and one-way repeated measures analysis of variance were used to compare the variables between and within two groups. In the indomethacin group, the percentage of progressive motile sperms, total motility, sperm MMP, and ATP content were increased significantly after 3-month treatment (P sperm count, sperm concentration, the percentage of progressive motile sperms, total motility, serum and seminal plasma TAC, sperm MMP, and ATP content were significantly improved or increased (P sperm count, sperm concentration, serum TAC, seminal plasma TAC, spermatozoa intracellular ROS, and sperm SDH activity. TAM treatment can significantly improve sperm quality, which is achieved through alleviating oxidative stress, improving sperm mitochondrial functionality, and subsequently increasing sperm motility.

137Cs monitoring in the meat of wild boar population in Slovakia

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Katarina Beňová

2016-06-01

Full Text Available Currently, due to the elapsed time and the nature of the Chernobyl accident, the only artificial radionuclide present in the soil is 137Cs, with a physical half-life conversion of 30.17 years. The 137Cs is quickly integrated into a biological cycle, similar to potassium. Generally, radionuclides are characterized by their mobility in soil. Contamination of materials and food by radionuclides represent a serious problem and has a negative impact on human health. The threat of international terrorism and the inability to forestall the impact of natural disasters on nuclear energetic (Fukushima accident, are also reasons for continuous monitoring of food safety. According screening measurement performed in European countries, high radioactivity levels were reported in the wild boars muscles from Sumava (Czech Republic. Seasonal fluctuation of 137Cs activity in the wild boar meat samples was observed in the forests on the southern Rhineland. Monitoring of 137Cs activity in the wild boar meat samples in the hunting grounds in Slovakia was initiated based on the reports on exceeding limits of the content of radiocaesium in the meat of wild boar from the surrounding countries. The aim of this study was to determine the 137Cs post Chernobyl contamination of wild boars population in different hunting districts of Slovakia during 2013 - 2014. A total of 60 thigh muscle samples from wild boars of different age categories (4 months - 2 years were evaluated. 137Cs activity was measured by gamma spectrometry (Canberra. Despite the fact Slovakia is closer to Chernobyl as Czech Republic and Germany, the 137Cs activity measured was very low and far below the permitted limit. The highest radiocaesium activity level measured in muscle was 37.2 Bq.kg-1 ±4.7%. Wild boar originated from Zlate Moravce district. The measurement results show, that 137Cs contamination levels of game in Slovakia are low. Radiocaesium activity in examined samples was very low and

THE INFLUENCE OF SEXUAL USE OF THE PIGS MALES ON THEIR SPERMOPRODUCTIONS

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ELENA MARANDICI

2008-10-01

Full Text Available The pigs rearing depends of the level of hold selection work and of the reproduction as wile it depends of the artificial insemination. The efficiency of artificial insemination depends of the reproductions’ quality. The large number of the boars which have to be used in the reproduction process do not correspond to the enquirements because their low sperm quality. One of the most important cause of the boars is their wrong rearing, their late testing. The aim of the experiments was to elaborate the optimal methods of growing using, and effective methods of boars testing grown for the reproduction of the Moldovenesc meat type breed. The held experiments allow to develop the growing technology and boars using in industrial farm what include: the young boars males selection at the age of 3 month, the free feeding, their maintenance, in the groups by 4 – 6 piglets until six month age, and beginning with five month age the young boars males are trained for the sperm collecting.

Cryopreservation of boar semen. III: Ultrastructure of boar spermatozoa frozen ultra-rapidly at various stages of conventional freezing and thawing.

Science.gov (United States)

Bwanga, C O; Ekwall, H; Rodriguez-Martinez, H

1991-01-01

Ejaculated boar spermatozoa subjected to a conventional freezing and thawing process, were ultra-rapidly fixed, freeze-substituted and examined by electron microscopy to monitor the presence of real or potential intracellular ice and the degree of cell protection attained with the different extenders used during the process. Numerous ice crystal marks representing the degree of hydration of the cells were located in the perinuclear space of those spermatozoa not in proper contact with the extender containing glycerol (i.e. prior to freezing). The spermatozoa which were in proper contact with the extenders presented a high degree of preservation of the acrosomes, plasma membranes as well as the nuclear envelopes. No ice marks were detected in acrosomes before thawing, indicating that the conventional assayed cryopreservation method provided a good protection against cryoinjury. The presence of acrosomal changes (internal vesiculization, hydration and swelling) in thawed samples however, raises serious questions about the thawing procedure employed.

Recovery of sperm after epididymal refrigeration from domestic cats using ACP-117c and Tris extenders

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D.B.C. Lima

2016-08-01

Full Text Available ABSTRACT We aimed to compare fresh sperm and sperm cooled to 4ºC that had been recovered from the epididymides of cats using powdered coconut water (ACP-117c and Tris extenders. Sixty epididymides were divided into 6 groups: 10 fresh epididymides were recovered using Tris (T0h; 10 were kept at 4°C/2h and recovered using Tris (T2h; 10 were kept at 4°C/4h and recovered using Tris (T4h; 10 fresh were recovered using ACP-117c (A0h; 10 were kept at 4°C/2h and recovered using ACP-117c (A2h, and 10 were kept at 4°C/4h and recovered using ACP-117c (A4h. The testis-epididymis complexes (TEC control were not cooled. The others were cooled at 4°C for 2 or 4h. The epididymis was separated and the sperm was recovered by the modified flotation method. Sperm kinetic parameters were evaluated by a computer-system analysis, and vigor, viability, concentration, membrane function and morphology of the sperm were assessed under a light microscope. The progressive motility with ACP-117c declined after 2h of cooling, but did not differ between fresh and 4h. The vigor and membrane function were higher in A4h than A0h. The vigor at T2h and T4h were decreased compared to T0h. T0h was higher than A0h for vigor and sperm membrane function. However, after 4h of cooling, ACP-117c maintained a higher percentage of living cells. Feline epididymal sperm quality can be maintained to the degree necessary for artificial breeding programs following cooling and ACP-117c may be successfully used to recover cat sperm that have been cooled for up to 4h.

Association between SNPs within candidate genes and compounds related to boar taint and reproduction

DEFF Research Database (Denmark)

Moe, Maren; Lien, Sigbjørn; Aasmundstad, Torunn

2009-01-01

BACKGROUND: Boar taint is an unpleasant odour and flavour of the meat from some uncastrated male pigs primarily caused by elevated levels of androstenone and skatole in adipose tissue. Androstenone is produced in the same biochemical pathway as testosterone and estrogens, which represents...... of this study was to detect SNPs in boar taint candidate genes and to perform association studies for both single SNPs and haplotypes with levels of boar taint compounds and phenotypes related to reproduction. RESULTS: An association study involving 275 SNPs in 121 genes and compounds related to boar taint...... and reproduction were carried out in Duroc and Norwegian Landrace boars. Phenotypes investigated were levels of androstenone, skatole and indole in adipose tissue, levels of androstenone, testosterone, estrone sulphate and 17beta-estradiol in plasma, and length of bulbo urethralis gland. The SNPs were genotyped...

Effects of feeding omega-3-fatty acids on fatty acid composition and quality of bovine sperm and on antioxidative capacity of bovine seminal plasma.

Science.gov (United States)

Gürler, Hakan; Calisici, Oguz; Calisici, Duygu; Bollwein, Heinrich

2015-09-01

The aim of the present study was to examine the effects of feeding alpha-linolenic (ALA) acid on fatty acid composition and quality of bovine sperm and on antioxidative capacity of seminal plasma. Nine bulls (ALA bulls) were fed with 800 g rumen-resistant linseed oil with a content of 50% linolenic acid and eight bulls with 400 g palmitic acid (PA bulls). Sperm quality was evaluated for plasma membrane and acrosome intact sperm (PMAI), the amount of membrane lipid peroxidation (LPO), and the percentage of sperm with a high DNA fragmentation index (DFI). Fatty acid content of sperm was determined using gas chromatography. Total antioxidant capacity, glutathione peroxidase, and superoxide dismutase activity were determined in seminal plasma. Feeding ALA increased (P acid (DHA) content in bulls whereas in PA bulls did not change. PMAI increased after cryopreservation in ALA bulls as well as in PA bulls during the experiment period (P fatty acids affect the antioxidant levels in seminal plasma. Both saturated as well as polyunsaturated fatty acids had positive effects on quality of cryopreserved bovine sperm, although the content of docosahexaenoic acid in sperm membranes increased only in ALA bulls. Copyright © 2015 Elsevier B.V. All rights reserved.

Effect of dietary vitamin E on the sperm quality of turbot ( Scophthalmus maximus)

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Xu, Houguo; Huang, Lina; Liang, Mengqing; Zheng, Keke; Wang, Xinxing

2015-08-01

A 3-month feeding experiment was conducted in an in-door seawater system to investigate the effect of dietary vitamin E (Ve) on the sperm quality of turbot ( Scophthalmus maximus). D-α-tocopherol acetate was supplemented to the basal (control) diet (65.14 mg kg-1 Ve) to obtain low and high levels of dietary Ve (244.60 mg kg-1, LVe; 721.60 mg kg-1, HVe). Compared with the control, sperm concentration was significantly increased in Ve-supplemented groups (LVe and HVe); while relative sperm volume and testis-somatic index were significantly increased in group HVe only. Sperm motility duration was significantly longer in group HVe than in the control, but no significant difference was observed in percent motility among groups. Sperm size, the uniformity of mitochondrial size, and the integrity of mitochondria cristae and plasma membrane were improved by dietary Ve, especially in HVe. The content of Ve in testis and liver as well as polyunsaturated fatty acids in sperm increased with dietary Ve. These results suggested that dietary Ve, especially at the high level (721.60 mg kg-1), significantly improved sperm concentration and motility duration and maintained normal sperm morphology of turbot.

Parasitic helminths of the digestive system of wild boars bred in captivity.

Science.gov (United States)

da Silva, Diego Silva; Müller, Gertrud

2013-01-01

This study aimed to identify the parasites that inhabit the digestive system of Sus scrofa scrofa from a commercial breeding facility in southern Brazil, and reports the first occurrence of Trichostrongylus colubriformis in wild boars. The gastrointestinal tracts of 40 wild boars from a commercial breeding facility were collected and individualized during slaughter in a cold-storage slaughterhouse. Out of this total, 87.5% were parasitized by the helminths Ascaris suum, Trichostrongylus colubriformis, Oesophagostomum dentatum and Trichuris suis. T. colubriformis presented a prevalence of 45%, mean intensity of 28.4 and mean abundance of 12.8. The data from this study showed that T. colubriformis not only has a capacity to develop in the small intestines of wild boars, but also adapts well to animals raised in captivity, thus representing a possible cause of economic loss in commercial wild boar farming.

Effect of Vitamin E and Polyunsaturated Fatty Acids on Cryopreserved Sperm Quality in Bos taurus Bulls Under Testicular Heat Stress.

Science.gov (United States)

Losano, João D A; Angrimani, Daniel S R; Dalmazzo, Andressa; Rocha, Carolina C; Brito, Maíra M; Perez, Eduardo G A; Tsunoda, Roberta H; Góes, Paola A A; Mendes, Camilla M; Assumpção, Mayra E O A; Barnabe, Valquiria H; Nichi, Marcilio

2018-04-03

Taurine bulls are highly susceptible to heat stress, leading to increased oxidative stress (OS) and impaired sperm viability. Polyunsaturated fatty acids (PUFAs) supplementation can be an alternative to improve semen quality, which also results in more sperm susceptibility to lipid peroxidation. Moreover, this deleterious effect can be exacerbated in animals affected by heat stress. Vitamin E is a key antioxidant that counteracts lipid peroxidation of sperm membrane caused by OS. Thus, combining PUFAs with vitamin E may improve sperm quality. In this context, this study aimed to evaluate the effect of interaction between PUFAs and vitamin E on sperm quality in Bos taurus bulls under testicular heat stress. Sixteen taurine bulls under testicular heat stress were randomly assigned in four groups: Control, Vitamin E, PUFA, and PUFA + Vitamin E. All groups lasted for 60 days. Samples were cryopreserved/thawed and analyzed for motility variables (CASA), membrane and acrosome integrity, mitochondrial activity, susceptibility to oxidative stress, DNA integrity, and sperm-binding capacity. Results showed that vitamin E had a beneficial effect on some sperm characteristics, whereas PUFA supplementation had an adverse effect when the two treatments were evaluated separately. Finally, the association between PUFAs and vitamin E did not improve sperm quality.

Meat quality characteristics of DurocxYorkshire, DurocxYorkshirexwild boar and wild boar

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Ivanović Snežana D.

2013-01-01

Full Text Available Chemical composition, pH value, fatty acids profile, cholesterol content, color and sensory analysis of pork meat from Duroc x Yorkshire (D x Y, Duroc x Yorkshire x wild boar (D x Y x WB crossbreeds and wild boars (WB was investigated. Samples for all tests were taken from m. longissimus dorsi. Chemical composition and pH value were tested by ISO methods. Fatty acid and cholesterol determination was performed by gas chromatography technique with external standard method. Color was determined instrumentally using the thristimulus colourimeter. The overall sensoric quality (appearance, texture and smell of samples of raw meat was evaluated. In evaluation of results the scoring system was used. In chemical composition (moisture, fat, protein, ash and pH values statistically significant difference was noted (p<0,05 between each of the examined groups. Also, among all the examined groups statistically significant difference (p<0,05 was found for fatty acids and cholesterol content. Measurment of the color of meat from all three groups showed that the L*, a * b *, Chroma and Hue angle were also statistically significantly different (p<0,01.

Individual adjustment of sperm expenditure accords with sperm competition theory.

Science.gov (United States)

Pilastro, Andrea; Scaggiante, Marta; Rasotto, Maria B

2002-07-23

Sperm competition theory predicts that males should strategically allocate their sperm reserves according to the level of sperm competition, defined as the probability that the sperm of two males compete for fertilizing a given set of ova. Substantial evidence from numerous animal taxa suggests that, at the individual level, sperm expenditure increases when the risk of sperm competition is greater. In contrast, according to the "intensity model" of sperm competition [Parker, G. A., Ball, M. A., Stockley, P. & Gage, M. J. G. (1996) Proc. R. Soc. London Ser. B 263, 1291-1297], when more than two ejaculates compete during a given mating event, sperm expenditure should decrease as the number of competing males increases. Empirical evidence supporting this prediction, however, is still lacking. Here we measured sperm expenditure in two gobiid fishes, the grass (Zosterisessor ophiocephalus) and black goby (Gobius niger), in which up to six sneakers can congregate around the nest of territorial males and release their sperm when females spawn. We show that, in accordance with theory, sneaker males of both species release fewer sperm as the number of competitors increases.

How much does it cost to look like a pig in a wild boar group?

DEFF Research Database (Denmark)

Batocchio, Daniele; Iacolina, Laura; Canu, Antonio

2017-01-01

Hybridization between domestic and wild species is known to widely occur and it is reported to be oneof the major causes of the current biodiversity crisis. Despite this, poor attention has been deserved tothe behavioural ecology of hybrids, in particular in relation to their social behaviour. We...... carried out acamera trap study to assess whether phenotypically anomalous colouration in wild boar, i.e. potentiallyintrogressed with domestic pigs, affected the hierarchical structure of wild boar social groups. Chromat-ically anomalous wild boars (CAWs) were detected in 32 out of 531 wild boar videos...

Electrophoretic and zymographic characterization of proteins isolated by various extraction methods from ejaculated and capacitated boar sperms

Czech Academy of Sciences Publication Activity Database

Zigo, Michal; Jonáková, Věra; Maňásková-Postlerová, Pavla

2011-01-01

Roč. 32, č. 11 (2011), s. 1309-1318 ISSN 0173-0835 R&D Projects: GA ČR(CZ) GA303/09/1285; GA MZd(CZ) NS10009; GA MŠk(CZ) 1M06011 Institutional research plan: CEZ:AV0Z50520701 Keywords : Extraction methods * Sperm proteins * Substrate zymography Subject RIV: CE - Biochemistry Impact factor: 3.303, year: 2011

Seroprevalence of hepatitis E virus in domestic pigs and wild boars in Switzerland.

Science.gov (United States)

Burri, C; Vial, F; Ryser-Degiorgis, M-P; Schwermer, H; Darling, K; Reist, M; Wu, N; Beerli, O; Schöning, J; Cavassini, M; Waldvogel, A

2014-12-01

Hepatitis E is considered an emerging human viral disease in industrialized countries. Studies from Switzerland report a human seroprevalence of hepatitis E virus (HEV) of 2.6-21%, a range lower than in adjacent European countries. The aim of this study was to determine whether HEV seroprevalence in domestic pigs and wild boars is also lower in Switzerland and whether it is increasing and thus indicating that this zoonotic viral infection is emerging. Serum samples collected from 2,001 pigs in 2006 and 2011 and from 303 wild boars from 2008 to 2012 were analysed by ELISA for the presence of HEV-specific antibodies. Overall HEV seroprevalence was 58.1% in domestic pigs and 12.5% in wild boars. Prevalence in domestic pigs was significantly higher in 2006 than in 2011. In conclusion, HEV seroprevalence in domestic pigs and wild boars in Switzerland is comparable with the seroprevalence in other countries and not increasing. Therefore, prevalence of HEV in humans must be related to other factors than prevalence in pigs or wild boars. © 2014 Blackwell Verlag GmbH.

Changes in subcellular elemental distributions accompanying the acrosome reaction in sea urchin sperm

International Nuclear Information System (INIS)

Cantino, M.E.; Schackmann, R.W.; Johnson, D.E.

1983-01-01

Energy-dispersive x-ray microanalysis was used to analyze changes in the subcellular distributions of Na, Mg, P, S, Cl, K, and Ca associated with the acrosome reaction of sea urchin sperm. Within 5 sec after induction of the acrosome reaction, nuclear Na and mitochondrial Ca increased and nuclear and mitochondrial K decreased. Uptake of mitochondrial P was detected after several minutes, and increases in nuclear Mg were detected only after 5-10 min of incubation following induction of the reaction. The results suggest that sudden permeability changes in the sperm plasma membrane are associated with the acrosome reaction, but that complete breakdown of membrane and cell function does not occur for several minutes

Quantification of leptin in seminal plasma of buffalo bulls and its correlation with antioxidant status, conventional and computer-assisted sperm analysis (CASA) semen variables.

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Kumar, Pradeep; Saini, Monika; Kumar, Dharmendra; Jan, M H; Swami, Dheer Singh; Sharma, R K

2016-03-01

The present study is the first to quantify leptin in seminal plasma of buffalo and investigate its relationship with seminal attributes. Ten ejaculates each from 10 Murrah buffalo bulls were collected. Semen quality variables such as semen volume, sperm concentration, sperm abnormalities, membrane integrity, antioxidant enzyme activities (superoxide dismutase, catalase and total antioxidant capacity), malondialdehyde (MDA) concentration, as well as sperm kinetics and motility variables were evaluated. The leptin concentration in serum and seminal plasma were estimated by the ELISA method. Bulls were classified in two groups on the basis of sperm concentration with Group I having >800 million sperm/mL and Group II <500 million sperm/mL. Greater (P<0.05) mean sperm abnormalities, seminal leptin concentrations and MDA concentrations were recorded in Group II than Group I. The seminal leptin was positively correlated with sperm abnormalities and MDA concentration while being negatively correlated with sperm concentration, but there was no correlation with sperm kinetic and motility variables, sperm membrane integrity and seminal plasma antioxidant enzyme activity. Thus, the data suggest that seminal leptin has a role in spermatogenesis and can be used as a marker for spermatogenesis to predict the capacity of buffalo bulls for semen production. Copyright © 2016 Elsevier B.V. All rights reserved.

Niche conservatism and the invasive potential of the wild boar.

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Sales, Lilian Patrícia; Ribeiro, Bruno R; Hayward, Matt Warrington; Paglia, Adriano; Passamani, Marcelo; Loyola, Rafael

2017-09-01

Niche conservatism, i.e. the retention of a species' fundamental niche through evolutionary time, is cornerstone for biological invasion assessments. The fact that species tend to maintain their original climate niche allows predictive maps of invasion risk to anticipate potential invadable areas. Unravelling the mechanisms driving niche shifts can shed light on the management of invasive species. Here, we assessed niche shifts in one of the world's worst invasive species: the wild boar Sus scrofa. We also predicted potential invadable areas based on an ensemble of three ecological niche modelling methods, and evaluated the performance of models calibrated with native vs. pooled (native plus invaded) species records. By disentangling the drivers of change on the exotic wild boar population's niches, we found strong evidence for niche conservatism during biological invasion. Ecological niche models calibrated with both native and pooled range records predicted convergent areas. Also, observed niche shifts are mostly explained by niche unfilling, i.e. there are unoccupied areas in the exotic range where climate is analogous to the native range. Niche unfilling is expected as result of recent colonization and ongoing dispersal, and was potentially stronger for the Neotropics, where a recent wave of introductions for pig-farming and game-hunting has led to high wild boar population growth rates. The invasive potential of wild boar in the Neotropics is probably higher than in other regions, which has profound management implications if we are to prevent their invasion into species-rich areas, such as Amazonia, coupled with expansion of African swine fever and possibly great economic losses. Although the originally Eurasian-wide distribution suggests a pre-adaptation to a wide array of climates, the wild boar world-wide invasion does not exhibit evidence of niche evolution. The invasive potential of the wild boar therefore probably lies on the reproductive, dietary and

Sperm head's birefringence: a new criterion for sperm selection.

Science.gov (United States)

Gianaroli, Luca; Magli, M Cristina; Collodel, Giulia; Moretti, Elena; Ferraretti, Anna P; Baccetti, Baccio

2008-07-01

To investigate the characteristics of birefringence in human sperm heads and apply polarization microscopy for sperm selection at intracytoplasmic sperm injection (ICSI). Prospective randomized study. Reproductive Medicine Unit, Società Italiana Studi Medicina della Riproduzione, Bologna, Italy. A total of 112 male patients had birefringent sperm selected for ICSI (study group). The clinical outcome was compared with that obtained in 119 couples who underwent a conventional ICSI cycle (control group). The proportion of birefringent spermatozoa was evaluated before and after treatment in relation to the sperm sample quality. Embryo development and clinical outcome in the study group were compared with those in the controls. Proportion of birefringent sperm heads, rates of fertilization, cleavage, pregnancy, implantation, and ongoing implantation. The proportion of birefringent spermatozoa was significantly higher in normospermic samples when compared with oligoasthenoteratospermic samples with no progressive motility and testicular sperm extraction samples. Although fertilization and cleavage rates did not differ between the study and control groups, in the most severe male factor condition (oligoasthenoteratospermic with no progressive motility and testicular sperm extraction), the rates of clinical pregnancy, ongoing pregnancy, and implantation were significantly higher in the study group versus the controls. The analysis of birefringence in the sperm head could represent both a diagnostic tool and a novel method for sperm selection.

Parasitic helminths of the digestive system of wild boars bred in captivity

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Diego Silva da Silva

Full Text Available This study aimed to identify the parasites that inhabit the digestive system of Sus scrofa scrofa from a commercial breeding facility in southern Brazil, and reports the first occurrence of Trichostrongylus colubriformis in wild boars. The gastrointestinal tracts of 40 wild boars from a commercial breeding facility were collected and individualized during slaughter in a cold-storage slaughterhouse. Out of this total, 87.5% were parasitized by the helminths Ascaris suum,Trichostrongylus colubriformis, Oesophagostomum dentatum and Trichuris suis. T. colubriformis presented a prevalence of 45%, mean intensity of 28.4 and mean abundance of 12.8. The data from this study showed that T. colubriformis not only has a capacity to develop in the small intestines of wild boars, but also adapts well to animals raised in captivity, thus representing a possible cause of economic loss in commercial wild boar farming.

Sperm competition selects for sperm quantity and quality in the Australian Maluridae.

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Rowe, Melissah; Pruett-Jones, Stephen

2011-01-25

When ejaculates from rival males compete for fertilization, there is strong selection for sperm traits that enhance fertilization success. Sperm quantity is one such trait, and numerous studies have demonstrated a positive association between sperm competition and both testes size and the number of sperm available for copulations. Sperm competition is also thought to favor increases in sperm quality and changes in testicular morphology that lead to increased sperm production. However, in contrast to sperm quantity, these hypotheses have received considerably less empirical support and remain somewhat controversial. In a comparative study using the Australian Maluridae (fairy-wrens, emu-wrens, grasswrens), we tested whether increasing levels of sperm competition were associated with increases in both sperm quantity and quality, as well as an increase in the relative amount of seminiferous tubule tissue contained within the testes. After controlling for phylogeny, we found positive associations between sperm competition and sperm numbers, both in sperm reserves and in ejaculate samples. Additionally, as sperm competition level increased, the proportion of testicular spermatogenic tissue also increased, suggesting that sperm competition selects for greater sperm production per unit of testicular tissue. Finally, we also found that sperm competition level was positively associated with multiple sperm quality traits, including the proportion of motile sperm in ejaculates and the proportion of both viable and morphologically normal sperm in sperm reserves. These results suggest multiple ejaculate traits, as well as aspects of testicular morphology, have evolved in response to sperm competition in the Australian Maluridae. Furthermore, our findings emphasize the importance of post-copulatory sexual selection as an evolutionary force shaping macroevolutionary differences in sperm phenotype.

Method-related estimates of sperm vitality.

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Cooper, Trevor G; Hellenkemper, Barbara

2009-01-01

Comparison of methods that estimate viability of human spermatozoa by monitoring head membrane permeability revealed that wet preparations (whether using positive or negative phase-contrast microscopy) generated significantly higher percentages of nonviable cells than did air-dried eosin-nigrosin smears. Only with the latter method did the sum of motile (presumed live) and stained (presumed dead) preparations never exceed 100%, making this the method of choice for sperm viability estimates.

Freeze-tolerance of Trichinella muscle larvae in experimentally infected wild boars.

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Lacour, Sandrine A; Heckmann, Aurélie; Macé, Pauline; Grasset-Chevillot, Aurélie; Zanella, Gina; Vallée, Isabelle; Kapel, Christian M O; Boireau, Pascal

2013-05-20

Freeze-tolerance of encapsulated Trichinella muscle larvae (ML) is mainly determined by Trichinella species, but is also influenced by host species, the age of the infection and the storage time and temperature of the infected meat. Moreover, the freeze-tolerance of the encapsulated species appears to be correlated to the development of thick capsule walls which increases with age. An extended infection period and the muscle composition in some hosts (e.g. herbivores) may provide freeze-avoiding matrices due to high carbohydrate contents. The present experiment compares freeze-tolerance of Trichinella spiralis and Trichinella britovi ML in wild boar meat 24 weeks post inoculation (wpi). Three groups of four wild boars were infected with 200, 2000 or 20,000 ML of T. britovi (ISS 1575), respectively. Additionally, three wild boars were inoculated with 20,000 ML of T. spiralis (ISS 004) and two animals served as negative controls. All wild boars were sacrificed 24 wpi. Muscle samples of 70 g were stored at -21°C for 19, 30 and 56 h, and for 1-8 weeks. Larvae were recovered by artificial digestion. Their mobilities were recorded using Saisam(®) image analysis software and their infectivities were evaluated using mouse bioassays. Samples frozen for 19, 30 and 56 h allowed recovery of mobile ML, but samples frozen for 1 or 2 weeks did not. Correspondingly, only T. spiralis and T. britovi larvae isolated from wild boar meat frozen for 19, 30 and 56 h established in mice. This study showed that freezing at -21°C for 1 week inactivated T. spiralis and T. britovi ML encapsulated in wild boar meat for 24 weeks. Copyright © 2013 Elsevier B.V. All rights reserved.

Helminth parasites of wild boar, Sus scrofa, in Iran.

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Eslami, A; Farsad-Hamdi, S

1992-04-01

Fifty-seven wild boars (Sus scrofa) from protected regions of Iran were examined for helminths. Sixteen species of helminths were collected; there were ten nematodes, one acanthocephalan, two trematodes and three larval cestodes. New host and distribution records were established for all helminths except of Taenia solium cysticerci. Wild boar shared nine of these helminths with domestic pigs, six with ruminants and three with human beings in Iran. Metastrongylus pudendotectus and M. salmi are reported for the first time from Iran.

Sperm competition promotes diversity of sperm bundles in Ohomopterus ground beetles

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Takami, Yasuoki; Sota, Teiji

2007-07-01

Diversification of sperm morphology has been investigated in the context of sperm competition, but the adaptive significance of sperm bundles is still unclear. In analyzing 10 taxa of the genus Carabus subgenus Ohomopterus and one related Carabus ground beetles, we found that dimorphic sperm bundles occurred in most species with varied degrees of bimodality, whereas sperm were generally monomorphic. Comparative analyses with phylogenetically independent contrasts revealed that the sizes of large and small sperm bundles evolved more rapidly than, and were not correlated with, the length of sperm, suggesting more intense selection on sperm bundle sizes and their independent responses to different evolutionary forces. The size of large sperm bundles was positively correlated with male genital morphology (pertinent to displacement of rival spermatophores) and postcopulatory guarding duration as well as male body length, suggesting that larger sperm bundles have been favored when the risk of spermatophore displacement is high. Larger sperm bundles may be advantageous because of their ability to migrate more rapidly into the spermatheca. In contrast, no clear association was detected between the small sperm bundle size and mating traits despite its rapid diversification. The present study provides the first record of heteromorphic sperm bundles, the diversity of which may be promoted by sperm competition.

Rogue sperm indicate sexually antagonistic coevolution in nematodes.

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Ronald E Ellis

2014-07-01

Full Text Available Intense reproductive competition often continues long after animals finish mating. In many species, sperm from one male compete with those from others to find and fertilize oocytes. Since this competition occurs inside the female reproductive tract, she often influences the outcome through physical or chemical factors, leading to cryptic female choice. Finally, traits that help males compete with each other are sometimes harmful to females, and female countermeasures may thwart the interests of males, which can lead to an arms race between the sexes known as sexually antagonistic coevolution. New studies from Caenorhabditis nematodes suggest that males compete with each other by producing sperm that migrate aggressively and that these sperm may be more likely to win access to oocytes. However, one byproduct of this competition appears to be an increased probability that these sperm will go astray, invading the ovary, prematurely activating oocytes, and sometimes crossing basement membranes and leaving the gonad altogether. These harmful effects are sometimes observed in crosses between animals of the same species but are most easily detected in interspecies crosses, leading to dramatically lowered fitness, presumably because the competitiveness of the sperm and the associated female countermeasures are not precisely matched. This mismatch is most obvious in crosses involving individuals from androdioecious species (which have both hermaphrodites and males, as predicted by the lower levels of sperm competition these species experience. These results suggest a striking example of sexually antagonistic coevolution and dramatically expand the value of nematodes as a laboratory system for studying postcopulatory interactions.

Social genetic effects for growth in pigs differ between boars and gilts

DEFF Research Database (Denmark)

Nielsen, Hanne M.; Ask, Birgitte; Madsen, Per

2018-01-01

between boars and gilts and that accounting for these differences will improve the predictive ability of a social genetic effects model (SGM). Our data consisted of ADG from 30 to 94 kg for 32,212 uncastrated males (boars) and 48,252 gilts that were raised in sex-specific pens. Data were analyzed using......Average daily gain (ADG) in pigs is affected by the so-called social (or indirect) genetic effects (SGE). However, SGE may differ between sexes because boars grow faster than gilts and their social behaviours differ. We hypothesized that direct genetic effects (DGE) and SGE for ADG in pigs differ...

Effect of semen collection practices on sperm characteristics before and after storage and on fertility of stallions.

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Sieme, H; Katila, T; Klug, E

2004-02-01

This study analyzed effects of different methods and intervals of semen collection on the quantity and quality of fresh, cool-stored, and frozen-thawed sperm and fertility of AI stallions. In Experiment 1, ejaculates were obtained from six stallions (72 ejaculates per stallion) using fractionated versus non-fractionated semen collection techniques. Initial sperm quality of the first three jets of the ejaculate was not different from that of total ejaculates. Centrifugation of sperm-rich fractions before freezing improved post-thaw motility and sperm membrane integrity when compared to non-centrifuged sperm-rich fractions or non-fractionated centrifuged ejaculates (Psperm concentrations, percentages of progressively motile sperm (PMS) after storage for 24h at 5 degrees C and lower percentages of midpiece alterations than single daily ejaculates. Semen collected once daily showed significantly lower values of live sperm after freezing and thawing than the first ejaculate of two ejaculates collected 1h apart every 48 h. In Experiment 3, semen was collected from 36 stallions (> or =12 ejaculates per stallion) during the non-breeding season and the time to ejaculation and the number of mounts was recorded. When time to ejaculation and the number of mounts increased, volume and total sperm count (TSC) also increased (Psperm concentration, percentage of PMS after storage for 24 h at 5 degrees C, percentage of membrane-intact sperm in fresh semen (Psperm of frozen-thawed sperm (Psperm concentration and percentage of PMS after 24-h cool-storage decreased with increasing number of mounts on the phantom (P or =1 on an average per day) showed significantly higher FRs than mares inseminated with semen from stallions with a daily collection frequency of 0.5-1 or 2.5 days.

Oxidative Stress in Wild Boars Naturally and Experimentally Infected with Mycobacterium bovis.

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Diana Gassó

Full Text Available Reactive oxygen and nitrogen species (ROS-RNS are important defence substances involved in the immune response against pathogens. An excessive increase in ROS-RNS, however, can damage the organism causing oxidative stress (OS. The organism is able to neutralise OS by the production of antioxidant enzymes (AE; hence, tissue damage is the result of an imbalance between oxidant and antioxidant status. Though some work has been carried out in humans, there is a lack of information about the oxidant/antioxidant status in the presence of tuberculosis (TB in wild reservoirs. In the Mediterranean Basin, wild boar (Sus scrofa is the main reservoir of TB. Wild boar showing severe TB have an increased risk to Mycobacterium spp. shedding, leading to pathogen spreading and persistence. If OS is greater in these individuals, oxidant/antioxidant balance in TB-affected boars could be used as a biomarker of disease severity. The present work had a two-fold objective: i to study the effects of bovine TB on different OS biomarkers (namely superoxide dismutase (SOD, catalasa (CAT, glutathione peroxidase (GPX, glutathione reductase (GR and thiobarbituric acid reactive substances (TBARS in wild boar experimentally challenged with Mycobacterium bovis, and ii to explore the role of body weight, sex, population and season in explaining the observed variability of OS indicators in two populations of free-ranging wild boar where TB is common. For the first objective, a partial least squares regression (PLSR approach was used whereas, recursive partitioning with regression tree models (RTM were applied for the second. A negative relationship between antioxidant enzymes and bovine TB (the more severe lesions, the lower the concentration of antioxidant biomarkers was observed in experimentally infected animals. The final PLSR model retained the GPX, SOD and GR biomarkers and showed that 17.6% of the observed variability of antioxidant capacity was significantly correlated

Development of Domestic Cat Embryo Produced by Preserved Sperms

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KARTINI ERIANI

2008-12-01

Full Text Available The ability to mature and fertilize oocytes of endangered species may allow us to sustain genetic and global biodiversity. Epididymis sperms may be the last chance to ensure preservation of genetic materials after injury or death of a valuable animal. Studies have been conducted to determine wether both epididymis sperms and oocytes can be used to produce viable embryos and offspring. The purpose of this study was to determine how long cats sperms contained in epididymis were remain motile and had intact membranes when preserved at 4 ° C, and to determine whether such those preserved sperms are able to fertilize oocytes. Epididymis was preserved immediately in phosphate buffer saline at 4 ° C for 1, 3, and 6 days. The observation of sperm quality and viability after preservation was performed by vital staining acrosom and Hoechst-Propidium Iodine. Biological functions of sperms were evaluated by in vitro culture technique for fertilization, micro fertilization and embryonic development rate in CR1aa medium. The results showed that average motility of sperms collected from ductus deferens, cauda and corpus epididymis decreased not significantly (P > 0.05 from 0, 1, 3, and 6 days of preservation times (from 83.0%, 80.2%, 79.0%; 80.9%, 75.0%, 75.5%; 52.0%, 63.2%, 55.0% to 34.6%, 34.6%, 33.3%, respectively. The general results showed that sperms from epididymis preserved for 1, 3, and 6 days can be used for IVF. The rate of embryonal cleavage produced by IVF technique using sperms collected from epididymis preserved for 1-, 3- and 6-days were 33.3, 26.7, and 20.0%, respectively and significantly different (p < 0.05 from that of controll (50.0%. In conclusion, sperms contained in epididyimis preserved at 4 ° C in PBS (Phospate Buffer Saline for 1-6 days can be used to IVF and in vitro production of cat embryos.

Comparison of two different extenders for cryopreservation of epididymal dog sperm.

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Martins, Mim; Justino, R C; Sant'anna, M C; Trautwein, L G C; Souza, F F

2012-12-01

The collection of epididymal sperm is an option for preservation of germplasm of genetically superior animals that need to be orchiectomized or have died. The extender type used to freeze sperm is important to avoid spermatozoal membrane damage and to preserve semen quality after cryopreservation. The objective of this study was to verify the effects of a commercial bovine extender (Bovimix(®); Nutricell, Campinas) and a traditional TRIS-citric acid-glucose-egg yolk-7% glycerol extender on cryopreservation of canine epididymal sperm. The testes of 13 adult dogs were kept at 5 °C for 24 h in saline solution, and epididymal sperm was recovered in Ringers solution without lactate and were evaluated for motility. Samples with ≥ 80% motility were pooled and then divided before dilution and packaging in 0.5 ml plastic straws, equilibration at 4 °C for 1 h, freezing in nitrogen vapour for 20 min and storing at -196 °C. The straws were thawed at 56 °C for 10 s and were evaluated for motility by computer assisted analysis (CASA). The semen parameters, sperm movement index, linearity, total motility and rapid progressive motility were statistically higher in Bovimix(®) than TRIS. In contrast, amplitude of lateral head displacement, slow sperm and static sperm were lower in Bovimix(®). Despite the high percentage of sperm defects in epididymal cells, regardless of the extender, we concluded that Bovimix(®) is a viable alternative for the freezing of canine epididymal sperm. © 2012 Blackwell Verlag GmbH.

Expression and secretion of plasma membrane Ca2+-ATPase 4a (PMCA4a during murine estrus: association with oviductal exosomes and uptake in sperm.

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Amal A Al-Dossary

Full Text Available PMCA4, a membrane protein, is the major Ca(2+ efflux pump in murine sperm where its deletion leads to a severe loss of hyperactivated motility and to male infertility. We have previously shown that the PMCA4b splice variant interacts with CASK (Ca(2+/CaM-dependent serine kinase in regulating sperm Ca(2+. More recently we detected that PMCA4a isoform, in addition to its presence in testis, is secreted in the epididymal luminal fluid and transferred to sperm. Here we show that Pmca4 mRNA is expressed in both the 4a and 4b variants in the vagina, uterus, and oviduct. Immunofluorescence reveals that PMCA4a is similarly expressed and is elevated during estrus, appearing in the glandular and luminal epithelia. Western analysis detected PMCA4a in all tissues and in the luminal fluids (LF of the vagina (VLF, uterus (ULF, and the oviduct (OLF collected during estrus. It was ~9- and 4-fold higher in OLF than in VLF and ULF, and only marginally present in LF collected at metestrus/diestrus. Fractionation of the LF collected at estrus, via ultracentrifugation, revealed that 100% of the PMCA4a resides in the vesicular fraction of the ULF and OLF. Transmission electron microscopy (TEM revealed that OLF vesicles have an exosomal orientation (with the cytoplasmic-side inward, a size range of 25-100 nm, with the characteristic CD9 biomarker. Thus, we dubbed these vesicles "oviductosomes", to which PMCA4a was immunolocalized. Incubation of caudal sperm in the combined LF or exosomes resulted in up to a ~3-fold increase of sperm PMCA4a, as detected by flow cytometry, indicating in vitro uptake. Our results are consistent with the increased requirement of Ca(2+ efflux in the oviduct. They show for the first time the presence of oviductal exosomes and highlight their role, along with uterosomes and vaginal exosomes, in post-testicular sperm acquisition of PMCA4a which is essential for hyperactivated motility and fertility.

Expression and secretion of plasma membrane Ca2+-ATPase 4a (PMCA4a) during murine estrus: association with oviductal exosomes and uptake in sperm.

Science.gov (United States)

Al-Dossary, Amal A; Strehler, Emanuel E; Martin-Deleon, Patricia A

2013-01-01

PMCA4, a membrane protein, is the major Ca(2+) efflux pump in murine sperm where its deletion leads to a severe loss of hyperactivated motility and to male infertility. We have previously shown that the PMCA4b splice variant interacts with CASK (Ca(2+/)CaM-dependent serine kinase) in regulating sperm Ca(2+). More recently we detected that PMCA4a isoform, in addition to its presence in testis, is secreted in the epididymal luminal fluid and transferred to sperm. Here we show that Pmca4 mRNA is expressed in both the 4a and 4b variants in the vagina, uterus, and oviduct. Immunofluorescence reveals that PMCA4a is similarly expressed and is elevated during estrus, appearing in the glandular and luminal epithelia. Western analysis detected PMCA4a in all tissues and in the luminal fluids (LF) of the vagina (VLF), uterus (ULF), and the oviduct (OLF) collected during estrus. It was ~9- and 4-fold higher in OLF than in VLF and ULF, and only marginally present in LF collected at metestrus/diestrus. Fractionation of the LF collected at estrus, via ultracentrifugation, revealed that 100% of the PMCA4a resides in the vesicular fraction of the ULF and OLF. Transmission electron microscopy (TEM) revealed that OLF vesicles have an exosomal orientation (with the cytoplasmic-side inward), a size range of 25-100 nm, with the characteristic CD9 biomarker. Thus, we dubbed these vesicles "oviductosomes", to which PMCA4a was immunolocalized. Incubation of caudal sperm in the combined LF or exosomes resulted in up to a ~3-fold increase of sperm PMCA4a, as detected by flow cytometry, indicating in vitro uptake. Our results are consistent with the increased requirement of Ca(2+) efflux in the oviduct. They show for the first time the presence of oviductal exosomes and highlight their role, along with uterosomes and vaginal exosomes, in post-testicular sperm acquisition of PMCA4a which is essential for hyperactivated motility and fertility.

Investigations on the contamination of Saxonian wild boars with radiocaesium

International Nuclear Information System (INIS)

Heinrich, T.; Abraham, A.; Preusse, W.; Pianski, J.; Alisch-Mark, M.; Lange, S.

2016-01-01

As a result of the Chernobyl fallout some parts of the free state of Saxony were contaminated with radioactive caesium. Based on published maps of the soil contamination and on additional investigations some regions of elevated contamination could be localized. Parallel to soil investigations a game monitoring to wild boars and roe deer was performed. For both types of game typical seasonal variations of contamination were found. In Saxony only the contamination of wild boars is important. In the south of the Vogtland a region was found, where in all seasons the recommended high value of 600 Bq/kg was exceeded in game. In this region the investigation on radiocaesium is now obligatory for wild boars. The hunter can combine this analysis with the analysis on trichina. After three years measurements the region for obligatory analysis was adapted and expanded to neighbouring counties.

137Cs in the meat of wild boars: a comparison of the impacts of Chernobyl and Fukushima.

Science.gov (United States)

Steinhauser, Georg; Saey, Paul R J

The impact of Chernobyl on the 137 Cs activities found in wild boars in Europe, even in remote locations from the NPP, has been much greater than the impact of Fukushima on boars in Japan. Although there is great variability within the 137 Cs concentrations throughout the wild boar populations, some boars in southern Germany in recent years exhibit higher activity concentrations (up to 10,000 Bq/kg and higher) than the highest 137 Cs levels found in boars in the governmental food monitoring campaign (7900 Bq/kg) in Fukushima prefecture in Japan. The levels of radiocesium in boar appear to be more persistent than would be indicated by the constantly decreasing 137 Cs inventory observed in the soil which points to a food source that is highly retentive to 137 Cs contamination or to other radioecological anomalies that are not yet fully understood.

Molecular epidemiology of Mycoplasma hyopneumoniae from outbreaks of enzootic pneumonia in domestic pig and the role of wild boar.

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Kuhnert, Peter; Overesch, Gudrun

2014-11-07

Mycoplasma hyopneumoniae is the major cause of enzootic pneumonia (EP) in domestic pigs, a disease with low mortality but high morbidity, having a great economic impact for producers. In Switzerland EP has been successfully eradicated, however, sporadic outbreaks are observed with no obvious source. Besides the possibility of recurrent outbreaks due to persisting M. hyopneumoniae strains within the pig population, there is suspicion that wild boars might introduce M. hyopneumoniae into swine herds. To elucidate possible links between domestic pig and wild boar, epidemiological investigations of recent EP outbreaks were initiated and lung samples of pig and wild boar were tested for the presence of specific genotypes by multilocus sequence typing (MLST). Despite generally different genotypes in wild boar, outbreak strains could be found in geographically linked wild boar lungs after, but so far not before the outbreak. Recurrent outbreaks in a farm were due to the same strain, indicating unsuccessful sanitation rather than reintroduction by wild boar. In another case outbreaks in six different farms were caused by the same strain never found in wild boar, confirming spread between farms due to hypothesized animal transport. Results indicate the presence of identical lineages of wild boar and domestic pig strains, and possible transmission of M. hyopneumoniae between wild boar and pig. However, the role of wild boar might be rather one as a recipient than a transmitter. More important than contact to wild boar for sporadic outbreaks in Switzerland is apparently persistence of M. hyopneumoniae within a farm as well as transmission between farms. Copyright © 2014 Elsevier B.V. All rights reserved.

Flow cytometry application in the assessment of sperm DNA integrity of men with asthenozoospermia.

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A Brodowska

2008-04-01

Full Text Available Sperm genomic integrity and ultrastructural features of ejaculated spermatozoa contributing to the assessment of gamete fertility potential in patients with asthenozoospermia are discussed. The proportion of TUNEL-positive cells was significantly higher in the semen of patients with low sperm motility (n=40; p<0.01 as compared to men with normal sperm motility (n=54. Sperm DNA fragmentation negatively correlated (n=94 with sperm motility, sperm concentration, and integrity of the sperm cellular membrane (HOS-test. Two categories of patients were distinguished: (1 patients (23 out of 94 subjects with < or = 4% of TUNEL-positive cells and (2 patients (71 subjects with 4% of TUNEL-positive cells. A significant difference was noted in the sperm motility and HOS-test results between patients from both groups. Large numbers of immature spermatozoa with extensive cytoplasmic retention, ultrastructural chromatin and midpiece abnormalities, and conglomerates containing sperm fragments were present more frequently in the semen of asthenozoospermic subjects with >4% of TUNEL-positive sperm cells. Low sperm motility seems to be accompanied by serious defects of gamete chromatin expressed as diminished sperm genomic integrity and abnormal DNA condensation and by defects of sperm midpiece. These abnormalities may reflect developmental failure during the spermatogenic remodeling process. The DNA fragmentation test may be considered as an additional assay for the evaluation of spermatozoa beside standard analysis and taken together with electron microscopy may help to determine the actual number of "healthy" spermatozoa thereby playing an important role during diagnosis and treatment of male infertility.

137Cs activity concentration in wild boar meat may still exceed the permitted levels

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Rachubik J.

2012-04-01

Full Text Available The radiocaesium activity concentration may still remain high in natural products such as game meat, wild mushrooms, and forest berries even more than two decades after the Chernobyl accident. The results of regular control studies of game meat conducted in Poland showed wild boars as the most contaminated game animals. It is well documented that some mushrooms, readily consumed by animals, show high ability to accumulate caesium radioisotopes. Bay bolete, one of the most wide-spread mushroom species in Poland, reveals a unique radiocaesium accumulation feature. Moreover, deer truffle, containing also particularly high levels of radiocaesium, could be another radionu-clide source for wild boars. Furthermore, animals consuming deer truffles could digest contaminated soil components. Among 94 wild boar meat samples analysed in 2008–2009, two exceeded the permitted level. Hence, some precautions should be taken in the population with an elevated intake of wild boar meat. Moreover, since each hunted wild boar is examined for the presence of Trichinella larvae, regular measurements of radiocaesium concentrations in these animals may be advisable for enhancing consumer safety.

137Cs activity concentration in wild boar meat may still exceed the permitted levels

Science.gov (United States)

Rachubik, J.

2012-04-01

The radiocaesium activity concentration may still remain high in natural products such as game meat, wild mushrooms, and forest berries even more than two decades after the Chernobyl accident. The results of regular control studies of game meat conducted in Poland showed wild boars as the most contaminated game animals. It is well documented that some mushrooms, readily consumed by animals, show high ability to accumulate caesium radioisotopes. Bay bolete, one of the most wide-spread mushroom species in Poland, reveals a unique radiocaesium accumulation feature. Moreover, deer truffle, containing also particularly high levels of radiocaesium, could be another radionu-clide source for wild boars. Furthermore, animals consuming deer truffles could digest contaminated soil components. Among 94 wild boar meat samples analysed in 2008-2009, two exceeded the permitted level. Hence, some precautions should be taken in the population with an elevated intake of wild boar meat. Moreover, since each hunted wild boar is examined for the presence of Trichinella larvae, regular measurements of radiocaesium concentrations in these animals may be advisable for enhancing consumer safety.

Fertilisation rate obtained with frozen-thawed boar semen supplemented with rosmarinic acid using a single insemination timed according to vulvar skin temperature changes.

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Luño, Victoria; Gil, Lydia; Olaciregui, Maite; Grandía, Juan; Ansó, Trinidad; De Blas, Ignacio

2015-03-01

Artificial insemination (AI) of sows with frozen-thawed semen usually results in lower pregnancy rates and litter sizes than the use of liquid preserved semen. The present study evaluated the effectiveness of vulvar skin temperature changes as a predictor of ovulation in sows and determined the fertility rates obtained after AI with frozen-thawed semen supplemented with rosmarinic acid (RA). Semen was collected from mature boars and cryopreserved in experimental extenders supplemented with or without 105 μM of RA. Multiparous sows were inseminated with a single dose of semen when vulvar skin temperature decreased to a value below 35 °C. Intrauterine insemination was performed using 1.5 × 109 spermatozoa. The sows were slaughtered 48 h after AI and the embryos and oocytes were recovered from the oviducts. Total and progressive motility, viability and acrosome integrity were significantly (P semen samples compared with the control. Fertilisation occurred in all sows inseminated in the study, although there were no significant differences between the experimental groups. Sows inseminated with RA-supplemented semen showed a slight increase in the number of embryos recovered as compared to sows inseminated with control semen. In conclusion, insemination according to vulvar skin temperature changes resulted in successful fertilisation in all sows, although supplementation of the freezing media with RA did not improve the fertilising ability of frozen-thawed boar sperm.

Identification of Wild Boar-Habitat Epidemiologic Cycle in African Swine Fever Epizootic.

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Chenais, Erika; Ståhl, Karl; Guberti, Vittorio; Depner, Klaus

2018-04-01

The African swine fever epizootic in central and eastern European Union member states has a newly identified component involving virus transmission by wild boar and virus survival in the environment. Insights led to an update of the 3 accepted African swine fever transmission models to include a fourth cycle: wild boar-habitat.

Sperm competition and reproductive mode influence sperm dimensions and structure among snakes.

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Tourmente, Maximiliano; Gomendio, Montserrat; Roldan, Eduardo R S; Giojalas, Laura C; Chiaraviglio, Margarita

2009-10-01

The role of sperm competition in increasing sperm length is a controversial issue, because findings from different taxa seem contradictory. We present a comparative study of 25 species of snakes with different levels of sperm competition to test whether it influences the size and structure of different sperm components. We show that, as levels of sperm competition increase, so does sperm length, and that this elongation is largely explained by increases in midpiece length. In snakes, the midpiece is comparatively large and it contains structures, which in other taxa are present in the rest of the flagellum, suggesting that it may integrate some of its functions. Thus, increases in sperm midpiece size would result in more energy as well as greater propulsion force. Sperm competition also increases the area occupied by the fibrous sheath and outer dense fibers within the sperm midpiece, revealing for the first time an effect upon structural elements within the sperm. Finally, differences in male-male encounter rates between oviparous and viviparous species seem to lead to differences in levels of sperm competition. We conclude that the influence of sperm competition upon different sperm components varies between taxa, because their structure and function is different.

Association of sperm apoptosis and DNA ploidy with sperm chromatin quality in human spermatozoa.

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Mahfouz, Reda Z; Sharma, Rakesh K; Said, Tamer M; Erenpreiss, Juris; Agarwal, Ashok

2009-04-01

To examine the relationship among sperm apoptosis, sperm chromatin status, and DNA ploidy in different sperm fractions. Prospective study. Reproductive research center in a tertiary care hospital. Sperm prepared by density gradient were evaluated for sperm count, motility, apoptosis, and sperm chromatin assessment. Sperm count, sperm motility, toluidine blue (TB) results, DNA fragmentation index (%DFI), high DNA stainability, DNA cytometry, and early and late apoptosis. Sperm motility was related to late apoptotic and subhaploid apoptotic sperm (r = -0.56 and -0.53, respectively). The sperm %DFI showed significant correlation with late apoptotic and subhaploid sperm (r = 0.62 and 0.68). TB-stained sperm were significantly correlated with late apoptotic sperm (r = 0.51). Significantly higher proportions of haploid sperm and light blue TB-stained sperm were seen in mature compared with immature fractions. Even in semen samples with low %DFI, semen processing results in a lower incidence of nuclear immaturity and subhaploidy, but the incidence of late apoptotic sperm remains unchanged. Therefore, simultaneous evaluation of apoptosis and sperm chromatin status is important for processing sperm in assisted reproductive procedures.

Rapid evaporative ionization mass spectrometry for high-throughput screening in food analysis: The case of boar taint.

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Verplanken, Kaat; Stead, Sara; Jandova, Renata; Poucke, Christof Van; Claereboudt, Jan; Bussche, Julie Vanden; Saeger, Sarah De; Takats, Zoltan; Wauters, Jella; Vanhaecke, Lynn

2017-07-01

Boar taint is a contemporary off-odor present in meat of uncastrated male pigs. As European Member States intend to abandon surgical castration of pigs by 2018, this off-odor has gained a lot of research interest. In this study, rapid evaporative ionization mass spectrometry (REIMS) was explored for the rapid detection of boar taint in neck fat. Untargeted screening of samples (n=150) enabled discrimination between sow, tainted and untainted boars. The obtained OPLS-DA models showed excellent classification accuracy, i.e. 99% and 100% for sow and boar samples or solely boar samples, respectively. Furthermore, the obtained models demonstrated excellent validation characteristics (R 2 (Y)=0.872-0.969; Q 2 (Y)=0.756-0.917), which were confirmed by CV-ANOVA (phighly accurate and high-throughput (<10s) classification of tainted and untainted boar samples was achieved, rendering REIMS a promising technique for predictive modelling in food safety and quality applications. Copyright © 2017 Elsevier B.V. All rights reserved.

Effect of Mucuna pruriens on oxidative stress mediated damage in aged rat sperm.

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Suresh, Sekar; Prithiviraj, Elumalai; Prakash, Seppan

2010-02-01

Mucuna pruriens Linn., a leguminous plant, has been recognized as an aphrodisiac and spermatogenic agent. Protective efficacy of M. pruriens on reactive oxygen species (ROS)-induced pathophysiological alterations in structural and functional integrity of epididymal sperm in aged Wister albino rat was analysed. Animals were grouped as groups I, II, III and IV, i.e. young (control), aged, aged treated with ethanolic extract (200 mg/kg b.w.) of M. pruriens and young rats treated with M. pruriens, respectively. At the end of the experimental period, i.e. after 60 days animals were sacrificed, epididymal sperm were collected and subjected to count, viability, motility, morphology and morphometric analysis. Enzymatic and non-enzymatic antioxidants, ROS, lipid peroxidation (LPO), DNA damage, chromosomal integrity and mitochondrial membrane potential were estimated. Results obtained from the aged animals showed significant reduction in sperm count, viability and motility, increased morphological damage and an increase in the number of sperm with cytoplasmic remnant, and these alterations were significantly reversed in M. pruriens treated group. Significant increase in LPO, HO and H(2)O(2) production and significant decline in the levels of the enzymatic and non-enzymatic antioxidants were observed in the aged animals. Supplementation of M. pruriens significantly reduced ROS and LPO production and significant increase in both enzymatic and non-enzymatic antioxidant levels. There were significant DNA damage, loss of chromosomal integrity and increase in mitochondrial membrane permeability in aged rat sperm. This was significantly reduced in group III. Present observation indicates the antioxidant enhancing property, free radical quenching ability and spermatogenic efficacy of the M. pruriens. Collectively, sperm damage in ageing was significantly reduced by quenching ROS, improving antioxidant defence system and mitochondrial function.

Subversive practices of sperm donation - globalizing Danish sperm

DEFF Research Database (Denmark)

Willum Adrian, Stine

as the use of donated sperm continuously has been debated as an ethical issue, and increasingly been regulated. In this presentation I will discuss how Denmark became a destination for fertility travelling (sperm donation) as a result of various subversive strategies of family making. The article inquires......-sited ethnography drawing on ethnographic research including observations and interviews from fertility clinics and sperm banks in Denmark during 2002/2003 and 2011- 2013, legislative documents and websites of fertility clinics and sperm banks. The presentation is methodologically inspired by Adele Clarke...

First isolation of Trichinella britovi from a wild boar (Sus scrofa) in Belgium.

NARCIS (Netherlands)

Schynts, F; Giessen, Joke van der; Tixhon, S; Pozio, E; Dorny, P; Borchgrave, J de

2006-01-01

Since 1992, when the European Union Council Directive requires that wild boars (Sus scrofa) hunted in EU for commercial purpose should be examined for Trichinella, the infection has not been detected in wild boars from Belgium, despite serological evidence of the presence of anti-Trichinella

Sorbitol Can Fuel Mouse Sperm Motility and Protein Tyrosine Phosphorylation via Sorbitol Dehydrogenase1

Science.gov (United States)

Cao, Wenlei; Aghajanian, Haig K.; Haig-Ladewig, Lisa A.; Gerton, George L.

2008-01-01

Energy sources that can be metabolized to yield ATP are essential for normal sperm functions such as motility. Two major monosaccharides, sorbitol and fructose, are present in semen. Furthermore, sorbitol dehydrogenase (SORD) can convert sorbitol to fructose, which can then be metabolized via the glycolytic pathway in sperm to make ATP. Here we characterize Sord mRNA and SORD expression during mouse spermatogenesis and examine the ability of sorbitol to support epididymal sperm motility and tyrosine phosphorylation. Sord mRNA levels increased during the course of spermatogenic differentiation. SORD protein, however, was first detected at the condensing spermatid stage. By indirect immunofluorescence, SORD was present along the length of the flagella of caudal epididymal sperm. Furthermore, immunoelectron microscopy showed that SORD was associated with mitochondria and the plasma membranes of sperm. Sperm incubated with sorbitol maintained motility, indicating that sorbitol was utilized as an energy source. Sorbitol, as well as glucose and fructose, were not essential to induce hyperactive motility. Protein tyrosine phosphorylation increased in a similar manner when sorbitol was substituted for glucose in the incubation medium used for sperm capacitation. These results indicate that sorbitol can serve as an alternative energy source for sperm motility and protein tyrosine phosphorylation. PMID:18799757

Aqueous extract of Telfairia occidentalis leaves reduces blood sugar ...

African Journals Online (AJOL)

STORAGESEVER

2008-07-18

Jul 18, 2008 ... white blood cell counts), sperm parameters (sperm motility, viability and counts) and blood glucose were determined. ... cantly increased red blood cell count, white blood cell count, packed cell volume and .... improve sperm motility and fertility in smokers (Dawson et al., 1992) and boar (Ivos et al., 1971), ...

A Sperm Spawn-Inducing Pheromone in the Silver Lip Pearl Oyster (Pinctada maxima).

Science.gov (United States)

Taylor, A; Mills, D; Wang, T; Ntalamagka, N; Cummins, S F; Elizur, A

2018-04-28

Pheromones are considered to play an important role in broadcast spawning in aquatic animals, facilitating synchronous release of gametes. In oysters, the sperm has been implicated as a carrier for the spawn-inducing pheromone (SIP). In hatchery conditions, male pearl oysters (Pinctata maxima) can be stimulated to spawn through a variety of approaches (e.g. rapid temperature change), while females can only be induced to spawn through exposure to conspecific sperm, thus limiting development of targeted pairing, required for genetic research and management. The capacity for commercial production and improvement of genetic lines of pearl oysters could be greatly improved with access to a SIP. In this study, we prepared and sequenced crude and semi-purified P. maxima sperm extracts that were used in bioassays to localise the female SIP. We report that the P. maxima SIP is proteinaceous and extrinsically associated with the sperm membrane. Bioactivity from pooled RP-HPLC fractions, but not individual fractions, suggests that the SIP is multi-component. We conclude that crude sperm preparations, as described in this study, can be used as a sperm-free inducer of female P. maxima spawning, which enables for a more efficient approach to genetic breeding.

Seroepidemiologic survey for Chlamydia suis in wild boar (Sus scrofa) populations in Italy.

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Di Francesco, Antonietta; Donati, Manuela; Morandi, Federico; Renzi, Maria; Masia, Marco Antonio; Ostanello, Fabio; Salvatore, Daniela; Cevenini, Roberto; Baldelli, Raffaella

2011-07-01

We used serology to estimate the prevalence of exposure to chlamydiae in Italian populations of wild boars (Sus scrofa). Sera from 173 hunter-killed wild boars harvested during the 2006-2009 hunting seasons in three Italian regions were tested for antibodies to Chlamydia suis, Chlamydophila pecorum, Chlamydophila abortus, and Chlamydophila psittaci by the microimmunofluorescence test. Antibody titers to chlamydiae ≥ 1:32 were detected in 110 of the 173 samples tested (63.6%). Specific reactivity could be assessed only in 44 sera with antibody titers to C. suis that were two- to threefold higher than antibody titers against the other chlamydial species; the other 66 sera had similar reactivity against all the chlamydia species tested. Antibody to C. suis was detected in sera from wild boar populations with rare or no known contact with domestic pigs. These results suggest that the wild boar could be a chlamydia reservoir and may acquire chlamydiae independent of contacts with the domestic pig.

Assessment of Domestic Pigs, Wild Boars and Feral Hybrid Pigs as Reservoirs of Hepatitis E Virus in Corsica, France

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Ferran Jori

2016-08-01

Full Text Available In Corsica, extensive pig breeding systems allow frequent interactions between wild boars and domestic pigs, which are suspected to act as reservoirs of several zoonotic diseases including hepatitis E virus (HEV. In this context, 370 sera and 166 liver samples were collected from phenotypically characterized as pure or hybrid wild boars, between 2009 and 2012. In addition, serum and liver from 208 domestic pigs belonging to 30 farms were collected at the abattoir during the end of 2013. Anti-HEV antibodies were detected in 26% (21%–31.6% of the pure wild boar, 43.5% (31%–56.7% of hybrid wild boar and 88% (82.6%–91.9% of the domestic pig sera. In addition, HEV RNA was detected in five wild boars, three hybrid wild boars and two domestic pig livers tested. Our findings provide evidence that both domestic pig and wild boar (pure and hybrid act as reservoirs of HEV in Corsica, representing an important zoonotic risk for Corsican hunters and farmers but also for the large population of consumers of raw pig liver specialties produced in Corsica. In addition, hybrid wild boars seem to play an important ecological role in the dissemination of HEV between domestic pig and wild boar populations, unnoticed to date, that deserves further investigation.

Wild boar mapping using population-density statistics: From polygons to high resolution raster maps.

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Pittiglio, Claudia; Khomenko, Sergei; Beltran-Alcrudo, Daniel

2018-01-01

The wild boar is an important crop raider as well as a reservoir and agent of spread of swine diseases. Due to increasing densities and expanding ranges worldwide, the related economic losses in livestock and agricultural sectors are significant and on the rise. Its management and control would strongly benefit from accurate and detailed spatial information on species distribution and abundance, which are often available only for small areas. Data are commonly available at aggregated administrative units with little or no information about the distribution of the species within the unit. In this paper, a four-step geostatistical downscaling approach is presented and used to disaggregate wild boar population density statistics from administrative units of different shape and size (polygons) to 5 km resolution raster maps by incorporating auxiliary fine scale environmental variables. 1) First a stratification method was used to define homogeneous bioclimatic regions for the analysis; 2) Under a geostatistical framework, the wild boar densities at administrative units, i.e. subnational areas, were decomposed into trend and residual components for each bioclimatic region. Quantitative relationships between wild boar data and environmental variables were estimated through multiple regression and used to derive trend components at 5 km spatial resolution. Next, the residual components (i.e., the differences between the trend components and the original wild boar data at administrative units) were downscaled at 5 km resolution using area-to-point kriging. The trend and residual components obtained at 5 km resolution were finally added to generate fine scale wild boar estimates for each bioclimatic region. 3) These maps were then mosaicked to produce a final output map of predicted wild boar densities across most of Eurasia. 4) Model accuracy was assessed at each different step using input as well as independent data. We discuss advantages and limits of the method and its

Cryopreservation of boar semen in mini- and maxi-straws.

Science.gov (United States)

Bwanga, C O; de Braganca, M M; Einarsson, S; Rodriguez-Martinez, H

1990-10-01

Split ejaculates from four boars were frozen with a programmable freezing machine, in mini- (0.25 ml) and maxi- (5 ml) plastic straws with an extender at either acidic (6.3) or alkaline (7.4) pH. Glycerol (3%) was used as cryoprotectant. The freezing of the semen was monitored by way of thermocouples placed in the straws. Post-thaw motility and acrosome integrity were evaluated; the latter using phase contrast microscopy, eosin-nigrosin stain and electron microscopy. Post-thaw sperm motility was significantly higher when semen was frozen in mini-straws than in maxi-straws. For the mini-straws, the motility was better when semen was exposed to an acidic environment during freezing, but this beneficial effect of the low extracellular pH was not evident when maxi-straws were thawed. The motility of the spermatozoa diminished significantly during the thermoresistance test (0 h and 2 h time) at 37 degrees C in a similar way for both straws and extracellular pH's. The freezing procedure, no matter the extracellular pH, did not cause major acrosomal damages, but significantly more normal apical ridges were present in the mini-straws than in the maxi-straws. This in vitro evaluation indicated that the freezing method employed was better for mini- than for maxi-straws since the freezing of the 5 ml volumes was not homogeneous, due to the large section area between the surface and the core of the straw.

Reproductive and genetic effects of continuous prenatal irradiation in the pig

International Nuclear Information System (INIS)

Erickson, B.H.; Martin, P.G.

1984-01-01

The stem germ cells of the prenatal pig are highly vulnerable to the cytotoxic effects of ionizing irradiation. This study was conducted to determine whether sensitivity to killing was also marked by a sensitivity to mutation and how prenatal depletion of the germ-cell population affects reproductive performance. Germ-cell populations were reduced by continuously irradiating sows at dose rates of either 0.25 or 1.0 rad/day for the first 108 days of gestation. The prenatally irradiated boars were tested for sperm-producing ability, sperm abnormalities, dominant lethality, reciprocal translocations, and fertility. Prenatally irradiated females were allowed to bear and nurture one litter, then tested for dominant lethality in a second litter; germ cell survival and follicular development were assessed in their serially sectioned ovaries. Sperm production was not significantly affected in the 0.25-rad boars, but boars irradiated with 1.0 rad per day produced sperm at only 17% of the control level. Incidence of defective sperm was 4.9% and 11.1% in the 0.25 and 1.0 groups, respectively. Four of the 1.0-rad boars were infertile, but prenatal irradiation apparently caused neither dominant lethality nor reciprocal translocations in fertile males. Number of oocytes was reduced to 66 +/- 7% of control in the 0.25-rad gilts, but reproductive performance was unaffected and no dominant lethality was observed. Only 7 +/- 1% of the oocytes survived in the 1.0-rad group. Reproductive performance was normal for the first litter, but four of the 23 sows tested were infertile at the second litter and a significant incidence of dominant lethality was observed

Artificial insemination in pigs today.

Science.gov (United States)

Knox, R V

2016-01-01

Use of artificial insemination (AI) for breeding pigs has been instrumental for facilitating global improvements in fertility, genetics, labor, and herd health. The establishment of AI centers for management of boars and production of semen has allowed for selection of boars for fertility and sperm production using in vitro and in vivo measures. Today, boars can be managed for production of 20 to 40 traditional AI doses containing 2.5 to 3.0 billion motile sperm in 75 to 100 mL of extender or 40 to 60 doses with 1.5 to 2.0 billion sperm in similar or reduced volumes for use in cervical or intrauterine AI. Regardless of the sperm dose, in liquid form, extenders are designed to sustain sperm fertility for 3 to 7 days. On farm, AI is the predominant form for commercial sow breeding and relies on manual detection of estrus with sows receiving two cervical or two intrauterine inseminations of the traditional or low sperm doses on each day detected in standing estrus. New approaches for increasing rates of genetic improvement through use of AI are aimed at methods to continue to lower the number of sperm in an AI dose and reducing the number of inseminations through use of a single, fixed-time AI after ovulation induction. Both approaches allow greater selection pressure for economically important swine traits in the sires and help extend the genetic advantages through AI on to more production farms. Copyright © 2016 Elsevier Inc. All rights reserved.

Systems genomics study reveals expression quantitative trait loci, regulator genes and pathways associated with boar taint in pigs

DEFF Research Database (Denmark)

Drag, Markus; Hansen, Mathias B.; Kadarmideen, Haja N.

2018-01-01

Boar taint is an offensive odour and/or taste from a proportion of non-castrated male pigs caused by skatole and androstenone accumulation during sexual maturity. Castration is widely used to avoid boar taint but is currently under debate because of animal welfare concerns. This study aimed...... to identify expression quantitative trait loci (eQTLs) with potential effects on boar taint compounds to improve breeding possibilities for reduced boar taint. Danish Landrace male boars with low, medium and high genetic merit for skatole and human nose score (HNS) were slaughtered at similar to 100 kg. Gene...... monitoring of other overlapping QTL traits should be performed to avoid any negative consequences of selection....

The N-terminal of a heparin-binding sperm membrane mitogen possess lectin-like sequence

International Nuclear Information System (INIS)

Mor, Visesato; Chatterjee, Tapati

2007-01-01

Glycosaminoglycans like heparin and heparin sulfate in follicular fluid induce changes in the intracellular environment during the spermatozoal functional maturation. We previously reported the isolation, purification and partial characterization of a heparin binding sperm membrane protein (HBSM). In the present study, the amino acids analysis provided evidence of a single sequence, which suggest the homogeneity of the purified HBSM. Fourteen amino acids- 1 A D T I V A V E L D T Y P N 14 -correspond to the amino terminal sequence of Concanavalin A (Con A) and contain 45.2% carbohydrate by weight. HBSM possess mitogenic property on lymphocytes with comparable magnitude to the well-known mitogen; Con A, inducing 83% radiolabel thymidine incorporation in growing lymphocytes. Unlike Con A, there was no agglutination of cell by HBSM upto 5 ng/ml concentration. Interestingly, we found that heparin and chondroitin sulfate-conjugated HBSM inhibit the proliferative activity. Similar effect was also found with an in-house isolate sulfated glycans; G-I (28% sulfate). In contrast, there was no inhibition by the desulfated form; G-ID. Altogether, our data suggest that the mechanism of cell proliferative pathway may be different for HBSM and Con A

What use is an infertile sperm? A comparative study of sperm-heteromorphic Drosophila

DEFF Research Database (Denmark)

Holman, Luke; Freckleton, Robert P; Snook, Rhonda R

2007-01-01

Sperm size and number are important determinants of male reproductive success. The genus Drosophila exhibits a remarkable diversity of sperm production strategies, including the production of multiple sperm morphs by individual males, a phenomenon called sperm heteromorphism. Sperm-heteromorphic ......Sperm size and number are important determinants of male reproductive success. The genus Drosophila exhibits a remarkable diversity of sperm production strategies, including the production of multiple sperm morphs by individual males, a phenomenon called sperm heteromorphism. Sperm......-heteromorphic Drosophila species in the obscura group produce large numbers of infertile "parasperm" in addition to fertile eusperm. Parasperm have been hypothesized to perform a number of roles in place of fertilization, predominantly focused on their potential function in postcopulatory sexual selection. However...

Yersinia enterocolitica Isolates from Wild Boars Hunted in Lower Saxony, Germany.

Science.gov (United States)

von Altrock, Alexandra; Seinige, Diana; Kehrenberg, Corinna

2015-07-01

Yersiniosis is strongly associated with the consumption of pork contaminated with enteropathogenic Yersinia enterocolitica, which is harbored by domestic pigs without showing clinical signs of disease. In contrast to data on Y. enterocolitica isolated from conventionally reared swine, investigations into the occurrence of Y. enterocolitica in wild boars in Germany are rare. The objectives of the study were to get knowledge about these bacteria and their occurrence in wild boars hunted in northern Germany by isolation of the bacteria from the tonsils, identification of the bioserotypes, determination of selected virulence factors, macrorestriction analysis, multilocus sequence typing (MLST), and testing of antimicrobial susceptibility. Altogether, tonsils from 17.1% of 111 tested wild boars were positive for Y. enterocolitica by culture methods. All but two isolates belonged to biotype (BT) 1A, with the majority of isolates bearing a ystB nucleotide sequence which was revealed to have 85% identity to internal regions of Y. enterocolitica heat-stable enterotoxin type B genes. The remaining Y. enterocolitica isolates were identified to be BT 1B and did not carry the virulence plasmid. However, two BT 1A isolates carried the ail gene. Macrorestriction analysis and results from MLST showed a high degree of genetic diversity of the isolates, although the region where the samples were taken was restricted to Lower Saxony, Germany, and wild boars were shot during one hunting season. In conclusion, most Y. enterocolitica isolates from wild boars investigated in this study belonged to biotype 1A. Enteropathogenic Y. enterocolitica bioserotypes 4/O:3 and 2/O:9, usually harbored by commercially raised pigs in Europe, could not be identified. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Secrets of Success in a Landscape of Fear: Urban Wild Boar Adjust Risk Perception and Tolerate Disturbance

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Milena Stillfried

2017-12-01

Full Text Available In urban areas with a high level of human disturbance, wildlife has to adjust its behavior to deal with the so called “landscape of fear.” This can be studied in risk perception during movement in relation to specific habitat types, whereby individuals trade-off between foraging and disturbance. Due to its high behavioral plasticity and increasing occurrence in urban environments, wild boar (Sus scrofa is an excellent model organism to study adjustment to urbanization. With the help of GPS tracking, space use of 11 wild boar from Berlin's metropolitan region was analyzed: we aimed at understanding how animals adjust space use to deal with the landscape of fear in urban areas compared to rural areas. We compared use vs. availability with help of generalized linear mixed models. First, we studied landscape types selected by rural vs. urban wild boar, second, we analyzed distances of wild boar locations to each of the landscape types. Finally, we mapped the resulting habitat selection probability to predict hotspots of human-wildlife conflicts. A higher tolerance to disturbance in urban wild boar was shown by a one third shorter flight distance and by an increased re-use of areas close to the trap. Urban wild boar had a strong preference for natural landscapes such as swamp areas, green areas and deciduous forests, and areas with high primary productivity, as indicated by high NDVI (normalized difference vegetation index values. The areas selected by urban wild boar were often located closely to roads and houses. The spatial distribution maps show that a large area of Berlin would be suitable for urban wild boar but not their rural conspecifics, with the most likely reason being a different perception of anthropogenic disturbance. Wild boar therefore showed considerable behavioral plasticity suitable to adjust to human-dominated environments in a potentially evolutionarily adaptive manner.

Preliminary study on the detection of hepatitis E virus (HEV antibodies in pigs and wild boars in Poland

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Weiner Marcin

2016-12-01

Full Text Available Introduction: Although HEV infection in pigs does not pose a major economic risk to pork production, the risk of zoonotic transmission to humans is an important aspect of public health. HEV genotype 3 infections were reported in developed countries in individuals who had consumed raw meat or meat products from deer, wild boars, or pigs. The aim of the study was the analysis of the occurrence of HEV-specific antibodies among wild boars and domestic pigs in Poland. Material and Methods: A total of 290 samples from wild boars and 143 samples from pigs were tested. The antibodies were tested by ELISA. Results: The presence of anti-HEV IgG was demonstrated in 44.1% of pigs and 31.0% of wild boars. Anti-HEV IgG antibodies were detected in 1.4% of samples from pigs and in 2.1% of samples from wild boars at borderline level. The statistical analysis shows significant differences in the positive results for anti-HEV IgG between the groups of pigs and wild boars (P = 0.0263. Conclusion: Regular surveillance of the occurrence of HEV in swine and wild boars should be performed in the future.

Wild Boar Impact to the Natural Regeneration of Oak and Acorn Importance in its Diet

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Jaroslav Zeman

2016-01-01

Full Text Available In this study, the impact of wild boar on the natural regeneration of oak and importance of acorns in the wild boar diet were surveyed. The data were collected near Moravian Krumlov (Czech Republic in three types of oak stands differing in the canopy density: fully-stocked stand (1, open-canopy stand (2 and the forest stand in a conversion to the coppice forest (3. Within each stand 150 m long lines were set out. Seed traps to collect acorn harvest and control plots were installed on these lines. The plots were inspected at weekly intervals. After the end of acorn fall the average amount of fallen acorns was evaluated. The quantity of metabolizable energy in acorns was assessed and daily survival dose of energy for average weight of wild boar was counted. In spring 2014 the number of seedlings was counted at the same plots. Production of acorns per hectare and basic energy needs for one individual wild boar per day were evaluated for each chosen stand type. The found seedling density amounted to 29,600, 32,000 and 14,000 ind./ha in the first, second and third stand under study, respectively. Wild boar is a major consumer of acorns. In the studied area the production of acorns sufficiently supplied the local wild boar population during winter. Necessary amount of acorns remained for the natural regeneration.

The effect of birth weight of boars and litter size in which were 1 ...

African Journals Online (AJOL)

Eugenia

2017-05-22

May 22, 2017 ... 1.15 kg, 70 boars) and greater birth weight (GBW) (mean: 1.59 kg, 70 boars). Testes volume .... The data were subjected to analysis of variance (ANOVA) using the MIXED procedure. The model for all ..... Frontiers. 3,. 62-67.

The association between disease and profitability in individual finishing boars at a test station

DEFF Research Database (Denmark)

Jensen, Tina Birk; Baadsgaard, Niels Peter; Houe, Hans

2008-01-01

Endemic diseases in finisher herds are considered to be costly for the pig producer. We investigated the effect of diseases on the profit margin using data from a Danish boar test station (n = 5777) collected from July 2002 to December 2004. Boars reaching a target slaughter weight of at least 80...... kg were included in the study. Oral and parenteral treatments were used as indicator of disease in the finishing period and, pathological lesions were used as indicator of disease at slaughter. Profit margin was calculated individually for each boar as the difference between the total revenue......: profit margin. The results showed that treatment in the finishing period had a negative effect on the profit margin. According to the least square means estimates, boars that were treated parenterally had a reduction in the profit margin of 2.24 €. This corresponded to a reduction in the profit margin...

Single-layer centrifugation through colloid selects improved quality of epididymal cat sperm.

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Chatdarong, K; Thuwanut, P; Morrell, J M

2010-06-01

The objectives were to determine the: 1) extent of epithelial and red blood cell contamination in epididymal cat sperm samples recovered by the cutting method; 2) efficacy of simple washing, single-layer centrifugation (SLC), and swim-up for selecting epididymal cat sperm; and 3) effects of freezing and thawing on cat sperm selected by various techniques. Ten unit samples were studied; each contained sperm from the cauda epididymides of four cats (total, approximately 200 x 10(6) sperm) and was equally allocated into four treatments: 1) simple washing, 2) single-layer centrifugation through colloid prior to cryopreservation (SLC-PC), 3) single-layer centrifugation through colloid after cryopreservation (SLC-AC), and 4) swim-up. Centrifugation (300 x g for 20 min) was done for all methods. The SLC-PC had a better recovery rate than the SLC-AC and swim-up methods (mean+/-SD of 16.4+/-8.7, 10.7+/-8.9, and 2.3+/-1.7%, respectively; Pblood cell contamination than simple washed samples (0.02+/-0.01, 0.02+/-0.04, 0.03+/-0.04, and 0.44+/-0.22 x 10(6) cells/mL, respectively; P0.05), SLC-PC yielded the highest percentage of sperm with normal midpieces and tails (P0.05). In conclusion, both SLC-PC and swim-up improved the quality of epididymal cat sperm, including better morphology, membrane and DNA integrity, and removal of cellular contamination. However, SLC had a better sperm recovery rate than swim-up. 2010 Elsevier Inc. All rights reserved.

Penambahan Astaxanthin pada Pengencer Kuning Telur Berbagai Jenis Unggas Dapat Memproteksi Semen Babi Selama Penyimpanan (THE ADDITION OF ASTAXANTHIN ON SPERM DILUENTS PHOSPHATE EGGYOLK OF VARIOUS POULTRY CAN PROTECT QUALITY OF PIG SPERM DURING STORAGE

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Wayan Bebas

2017-01-01

Full Text Available A study was conducted to formulate semen diluent of pigswith a better quality, cheap and easy toprepare using egg yolk of various poultries such as chickens, ducks and quails in combinationwithastaxanthin, a potent antioxidant. The research design used was a completely randomized factorialdesign with three different types of sperm diluents and four levels of astaxanthin concentration. Spermdiluents used were phosphate duck egg yolk, phosphate quail egg yolk and yolks phosphate supplementedrespectivelywith 0,002%, 0,004% and 0,008%astaxanthine. The treated sperm were strored at 5oC for 48hours. The sperm qualities were examined for progressive motility, spermatozoa abnormalities, viabilityand plasma membrane integrity. The result showed that sperm diluents of using duck egg yolk phosphatein combination with 0.002% astaxanthinresulted in the highest progressive motility, viability and plasmamembrane intact while abnormalities spermatozoa is lowest. It can be concluded that phosphate duck eggyolk sperm diluents with the addition of 0,002% astaxanthinappeared to be able to maintain the qualityof pig sperm stored at 5oC for 48 hours.

The Pattern of Tyrosine Phosphorylation in Human Sperm in Response to Binding to Zona Pellucida or Hyaluronic Acid

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Sati, Leyla; Cayli, Sevil; Delpiano, Elena; Sakkas, Denny

2014-01-01

In mammalian species, acquisition of sperm fertilization competence is dependent on the phenomenon of sperm capacitation. One of the key elements of capacitation is protein tyrosine phosphorylation (TP) in various sperm membrane regions. In previous studies performed, the pattern of TP was examined in human sperm bound to zona pellucida of oocytes. In the present comparative study, TP patterns upon sperm binding to the zona pellucida or hyaluronic acid (HA) were investigated in spermatozoa arising from the same semen samples. Tyrosine phosphorylation, visualized by immunofluorescence, was localized within the acrosomal cap, equatorial head region, neck, and the principal piece. Tyrosine phosphorylation has increased in a time-related manner as capacitation progressed, and the phosphorylation pattern was identical within the principal piece and neck, regardless of the sperm bound to the zona pellucida or HA. Thus, the data demonstrated that the patterns of sperm activation-related TP were similar regardless of the spermatozoa bound to zona pellucida or HA. Further, sperm with incomplete development, as detected by excess cytoplasmic retention, failed to exhibit TP. PMID:24077441

Involvement of complexin 2 in docking, locking and unlocking of different SNARE complexes during sperm capacitation and induced acrosomal exocytosis.

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Pei-Shiue J Tsai

Full Text Available Acrosomal exocytosis (AE is an intracellular multipoint fusion reaction of the sperm plasma membrane (PM with the outer acrosomal membrane (OAM. This unique exocytotic event enables the penetration of the sperm through the zona pellucida of the oocyte. We previously observed a stable docking of OAM to the PM brought about by the formation of the trans-SNARE complex (syntaxin 1B, SNAP 23 and VAMP 3. By using electron microscopy, immunochemistry and immunofluorescence techniques in combination with functional studies and proteomic approaches, we here demonstrate that calcium ionophore-induced AE results in the formation of unilamellar hybrid membrane vesicles containing a mixture of components originating from the two fused membranes. These mixed vesicles (MV do not contain the earlier reported trimeric SNARE complex but instead possess a novel trimeric SNARE complex that contained syntaxin 3, SNAP 23 and VAMP 2, with an additional SNARE interacting protein, complexin 2. Our data indicate that the earlier reported raft and capacitation-dependent docking phenomenon between the PM and OAM allows a specific rearrangement of molecules between the two docked membranes and is involved in (1 recruiting SNAREs and complexin 2 in the newly formed lipid-ordered microdomains, (2 the assembly of a fusion-driving SNARE complex which executes Ca(2+-dependent AE, (3 the disassembly of the earlier reported docking SNARE complex, (4 the recruitment of secondary zona binding proteins at the zona interacting sperm surface. The possibility to study separate and dynamic interactions between SNARE proteins, complexin and Ca(2+ which are all involved in AE make sperm an ideal model for studying exocytosis.

PREVALENCE AND RISK FACTORS ASSOCIATED WITH THE PRRS VIRUS IN SEMEN OF BOARS IN PIG FARMS OF YUCATAN

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Aremi Jordan-Craviotto

2009-07-01

Full Text Available The objectives of the present study were to estimate the prevalence of and to determine the risk factors associated with the porcine reproductive and respiratory syndrome virus (PRRSV, American strain in semen of boars in pig herds of Yucatan, Mexico. Ninety two boars from 26 herds were ejaculated once. Semen samples were processed by the RT-nPCR test using the ORF7 primer to detect the PRRS virus. The true prevalence estimated was 10.1% (95% CI = 4.1-16.1%. Significance of risk factors was determined by Fisher-exact test. The odds of detecting genetic material of the PRRSV was greater (OR = 9.2 in semen of boars used under natural mating than those used in artificial insemination. In herds where boar’s acclimatization was not practiced the odds of a positive boar was 4.3. Another risk factor (P < 0.05 was the origin of the animals. In conclusion, the prevalence of the PRRSV in boar semen was smaller to the notified in the literature and determinate in blood serum. Management practices, such as the use of the artificial insemination and acclimatization of the boar, could be useful in reducing the prevalence of the PRRS virus in the pig farms.

Zoonotic intestinal protozoan of the wild boars, Sus scrofa, in Persian Gulf's coastal area (Bushehr province), Southwestern Iran.

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Yaghoobi, Kambiz; Sarkari, Bahador; Mansouri, Majid; Motazedian, Mohammad Hossein

2016-10-01

Wild boars, Sus scrofa , are potential reservoirs of many zoonotic diseases, and there are a possibility of transmission of the zoonotic diseases from these animals to humans and also domestic animals. This study aimed to evaluate the protozoan contamination of wild boars in the Persian Gulf's coastal area (Bushehr Province), southwestern Iran. A total of 25 crossbred boars were collected during a course of vertebrate pest control in Bushehr province, in 2013. Samples were collected from the gastrointestinal tracts of each boar in 5% formalin, Bouin's solution, sodium acetate-acetic acid-formalin, and polyvinyl alcohol fixatives. Fixed stool smears examined by trichrome and Ziehl-Neelsen staining. Each of the 25 wild boars was infected with at least one of the intestinal protozoans. The rate of contamination with intestinal protozoan was 64% for Balantidium coli , 76% for Iodamoeba sp., 52% for Entamoeba polecki , 44% for Blastocystis sp. and 8% for Chilomastix sp. No intestinal coccidian was detected in studied boars when the stool samples were evaluated by Ziehl-Neelsen staining method. Findings of this study demonstrated that wild boars in the Persian Gulf coastal area are contaminated by many protozoans, including zoonotic protozoan, which poses a potential risk to locals as well as the domestic animals of the area.

PRESENCE OF TRICHINELLA BRITOVI IN WILD BOAR IN THE MARCHE REGION REGULARLY SLAUGHTERED

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L. Di Giacomo

2011-01-01

Full Text Available Trichinellosis is a zoonotic disease caused by a nematode belonging to the genus Trichinella. Numerous mammal species as well as birds and crocodiles can harbour the parasite worldwide, but the wild cycle is mainly maintained by wild carnivores. Human represents only a possible host and the parasite is exclusively transmitted through consumption of undercooked or raw meat. In Italy, pork, wild boar meat and horse meat are the main sources for human infection. This article describe a presence of Trichinella britovi in wild boar in the Marche region regularly slaughtered. These case confirms the occurrence of Trichinella britovi in wild boar and the important role of slaughterhouse in epidemiological surveillance of zoonotic diseases.

Why small males have big sperm: dimorphic squid sperm linked to alternative mating behaviours.

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Iwata, Yoko; Shaw, Paul; Fujiwara, Eiji; Shiba, Kogiku; Kakiuchi, Yasutaka; Hirohashi, Noritaka

2011-08-10

Sperm cells are the target of strong sexual selection that may drive changes in sperm structure and function to maximize fertilisation success. Sperm evolution is regarded to be one of the major consequences of sperm competition in polyandrous species, however it can also be driven by adaptation to the environmental conditions at the site of fertilization. Strong stabilizing selection limits intra-specific variation, and therefore polymorphism, among fertile sperm (eusperm). Here we analyzed reproductive morphology differences among males employing characteristic alternative mating behaviours, and so potentially different conditions of sperm competition and fertilization environment, in the squid Loligo bleekeri. Large consort males transfer smaller (average total length = 73 μm) sperm to a female's internal sperm storage location, inside the oviduct; whereas small sneaker males transfer larger (99 μm) sperm to an external location around the seminal receptacle near the mouth. No significant difference in swimming speed was observed between consort and sneaker sperm. Furthermore, sperm precedence in the seminal receptacle was not biased toward longer sperm, suggesting no evidence for large sperm being favoured in competition for space in the sperm storage organ among sneaker males. Here we report the first case, in the squid Loligo bleekeri, where distinctly dimorphic eusperm are produced by different sized males that employ alternative mating behaviours. Our results found no evidence that the distinct sperm dimorphism was driven by between- and within-tactic sperm competition. We propose that presence of alternative fertilization environments with distinct characteristics (i.e. internal or external), whether or not in combination with the effects of sperm competition, can drive the disruptive evolution of sperm size.

Why small males have big sperm: dimorphic squid sperm linked to alternative mating behaviours

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Shiba Kogiku

2011-08-01

Full Text Available Abstract Background Sperm cells are the target of strong sexual selection that may drive changes in sperm structure and function to maximize fertilisation success. Sperm evolution is regarded to be one of the major consequences of sperm competition in polyandrous species, however it can also be driven by adaptation to the environmental conditions at the site of fertilization. Strong stabilizing selection limits intra-specific variation, and therefore polymorphism, among fertile sperm (eusperm. Here we analyzed reproductive morphology differences among males employing characteristic alternative mating behaviours, and so potentially different conditions of sperm competition and fertilization environment, in the squid Loligo bleekeri. Results Large consort males transfer smaller (average total length = 73 μm sperm to a female's internal sperm storage location, inside the oviduct; whereas small sneaker males transfer larger (99 μm sperm to an external location around the seminal receptacle near the mouth. No significant difference in swimming speed was observed between consort and sneaker sperm. Furthermore, sperm precedence in the seminal receptacle was not biased toward longer sperm, suggesting no evidence for large sperm being favoured in competition for space in the sperm storage organ among sneaker males. Conclusions Here we report the first case, in the squid Loligo bleekeri, where distinctly dimorphic eusperm are produced by different sized males that employ alternative mating behaviours. Our results found no evidence that the distinct sperm dimorphism was driven by between- and within-tactic sperm competition. We propose that presence of alternative fertilization environments with distinct characteristics (i.e. internal or external, whether or not in combination with the effects of sperm competition, can drive the disruptive evolution of sperm size.

Boar taint detection using parasitoid biosensors

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To evaluate the potential for a non-stinging wasp to be used as a biosensor in the pig industry, we trained wasps to 3 individual chemicals associated with boar taint. Training consisted of presenting the odors to hungry wasps while they were feeding on sugar. This associates the chemical with a fo...

A genetic variant of the sperm-specific SLO3 K+ channel has altered pH and Ca2+ sensitivities.

Science.gov (United States)

Geng, Yanyan; Ferreira, Juan J; Dzikunu, Victor; Butler, Alice; Lybaert, Pascale; Yuan, Peng; Magleby, Karl L; Salkoff, Lawrence; Santi, Celia M

2017-05-26

To fertilize an oocyte, sperm must first undergo capacitation in which the sperm plasma membrane becomes hyperpolarized via activation of potassium (K + ) channels and resultant K + efflux. Sperm-specific SLO3 K + channels are responsible for these membrane potential changes critical for fertilization in mouse sperm, and they are only sensitive to pH i However, in human sperm, the major K + conductance is both Ca 2+ - and pH i -sensitive. It has been debated whether Ca 2+ -sensitive SLO1 channels substitute for human SLO3 (hSLO3) in human sperm or whether human SLO3 channels have acquired Ca 2+ sensitivity. Here we show that hSLO3 is rapidly evolving and reveal a natural structural variant with enhanced apparent Ca 2+ and pH sensitivities. This variant allele (C382R) alters an amino acid side chain at a principal interface between the intramembrane-gated pore and the cytoplasmic gating ring of the channel. Because the gating ring contains sensors to intracellular factors such as pH and Ca 2+ , the effectiveness of transduction between the gating ring and the pore domain appears to be enhanced. Our results suggest that sperm-specific genes can evolve rapidly and that natural genetic variation may have led to a SLO3 variant that differs from wild type in both pH and intracellular Ca 2+ sensitivities. Whether this physiological variation confers differences in fertility among males remains to be established. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Relationship between sperm parameters and intracytoplasmic sperm injection outcome

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Shahla Chaichian

2015-12-01

Full Text Available Objectives: With the adventure of intracytoplasmic sperm injection (ICSI technique, great progresses have developed in the treatment of infertility. Concentration on the properties of male’s gamete has been encouraged by the increasing concerns about the causes of ICSI failure. We hence conducted this study to investigate the probable association of sperm parameters with ISCI outcome. Methods: A total of 523 couples referred to Isfahan Fertility and Sterility Center from January 2007 to June 2008 for ICSI. Semen analysis was performed before ICSI procedure according to the WHO criteria. Patients were assigned into successful ICSI (case and failed ICSI (control groups. Sperm parameters were then compared between the 2 groups. Results: One hundred and six patients (20% had successful ICSI results (case group compared with 417 couples (80% with undesirable ICSI outcomes (control group. Among evaluated factors, sperm agglutination (p = 0.007, sperm concentration (p = 0.043, leukocytospermia (p = 0.026 and head abnormality of sperm (p = 0.019 showed statistically significant differences between two groups with differing ICSI results. None of the other semen parameters revealed significant differences between these two groups. Conclusion: Our study showed that some sperm parameters are associated with desirable ICSI outcome. However, it is unclear whether these associations are causal.

Thermodynamics of the interaction between bovine binder of sperm BSP1 and low-density lipoprotein from hen's egg yolk

Energy Technology Data Exchange (ETDEWEB)

Lusignan, Marie-France; Manjunath, Puttaswamy [Maisonneuve-Rosemont Hospital Research Center and Department of Medicine, Universite de Montreal, C.P. 6128, Succ. Centre Ville, Montreal, Quebec, Canada H3C 3J7 (Canada); Lafleur, Michel, E-mail: michel.lafleur@umontreal.ca [Department of Chemistry, Centre for Self-Assembled Chemical Structures (CSACS), Universite de Montreal, C.P. 6128, Succ. Centre Ville, Montreal, Quebec, Canada H3C 3J7 (Canada)

2011-03-20

Research highlights: {yields} Isothermal titration calorimetry revealed Binder-of-SPerm BSP1 protein has high affinity for egg yolk low density lipoproteins. {yields} It is estimated that 104 BSP1 molecules could bind one LDL particle. {yields} BSP1 has 2 phosphocholine binding sites and the BSP1/LDL ratio corresponds to 1.6 phosphatidylcholine per bound BSP1. {yields} The strong interaction between LDL particles and BSP1 is proposed to be important for sperm protection by egg yolk extender. - Abstract: Egg yolk is used in extender to protect sperm from cold shock and freezing during preservation. It is the low-density lipoprotein (LDL) fraction of egg yolk that protects sperm. Even though essential for sperm capacitation, the major proteins from bull seminal plasma, the Binder of SPerm (BSP) proteins, are detrimental for sperm preservation because they induce a continual phospholipids and cholesterol efflux from sperm membranes. The BSP proteins were proposed to bind to egg yolk LDL, preventing the sperm membrane damage. We characterized the binding between the BSP proteins and the LDL by isothermal titration calorimetry, providing the thermodynamics and quantitative description of this putative association. The association between BSP1 (major BSP proteins) and LDL is characterized by an affinity constant (K{sub a}) of 3.4 {+-} 0.4 {mu}M{sup -1}. A protein/LDL ratio of 104 {+-} 5 was determined indicating that 104 molecules of BSP1 would bind to one LDL particle. This stoichiometry leads to proposing that the association involves 1.6 {+-} 0.1 phosphatidylcholines (PC) per BSP protein. This finding is satisfactorily consistent with the fact that each BSP1 protein has 2 binding sites for choline group. In conclusion, the formation of a high affinity complex between BSP1 and LDL is proposed to be important for the protection of sperm by egg yolk extender.

Natural and experimental hepatitis E virus genotype 3-infection in European wild boar is transmissible to domestic pigs.

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Schlosser, Josephine; Eiden, Martin; Vina-Rodriguez, Ariel; Fast, Christine; Dremsek, Paul; Lange, Elke; Ulrich, Rainer G; Groschup, Martin H

2014-11-26

Hepatitis E virus (HEV) is the causative agent of acute hepatitis E in humans in developing countries, but sporadic and autochthonous cases do also occur in industrialised countries. In Europe, food-borne zoonotic transmission of genotype 3 (gt3) has been associated with domestic pig and wild boar. However, little is known about the course of HEV infection in European wild boar and their role in HEV transmission to domestic pigs. To investigate the transmissibility and pathogenesis of wild boar-derived HEVgt3, we inoculated four wild boar and four miniature pigs intravenously. Using quantitative real-time RT-PCR viral RNA was detected in serum, faeces and in liver, spleen and lymph nodes. The antibody response evolved after fourteen days post inoculation. Histopathological findings included mild to moderate lymphoplasmacytic hepatitis which was more prominent in wild boar than in miniature pigs. By immunohistochemical methods, viral antigens were detected mainly in Kupffer cells and liver sinusoidal endothelial cells, partially associated with hepatic lesions, but also in spleen and lymph nodes. While clinical symptoms were subtle and gross pathology was inconspicuous, increased liver enzyme levels in serum indicated hepatocellular injury. As the faecal-oral route is supposed to be the most likely transmission route, we included four contact animals to prove horizontal transmission. Interestingly, HEVgt3-infection was also detected in wild boar and miniature pigs kept in contact to intravenously inoculated wild boar. Given the high virus loads and long duration of viral shedding, wild boar has to be considered as an important HEV reservoir and transmission host in Europe.

Relationship between Porcine Sperm Motility and Sperm Enzymatic Activity using Paper-based Devices

Science.gov (United States)

Matsuura, Koji; Huang, Han-Wei; Chen, Ming-Cheng; Chen, Yu; Cheng, Chao-Min

2017-04-01

Mammalian sperm motility has traditionally been analyzed to determine fertility using computer-assisted semen analysis (CASA) systems. To develop low-cost and robust male fertility diagnostics, we created a paper-based MTT assay and used it to estimate motile sperm concentration. When porcine sperm motility was inhibited using sperm enzyme inhibitors for sperm enzymes related to mitochondrial activity and glycolysis, we simultaneously recorded sperm motility and enzymatic reactivity using a portable motility analysis system (iSperm) and a paper-based MTT assay, respectively. When using our paper-based MTT-assay, we calculated the area mean value signal intensity (AMV) to evaluate enzymatic reactivity. Both sperm motility and AMV decreased following treatment with iodoacetamide (IODO) and 3-bromopyruvic acid (3BP), both of which are inhibitors of glycolytic enzymes including glyceraldehyde-3-phosphate dehydrogenase (GAPDH). We found a correlation between recorded motility using iSperm and AMV from our paper-based assay (P Based on this inhibitor study, sperm motility can be estimated using our paper-based MTT-assay.

First isolation of Haemophilus parasuis and other NAD-dependent Pasteurellaceae of swine from European wild boars.

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Olvera, A; Cerdà-Cuéllar, M; Mentaberre, G; Casas-Diaz, E; Lavin, S; Marco, I; Aragon, V

2007-11-15

Haemophilus parasuis is a colonizer of the upper respiratory tract of pigs and the etiological agent of Glässer's disease, which is characterized by a fibrinous polyserositis, meningitis and arthritis. Glässer's disease has never been reported in wild boar (Sus scrofa), although antibodies against H. parasuis have been detected. The goal of this study was to confirm the presence of this bacterium in wild boar by bacterial isolation and to compare the strains to H. parasuis from domesticated pigs. Therefore, nasal swabs from 42 hunted wild boars were processed for bacterial isolation and subsequent H. parasuis identification by specific PCR, biochemical tests and 16S rRNA gene sequencing. Two different strains of H. parasuis from two wild boars were isolated. These strains belonged to serotype 2 and were included by 16S rRNA gene sequencing and MLST analysis in a cluster with other H. parasuis strains of nasal origin from domestic pigs. During this study, Actinobacillus minor and Actinobacillus indolicus, which are NAD-dependent Pasteurellaceae closely related to H. parasuis, were also isolated. Our results indicate similarities in the respiratory microbiota of wild boars and domestic pigs, and although H. parasuis was isolated from wild boars, more studies are needed to determine if this could be a source of H. parasuis infection for domestic pigs.

Evaluation of swine fertilisation medium (SFM) efficiency in preserving spermatozoa quality during long-term storage in comparison to four commercial swine extenders.

Science.gov (United States)

Fantinati, P; Zannoni, A; Bernardini, C; Forni, M; Tattini, A; Seren, E; Bacci, M L

2009-02-01

In pig production, artificial insemination is widely carried out and the use of fresh diluted semen is predominant. For this reason, there are increasing interests in developing new extenders and in establishing the optimal storage conditions for diluted spermatozoa. In the last few decades, we utilised a homemade diluent (swine fertilisation medium (SFM)) for spermatozoa manipulation and biotechnological application as the production of transgenic pigs utilising the sperm-mediated gene transfer technique. The purpose of the present study is therefore to analyse the ability of SFM, in comparison to four commercial extenders, in preserving the quality of diluted boar semen stored at 16.5°C till 15 days. We utilised some of the main predictive tests as objectively measured motility, acrosome and sperm membrane integrity, high mitochondrial membrane potential and pH. Based on our in vitro study, SFM could be declared as a good long-term extender, able to preserve spermatozoa quality as well as Androhep Enduraguard for up to 6 to 9 days and more.

Wild boars (Sus scrofa scrofa seminiferous tubules morphometry

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Deiler Sampaio Costa

2006-09-01

Full Text Available The aim of this data was to analyze morphology and function of the seminiferous tubule in adult wild boars. Testes removed by unilateral castration of five animals were used. The testicular parenchyma was composed by 82.1±2.2% of seminiferous tubule and 17.9±2.2% of intertubular tissue. The tubular diameter was 249.2±33.0 µm and the seminiferous tubule lenght per gram of testis was 19.3±4.9m. The spermatogonial mitoses efficiency coefficient, meiotic index and spermatogenesis efficiency were 10.34, 2.71 and 30.5 respectively. Each Sertoli cell supported about 13 germinatives cells. The hystometric parameters studied were very similar to those related for domestic boars, however, the wild boars intrinsic efficiency of spermatogenesis and Sertoli cells indexes were smaller than in domestic boars.Objetivou-se com esta pesquisa estudar as características morfométricas e funcionais dos túbulos seminíferos de javalis adultos. Utilizaram-se testículos de cinco animais submetidos a orquiectomia unilateral. O parênquima testicular foi composto por 82,1 ± 2,2% de túbulos seminíferos e 17,9 ± 2,2% de tecido intertubular. O diâmetro tubular foi de 249,2 ± 33,0µm e o comprimento dos túbulos seminíferos por grama de testículo foi de 19,3 ± 4,9m. O coeficiente de eficiência das mitoses espermatogônias, o rendimento meiótico e o rendimento geral da espermatogênese foram, respectivamente, 10,34, 2,71 e 30,50. Cada célula de Sértoli suportou cerca de 13 células germinativas. Conclui-se que os parâmetros histométricos estudados nesta pesquisa foram muito semelhantes aos valores relatados para suínos domésticos, entretanto, o rendimento intrínseco da espermatogênese e os índices de células de Sértoli de javalis foram relativamente baixos quando comparados com aqueles animais.

Systems genomics study reveals expression quantitative trait loci, regulator genes and pathways associated with boar taint in pigs

DEFF Research Database (Denmark)

Drag, Markus; Hansen, Mathias B.; Kadarmideen, Haja N.

2018-01-01

Boar taint is an offensive odour and/or taste from a proportion of non-castrated male pigs caused by skatole and androstenone accumulation during sexual maturity. Castration is widely used to avoid boar taint but is currently under debate because of animal welfare concerns. This study aimed...... to identify expression quantitative trait loci (eQTLs) with potential effects on boar taint compounds to improve breeding possibilities for reduced boar taint. Danish Landrace male boars with low, medium and high genetic merit for skatole and human nose score (HNS) were slaughtered at similar to 100 kg. Gene...... and SSC14. Functional characterisation of eQTLs revealed functions within regulation of androgen and the intracellular steroid hormone receptor signalling pathway and of xenobiotic metabolism by cytochrome P450 system and cellular response to oestradiol. A QTL enrichment test revealed 89 QTL traits...

Sperm length, sperm storage and mating system characteristics in bumblebees

DEFF Research Database (Denmark)

Baer, Boris; Schmid-Hempel, Paul; Høeg, Jens Thorvald

2003-01-01

-term storage of sperm, using three bumblebee species with different mating systems as models. We show that individual males produce only one size-class of sperm, but that sperm length is highly variable among brothers, among unrelated conspecific males, and among males of different species. Males of Bombus...

Electron probe X-ray microanalysis of boar and inobuta testes after the Fukushima accident

International Nuclear Information System (INIS)

Yamashiro, Hideaki; Abe, Yasuyuki; Hayashi, Gohei; Urushihara, Yusuke; Kuwahara, Yoshikazu; Suzuki, Masatoshi; Kobayashi, Jin; Kino, Yasuyuki; Fukuda, Tomokazu; Tong, Bin; Takino, Sachio; Sugano, Yukou; Sugimura, Satoshi; Yamada, Takahisa; Isogai, Emiko; Fukumoto, Manabu

2015-01-01

We aimed to investigate the effect of chronic radiation exposure associated with the Fukushima Daiichi Nuclear Power Plant (FNPP) accident on the testes of boar and inobuta (a hybrid of Sus scrofa and Sus scrofa domestica). This study examined the contamination levels of radioactive caesium (Cs), especially 134 Cs and 137 Cs, in the testis of both boar and inobuta during 2012, after the Fukushima accident. Morphological analysis and electron-probe X-ray microanalysis (EPMA) were also undertaken on the testes. The 134 Cs and 137 Cs levels were 6430 ± 23 and 6820 ± 32 Bq/kg in the boar testes, and 755 ± 13 and 747 ± 17 Bq/kg in the inobuta testes, respectively. The internal and external exposure of total 134 Cs and 137 Cs in the boar testes were 47.1 mGy and 176.2 mGy, respectively, whereas in the inobuta testes, these levels were 6.09 mGy and 59.8 mGy, respectively. Defective spermatogenesis was not detected by the histochemical analysis of radiation-exposed testes for either animal. In neither animal were Cs molecules detected, using EPMA. In conclusion, we showed that adverse radiation-induced effects were not detected in the examined boar and inobuta testes following the chronic radiation exposure associated with the FNPP accident

Protein-Carbohydrate Interaction between Sperm and the Egg-Coating Envelope and Its Regulation by Dicalcin, a Xenopus laevis Zona Pellucida Protein-Associated Protein

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Naofumi Miwa

2015-05-01

Full Text Available Protein-carbohydrate interaction regulates multiple important processes during fertilization, an essential biological event where individual gametes undergo intercellular recognition to fuse and generate a zygote. In the mammalian female reproductive tract, sperm temporarily adhere to the oviductal epithelium via the complementary interaction between carbohydrate-binding proteins on the sperm membrane and carbohydrates on the oviductal cells. After detachment from the oviductal epithelium at the appropriate time point following ovulation, sperm migrate and occasionally bind to the extracellular matrix, called the zona pellucida (ZP, which surrounds the egg, thereafter undergoing the exocytotic acrosomal reaction to penetrate the envelope and to reach the egg plasma membrane. This sperm-ZP interaction also involves the direct interaction between sperm carbohydrate-binding proteins and carbohydrates within the ZP, most of which have been conserved across divergent species from mammals to amphibians and echinoderms. This review focuses on the carbohydrate-mediated interaction of sperm with the female reproductive tract, mainly the interaction between sperm and the ZP, and introduces the fertilization-suppressive action of dicalcin, a Xenopus laevis ZP protein-associated protein. The action of dicalcin correlates significantly with a dicalcin-dependent change in the lectin-staining pattern within the ZP, suggesting a unique role of dicalcin as an inherent protein that is capable of regulating the affinity between the lectin and oligosaccharides attached on its target glycoprotein.

Improving sperm banking efficiency in endangered species through the use of a sperm selection method in brown bear (Ursus arctos) thawed sperm.

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Anel-Lopez, L; Ortega-Ferrusola, C; Álvarez, M; Borragán, S; Chamorro, C; Peña, F J; Morrell, J; Anel, L; de Paz, P

2017-06-26

Sperm selection methods such as Single Layer Centrifugation (SLC) have been demonstrated to be a useful tool to improve the quality of sperm samples and therefore to increase the efficiency of other artificial reproductive techniques in several species. This procedure could help to improve the quality of genetic resource banks, which is essential for endangered species. In contrast, these sperm selection methods are optimized and focused on farm animals, where the recovery task is not as important as in endangered species because of their higher sperm availability. The aim of this study was to evaluate two centrifugation methods (300 x g/20 min and 600 x g/10 min) and three concentrations of SLC media (Androcoll-Bear -80, 65 and 50%) to optimise the procedure in order to recover as many sperm with the highest quality as possible. Sperm morphology could be important in the hydrodynamic relationship between the cell and centrifugation medium and thus the effect of sperm head morphometry on sperm yield and its hydrodynamic relationship were studied. The samples selected with Androcoll-Bear 65% showed a very good yield (53.1 ± 2.9) although the yield from Androcoll-Bear 80% was lower (19.3 ± 3.3). The latter showed higher values of motility than the control immediately after post-thawing selection. However, both concentrations of colloid (65 and 80%) showed higher values of viable sperm and viable sperm with intact acrosome than the control. After an incubation of 2 h at 37 °C, the samples from Androcoll-Bear 80% had higher kinematics and proportion of viable sperm with intact acrosome. In the morphometric analysis, the sperm selected by the Androcoll-Bear 80% showed a head with a bigger area which was more elongated than the sperm from other treatments. We conclude that sperm selection with Androcoll-Bear at either 65% or 80% is a suitable technique that allows a sperm population with better quality than the initial sample to be obtained. We recommend the

BSp66 protease is widespread in the acrosomal region of sperm from several mammalian species

International Nuclear Information System (INIS)

Cesari, A.; Katunar, M.R.; Monclus, M.A.; Vincenti, A.; Fornes, M.W.

2004-01-01

Fertilization in mammals comprises a sequence of events leading to the fusion of sperm and oocyte membranes. Although proteases are known to be involved in this process, their role in fertilization is controversial. There is extensive work on the characterization of proteolytic systems, including serine proteases, which demonstrates that acrosomal proteases can be distinguished among the sperm of different mammalian species on the basis of the gelatin-hydrolyzing activity on SDS-PAGE by the quantity and variety of the enzymes. In this report, we investigated the occurrence and activity of the serine protease BSp66, previously characterized in bovine spermatozoa, in various mammalian sperm. A protein with a molecular mass of 66 kDa cross-reacted with heterologous antibodies against bovine BSp66 when sperm extracts of several mammalian species were analyzed by Western blot. In agreement, proteolytic activity corresponding to the molecular mass of BSp66 was detected by gelatin zymography in all the species analyzed. This protein was located on the acrosomal region of sperm cells by immunofluorescence methods. We concluded that BSp66 is widespread in mammalian sperm, with a conserved location in the acrosomal region

Selection of Sperm Based on Hypo-Osmotic Swelling May Improve ICSI Outcome: A Preliminary Prospective Clinical Trial

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Nasim Charehjooy

2014-03-01

Full Text Available Background: The intra-cytoplasmic sperm injection (ICSI technique selects sperm according to morphology and motility. However, these parameters cannot predict the chromatin integrity of sperm. Considering the detrimental effects of DNA-damaged sperm on reproductive outcomes, novel sperm selection procedures have been proposed to circumvent the possibility of inseminating DNA-damaged sperm. It has been shown that different potential hypo-osmotic swelling test (HOST patterns possess the potential to differentiate between sperm that have intact or damaged chromatin. Therefore, for the first time, this preliminary study evaluates the role of HOST as a sperm selection procedure in a clinical setting. Materials and Methods: In this preliminary prospective clinical trial study, we divided infertile couples diagnosed with male infertility into two groups. In the treatment group (n=39, half of the oocytes were inseminated by sperm selected following density gradient centrifugation (DGC group. The remaining oocytes from the treatment group were inseminated by sperm chosen according to HOST pattern (c, d or e following DGC processing (HOST group. In the control group (n=63, all oocytes were inseminated by sperm chosen after DGC. Results: There was a significantly higher percentage of embryos that had good quality, implantation, and chemical pregnancy rates in the HOST group compared to the DGC group (p≤0.05. Conclusion: This study has shown that selecting sperm according to membrane functionality (HOST pattern rather morphology and viability may open a new window in our approach for determining the appropriate sperm for ICSI, particularly in individuals with severe male infertility (Registration Number: IRCT201307087223N2.

Analysis of limited fertility in intracytoplasmic sperm injection of sperm obtained by electroejaculation

Science.gov (United States)

Nakamura, Yoshihiro; Kitamura, Masaya; Nishimura, Kenji; Tsujimura, Akira; Takeyama, Masami; Kondoh, Nobuyuki; Miyazaki, Kazunori; Okuyama, Akihiko

2004-01-01

Background and Aims:  We correlated findings in semen from patients with ejaculatory dysfunction with results of in vitro fertilization using their electroejaculated sperm. Methods and Results:  Electroejaculation was carried out in six patients with the above‐mentioned criteria for a total of eight times. Sperm was obtained in six attempts. Intracytoplasmic injection of these sperm was performed in 156 eggs. Sixty‐seven eggs were fertilized; most of these were injected with motile sperm. Two women became pregnant, both after injection with motile sperm. As previously reported, electroejaculated sperm showed low motility and a low fertilization rate, but even motile sperm had a low fertilization rate. Conclusion:  The results of the present study suggest the importance in fertilization of undetermined factors in addition to sperm motility. (Reprod Med Biol 2004; 3: 9–12) PMID:29662380

The E3 Ubiquitin Ligase MIB-1 Is Necessary To Form the Nuclear Halo in Caenorhabditis elegans Sperm.

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Herrera, Leslie A; Starr, Daniel A

2018-05-18

Unlike the classical nuclear envelope with two membranes found in other eukaryotic cells, most nematode sperm nuclei are not encapsulated by membranes. Instead, they are surrounded by a nuclear halo of unknown composition. How the halo is formed and regulated is unknown. We used forward genetics to identify molecular lesions behind three classical fer (fertilization defective) mutations that disrupt the ultrastructure of the Caenorhabditis elegans sperm nuclear halo. We found fer-2 and fer-4 alleles to be nonsense mutations in mib-1. fer-3 was caused by a nonsense mutation in eri-3 GFP::MIB-1 was expressed in the germline during early spermatogenesis, but not in mature sperm. mib-1 encodes a conserved E3 ubiquitin ligase homologous to vertebrate Mib1 and Mib2, which function in Notch signaling. Here, we show that mib-1 is important for male sterility and is involved in the regulation or formation of the nuclear halo during nematode spermatogenesis. Copyright © 2018, G3: Genes, Genomes, Genetics.

No evidence of trade-offs in the evolution of sperm numbers and sperm size in mammals.

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Tourmente, M; Delbarco Trillo, J; Roldan, E R S

2015-10-01

Post-copulatory sexual selection, in the form sperm competition, has influenced the evolution of several male reproductive traits. However, theory predicts that sperm competition would lead to trade-offs between numbers and size of spermatozoa because increased costs per cell would result in a reduction of sperm number if both traits share the same energetic budget. Theoretical models have proposed that, in large animals, increased sperm size would have minimal fitness advantage compared with increased sperm numbers. Thus, sperm numbers would evolve more rapidly than sperm size under sperm competition pressure. We tested in mammals whether sperm competition maximizes sperm numbers and size, and whether there is a trade-off between these traits. Our results showed that sperm competition maximizes sperm numbers in eutherian and metatherian mammals. There was no evidence of a trade-off between sperm numbers and sperm size in any of the two mammalian clades as we did not observe any significant relationship between sperm numbers and sperm size once the effect of sperm competition was taken into account. Maximization of both numbers and size in mammals may occur because each trait is crucial at different stages in sperm's life; for example size-determined sperm velocity is a key determinant of fertilization success. In addition, numbers and size may also be influenced by diverse energetic budgets required at different stages of sperm formation. © 2015 European Society For Evolutionary Biology. Journal of Evolutionary Biology © 2015 European Society For Evolutionary Biology.

Daily Sperm Production, Gonadal and Extra-Gonadal Sperm ...

African Journals Online (AJOL)

SH

animals fed diets 2 and 3 were similar to the control animals but they were significantly (P<0.05) lower than those fed ... Keywords: Prebiotics, probiotics, rabbits, sperm reserves, sperm production. ... Materials and methods .... In: Handbook of.

Role of sperm surface proteins in reproduction

Czech Academy of Sciences Publication Activity Database

Jonáková, Věra; Maňásková, Pavla; Davidová, Nina; Tichá, M.; Pěknicová, Jana

2009-01-01

Roč. 30, Supplement (2009), s. 63-64 ISSN 0196-3635. [9th International Congress of Andrology. 07.03.2009-10.03.2009, Barcelona] R&D Projects: GA MŠk(CZ) 1M06011; GA ČR(CZ) GA523/08/H064; GA ČR(CZ) GA303/06/0895 Institutional research plan: CEZ:AV0Z50520701 Keywords : boar seminal plasma proteins * spermadhesins * proteinase inhibitor * DQH * boar spermatozoa Subject RIV: CE - Biochemistry

Influence of growth rate and onset of boar contact on puberty attainment of replacement gilts raised in Thailand.

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Roongsitthichai, Atthaporn; Olanratmanee, Em-On; Tummaruk, Padet

2014-10-01

This study aimed to investigate the influence of growth rate and onset of boar contact on age at first observed estrus of the replacement gilts raised in Thailand. In total, 766 gilts were measured for body weight and backfat thickness prior to insemination. Body weight was further calculated for growth rate. Estrus detection was performed twice a day by back pressure test with an existence of mature boars with high libido. The first date of boar exposure and that of first observed estrus were individually recorded. Due to growth rate, they were classified into three groups: high (>700 g/day), moderate (600-700 g/day), and low (<600 g/day). According to onset of boar contact, the gilts were grouped into two categories: early (<150 days) and late (≥150 days). The results revealed that the gilts expressed first observed estrus, averagely, at age 205.1 ± 34.1 days, had a growth rate of 615.5 ± 57.6 g/day, and first contact with boars at 160.7 ± 19.9 days of age. The gilts with low growth rate expressed first estrus later than those with moderate (208.6 ± 2.0 vs 198.0 ± 3.2 days, P = 0.033) and high growth rate (208.6 ± 2.0 vs 193.9 ± 6.7 days, P = 0.005) groups. Together with the influence of boar exposure, the gilts contacted boar earlier with high growth rate showed first estrus at age 180.3 ± 10.1 days, whereas those with later boar contact with low growth rate showed first estrus at age 197.9 ± 3.2 days. In summary, the replacement gilts should have high growth rate and contact boar early to attain puberty faster and possess decent subsequent reproductive performance.

Analysis of hygienic critical control points in boar semen production.

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Schulze, M; Ammon, C; Rüdiger, K; Jung, M; Grobbel, M

2015-02-01

The present study addresses the microbiological results of a quality control audit in artificial insemination (AI) boar studs in Germany and Austria. The raw and processed semen of 344 boars in 24 AI boar studs were analyzed. Bacteria were found in 26% (88 of 344) of the extended ejaculates and 66.7% (18 of 24) of the boar studs. The bacterial species found in the AI dose were not cultured from the respective raw semen in 95.5% (84 of 88) of the positive samples. These data, together with the fact that in most cases all the samples from one stud were contaminated with identical bacteria (species and resistance profile), indicate contamination during processing. Microbiological investigations of the equipment and the laboratory environment during semen processing in 21 AI boar studs revealed nine hygienic critical control points (HCCP), which were addressed after the first audit. On the basis of the analysis of the contamination rates of the ejaculate samples, improvements in the hygiene status were already present in the second audit (P = 0.0343, F-test). Significant differences were observed for heating cabinets (improvement, P = 0.0388) and manual operating elements (improvement, P = 0.0002). The odds ratio of finding contaminated ejaculates in the first and second audit was 1.68 (with the 95% confidence interval ranging from 1.04 to 2.69). Furthermore, an overall good hygienic status was shown for extenders, the inner face of dilution tank lids, dyes, and ultrapure water treatment plants. Among the nine HCCP considered, the most heavily contaminated samples, as assessed by the median scores throughout all the studs, were found in the sinks and/or drains. High numbers (>10(3) colony-forming units/cm(2)) of bacteria were found in the heating cabinets, ejaculate transfer, manual operating elements, and laboratory surfaces. In conclusion, the present study emphasizes the need for both training of the laboratory staff in monitoring HCCP in routine semen

Improvement of post-thawed sperm quality and fertility of Arian rooster by oral administration of d-aspartic acid.

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Ansari, Mahdi; Zhandi, Mahdi; Kohram, Hamid; Zaghari, Mojtaba; Sadeghi, Mostafa; Sharafi, Mohsen

2017-04-01

This study was conducted to investigate the effect of d-Aspartic acid (D-Asp) on post-thawed sperm quality, fertility and hatchability outcomes in male broiler breeders. Twenty 55-week-old roosters were selected and equally split into four groups (n = 5 rooster/group). Different daily D-Asp doses including 0 (D-0), 100 (D-100), 200 (D-200) or 300 (D-300) mg/kg BW were capsulated and individually administered for 12 weeks to roosters in each group. Semen samples were weekly collected from 7th to 12th week of experiment. Sperm quality from 7th to 11th week was evaluated in both fresh (total and forward motility and plasma membrane functionality) and post-thawed (total and forward motility, plasma membrane functionality, apoptosis status and mitochondrial activity) conditions. Also, collected semen samples on the 12th week were frozen and artificially inseminated to evaluate fertility and hatchability. The results from fresh condition showed that total and forward motility and plasma membrane functionality were significantly higher in D-200 compared to other groups. Also, interaction effect of time and treatment was not significant for all assessed parameters in fresh condition. In post-thawed condition, D-200 showed significantly higher total and forward motility, fertility and hatchability compared to other groups. The higher value for plasma membrane functionality and mitochondrial activity was observed in D-200 compared to D-0 and D300 groups. However, the percentage of live, early apoptotic and dead spermatozoa were not significantly affected by applied treatment in the current study. No significant difference for time and treat interaction effect was observed for all assessed parameters except forward motility. In conclusion, it seems that D-Asp administration could improve fresh and post-thawed sperm quality and post-thawed sperm fertility in male broiler breeders. Copyright © 2017 Elsevier Inc. All rights reserved.

Detection of atypical porcine pestivirus in semen from commercial boar studs in the United States.

Science.gov (United States)

Gatto, I R H; Arruda, P H; Visek, C A; Victoria, J G; Patterson, A R; Krull, A C; Schwartz, K J; de Oliveira, L G; Arruda, B L

2018-04-01

Atypical porcine pestivirus (APPV) has recently been identified as a cause of congenital tremor (CT) in pigs and has been detected in semen and preputial swabs from boars that were known to be clinically affected with CT. Accordingly, the objectives of this study were to 1) detect the presence of APPV in semen, preputial fluids and preputial swabs from adult boars by quantitative reverse transcription PCR (qRT-PCR) and 2) genetically characterize a subset of positive samples to better understand the ecology of APPV in commercial boar studs and the potential risk of transmission of APPV via semen. A total of 597 samples of semen, preputial fluid and preputial swabs each representing a different boar were obtained from four commercial boar studs located in three different states in the United States. Viral RNA was detected by qRT-PCR in 90 samples (15.08%; 90/597), with the greatest per cent positive from preputial swabs (23.81%; 5/21) followed by preputial fluid (22.81%; 26/114) and semen (12.91%; 59/457). The mean cycle quantification (Cq) between sample types was similar while eleven semen samples had Cq values lower than 27.0 corresponding to approximately 2 × 10 6  copies/ml. Based on phylogenetic analysis of the Npro gene, different viral strains can be on the same farm at the same and different times. This is the first report of detection of APPV in semen from commercial boar studs. Studies investigating the role of semen in the transmission of APPV and production of CT are needed. © 2017 Blackwell Verlag GmbH.

Toxic indole alkaloids avrainvillamide and stephacidin B produced by a biocide tolerant indoor mold Aspergillus westerdijkiae.

Science.gov (United States)

Mikkola, Raimo; Andersson, Maria A; Hautaniemi, Maria; Salkinoja-Salonen, Mirja S

2015-06-01

Toxic Aspergillus westerdijkiae were present in house dust and indoor air fall-out from a residence and a kindergarten where the occupants suffered from building related ill health. The A. westerdijkiae isolates produced indole alkaloids avrainvillamide (445 Da) and its dimer stephacidin B (890 Da). It grew and sporulated in presence of high concentrations of boron or polyguanidine (PHMB, PHMG) based antimicrobial biocides used to remediate mold infested buildings. The boar sperm cells were used as sensor cells to purify toxins from HPLC fractions of the fungal biomass. Submicromolar concentrations (EC50 0.3-0.4 μM) blocked boar spermatozoan motility and killed porcine kidney tubular epithelial cells (PK-15). Plate grown hyphal mass of the A. westerdijkiae isolates contained 300-750 ng of avrainvillamide and 30-300 ng of stephacidin B per mg (wet weight). The toxins induced rapid (30 min) loss of boar sperm motility, followed (24 h) by loss of mitochondrial membrane potential (ΔΨm). Apoptotic cell death was observed in PK-15 cell monolayers, prior to cessation of glucose uptake or loss of ΔΨm. Avrainvillamide and stephacidin B were 100-fold more potent towards the porcine cells than the mycotoxins stephacidin A, ochratoxin A, sterigmatocystin and citrinin. The high toxicity of stephacidin B indicates a role of nitrone group in the mechanism of toxicity. Avrainvillamide and stephacidin B represent a new class of toxins with possible a threat to human health in buildings. Furthermore, the use of biocides highly enhanced the growth of toxigenic A. westerdijkiae. Copyright © 2015 Elsevier Ltd. All rights reserved.

Zoonotic intestinal protozoan of the wild boars, Sus scrofa, in Persian Gulf’s coastal area (Bushehr province, Southwestern Iran

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Kambiz Yaghoobi

2016-10-01

Full Text Available Aim: Wild boars, Sus scrofa, are potential reservoirs of many zoonotic diseases, and there are a possibility of transmission of the zoonotic diseases from these animals to humans and also domestic animals. This study aimed to evaluate the protozoan contamination of wild boars in the Persian Gulf’s coastal area (Bushehr Province, southwestern Iran. Materials and Methods: A total of 25 crossbred boars were collected during a course of vertebrate pest control in Bushehr province, in 2013. Samples were collected from the gastrointestinal tracts of each boar in 5% formalin, Bouin’s solution, sodium acetate-acetic acid-formalin, and polyvinyl alcohol fixatives. Fixed stool smears examined by trichrome and Ziehl–Neelsen staining. Results: Each of the 25 wild boars was infected with at least one of the intestinal protozoans. The rate of contamination with intestinal protozoan was 64% for Balantidium coli, 76% for Iodamoeba sp., 52% for Entamoeba polecki, 44% for Blastocystis sp. and 8% for Chilomastix sp. No intestinal coccidian was detected in studied boars when the stool samples were evaluated by Ziehl–Neelsen staining method. Conclusion: Findings of this study demonstrated that wild boars in the Persian Gulf coastal area are contaminated by many protozoans, including zoonotic protozoan, which poses a potential risk to locals as well as the domestic animals of the area.

The stochastic dance of circling sperm cells: sperm chemotaxis in the plane

Energy Technology Data Exchange (ETDEWEB)

Friedrich, B M; Juelicher, F [Max Planck Institute for the Physics of Complex Systems, Noethnitzer Strasse 38, 01187 Dresden (Germany)], E-mail: ben@pks.mpg.de, E-mail: julicher@pks.mpg.de

2008-12-15

Biological systems such as single cells must function in the presence of fluctuations. It has been shown in a two-dimensional experimental setup that sea urchin sperm cells move toward a source of chemoattractant along planar trochoidal swimming paths, i.e. drifting circles. In these experiments, a pronounced variability of the swimming paths is observed. We present a theoretical description of sperm chemotaxis in two dimensions which takes fluctuations into account. We derive a coarse-grained theory of stochastic sperm swimming paths in a concentration field of chemoattractant. Fluctuations enter as multiplicative noise in the equations for the sperm swimming path. We discuss the stochastic properties of sperm swimming and predict a concentration-dependence of the effective diffusion constant of sperm swimming which could be tested in experiments.

The stochastic dance of circling sperm cells: sperm chemotaxis in the plane

International Nuclear Information System (INIS)

Friedrich, B M; Juelicher, F

2008-01-01

Biological systems such as single cells must function in the presence of fluctuations. It has been shown in a two-dimensional experimental setup that sea urchin sperm cells move toward a source of chemoattractant along planar trochoidal swimming paths, i.e. drifting circles. In these experiments, a pronounced variability of the swimming paths is observed. We present a theoretical description of sperm chemotaxis in two dimensions which takes fluctuations into account. We derive a coarse-grained theory of stochastic sperm swimming paths in a concentration field of chemoattractant. Fluctuations enter as multiplicative noise in the equations for the sperm swimming path. We discuss the stochastic properties of sperm swimming and predict a concentration-dependence of the effective diffusion constant of sperm swimming which could be tested in experiments.

Use of testicular sperm for intracytoplasmic sperm injection in men with high sperm DNA fragmentation: a SWOT analysis.

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Esteves, Sandro C; Roque, Matheus; Garrido, Nicolás

2018-01-01

Spermatozoa retrieved from the testis of men with high levels of sperm DNA fragmentation (SDF) in the neat semen tend to have better DNA quality. Given the negative impact of SDF on the outcomes of Assisted Reproductive Technology (ART), an increased interest has emerged about the use of testicular sperm for intracytoplasmic sperm injection (Testi-ICSI). In this article, we used a SWOT (strengths, weaknesses, opportunities, and threats) analysis to summarize the advantages and drawbacks of this intervention. The rationale of Testi-ICSI is bypass posttesticular DNA fragmentation caused by oxidative stress during sperm transit through the epididymis. Hence, oocyte fertilization by genomically intact testicular spermatozoa may be optimized, thus increasing the chances of creating a normal embryonic genome and the likelihood of achieving a live birth, as recently demonstrated in men with high SDF. However, there is still limited evidence as regards the clinical efficacy of Testi-ICSI, thus creating opportunities for further confirmatory clinical research as well as investigation of Testi-ICSI in clinical scenarios other than high SDF. Furthermore, Testi-ICSI can be compared to other laboratory preparation methods for deselecting sperm with damaged DNA. At present, the available literature supports the use of testicular sperm when performing ICSI in infertile couples whose male partners have posttesticular SDF. Due to inherent risks of sperm retrieval, Testi-ICSI should be offered when less invasive treatments for alleviating DNA damage have failed. A call for continuous monitoring is nonetheless required concerning the health of generated offspring and the potential complications of sperm retrieval.

Recovery and cryopreservation of epididymal sperm from agouti (Dasiprocta aguti) using powdered coconut water (ACP-109c) and Tris extenders.

Science.gov (United States)

Silva, M A; Peixoto, G C X; Santos, E A A; Castelo, T S; Oliveira, M F; Silva, A R

2011-10-01

The objective was to compare the use of powdered coconut water (ACP-109c; ACP Biotecnologia, Fortaleza, CE, Brazil) and Tris extenders for recovery and cryopreservation of epididymal sperm from agouti. The caudae epididymus and proximal ductus deferens from 10 sexually mature agoutis were subjected to retrograde washing using ACP-109c (ACP Biotecnologia) or Tris. Epididymal sperm were evaluated for motility, vigor, sperm viability, membrane integrity, and morphology. Samples were centrifuged, and extended in the same diluents plus egg yolk (20%) and glycerol (6%), frozen in liquid nitrogen, and subsequently thawed at 37°C for 1 min, followed by re-evaluation of sperm characteristics. The two extenders were similarly efficient for epididymal recovery, with regard to the number and quality of sperm recovered. However, for both extenders, sperm quality decreased (P Biotecnologia) group, which was significantly better than 9.7 ± 2.6% motile sperm with 1.2 ± 0.3 vigor in Tris. In conclusion, agouti epididymal sperm were successfully recovered using either ACP-109c (ACP Biotecnologia) or Tris extenders; however, ACP-109c (ACP Biotecnologia) was a significantly better extender for processing and cryopreserving these sperm. Copyright © 2011 Elsevier Inc. All rights reserved.

Protective effects of Opuntia ficus-indica extract on ram sperm quality, lipid peroxidation and DNA fragmentation during liquid storage.

Science.gov (United States)

Allai, Larbi; Druart, Xavier; Öztürk, Mehmet; BenMoula, Anass; Nasser, Boubker; El Amiri, Bouchra

2016-12-01

The present study aimed to assess the phenolic composition of the acetone extract from Opuntia ficus indica cladodes (ACTEX) and its effects on ram semen variables, lipid peroxidation and DNA fragmentation during liquid storage at 5°C for up to 72h in skim milk and Tris egg yolk extenders. Semen samples from five rams were pooled extended with Tris-egg yolk (TEY) or skim milk (SM) extenders containing ACTEX (0%, 1%, 2%, 4% and 8%) at a final concentration of 0.8×10 9 sperm/ml and stored for up to 72h at 5°C. The sperm variables were evaluated at different time periods (8, 24, 48 and 72h). Sperm total motility and viability were superior in TEY than in SM whereas the progressive motility, membrane integrity, abnormality and spontaneous lipid peroxidation were greater in SM compared to TEY (P<0.05). The results also indicated that the inclusion of 1% ACTEX in the SM or TEY extender increased the sperm motility, viability, membrane integrity, and decreased the abnormality, lipids peroxidation up to 72h in storage compared to control group. Similarly, even at 72h of storage, 1% ACTEX can efficiently decrease the negative effects of liquid storage on sperm DNA fragmentation (P<0.05). In conclusion, SM and TEY supplemented with 1% of ACTEX can improve the quality of ram semen. Further studies are required to identify the active components in ACTEX involved in its effect on ram sperm preservation. Copyright © 2016 Elsevier B.V. All rights reserved.

Sperm competition leads to functional adaptations in avian testes to maximize sperm quantity and quality.

Science.gov (United States)

Lüpold, Stefan; Wistuba, Joachim; Damm, Oliver S; Rivers, James W; Birkhead, Tim R

2011-05-01

The outcome of sperm competition (i.e. competition for fertilization between ejaculates from different males) is primarily determined by the relative number and quality of rival sperm. Therefore, the testes are under strong selection to maximize both sperm number and quality, which are likely to result in trade-offs in the process of spermatogenesis (e.g. between the rate of spermatogenesis and sperm length or sperm energetics). Comparative studies have shown positive associations between the level of sperm competition and both relative testis size and the proportion of seminiferous (sperm-producing) tissue within the testes. However, it is unknown how the seminiferous tissue itself or the process of spermatogenesis might evolve in response to sperm competition. Therefore, we quantified the different germ cell types and Sertoli cells (SC) in testes to assess the efficiency of sperm production and its associations with sperm length and mating system across 10 species of New World Blackbirds (Icteridae) that show marked variation in sperm length and sperm competition level. We found that species under strong sperm competition generate more round spermatids (RS)/spermatogonium and have SC that support a greater number of germ cells, both of which are likely to increase the maximum sperm output. However, fewer of the RS appeared to elongate to mature spermatozoa in these species, which might be the result of selection for discarding spermatids with undesirable characteristics as they develop. Our results suggest that, in addition to overall size and gross morphology, testes have also evolved functional adaptations to maximize sperm quantity and quality.