Does sperm quality and DNA integrity differ in cryopreserved semen samples from young,adult, and aged Nellore bulls?

Background

In humans, it is now well documented that rising paternal age is correlated with decreased sperm DNA integrity and embryonic developmental failures. On the other side of the coin, it is also reported that very young fathers such as teenagers carry an increased risk of adverse birth outcomes. These observations suggest that, at least in humans, there is an age window for optimal sperm DNA integrity. In bovine, little is known about sperm DNA quality in young bulls and how it evolves with age. This study aimed to fill in this gap as it may be of importance for the bovine industry to know when exactly a bull is an optimal performer for reproductive programs.

Methods

Forty Nellore bulls were divided into three age groups: 1.8 to 2 years – young bulls; 3.5 to 7 years – adult bulls; and 8 to 14.3 years – aged bulls. Three ejaculates were collected from each bull, cryopreserved and evaluated for various parameters including: computer-assisted sperm analysis (CASA), plasma membrane and acrosome integrity, mitochondrial potential, sperm nuclear protamination, DNA oxidative damage, and Sperm Chromatin Structure Assay (SCSA).

Results

We report here that young bulls presented superior values for motility, plasma and acrosomal membrane integrity, and high mitochondrial potential. However, they also presented higher values for sperm morphological abnormalities compared to adult and aged animal groups (p?<?0.05). In addition, young bulls exhibited more defective protamination than older animals did. The oldest bulls showed more nuclear oxidative damage than the younger groups of bulls while both the young and aged groups were found more susceptible to DNA denaturation as revealed with the SCSA test (p?<?0.05).

Conclusion

These results indicate that young bulls spermatozoa best survived the freezing procedure, followed by adult and aged bulls. However, young and aged bulls were found to be more susceptible to DNA damage, respectively caused by protamine deficiency and oxidation. Therefore, although young bulls have correct semen parameters according to classical evaluation, our results indicate that they may show some structural nuclear immaturity.

     

What affects fertility of sexed bull semen more, low sperm dosage or the sorting process?

Until now it has been unclear to what extent the reduced fertility with sexed semen in the dairy industry is caused by too few sperm per AI dose, or by the effect of flow cytometric sorting, which is the established procedure for sexing semen. Therefore, we evaluated the effects of low sperm numbers per dose with and without sorting on non-return rates after 56 days (NRR56); in addition, we evaluated the effects of bulls, in order to further optimize use of sexed semen.Based on results of using sexed semen from seven Holstein bulls, an overall numerical decline of 13.6% in NRR56 was observed (P < 0.05). About two-thirds of this decline (8.6%) was due to the low dose (P < 0.05), and a third (5.0%) due to the process of sorting (P < 0.05). The effect of low dosage and sorting differed among bulls. We observed a sex ratio of 91.6% females for sexed semen from the first 131 calves born.Currently the best way to increase fertility of sexed semen is by closely monitoring fertility so that the highest fertility bulls are used, and by improving farm animal management. However, to make substantial progress, more in depth studies are needed on the sexing technology, especially on aspects such as sorting procedures and sperm dosage.     

Does attractiveness in men provide clues to semen quality?

The psychological mechanisms underlying attractiveness judgements in humans are thought to be evolved adaptations for finding a high quality mate. The phenotype-linked fertility hypothesis proposes that females obtain reliable information on male fertility from male expression of sexual traits. A previous study of Spanish men reported that facial attractiveness was positively associated with semen quality. We aimed to determine whether this effect was widespread by examining a large sample of Australian men. We also extended our study to determine whether cues to semen quality are provided by components of attractiveness: masculinity, averageness and symmetry. Each male participant was photographed and provided a semen sample that was analyzed for sperm morphology, motility and concentration. Two independent sets of women rated the male photographs for attractiveness, and three further sets of 12 women rated the photographs for masculinity, symmetry or averageness. We found no significant correlations between semen quality parameters and attractiveness or attractive traits. Although male physical attractiveness may signal aspects of mate quality, our results suggest that phenotype-linked cues to male fertility may not be general across human populations.     

Motility and fertility of cryopreserved semen in Persian sturgeon,Acipenser persicus,stored for 30–60 min after thawing

We investigated the effect of storage times of frozen–thawed Persian sturgeon (Acipenser persicus) semen on the duration of sperm motility, percentage of motile sperm, and fertilization and hatching rates of fresh sperm and sperm stored for 0, 30, and 60 min at 4 °C post-thawing. Frozen thawed semen analyzed immediately after thawing had similar quality characteristics as fresh semen. For cryopreserved semen stored for 30 min after thawing the characteristics did not differ to fresh semen and cryopreserved semen. For cryopreserved semen stored for 60 min a significant decline in the parameters was observed. Fertilization and hatching rates were not affected by storage times of maximally 30 min of storage.     

Declining semen quality: Can the past inform the present?

By using instrumentation initially designed for counting white blood cells, sperm counts have been utilized by clinicians since 1929, particularly to evaluate cases of suspected infertility. Although this basic biological parameter might be assumed to be stable over time, several studies over the past 20 years have suggested a decline in sperm count or density. The most controversial of these analyses was published in 1992. A flood of criticism followed this analysis of 61 studies that found a 50% decline in sperm density between 1938 and 1990. Critics suggested that historical methods (of counting sperm or conducting studies) were variable and unreliable, differing from modern methods both qualitatively and quantitatively. To address this issue we analyzed these studies for trends in counting methods or their variability. We found neither. Alternative analyses produced some differences in trend estimates, but statistical factors alone could not account for the total decline in sperm density. We reviewed study populations to identify trends in population characteristics, such as abstinence time, that might explain the decline. However, controlling analytically for such factors only increased the rate of decline. We conclude that historical data on sperm density, despite large random error, are surprisingly reliable. Nonetheless, understanding causes of temporal and geographic differences in sperm density must await contemporary data. BioEssays 21:614–621, 1999. © 1999 John Wiley & Sons, Inc.     

Morphologic,endocrine and thermographic measurements of testicles in comparison with semen characteristics in mature Holstein–Friesian breeding bulls

Twenty Holstein–Friesian breeding bulls (62–79 months of age) were examined 3 times, at 30-day intervals. Scrotal thermograms for assessment of scrotal surface temperature (SST) and blood samples for plasma testosterone concentrations were taken just before and then 45 and 90 min, respectively, after treatment with GnRH (50 μg, Gonavet, i.m. per bull). Following GnRH treatment, there generally were significant increases in mean values of both top SST (range, −0.1 to 1.4°C) and bottom SST (range, 0.3 to 1.8°C). Scrotal circumference was highly repeatable but SST and video-measurements of scrotal dimensions were less repeatable, because apparently they were affected by ambient temperature. Plasma testosterone concentrations before GnRH treatment were more repeatable than those after GnRH treatment. Correlations between examinations of 0.67 to 0.81 and −0.14 to 0.47, respectively, but the converse was true for SST measurements. Semen was collected with an artificial vagina 3 times per week for 12 weeks starting 2 weeks before the first examination. The total number of spermatozoa per ejaculate was highly repeatable and the percentage of motile and live spermatozoa were relatively consistent. Separate regressions for each variable and for each examination were conducted for these 3 semen characteristics as dependent variables. For the number of spermatozoa per ejaculate and for the percentage of motile spermatozoa, significant independent variables were plasma testosterone concentrations and difference between top and bottom SST, respectively. The slopes of these equations were nearly all negative and the R² was from 0.15 to 0.42. For prediction of the percentage of live spermatozoa, both SST gradient and plasma testosterone concentrations were significant independent variables. For these regressions, the slopes were negative and the regression coefficients were generally lower than for the other 2 dependent variables (range, 0.16 to 0.25). Treatment with GnRH and assessment of SST and plasma testosterone concentrations have some correlation with the semen production in the mature bull.     

Can detomidine replace medetomidine for pharmacological semen collection in domestic cats?

Among the different methods used for semen collection from domestic cats, the pharmacological collection by urethral catheterization becomes disruptive. Medetomidine is the elected α²-adrenoceptor agonist for that, but in several countries, it is not commercially available. This study aimed to evaluate the efficacy of detomidine compared to medetomidine in collecting semen by urethral catheterization in domestic cats. Urethral catheterization was performed on 13 mongrel cats using a disposable semi-rigid tomcat urinary catheter. Of the 19 semen collections performed with medetomidine induction, 94.7% were successful, while with detomidine induction, only 56.3% of 16 were successful. The values semen samples variables were as follows for volume - 10.56 ± 0.4 vs 8.88 ± 0.5 mL, motility - 171.67 ± 0.79 vs 49.77 ± 3.45%, vigor – 4.1 ± 0.03 vs 3.10 ± 0.1 and concentration - 3.24 ± 0.19 vs 2.15 ± 0.13 ×10⁹ sperm/mL respectively for medetomidine and detomidine group. The failure in semen collections with detomidine was mainly due to azoospermic samples, poor urethral relaxation, insufficient volume, or contamination of urine. The sperm concentration was also lower in the detomidine group (P <0.05) when compared to medetomidine. However, when the volume of semen collected was compared, we found no statistical differences. Despite its low performance in collecting semen from cats, detomidine may be an alternative when medetomidine is not accessible.     

Can amides be alternative cryoprotectors for the preservation of feline semen?

Sperm cryopreservation is a tool for the conservation of the genetic material of animals of genetic importance or for species preservation. In the case of domestic cats, this can be used to generate information about seminal harvest, evaluation and preservation, which is especially important due to its applicability to wild felids. This study evaluated seminal samples harvested by urethral catheterisation from 13 adult domestic cats. Samples were cryopreserved with experimental groups of extenders were defined by the penetrating cryoprotectant: 6% glycerol (GLY6%), 3% dimethylacetamide (DMA3%) and 3% dimethylformamide (DMF3%). The samples were thawed and evaluated by conventional microscopy and by computer-assisted sperm analysis (CASA). The structural and functional membrane integrity was assessed by supravital tests (EOS), hypoosmotic swelling tests (HOST) and flow cytometry (FC). There was a correlation (P < 0.05) between total motility and EOS (r = 0.54), HOST and FC (r = −0.62) and total motility and flow cytometry (r = 0.63), indicating that these are complementary parameters that increase the accuracy of the feline sperm quality evaluation post-thaw. The results regarding the structural and functional integrity of the sperm plasma membrane did not differ (P > 0.05) among groups. However, the DMA3% group had a lower (P < 0.05) percentage of morphological changes in the sperm tail compared to samples cryopreserved with GLY6% and DMF3%. Additionally, DMA3% provided lower values of immobile sperm post-thaw when compared to DMF3%. DMA is an interesting alternative to GLY and superior to DMF for the cryopreservation of feline semen at the studied concentrations.     

Effects of 72—hour heat stress on semen quality in boars

Three sibling pairs of six-month old boars of each of four breeding groups (Yorkshire, Poland, China and the reciprocal crosses) were maintained in an area held at or below 21.5°C with a 12-hour daily light cycle. After a three month acclimation period, a randomly selected boar of each pair was subjected to 33°C, 50% RH for 72 hours. Semen was collected from each boar once during the treatment and every fourth day for 64 days post-treatment (PT). There were significant (p < 0.05) decreases in sperm concentration per ml, and total sperm number. The inverse of the methylene blue reduction time and the per cent motile sperm and total motile sperm in the ejaculate were significantly decreased (p < 0.01). The percentages of primary and secondary morphologically abnormal spermatozoa were increased significantly (p < 0.01). Maximal pair differences occurred at days 28 to 32 PT; all had returned to pretreatment levels by day 64 PT.

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Zusammenfassung Drei Geschwisterpaare 6-Monate alter Saubären aus 4 Zuchtgruppen (Yorkshire, Polen, China und deren reziproken Kreuzungen) wurden bei 21, 5°C in einem 12-Stunden Lichtzyklus gehalten. Nach einer Akklimatisationsperiode von 3 Monaten wurde ein zufällig ausgewählter Saubären jedes Paares 72 Stunden bei 33°C und 50% RF exponiert und von jedem Tier Samenproben einmal während der Hitzebelastung und an jedem 4. Tag bis zum 64. Tag nach der Behandlung gesammelt. Die Samenkonzentration im ml und die Gesamtspermienzahl waren signifikant reduziert (p < 0,05). Die Umkehrung der Methylenblau-Reduktionszeit und die prozentuale und totale Anzahl motiler Spermien im Ejakulat waren signifikant reduziert (p < 0,01). Der prozentuale Anteil primär und sekundär morphologisch abnormer Spermien war signifikant erhöht (p < 0,01). Maximale Unterschiede zwischen den Paaren traten zwischen dem 28. und 32. Tag auf. Am 64. Tag waren die Unterschiede verschwunden.

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Resume On a soumis 3 paires de verrats frères âgés de 6 mois à une atmosphère de 21, 5°C et à un cycle lumineux de 12 heures. Ces animaux provenaient de 4 groupes d'élevage (Yorkshire, Polen, China et leurs croisements réciproques). Après une période d'adaptation de 3 mois, on a choisi au hasard une bête de chaque paire qui fut placée durant 72 heures à 33°C et à 50% d'humidité relative. On a alors prélevé de la semence de chaque animal et cela durant la période de contrainte thermique, puis tous les 4 jours jusqu'au 64ème après l'essai. La concentration des semences par ml et le nombre total de spermatozoïdes furent réduits de façon significative (p < 0,05). L'inverse du temps de réduction au bleu de Méthylène, le taux et le nombre total de spermatozoïdes mobiles dans l'éjaculat furent également réduits de façon significative (p < 0,01). Le taux des spermatozoïdes présentant des abnormités morphologiques primaires et secondaires fut plus élevé de façon significative (p < 0,01). Les plus grandes différences entre les individus d'une même paire furent relevées du 28ème au 32ème jour après le traitement. Au 64ème jour, toute différence avait disparu.

     

Improved cryopreservation protocol for Blanca-Celtibérica buck semen collected by electroejaculation

The collection of sperm samples by electroejaculation (EE) leads to an increase of the production of seminal plasma which could modify the tolerance of spermatozoa to the cryopreservation procedure. This study aims to compare a standard sperm cryopreservation protocol for samples collected by artificial vagina (AV) with the same protocol and modifications to this for samples obtained by EE. Semen from six males of Blanca-Celtibérica goat breed was collected by AV (control) and EE, and three experiments were conducted. In Experiment 1, it was examined the effects of egg yolk concentration contained in freezing extender (0%, 1.5%, 10% and 20% of egg yolk); in Experiment 2, it was evaluated the cooling rate from 30 to 5 °C (fast: 10 min and slow: 90 min) and the temperature of glycerol addition (30 and 5 °C); and in Experiment 3, it was examined the time of equilibration at 5 °C (0, 1, 2 or 3 h). A heterologous in vitro fertilization test was carried out in order to compare the fertility of control samples with that resulting from the EE protocol which showed the highest sperm quality. Results showed greater sperm motility parameters after thawing for control samples cryopreserved in standard conditions in the three experiments. For samples collected by EE, extender with 20% egg yolk, a slow cooling rate and a longer equilibration time (3 h) provided higher sperm quality, and no differences were observed between temperatures of glycerol addition. Samples collected by EE and cryopreserved with the protocol which yielded the best sperm quality after thawing showed higher fertility compared to AV.     

Centrifugation on PureSperm(®) density-gradient improved quality of spermatozoa from frozen-thawed dog semen

The main objective of this study was to investigate if centrifugation through PureSperm^® density-gradient can improve the post-thaw semen quality of dog semen. Semen from 5 dogs was collected and cryopreserved following a standard protocol. After thawing, semen samples were selected by centrifugation on PureSperm^®. Assessments of sperm motility (assessed by computerized-assisted semen analysis), morphology (Diff-Quick staining) and viability (triple fluorescent stain of Propidium iodine/isothiocyanate-labeled peanut (Arachis hypogaea) agglutinin/Rhodamine 123), were performed on aliquots of fresh semen, unselected samples and selected preparations. Cryopreservation had a significant (P < 0.001) effect on all studied semen parameters. PureSperm^® centrifugation yielded sperm suspensions with improved motility and viability (P < 0.001). The washing step significantly reduced (P < 0.001) all of the kinematics parameters evaluated as well as reduced the proportion of viable spermatozoa with intact acrosomes (P < 0.05). We concluded that PureSperm^® centrifugation is a successful method for improving the quality of frozen-thawed dog spermatozoa. However, washing after density-gradient centrifugation dramatically reduces the post-thaw semen quality, indicating that the inclusion of such a washing step is unnecessary.     

Does artificial insemination with chilled, extended semen reduce the antigenic challenge to the mare's uterus compared with natural service?

Uterine response in infection-resistant mares (n = 5) at 48 h after AI was compared with that following natural service in these same mares, and after AI in infection-susceptible mares (n = 6). In the resistant mares, small amounts of uterine fluid were detected and bacteria were isolated infrequently at 48 h after breeding, but cytological examination of uterine flushes revealed that a significant degree of endometritis was present. There was no difference in the degree of inflammatory response by 48 h after AI or natural mating. In 4 of the 6 susceptible mares moderate to large accumulations of intrauterine fluid were detected at 48 h after AI, and massive uterine neutrophilia was present in all 6 mares. It was concluded that there was no evidence that using conventional AI techniques limited the inflammatory response of the mare's uterus.     

Seminal plasma zinc concentration and α-glucosidase activity with respect to semen quality

This study was conducted to examine the relationship between the concentration of zinc and neutral α-glucosidase (NAG) with semen quality. Semen samples from 75 male partners of couples who were attending the Obstetrics and Gynecology Department were analyzed for semen quality. Based on sperm count, the subjects were divided into three groups. Zinc and neutral α-glucosidase activity were estimated in seminal plasma. Results showed that mean the α-glucosidase activity was lowest among the azoospermic group with respect to oligozoospermic and normozoospermic groups. Mean zinc levels were also lower among azoospermics compared to oligozoospermic and normospermic groups. The difference was statistically significant (p<0.05). A significant positive correlation was observed between zinc levels and sperm count (r=0.29, p<0.05) and zinc and α-glucosidase activity (r=0.31, p<0.05) in seminal plasma. These results suggest that zinc and neutral α-glucosidase seem to play an important role in human reproduction.     

Is photometry an accurate and reliable method to assess boar semen concentration?

Sperm concentration assessment is a key point to insure appropriate sperm number per dose in species subjected to artificial insemination (AI). The aim of the present study was to evaluate the accuracy and reliability of two commercially available photometers, AccuCell™ and AccuRead™ pre-calibrated for boar semen in comparison to UltiMate™ boar version 12.3D, NucleoCounter SP100 and Thoma hemacytometer. For each type of instrument, concentration was measured on 34 boar semen samples in quadruplicate and agreement between measurements and instruments were evaluated. Accuracy for both photometers was illustrated by mean of percentage differences to the general mean. It was −0.6% and 0.5% for Accucell™ and Accuread™ respectively, no significant differences were found between instrument and mean of measurement among all equipment. Repeatability for both photometers was 1.8% and 3.2% for AccuCell™ and AccuRead™ respectively. Low differences were observed between instruments (confidence interval 3%) except when hemacytometer was used as a reference. Even though hemacytometer is considered worldwide as the gold standard, it is the more variable instrument (confidence interval 7.1%).The conclusion is that routine photometry measures of raw semen concentration are reliable, accurate and precise using AccuRead™ or AccuCell™. There are multiple steps in semen processing that can induce sperm loss and therefore increase differences between theoretical and real sperm numbers in doses. Potential biases that depend on the workflow but not on the initial photometric measure of semen concentration are discussed.     

Effect of anti-oxidants and oxidative stress parameters on ram semen after the freeze–thawing process

Oxidative damage to sperm resulting from reactive oxygen species generated by the cellular components of semen is one of the main causes for the decline in motility and fertility of sperm during the freeze–thawing process. The aim of this study was thus to determine the effects of anti-oxidants on standard semen parameters, lipid peroxidation (LPO) and anti-oxidant activities after the freeze–thawing of ram semen. Ejaculates collected from four Akkaraman rams, were pooled and evaluated at 33 °C. Semen samples were diluted in a Tris-based extender containing the anti-oxidants glutathione (GSH) (5 mM), oxidized glutathione (GSSG) (5 mM) or cysteine (5 mM) and an extender containing no anti-oxidants (control), cooled to 5 °C and frozen in 0.25 ml French straws. Frozen straws were thawed individually for 20 s in a water bath (37 °C) for microscopic evaluation. The use of an extender supplemented with cysteine led to the highest (P < 0.01) post-thaw motility (61.0 ± 1.9%), compared to the other treatment groups. No significant differences were observed in viability, acrosome damage and total abnormalities, and following the hypo-osmotic swelling test (HOST), following supplementation with anti-oxidants after the thawing of the semen. Following the thawing process, the levels of malondialdehyde (MDA) did not change with the addition of anti-oxidants, compared to the control. The GSH level and glutathione peroxidase (GSH-PX) activity remained significantly higher upon the addition of GSH (3.33 ± 0.14 nmol/ml and 22.02 ± 1.27 IU/g protein) and GSSG (3.24 ± 0.08 nmol/ml and 20.17 ± 3.38 IU/g protein) compared to the other treatment (P < 0.001) groups. Only cysteine significantly elevated the activity of catalase (CAT, 842.40 ± 90.42 kU/l) following the freeze–thawing process. The Vitamin E (VitE) level was significantly higher, when compared to GSSG, cysteine and the control, when GSH (4.21 ± 0.20 mg/dl) was added to the freezing extender (P < 0.001). It could be concluded that future efforts aimed on improving the efficiency of cryopreservation of ram sperm should concentrate on the use of anti-oxidant additives. The results obtained provide a new approach to the cryopreservation of ram semen, and could positively contribute to intensive sheep production.     

Do sperm contribute to the buffering capacity of steelhead trout (Oncorhynchus mykiss) semen?

The motility of salmonid sperm is pH-sensitive and the buffering capacity of the seminal plasma is low. The objective of the present study was to determine the extent to which sperm contribute to the buffering capacity of whole steelhead (Oncorhynchus mykiss) semen. To determine the buffering capacity, semen and seminal plasma samples were titrated with HCl and pH measurements taken at 1-2 min. The buffering capacity of semen was not different from that of seminal plasma over the pH range 7.5 to 8.5 and was approximately 15% to 20% less over the range 6.0 to 7.0. Comparable results were obtained for the semen and seminal plasma of the chinook salmon (Oncorhynchus tshawytscha). To assess whether the intracellular environment could influence the buffering capacity, the effects of cell disruption with n-butanol and Triton X-100 (TX-100) were determined. Over the pH range 7.5 to 8.5, the presence of n-butanol or TX-100 resulted in a doubling of the buffering capacity of the semen; TX-100, but not n-butanol, increased semen buffering capacity over the pH range 6.0 to 7.0. To determine whether the sperm's intracellular compartment might contribute to the buffering capacity over a longer duration, semen and seminal plasma samples were acidified with HCl and the pH measured over several hours. These data suggest that intact sperm contribute no more than about 25% to the buffering capacity of whole semen. The buffering capacity of steelhead semen and seminal plasma were comparably and modestly temperature sensitive. The results suggest that the sperm may contribute to the buffering capacity of the semen over a physiological pH range, however, if so, the effect is relatively small.     

Comparison of various semen extenders and addition of prostaglandin F2α on pregnancy rate in cows

This investigation comprises three trials. Trial 1 consists of an in vitro comparison of three semen extenders: two egg yolk based (customized Tris-egg yolk-glycerol and Triladyl®), the third (AndroMed®) soybean lecithin based. With regard to post-thaw motility, the phytoextender AndroMed® proved to be superior (59±3% v. 53±2% and 53±2%, P<0.05). It had earlier been shown that addition of the commercial prostaglandin F_2α preparation Dinolytic® before freezing compromises post-thaw motility; therefore, in Trial 2, Dinolytic® was added after thawing. Frozen-thawed spermatozoa tolerated addition of Dinolytic® at a concentration of 30% (v/v). In Trial 3, cows were inseminated using straws in which diluted semen and Dinolytic® were frozen in the same straw, separated by an air bubble, so intermingling could only take place in the course of insemination. Pregnancy rates at Dinolytic® dosages of 0%, 30% or 60% amounted to 44%, 41% and 56%, respectively (P>0.05), a result that encourages a large-scale field study, which is envisioned.     

Studies on the glycosidases of semen: Purification and properties of α-D-mannopyranosidase from goat seminal plasma

Summary Alpha D-mannosidase activity in goat semen was observed to be distributed in sperm and seminal plasma. In sperm the enzyme, present in soluble and bound forms, was located within the acrosome. The bound enzyme was associated with the denuded sperm. Seminal plasma -mannosidase was purified 100-fold and the final preparation was shown to be homogeneous by polyacrylamide and SDS gel electrophoresis and on isoelectric focusing. The molecular weight of the enzyme, determined by gel filtration and disc electrophoresis in the presence of SDS, was 220,000. The isoelectric pH was 7.42 and the amino acid composition is reported.-Mannosidase catalyzed the hydrolysis of both synthetic and natural substrates. The K_m of p-nitrophenyl -D-mannoside and -methyl D-mannoside were 0.695 mm and 71.9 mm at pH 4.0, the optimum pH. The natural substrates were hydrolysed to varying degrees. Zn²⁺ was not essential though it activated the enzyme activity over longer incubations. The enzyme was observed to be more stable at wider pH range in the presence of Zn²⁺ than in its absence. EDTA which did not affect the enzyme activity has effect on enzyme stability similar to Zn.²⁺ Seminal -mannosidase is not a zinc metalloenzyme but is activated by Zn²⁺.NDRI-publication no. 77-145.     

Adeno-associated virus (AAV) in semen and testis: a role in infertility? An overview

In the past decade, infection of the female genital tract with the ubiquitous adeno-associated viruses (AAV) has been reported by a number of authors. Though the virus is thought to be non-pathogenic, there were hints on an association of AAV with miscarriage and problems in pregnancy. In more recent years it has been discovered that there is also a rather frequent infection of male genital tissues supporting the hypothesis that this virus may be sexually transmitted and may play a role in infertility.     

Does cleansing of frozen-thawed bull semen before assessment provide samples that relate better to potential fertility?

The present study estimated, in vitro, the influence of two cleansing methods on sperm parameters post-thaw and their relation to the fertility of the frozen-thawed semen after AI. Frozen semen from six 1-year-old Swedish Red and White dairy bulls with a range in fertility (as 56d-Non-Return Rates, i.e., 56d-NRR) of 62.2-70.7% among batches was tested, using three batches of semen per bull. From each batch, individual straws were analyzed immediately after thawing (PT, control) or pooled and subjected to a swim-up procedure (SU) or washing by centrifugation/re-suspension (W) prior to in vitro assessments. Subjective and computerized measurements of sperm motility and of concentration, morphology, and membrane integrity were recorded. SU provided spermatozoa with significantly better motility, acrosome-, midpiece- and tail morphology and membrane integrity compared to either control or W treatment. Significant, albeit low, correlations among single sperm parameters and NRR were found (after PT for tail abnormalities (r = 0.49) and average path velocity, VAP (r = 0.47), after SU for total sperm motility with CASA (r = 0.50) and after W only for non-linear motility (r = -0.69)). SU of frozen-thawed bull semen is a simple preparation procedure that selects for sperm motility and membrane integrity, essential parameters for fertilization. It helps in vitro assessment of the semen and provides a significant, although low, relationship to the fertility of the assayed semen.