Sperm Quality Variables Research Articles

Predictive capacity of sperm quality parameters and sperm subpopulations on field fertility after artificial insemination in sheep

This study was designed to evaluate the relevance of several sperm quality parameters and sperm population structure on the reproductive performance after cervical artificial insemination (AI) in sheep. One hundred and thirty-nine ejaculates from 56 adult rams were collected using an artificial vagina, processed for sperm quality assessment and used to perform 1319 AI. Analyses of sperm motility by computer-assisted sperm analysis (CASA), sperm nuclear morphometry by computer-assisted sperm morphometry analysis (CASMA), membrane integrity by acridine orange-propidium iodide combination and sperm DNA fragmentation using the sperm chromatin dispersion test (SCD) were performed. Clustering procedures using the sperm kinematic and morphometric data resulted in the classification of spermatozoa into three kinematic and three morphometric sperm subpopulations. Logistic regression procedures were used, including fertility at AI as the dependent variable (measured by lambing, 0 or 1) and farm, year, month of AI, female parity, female lambing-treatment interval, ram, AI technician and sperm quality parameters (including sperm subpopulations) as independent factors. Sperm quality variables remaining in the logistic regression model were viability and VCL. Fertility increased for each one-unit increase in viability (by a factor of 1.01) and in VCL (by a factor of 1.02). Multiple linear regression analyses were also performed to analyze the factors possibly influencing ejaculate fertility (N=139). The analysis yielded a significant (P<0.05) relationship between sperm viability and ejaculate fertility. The discriminant ability of the different semen variables to predict field fertility was analyzed using receiver operating characteristic (ROC) curve analysis. Sperm viability and VCL showed significant, albeit limited, predictive capacity on field fertility (0.57 and 0.54 Area Under Curve, respectively). The distribution of spermatozoa in the different subpopulations was not related to fertility.

Successful ultrarapid cryopreservation of wild Iberian ibex (Capra pyrenaica) spermatozoa

A method for cryopreserving wild ibex sperm at high cooling rates was developed. To design a freezing solution based on Tris, citric acid, and glucose (TCG), two preliminary experiments were performed using glycerol (GLY) and dimethyl sulfoxide (DMSO) at different concentrations (5%, 10%, 20%). The 10% GLY + 10% DMSO combination reduced (P < 0.05) frozen-thawed sperm motility, which reached a minimum when 20% GLY + 20% DMSO was used. In the second experiment, sperm tolerance to three sucrose concentrations was evaluated (100-mM sucrose, 300-mM sucrose, 500-mM sucrose). Frozen-thawed sperm motility and sperm viability decreased (P < 0.05) at concentrations above 300 mM. The ultrarapid cooling procedure finally used involved a TCG egg yolk (ey)–based extender with 100-mM sucrose, either alone or with 5% GLY with or without BSA. Two warming procedures (37 °C vs. 60 °C) were also evaluated. The TCG ey with 100-mM sucrose but without GLY/BSA returned the best sperm quality variables. Slow warming at 37 °C strongly affected (P < 0.05) sperm motility and viability in all groups. Sperm selection by density gradient centrifugation produced no motile sperm when slow warming was performed. In contrast, when fast warming was used, sperm selection increased (P < 0.05) percentage of motility, viability, and the percentage of sperms with intact acrosomes. Heterologous in vivo fertilization involving domestic goats was performed to evaluate the in vivo fertilization capacity of the ultrarapidly cooled cryopreserved sperm (in TCG-ey + 100 mM sucrose), with warming undertaken at 60 °C. Inseminations of domestic goats resulted in three pregnancies (3 of 16, 18.7% fertility). In conclusion, ibex spermatozoa are strongly sensitive to high concentrations of permeable cryoprotectants and sucrose. However, the combination of ultrarapid cooling, using TCG-ey + 100-mM sucrose, and fast warming at 60 °C, followed by sperm selection by density gradient centrifugation to collect the motile sperm, has a positive effect on sperm viability.

Blood and seminal plasma concentrations of selenium, zinc and testosterone and their relationship to sperm quality and testicular biometry in domestic cats

The aim of this study was to assess seminal plasma (SP) and serum concentrations of zinc (Zn), selenium (Se) and testosterone (T) in domestic cats and determine whether these are related to sperm quality and testicular biometry. Six tomcats were collected using an artificial vagina and sperm analysis included motility by CASA, morphology, plasma membrane integrity, and sperm count. Serum and SP were submitted to total T concentration determination using a solid-phase radioimmunoassay technique while Zn and Se were measured by atomic absorption spectroscopy. Serum T concentrations were greater compared to SP concentrations, but both values were significantly correlated. Se concentrations were higher in serum, whereas SP had greater Zn values. Concentrations of Se, Zn and T were not correlated with each other either in serum or SP. Negative correlations were detected between Se concentrations in SP and total sperm head defects, and between Se concentrations in serum and VAP, VSL, STR, and LIN. Serum concentrations of Zn were negatively correlated with total abnormal sperm and midpiece defects and positively related to progressive motility. Both serum and SP concentrations of T had no relationship with sperm quality. Concentrations of Se exhibited a negative correlation with total testicular weight, whereas T concentrations in SP and serum were correlated with total testicular volume and weight. In conclusion, both Se and Zn concentrations in serum were correlated to sperm quality variables in the domestic cat, thus, making these potential candidates for fertility markers.

Effects of tetrabrombisphenol A on DNA integrity, oxidative stress, and sterlet (Acipenser ruthenus) spermatozoa quality variables

The sperm of sterlet (Acispenser ruthenus) was used to investigate the effect of the xenobiotic tetrabrombisphenol A (TBBPA) on sperm quality variables (ATP content, spermatozoa motility, and velocity), DNA integrity, and oxidative stress indices. Sperm was diluted to obtain a spermatozoa density of 5 × 10(8) cells/mL and exposed for 2 h to final concentrations of TBBPA (0.5, 1.75, 2.5, 5, and 10 μg/L). The oxidative stress indices, including lipid peroxidation, carbonyl derivatives of proteins, and antioxidant activity were significantly higher with increased concentrations of TBBPA. There was significantly less intracellular ATP in sperm samples at TBBPA concentrations of 2.5 μg/L and above. Spermatozoa velocity and percent motile sperm were significantly lower at each sampling time post-activation compared to controls. DNA damage expressed as percent DNA in Tail and Olive Tail moment was significantly higher with exposures ≥2.5 μg/L TBBPA. The results demonstrated that TBBPA and other xenobiotics can induce reactive oxygen species stress in fish spermatozoa, which could impair the sperm quality, DNA integrity, ATP content, and the antioxidant defense system. This study confirmed that fish spermatozoa can be used in in vitro assays for monitoring residual pollution in aquatic environments.

Effect of Short‐Term Scrotal Hyperthermia on Spermatological Parameters, Testicular Blood Flow and Gonadal Tissue in Dogs

The objective was to assess the effect of a short-term scrotal hyperthermia in dogs on quantitative and qualitative ejaculate parameters, testicular blood flow and testicular and epididymal histology. After a control period, the scrotum of seven normospermic adult beagle dogs was insulated with a self-made suspensory for 48h. Nine weeks later, two animals were castrated, while in five animals, scrotal hyperthermia was repeated. Dogs were castrated either 10 or 40days thereafter. In each phase of scrotal insulation, average scrotal surface temperature increased by 3.0°C. Semen was collected twice weekly throughout the experiment. Total sperm count did not change after the first hyperthermia, but it slightly decreased after the second (p<0.05). Profiles of sperm morphology and velocity parameters (CASA) rather indicated subtle physiological variations in sperm quality than effects of a local heat stress. Chromatin stability of ejaculated spermatozoa as indicated by SCSA remained constant throughout the experiment. Perfusion characteristics of the gonads, that is, systolic peak velocity, pulsatility and resistance index at the marginal location of the testicular artery, did not change due to hyperthermia (p>0.05). Histological examination of excised testes and epididymides for apoptotic (TUNEL and activated caspase-3) and proliferating cells (Ki-67 antigen) indicated only marginal effects of scrotal insulation on tissue morphology. In conclusion, a mild short-term scrotal hyperthermia in dogs does not cause substantial changes in sperm quantity and quality. In contrast to other species, canine testes and epididymides may have a higher competence to compensate such thermal stress.

Sperm Quality of Hatchery‐Reared Lake Trout Throughout the Spawning Season

AbstractThe objective of this study was to investigate variation in sperm quality metrics (motility, velocity, linearity, longevity, and density) of hatchery‐reared Lake Trout Salvelinus namaycush throughout the spawning season. Seasonal variation in sperm quality was investigated using both a regression and repeated‐measures approach. Sperm was collected from the same 16 individuals over four sampling periods, separated by 3‐week intervals. Regression analyses showed that 7–27% of the variation in sperm traits could be explained by seasonal variation, indicating that seasonality can have a significant impact on the quality of sperm. Significant positive linear relationships were found for percent motility and linearity at 5 s postactivation. Significant negative quadratic relationships were found for velocity at 5 s postactivation, longevity, and density, whereas a positive quadratic relationship was found for linearity at 10 s postactivation. Repeated measures ANOVAs showed a significant effect of season for percent motility and linearity at 5 and 10 s postactivation, velocity at 10 s postactivation, and longevity. Our findings are important for optimizing fertilization protocols for hatchery production and can also be used to understand reproductive biology and ecology of wild Lake Trout stocks.

How Multivariate Ejaculate Traits Determine Competitive Fertilization Success in Drosophila melanogaster

Success in sperm competition, occurring whenever females mate with multiple males, is predicted to be influenced by variation in ejaculate quality and interactions among competing sperm. Yet, apart from sperm number, relevant ejaculate characteristics and sperm-sperm interactions are poorly understood, particularly within a multivariate framework and the natural selective environment of the female reproductive tract. Here, we used isogenic lines of Drosophila melanogaster with distinguishable sperm to demonstrate and partition genetic variation in multiple sperm quality and performance traits. Next, by competing males from different lines, we show how rival sperm significantly influence each other's velocity and reveal that males with relatively slow and/or long sperm better displace rival sperm and resist displacement, thus avoiding ejection by the female from her reproductive tract. Finally, we establish fitness consequences of genetic variation in sperm quality and its role in securing a numerical advantage in storage by showing that offspring paternity is determined strictly by the representation of stored, competing sperm. These results provide novel insight into complex postcopulatory processes, illustrate that different ejaculate traits are critical at different biologically relevant time-points, and provide a critical foundation for elucidating the role of postcopulatory sexual selection in trait diversification and speciation.

Sperm lipid peroxidation is correlated with differences in sperm quality during the reproductive season in precocious European sea bass (Dicentrarchus labrax) males

Precocious maturation is a common feature in European sea bass males which occurs just before reaching the marketable size, reducing growth and feed conversion and representing an important problem that reduces the commercial value of fish. The sperm produced by precocious sea bass males could be useful for reproductive purposes since these males could be selected for breeders and used earlier as a new broodstock. However the characterization of sperm quality in these males during the reproductive season is fundamental in determining its possible use in reproduction or cryopreservation. Thus, in the present work, sperm from precocious European sea bass (Dicentrarchus labrax) males was characterized to determine the effect of seasonality on parameters influencing quality throughout the reproductive period. Several characteristics associated with sperm quality were determined: motility, cell concentration, sperm pH and osmolality, cell viability, lipid peroxidation (malondialdehyde-MDA), phospholipid, cholesterol and fatty acid composition of the plasma membrane. The results demonstrated seasonal variation in some of the parameters tested, the quality of samples being relatively poor at the beginning of the spawning season, corresponding with the highest content of MDA and the lowest motility parameters. Cell viability, phosphatidylcholine and phosphatidylethanolamine, polyunsaturated fatty acids n−3 and saturated fatty acids did not show significant differences during the reproductive period. However, monounsaturated fatty acids and polyunsaturated fatty acids n−6 showed a reduction at the end of the reproductive period. Parameter correlations showed that motility was negatively correlated with lipid peroxidation (−0.394, p<0.05), cholesterol/phospholipid ratio (−0.449, p<0.05) and osmolality (−0.478, p<0.01). Our results demonstrated that lipid peroxidation could explain the seasonal variation in sperm quality, since the highest malondialdehyde value quantified in November is the only parameter associated with the lowest motility recorded at the beginning of the season. Thus, approaches to reduce lipid peroxidation as a consequence of oxidative stress could be useful for improving sperm quality in precocious sea bass males. The parameters quantified are consistent with the interpretation that January is the optimal month in the spawning season to collect sperm from precocious sea bass males for further reproductive purposes.

Relationship among fluctuating asymmetry, morphological traits, and sperm quality in layers

The aim of the present study was to analyze the relationship between fluctuating asymmetry and the mean value of several bilateral traits (toe, leg, wing, and wattle lengths, and leg width) and sperm quality variables (volume, appearance, concentration, motility, morphological abnormalities, and acrosome and membrane integrities) in different breeds of layers. In total, 165 roosters were sampled at 36 wk of age. There was no significant correlation between fluctuating asymmetry and sperm quality variables. There was a positive association (P < 0.01) between sperm motility and concentration, and between sperm concentration and volume. Sperm motility was positively correlated (P < 0.001) with morphologically normal sperm. The membrane integrity did not correlate with motility. This study shows different results for the association between sperm quality variables and morphological traits. There was a negative association (P < 0.05) between toe length and membrane integrity and between leg length and sperm motility. In disagreement with the expected results, wattle length did not correlate with any sperm quality parameter. In general, roosters with longer legs had longer toes and wings, whereas those with longer wattles had longer toes, legs, and wings. Breed effect was significant for the sperm motility index (P < 0.001), concentration and membrane integrity (P < 0.05), and morphologically normal sperm (P < 0.01). Spanish breeds had similar sperm quality variables to the White Leghorn population, although the Blue Andaluza (a Spanish breed) showed smaller values for sperm motility index and concentration and membrane integrity. Breed effect was not significant for any fluctuating asymmetry, with values ranging from 1 to 6% of the trait value; however, breed effect was significant for all of the morphological traits (P < 0.001). The CV was higher for ejaculate volume and concentration, intermediate for semen appearance, motilities, and morphologically normal sperm, and smaller for membrane and acrosome integrity. In conclusion, the results suggest that fluctuating asymmetry is of limited use for assessment of rooster sperm quality. Some morphological traits (toe length, leg length, and leg width) might be associated with some sperm quality variables (motility and membrane integrity).

Relationship between sperm motility, morphology and the fertility of stallions

Sperm quality has an important role in determining fertility. Although there have been numerous studies to document the relationship between sperm quality and fertility, the methods of determining this association and conclusions vary. In the present study, computer-assisted sperm analysis (CASA) was used for evaluation of sperm motility, and differential interference contrast (DIC) microscopy was used for evaluating sperm morphologic features of breeding stallions. Fertility was measured using three endpoints: seasonal pregnancy rate (PR), percent pregnant/cycle (PC), and percent pregnant/first cycle (FCP). Increased total sperm motility (P = 0.08) and progressive path velocity (P = 0.06) tended to be associated with higher PR, whereas percent coiled tails (P = 0.02) was associated with a lower PR. Sperm motility variables associated with an increase in PC and FCP included total, progressive, and rapid sperm motility, and increased path and progressive velocity. Percent pregnant/first cycle was the only fertility measure able to discriminate among high, average, and low fertility groups, based on total and progressive sperm motility. Percent normal sperm was the only morphology variable associated with an increased PC and FCP, whereas increased levels of most sperm morphologic abnormalities (including abnormal and detached heads, proximal and distal droplets, general midpiece abnormality, and coiled tails) were associated with a decline in PC and FCP. Sperm quality variables most highly correlated with fertility included percent total sperm motility (PR, r = 0.37, P < 0.05; PC, r = 0.59, P < 0.05; and FCP, r = 0.64, P < 0.05), and percent morphologically normal sperm (PC, r = 0.42, P < 0.05; and FCP, r = 0.39, P < 0.05).

Seasonal variation in seminal parameters, testicular size, and plasma testosterone concentration in Iranian Moghani rams

The aim of the present study was to monitor seasonal variations in semen characteristics, testicular size, and plasma testosterone concentration in Iranian Moghani rams. An artificial vagina was used to collect semen from 10 rams (3–4 years of age), from November (autumn) 2008, to October (autumn) 2009. Testicular circumference was larger ( P < 0.05) during late summer (September) to early autumn (November). Ejaculate volume, sperm density, progressive motility and the percentage normal sperm on the other hand were higher ( P < 0.05) during autumn and lowest during winter ( P < 0.05). The plasma testosterone concentration was higher during autumn ( P < 0.05) than at any other time of the year, while the testicular circumference was positively correlated with the testosterone concentration, percentage live/normal sperm and semen concentration. The lowest level of lactate dehydrogenase (LDH) in the seminal fluid was recorded during early October (autumn) (1.52 U/mL) and the highest level in early spring (2.57 U/mL). The lowest levels of K + and Na + in the seminal fluid were recorded in spring (72.7 and 70.2 mg/dL, respectively). However, the variation of the highest levels was in late autumn (94.1 and 72.4 mg/dL, respectively), with the variation in the Na + concentration being much smaller than that of K +. The correlation coefficient of the percentage live/normal sperm and sperm motility with seminal fluid K + level ( r = 0.61 and 0.49, respectively) and Na + level ( r = 0.48 and 0.34, respectively) were positive, with the LDH correlation being negative ( r = −0.51 and −0.43, respectively) ( P < 0.01). In spite of monthly variations in sperm quality and testicular measurements, the semen quality of the Moghani rams was within the range regarded as satisfactory for normal fertility. It would seem as if the best quality semen for long-term preservation of Moghani sperm is produced during autumn.

Sperm Swimming Velocity Predicts Competitive Fertilization Success in the Green Swordtail Xiphophorus helleri

Sperm competition is expected to favour the evolution of traits that influence the performance of sperm when they compete to fertilize a female's eggs. While there is considerable evidence that selection favours increases in sperm numbers, much less is known about how sperm quality contributes towards competitive fertilization success. Here, we determine whether variation in sperm quality influences competitive fertilization success in the green swordtail Xiphophorus helleri, a highly promiscuous livebearing fish. We use artificial insemination as a method of controlled sperm delivery and show that sperm swimming velocity is the primary determinant of fertilization success when ejaculates from two males compete to fertilize a female's eggs. By contrast, we found no evidence that sperm length had any effect on siring success. We also found no evidence that pre- and postcopulatory sexual traits were phenotypically integrated in this species, suggesting that the previous observation that reproductive skew favours males with high mating rates is unlikely to be due to any direct association between sperm quality and male sexual ornamentation.

Effect of season on fresh and cryopreserved stallion semen

The objective of this study was to determine the effect of season on sperm quality variables, expression of the fertility-related protein SP22 and selected mRNA transcripts in fresh and cryopreserved stallion sperm. Four stallions were collected in each of the four seasons: summer, fall, winter and spring. Ejaculates were divided and then evaluated for motility, morphology, SP22 staining and expression of selected mRNAs as either fresh semen samples or cryopreserved samples. A significant interaction between season and cryopreservation status was found for total and progressive sperm motility. RNA yield from sperm was not affected by any variable examined. There was no effect of season or cryopreservation on the relative amounts of mRNA for PGK2, TPX1, TIMP3 or ACTB. There was a tendency ( P = 0.1) for an effect of stallion on the relative amount of ACTB mRNA. The proportion of sperm immunostained for SP22 over the equatorial segment was affected ( P < 0.05) by stallion. In addition, there was an interaction ( P < 0.05) between season and cryopreservation status on the percentage of sperm staining for SP22 on the equatorial segment. The correlation among total motility, progressive motility and SP22 immunostaining was much greater ( P < 0.05) during the breeding season (March and June) than during the non-breeding season (September and December). Based on data analyzed, semen collected in the Northern Hemisphere between March and June may be best suited for cryopreservation.

Seasonal functional relevance of sperm characteristics in equine spermatozoa

A group of stallions with different reproductive indexes were used to study seasonal variations in sperm quality (Equus caballus). Semen samples were collected from late September to July and analyzed according to four seasonal periods: late September–December, January–March, late March–May, and June–July. Parameters monitored included sperm concentration, sperm motility, sperm morphology, sperm viability, acrosomal status, plasma membrane stability, and sperm mitochondrial membrane potential. Overall, seminal parameters monitored are affected mostly by time period, followed by animal and lastly by fertility, stressing the importance of individual variations in out-bred animal models. The analysis of multiple ejaculates from the same animals showed clear seasonal-based differences (P < 0.05) with poor semen quality in winter and a noticeable improvement in sperm quality with increasing photoperiod. Better semen quality was observed between late March and May. Interactions between month period, animal, and fertility were evident (P < 0.05) for sperm concentration, head and tail sperm anomalies, and acrosomal integrity. Thus, it may be advisable to adjust the use of stallion semen according to seasonal variations.

Sperm viability staining in ecology and evolution: potential pitfalls

The causes and consequences of variation in sperm quality, survival and ageing are active areas of research in ecology and evolution. In order to address these topics, many recent studies have measured sperm viability using fluorescent staining. Although sperm viability staining has produced a number of interesting results, it has some potential pitfalls that have rarely been discussed. In the present paper, I review the major findings of ecology and evolution studies employing sperm viability staining and outline the method’s principle limitations. The key problem is that the viability assay may itself kill sperm, which is likely to confound many common experimental designs in addition to producing artificially low estimates of sperm viability. I further suggest that sperm number should be routinely measured in sperm viability studies, as it may be an important but overlooked source of spurious results. I provide methodological advice on sperm viability staining aimed at minimising artefacts and producing robust conclusions, and discuss possible avenues for future research.

Sperm Quantity Explains Age‐Related Variation in Fertilization Success in the Hide Beetle

AbstractAge‐related variation in sperm quality and quantity may have a dramatic impact on female fecundity and fertilization success. Despite this, the relationships between these parameters are rarely investigated. Moreover, studies exploring age‐related variation in male mating success generally fail to consider the entire lifespan of an individual male; instead they restrict analyses to a small number of defined age classes. As a consequence they are unable to assess the impact of early and late life history components on male reproductive success. In this study, we explore these questions using the hide beetle, Dermestes maculatus, a model species for investigating age‐related male mating success. We first employed a longitudinal design to explore whether patterns of sperm transfer and subsequent reproductive success varied over the reproductive lives of a cohort of males. Secondly, we investigated age‐related variation in sperm viability, a surrogate measure of sperm quality. Finally, we combined these data and assessed whether the observed patterns of sperm transfer were correlated with fertilization success. We found that the quantity of sperm varied with male age: the amount of sperm transferred to a female increased with male age until 9 wk and then started to decline. Similarly, female fecundity and fertilization success were related to male age: females mating with males when they were relatively young (1 wk) or relatively old (13 wk) suffered reductions in fecundity and fertilization. Our data suggest that fertilization success is driven at least in part by the quantity of sperm transferred during mating.

Collection and evaluation of semen from the three-toed sloth ( Bradypus tridactylus)

Sloths ( Bradypus sp.) are extremely sensitive animals that suffer with the destruction and fragmentation of forests. They present a low population growth rate and need to be further studied for the preservation of the specie. Thus, the aim of this study was to establish an efficient semen collection protocol as well as characterize sperm concentration, motility and morphology in order to contribute with information about the reproductive traits of this specie, which has never been described in the literature before. For that, nine Bradypus tridactylus males were captured during the wet season and six during the dry season, in Manaus (AM), located in the north region of Brazil, semen was collected by electroejaculation with shocks given in sequences of progressive intensities (minimum 20 mA and maximum 60 mA). All animals ejaculated small volumes of semen and in some of them, the volume ejaculated was not enough for a complete spermiogram. Physical characteristics observed on the collections of the wet season were different from those seen in the specimen collected in the dry season. Motility an vigor was very low and did not show forward progression, only oscillatory movement. After Spermac stain, spermatozoa presented a wide variety of defects; however, the differences in morphology were not significant between seasons. The morphology assessed by scanning electron microscopy shows that the head in both groups could be elongated, short or could have a base narrower than the apex and the midpiece narrowed abruptly, forming a nip in its transition to the tail. Although further studies are necessary to verify our preliminary findings concerning seasonal variation in sperm quality, these results demonstrate that semen can be safely collected from sloths by electroejaculation and provide the first reports of semen characteristics in this species.

Sperm morphology and chromatin integrity in Swedish warmblood stallions and their relationship to pregnancy rates

BackgroundArtificial insemination is not as widely used in horses as in other domestic species, such as dairy cattle and pigs, partly because of the wide variation in sperm quality between stallion ejaculates and partly due to decreased fertility following the use of cooled transported spermatozoa. Furthermore, predictive tests for sperm fertilising ability are lacking. The objective of the present study was to assess sperm morphology and chromatin integrity in ejaculates obtained from 11 warmblood breeding stallions in Sweden, and to evaluate the relationship of these parameters to pregnancy rates to investigate the possibility of using these tests predictively.MethodsAliquots from fortyone ejaculates, obtained as part of the normal semen collection schedule at the Swedish National Stud, were used for morphological analysis by light microscopy, whereas thirtyseven were used for chromatin analysis (SCSA) by flow cytometry. The outcome of inseminations using these ejaculates was made available later in the same year.ResultsRanges for the different parameters were as follows; normal morphology, 27–79.5%; DNA-fragmentation index (DFI), 4.8–19.0%; standard deviation of DNA fragmentation index (SD_DFI) 41.5–98.9, and mean of DNA fragmentation index (mean_DFI), 267.7–319.5. There was considerable variation among stallions, which was statistically significant for all these parameters except for mean_DFI (P < 0.001, P < 0.01, P < 0.001 and P < 0.2 respectively). There was a negative relationship between normal morphology and DFI (P < 0.05), between normal morphology and SD_DFI (P < 0.001), and between normal morphology and mean_DFI (P < 0.05). For specific defects, there was a direct relationship between the incidence of pear-shaped sperm heads and DFI (P < 0.05), and also nuclear pouches and DFI (P < 0.001), indicating that either morphological analysis or chromatin analysis was able to identify abnormalities in spermiogenesis that could compromise DNA-integrity. A positive relationship was found between normal morphology and pregnancy rate following insemination (r = 0.789; P < 0.01) and a negative relationship existed between DFI and pregnancy rate (r = -0.63; P < 0.05). Sperm motility, assessed subjectively, was not related to conception rate.ConclusionEither or both of the parameters, sperm morphology and sperm chromatin integrity, seem to be useful in predicting the fertilising ability of stallion ejaculates, particularly in determining cases of sub-fertility.

Factors influencing boar sperm cryosurvival1

Optimal sperm cryopreservation is a prerequisite for the sustainable commercial application of frozen-thawed boar semen for AI. Three experiments were performed to identify factors influencing variability of postthaw sperm survival among 464 boar ejaculates. Sperm-rich ejaculate fractions were cryopre-served using a standard freezing-thawing procedure for 0.5-mL plastic straws and computer-controlled freezing equipment. Postthaw sperm motility (assessed with a computer-assisted semen analysis system) and viability (simultaneously probed by flow cytometry analysis after triple-fluorescent stain), evaluated 30 and 150 min postthaw, were used to estimate the success of cryopreservation. In the first experiment, 168 unselected ejaculates (1 ejaculate/boar), from boars of 6 breeds with a wide age range (8 to 48 mo), were cryopreserved over a 12-mo period to evaluate the predictive value of boar (breed and age), semen collection, transport variables (season of ejaculate collection, interval between collections, and ejaculate temperature exposure), initial semen traits, and sperm quality before freezing on sperm survival after freezing-thawing. In Exp. 2, 4 ejaculates from each of 29 boars, preselected according to their initial semen traits and sperm quality before freezing, were collected and frozen over a 6-mo period to evaluate the influence of interboar and intraboar ejaculate variability in the survival of sperm after cryopreservation. In Exp. 3, 12 ejaculates preselected as for Exp. 2, from each of 15 boars with known good sperm cryosurvival, were collected and frozen over a 12-mo period to estimate the sustainability of sperm cryosurvival between ejaculates over time. Boar and semen collection and transport variables were not predictive of sperm cryosurvival among ejaculates. Initial semen traits and sperm quality variables observed before freezing explained 23.2 and 10.9%, respectively, of the variation in postthaw sperm motility and viability. However, more that 70% of total variance observed in postthaw sperm quality variables among ejaculates was explained by boar. This indicates that boar is the most important (P < 0.001) factor explaining the variability among ejaculates in sperm cryosurvival, with most (14 of the 15 boars in Exp. 3) showing consistent (P > 0.05) sperm cryosurvival over time.

Sperm quality influences male fertilization success in walleye (Sander vitreus)

We examined how variation in sperm quality influences a male's success at fertilizing ova (male fertilization success) in a wild population of walleye (Sander vitreus). To do this, we conducted controlled fertilization trials using milt and eggs (ova) from wild-spawning fish and measured male fertilization success (percentage of ova fertilized) by examining eggs after 24 h of incubation. We found that both the number of sperm and sperm swimming speed (at 10 s after activation) were significantly related to fertilization success. There was, with respect to fertilization success, a relatively large return on male investment in the number of sperm, but this return diminished as the percentage of ova fertilized increased above 50%. This is in agreement with theoretical predictions based on external fertilization dynamics. When the number of sperm used in the experimental trials was kept constant, variation in sperm swimming speed (at 10 s after activation) explained approximately 90% of the variation in a male's fertilization success. These findings demonstrate that the variation in sperm quality found in wild spawning populations has the potential to dramatically influence male reproductive success.