Proportion Of Motile Spermatozoa Research Articles

The impact of diet quality on the velocity, morphology and normality of sperm in the zebra finch Taeniopygia guttata

ABSTRACTSperm traits can influence fertilisation success, but there is still much we do not understand about sperm condition dependence, that is, how much sperm traits depend on the male's energy acquisition and allocation. This is especially pronounced in avian taxa, despite extensive observational studies and sampling in wild populations. In this study, we collected sperm samples before and after experimentally reducing diet quality of wild-derived captive zebra finches in small mixed-sex groups, which we compared with individuals on a control diet. We measured the length of sperm components (head, midpiece, flagellum and total sperm length), the proportion of sperm with normal morphology, the proportion of sperm that were progressively motile and sperm swimming velocity (curvilinear velocity; VCL). The only sperm trait we found to be impacted by reduced diet quality was a significant decrease in sperm midpiece length. This is consistent with emerging evidence in other non-model systems, as well the fact that diet can alter mitochondrial density and structure in other tissue types. There was also a significant decrease in sperm velocity and the proportion of motile sperm over the course of the experiment for both experimental groups (i.e. unrelated to diet). This decrease in sperm velocity with largely unchanged sperm morphology emphasizes that there are other important determinants of sperm velocity, likely including seminal fluid composition.

Black Sea turbot sperm motility depending on the dilution of seawater

The sperm motility of fish reflects their fertilizing ability. Sperm activation in fish with external fertilization occurs in an isotonic medium (sea or freshwater). The duration of sperm motility is a compromise between the level of energy reserves possessed by the sperm and the process of osmotic damage it experiences in the activating medium. Under natural conditions, various biotic and abiotic factors can affect the characteristics of motility. At the same time, when assessing the reproductive potential of males in laboratory conditions, the dilution and methods of its activation have a significant effect on the characteristics of sperm quality (namely, speed, the proportion of motile spermatozoa, and the time of their activity). In our work, we determined the main characteristics of the motility of the Black Sea turbot sperm at a dilution of 1:10 and 1: 100 with seawater. We have shown that, the average speed and percentage of motile sperm significantly decrease with higher dilution of seawater.

Supplementing extender with anandamide enhances quality of low sperm doses during cryopreservation in bulls.

The present study explored the effect of anandamide supplementation in the extender on quality of low sperm doses during cryopreservation in Sahiwal bulls. Each fresh semen sample was split into eight aliquots (I, II, III, IV, V, VI, VII and VIII). The aliquots I, II, III and IV were taken as control and diluted to 20, 15, 10 and 5 million spermatozoa/0.25ml respectively. The aliquots V, VI, VII and VIII were diluted with extender (supplemented with anandamide at 1µM/ml of extender) to 20, 15, 10 and 5 million spermatozoa/0.25ml respectively. This was followed by filling of diluted semen into French mini straws, equilibrated at 4°C of 4hr and cryopreserved. The results revealed that the proportions of motile spermatozoa, live spermatozoa and live acrosome intact spermatozoa were significantly (p<.05) higher in all anandamide-treated sperm doses compared to control. The proportions of moribund spermatozoa, dead acrosome intact spermatozoa and capacitated spermatozoa were significantly (p<.05) reduced in all anandamide-treated sperm doses compared to control, with no difference in proportion of dead acrosome-reacted spermatozoa. In conclusion, anandamide supplementation in the extender increases the post-thaw quality of low sperm doses during cryopreservation in bulls.

Post-glacial establishment of locally adapted fish populations over a steep salinity gradient.

Studies of colonization of new habitats that appear from rapidly changing environments are interesting and highly relevant to our understanding of divergence and speciation. Here, we analyse phenotypic and genetic variation involved in the successful establishment of a marine fish (sand goby, Pomatoschistus minutus) over a steep salinity drop from 35 PSU in the North Sea (NE Atlantic) to two PSU in the inner parts of the post-glacial Baltic Sea. We first show that populations are adapted to local salinity in a key reproductive trait, the proportion of motile sperm. Thereafter, we show that genome variation at 22,190 single nucleotide polymorphisms (SNPs) shows strong differentiation among populations along the gradient. Sequences containing outlier SNPs and transcriptome sequences, mapped to a draft genome, reveal associations with genes with relevant functions for adaptation in this environment but without overall evidence of functional enrichment. The many contigs involved suggest polygenic differentiation. We trace the origin of this differentiation using demographic modelling and find the most likely scenario is that at least part of the genetic differentiation is older than the Baltic Sea and is a result of isolation of two lineages prior to the current contact over the North Sea-Baltic Sea transition zone.

Comparative Evaluation of Different Methods for Assessing Stallion Sperm Membrane Integrity

Comparative evaluation of three different methods for assessing stallion sperm-membrane integrity, including the eosin staining technique, the fluorochrome staining (SYBR-14 + PI) method, and the hypo-osmotic test, has been made in order to reveal the most efficient method of measurement for this item. The stallion sperm showed high coefficients of correlation between sperm motility and intact membrane proportion with all the methods of measurement. The best coincidence result was attained with the eosin staining technique, assessed by a correlation coefficient of 0.91. This technique has the advantages of simplicity, accessibility, and efficiency. Accurate assessment requires performing a method of subtracting a number of unstained damaged sperm cells, which are nonfunctional, from a total number of unstained spermatozoa. The proportion of motile sperm does not coincide with the proportion of sperm intact membranes during the stallion-semen storage process according to all the methods of measurement. A significant sperm count remains with intact cell membranes at the loss of total motility at the end of a storage process. This indicates that the sperm cells become immotile because of layout of the energetic material in flagellar mitochondria, which is not associated with membrane integrity. Therefore, it is recommended to assess the stallion sperm membranes with the use of fresh or thawed sperm. It is suggested to use assessment of membrane integrity in clarifying the sperm motility, particularly, in especially doubtful cases.

Single layer colloid centrifugation technique improves motility, viability and chromatin integrity of ram spermatozoa after thawing

The cell membrane of ram spermatozoa is more sensitive to the freezing process than in other species due to its composition. As a result, the quality and viability of frozen thawed ram spermatozoa are often poor, which together with the specific structure of the ewe's cervix are the main reasons for lower fertility in ewes after intracervical insemination. In the present study we investigated the effects of semen centrifugation through a single layer of a species-specific colloid (Androcoll-O) on post-thaw quality of ram spermatozoa. Motility, viability and morphology were analysed 0, 6, 12 and 24 h after thawing. DNA fragmentation index (%DFI) of the samples was assessed 0 h after thawing, by SCSA™. Membrane and acrosome integrity of spermatozoa were analysed by Sybr-14/PI/PNA test 0 h after thawing.The proportion of motile spermatozoa was significantly higher in SLC – selected samples in comparison to control (not SLC – selected) samples at 0, 6, 12 (P < 0.001) and 24 h (P < 0.05). The proportion of viable spermatozoa was also significantly higher in SLC - selected samples in comparison to control samples at all times (P < 0.001). The proportion of abnormal acrosomes and morphologically abnormal spermatozoa (MAS) were significantly lower in SLC – selected samples compared to control samples at all times (P < 0.001). Analysis of chromatin stability revealed significantly lower %DFI values in SLC – selected samples compared to control samples (P < 0.001). The SYBR-14/PI/PNA test also revealed significantly better values in SLC – selected compared to control samples (P < 0.05). In conclusion, single layer colloid centrifugation significantly improved post-thaw quality and longevity of ram spermatozoa, making it suitable for artificial insemination initiatives.

Rival seminal fluid induces enhanced sperm motility in a polyandrous ant

BackgroundPromiscuous mating and sperm competition often induce arms races between the sexes with detrimental outcomes for females. However, ants with multiply-inseminated queens have only a single time-window for sperm competition and queens are predicted to gain control over the outcome of sperm storage quickly. The seminal fluid of Acromyrmex leaf-cutting ants reduces the viability of rival sperm, but how confrontations between unrelated ejaculates affect sperm storage remains unknown.ResultsWe investigated the effects of ejaculate admixture on sperm motility in A. echinatior and found that the proportion of motile spermatozoa, sperm swimming speed, and linearity of sperm movement increased when rival ejaculates were mixed in vitro. Major effects induced by the seminal fluid of rival males were of similar magnitude to those generated by queen reproductive tract secretions, whereas own seminal fluid induced lower sperm activation levels.ConclusionsOur results suggest that ant sperm respond via a self–non-self recognition mechanism to similar or shared molecules expressed in the reproductive secretions of both sexes. Lower sperm motility in the presence of own seminal fluid indicates that enhanced motility is costly and may trade-off with sperm viability during sperm storage, consistent with studies in vertebrates. Our results imply that ant spermatozoa have evolved to adjust their energetic expenditure during insemination depending on the perceived level of sperm competition.

207 Investigating the Cryotolerance of Boar Spermatozoa Subjected to Prior Selection

The proportion of non-viable spermatozoa in insemination semen doses poses limitations to breeding. Recent studies have shown the potential of removing moribund spermatozoa from semen doses through their interactions with specifically designed magnetic nanoparticles (MNP). This specific retrieval leads to substantial enrichment of insemination doses with robust spermatozoa (nanoselected), which would have advantageous effects on the animal husbandry industry. Here, we explored the potential of these MNP for semen dose enrichment and freezability improvement. The MNP were synthesised to target acrosome reacted (lectin) and apoptotic (annexin-V) spermatozoa. Freshly harvested semen of at least 2 boars were pooled and extended in a commercial diluent at a local stud. Extended semen (40 mL) were mixed with varying amounts of MNP (0, 87.5, and 175 μg) and incubated at 37°C for 60 min. Thereafter, the tube mixtures were placed under a magnetic field to trap moribund spermatozoa bound to MNP. The intact or MNP-free (nanoselected) spermatozoa were eluted by pouring the untrapped spermatozoa into new tubes. Both fresh non-selected (control) and nanoselected spermatozoa were mixed with the cryoguard-cooling extender and equilibrated for 120 min at 4°C, followed by the addition of an equal volume of cryoguard-freezing extender. Spermatozoa were frozen according to the preset, porcine protocol (4 to 1°C in 1.3 min, to −10°C in 2.36 min, to –30°C in 3.54 min, and to –140°C in 14.56 min), using the IceCube 14S-B freezer. Frozen spermatozoa were stored at –196°C until analyses. Sperm motion characteristics were assessed before and after freezing (computer-assisted sperm analyzer), and their quality was evaluated post-thaw through plasma and acrosome membrane integrities (flow cytometry). Data were analysed (ANOVA-1), and P &lt; 0.05 indicated significant difference. Motion analyses revealed dose-dependent and significantly increased proportions of motile (75 ± 5%, 77 ± 3%, and 84 ± 2%) and progressive (44 ± 4%, 49 ± 2%, and 64 ± 5%) spermatozoa, following exposure to MNP (0, 87.5, 175 μg, respectively; P &lt; 0.05). The proportions of nanoselected spermatozoa with bent tails and distal droplets decreased significantly (P &lt; 0.05). Freeze-thawing led to poor proportions of motile spermatozoa (&lt;10%), regardless of the MNP concentrations. Control and nanoselected spermatozoa had similar post-thaw motility (P &gt; 0.05). Flow cytometry analyses indicated comparable proportions of spermatozoa with intact membranes (P &gt; 0.05). However, fluorescence intensities of staining dyes were decreased in nanoselected spermatozoa, indicating possible reduced sensitivity to further insults. We concluded that (1) the designed MNP allowed for high-throughput semen enrichment with highly motile spermatozoa, (2) nanoselection was more likely beneficial to semen with poor sperm motility (&lt;70%), and (3) nanoselected spermatozoa may have greater robustness that did not show through cryotolerance. Our freeze-thawing protocol is under optimization for further analyses of nanoselected spermatozoa. This research was supported by USDA-ARS Grant# 58-6402-3-018.

Environmental Tipping Points for Sperm Motility, Fertilization, and Embryonic Development in the Crown-of-Thorns Starfish

For broadcast spawning invertebrates such as the crown-of-thorns starfish, early life history stages (from spawning to settlement) may be exposed to a wide range of environmental conditions, and could have a major bearing on reproductive success and population replenishment. Arrested development in response to multiple environmental stressors at the earliest stages can be used to define lower and upper limits for normal development. Here, we compared sperm swimming speeds and proportion of motile sperm and rates of fertilization and early development under a range of environmental variables (temperature: 20–36 °C, salinity: 20–34 psu, and pH: 7.4–8.2) to identify environmental tipping points and thresholds for reproductive success. We also tested the effects of water-soluble compounds, derived from eggs, on sperm activity. Our results demonstrate that gametes, fertilization, and embryonic development are robust to a wide range of temperature, salinity, and pH levels that are outside the range found at the geographical limits of adult distribution and can tolerate environmental conditions that exceed expected anomalies as a result of climate change. Water-soluble compounds derived from eggs also enhanced sperm activity, particularly in environmental conditions where sperm motility was initially limited. These findings suggest that fertilization and embryonic development of crown-of-thorns starfish are tolerant to a wide range of environmental conditions, though environmental constraints on recruitment success may occur at later ontogenic stages.

Computer assisted sperm analysis: Relationship between the movement characteristics of buffalo spermatozoa and sire fertility

The present study was undertaken to identify the differences in sperm kinematics between buffalo bulls with different fertility ratings. Murrah buffalo bulls (n=9) that were routinely used for breeding purpose under progeny testing programme were utilized for the study. Bull fertility was determined based on in vivo fertility trials and the conception rates (CR) were adjusted for different non-genetic parameters. Based on the adjusted CR, bulls were classified into high, medium and low fertile group. Frozen semen samples of these bulls were obtained and sperm kinematic parameters were assessed using a computer assisted sperm analyzer. The kinematic parameters analyzed included the curvilinear velocity (VCL), the linear velocity (VSL), the average path velocity (VAP), the amplitude of lateral head displacement (ALH), the linearity (LIN), the straightness coefficient (STR) and the beat cross frequency (BCF). In high fertile bulls, the proportion of motile spermatozoa was higher (p&lt;0.001) than the medium and low fertile bulls. The VAP and VCL of sperm motion were significantly higher (P&lt;0.05) in high fertile bulls compared to either medium or low fertile bulls. The VSL was significantly lower in low fertile bulls (P&lt;0.005) compared to either high or medium fertile bulls. Spermatozoa from high fertile bulls had significantly higher (P&lt;0.05) BCF, STR, ALH and LIN compared to either medium or low fertile bulls. Buffalo bull fertility was significantly and positively correlated with sperm motility, VAP, VSL, VCL and ALH.

A multilaboratory study on the variability of bovine semen analysis

To evaluate the variability of semen analysis, five replicates of 10 different bovine frozen semen batches were coded with different identification numbers and submitted to various laboratories for evaluation. Three studies were conducted: study I included eight laboratories in semen processing centers in the United States; study II included one laboratory in one semen processing center and five veterinary university laboratories in the United States; and study III included five veterinary university laboratories in Brazil. Evaluation methodology, sample classification criteria, and reporting format varied considerably among laboratories. There were laboratory effects (P < 0.05) on sperm concentration, motility, and morphology results in all studies. When Bland–Altman plots were evaluated, differences in sperm concentration were approximately between −5 and +5 × 106 sperm/mL in study I, when the same method of evaluation was used by all laboratories but ranged between −30 and +30 × 106 sperm/mL in studies II and III. Differences in the proportions of motile sperm were approximately −30% to +30%, and differences in the proportion of normal sperm were −15% to +15% in studies I and II; these differences were −15% to +15% and −10% to +10%, respectively, in study III. Mean absolute (one tail) proportional differences in estimates across all laboratories ranged from 9% to 31%, 16% to 37%, and 9% to 14% for sperm concentration, motile sperm, and normal sperm across studies; much larger (48%–86%) differences were observed for sperm abnormality categories. Intralaboratory and interlaboratory precision varied considerably across laboratories and seemed to be at least in part related to methods used for evaluation; precision was better when the NucleoCounter was used for evaluation of sperm concentration, whereas the use of computer-assisted sperm analysis for evaluation of sperm motility resulted in greater precision in some but not all laboratories. None of the laboratories that classified samples as satisfactory or unsatisfactory achieved complete consistency for all replicates within all batches. In addition, consistent classification among laboratories was observed for just three batches in studies II and III. These observations put the reliability of semen analysis in check and make it very difficult, if not impossible, to meaningfully interpret evaluation results.

Identification and Characterization of an Oocyte Factor Required for Porcine Nuclear Reprogramming

Nuclear reprogramming of somatic cells can be induced by oocyte factors. Despite numerous attempts, the factors responsible for successful nuclear reprogramming remain elusive. In the present study, we found that porcine oocytes with the first polar body collected at 42 h of in vitro maturation had a stronger ability to support early development of cloned embryos than porcine oocytes with the first polar body collected at 33 h of in vitro maturation. To explore the key reprogramming factors responsible for the difference, we compared proteome signatures of the two groups of oocytes. 18 differentially expressed proteins between these two groups of oocytes were discovered by mass spectrometry (MS). Among these proteins, we especially focused on vimentin (VIM). A certain amount of VIM protein was stored in oocytes and accumulated during oocyte maturation, and maternal VIM was specifically incorporated into transferred somatic nuclei during nuclear reprogramming. When maternal VIM function was inhibited by anti-VIM antibody, the rate of cloned embryos developing to blastocysts was significantly lower than that of IgG antibody-injected embryos and non-injected embryos (12.24 versus 22.57 and 21.10%; p < 0.05), but the development of in vitro fertilization and parthenogenetic activation embryos was not affected. Furthermore, we found that DNA double strand breaks dramatically increased and that the p53 pathway was activated in cloned embryos when VIM function was inhibited. This study demonstrates that maternal VIM, as a genomic protector, is crucial for nuclear reprogramming in porcine cloned embryos.

Post-testicular sperm maturation and identification of an epididymal protein in the Japanese quail (Coturnix coturnix japonica)

The role of the avian epididymis in post-testicular development and capacitation was examined to assess whether avian spermatozoa undergo any processes similar to those characteristic of mammalian sperm development. We found no evidence of a need for quail sperm to undergo capacitation and 90% of testicular sperm could bind to a perivitelline membrane and acrosome react. However, computer-assisted sperm analysis showed that 20% of testicular sperm from the quail were capable of movement and only about 12% of the motile sperm would have a curvilinear velocity greater than the mean for sperm from the distal epididymis. Nevertheless, epididymal transit was associated with increases in mean sperm velocity and the proportion of motile sperm. Together, these findings explain why earlier workers have achieved some fertilizations following inseminations of testicular spermatozoa and also demonstrate the need for some epididymal maturation of avian spermatozoa. Analysis of the electrophoretic profile of quail epididymal luminal proteins revealed that only one major protein (∼16 kDa) is secreted by the epididymis and it was virtually the only protein secreted by the ipsilateral epididymis following unilateral orchidectomy. Mass spectrometry showed that this protein is hemoglobin; this finding was confirmed using anti-hemoglobin antibodies. It is suggested that hemoglobin may support sperm metabolism in the quail epididymis, aid in motility, and/or serve as an antioxidant.

Combined n-3 Fatty Acids and α-Tocopherol Supplementation Improved the Ovine Sperm Cryosurvival

Background: Spermatozoa preservation is an approach to improve the fertility rate to pass on the valuable genetic material from sire to their offspring. During the last 20 years, reproductive biotechnologists have focused on the approaches that improve spermatozoa cryosurvival. One of the possible mechanisms is supplementation in semen extender. Objectives: The aim of this study was to evaluate the combined effects of healthy ram semen with n-3 fatty acids and α-tocopherol (Vitamin E) on freezing ability and fatty acid (FA) content of sperm cell. Materials and Methods: Semen collection was performed on six mature Zandi rams by an artificial vagina. In the present study, two experiments were carried out. In Experiment 1, the specimen quality was assessed. Then the samples were pooled. The pooled specimens were allocated into 12 groups, in a 3 × 4 factorial design, including four levels of n-3 FA (0, 0.1, 1, 10 ng.mL -1) and three levels of α-tocopherol (0. 0.1, 0.2 mM). Then sperm critical characteristics such as proportion of motile sperm, progressive motile sperm, viable and abnormal sperms were measured. Furthermore, after freezing-thawing procedure, the recovery rate was considered as a vital indicator of semen quality. After thawing, the highest progressive motility was obtained when treated with 0.1 mM α-tocopherol and 1 ng.mL -1 n-3 FA. So, the second experiment was designed to measure the content of FA in specimens that fortified with 0.1 mM α-tocopherol, 1 ng.mL -1 n-3 FA and also in groups without α-tocopherol and FA. Results: The data showed that before freezing, docosahexaenoic acid (DHA) level of sperm was increased when the FA introduced into extender (P ≤ 0.01). On the other hand, in the FA group, the n-3 FA and polyunsaturated fatty acid content were significantly higher compared with n-6 FA and saturated fatty acid level. However, in other groups, there were no significant alteration in the overall proportion of n-3 FA and n-6 FA were recorded (P ≤ 0.01). Conclusion: It was concluded that the cryosurvival of ram semen could be improved by adding DHA along with alfa-tocopherol as an antioxidant.

Functional activity of adenylyl and guanylyl cyclases in human spermatozoa with different motility

The most important functional characteristics of ejaculated spermatozoa is their ability to directional sustained movement that to a large extent determines their fertility. It is assumed that the enzymes with cyclase activity--adenylyl cyclase (AC) and guanylyl cyclase (GC), soluble and membrane-bound forms of which are found in human and mammalian sperm, play a key role in the regulation of motility. However, the functional activity of the cyclases in ejaculated spermatozoa with different motility and their contribution to the regulation of this process is virtually unexplored. The aim of this work was the functional characteristics of AC and GC in human sperm ejaculates with different content of motile forms of spermatozoa, and the study of regulation of these enzymes by hormones and non-hormonal agents. We found the differences in the activity and regulatory properties of AC and GC in the ejaculates differing in the content of motile forms of sperm. The basal AC activity and its sensitivity to bicarbonate anion and manganese cations, activators of cytosolic AC (cAC), were increased in ejaculates with a high proportion of motile spermatozoa. At the same time, AC effect of forskolin, GppNHp and adrenergic receptor agonists acting via membrane-bound AC (mAC) were significantly reduced in this case. Cytosolic GC in the ejaculates with a high proportion of motile spermatozoa was more sensitive to manganese cations, but the basal activity of GCs was altered slightly. The increase in the content of motile spermatozoa in the ejaculates led to decrease in the sensitivity of receptor GC to CNP, while the sensitivity to ANP was maintained, which indicated a change in the pattern of enzyme regulation with natriuretic peptides in favor of ANP, an important regulator of sperm chemotaxis. Thus, we have concluded that the change in the proportion of motile spermatozoa in the ejaculate induces changes in the functional activity and regulatory properties of soluble and membrane-bound forms of AC and GC, which can be used to control the motility, chemotaxis, acrosomal reaction and other processes determining the fertility of male germ cells.

Spectrophotometric determination of sperm concentration and short-term cold-storage of sperm in Atlantic croakerMicropogonias undulatusL. broodstock

The objective of this study was to optimize the methodology for spectrophotometric determination of sperm concentration in Atlantic croaker Micropogonias undulatus L. milt and to estimate its potential for short-term cold-storage. The spectrophotometric determination of sperm concentration was evaluated using milt samples from six males serially diluted in Hank's balanced salt solution at 200 mOsm kg−1 (HBSS). The predictive power of regression models between sperm concentration and absorbance was determined from 200 to 500 nm and found to be highest within the visible spectrum despite a peak of milt absorbance at 288 nm. Absorbance reading at 400 nm was selected for further analysis to maximize the absorbance of the sample hence the sensitivity of the method while minimizing the impact of potential sample contamination with blood. The standard-curve of correlation between sperm absorbance at 400 nm and concentration was validated and held an accuracy ranging between −7.40% and +4.56% across males. Total sperm motility duration and the proportion of motile spermatozoa were significantly higher in milt samples diluted 1:3 in HBSS than in the undiluted control during up to 30 h of cold-storage.The methodologies investigated in this study can be applied to optimize sperm usage and achieve predictable artificial fertilization protocols in Atlantic croaker.

Comparison of assessment of pigeon sperm viability by contrast-phase microscope (eosin-nigrosin staining) and flow cytometry (SYBR-14/propidium iodide (PI) staining) [evaluation of pigeon sperm viability

The aim of these experiments was to compare the conventional, microscopic method of evaluating pigeon sperm viability to sperm assessed by flow cytometry. Semen was collected twice a week from two groups of pigeons. In every group were 20 males (Group I: meat-type breed; Group II: fancy pigeon breed). Semen was collected using the lumbosacral and cloacal region massage method. Ejaculates collected from each group were pooled and diluted to 10 × 10 6 sperm/ml in BPSE solution. Samples were divided into three equal parts and estimated after collection as well as after in vitro storage for 3, 6 and 24 h. The first part was using for semen motility evaluation. The proportion of motile spermatozoa (MOT) and progressive movement (PMOT) of fresh and stored semen were evaluated using the CASA-system. The second part was examined subjectively by microscope (eosin-nigrosin (EN), eosin-nigrosin staining), the third one was assessed using dual fluorescence SYBR-14/propidium iodide (PI) and flow cytometry (FC). There were not any significant differences in sperm viability and motility between the groups at 0, 3, 6, and 24 h post collection. The percentage of viable spermatozoa in fresh semen determined by EN and FC was not different in Groups I and II (I - 88.71 ± 5.42 and 84.01 ± 3.19, respectively; II-90.87 ± 6.01 and 87.38 ± 5.57, respectively). Significantly lower percentages of viable spermatozoa were detected by FC compared to the EN method in both groups after 6 h (P ≤ 0.05) as well as 24 h (P ≤ 0.01) of storage. Moreover, the dual fluorescent SYBR-14/PI staining allowed for the identification a third population of double stained, moribund spermatozoa. High positive correlations in percentage of live spermatozoa were noted between EN and FC methods in both groups of birds. Evaluation of sperm viability by FC is a rapid, accurate, sensitive, and objective method for the assessment of pigeon sperm viability in fresh as well as stored semen.

Percoll gradient separation of cryopreserved common carp spermatozoa to obtain a fraction with higher motility, velocity and membrane integrity

We attempted to select a fraction of common carp, Cyprinus carpio spermatozoa that best survived a conventional freeze/thaw procedure, by centrifugation of frozen/thawed sperm through a Percoll gradient (45% and 90%). The proportion of motile spermatozoa (65.81 ± 5.19%), their velocity (77.58 ± 31.07 μm/sec), and membrane integrity (83.66 ± 4.38% intact) were significantly higher in separated sperm than in whole samples (motility 23.36 ± 2.98%, velocity 55.55 ± 19.03 μm/sec, and membrane integrity 57.92 ± 4.65%). Our results demonstrated that Percoll gradient centrifugation shows promise as a technique for selecting high quality cryopreserved fish spermatozoa, which could be useful for cryobiological research. Further studies are needed to evaluate the potentially higher fertilizing ability of the separated spermatozoa.

The Effect of the Partial Obstruction Site of the Renal Vein on Testis and Kidney in Rats: Is the Traditional Animal Model Suitable for Varicocele Research?

PurposeWe investigated the influence of the location of the partial renal vein obstruction on the left kidney, the bilateral testes, and cauda epididymal sperm quality and determined whether this animal model is suitable for varicocele study.Materials and MethodsA total of 25 adult male Sprague-Dawley rats were assigned to three groups: group 1 (experimental varicocele by partial ligation medial to the internal spermatic vein for 8 weeks, n=8), group 2 (partial ligation lateral to the internal spermatic vein for 8 weeks, n=10), and group 3 (sham operation for 8 weeks, n=7). Rats in groups 1, 2, and 3 underwent a left nephrectomy and bilateral orchiectomy at 8 weeks after the operation. Histological changes and Johnsen score in both testes were analyzed. Fibrotic changes in the left kidney were assessed by quantitative image analysis. Numbers of sperm and proportions of motile sperm in the cauda epididymides were determined.ResultsSignificant histological abnormalities and Johnsen score changes were observed in the testes in group 1. Renal fibrosis did not differ significantly among the groups. The proportions of motile sperm were significantly lower bilaterally in group 1 than in groups 2 and 3. However, the mean bilateral epididymal sperm count in group 1 was not significantly lower than in groups 2 and 3.ConclusionsOur results showed that experimental varicocele in the rat, induced by partial ligation medial to the internal spermatic vein, influences epididymal sperm quality without harmful effects on the left kidney. The present study certifies that this traditional animal model is suitable for varicocele research.

Sperm morphology, motility and fertilisation capacity in the myobatrachid frog Crinia georgiana

Sperm traits have been found to vary between individuals within populations in a variety of taxa. Sperm motility, morphometry and viability may be expected to have important effects on male fertility, although previous studies have found varying patterns, especially in external fertilisers. In the present study, we examined the effects of sperm swimming velocity, the proportion of motile spermatozoa, sperm head and tail length and the proportion of live spermatozoa on fertilisation success in the externally fertilising myobatrachid frog Crinia georgiana using IVF techniques and by controlling sperm numbers. We found no effect of any of the sperm traits we measured on IVF success. Neither did we find any relationship between sperm morphology and sperm performance. There was a negative relationship between sperm viability and male body size, which could be a function of age or an alternative tactic of differential investment in spermatozoa by smaller-sized males using sneak tactics in multiple matings. In contrast with most externally fertilising aquatic organisms, high rates of fertilisation appear to be achieved in C. georgiana with relatively low sperm swimming speeds.