Do parameters of seminal quality correlate with the results of on-farm inseminations in rabbits?

This study was conducted to determine if different sperm characteristics correlate with the in vivo fertility of rabbit sperm. A total of 2,765 heterospermic inseminations were performed in commercial rabbitries using 50-pooled samples of fresh semen. Sperm motility and morphological evaluations were performed on each of the heterospermic pooled samples to asses the seminal quality, and the percentage of kindling rate (76.2%) and number of kits born alive (9.3) were recorded. Sperm motility parameters, assessed using a computer-assisted sperm analysis (CASA) system (Sperm Class Analyzer, Microptic, Barcelona, Spain), were: average path velocity, curvilinear velocity, straight-line velocity, linearity, amplitude of lateral head displacement, beat cross-frequency, wobble and percentage of total motile spermatozoa. Morphological analyses included the percentage of sperm with a normal apical ridge, the percentage of sperm with cytoplasmatic droplets and the percentage of abnormal sperm. Significant correlations were observed between kindling rate and the percentage of total motile cells (r=0.31; P<0.05), linearity index (r=-0.32; P<0.05) and the percentage of abnormal sperm in the sample (r=-0.32; P<0.05). Regression models including motility and the morphological parameters explained 45% of the variation in kindling rate. These results indicate that motility parameters, determined by CASA systems, in combination with sperm morphology analyses can provide some information about the fertilizing potential of rabbit sperm.     

Regression analyses and motile sperm subpopulation structure study as improving tools in boar semen quality analysis

A precise estimation of the fertilizing ability of a boar ejaculate would be very useful to improve pig assisted reproduction results. For this purpose, we tested the mathematical combination of several parameters of the boar semen quality analysis, including the computer-assisted semen motility analysis (CASA), as a predictive fertility tool. The utilized mathematical relations among parameters were logistic and linear regressions. Two mathematical models obtained by logistic regression involving Osmotic Resistance Test (ORT Test), Hyperosmotic Resistance Test (HRT Test) and viability of fresh samples, showed a significant (P<0.05) correlation between semen characteristics and conception rate. However, none of the obtained models produced a significant correlation model between semen characteristics and prolificacy. The CASA analyses show that three separate subpopulations of spermatozoa with different motility characteristics coexist in boar ejaculates. There were significant (P<0.001) differences in the distribution of these subpopulations among boars, but no clear relationship between motile subpopulation structure and fertility was obtained. Our results support the belief that the predictive use of the results obtained in a standard boar semen quality analysis can reasonably be achieved by applying logistic correlation analyses among several function parameters of boar semen quality analysis and in vivo conception rates obtained after artificial insemination (AI).     

Evaluation of Spermatological Parameters Used to Predict the Fertility of Frozen Bull Semen

Post-thaw motility, velocity and acrosome integrity of frozen semen were determined in 18 bulls with varying fertility (average non-return rates: 71.3 (± 2.8) - range: 65.2-75.7). Five semen straws were investigated from each bull. The average values for sperm motility (percentage motile spermatozoa), sperm velocity (graded from 0-3) and acrosome integrity (proportion of spermatozoa with intact acrosome) were 67.5%, 2.5 and 79.3%, respectively. Significant correlations were found between sperm motility and velocity, but not between sperm motility and acrosome integrity. Both sperm motility and velocity were significantly related to bull fertility. It was concluded that of the post-thaw semen characteristics investigated in this study these 2 parameters provided a reliable basis for prediction of bull fertility.     

The relationship between rabbit semen characteristics and reproductive performance after artificial insemination

The relationships between several rabbit buck semen traits, concerning either the ejaculate or the dose inseminated (volume, mass motility, pH, percentage of motile sperm (PMS), concentration, number of total and motile sperm per ejaculate and per insemination dose) and the reproductive performance of does was investigated in 839 inseminations involving 54 bucks and 111 does. Four genetic types were involved: INRA1601 strain (A), INRA2066 strain (B) and the two reciprocal crossbreds (AB and BA). The mating design was A x A, B x B, (AB or BA) x (AB or BA). Semen was diluted (1:9) and a constant volume of 0.5 ml was inseminated 2-4h after collection. Therefore, the total number of spermatozoa per dose was variable and proportional to the initial concentration. Mass motility significantly influenced the kindling rate. Taken separately, volume, PMS and concentration did not influence the kindling rate but their product, the number of motile sperm per ejaculate, did. Litter size (total born) was significantly influenced by concentration and all variables depending on it, particularly the number of total and motile sperm per dose. However, reproductive performances were predominantly influenced by the physiological status of the does at insemination (lactation stage and receptivity).     

Motility and fertility of equine spermatozoa in a milk extender after 12 or 24 hours at 20 degrees C

The effects of extender and storage at 20 degrees C on equine spermatozoa were evaluated in two experiments using embryo recovery as the end point. In both experiments, inseminations were every other day, starting on Day 2 or 3 of estrus or after a 35-mm follicle was detected, with 250 x 10(6) progressively motile cells (based on initial evaluation). In Experiment 1, semen from two stallions was used to compare the motility and fertility of spermatozoa maintained in a) heated skim milk extender at 37 degrees C with insemination in <1 h; b) E-Z Mixin extender at 37 degrees C with insemination in <1 h; and c) E-Z Mixin extender at 37 degrees C with cooling to 20 degrees C and insemination after storage for 12 h at 20 degrees C. The percentage of motile spermatozoa was 34% after 12 h compared to 55% at 0 h (P < 0.05). However, the percentage of mares from which an embryo was recovered 6.5 d after ovulation was 62, 56, and 50% for Treatments A, B, and C (P > 0.05). In Experiment 2, semen from three stallions was used to compare the motility and fertility of spermatozoa in a) E-Z Mixin extender at 37 degrees C with insemination in <1 h or b) E-Z Mixin extender at 37 degrees C with cooling to 20 degrees C and insemination after storage for 24 h at 20 degrees C. The percentage of motile spermatozoa was 17% after 24 h compared to 54% at 0 h (P < 0.05). There was no difference between treatments (P > 0.05) in the percentage of mares from which an embryo was recovered 6.0 d after ovulation (68 vs 62%) or among stallions. Thus, stallion semen extended in E-Z Mixin was held at 20 degrees C for 24 h without a marked decline in fertility.     

Predictors of success of semen cryopreservation in chickens

Semen cryopreservation is very important for the ex situ management of genetic diversity in birds but it is rarely used. This is partly because of the highly variable success rates, and this emphasizes the need for predictors of semen freezability. This study evaluated the ability of semen quality tests to predict the success rates of semen cryopreservation in chickens and the relationships between each test. Individual variations of in vitro quality tests of semen were compared to the fertility obtained with fresh and cryopreserved semen. The in vitro semen quality tests represented viability, integrity, motility (percentage of viable and morphologically normal cells (PVN); mass motility (MMOT) and different motion parameters including percentage of motile spermatozoa (PMOT)) and biophysical tests (OSM, resistance to osmotic stress; membrane fluidity (FLUID)). Different in vitro tests were significantly correlated between each other for fresh (MMOT, PVN and FLUID, many criteria of objective motility) and cryopreserved semen (MMOT, different objective motility parameters, PVN). Fertility was significantly correlated with PVN for fresh semen and PVN and different objective motility criteria for cryopreserved semen. Membrane fluidity, followed by PVN, PMOT and MMOT, measured on fresh semen samples was positively correlated with fertility obtained with cryopreserved semen. The combination of the first three tests explained 85% of the variability of fertility observed with cryopreserved semen. In conclusion, we showed that different in vitro tests of semen quality are of predictive value for the success rate of semen cryopreservation in the chicken, the most accurate being membrane fluidity.     

The importance of porcine sperm parameters on fertility in vivo

It would be desirable to use semen parameters to predict the in vivo fertilizing capacity of a particular ejaculate. In animal production, an ejaculate is divided into multiple doses for artificial insemination (AI); therefore, it would be economically beneficial to know the functional quality (i.e., fertility) of the semen before it is inseminated. To identify a predictive assay of the fertilizing capacity of a porcine ejaculate, we performed 4 rapid assays of sperm quality (motility, viability, physiological status as assessed by chlortetracycline fluorescence, and ATP content) on samples from 9 ejaculates, before and after a thermal stress test (42.5 degrees C, 45 min). These parameters were subsequently correlated with in vivo fertility resulting from AI with 2 sperm doses, 3 x 10(9) or 0.3 x 10(9) motile cells in 70 mL (optimal or suboptimal sperm number per insemination, respectively) from these same ejaculates. No parameter was correlated to the fertility rates obtained after inseminating with the optimal semen doses, either before or after the thermal stress test (P > 0.05). However, with respect to the animals inseminated with the suboptimal semen dose, sperm motility (the percentage of motile spermatozoa as assessed visually by microscopy) prior to thermal stress was well-correlated to fertility rates (r = 0.783, P = 0.01). The percentage of spermatozoa displaying the chlortetracycline Pattern AR (acrosome reaction) was also statistically related to fertility (r = 0.05, P = 0.04), but the biological importance of this relationship is questionable given the small variation among ejaculates (range: 0 to 2%). No other sperm parameter was significantly related to fertility rates in this group (P > 0.05). These data, therefore, indicate that sperm motility is a useful indicator of sperm fertilizing capacity in vivo. Moreover, to identify a predictor of semen fertility it is critical that the number of spermatozoa used during insemination is sufficiently low to detect differences in sperm fertilizing efficiency.     

Identification of sperm subpopulations with defined motility characteristics in ejaculates from Florida goats

The aims of this study were to test the presence of discrete sperm subpopulations in Florida goat ejaculates using a computer-assisted sperm analysis (CASA) system and to establish the relationship between the distribution of the subpopulations found and individual buck, total motility, and sperm concentration. Clustering methods and discriminant analysis were applied to identify motile sperm subpopulations within the semen samples. Principal component analysis revealed that three principal components represented more than the 88% of the variance. After the cluster analysis was performed four motile sperm subpopulations were identified. Subpopulation 1 consisted of rapid and linear sperm (39.84%), Subpopulation 2 consisted of slow but linear spermatozoa (33.23%), Subpopulation 3 consisted of rapid, high ALH but non-linear spermatozoa (14.63%), and Subpopulation 4 consisted of slow and non-linear spermatozoa (12.31%). There were significant differences in the distribution of the four subpopulations (P < 0.001) as well as in the percentage of total motility and the overall sperm concentration (P < 0.05) in fresh ejaculates among the four bucks tested. In conclusion, four well-defined motile sperm subpopulations were identified in Florida goat ejaculates. The relationship between the distribution of the sperm subpopulations and individual buck, total motility, and sperm concentration shows that the spermatozoa of each have different motility patterns. Therefore, the study of discrete subpopulations of motile spermatozoa could lead to a substantial increase in information acquired during caprine semen analysis.     

Effect of season on fertility of frozen buffalo semen

Twenty double ejaculates from each of ten water-buffalo bulls were collected in June (non-breeding season) and again in November (breeding season). Fresh semen was screened for sperm quantity, motility, eosin uptake, and sperm morphology and was frozen using lactose, skim-milk, and Tris extenders. Thawed semen was checked for motility and Sephadex filtration. Half of each semen batch was used for artificial insemination in the breeding season and the other half during the non-breeding season.Laboratory screening revealed that June semen had a significantly lower Sephadex filtration rate and a higher percentage of abnormal sperm cells, and three June ejaculates were excluded from further processing due to poor sperm motility. In the remaining ejaculates the motility before freezing and the sperm cell quantity were higher in June semen than in November semen. Eosin uptake, mass motility, and post-freeze-motility did not vary with season. November semen produced significantly higher pregnancy rates than June semen over a total of 3220 inseminations in both seasons. Forty percent of the observed seasonality of buffalo fertility was attributable to the male. No fertility differences appeared between extenders used. When November semen was used, the fertility in adult buffaloes in both seasons was higher than in heifers.     

The effect of dialysis of extended ram semen prior to freezing on post-thaw survival and fertility

The effect of dialysis on extended ram semen prior to cryopreservation was studied. Techniques were developed to improve post-thaw recovery of dialyzed semen and a fertility trial was used to evaluate the viability of dialyzed and frozen semen. Dialysis prior to freezing was shown to increase post-thaw recovery of motile cells and percentage of cells passing through a Sephadex filter. Freezing semen in pellets on dry ice was superior to freezing in French straws. Pellets were thawed in an aluminum thaw block at 42 to 45 degrees C before insemination of progestagen-PMSG synchronized ewes. Double inseminations were made at 12-hr intervals. Natural service of synchronized ewes was also made at 12-hr intervals as a control. There was no significant difference (P greater than 0.05) in fertility between naturally serviced ewes (44.4%) and ewes inseminated with frozen semen (44.7%).     

The contribution of bulls and cows to the seasonal differences in the fertility of dairy cattle in Israel

Semen collected from dairy Israeli-Friesian bulls in the winter and summer was pelleted-frozen and used for inseminations in the collection season, in the other season, and after a 12-month storage period.The semen quality, as assessed by the percentage of ejaculates with initial poor motility and low sperm density and the percentage of spermatozoa surviving the freezing-thawing process, and those revealing acrosomal and tail abnormalities after thawing, was slightly and non-significantly lower in summer than in winter.The fertility of semen collected in both seasons was not affected by a 12-month storage period. The use of winter semen in summer did not improve the low fertility of cows in that season compared with those inseminated with summer semen.It appears from the results of this study that the summer decrease in the fertility of the inseminations is due to the adverse effect of this season on the fertility of the cows.     

Bull semen in vitro fertility after cryopreservation using egg yolk LDL: a comparison with Optidyl, a commercial egg yolk extender

Low-density lipoproteins (LDL) have been previously isolated and identified as the cryoprotective fraction of yolk. The effect of LDL on sperm motility after freezing-thawing has been reported, but no study has been made to assess the effect of LDL on bull semen fertility. The aim of this study was to evaluate the fertility of bull semen cryopreserved in the presence of LDL. Motility of semen cryopreserved in LDL was analyzed and compared to semen cryopreserved with Optidyl, a commercial extender containing egg yolk. To evaluate the fertilizing ability of semen, we used in vitro fertilization test, whereas acrosome and plasma membrane integrity were also evaluated. The percentage of motile spermatozoa was two fold higher after freezing in LDL than in Optidyl 54.4% versus 30.2% (P < 0.05). The cleavage rate was significantly higher after fertilization with semen frozen in LDL than with Optidyl 63.0% versus 54.8% (P < 0.05). No significant difference was observed on the blastocyst rate after in vitro culture. Integrity of the acrosome and the plasma membrane were maintained in both extenders. In conclusion, LDL preserve bull semen quality and fertilizing ability, allowing also better semen motility, after the freeze-thaw process.     

Effect of seminal plasma on the characteristics and fertility of rabbit spermatozoa

The effects of different dilutions of seminal plasma (SP) on the qualitative characteristics of rabbit spermatozoa and on their fertilising ability were analysed. Ejaculated semen was centrifuged twice and the sperm resuspended in media with decreasing ratios of SP/Tris: (1/2; 1/5; 1/10; 1/20; 1/30; 1/100) until the complete substitution was with SP. The control constituted sperm in undiluted SP. Samples were maintained at 37 degrees C and kinetic analysis done at fixed intervals (0-6h). Also the thiobarbituric reactive substances (TBA-RS) values were determined. Rabbit sperm suspended in Tris, or with extremely low content of SP, lost motility and viability within 1-3h, while sperm suspended in SP either undiluted or diluted up to 10-fold, showed similar motility during the 6h period (from 39 to 49%). Further dilutions of SP (1/20-1/30) had no effect during the initial 2h of storage but thereafter the decline of motility was more marked (after 6h: from 0 to 17%). Kinetic parameters followed the same trend and differences were particularly marked after storage: the highest values were in samples diluted up to 1/10; a sharp decline in motility characteristics was observed at higher dilutions. The addition of SP (1/2 v/v) to immotile sperm reactivated 35.5% of cells. However, SP did not significantly affect fertility rate or litter size possible involving an interaction with the female reproductive tract. SP reduced lipid oxidation (TBA-RS) of semen only after storage. A positive correlation between final TBA-RS and cell viability indicated that peroxidation was one of the cause of rabbit sperm deterioration during conservation.     

Effect of filtration on post-thaw quality of bull semen

Semen from 4 Holstein bulls was diluted in 4 different extenders, filtered with Sephadex ion-exchange column, and frozen in liquid nitrogen. Sperm motility, progressive motility, path velocity, progressive velocity and the percentage of normal acrosomes of filtered and nonfiltered semen were recorded before and after freezing. Semen characteristics were significantly influenced by extender, filtration and freezing. Before and after freezing, motility measurements and the percentage of normal acrosomes were higher (P < 0.001) in filtered than in nonfiltered spermatozoa. Post-thaw recovery rate of motile spermatozoa was higher in filtered semen than nonfiltered (68 vs 39%, P < 0.0001). The reduction in motility, progressive motility and the percentage of normal acrosomes during freezing and thawing processes were significantly lower (P < 0.0001) in filtered semen (34, 34 and 4%, respectively) than nonfiltered (59, 54 and 15%, respectively). Post-thaw viability of spermatozoa was significantly affected by extender, filtration and time (P < 0.0001). Immediate (0 h) post-thaw motility of nonfiltered semen (29%) was similar to 4-h post-thaw motility of filtered semen (25%; P > 0.05). In conclusion, bull spermatozoa recovered by Sephadex ion-exchange filtration showed better post-thaw viability.     

Fertility of stallion semen processed, frozen and thawed by a new procedure

The fertility of frozen-thawed and fresh semen from three stallions was compared in a trial using a randomized block design and 90 mares for 108 cycles. Semen was collected every third day, diluted to 50 x 10(6) sperm/ml with a citrate-based centrifugation medium, and centrifuged. The cells were resuspended at 700 x 10(6) progressively motile sperm/1.0 ml of added lactose-EDTA-egg yolk extender containing 4% glycerol, packaged by placing 0.55 ml into polypropylene straws, and frozen. Semen was thawed by immersion in 75 degrees C water for 10 sec. All of the 43 ejaculates collected were frozen, but 21 were discarded because progressive sperm motility was <35% immediately after thawing or <40% after 30 min of incubation at 37 degrees C. semen from the same stallions was collected daily for inseminations with fresh semen. Semen containing 200 x 10(6) progressively motile sperm was added to 10 ml of heated skimmilk extender. Mares were inseminated daily starting on the third day of estrus or when a >/=4-cm follicle was detected, whichever came later, and continuing through the end of estrus or for nine days. Based on palpation per rectum on day 50 postovulation, the pregnancy rates from inseminations during one estrus were 50, 56 and 61% with frozen semen and 67, 67 and 61% with fresh semen (P>0.05) from the three stallions, respectively. Thus, mean pregnancy rate with frozen semen was 86% of the rate attained with fresh semen.     

Effect of number of motile sperms inseminated on reproductive performance of rabbit does

The relationship between the number of motile sperms inseminated and fertility rate and litter size at birth in rabbits was investigated. A total of 651 artificial inseminations on multiparous rabbit does in different physiological states (lactating or not; sexually receptive or not) were inseminated with a number of motile cells/dose varying from about 1-20 x 10(6). The statistical model computed the following traits: maximum expected value (a), rate of approach to this value (b) and number of motile spermatozoa/dose needed to obtain threshold level (95% of the maximum). The physiological condition of the doe affected the reproductive performance. The sexual receptivity had significant effect during lactation: non-receptive does had the lowest fertility (a: 43.70%; P < 0.05) while no significant differences were found in non-lactating ones (73.80 vs. 89.11%). Also the rate of approach to this maximum was different: non-receptive does showed a higher dependence on spermatozoa number than receptive females (b: 0.69 and 0.58, respectively, in lactating and non-lactating does) and consequently more spermatozoa (13.1 and 11.1 x 10(6) motile/dose) were needed to reach the threshold value. The physiological state of the does did not affect litter size and only 4.1 x 10(6) of motile spermatozoa/dose are needed to reach the threshold value.     

Semen assessment, fertility and the selection of Hereford bulls for use in AI

Nineteen young Hereford bulls were used to study the relationship between semen characteristics and fertility in artificial insemination following 15 320 inseminations. Seven measures of sperm motility, morphological abnormalities, the release of hyaluronidase, ATP content and sperm head measurements were examined as predictors of fertility (49-day fixed-interval non-return rate). Two assessments of motility, three categories of abnormal spermatozoa, acrosomal changes and the release of hyaluronidase had predictive power. Multiple regression analysis showed that a combination of sperm motility after dilution in saline, motility after thawing and the proportion of coiled tails and proximal protoplasmic droplets provided the best prediction of fertility and allowed bulls to be ranked in order of observed non-return rate (%) with a Spearman correlation better than +0.80.     

Sperm motion characteristics of GarolexMalpura sheep evolved in a semi-arid tropical environment through introgression of FecB gene

The arid and semi-arid tropical climates of India are endowed with vast diversity of non-prolific sheep breeds. The GarolexMalpura sheep has been evolved in a semi-arid tropical environment by introgression of FecB gene via artificial insemination of Malpura ewes using diluted semen of prolific microsheep Garole and subsequently multiplied by inter se mating among GarolexMalpura halfbreds. The aim of the present study was to identify FecB mutation in sexually mature GarolexMalpura rams by forced RFLP-PCR of BMPR-1B gene and evaluate: (i) semen production and sperm motion characteristics of GM rams and (ii) influence of age and FecB genotype on their semen attributes. Semen was collected during autumn season from 12 donor rams by artificial vagina on 8 occasions at weekly interval. The overall means of traits which did not differed significantly with age or FecB genotyping were volume (0.72 ml), mass motility (4.44), sperm concentration (2721.56 x 10(6)ml(-1)), curvilinear velocity (134.51 microm/s), motility (81.3%), amplitude of lateral head displacement (6.24 microm), beat frequency (44.43 Hz), sperm head elongation (48.9%) and sperm head area (10.01 microm(2)). The FecB genotyping had a significant effect (P<0.05) on percent linearity and rapid motile sperms, which did not vary significantly with age. Although sperm concentration was higher in FecB(BB) and FecB(B+), compared to FecB(++) genotypes but the effect was non-significant. The age and FecB genotyping had significant effects (P<0.05) on straightness, average path velocity, straight-line velocity and percentage of medium or slow motile sperms. It is concluded that GarolexMalpura rams with introgressed FecB gene are capable of producing good quality semen in a semi-arid tropical climate.     

The relationship of semen quality to pregnancy rate and litter size following artificial insemination in the bitch

Artificial insemination of 31 bitches with fresh, undiluted, semen resulted in 22 31 conceptions (71%) and the birth of 124 puppies. Inseminations using at least 220 x 10(6) spermatozoa of normal morphology resulted in a mean pregnancy rate of 81.5% (22 27 ). The pregnancy rates were significantly influenced by the total number of progressively motile or morphologically normal sperm cells per ejaculate (P < 0.05). The pregnancy rate was not influenced by the percentage of progressively motile or morphologically normal sperm cells per ejaculate (P > 0.1). Litter size was not influenced by sperm motility or morphology (P > 0.1).     

Effects of added caffeine on results following artificial insemination with fresh and refrigerated rabbit semen

Lactating rabbits (n=1335) were artificially inseminated to study the effect of the addition of caffeine on rabbit semen stored for up to 96 h. Concentration of 0-5 mM/l were tested. Whereas a concentration of 0.2 mM/l increased spermatozoa motility, higher concentration values adversely affected reproductive parameters. Spermatozoa stored at 18 degrees C for 72-96 h did not have the capacity to react with caffeine when it was added before insemination. Caffeine did not enhance fertility or prolificacy, regardless of its ability to increase sperm motility.