Practical applications of sperm selection techniques for improving reproductive efficiency

Several selection techniques are available for processing spermatozoa. Apart from sperm washing to remove seminal plasma, only “swim-up” and colloid centrifugation have been used to any extent to prepare spermatozoa for in vitro fertilization, and only colloid centrifugation has been used to prepare sperm samples for artificial insemination. Single-layer centrifugation (SLC) through a species-specific colloid has been shown to be effective in selecting spermatozoa with good motility, normal morphology and intact chromatin in a range of species. This method is less timeconsuming than swim-up, and has been scaled-up to allow whole ejaculates to be processed in a practical manner. The applications of SLC are as follows: to improve sperm quality in insemination doses or in samples for in vitro fertilization, to increase the shelf life of normal sperm doses, to remove pathogens (viruses, bacteria), to improve cryosurvival by removing dead and dying spermatozoa before freezing or after thawing, to select spermatozoa for intracytoplasmic sperm injection, and to aid conservation breeding.

Practical applications of sperm selection techniques for improving reproductive efficiency

Several selection techniques are available for processing spermatozoa. Apart from sperm washing to remove seminal plasma, only “swim-up” and colloid centrifugation have been used to any extent to prepare spermatozoa for in vitro fertilization, and only colloid centrifugation has been used to prepare sperm samples for artificial insemination. Single-layer centrifugation (SLC) through a species-specific colloid has been shown to be effective in selecting spermatozoa with good motility, normal morphology and intact chromatin in a range of species. This method is less timeconsuming than swim-up, and has been scaled-up to allow whole ejaculates to be processed in a practical manner. The applications of SLC are as follows: to improve sperm quality in insemination doses or in samples for in vitro fertilization, to increase the shelf life of normal sperm doses, to remove pathogens (viruses, bacteria), to improve cryosurvival by removing dead and dying spermatozoa before freezing or after thawing, to select spermatozoa for intracytoplasmic sperm injection, and to aid conservation breeding.(AU)

Androcoll™-P-Large selects boar spermatozoa with good membrane integrity from the sperm-rich fraction of the ejaculate

The study investigated sperm membrane integrity (as a measure of sperm viability) and sperm motility in spermatozoa taken from different portions of the ejaculate, namely the first 10 ml (P1) of sperm-rich fraction (SRF) and from the rest of the ejaculate up to the appearance of gel (P2), both before and after centrifugation on a single layer of Androcoll-P-Large (SLC). Thus there were 4 treatment groups: P1, P2, SLC P1 and SLC P2. Sperm motilities were not different between the various treatment groups, except that the SLC samples had higher linear + non-linear motility than the non-SLC-selected samples. Sperm membrane integrity, in contrast, was significantly higher in P2 than in P1 (P < 0.001), and was also higher in both SLC-selected groups than in the uncentrifuged groups (P < 0.001). There was a significant correlation between membrane integrity and linear + non-linear motility (P < 0.001). These results indicate that the spermatozoa found in P2 have better membrane integrity than those in P1 when used as fresh spermatozoa, and furthermore, that SLC selects the most robust spermatozoa regardless of their origin in the ejaculate. Thus, in situations where P1 is collected separately for sperm cryopreservation purposes, the remainder of the SRF could be used for fresh AI doses, particularly where SLC can be used to select the most robust spermatozoa. These findings have practical importance for the swine insemination industry.(AU)

Androcoll™-P-Large selects boar spermatozoa with good membrane integrity from the sperm-rich fraction of the ejaculate

The study investigated sperm membrane integrity (as a measure of sperm viability) and sperm motility in spermatozoa taken from different portions of the ejaculate, namely the first 10 ml (P1) of sperm-rich fraction (SRF) and from the rest of the ejaculate up to the appearance of gel (P2), both before and after centrifugation on a single layer of Androcoll-P-Large (SLC). Thus there were 4 treatment groups: P1, P2, SLC P1 and SLC P2. Sperm motilities were not different between the various treatment groups, except that the SLC samples had higher linear + non-linear motility than the non-SLC-selected samples. Sperm membrane integrity, in contrast, was significantly higher in P2 than in P1 (P < 0.001), and was also higher in both SLC-selected groups than in the uncentrifuged groups (P < 0.001). There was a significant correlation between membrane integrity and linear + non-linear motility (P < 0.001). These results indicate that the spermatozoa found in P2 have better membrane integrity than those in P1 when used as fresh spermatozoa, and furthermore, that SLC selects the most robust spermatozoa regardless of their origin in the ejaculate. Thus, in situations where P1 is collected separately for sperm cryopreservation purposes, the remainder of the SRF could be used for fresh AI doses, particularly where SLC can be used to select the most robust spermatozoa. These findings have practical importance for the swine insemination industry.

Routine assessment of motility of ejaculated stallion spermatozoa using a novel computer-assisted motility analyzer (Qualisperm™)

Visual motility analysis is the basis for routine quality evaluation of stallion semen, although its prognostic value for fertilizing ability is considered low. The present study evaluated the ability of a novel computer-assisted motility analyzer (QualiSperm™) to determine the motility and velocity of ejaculated, extended stallion spermatozoa (collected from 10 stallions, 3 ejaculates/stallion) and following two different colloidal centrifugation methods (one- or two- layer), compared to visual evaluation by two independet operators. The Qualisperm™ instrument was able to retrieve and analyze ~10 times more spermatozoa per sample compared to routine visual estimation on the same time frame (~1,100 vs ~100 spermatozoa).The proportion of motile spermatozoa increased after the colloid-separation, compared to the extended ejaculates (P < 0.05) in some stallions. However, owing to the large variation seen among ejaculates and stallions, both for extended ejaculate (P < 0.05) as well as for the colloid centrifugations (P < 0.01), the differences were lost when the entire population was examined statistically. Interestingly, significant differences were seen for individual stallions between the measurements ofQualisperm™ and observers, as well as between observers (P < 0.05). Apart from the significantly higher number of spermatozoa analyzed at one time, the Qualisperm™ system provided a parameter that could simply not be estimated by visual assessment; mean sperm velocity (in µm/sec). Sperm velocity, upon which every computer assisted instrumentation base their evaluations, varied among stallions (and ejaculates within stallions, P < 0.05), with a tendency to increase after colloid-separation, thus suggesting the Qualisperm™ system might be able to differentiate sperm sub-populations. Due to its higher accuracy (in terms of sperm numbers examined) and speed, the Qualisperm™ system appears to be a suitable instrument for routine evaluation of equine semen.(AU)

Routine assessment of motility of ejaculated stallion spermatozoa using a novel computer-assisted motility analyzer (Qualisperm™)

Visual motility analysis is the basis for routine quality evaluation of stallion semen, although its prognostic value for fertilizing ability is considered low. The present study evaluated the ability of a novel computer-assisted motility analyzer (QualiSperm™) to determine the motility and velocity of ejaculated, extended stallion spermatozoa (collected from 10 stallions, 3 ejaculates/stallion) and following two different colloidal centrifugation methods (one- or two- layer), compared to visual evaluation by two independet operators. The Qualisperm™ instrument was able to retrieve and analyze ~10 times more spermatozoa per sample compared to routine visual estimation on the same time frame (~1,100 vs ~100 spermatozoa).The proportion of motile spermatozoa increased after the colloid-separation, compared to the extended ejaculates (P < 0.05) in some stallions. However, owing to the large variation seen among ejaculates and stallions, both for extended ejaculate (P < 0.05) as well as for the colloid centrifugations (P < 0.01), the differences were lost when the entire population was examined statistically. Interestingly, significant differences were seen for individual stallions between the measurements ofQualisperm™ and observers, as well as between observers (P < 0.05). Apart from the significantly higher number of spermatozoa analyzed at one time, the Qualisperm™ system provided a parameter that could simply not be estimated by visual assessment; mean sperm velocity (in µm/sec). Sperm velocity, upon which every computer assisted instrumentation base their evaluations, varied among stallions (and ejaculates within stallions, P < 0.05), with a tendency to increase after colloid-separation, thus suggesting the Qualisperm™ system might be able to differentiate sperm sub-populations. Due to its higher accuracy (in terms of sperm numbers examined) and speed, the Qualisperm™ system appears to be a suitable instrument for routine evaluation of equine semen.

Prolongation of stallion sperm survival by centrifugation through coated silica colloids: a preliminary study

Difficulties to be overcome in the widespread use of artificial insemination (AI) in mares are low sperm survival and poor sperm quality, which are encountered frequently among breeding stallions. Therefore, a method is needed to prolong the useable life of stallion spermatozoa destined for AI. In a preliminary study using 8 ejaculates from one stallion, density gradient centrifugation or centrifugation through a single layer of silica colloid appeared to prolong sperm motility compared to uncentrifuged spermatozoa, thereby potentially extending the useable life of treated stallion spermatozoa for AI. Furthermore, there was an improvement in sperm morphology, with the number of morphologically normal spermatozoa increasing from 42 to 60.5% and with the removal of approximately 60% spermatozoa with head or tail defects from the original population. No difference between the two centrifugation methods, in terms of yield or duration of spontaneous motility, could be detected in this study. Either of these methods of colloidal centrifugation could be a useful aid to preparing stallion spermatozoa for artificial breeding techniques, including AI.(AU)

Prolongation of stallion sperm survival by centrifugation through coated silica colloids: a preliminary study

Difficulties to be overcome in the widespread use of artificial insemination (AI) in mares are low sperm survival and poor sperm quality, which are encountered frequently among breeding stallions. Therefore, a method is needed to prolong the useable life of stallion spermatozoa destined for AI. In a preliminary study using 8 ejaculates from one stallion, density gradient centrifugation or centrifugation through a single layer of silica colloid appeared to prolong sperm motility compared to uncentrifuged spermatozoa, thereby potentially extending the useable life of treated stallion spermatozoa for AI. Furthermore, there was an improvement in sperm morphology, with the number of morphologically normal spermatozoa increasing from 42 to 60.5% and with the removal of approximately 60% spermatozoa with head or tail defects from the original population. No difference between the two centrifugation methods, in terms of yield or duration of spontaneous motility, could be detected in this study. Either of these methods of colloidal centrifugation could be a useful aid to preparing stallion spermatozoa for artificial breeding techniques, including AI.