Biomedical subjects

W V Holt

Publications and source records attributed to W V Holt.

At least 19 recordsLinked to original sources

PubMed2006

An investigation into the similarities and differences governing the cryopreservation success of koala (Phascolarctos cinereus: goldfuss) and common wombat (Vombatus ursinus: shaw) spermatozoa.

The aim of this study was to determine the relative cryopreservation success of koala and wombat spermatozoa and to investigate reasons for their respective post-thaw survival by examining the sperm's response to a range of osmotic media and determining the presence and distribution of F-actin. An hypothesis was proposed that F-actin may be imparting a degree of structural inflexibility to the koala sperm plasma membrane; hence, exposure of spermatozoa to cytochalasin D (5 microM), a F-actin depolymerisation agent, should result in increased plasticisation of the membrane and greater tolerance of cell volume changes that typically occur during cryopreservation. In experiment 1, koala (n = 4) and wombat (n = 4) spermatozoa packaged in 0.25 mL straws were cryopreserved using two freezing rates (fast-3 cm above liquid N2 interface; slow-6 degrees C/min in a freezing chamber) and two glycerol concentrations (8 and 14% v/v) in a tris-citrate glucose buffer with 15% (v/v) egg yolk. Wombat spermatozoa showed better (P < 0.01) post-thaw survival (% motile, % intact plasma membranes, % decondensed sperm heads) than koala spermatozoa. When exposed to media of varying osmolality, koala spermatozoa were less tolerant (% intact plasma membrane) of hyper-osmotic conditions (920 and 1410 mOsmol/kg) than wombat spermatozoa. F-actin was localised using a monoclonal antibody but only found in the wombat sperm head. When koala and wombat spermatozoa were exposed to media of varying osmolality, cytochalasin D had no beneficial effect on sperm survival (% intact plasma membranes). This study has demonstrated that wombat spermatozoa are highly tolerant of cryopreservation when compared to koala sperm but that spermatozoa from both species show greatest post-thaw survival when frozen slowly in 14% glycerol. Koala sperm are also particularly susceptible to hyper-osmotic environments but lack of detectable F-actin in the koala spermatozoan suggests that poor cryopreservation success in this species is unlikely to be associated with F-actin induced plasma membrane inflexibility.

Actins↗

PubMed2006

Impact of antifreeze proteins and antifreeze glycoproteins on bovine sperm during freeze-thaw.

There are no reports on the use of antifreeze proteins (AFP) and antifreeze glycoproteins (AFGP) for the use of bull sperm cryopreservation despite studies in the ram, mouse and chimpanzee. The effect of freezing and thawing on bull sperm viability, osmotic resistance and acrosome integrity were observed following the addition of AFP1, AFPIII and AFGP at four concentrations (0.1, 1, 10 and 100 microg/ml). In a second part of the experiment, fluorescein was conjugated to the AFPs and AFGP and observations were made using fluorescence microscopy to determine whether binding occurred between the sperm cell membranes and the proteins. In the final part of the study the cryopreservation media were cooled in the presence of the AFPs and AFGPs at the four concentrations on a cryomicroscope to mimic similar cooling curves as those used in the presence of sperm. Following freeze-thaw, AFPI resulted in increased osmotic resistant cells at 0.1-10 microg/ml compared to the control (P<0.01). AFPI and AFPIII did bind to the sperm cells. There was no visual difference in ice structure between the control, AFPIII and AFGP but AFPI resulted in parallel crystals at 0.1, 1 and 10 microg/ml. We suggest that the increased osmotic resistance in the spermatozoa cryopreserved in AFPI is due to the cells orientating between the ice crystals, reducing mechanical stress to the cell membrane. Previous research has shown that osmotic resistance correlates with bull fertility, suggesting that bull spermatozoa cryopreserved in the presence of AFPI may have increased fertility in vivo.

Acrosome↗

PubMed2006

Dilution of spermatozoa results in improved viability following a 24 h storage period but decreased acrosome integrity following cryopreservation.

This study was designed to investigate the physiological factors affecting the reduced viability of cryopreserved spermatozoa following dilution. Ninety-six ejaculates were collected from 13 bulls and diluted to 10 x 10(6) and 60 x 10(6) sperm/ml in a commercial long term extender (Eqcellsire; IMV) and in an egg yolk extender. Samples diluted in the egg yolk extender were frozen in 0.25 ml straws. Samples diluted in the Eqcellsire were stored at room temperature for 24 h and assessed for sperm cell viability using SYBR14 and PI (Molecular Probes) and osmotic resistance. Frozen samples were thawed and assessed for viability, osmotic resistance and acrosome intergrity. Acrosome integrity was measured using Mitotracker, PI and PNA-FITC (Molecular Probes). Spermatozoa diluted to 10 x 10(6) sperm/ml and stored at ambient temperature had a higher proportion of viable spermatozoa (P < 0.01) and were less susceptible to osmotic stress (P < 0.01) than sperm diluted to 60 x 10(6) sperm/ml. Following cryopreservation there was no concentration-related difference in the proportion of viable spermatozoa and their relative susceptibility to osmotic stress. Spermatozoa diluted to lower cell concentrations had a higher proportion of viable cells that were acrosome reacted (P < 0.001). It is suggested that the higher proportion of acrosome reacted cells may result from an increased proportion of cells in a capacitated-like state in the spermatozoa diluted to lower concentrations. A Spearmans ranked correlation demonstrates a relationship between individual bull spermatozoa following dilution or cryopreservation for viability (r2 = 0.98; P < 0.001) or osmotic resistance (r2 = 0.87; P < 0.001) suggesting a variation in these characteristics between bulls.

Acrosome↗

PubMed2006

Harnessing the biology of the oviduct for the benefit of artificial insemination.

Spermatozoa fulfil a single role, namely achieving syngamy by transporting the haploid genome to their counterpart gamete, the oocyte. Simple as this may seem, it is fraught with many difficulties, especially in the face of biological processes that enable females to select spermatozoa after they have mated multiply with several males. Conversely, the female reproductive tract sequesters a privileged sperm subpopulation in the oviductal isthmus for variable periods of time, releasing them when the time is opportune for fertilisation. Recent studies of sperm transport in the female reproductive tract suggest that these phenomena involve signalling dialogues between spermatozoa and the female reproductive tract environment. Opportunities for mutual signalling are immense but have received relatively little attention. The oviduct is an organ of crucial significance in modulating sperm function and may be one of the most important sites for determining many aspects of sperm selection and competition. The oviductal environment possesses the potential for enhancing sperm survival, suppressing and activating sperm motility as required, and responds to the arrival of spermatozoa by producing novel proteins. While the biological nature of the sperm-oviduct dialogue is interesting for its own sake, the mechanisms that govern these processes offer opportunities for the improvement of artificial insemination procedures. If oviductal proteins enhance sperm survival, they offer opportunities for the development of long-life semen diluents. Conversely, if we understood the basis of sperm selection we may be able to concentrate on identifying and using only the best sperm subpopulations for improved animal breeding efficiency.

Animals↗

PubMed2005

Is quality assurance in semen analysis still really necessary? A spermatologist's viewpoint.

In a provocative article to this Journal, Anne Jecquier, an eminent andrologist who, more than 20 years ago, was a prime mover in suggesting the need for quality assurance (QA) in andrology laboratories, has now proposed that the QA schemes may no longer be needed. Here I reply to that proposition, largely by agreeing that, since the QA schemes have brought about higher technical standards in laboratories, Anne Jecquier's assertion is possibly true. However, vigilance is still needed in discriminating between unproductive investment of time and energy in the refinement of tests that may offer little information about fertility, and maintaining technical standards such that where necessary they provide the requisite information. Thus, although it may not matter in practice whether a sperm concentration is estimated as 100 or 200 x 10(6)/ml, distinguishing between 25 and 100 x 10(6)/ml would probably influence a clinician's treatment decisions. Anne Jecquier also suggested that sperm function tests have limited predictive value in terms of fertility assessment. While I agree that this is largely true at present, I also argue that these tests are probably not developed to their full potential. I am optimistic that tests to distinguish and quantify the population of fertilization-competent sperm within an ejaculate will eventually become available.

Andrology↗

PubMed2005

Validation of an experimental strategy for studying surface-exposed proteins involved in porcine sperm-oviduct contact interactions.

Previous experiments have shown that boar sperm survival in vitro is enhanced when co-incubated with a solubilised protein extract of porcine oviducal apical plasma membrane proteins. Here, we examine the hypothesis that the effects are mediated by direct oviduct-sperm contact and use in situ biotinylation of the oviducal epithelial surface to trace the surface-exposed biotinylated proteins through purification and solubilisation steps. We have also examined the effectiveness of mechanical scraping as a method of recovering oviducal epithelial proteins. We show that a subset of proteins originally exposed at the oviducal surface eventually bind to spermatozoa during incubation in vitro, but also show that a different protein subset is implicated if the sperm incubation is performed with proteins that had been biotinylated after (ex situ) extraction from the oviduct. Apical plasma membrane fractions biotinylated after purification contained many more biotinylated protein bands than preparations labelled before purification and multiple protein bands were eventually found to associate with spermatozoa. Although the evidence presented here supports the hypothesis that protein(s) anchored to the oviducal epithelium bind populations of spermatozoa directly and may have a role in the enhancement of sperm viability, it also shows that the choice of investigative technique exerts a major influence on experimental outcomes.

Animals↗

PubMed2004

Smooth muscle actin and vimentin as markers of testis development in the harbour porpoise (Phocoena phocoena).

Testicular development in the harbour porpoise Phocoena phocoena was examined using animals (n = 192) stranded or by-caught off the coast of England, Wales and Scotland. Classification of animals according to their stage of sexual development was undertaken using gonadal morphology and the distribution of cytoskeletal proteins. Smooth muscle actin (SMA) and vimentin proved particularly useful in this respect; SMA was prominent in the myoid peritubular cells of the adult testis, and two stages of peritubular cell SMA expression could be recognized ('absent' or 'incomplete'). The initial appearance of SMA in peritubular cells was associated with significant increases in body length and body weight (P < 0.001), and occurred during the second year of life. Vimentin, which was prominent in prespermatogonia and spermatogonia, sometimes showed a polarized cytoplasmic distribution. This correlated with a developmental stage at which the seminiferous tubule epithelium becomes populated by germ cells (mean age 1.8 years). Several antibodies were tested for their utility as Sertoli cell markers, but none was found to be specific or useful. Nevertheless, immunohistochemical localization of desmin, GATA-4, Ki67 and androgen receptor was possible despite the poor quality of tissue preservation. This study showed that immunohistochemical classification of these individuals provides a robust basis for the recognition of key physiological stages of sexual development in the male harbour porpoise. This may provide an alternative to the estimation of age, body weight and body length in future analyses aimed at detecting possible adverse effects of environmental pollutants on the reproductive potential of wild marine mammals.

Actins↗

PubMed2003

Detrimental effects of cryopreservation of loach (Misgurnus fossilis) sperm on subsequent embryo development are reversed by incubating fertilised eggs in caffeine.

Cryopreservation can cause changes to the genetic material of cells, but the mechanism and significance of these changes are still unknown. It has been suggested that some damage to the sperm genome could be repaired by the DNA repair system of the oocyte after fertilisation. Caffeine has been reported to be an inhibitor of such repair processes. In this study the effect of caffeine on the repair system of Loach (Misgurnus fossilis) oocytes was investigated. Loach eggs were fertilised using cryopreserved sperm. Embryos derived from cryopreserved sperm were exposed to 2.6mM caffeine for 1h after fertilisation. The experiments were carried out using 32313 embryos from four females and eight males. Embryo survival was evaluated for 46 h until the hatching stage. Reduction in embryo survival after 20th stage is generally believed to result from the failure in the genome function of embryos. Cryopreservation of sperm significantly decreased embryo survival (53.4+/-2.8% compared to 68.4+/-2.8% of control) after the 20th stage. However, the addition of caffeine to the embryos derived from cryopreserved sperm, in contrast to our expectation, significantly increased survival of loach embryos (70.9+/-2.8% compared to 53.4+/-2.8% of embryos derived from cryopreserved sperm in the absence of caffeine). The effect of individual donors of sperm and eggs on overall embryo survival was also studied. Whilst no significant differences were observed between males, the effect of individual females on embryo survival was significant. The analysis of embryo survival at different developmental stages showed that embryo survival both before and after 20th stage decreased with embryo development. When fresh sperm were used the decline of embryo survival with development was more pronounced compared with those embryos derived from cryopreserved sperm. Possible explanations of these effects are presented.

Animals↗

PubMed2003

In vitro maintenance of boar sperm viability by a soluble fraction obtained from oviductal apical plasma membrane preparations.

Oviductal apical plasma membrane fractions have been successfully used to provide an in vitro model to study the role of direct membrane contact in sperm-oviduct interactions. Apical plasma membrane preparations from pig oviductal tissues show a dose-response in their ability to maintain boar sperm viability in vitro. Membrane preparations obtained from other tissues (lung and duodenum) are incapable of maintaining boar sperm viability to the same extent as oviductal tissue. The present study examined the validity of two hypotheses that arise from current knowledge of sperm-oviduct interactions, namely, that (i) apical plasma membranes prepared from ampullar regions of the oviduct are less effective than those from isthmus regions, and (ii) sperm survival is more effective in apical plasma membrane preparations derived from follicular phase oviducts than those derived from luteal phase oviducts. Both hypotheses were proved false. The nature of the active component(s) in the oviductal apical plasma membrane fractions was further investigated. Heat treatment (100 degrees C for 20 min) diminished the capacity of membranes to support boar sperm viability. Furthermore, a soluble salt-extracted fraction obtained from oviductal apical plasma membrane preparations was biologically active and supported boar sperm viability in vitro. This may indicate that the active factor(s) responsible for the maintenance of boar sperm viability is not an integral part of oviductal membranes and is peripherally bound to these membranes.

Animals↗

PubMed2002

Non-invasive assessment of oestrous cycles and evaluation of reproductive seasonality in the female wild black rhinoceros (Diceros bicornis minor).

Female wild black rhinoceroses in Zimbabwe were monitored non-invasively using faecal progesterone metabolite analysis and observation of reproductive behaviour. A postpartum period of reproductive inactivity of at least 4 months, followed by a period of 4-7 months of oestrous cyclicity, was detected in six multiparous females. Three-quarters of the oestrous cycles (n = 21) had a total duration (mean +/- SEM) of 26.8 +/- 1 days. Other types of cycle were characterized either by an extended luteal phase, lasting on average twice as long as the normal cycle, or by an extended follicular phase. These extended cycles may have resulted from early embryo loss and heat stress. Female rhinoceroses did not conceive before 8 months after giving birth and some females (n = 2) most likely aborted after 3.0-3.5 months of gestation. The detected period of cyclic oestrus occurred between May and March in females (n = 9), and there was a 3 month extended interoestrous interval in nulliparous females during the period of decreasing daylengths that can be presumed to be the period of poorest fertility for the black rhinoceros under tropical latitudes. In contrast, the period of optimum fertility in the Southern hemisphere coincided with the late spring and early summer, and corresponded to the early rainy season. As a result, a higher incidence of births was detected in the late rainy season, providing the lactating female with the most suitable environment in terms of nutritional requirements.

Animals↗

PubMed2002

Importance of cooling rate and animal variability for boar sperm cryopreservation: insights from the cryomicroscope.

A series of experiments was set up to investigate the effect of different cooling rates on boar sperm cryosurvival using cryomicroscopy. The cooling protocols were split into two stages: (i) from +5 degrees C to -5 degrees C and (ii) from -5 degrees C to -50 degrees C. Fluorescent probes (SYBR14 and propidium iodide) were used to monitor plasma membrane integrity during the entire process. Cooling rates in the range 3 degrees C min(-1) to 12 degrees C min(-1) did not cause significant damage to the sperm plasma membrane between +5 degrees C and -5 degrees C; however, spermatozoa cooled at 24 degrees C min(-1) to -5 degrees C were slightly damaged. Motility was not particularly sensitive to variations in cooling rate. Cooling rates in the range 15 degrees C min(-1) to 60 degrees C min(-1) did not produce differences in sperm cryosurvival during freezing between -5 degrees C and -50 degrees C, or after thawing. In addition, cooling rates in the range 3 degrees C min(-1) to 80 degrees C min(-1) did not produce significant differences in sperm cryosurvival. However, slow freezing (3 degrees C min(-1)) induced a slight increase in the percentage of plasma membrane-damaged spermatozoa (propidium iodide-positive) at -50 degrees C. Inter-ejaculate and inter-boar differences in sperm cryosurvival were manifested independently of cooling rate. The sperm plasma membrane remained intact (SYBR14-positive) during cooling and freezing, but upon rewarming, the plasma membrane of a high proportion of spermatozoa was damaged (propidium iodide-positive), indicating that rewarming is a critical step of the freezing-thawing process.

Analysis of Variance↗

PubMed2001

Use of computer-assisted sperm motility assessment and multivariate pattern analysis to characterize ejaculate quality in Mohor gazelles (Gazella dama mhorr): effects of body weight, electroejaculation technique and short-term semen storage.

Subjective and objective semen assessments were performed on 18 male Mohor gazelles (Gazella dama mhorr). Sperm motility assessments combined with sperm plasma membrane and acrosomal integrity evaluations were undertaken as part of a captive breeding programme. The primary objective was to test methodology for short-term preservation of gazelle semen for artificial insemination (storage in N-[Tris(hydroxymethyl)methyl]-2-aminoethanesulphonic acid-Tris diluent (TEST) for up to 96 h at 17 degrees C). However, the secondary objective was to investigate phenotypic and genotypic influences on semen quality within this small population, which was established in 1971 with only 12 genetic founders. Sperm motility was measured by computer-assisted semen assessment and the data were analysed using a pattern analysis technique to detect and quantify naturally occurring sperm subpopulations within the semen samples. Four sperm subpopulations distinguishable by their motion characteristics were detected. The relative frequencies of two subpopulations (population 2: highly motile, non-linear; and population 4: poorly motile, non-linear) in fresh semen were correlated with the maximum voltage used during electroejaculation. The frequency of subpopulation 2 was negatively correlated with maximum voltage (r = -0.875, P < 0.0001) and the frequency of subpopulation 4 was positively correlated (r = 0.727, P < 0.005). The frequencies of all subpopulations varied significantly among the animals sampled (chi-squared = 2577.6, degrees of freedom = 54, P < 0.0001) and subpopulation 4 was also correlated with body weight (r = -0.59, P < 0.005). Semen stored at 17 degrees C retained motility, plasma membrane and acrosomal integrity for 48 h, but these measures decreased thereafter. The frequency of a sperm subpopulation showing uncoordinated but active motility increased significantly over the first 48 h and then decreased.

Animals↗

PubMed2001

Carbohydrate mediation of boar sperm binding to oviductal epithelial cells in vitro.

After mating, mammalian spermatozoa are transported to the lower oviductal isthmus. Spermatozoa are sequestered at the isthmus by attaching and interacting with oviductal epithelial cells, hence forming a sperm reservoir. In several mammalian species, specific carbohydrates mediate sperm-oviductal epithelial cell binding. A quantitative in vitro free cell bioassay was developed to investigate the involvement of carbohydrate recognition in pig sperm-oviductal epithelial cell interactions. This assay was validated. The sensitivity of the assay was such that it was possible to discriminate between different sperm concentrations and sperm-oviductal epithelial cell co-incubation periods, spermatozoa with damaged plasma membranes and epithelial cells of non-reproductive origin. Optimal conditions were used to incubate spermatozoa and oviductal epithelial cells in the presence of six hexose sugars at concentrations of 0, 2, 10 and 50 mmol l(-1). A significant (P < or = 0.05) reduction in the binding of spermatozoa to the oviductal epithelium was detected with 2, 10 and 50 mmol maltose l(-1), 50 mmol lactose l(-1) and 50 mmol mannose l(-1). These findings support the hypothesis that attachment of pig spermatozoa to oviductal epithelium before fertilization is mediated by carbohydrate recognition.

Animals↗

PubMed2001

Hormonal characterization of the reproductive cycle and pregnancy in the female Mohor gazelle (Gazella dama mhorr).

The oestrous cycles of seven captive Mohor Gazelles (Gazella dama mhorr) were investigated. Hormone profiles obtained from faecal samples collected each day from cyclic females indicated that the mean duration of the oestrous cycle was 18.62 +/- 0.26 days (range 16-22 days; n = 37 oestrous cycles). No inter-individual differences in the concentration of faecal progestagen metabolites excreted were observed, but mean faecal oestrogen excretion during both the luteal and inter-luteal phases of the oestrous cycle varied among females (P < 0.001 and P = 0.070, respectively). Oestrous cycles were synchronized using controlled internal drug release (CIDR) devices, before natural mating with an intact male. Concentrations of faecal progestagen metabolites remained approximately constant for the first 10 weeks of gestation (mean +/- SEM = 4048 +/- 407 ng g(-1) faeces), before increasing to a mean of 23 556 +/- 1176 ng g(-1) faeces. Two of seven female gazelles conceived immediately after removal of the CIDR device, a similar proportion to that conceived at the postpartum oestrus under natural conditions. Life history data for these individuals indicated that the mean time to conception in female gazelles is positively correlated with peak values in the ratio of excreted oestrogen : progestagen during the inter-luteal period of their oestrous cycles (R(2) = 0.58; P < 0.05). This finding indicates that interactions between steroid production and metabolism may influence the likelihood of conception occurring in this species.

Animals↗

PubMed2000

Basic aspects of frozen storage of semen.

Basic concepts of cryopreservation and the causes of cryoinjury are reviewed. The possible roles of cryoprotectants and additives are considered in the context of their putative interactions with the sperm plasma membrane. Modern approaches to the laboratory assessment of spermatozoa after freeze-thawing are also briefly discussed.

Animal Diseases↗

PubMed2000

Fundamental aspects of sperm cryobiology: the importance of species and individual differences.

While semen cryopreservation is successfully used for a few species, application to other species can be problematic. Here, I argue that species differences in female tract anatomy, subtle differences in sperm transport mechanisms, ability to time inseminations and deliver spermatozoa effectively are powerful determinants of fertility with cryopreserved spermatozoa. Poor sperm survival represents one major aspects of the problem and determining biophysical characteristics of the sperm plasma membrane is an established approach to solving it. However, this approach is unable to account for the consistent differences in post-cryopreservation sperm quality between individual males, an effect that is recognized is many species although only documented in a few. Searching for genetic differences between these individuals might offer a genomically-based direction in semen cryopreservation research. Cryopreservation of spermatozoa and spermatogenic cells for intracytoplasmic sperm injection has been developed primarily to deliver an intact genome and presents a very different set of technical problems.

Animals↗

PubMed1999

Cryopreservation of macropodid spermatozoa: new insights from the cryomicroscope.

This study examined the effects of cooling and cryopreservation upon macropod spermatozoa (eastern grey kangaroo, Macropus giganteus and red-necked wallaby, Macropus rufogriseus). Sperm survival during and after freezing to -30 degrees C or 70 degrees C in minimum essential medium (MEM) + 5, 10, 20 or 30% (v/v) glycerol, MEM + 10 or 20% (v/v) ethylene glycol and MEM containing a mixture of 7.5% (v/v) glycerol + 10% (v/v) dimethylsulphoxide was examined by cryomicroscopy. The MEM/glycerol mixtures permitted better post-thaw sperm recovery than the other cryoprotectants. After freezing to -30 degrees C at 10 degrees C min(-1) in 20% glycerol, then rewarming at 20 degrees C min(-1), flagellar activity resumed in more than 50% of spermatozoa when the temperature increased into the range 5-10 degrees C. However, as the temperature increased, into the range 20-25 degrees C, motility declined rapidly so that less than 5% motile cells were seen at 35 degrees C. Spermatozoa in MEM without cryoprotectant were also examined by cryomicroscopy to evaluate changes in flagellar configuration, swimming behaviour and viability during cooling from 35 degrees C to approximately -7 degrees C, and rewarming to 35 degrees C. Cooling from 35 to 28 degrees C induced kangaroo spermatozoa to exhibit rigid principal-piece bending and non-linear motility, which was reversed by further cooling and the spermatozoa resumed their normal linear movement. Rewarming induced principal-piece bending in the range of 20-30 degrees C, but this effect was reversed by further warming. Although red-necked wallaby spermatozoa showed these effects, they also exhibited a tendency to form rosette-like clusters during rewarming, especially when the temperature reached approximately 14 degrees C. The clusters were induced when the flagellar end-pieces became anteriorly reflected, producing hook-like flagellar conformations, which then became interlinked.

Animals↗

PubMed1999

Sperm subpopulations in boar (Sus scrofa) and gazelle (Gazella dama mhorr) semen as revealed by pattern analysis of computer-assisted motility assessments.

The aim of this study was to test the suitability of "pattern analysis" for the exploration of data provided by computer-assisted semen analysis methods. Data sets derived from the examination of boar sperm responses to bicarbonate and caffeine (measurements on 3208 spermatozoa) and from studies of semen cryopreservation in Mohor gazelles (7278 spermatozoa) were reanalyzed. A nonhierarchical classification method was used to generate initial subgroups of spermatozoa (9 for boar, 13 for gazelle). The subgroup centroids were fused, yielding three boar sperm subpopulations and four gazelle sperm subpopulations distinguished by sperm behaviors. Bicarbonate and caffeine both induced major transitions (p < 0.0001) of boar sperm behavior, detected as shifts in group membership (from group 2, i.e., active but nonlinear movement, into group 1, i.e., linear, rapid movement). Some spermatozoa (approximately 3%) were refractory to both caffeine and bicarbonate. The gazelle sperm subpopulation structure was affected by the inclusion of equex (sodium triethanolamine lauryl sulfate) in the cryoprotective diluents. Equex suppressed the appearance of spermatozoa with erratic behavior (p < 0.0001; high curvilinear velocity, low linearity, low straight-line velocity) after cryopreservation. The proportion of these erratic spermatozoa was positively correlated with animal age (r = 0.68, p = 0.029). Pattern analysis revealed novel aspects of the data not seen in the original investigations and usefully supplemented the more standard data analysis approaches.

Animals↗