Semen collection,characterization, and cryopreservation in a Magellanic penguin (Spheniscus magellanicus)

A cooperative method was developed for collecting semen from a Magellanic penguin. Ejaculate parameters and semen production during a breeding season were characterized. Experiments were performed to study the effect on penguin spermatozoa of two temperatures (4°C and 21°C) for short‐term storage, and two cryoprotectants (dimethylsulfoxide [DMSO] and ethylene glycol [EG]) for long‐term storage (cryopreservation). All dilutions were made using modified Beltsville Poultry Semen Extender. Sperm quality was assessed by evaluating motility and forward progression (sperm motility index [SMI]), viability, and morphology. A total of 39 ejaculates was collected over the 40‐day study period. Thirty‐eight ejaculates contained spermatozoa, but semen quality decreased toward the end of the study period. Varying levels of urate contamination were present in all ejaculates. Sperm quality parameters were similar for diluted samples held at 4°C and 21°C, and samples maintained high numbers of viable (77.8 ± 5.4%) and morphologically normal (67.9 ± 2.5%) spermatozoa at 3 hr. SMI and percentage of viable sperm decreased (P < 0.05) and the number of spermatozoa with a bent head or midpiece increased (P < 0.05) for both temperature groups over the 3‐hr storage interval. DMSO and EG were equally effective in maintaining penguin sperm quality parameters during the cryopreservation and thawing process. Frozen‐thawed semen maintained 69 ± 5 and 78 ± 3% of its pre‐freeze SMI and viability, respectively. SMI and viability decreased slightly during the cooling and equilibration phases but remained relatively stable during the 3‐hr storage interval post‐thaw. Frozen‐thawed semen also exhibited an increase (P < 0.05) in spermatozoa with a bent head or midpiece over time. The pre‐freeze SMI was higher (P < 0.05) for ejaculates with low levels of urates (clean ejaculates) compared with ejaculates with high levels of urate contamination, but sperm viability and morphology were similar (P > 0.05). Both SMI and viability of frozen‐thawed spermatozoa were higher (P < 0.05) for clean than for contaminated ejaculates. This is the first report on penguin ejaculate parameters, semen production, and preliminary methods for short‐ and long‐term semen storage. Zoo Biol 18:199–214, 1999. © 1999 Wiley‐Liss, Inc.     

Semen evaluation of giant pandas (Ailuropoda melanoleuca) at the Wolong Reserve

Semen from 4 wild-caught giant pandas (Ailuropoda melanoleuca) held at the China Conservation and Research Centre for the Giant Panda was collected (22 samples during 1991–1993) by electroejaculation, and evaluated for use in artificial insemination. Semen characteristics (mean ± SD) recorded were as follows: semen volume–1.5 ± .9 ml (range—0.3–3.5); sperm density 1.5 ± 0.1 × 10⁹/ml (range—0.24–4.2); motility 79 ± 10% (range—60–95); abnormal sperm 14 ± 5% (range—10–27); and pH 7.1 ± 0.2 (range—6.7–7.5). There were significant differences from year to year (P < 0.05) in semen volume collected and in the percentage abnormal sperm in 1993 compared to other years. There were no significant differences among semen produced from the four different pandas. Data collected were similar to reports for other giant pandas, and semen from all 4 giant pandas was considered suitable for use in artificial insemination. © 1994 Wiley-Liss, Inc.     

Reproductive status of endemic felid species in Latin American zoos and implications for ex situ conservation

Reproductive evaluations were conducted on 185 male cats representing eight endemic Latin American species that were maintained in 44 zoos and private facilities in 12 Latin American countries. Reproductive assessments (testicular measures, ejaculate quality, and blood testosterone/cortisol concentration) were used to establish normative values for large‐ and small‐sized cats in Latin American collections. Data also were analyzed using multiple regression to study the impact of proven breeder status, diet, and various animal housing combinations. Most felids (>95%) in the survey were of wild‐born origin, and <20% had produced offspring in captivity. Larger felids had bigger testes and produced more semen, but tended to produce low‐sperm‐density ejaculates. The ejaculates of small felids were more sperm‐concentrated, but contained fewer total spermatozoa. Sperm motility was unrelated to species size, and certain species (puma, margay, tigrina, and jaguarundi) consistently produced few (<40%) normal sperm forms. Across species, >50% of males had low sperm counts (<1 million total sperm per ejaculate). Among large cats (jaguars and pumas), proven breeders had larger (P<0.05) testes, greater semen volume, and more normal sperm than nonbreeders. Males on adequate diets had higher (P<0.05) circulating cortisol. Among small‐sized felids, proven breeders had higher (P<0.05) testosterone, and males housed alone or paired with a conspecific female had more (P<0.05) total sperm per ejaculate and greater (P<0.05) seminal and testicular volumes. Fifty‐nine ejaculates (potentially representing ～100 artificial insemination (AI) or 26,000 in vitro fertilization (IVF) procedures) were cryopreserved for a felid genome resource bank. In conclusion, breeding success and reproductive traits for many endemic felids in Latin American zoos appear to be suboptimal, and likely would benefit from improvements in diet and exhibitry. Technology transfer and continued training of zoo staff and scientists in Latin American countries are essential if these zoos are to achieve their tremendous conservation potential for felids and other threatened endemic species. Zoo Biol 22:421–441, 2003. © 2003 Wiley‐Liss, Inc.     

Freezability and Fertility Results with Uncentrifuged Stallion Semen

The first (1 to 3) sperm-rich fractions of the ejaculate were collected from 4 stallions using an open-ended vagina. The volume of the collected fractions was 12 ± 8 ml with a density of 475 ± 200 million spermatozoa/ml. Before freezing, the semen was diluted with a skim-milk based extender 1:1 to 1: 8 (volume of semen: volume of extender), depending on the initial sperm concentration to achieve a final concentration of 100 million/ml. The total number of spermatozoa in an insemination dose ranged from 0.7 to 1 billion spermatozoa. Within 12 h after ovulation, 48 mares were inseminated in 70 cycles. The total single-cycle pregnancy rate at day 21 was 24%, but varied from 10% to 33% per cycle among the stallions.     

Collection and fertilization potential of sperm from the Sulawesi crested black macaque (Macaca nigra)

In this study, semen was obtained by rectal probe electrostimulation (RPE) from the Sulawesi crested black macaque (Macaca nigra). Three experimental series were conducted. First, semen was collected from four animals anesthetized with either tiletamine-zolazepam (Telazol®) or ket-amine-HCl (Vetalar®) (five collections from each animal with each drug). Because of greater muscle relaxation and analgesia, we found tiletaminezolazepam to be an attractive alternative to ketamine-HCl as an anesthetic agent for RPE in M. nigra. Second, semen was collected from another four animals at stimulation frequencies of either 30 Hz or 60 Hz (five collections from each animal at each frequency). There were no significant differences in sperm number, in percentage of sperm with progressive motility, in the current required for sample recovery between tiletamine-zolazepam or ketamine-HCl anesthesia, or between a 30 Hz or 60 Hz stimulation frequency. Third, to check for retrograde sperm loss, the bladders of four animals were emptied, flushed with sterile saline, and then infused with TALP-Hepes medium. After RPE, sperm numbers in the bladder were compared with those in the ejaculate. Although sperm were recovered from the bladder [1.6 (± 0.9) × 10⁶] (mean ± SEM), the numbers were significantly less (P < 0.05) than those in the ejaculate [49 (± 18) × 10⁶]. The percentage of sperm with normal morphology in these samples was high (96.8 ± 1.0%). The average sperm number in the 84 samples collected for this study was 33.8 (± 4.1) × 10⁶. In preliminary experiments, we found that M. nigra sperm will fertilize rhesus monkey oocytes (Macaca mulatta) in vitro. © 1992 Wiley-Liss, Inc.     

Noninvasive semen collection from an adult orangutan

Preservation of the genetic diversity of the captive orangutan, especially the wild-caught founders, is critical in maintaining a long-term population in zoological parks. One solution to the problem of maintaining maximum genetic diversity would be to initiate a program of artificial insemination for genetically underrepresented individuals through the banking and interinstitutional use of cryopreserved semen. However, little is known about basic orangutan semen characteristics, and current methodology is inadequate to support such a program. In this paper, we report the results of semen collection from an adult Sumatran orangutan (Pongo pygmaeus abelli), using an artificial vagina without anesthesia or electrical stimulation. A total of 27 ejaculates were evaluated during a 1-year period. The total and liquid volumes of the ejaculates at 1 h following collection were 6.1 ± 0.6 ml and 2.6 ± 0.4 ml, respectively (mean ± SEM). The liquid portion continued to exude semen for 2 h; however, 90% of the motile sperm was exuded within the first 30 min. The total number of sperm in the ejaculate was 164 ± 10⁶ ± 16.5, and the percentage of motile cells was 60 ± 2.7%. We conclude that the artificial vagina provides a promising technique for semen collection in the orangutan, and view these results as an initial step in developing methods for in vitro sperm capacitation, sperm cryopreservation, and artificial insemination. © 1993 Wiley-Liss, Inc.     

Artificial insemination in captive whooping cranes: Results from genetic analyses

Artificial insemination has been used frequently in the captive whooping crane (Grus americana) population. In the 1980s, it was necessary at times to inseminate females with semen from several males during the breeding season or with semen from multiple males simultaneously due to unknown sperm viability of the breeding males. The goals of this study were to apply microsatellite DNA profiles to resolve uncertain paternities and to use these results to evaluate the current paternity assignment assumptions used by captive managers. Microsatellite DNA profiles were successful in resolving 20 of 23 paternity questions. When resolved paternities were coupled with data on insemination timing, substantial information was revealed on fertilization timing in captive whooping cranes. Delayed fertilization from inseminations 6+ days pre‐oviposition suggests capability of sperm storage. Zoo Biol 20:331–342, 2001. © 2001 Wiley‐Liss, Inc.     

Seasonality of serum testosterone levels and sperm density in Tursiops truncatus

We trained a mature male bottlenose dolphin, Tursiops truncatus, to provide semen samples on command. After completion of the 10-week training period, semen was collected twice weekly and blood was sampled twice monthly for a period of 28 months. Total sperm per ejaculate ranged from near 0 to 54.6 x 10(9) (n = 1332). Sperm densities from each session ranged from no sperm to 1,587 x 10(6)/ml (n = 241). Testosterone levels ranged from 1.1 to 54.4 ng/ml (n = 79). Seasonal variations were observed in total sperm per ejaculate, sperm density per ml of ejaculate, and in serum testosterone levels. Peak sperm densities were detected during September and October of three consecutive breeding seasons. Serum testosterone levels peaked in June, decreased during July and August, and were lowest in September and October, the period of greatest sperm density. Peak sperm production and density were coincident with the peak period of breeding activity but at a time when serum testosterone levels were lowest.     

Electroejaculation in chimpanzees and gorillas and artificial insemination in chimpanzees

Electroejaculation was performed in 3 chimpanzees, 1 pygmy chimpanzee, and 2 gorillas with an instrument that delivers a modified sine wave current with a frequency of 24 Hz. The current stimuli were applied by a rectal probe with longitudinal electrodes. The electrical parameters varied from 6 to 12 V and from 30 to 40 mA for response of erection and lay between 8 and 18 V and between 40 and 145 mA during semen emission. Eleven chimpanzee semen samples showed the following data (x ± SD): total volume 1.9 ± 1.3 ml, volume of the liquid fraction 0.3 ± 0.2 ml, spermatozoa per ejaculate 743 ± 376 × 10⁶, sperm motility 52.7 ± 9.6%, morphologically abnormal spermatozoa 12.2 ± 7.5%. From an adult gorilla, three semen samples were collected, in each case without spermatozoa. The electrostimulation of a 6-year-old gorilla led to an erection, but not to semen emission. Three female chimpanzees were inseminated with fresh or frozen semen, each of them within three different estrous cycles. None of these inseminations led to a pregnancy.     

Comparison of conventional and computer-assisted semen analysis in cockatiels (Nymphicus hollandicus) and evaluation of different insemination dosages for artificial insemination

Many psittacine species are threatened in the wild and also rare in captivity. Therefore, successful conservation and breeding programs are important to save these species. Unfortunately, clutches in conservation programs are frequently infertile. Semen evaluation is beneficial to investigate the causes of infertility and is advisable before artificial insemination (AI). In this study, we analyzed the semen of cockatiels (Nymphicus hollandicus) using two different methods and investigated different insemination dosages for AI. Cockatiels (n = 30) were divided into two groups (group A: nine males; group B: six males). The males in group B were endoscopically sterilized, whereas the males in group A were used as semen donors. In the first part of the study, the semen of males in group A was evaluated by semen analysis. Semen samples were collected by the massage technique and examined using a conventional light microscope and a computer-assisted semen analyzer for comparison. Results demonstrated that the evaluations of motility, progressive motility, and sperm concentration, but not of live/dead ratio, correlated strongly for both methods. However, the results for sperm concentration, progressive motility, and live/dead ratio differed significantly. In the second part of our study, the volume and quantity of spermatozoa of the semen samples were adjusted and used for AI of females of group B. Intravaginal insemination with 250,000 spermatozoa resulted in five of 17 (29%) eggs fertilized; however, intracloacal insemination resulted in only four of 57 (7%) eggs fertilized at 232,000 and 250,000 spermatozoa but none at higher or lower dosages.     

Case studies in antelope aggression control using a GnRH agonist

Maintaining surplus captive male antelope in bachelor groups can result in aggression in some species, leading to injury or death. Suppressing endogenous testosterone using gonadotropin‐releasing hormone (GnRH) analogs has been used in primates to control aggressive behavior, but little information is available on the use of GnRH analogs in nondomestic ruminant species. The aim of this study was to investigate the effect of a slow‐release GnRH agonist (deslorelin) on circulating hormone concentrations, semen and sperm characteristics and behavior in male gerenuk, dorcas gazelle, and scimitar horned oryx. Body weight, testicular volume, circulating hormone concentrations, ejaculate traits, and behavior were recorded before and during deslorelin treatment. A GnRH challenge (with serial blood sampling) was administered to gerenuk and dorcas gazelles before and during GnRH analog treatment. Quantitative behavioral data were collected for gerenuk and dorcas gazelles for 30 min three times a week, starting 1 month before deslorelin treatment, and the mean incidence of combined aggressive behaviors (supplanting, foreleg kicking, sparring, marking, and mounting) was compared before and during deslorelin treatment. No statistical difference (P>0.05) in body weight, semen volume, sperm concentration, percent sperm motility, percent sperm plasma membrane integrity, or percent normal sperm morphology was found before or during deslorelin treatment. The characteristic rise in luteinizing hormone (LH), occurring ～10 min following administration of a GnRH challenge in untreated males, was not evident during deslorelin treatment, although tonic LH concentrations were maintained. No differences (P>0.05) in the mean incidence of any aggressive behavioral traits in gerenuk or dorcas gazelle were detected before and during deslorelin. The absence of a GnRH‐induced increase in serum LH in treated males indicated that deslorelin suppressed pituitary responsiveness to endogenous GnRH, but that the continued tonic production of LH was sufficient to maintain testosterone production, aggressive behavior, and subsequent semen production. Zoo Biol 21:435–448, 2002. © 2002 Wiley‐Liss, Inc.     

The breeding management affects fresh and cryopreserved semen characteristics in Melopsittacus undulatus

Melopsittacus undulatus is a companion parrot worldwide diffused. Many parrots are considered endangered or vulnerable. The preservation of semen is crucial in endangered species, thus, M. undulatus could be a good model to study sperm characteristics and semen cryopreservation in these other endangered parrots. In this study the effect of the breeding management (males bred in promiscuous aviary or in couple) on sperm characteristics (motility, membrane integrity and morphometry) of fresh and cryopreserved semen was evaluated. The computer-assisted sperm analysis (CASA) revealed a significant effect of the husbandry method on semen characteristics in budgerigars: male housed in couple with the female in individual cages allowed the higher results in term of both semen quantity and sperm quality. Total and progressive motility were significantly higher in males bred in couple (68.7 ± 8.9% and 54 ± 15.9%, respectively) than in promiscuous aviary (48.3 ± 15.1% and 24.4 ± 12.4%, respectively), such as sperm velocity (average path velocity, straight line velocity, and curvilinear velocity). The type of sperm movement (amplitude of lateral head displacement, beat cross frequency, straightness, and linearity), sperm membrane integrity and morphometry parameters seemed not affected by the husbandry method. The standardization of a CASA procedure for the semen analysis in M. undulatus allow further studies on parrot semen manipulation and cryopreservation, but the method used for the breeding of the male could have a significant effect on the semen quality.     

Transaminases in cattle and buffalo (Bubalus bubalis) semen in relation to fertility and seminal characteristics during moderate and colder seasons

Semen from eight (four cattle and four buffalo) bulls was collected and analysed for physical characteristics and transaminase activity. The values of glutamic-oxaloacetic transaminase (GOT) and glutamic-pyruvic transaminase (GPT) activity were 1567±63 and 368±25 SF units¹ in the whole semen and 807±31 and 121±11 SF units in the seminal plasma for cattle and 1359±48 and 279±29 in the whole semen and 635±28 and 94±11 in the seminal plasma for buffalo. Cattle semen had significantly (P<0.01) higher mass motility, sperm concentration and transaminase activity than did buffalo semen. The enzyme activity was positively correlated with mass motility, sperm concentration and fertility, while only GOT activity was inversely related to the ambient temperature.     

Hormonal induction and semen characteristics of tambaqui Colossoma macropomum

In the hatchery-bred tambaqui Colossoma macropomum, spontaneous semen release does not occur, and hand-stripping produces reduced semen volume. The goal of this work is to evaluate the effects of hormonal induction with carp pituitary extract (CPE) on both qualitative (visual aspect, pH, motility, viability and morphological abnormalities) and quantitative (volume, concentration and number of spermatozoa per ejaculate) traits of tambaqui semen. Eleven males were treated with CPE (induced), and 11 were left untreated as a control (non-induced). All analysed parameters except motility and percentage of viable spermatozoa presented significant differences (p < 0.05) between the induced and non-induced treatments. CPE induction resulted in a 25-fold increase in semen volume and a 10-fold increase in the number of spermatozoa collected. However, both sperm concentration and the frequency of sperm with morphological abnormalities (commonly detached heads or bent tails) were significantly lower in CPE-induced fish. The hormonal induction of tambaqui males with CPE is efficient and positively influences some qualitative and quantitative properties of semen. Additionally, semen collection via gentle abdominal massage occurs more readily in CPE-induced fish.     

Comparison of semen collection techniques in goats

Characteristics of goat semen collected with the artificial vagina (AV), electroejaculator (EE), and Bailey ejaculator (BE) were compared. Semen was collected three times by each method from four bucks for a total of 36 separate collections. The semen was evaluated for volume, sperm concentration, mass activity (0 to 4, no activity to rapid wave motion), sperm motility (%), pH, and acrosome morphology (% normal). The means (± SEM) of semen samples collected with the AV, EE, and BE were volume-0.4 (±0.1), 0.7 (± 0.2), 0.6 (± 0.1) ml; concentration-4.6 (± 0.5), 2.1 (± 0.3), 1.4 (± 0.2) 10⁹/ml; mass activity-4.0 (± 0), 3.3 (± 0.3), 2.8 (± 0.2); motility-81 (± 2), 78 (± 2), 76 (± 2) %; pH-6.2 (± 0.1), 7.0 ± (0.2), 7.0 ± (0.2); and normal acrosomes-96 (± 1), 92 (± 3), and 88 (± 3) %, respectively. Semen collected with the AV was lower in volume than that collected with the BE (P < 0.05). The sperm concentration in semen samples collected with the AV was greater than those collected with the EE and BE (P < 0.05). Mass activity was greater and pH was less in AV samples than in EE or BE samples (P < 0.05). There was no difference in sperm motility of semen samples collected by the AV, EE, or BE. The average percentage of normal acrosomes was greater for AV than for BE samples (P < 0.05). Use of the EE and especially the BE resulted in increased vocalization by the goats and excessive muscular contractions of the rear limbs. The BE, in its present form (11.2 volts), is not recommended for semen collection in goats.     

Effect of progestins on serum hormones,semen production,and agonistic behavior in the gerenuk (Litocranius walleri walleri)

The use of progestins to suppress endogenous testosterone production to reduce agonistic behavior and prevent semen production was studied in gerenuk. Five male gerenuk (20 months to 3 years of age), housed as a bachelor group, were treated with 3 monthly injections of medroxyprogesterone acetate (MPA; 2.5–20 mg/kg), followed by a melengestrol acetate implant (MGA; 0.3 g/kg) for 2 months. Blood samples collected monthly were assayed for serum testosterone and cortisol using enzyme‐linked immunoassays. Quantitative behavioral data were collected for 30 min 3/week starting 1 month before treatment. Body weight, testes volume, and semen traits were measured before treatment, after MPA treatment, and after MGA treatment. Results showed lower (P<0.05) mean serum testosterone concentrations after MPA (4.34 pg/ml) and MGA (5.02 pg/ml) treatment compared to pre‐treatment values (65.9 pg/ml) in four of five gerenuk. The remaining sub‐adult gerenuk had low testosterone initially (4.9 pg/ml) that did not decrease further with treatment (1.4 and 7.8 pg/ml for MPA and MGA, respectively). Mean serum cortisol concentrations decreased markedly after treatment with MPA (6.0±3.7 ng/ml) and MGA (0.8±0.3 ng/ml). Cortisol concentrations were regained rapidly post‐treatment (42.8±4.8 ng/ml) and were not significantly different from the pre‐treatment value (60.6±12.6 ng/ml; P>0.05). The mean incidence of combined aggressive/dominant behaviors (horning, sparring, supplanting, threat) was not different before and after treatment. Body weight, total numbers of spermatozoa produced per ejaculate, percent motility, and percent normal spermatozoa declined maximally 8 months after treatment. Mean testes volume decreased (P<0.05) after MGA treatment (10.53 cm³vs. 11.96 cm³ pre‐treatment). Elevated hepatic enzymes and bile acids were seen in three of five animals after progestin treatment and anorexia was noted in two males after MGA implant removal, however two of three males had elevated liver enzymes before progestin treatment began. Results show that reducing serum testosterone concentration does not seem to modify agonistic behavior in bachelor gerenuk groups. Zoo Biol 26:245–257, 2007. © 2007 Wiley‐Liss, Inc.     

Comparison of electroejaculation and transrectal massage for semen collection in range and yearling feedlot beef bulls

Two experiments were conducted to compare electroejaculation (EE) and transrectal massage (RM) of the ampullary region for semen collection from beef bulls, and to determine the effect of semen collection method on semen traits. In experiment 1, semen was collected either by EE or RM randomly assigned on an alternate basis in 137 range beef bulls unaccustomed to being handled. The maximum time allowed for RM was 4 min and if no semen was obtained, EE was used. In experiment 2, semen was collected from 39 yearling feedlot beef bulls that were accustomed to being handled, by RM followed immediately by EE. The maximum time allowed for semen collection by both methods was 4 min. In both experiments, sperm concentration, percent of progressively motile sperm, percent of sperm staining alive, and sperm morphology were determined. In experiment 1, RM resulted in fewer (P<0.001) successful semen collections and fewer bulls with penile protrusion than EE (80.9% versus 100% and 54.4% versus 91.5%, respectively). The success of RM was not influenced by bull age or breed, or by the veterinarian performing the massage. Transrectal massage required more time (30s, P<0.001) for obtaining a semen sample and resulted in samples with lower sperm concentration (P<0.001), percent motile sperm (P<0.05) and percent live sperm (P<0.001) when compared to EE. In experiment 2, EE and RM were equally effective for obtaining a semen sample (97.4 and 94.9%, respectively), but the proportion of bulls exhibiting penile protrusion during semen collection was lower (P<0.0001) with RM compared to EE. Percent of sperm staining alive was also lower (P<0.01) in samples collected by RM. Sperm morphology (normal sperm, head defects, midpiece defects, proximal cytoplasmic droplets, and detached sperm heads) did not differ between samples collected by EE and RM. In conclusion, semen could be collected by transrectal massage from approximately 80% of range beef bulls and from 95% of yearling beef bulls accustomed to handling. Sperm morphology was not affected by the method of semen collection, but percent of motile sperm and live sperm were lower in samples collected by RM. A reduced ability to stimulate penile protrusion with RM precluded examination of the penis in a large proportion of bulls.     

Seasonal variation in the semen characteristics of Muturu (Bos brachyceros) bulls

Semen was collected by electroejaculation from seven mature Muturu bulls at weekly intervals for 9 weeks in each of three seasons (‘Dry’, ‘Rainy’, and ‘Late Rainy/Early Dry’) to study the effect of season on semen characteristics. The respective mean values for the ‘Dry’, ‘Rainy’ and ‘Late Rainy/Early Dry’ seasons were: volume 1.8 ± 0.1, 2.3 ± 0.1 and 2.0 ± 0.1 ml; percentage motility 36.2 ± 2.6, 37.7 ± 2.8 and 27.5±2.9; and percent morphologically normal sperm 70.0 ± 3.1, 79.1 ± 2.6 and 79.8 ± 2.3. No seasonal differences were found for sperm concentration/ml (overall mean 1.92 ± 0.16 × 10⁸) and for sperm output/ejaculate (overall mean 4.14 ± 0.39 × 10⁸). The chemical composition of seminal plasma (mg %) for the respective seasons was calcium: 4.8 ± 1.0, 7.9 ± 0.9 and 2.5 ± 0.7; magnesium: 4.8 ± 0.4, 4.3 ± 0.7 and 2.5 ± 0.7; and chloride: 330.7 ± 33.7, 136.0 ± 9.6 and 344.3 ± 31.8. No seasonal differences were found in sodium or potassium concentrations. Fructose was found in the semen of only one bull and only during the ‘Dry’ season.     

Correlation between sperm creatine phosphokinase activity and sperm concentrations in normospermic and oligospermic men

Toward the development of biochemical probes for the assessment of sperm function we have measured the activities of sperm creatine-N-phosphotransferase (CPK). There was a highly significant inverse correlation (P < 0.001 in all comparisons) between sperm CPK activities and sperm concentrations in specimens of normospermic and oligospermic men with > 30 million sperm/ml (0.106 ± 0.01 SEM, Nequals;90, expressed as CPK U/100 million sperm), 20–30 million sperm/ml (0.333 ± 0.07 SEM, Nequals;30) and 10–20 million sperm/ml (0.583 ± 0.12 SEM, Nequals;30) when compared with the CPK values of the < 10 million/ml specimens (2.242 ± 0.46 SEM, Nequals;30). Furthermore, the distribution of CPK activities within these four groups showed that 96%, 67%, 43%, and 4% of the samples, respectively, were in the < 0.250 CPK U/100 million sperm normal range (mean + 2 SD of the > 30 million sperm/ml group). However, there was no relationship between sperm CPK activities and the values of sperm motility (P > 0.15) or morphology (Pequals;0.38) in the samples. The migrated sperm fractions (significantly improved in motility and velocity parameters) showed CPK activities lower than the initial semen specimens (P < 0.01, Nequals;150). In fact, in some oligospermic men the CPK activities of the migrated sperm fractions were within the range of normospermic samples. The data suggest that sperm CPK values in the initial specimens and the degree of improvement in the migrated sperm fractions reflect the relative concentrations of a “normal” sperm subpopulation. We propose that CPK activities and similar objective biochemical parameters may be important in predicting sperm quality and the fertilizing potential of oligospermic men.     

Prediction of porcine semen fertility by homologous in vitro penetration (hIVP) assay

A field trial was conducted to compare the fertility predicting capacity of different sperm assays applying classical semen analysis, sperm function and the homologous in vitro penetration test (hIVP) to 60 ejaculates from four boars collected over a period of 15 weeks. No differences were found between the groups of fertility (Low Fertility: <20%; Intermediate: 40–60% and High: >80%) for sperm-rich fraction volume collection, sperm concentration, total sperm number, cationic contents in seminal plasma and ATP concentration. Partial differences were found in the parameters of motility, normal morphology, normal apical ridge (NAR), viability with eosin–nigrosin stain, hypo-osmotic swelling test (HOS), osmotic resistance test (ORT) and functional membrane integrity (with carboxyfluorescein diacetate, DCF). These parameters would be useful for detecting sperm with poor fertility, but they are not precise enough to discriminate an ejaculate with higher fertility than the herd median. Only the penetration percentage (10.24±1.45 vs. 55.13±3.35 vs. 84.72±1.73) and sperm number per oocyte (1.29±0.07 vs. 11.29±1.79 vs. 25.86±1.43) in a hIVP system were parameters with a predictive capacity to discriminate between the three fertility groups. Consequently, hIVP was found to be the best seminal assay and it may improve the in vitro assessment of sperm fertilizing ability.