Semen characteristics and sperm morphology in the Arabian leopard (Panthera pardus nimr) and how these vary with age and season

The Arabian leopard is a critically endangered species. Since there are only an estimated 200 animals remaining in the wild, careful management of the captive population is necessary to minimise inbreeding. The objective of this study was to characterise sperm morphology and ejaculate quality in captive males. Semen was collected by electroejaculation from 8 adult captive male leopards (aged 2-16 years) during the summer and winter months, and semen parameters, including sperm morphology, were assessed. Two-year-old leopards showed lower total sperm counts per ejaculate than older animals and these counts declined at > 8 years. Ejaculates collected during the hot summer showed significantly lower sperm concentrations, total sperm counts, sperm motility and viability and percentage of spermatozoa showing normal morphology than ejaculates collected in the cooler winter. The results showed that the male leopard attains sexual maturity between 2 and 3 years of age and exhibits good semen quality until 8 years. Collection of semen for artificial breeding or banking would best be carried out in the cooler winter months.     

Reproductive physiology and artificial insemination studies in wild and captive gerenuk ( Litocranius walleri walleri )

Gerenuk antelope in North American zoos are descended from 28 founders imported from Kenya approximately 20 years ago. Intensive management is required to prevent inbreeding depression. Artificial insemination has potential for augmenting genetic management, but successful application requires a thorough understanding of species' reproductive norms. Semen collected from captive (n = 10) and wild (n = 6) gerenuk contained low numbers of morphologically normal spermatozoa (approximately 40%). Age, but not season, influenced (P < 0.05) the proportion of morphologically normal spermatozoa (mean +/- s.e.m., 12-17 months of age, 10.3 +/- 1.9%; 18-26 months of age, 34.4 +/- 6.2%; 3-6 years of age, 40.0 +/- 4.7%). Seasonality was investigated by analysing faecal testosterone and progesterone in males and females, respectively, by radioimmunoassays. Females cycled all year (ovarian cycle length, 18.7 +/- 0.9 days). Testosterone in males did not vary (P > 0.05) with time of year. Three females (3/9, 33%) became pregnant by insemination with 9.75-54.0 x 0(6) motile fresh or frozen sperm after oestrus synchronisation with two prostaglandin F(2alpha) injections, 12 days apart. One female inseminated with frozen-thawed sperm delivered a full-term stillborn calf after 213 days gestation. These results characterise gerenuk reproductive norms and indicate that artificial insemination may be a useful tool in the genetic management of gerenuk.     

Assessment of basic seminal characteristics, sperm cryopreservation and heterologous in vitro fertilisation in the fishing cat (Prionailurus viverrinus)

Conservation of the fishing cat, a threatened south-east Asian felid, could benefit from effective ex situ genetic management and breeding programmes, including the use of assisted reproduction. The aims of the present study were to: (1) characterise basal seminal traits of fishing cats in Thailand zoos; and (2) investigate the effect of cryopreservation on sperm motility, acrosomal integrity and in vitro function. Seminal traits were evaluated in electroejaculates collected from eight males. Spermatozoa were diluted in n-tris(hydroxymethyl)-methyl-2-aminoethanesulfonic acid Tris (TEST)-yolk buffer (TYB) without glycerol, then diluted further with TYB with glycerol (4% final concentration) at either 25 degrees C or after slow cooling to 5 degrees C and frozen in straws over liquid nitrogen vapour. After thawing, sperm function was assessed by insemination of viable domestic cat oocytes. Fishing cat ejaculates averaged (+/- s.e.m.) 43.6 +/- 14.2 x 10(6) motile spermatozoa with 33.5 +/- 6.8% normal sperm morphology. Semen processing had a negligible effect (P > 0.05) on sperm motility and acrosomal integrity, but values were reduced (P < 0.05) after thawing. All thawed samples fertilised domestic cat oocytes, with 62.1% (36/58) of mature oocytes cleaving. Glycerol addition at 5 degrees C resulted in higher (P < 0.05) post-thaw motility and intact acrosomes than glycerol addition at 25 degrees C. In conclusion, good-quality ejaculates can be obtained from Thai fishing cats and their spermatozoa exhibit adequate function after cryopreservation for in vitro fertilisation procedures.     

Semen quality in captive Houbara bustard,Chlamydotis undulata undulata

Semen quality in captive-bred Houbara bustards, Chlamydotis undulata undulata, was assessed during three consecutive breeding seasons. In any one season, sperm quality, in terms of the proportion of eosin-permeable spermatozoa and of spermatozoa with abnormally large nuclei, varied among individual males, but not among their ejaculates. Neither the proportion of spermatozoa with large nuclei, nor those permeable to eosin were related to the total sperm output of males. The fertilizing ability of males was related to their mean seasonal proportion of eosin-permeable spermatozoa, but not the proportion of spermatozoa with large nuclei. The ranking of males on the basis of the proportion of spermatozoa with large nuclei in their ejaculates was significantly positively correlated between seasons, although ranking on the basis of sperm eosin-permeability was not. The cause or consequence of producing spermatozoa with large nuclei (and excess DNA) remains to be elucidated, but appears to be a trait that is characteristic of houbara bustard males that is maintained between breeding seasons.     

Flow cytometric sorting of frozen-thawed spermatozoa in sheep and non-human primates

Research was conducted in sheep to determine an effective preparation method for high-purity sorting of frozen-thawed spermatozoa. The efficacy of sorting frozen-thawed spermatozoa was then investigated in several non-human primate species. An aliquot of each ejaculate (three rams, three ejaculates per ram) was processed as a fresh control (FRESH). Frozen spermatozoa were thawed and prepared for sorting by no further processing (FT-NEAT), washing (FT-WASH) or gradient centrifugation (FT-GRADIENT) and evaluated for motility at 1 h post-staining and motility and acrosomal status at 0 and 4 h post-sorting. Samples were analysed using a high-speed cell sorter. High levels of purity for X- and Y-enriched samples were achieved for all treatments (85-92%). The percentage of motile spermatozoa before sorting was lower (P < 0.05) for frozen-thawed samples (FT-NEAT: 32.7 +/- 2.5%; FT-WASH: 32.2 +/- 3.3%; FT-GRADIENT: 73.9 +/- 3.7%) compared with FRESH (83.3 +/- 1.2%). Post-sorting, the percentage of motile spermatozoa before and after incubation for FT-NEAT (60.0 +/- 5.1% and 27.2 +/- 6.1% for 0 and 4 h, respectively) was lower than that for FRESH (87.8 +/- 0.9% and 83.3 +/- 1.2% for 0 and 4 h, respectively; P < 0.05), FT-WASH (80.0 +/- 2.4% and 71.7 +/- 3.6% for 0 and 4 h, respectively; P < 0.05) and FT-GRADIENT (84.4 +/- 1.3% and 77.2 +/- 1.7% for 0 and 4 h, respectively; P < 0.05). Vanguard sperm migration distance through artificial cervical mucus was lower (P < 0.05) for FT-NEAT (17.7 +/- 1.7 mm) compared with FT-WASH (29.1 +/- 3.8 mm) and FT-GRADIENT (28.4 +/- 2.0 mm) and similar (P < 0.05) to FRESH (23.7 +/- 1.8 mm). Sample preparation using a modified wash method enabled high-purity sorting (range 86-97% purity) of frozen-thawed epididymal spermatozoa in the baboon (Papio hamadryas), common marmoset (Callithrix jacchus) and common chimpanzee (Pan troglodytes). For all non-human primate species, sorted spermatozoa were progressively motile (marmoset: 20.5 +/- 5.5%; baboon: 37.5 +/- 2.5%; chimpanzee: 73.0 +/- 2.0%), acrosome intact (marmoset: 68.5 +/- 7.5%; baboon: 89.5 +/- 1.5%; chimpanzee: 84.0 +/- 1.0%) and able to penetrate an artificial cervical mucus. In summary, high-purity sorting of frozen-thawed ram and non-human primate spermatozoa with recovery of progressively motile, acrosome-intact spermatozoa was possible after processing to remove cryodiluent.     

Sperm transport, size of the seminal plug and the timing of ovulation after natural mating in the female tammar wallaby Macropus eugenii

The distribution of spermatozoa and seminal plug in the reproductive tract and the timing of ovulation were examined at various times in a naturally mated monovular macropodid marsupial, namely the tammar wallaby (Macropus eugenii). After the first post partum (p.p.) mating, 28 females were isolated and their reproductive tracts dissected at 0.5, 6, 18, 36 and 40 h post coitum (p.c.). Each tract was ligated into 13 major anatomical sections and spermatozoa and eggs were recovered by flushing. Mating was possibly delayed by handling and occurred 21.7 +/- 2.5 h p.p. in these animals. Copulation lasted 7.8 +/- 0.7 min. Within 0.5 h after a single mating, the tract contained 25.8 +/- 10.2 x 10(6) spermatozoa and 21.6 +/- 8.8 g of seminal plug, 96% and 70% of which was lost within 6 h p.c. respectively. Spermatozoa reached the uterus, isthmus and ampulla of the oviduct on the side of the developing follicle within 0.5, 6 and 18 h p.c., respectively, and a uterine population of 26.1 +/- 12.10(3) spermatozoa was maintained for over 40 h. Sperm numbers were reduced at the cervix (up to 57-fold) and uterotubule junction (eight-fold) and only one in approximately 7500 ejaculated spermatozoa (3.4 +/- 0.9 x 10(3)) reached the oviduct on the follicle side. Differential transport of spermatozoa was not observed. Although the numbers of spermatozoa were reduced in the parturient uterus, they were highly variable and were not significantly different to those in the non-parturient uterus. Ovulation and recovery of sperm-covered eggs from the isthmus occurred 36-41 h p.c. (49-72 h p.p.). In contrast with the polyovular dasyurid and didelphid marsupials, the tammar wallaby ejaculates large numbers of spermatozoa, but transport is relatively inefficient and sperm storage in the tract before ovulation is limited.     

Development and evaluation of electroejaculation techniques in the Tasmanian devil (Sarcophilus harrisii)

Electroejaculation (EEJ) has been used successfully to collect samples suitable for genome resource banking from a variety of endangered wildlife species. Ejaculates can also be used to evaluate the reproductive potential of individuals and provide information on seminal characteristics to aid in the development of sperm cryopreservation techniques. Electroejaculation techniques used for marsupial and eutherian species were tested on Tasmanian devils (n=35). Spermic ejaculates were collected in 54% (19/35) of EEJ attempts. Spermic ejaculates were low in volume (3.9±6.5×10(2)μL, range 10-3000μL) and contained low numbers of spermatozoa (3.3±7.8×10(3) spermatozoa per ejaculate, range 6-33000). The osmolality and pH of presumptive urine-free ejaculates were 389±130mOsmkg(-1) (range 102-566) and 7.0±0.9 (range 6.0-8.0), respectively. Prostatic bodies were observed in 79% (26/33) of ejaculates. Episodic fluctuations in serum testosterone concentrations were not detected during the EEJ procedure (P>0.05). Increases observed in serum cortisol concentrations during EEJ were less (P<0.05) than those observed after an adrenalcorticotropic hormone challenge and diurnal variation suggested that cortisol concentrations are greater during the day than at night (P<0.05). This information can be used to provide range values for the future examination of basic endocrine responses and the adrenal-pituitary axis of this species. This study also demonstrated that spermatozoa-rich devil electroejaculates are more difficult to obtain and poorer in quality than those of other marsupials.     

Boar semen can tolerate rapid cooling rates prior to freezing

Two experiments were performed in the present study that demonstrated that boar spermatozoa are capable of surviving rapid cooling rates within a range of 15-5 °C before freezing. Boar ejaculates diluted in Beltsville thawing solution (BTS) (1:1, v/v) were held at 17-20 °C and shipped over a 24-h time period from two AI centres to a cryobiology laboratory, where they were pooled (Experiment 1) or cryopreserved individually (Experiment 2) using a standard 0.5-mL straw freezing protocol. The effects of cooling before freezing were assessed after thawing through the objective evaluation of sperm motility and flow cytometric analysis of membrane integrity, acrosomal status, changes in membrane lipid architecture monitored by merocyanine and annexin V binding and intracellular production of reactive oxygen species. In Experiment 1 (six replicates), two semen pools (five ejaculates per pool) were cooled from 15 to 5 °C at rates of 0.08, 0.13, 0.40 and 1.50 °C min(-1). These cooling rates did not result in any significant differences (P>0.05) in any of the post-thaw sperm assessments, even in thawed samples incubated under capacitation conditions. In Experiment 2, three individual ejaculates from 16 boars were slowly (0.08 °C min(-1)) or rapidly (1.5 °C min(-1)) cooled before freezing. A consistent interboar variability (P<0.01) was detected, which was independent of the cooling rate used. Cooling rate only significantly influenced (P<0.05) sperm assessments in four of 16 boars, which exhibited slightly higher percentages of motile cells and intact plasma and acrosomal membranes in the samples that had been cooled slowly. These findings demonstrate that boar spermatozoa undergoing cryopreservation can withstand rapid cooling rates before freezing.     

Characterization of viability,mitochondrial activity,acrosomal integrity and capacitation status in boar sperm during in vitro storage at different ambient temperatures

Extended storage of unfrozen boar semen becomes an alternative because the use of frozen-thawed boar sperm results in low fertility. Sperm viability, mitochondrial activity, capacitation and acrosome integrity of freshly ejaculated boar semen stored in vitro for up to 48 h at 4 degrees C, 15 degrees C, 20 degrees C and 39 degrees C was characterized during the study. The viability of boar sperm was assessed by both Hoechst 33258 and SYBR-14/PI staining. Mitochondrial function was assessed by JC-1 staining. Capacitation status was determined by chlortetracycline (CTC)/Hoechst 33258 staining. The acrosome integrity was analysed with Coomassie blue staining. These data were derived from three ejaculates each from three crossbred boars. The viabilities assessed with SYBR-14/PI, Hoechst 33258 and JC-1 staining correlated highly (r > 0.980). In freshly ejaculated boar semen, 96 +/- 1% of the sperm did not take up the Hoechst 33258, whereas 95 +/- 2% were stained by SYBR-14 and 96 +/- 2% of the sperm had mitochondria exhibiting positive JC-1 staining. Staining with CTC/Hoechst 33258 suggested that a high percentage of sperm became capacitated after 24 h storage at 15 degrees C and 20 degrees C. There were 62 +/- 2% (15 degrees C) and 89 +/- 2% (20 degrees C) capacitated sperm by 48 h. Moreover, most of the capacitated sperm were acrosome intact. These results suggest that SYBR-14/PI, Hoechst 33258 or JC-1 staining can be used to effectively evaluate the quality of boar sperm during in vitro storage.     

Changes in the Quality of Rabbit Semen in 14 Consecutive Ejaculates Obtained Every 15 Minutes

Modifications of semen quality related to ejaculation frequency is one of the most important and neglected factors from the standpoint of artificial insemination or sperm competition. New Zealand white rabbits ( Oryctolagus cuniculus ) offer an advantageous experimental model because they have characteristic sexual behavior, they present rapid ejaculation after a single intromission, they have a very short interval between successive ejaculations, and semen can be easily collected. The authors studied the modifications on sperm quality (semen volume, sperm concentration, sperm motility) produced by 14 consecutive ejaculations recovered every 15 min using stimulus females and an artificial vagina. Bucks were exposed every 15 min to a sexually receptive female. After each ejaculation the female was removed and reintroduced 15 min later. Sperm concentration showed a clear biphasic conduct. The amount of spermatozoa per milliliter decreased rapidly until ejaculate number 6, showed a highly significant increase in ejaculates 7-9, and decreased to nil in the last 2 ejaculates. Total number of ejaculated spermatozoa was 557 &#50 10 6, 76% of which were recovered from the first 4 ejaculates. Ejaculate volume also showed a biphasic conduct. In the first ejaculates the volume decreased linearly until ejaculate number 6, showed a significant increase in ejaculates 7-10, and then decreased. The total semen volume recovered during the experiment was 2.44mL, 40% of which (0.98mL) was recovered from the first 2 ejaculates. Individual motility in the first 6 ejaculates was preferentially progressive (60% of the sperms) and turned to random or in situ from the seventh ejaculate up. The proportion of spermatozoa with cytoplasmic droplets increased from ejaculates 6 and 7 up. The results seem to reflect an acceleration of semen transport through the epididymis when the demands for spermatozoa increase.     

Development of sperm sexing and associated assisted reproductive technology for sex preselection of captive bottlenose dolphins (Tursiops truncatus)

Research was conducted to develop sperm sorting and novel sperm preservation methodologies for sex predetermination in the bottlenose dolphin (Tursiops truncatus) using artificial insemination. In Study 1, the effect of seminal plasma (SP), sperm concentration and freezing rate (FR) on in vitro sperm quality of liquid-stored, non-sorted spermatozoa was examined. There was no effect (P > 0.05) of prefreeze SP addition on post-thaw quality (progressive motility, kinetic rating, sperm motility index (SMI), viability and acrosome integrity). Post-thaw motility parameters and viability were higher (P < 0.05) for slow FR than fast FR samples. In Study 2 investigating the effects of liquid storage and sorting on sperm quality, motility and SMI after sorting and centrifugation were lower (P < 0.05) than those of the initial ejaculate. The sort rate for enrichment (91 +/- 4% purity) of X- and Y-bearing spermatozoa was 3400 +/- 850 spermatozoa sex(-1) s(-1). In Study 3, compared with a modified straw method, directional freezing resulted in enhanced in vitro quality of sorted and non-sorted spermatozoa derived from liquid-stored semen (P < 0.05). In Study 4, endoscopic insemination of three dolphins with sorted, frozen-thawed X-bearing spermatozoa resulted in one conception and the birth of a female calf. High-purity sorting of dolphin spermatozoa, derived from liquid-stored semen, can be achieved with minimal loss of in vitro sperm quality and samples are functional in vivo.     

Optimum Measurement Time for Human Sperm Velocity Determination

The optimal time-duration required by spermatozoa to travel distances from which their average velocity can be accurately determined was investigated. For this purpose, 180 motile spermatozoa, from 12 seminal specimens, were photographed while being illuminated by six light pulses, given at 1-sec intervals. From these photographs, distances traveled by each spermatozoon were measured over identical time periods. Most individual spermatozoa showed slight deviation of their velocity when measured sequentially for 5 sec. Nevertheless, when mean sperm velocity of grouped spermatozoa were determined from 1- or 5-sec measurement time, the results were almost identical. This was attributed to a pure statistical law, that is, the mean deviation decreases as the number of spermatozoa in the sample increases. It was concluded that the sperm velocity of a certain specimen can be determined properly using a 1-sec measurement time. An extended period of time does not seem to contribute to the accuracy of sperm velocity determination.     

Trace elements and sperm parameters in semen of male partners of infertile couples

OBJECTIVE: The relationships between element concentrations and sperm parameters in semen samples were investigated. METHODS: Semen samples (n = 113) were donated voluntarily by male partners of infertile couples. The concentrations of fourteen elements (Na, K, P, Ca, Zn, Mg, Fe, Cu, Se, Mn, Sn, Co, Ni, and Cd) in semen were determined by atomic absorption spectrometry, fluorometry, or colorimetry. Element concentrations in seminal plasma and in sperm were also measured. RESULTS: Element concentrations in semen were in the order Na > P, K > Ca > Zn > Mg > > Fe> Cu, Se > Mn > Sn, Ni, Co, Cd. When the samples were divided into two groups in terms of sperm concentration and number, the Se concentration in semen with normal parameter values (sperm concentration > or = 20 x 10(6) and sperm number > or = 40 x 10(6)), 99.4 +/- 37.4 ng/ml, was higher than that in semen with abnormal parameter values (sperm concentration < or = 20 x 10(6) and/or sperm number < or = 40 x 10(6)), 72.1 +/- 33.9 ng/ml (p < 0.001). A clearer positive correlation between the Se concentration and the sperm concentration was observed in the sperm portion (r = 0.853, p < 0.001) than in semen (r = 0.512, p < 0.001) and seminal plasma (r = 0.292, p = 0.003). Statistically significant correlations were also observed between the concentration of Se, P, Zn, Cu, Fe, or Mn in semen, the sperm portion or seminal plasma and the sperm concentration, semen volume or abnormal morphology, although correlation coefficients were small. CONCLUSION: Among biologically essential elements in semen of infertile males, Se was a good indicator of sperm concentration; however, other trace elements did not indicate clear relationships between their concentrations and sperm parameters.     

Giant panda (Ailuropoda melanoleuca) spermatozoon decondensation in vitro is not compromised by cryopreservation

Natural breeding of giant pandas in captivity is compromised, making artificial insemination and spermatozoa cryopreservation essential for genetic management. This study examined the influence of freeze-thawing on traditional parameters such as motility and spermatozoon functionality, specifically decondensation in vitro. Giant panda spermatozoa were assessed before and after rapid cryopreservation (4 degrees C to -130 degrees C over 2 min) in liquid nitrogen vapour. Spermatozoa pre-incubated in medium for 6 h were co-incubated with cat zonae (2 zonae microL(-1)) for 30 min to effect capacitation and an acrosome reaction. Spermatozoa were then mixed with mature cat oocyte cytoplasm (2 cytoplasm microL(-1)) for 4 h and evaluated for decondensation. Frozen spermatozoa were less motile (P < 0.05) than fresh counterparts immediately post-thawing, but not after 6 h incubation. There were more (P < 0.05) spermatozoa with completely diffused chromatin post-thaw (10.4 +/- 1.3%; mean +/- s.e.m.) compared to fresh counterparts (5.1 +/- 1.0%). However, there was no overall difference (P > 0.05) in the incidence of decondensation between fresh (4 h, 69.8 +/- 5.9%) and thawed (4 h, 71.5 +/- 4.9%) spermatozoa after exposure to cat oocyte cytoplasm. It is concluded that the 'rapid' method now used to cryopreserve giant panda spermatozoa has little impact on spermatozoon decondensation.     

Development of infertility at young adult age in a mouse model of human Sandhoff disease

Sandhoff disease is a human lysosomal storage disease. In a knockout mouse model of Sandhoff disease, which lacks the beta-subunit of beta-hexosaminidase A (Hex A, alphabeta subunits) and B (Hex B, betabeta subunits), the mutant homozygous mice (Hexb(-/-)) are healthy until 15 weeks of age when they develop neurodegenerative symptoms. This study was designed to analyse the fertility profile of male and female Hexb(-/-) mice. Mating behaviour of Hexb(-/-) mice was assessed at different ages. The ovarian function of Hexb(-/-) females was determined by superovulation studies. The quality of spermatozoa and ova was assessed by an in vitro fertilization (IVF) procedure. Hexb(-/-) mice were fertile at a young age. Males were fertile up to the age of 69.3 +/- 6.3 days (mean +/- SD) and females were fertile up to the age of 56-63 days. Since both the Hexb (-/-) sexes showed fertility, the results indicate that Hex A and Hex B (major isozymes of beta-hexosaminidase) may not be required for sperm-ovum interactions, in contrast to the widely accepted belief. On the other hand, young adult Hexb(-/-) males showed a reduction in mating behaviour at the age of 84.8 +/- 2.2 days and an absence of mating behaviour at 94.2 +/- 2.0 days. Spermatozoa from Hexb(-/-) mice (aged 109.2 +/- 1.8 days) showed a lower IVF rate. Among Hexb (-/-) females aged 85.6 +/- 2.1 days, no mice became pregnant although they were positive for a vaginal plug when caged with fertile males. The number of ova recovered from Hexb(-/-) females (aged 111.0 +/- 3.1 days) and the IVF rate of ova were lower than those of controls. In conclusion, Hex A and Hex B may not be required for sperm-ovum interactions. Mice lacking Hex A and Hex B activities develop infertility at a young adult age in an age-dependent manner.     

Semen quality and reproductive endocrine function in relation to biomarkers of lead, cadmium, zinc, and copper in men

Blood lead (BPb), activity of delta-aminolevulinic acid dehydratase (ALAD), erythrocyte protoporphyrin (EP), blood cadmium (BCd), serum zinc (SZn), seminal fluid zinc (SfZn), serum copper (SCu), and parameters of semen quality and of reproductive endocrine function were measured in 149 healthy male industrial workers 20-43 years of age. The group contained 98 subjects with slight to moderate occupational exposure to Pb and 51 reference subjects. All of the subjects lived in Zagreb, Croatia. Significant (p < 0.05) correlations of BPb, ALAD, and/or EP with reproductive parameters indicated a Pb-related decrease in sperm density, in counts of total, motile, and viable sperm, in the percentage and count of progressively motile sperm, in parameters of prostate secretory function (SfZn, acid phosphatase, and citric acid in seminal fluid), and an increase in abnormal sperm head morphology, serum testosterone, and estradiol. These associations were confirmed by results of multiple regression, which also showed significant (p < 0. 05) influence of BCd, SZn, SCu, smoking habits, alcohol consumption, or age on certain reproductive parameters. These effects were mainly of lower rank and intensity as compared to Pb-related reproductive effects, whereas BCd contributed to a decrease in sperm motility and an increase in abnormal sperm morphology and serum testosterone. No significant Pb- or Cd-related influence was found on levels of the lactate dehydrogenase isoenzyme LDH-C(4) and fructose in seminal fluid or on follicle-stimulating hormone, luteinizing hormone, and prolactin in serum. The seminal fluid concentrations of Pb (SfPb) and Cd (SfCd) were measured in 118 of the 149 subjects, and a highly significant (p < 0.0001) correlation was found between BPb and SfPb levels (r = 0.571) and between BCd and SfCd levels (r = 0.490). The overall study results indicate that even moderate exposures to Pb (BPb < 400 microg/L) and Cd (BCd < 10 microg/L) can significantly reduce human semen quality without conclusive evidence of impairment of male reproductive endocrine function.     

精索静脉曲张不育患者8-羟基脱氧鸟苷水平的检测分析

目的:检测人血清及精浆中8-羟基脱氧鸟苷(8-OHdG)水平,探讨男性精索静脉曲张(VC)患者体内氧化损伤情况。方法:对41例VC不育患者、33例正常生育者进行精液分析,用8-OHdG酶联免疫试剂盒检测两组对象血清及精浆中8-OHdG水平,采用Wilcoxon两样本比较法对数据进行统计分析。结果:正常生育组的精子浓度、前向活动率、存活率均高于VC不育组(P<0.05),血清8-OHdG水平(中位数34.83ng/ml)低于VC组(中位数76.87ng/ml,P<0.05),精浆8-OHdG水平(中位数18.51ng/ml)低于VC组(中位数22.19ng/ml,P<0.05)。血清中8-OHdG水平高于精浆(P<0.05)。血清8-OHdG水平与精子存活率、前向活动率呈负相关(γ=-0.293,P<0.05;γ=-0.243,P<0.05);精浆8-OHdG水平与精子存活率、前向活动率呈负相关(γ=-0.327,P<0.05;γ=-0.275,P<0.05);血清及精浆中8-OHdG水平与精子浓度、正常形态率不相关。结论:VC患者血清和精浆中8-OHdG水平高于正常生育组,表明VC患者可能存在DNA氧化损伤,导致精子活力下降。     

Inorganic lead exposure in battery and paint factory: effect on human sperm structure and functional activity

Lead is one of the industrially important heavy metals that causes male reproductive impairment among battery and paint factory workers, but information on the structure-function integrity of human spermatozoa is still limited. Therefore, it was necessary to investigate the effect of lead on sperm structure and functional activity in these workers. Oligozoospermia with concomitant lowering of sperm protein and nucleic acid content and the percentage of sperm DNA hyploidy (P <0.001) suggested the diminution of sperm cell production after occupational lead exposure. Low sperm vitality and hypoosmotic swelling percentage along with high malondialdehyde content and altered seminal plasma ascorbate level (P<0.001) indicating damage of sperm cell surface, might be due to high membrane lipid peroxidation and failure of non-enzymatic antioxidant protection after lead exposure. Alteration of sperm membrane surface was also evidenced from scanning electron microscopy and further authenticated by atomic and lateral force microscopy. Lowering of sperm velocity, gross and forward progressive motility with high stationary motile spermatozoa (P<0.001) suggested retarded sperm activity among the exposed workers, which was supported by high seminal plasma fructose level and reduced activity of sperm ATPase (P < 0.001). Increased incidence of teratozoospermia was also associated with high blood and semen lead level (PbB, PbS) (P<0.001). Therefore, the results suggested that lead not only affects the sperm count, but also damages the sperm structure and membrane integrity, motility and functional activity among the battery and paint factory workers.     

Continuous oral low-dosage cyproterone acetate for fertility regulation in the male? — A trend analysis in 15 volunteers —

As part of the “WHO Task Force Program on Regulation of Male Fertility”, a study has been carried out to investigate the effects of 10 or 20 mg cyproterone acetate (CPA) daily over 26 weeks in 15 healthy young males. Control periods of 18 and 16 weeks, respectively, preceded and followed the phase of CPA medication. The salient observations under treatment were: A decrease in sperm density and in normal-shaped sperm with conversely an increase of pathological and immature forms, an increase in dead spermatozoa, a relative reduction of motile spermatozoa, a distinct drop in the speed of spermatozoa and a distinct decrease in the ability of the spermatozoa to migrate through the cervical mucus of ovulation quality $\text{in vitro}$. Androgen levels (mainly testosterone) in blood plasma dropped significantly to about 40% of the basal values, without concurrent effects on sexual behaviour; no changes in plasma LH and FSH were observed; alkaline phosphatase values in seminal plasma were found to be considerably elevated during treatment; no changes in fructose and sialic acid levels in seminal plasma were seen. CPA also increased significantly the cortisol-binding capacity of transcortin. The investigations clearly demonstrate a distinct inhibitory influence of low-dosage CPA on several parameters of male fertility. These reversible effects appear to indicate that low doses of CPA might be used for fertility control in the male, but final conclusions in this respect cannot be drawn before a large scale phase three study on this issue has been performed.     

Fertility of male and female emus (Dromaius novaehollandiae) as determined by spermatozoa trapped in eggs

Changes in the fertility status of 10 pairs of emus were investigated using egg break-out and numbers of sperm in the perivitelline membrane of the germinal disc (GD) region. After the sexes were separated, sperm in consecutive eggs declined approximately logarithmically at a mean (+/-SEM, n = 10 females) rate of -0.148 +/- 0.021 per log day. Sperm continued to be detected in eggs for 16.5 +/- 1.7 days during which 5.6 +/- 0.6 fertilized eggs were laid. Fertilized eggs that did not contain detectable sperm were laid by five females for a further 2.2 +/- 0.9 days. Based on break-out fertility, the fertile period continued for up to 18.7 +/- 2.1 days, for which the mean number of laid eggs was 6.3 +/- 0.8. An egg with a 50:50 chance of being fertilized would contain 3.5 sperm mm(-2) of GD. Based on the sperm decline model, an egg containing that many sperm would be laid 21 days after the last copulation. In emus that were not separated and allowed to incubate their eggs (n = 3 pairs), the number of sperm in eggs laid before and during incubation declined in a manner similar to that after the last copulation and egg-laying stopped after the females had laid 3.3 +/- 0.3 eggs. After incubation was terminated, females resumed laying within 8.3 +/- 1.2 days and the number of sperm in eggs gradually increased but it did not return to pre-incubation levels. In non-incubating emus (three pairs), the number of sperm in eggs declined as laying progressed, although lit was higher during the period when the first seven eggs were laid than during the period when the rest of eggs were laid (214 +/- 39 v.100 +/- 16 sperm mm(-2) of GD). Sperm numbers varied between successive eggs but a sharp increase followed by a decrease acted as an indicator of recent copulation. There were 8.7 +/- 0.3 such increases per laying period (one per 2.8 +/- 0.2 eggs), a frequency that suggests that emus copulate once weekly. In conclusion, as long as a female emu is supplied with sperm on a weekly basis, she will be fertile but, when copulations stop, she will stop laying soon after. Male fertility appears to fall towards the end of the laying season and it can be affected by egg incubation at any time of the season.