Freezing African Elephant Semen as a New Population Management Tool

Background

The captive elephant population is not self-sustaining and with a limited number of breeding bulls, its genetic diversity is in decline. One way to overcome this is to import young and healthy animals from the wild. We introduce here a more sustainable alternative method - importation of semen from wild bulls without removing them from their natural habitat. Due to the logistics involved, the only practical option would be to transport cryopreserved sperm. Despite some early reports on African elephant semen cryopreservation, the utility of this new population management tool has not been evaluated.

Methodology/Principal Findings

Semen was collected by electroejaculation from 14 wild African savanna elephant (Loxodonta africana) bulls and cryopreserved using the directional freezing technique. Sperm treatments evaluated included the need for centrifugation, the use of hen or quail yolk, the concentration of glycerol (3%, 5% or 7%) in the extender, and maintenance of motility over time after thawing. Our results suggest that dilution in an extender containing hen yolk and 7% glycerol after centrifugation best preserved post-thaw sperm motility when compared to all other treatments (P≤0.012 for all). Using this approach we were able to achieve after thawing (mean ± SD) 54.6±3.9% motility, 85.3±2.4% acrosome integrity, and 86.8±4.6% normal morphology with no decrease in motility over 1 h incubation at 37°C. Sperm cryopreserved during this study has already lead to a pregnancy of a captive female elephant following artificial insemination.

Conclusions/Significance

With working techniques for artificial insemination and sperm cryopreservation of both African and Asian elephants in hand, population managers can now enrich captive or isolated wild elephant populations without removing valuable individuals from their natural habitat.     

The Effect of Low-Level Laser Irradiation on Sperm Motility,and Integrity of the Plasma Membrane and Acrosome in Cryopreserved Bovine Sperm

Background and Objective

Freezing changes sperm integrity remarkably. Cryopreservation involves cooling, freezing, and thawing and all these contribute to structural damage in sperm, resulting in reduced fertility potential. Low-level laser irradiation (LLLI) could increase energy supply to the cell and cause reactive oxygen species reduction (ROS), contributing to the restoration of oxygen consumption and adenosine triphosphate synthesis (ATP) in the mitochondria. Our goal was to analyze the effects of low-level laser irradiation on sperm motility and integrity of the plasma membrane and acrosome in cryopreserved bovine sperm.

Study Design/Materials and Methods

We analyzed 09 samples of bull semen (Bos taurus indicus), divided into three groups: a control group without laser irradiation, a 4J group subjected to a laser irradiation dose of 4 joules, and a 6J group subjected to dose of 6 joules. Samples were divided for the analysis of cell viability and acrosomal membrane integrity using flow cytometry; another portion was used for motion analysis. Irradiation was performed in petri dishes of 30 mm containing 3 ml of semen by an aluminum gallium indium phosphide laser diode with a wavelength of 660 nm, 30 mW power, and energy of 4 and 6 joules for 80 and 120 seconds respectively. Subsequently, the irradiated and control semen samples were subjected to cryopreservation and analyzed by flow cytometry (7AAD and FITC-PSA) using the ISAS - Integrated Semen Analysis System.

Results

Flow cytometry showed an increase in the percentage of live sperm cells and acrosome integrity in relation to control cells when subjected to irradiation of low-power laser in two different doses of 4 and 6 joules (p < 0.05). In the analysis of straightness, percentage of cell movement, and motility, a dose of 4 joules was more effective (p < 0.05).

Conclusion

We conclude that LLLI may exert beneficial effects in the preservation of live sperm. A dose of 4 joules prior to cryopreservation was more effective than a dose of 6 joules in preserving sperm motility.     

Widespread epigenetic abnormalities suggest a broad DNA methylation erasure defect in abnormal human sperm

Background

Male-factor infertility is a common condition, and etiology is unknown for a high proportion of cases. Abnormal epigenetic programming of the germline is proposed as a possible mechanism compromising spermatogenesis of some men currently diagnosed with idiopathic infertility. During germ cell maturation and gametogenesis, cells of the germ line undergo extensive epigenetic reprogramming. This process involves widespread erasure of somatic-like patterns of DNA methylation followed by establishment of sex-specific patterns by de novo DNA methylation. Incomplete reprogramming of the male germ line could, in theory, result in both altered sperm DNA methylation and compromised spermatogenesis.

Methodology/Principal Finding

We determined concentration, motility and morphology of sperm in semen samples collected by male members of couples attending an infertility clinic. Using MethyLight and Illumina assays we measured methylation of DNA isolated from purified sperm from the same samples. Methylation at numerous sequences was elevated in DNA from poor quality sperm.

Conclusions

This is the first report of a broad epigenetic defect associated with abnormal semen parameters. Our results suggest that the underlying mechanism for these epigenetic changes may be improper erasure of DNA methylation during epigenetic reprogramming of the male germ line.     

Semen Abnormalities,Sperm DNA Damage and Global Hypermethylation in Health Workers Occupationally Exposed to Ionizing Radiation

Background

Cytogenetic studies have demonstrated that low levels of chronic radiation exposure can potentially increase the frequency of chromosomal aberrations and aneuploidy in somatic cells. Epidemiological studies have shown that health workers occupationally exposed to ionizing radiation bear an increased risk of hematological malignancies.

Objectives

To find the influence of occupational radiation exposure on semen characteristics, including genetic and epigenetic integrity of spermatozoa in a chronically exposed population.

Methods

This cross sectional study included 134 male volunteers of which 83 were occupationally exposed to ionizing radiation and 51 were non-exposed control subjects. Semen characteristics, sperm DNA fragmentation, aneuploidy and incidence of global hypermethylation in the spermatozoa were determined and compared between the non-exposed and the exposed group.

Results

Direct comparison of the semen characteristics between the non-exposed and the exposed population revealed significant differences in motility characteristics, viability, and morphological abnormalities (P<0.05–0.0001). Although, the level of sperm DNA fragmentation was significantly higher in the exposed group as compared to the non-exposed group (P<0.05–0.0001), the incidence of sperm aneuploidy was not statistically different between the two groups. However, a significant number of hypermethylated spermatozoa were observed in the exposed group in comparison to non-exposed group (P<0.05).

Conclusions

We provide the first evidence on the detrimental effects of occupational radiation exposure on functional, genetic and epigenetic integrity of sperm in health workers. However, further studies are required to confirm the potential detrimental effects of ionizing radiation in these subjects.     

Consistent Paternity Skew through Ontogeny in Peron's Tree Frog (Litoria peronii)

Background

A large number of studies in postcopulatory sexual selection use paternity success as a proxy for fertilization success. However, selective mortality during embryonic development can lead to skews in paternity in situations of polyandry and sperm competition. Thus, when assessment of paternity fails to incorporate mortality skews during early ontogeny, this may interfere with correct interpretation of results and subsequent evolutionary inference. In a previous series of in vitro sperm competition experiments with amphibians (Litoria peronii), we showed skewed paternity patterns towards males more genetically similar to the female.

Methodology/Principal Findings

Here we use in vitro fertilizations and sperm competition trials to test if this pattern of paternity of fully developed tadpoles reflects patterns of paternity at fertilization and if paternity skews changes during embryonic development. We show that there is no selective mortality through ontogeny and that patterns of paternity of hatched tadpoles reflects success of competing males in sperm competition at fertilization.

Conclusions/Significance

While this study shows that previous inferences of fertilization success from paternity data are valid for this species, rigorous testing of these assumptions is required to ensure that differential embryonic mortality does not confound estimations of true fertilization success.     

The Effect of Chronic Prostatitis/Chronic Pelvic Pain Syndrome (CP/CPPS) on Semen Parameters in Human Males: A Systematic Review and Meta-Analysis

Background

Chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) is one of the risk factors of impaired male fertility potential. Studies have investigated the effect of CP/CPPS on several semen parameters but have shown inconsistent results. Hence, we performed a systematic literature review and meta-analysis to assess the association between CP/CPPS and basic semen parameters in adult men.

Methods

Systematic literature searches were conducted with PubMed, EMBASE and the Cochrane Library up to August 2013 for case-control studies that involved the impact of CP/CPSS on semen parameters. Meta-analysis was performed with Review Manager and Stata software. Standard mean differences (SMD) of semen parameters were identified with 95% confidence intervals (95% CI) in a random effects model.

Results

Twelve studies were identified, including 999 cases of CP/CPPS and 455 controls. Our results illustrated that the sperm concentration and the percentage of progressively motile sperm and morphologically normal sperm from patients with CP/CPPS were significantly lower than controls (SMD (95% CI) −14.12 (−21.69, −6.63), −5.94 (−8.63, −3.25) and −8.26 (−11.83, −4.66), respectively). However, semen volume in the CP/CPPS group was higher than in the control group (SMD (95% CI) 0.50 (0.11, 0.89)). There was no significant effect of CP/CPPS on the total sperm count, sperm total motility, and sperm vitality.

Conclusions

The present study illustrates that there was a significant negative effect of CP/CPPS on sperm concentration, sperm progressive motility, and normal sperm morphology. Further studies with larger sample sizes are needed to better illuminate the negative impact of CP/CPPS on semen parameters.     

The Adaptive Function of Masturbation in a Promiscuous African Ground Squirrel

Background

Studies of animal mating systems increasingly emphasize female multiple mating and cryptic sexual selection, particularly sperm competition. Males under intense sperm competition may manipulate sperm quantity and quality through masturbation, which could waste sperm and decrease fertility. I examined the factors influencing masturbation by male Cape ground squirrels (Xerus inauris) in light of a number of functional hypotheses.

Methodology

Observational data on a marked population of squirrels were collected in east-central Namibia using scan and all-occurrences sampling.

Findings

Masturbation was far more frequent on days of female oestrus and mostly occurred after copulation. Masturbation rates were higher in dominant males, which copulate more, than in subordinates and increased with number of mates a female accepts.

Conclusions

These results suggest that masturbation in this species was not a response to sperm competition nor a sexual outlet by subordinates that did not copulate. Instead masturbation could function as a form of genital grooming. Female Cape ground squirrels mate with up to 10 males in a 3-hr oestrus, and by masturbating after copulation males could reduce the chance of infection. Sexually transmitted infections (STIs) can profoundly affect fertility, and their consequences for mating strategies need to be examined more fully.     

The Impact of Paternal and Maternal Smoking on Semen Quality of Adolescent Men

Background

Maternal smoking during pregnancy has been reported to negatively impact sperm counts of the sons. Sufficient data on the effect of paternal smoking is lacking.

Objectives

We wished to elucidate the impact of maternal and paternal smoking during pregnancy and current own smoking on reproductive function of the male offspring.

Methods

Semen parameters including sperm DNA integrity were analyzed in 295 adolescents from the general population close to Malmö, Sweden, recruited for the study during 2008–2010. Information on maternal smoking was obtained from the Swedish Medical Birth Register, and regarding own and paternal smoking from questionnaires. The impacts of maternal, paternal and own smoking were evaluated in a multivariate regression model and by use of models including interaction terms. Totally, three exposures and five outcomes were evaluated.

Results

In maternally unexposed men, paternal smoking was associated with 46% lower total sperm count (95%CI: 21%, 64%) in maternally unexposed men. Both paternal and maternal smoking were associated with a lower sperm concentration (mean differences: 35%; 95%CI: 8.1%, 55% and 36%; 95%CI: 3.9%, 57%, respectively) if the other parent was a non-smoker. No statistically significant impact of own smoking on semen parameters was seen.

Conclusions

Prenatal both maternal and paternal smoking were separately associated with some decrease in sperm count in men of whom the other parent was not reported to smoke.     

The relationship between anogenital distance, fatherhood, and fertility in adult men

Background

Anogenital distance (AGD), a sexually dimorphic measure of genital development, is a marker for endocrine disruption in animal studies and may be shorter in infant males with genital anomalies. Given the correlation between anogenital distance and genital development, we sought to determine if anogenital distance varied in fertile compared to infertile adult men.

Methods

A cross sectional study of consecutive men being evaluated for infertility and men with proven fertility was recruited from an andrology clinic. Anogenital distance (the distance from the posterior aspect of the scrotum to the anal verge) and penile length (PL) were measured using digital calipers. ANOVA and linear regression were used to determine correlations between AGD, fatherhood status, and semen analysis parameters (sperm density, motility, and total motile sperm count).

Findings

A total of 117 infertile men (mean age: 35.3±17.4) and 56 fertile men (mean age: 44.8±9.7) were recruited. The infertile men possessed significantly shorter mean AGD and PL compared to the fertile controls (AGD: 31.8 vs 44.6 mm, PL: 107.1 vs 119.5 mm, p<0.01). The difference in AGD persisted even after accounting for ethnic and anthropomorphic differences. In addition to fatherhood, on both unadjusted and adjusted linear regression, AGD was significantly correlated with sperm density and total motile sperm count. After adjusting for demographic and reproductive variables, for each 1 cm increase in a man''s AGD, the sperm density increases by 4.3 million sperm per mL (95% CI 0.53, 8.09, p = 0.03) and the total motile sperm count increases by 6.0 million sperm (95% CI 1.34, 10.58, p = 0.01). On adjusted analyses, no correlation was seen between penile length and semen parameters.

Conclusion

A longer anogenital distance is associated with fatherhood and may predict normal male reproductive potential. Thus, AGD may provide a novel metric to assess reproductive potential in men.     

Do Genetic Diversity Effects Drive the Benefits Associated with Multiple Mating? A Test in a Marine Invertebrate

Background

Mothers that mate with multiple males often produce higher quality offspring than mothers that mate with a single male. By engaging in polyandry, mothers may increase their chances of mating with a compatible male or promote sperm competition - both of which act to increase maternal fitness via the biasing of the paternity of offspring. Surprisingly, mating with multiple males, can carry benefits without biasing paternity and may be due simply to differences in genetic diversity between monandrous and polyandrous clutches but role of genetic diversity effects in driving the benefits of polyandry remains poorly tested. Disentangling indirect, genetic benefits from genetic diversity effects is challenging but crucial if we are to understand the selection pressures acting to promote polyandry.

Methodology/Principal Findings

Here, we examine the post-fertilisation benefits of accessing the sperm of multiple males in an externally fertilising polychaete worm. Accessing the sperm of multiple males increases offspring performance but this benefit was driven entirely by genetic diversity effects and not by the biasing of paternity at fertilisation.

Conclusions/Significance

Previous studies on polyandry should be interpreted cautiously as genetic diversity effects alone can explain the benefits of polyandry yet these diversity effects may be difficult to disentangle from other mechanisms. We suggest that future studies use a modified experimental design in order to discriminate between genetic diversity effects and indirect, genetic benefits.     

Genetic integrity of black sea bream (Acanthopagrus schlegeli) sperm following cryopreservation

Although semen cryopreservation has been applied successfully in many fish species, extensive variation in post‐thaw semen quality exists between species and individuals. AFLP (amplified restriction fragment length polymorphism) is a powerful method for detecting DNA polymorphisms at the individual, population, and species levels. The method has been successfully applied to boars (Sus domestica, Suidae, Artiodactyla, Mammalia) to detect and evaluate differences in DNA sequences that correspond with semen integretiy after employing various freezing techniques. Freezing and thawing of semen has also an effect of selecting for freezing‐resistant (or intact) and eliminating non‐viable or defective sperm. Only the fully intact and functional sperm, despite potential compromise by adverse freezing and osmotic stresses, retain fertility after thawing. The objective of this study was to use AFLP to assess any genetic changes associated with the effect of employed cryo‐methodology on the genetic integrity of sperm of the black sea bream (Acanthopagrus schlegeli) under different cryopreservation treatments. The cryopreservation protocols had no significant effect on sperm motility or survival rate of fertilized ova regardless of using fresh (% motile sperm 89.6 ± 3.0; % embryonic survival rate 54.4 ± 2.9) and frozen‐thawed semen (% motile sperm 80.2 ± 2.0; % embryonic survival rate 51.8 ± 2.0). The post‐thaw sperm motility and survival rates were not significantly different among the sperm samples of the five black sea bream males examined. In the present study, the remaining black sea bream sperm that survive the cryopreservation limit the power of AFLP to trace the genetic markers which correlate with the differences in the sensitivity of sperm to cryo‐injury. It is also possible that point mutations outside the AFLP priming sites may not have been detected. More thorough investigations are needed to determine whether such DNA fingerprints would be found in fish species.     

Female choice for males with greater fertilization success in the Swedish Moor frog, Rana arvalis

Background

Studies of mate choice in anuran amphibians have shown female preference for a wide range of male traits despite females gaining no direct resources from males (i.e. non-resource based mating system). Nevertheless, theoretical and empirical studies have shown that females may still gain indirect genetic benefits from choosing males of higher genetic quality and thereby increase their reproductive success.

Methodology/Principal Findings

We investigated two components of sexual selection in the Moor frog (Rana arvalis), pre-copulatory female choice between two males of different size (‘large’ vs. ‘small’), and their fertilization success in sperm competition and in isolation. Females'' showed no significant preference for male size (13 small and six large male preferences) but associated preferentially with the male that subsequently was the most successful at fertilizing her eggs in isolation. Siring success of males in competitive fertilizations was unrelated to genetic similarity with the female and we detected no effect of sperm viability on fertilization success. There was, however, a strong positive association between a male''s innate fertilization ability with a female and his siring success in sperm competition. We also detected a strong negative effect of a male''s thumb length on his competitive siring success.

Conclusions/Significance

Our results show that females show no preference for male size but are still able to choose males which have greater fertilization success. Genetic similarity and differences in the proportion of viable sperm within a males ejaculate do not appear to affect siring success. These results could be explained through pre- and/or postcopulatory choice for genetic benefits and suggest that females are able to perceive the genetic quality of males, possibly basing their choice on multiple phenotypic male traits.     

Extreme Female Promiscuity in a Non-Social Invertebrate Species

Background

While males usually benefit from as many matings as possible, females often evolve various methods of resistance to matings. The prevalent explanation for this is that the cost of additional matings exceeds the benefits of receiving sperm from a large number of males. Here we demonstrate, however, a strongly deviating pattern of polyandry.

Methodology/Principal Findings

We analysed paternity in the marine snail Littorina saxatilis by genotyping large clutches (53–79) of offspring from four females sampled in their natural habitats. We found evidence of extreme promiscuity with 15–23 males having sired the offspring of each female within the same mating period.

Conclusions/Significance

Such a high level of promiscuity has previously only been observed in a few species of social insects. We argue that genetic bet-hedging (as has been suggested earlier) is unlikely to explain such extreme polyandry. Instead we propose that these high levels are examples of convenience polyandry: females accept high numbers of matings if costs of refusing males are higher than costs of accepting superfluous matings.     

The Long-Term Effects of Stress on Partner Weight Characteristics

Background

Recent experimental evidence suggests that stressed males find heavier women more attractive than non-stressed males. The aim of this study is to examine whether these results also appear in actual mating patterns of adults from a national sample.

Methods

Regression analysis linking partner weight measures to own measures of childhood stress, as measured by mistreatment. Cross-sectional data from the National Longitudinal Study of Adolescent Health, Romantic Partners Sample is used to measure partner weight, childhood stressful events, and socio-demographic characteristics. Childhood experiences of adult mistreatment are retrospectively collected.

Results

Men who experienced childhood mistreatment are more likely to have obese female partners during young adulthood. The results are strongest for interactions with social services, adult neglect and physical abuse. We also present novel evidence of the opposite association in similarly stressed women whose male partners are more likely to be thin.

Conclusions

These results suggest that preferences for partner characteristics are sensitive to histories of stress and that previously hypothesized patterns occur outside the experimental setting.     

Establishment and Characterization of Primary Glioblastoma Cell Lines from Fresh and Frozen Material: A Detailed Comparison

Background

Development of clinically relevant tumor model systems for glioblastoma multiforme (GBM) is important for advancement of basic and translational biology. High molecular heterogeneity of GBM tumors is well recognized, forming the rationale for molecular tests required before administration of several of the novel therapeutics rapidly entering the clinics. One model that has gained wide acceptance is the primary cell culture model. The laborious and time consuming process is rewarded with a relative high success rate (about 60%). We here describe and evaluate a very simple cryopreservation procedure for GBM tissue prior to model establishment that will considerably reduce the logistic complexity.

Methods

Twenty-seven GBM samples collected ad hoc were prepared for primary cell culture freshly from surgery (#1) and after cryopreservation (#2).

Results

Take rates after cryopreservation (59%) were as satisfactory as from fresh tissue (63%; p = 1.000). We did not observe any relevant molecular or phenotypic differences between cell lines established from fresh or vitally frozen tissue. Further, sensitivity both towards standard chemotherapeutic agents (Temozolomide, BCNU and Vincristine) and novel agents like the receptor tyrosine kinase inhibitor Imatinib did not differ.

Conclusions

Our simple cryopreservation procedure facilitates collection, long-time storage and propagation (modeling) of clinical GBM specimens (potentially also from distant centers) for basic research, (pre-) clinical studies of novel therapies and individual response prediction.     

Sex Differences in the Rapid Detection of Emotional Facial Expressions

Background

Previous studies have shown that females and males differ in the processing of emotional facial expressions including the recognition of emotion, and that emotional facial expressions are detected more rapidly than are neutral expressions. However, whether the sexes differ in the rapid detection of emotional facial expressions remains unclear.

Methodology/Principal Findings

We measured reaction times (RTs) during a visual search task in which 44 females and 46 males detected normal facial expressions of anger and happiness or their anti-expressions within crowds of neutral expressions. Anti-expressions expressed neutral emotions with visual changes quantitatively comparable to normal expressions. We also obtained subjective emotional ratings in response to the facial expression stimuli. RT results showed that both females and males detected normal expressions more rapidly than anti-expressions and normal-angry expressions more rapidly than normal-happy expressions. However, females and males showed different patterns in their subjective ratings in response to the facial expressions. Furthermore, sex differences were found in the relationships between subjective ratings and RTs. High arousal was more strongly associated with rapid detection of facial expressions in females, whereas negatively valenced feelings were more clearly associated with the rapid detection of facial expressions in males.

Conclusion

Our data suggest that females and males differ in their subjective emotional reactions to facial expressions and in the emotional processes that modulate the detection of facial expressions.     

Variation in Apical Hook Length Reflects the Intensity of Sperm Competition in Murine Rodents

Background

Post-copulatory sexual selection has been shown to shape morphology of male gametes. Both directional and stabilizing selection on sperm phenotype have been documented in vertebrates in response to sexual promiscuity.

Methodology

Here we investigated the degree of variance in apical hook length and tail length in six taxa of murine rodents.

Conclusions

Tail sperm length and apical hook length were positively associated with relative testis mass, our proxy for levels of sperm competition, thus indicating directional post-copulatory selection on sperm phenotypes. Moreover, our study shows that increased levels of sperm competition lead to the reduction of variance in the hook length, indicating stabilizing selection. Hence, the higher risk of sperm competition affects increasing hook length together with decreasing variance in the hook length. Species-specific post-copulatory sexual selection likely optimizes sperm morphology.     

Female control of mate plugging in a female-cannibalistic spider (Micaria sociabilis)

Background

Sperm competition imposes a strong selective pressure on males, leading to the evolution of various physiological, morphological and behavioral traits. Sperm competition can be prevented by blocking or impeding the access to female genitalia by means of a mating plug. We investigated the factors responsible for plug production and function in the promiscuous female-cannibalistic spider Micaria sociabilis (Gnaphosidae).

Results

We performed mating trials using females with and without a plug that consists of an amorphous mass. The mating trials demonstrated that the probability of male plugging increased non-linearly with the duration of copulation. Copulation duration and plug production seem to be controlled by the female. We found that females terminated matings later if males were fast at genital coupling. Whereas incomplete plugs had disappeared on the day following copulation, complete plugs persisted (40%). In matings with females with complete plugs, only a small proportion of males (7%) were able to remove the plug, indicating the high effectiveness of plugging. Moreover, males ceased attempts to copulate with plugged females with higher probability. 3D X-ray microscopy of the female and male genitalia showed that the plug material can extend far into the female genital tract and that the plug material is produced by a massive gland inside the palpal organ of the modified male pedipalps.

Conclusions

Our study demonstrates that the mating plug in M. sociabilis constitutes an effective male strategy to avoid sperm competition that seems to be under female control.

Electronic supplementary material

The online version of this article (doi:10.1186/s12862-014-0278-9) contains supplementary material, which is available to authorized users.     

Towards a semen proteome of the dengue vector mosquito: protein identification and potential functions

Background

No commercially licensed vaccine or treatment is available for dengue fever, a potentially lethal infection that impacts millions of lives annually. New tools that target mosquito control may reduce vector populations and break the cycle of dengue transmission. Male mosquito seminal fluid proteins (Sfps) are one such target since these proteins, in aggregate, modulate the reproduction and feeding patterns of the dengue vector, Aedes aegypti. As an initial step in identifying new targets for dengue vector control, we sought to identify the suite of proteins that comprise the Ae. aegypti ejaculate and determine which are transferred to females during mating.

Methodology and Principal Findings

Using a stable-isotope labeling method coupled with proteomics to distinguish male- and female-derived proteins, we identified Sfps and sperm proteins transferred from males to females. Sfps were distinguished from sperm proteins by comparing the transferred proteins to sperm-enriched samples derived from testes and seminal vesicles. We identified 93 male-derived Sfps and 52 predicted sperm proteins that are transferred to females during mating. The Sfp protein classes we detected suggest roles in protein activation/inactivation, sperm utilization, and ecdysteroidogenesis. We also discovered that several predicted membrane-bound and intracellular proteins are transferred to females in the seminal fluids, supporting the hypothesis that Ae. aegypti Sfps are released from the accessory gland cells through apocrine secretion, as occurs in mammals. Many of the Ae. aegypti predicted sperm proteins were homologous to Drosophila melanogaster sperm proteins, suggesting conservation of their sperm-related function across Diptera.

Conclusion and Significance

This is the first study to directly identify Sfps transferred from male Ae. aegypti to females. Our data lay the groundwork for future functional analyses to identify individual seminal proteins that may trigger female post-mating changes (e.g., in feeding patterns and egg production). Therefore, identification of these proteins may lead to new approaches for manipulating the reproductive output and vectorial capacity of Ae. aegypti.     

Genetic Variants in Meiotic Program Initiation Pathway Genes Are Associated with Spermatogenic Impairment in a Han Chinese Population

Background

The meiotic program initiation pathway genes (CYP26B1, NANOS1 and STRA8) have been proposed to play key roles in spermatogenesis.

Objective

To elucidate the exact role of the genetic variants of the meiosis initiation genes in spermatogenesis, we genotyped the potential functional genetic variants of CYP26B1, NANOS1 and STRA8 genes, and evaluated their effects on spermatogenesis in our study population.

Design, Setting, and Participants

In this study, all subjects were volunteers from the affiliated hospitals of Nanjing Medical University between March 2004 and July 2009 (NJMU Infertile Study). Total 719 idiopathic infertile cases were recruited and divided into three groups according to WHO semen parameters: 201 azoospermia patients (no sperm in the ejaculate even after centrifugation), 155 oligozoospermia patients (sperm counts <20×10⁶/ml) and 363 infertility/normozoospermia subjects (sperm counts >20×10⁶/ml). The control group consisted of 383 subjects with normal semen parameters, all of which had fathered at least one child without assisted reproductive technologies.

Measurements

Eight single nucleotide polymorphisms (SNPs) in CYP26B1, NANOS1 and STRA8 genes were determined by TaqMan allelic discrimination assay in 719 idiopathic infertile men and 383 healthy controls.

Results and Limitations

The genetic variant rs10269148 of STRA8 gene showed higher risk of spermatogenic impairment in the groups of abnormospermia (including azoospermia subgroup and oligozoospermia subgroup) and azoospermia than the controls with odds ratios and 95% confidence intervals of 2.52 (1.29–4.94) and 2.92 (1.41–6.06), respectively (P = 0.006, 0.002 respective). Notably, larger sample size studies and in vivo or in vitro functional studies are needed to substantiate the biological roles of these variants.

Conclusions

Our results provided epidemiological evidence supporting the involvement of genetic polymorphisms of the meiotic program initiation genes in modifying the risk of azoospermia and oligozoospermia in a Han-Chinese population.