Culture media affect sex after IVF treatment-a detailed analysis of explanatory variables.

PURPOSE: The purpose of this study was to provide a detailed analysis of clinical and laboratory factors associated with skewed secondary sex ratio (SSR) after ART. METHOD: Retrospective cohort study of embryos resulting in live births, from frozen and fresh single blastocyst transfers. Embryos were cultured in either G-TL (n = 686) or Sage media (n = 685). Data was analyzed using a multivariate logistic regression model and a mixed model analysis. RESULTS: Significantly more male singletons were born after culture in Sage media compared to G-TL media (odds ratio (OR) 1.34, 95% CI (1.05, 1.70), P = 0.02). Inner cell mass grade B vs A (OR 1.36 95% CI (1.05, 1.76), P = 0.02) and one previous embryo transfer (OR 1.49, 95% CI (1.03, 2.16), P = 0.03) were associated with a significantly higher probability of male child at birth. Factors associated with a reduced probability of male child were expansion grade 3 vs 5 (OR 0.66, 95% CI (10.45, 0.96), P = 0.03) and trophectoderm grade B vs A (OR 0.57, 95% CI (0.44, 0.74), P = 0.00). Male embryos developed significantly faster in Sage media compared to G-TL media for the stages of blastocyst (- 1.12 h, 95% CI (- 2.12, - 0.12)), expanded blastocyst (- 1.35 h, 95% CI (- 2.34, - 0.35)), and hatched blastocyst (- 1.75 h, 95% CI (- 2.99, - 0.52)). CONCLUSION: More male children were born after culture in Sage media compared to G-TL media. Male embryo development was affected by culture media. Our observations suggest that culture media impact male embryo quality selectively, thus potentially favoring the selection of male embryos.

Effects of Boric acid as Maternal Feed Additives on the Development and Sex Ratio of Mouse pups.

Boron is primarily used in industrial applications, with recent interest revolving around its effects on metabolism. In this study, we administered boric acid (BA), which has positive effects on reproduction, in conjunction with feed supplementation to serve as a model for experimental animal development and breeding. The pregnancy performance, offspring development, and biochemical effects of mice given feed supplemented with BA at concentrations of 0 (control group), 250, and 500 ppm (BA groups) were investigated. A total of 18 female Balb-C mice were utilized for pregnancy. The mice were given the BA-supplemented feed during a period encompassing three weeks of pregnancy and three weeks of lactation. The numbers and weights of offspring born in cages on days 19-21 were determined. Blood and tissue samples were collected from the offspring during the third week postnatal, and the malondialdehyde (MDA) and total antioxidant and oxidant status (TAS, TOS, and OSI) levels were determined. A significant increase in female offspring was observed in the groups born to mice fed with BA compared to the control group. Positive development in organ weights was observed in the 250-ppm BA group. The 250-ppm group exhibited a significant increase in TAS compared to the control group, while TOS and MDA levels showed a decrease. Also, the levels of BA groups were found to decrease in both the OSI index serum and organ samples compared to the control group. Thus, the use of 250-ppm BA demonstrated positive effects on female offspring production, organ development, and antioxidant levels.

17ß-Estradiol Induced Sex Reversal and Gonadal Transcriptome Analysis in the Oriental River Prawn (Macrobrachium nipponense): Mechanisms, Pathways, and Potential Harm.

Sex reversal induced by 17ß-estradiol (E2) has shown the potential possibility for monoculture technology development. The present study aimed to determine whether dietary supplementation with different concentrations of E2 could induce sex reversal in M. nipponense, and select the sex-related genes by performing the gonadal transcriptome analysis of normal male (M), normal female (FM), sex-reversed male prawns (RM), and unreversed male prawns (NRM). Histology, transcriptome analysis, and qPCR were performed to compare differences in gonad development, key metabolic pathways, and genes. Compared with the control, after 40 days, feeding E2 with 200 mg/kg at PL25 (PL: post-larvae developmental stage) resulted in the highest sex ratio (female: male) of 2.22:1. Histological observations demonstrated the co-existence of testis and ovaries in the same prawn. Male prawns from the NRM group exhibited slower testis development without mature sperm. RNA sequencing revealed 3702 differentially expressed genes (DEGs) between M vs. FM, 3111 between M vs. RM, and 4978 between FM vs. NRM. Retinol metabolism and nucleotide excision repair pathways were identified as the key pathways for sex reversal and sperm maturation, respectively. Sperm gelatinase (SG) was not screened in M vs. NRM, corroborating the results of the slice D. In M vs. RM, reproduction-related genes such as cathepsin C (CatC), heat shock protein cognate (HSP), double-sex (Dsx), and gonadotropin-releasing hormone receptor (GnRH) were expressed differently from the other two groups, indicating that these are involved in the process of sex reversal. Exogenous E2 can induce sex reversal, providing valuable evidence for the establishment of monoculture in this species.

Dietary crude oil exposure during sex differentiation skewed adult sex ratio towards males in the zebrafish.

Dietary crude oil exposure has detrimental morpho-physiological effects in fishes, including endocrine disruption. However, little is known about how it influences sex differentiation and its potential for skewing sex ratios of populations. Appropriate sex ratio is important for maintaining effective population size and structure. Deviations of these ratios can compromise population growth and maintenance and may induce changes in a species' evolutionary trajectory. We assessed the potential of dietary exposure to crude oil (6.5, 11.4, and 17.5 mg/kg food) to alter sex differentiation in the zebrafish (Danio rerio) (20-35 days post fertilization (dpf)) and subsequently skew the adult (90 dpf) sex ratio. Multiple health- and fitness-related phenotypic traits (i.e., body mass and length, condition factor, heart rate, oxygen consumption, and their capacity to cope with hypoxia) were also assessed to better understand the effects of dietary crude oil exposure. We showed that dietary exposure to crude oil during the process of sex differentiation skewed sex ratio towards males (up to 0.34:1 female to male ratio in the highest oil concentration). Remarkably, this effect occurred independently of affecting physiological variables and female gonad characteristics, thus highlighting just how subtle the effects of dietary crude oil exposure can be. Our results suggest that, although fish were in an apparently healthy state during experimentation, sex ratio was still impacted, potentially compromising the resilience of the population. Therefore, considering how complex chemical mixtures affect organisms at several levels (molecular-individual) in experimental designs is warranted to better understand the implications of the exposures and the hazards that populations face in the wild.

The intersection of bee and flower sexes: pollen presence shapes sex-specific bee foraging associations in sunflower.

Foraging preferences are known to differ among bee taxa, and can also differ between male and female bees of the same species. Similarly, bees can prefer a specific flower sex, particularly if only one sex provides pollen. Such variation in foraging preferences could lead to divergent bee communities visiting different flower sexes of a plant species. We sampled bees visiting sunflowers to characterize bee species richness, abundance, and sex ratios on pollen-fertile and pollen-sterile cultivars. We asked whether female or male bees were more abundant on sunflowers, whether female bees were more abundant on pollen-fertile or pollen-sterile cultivars, and whether pollen presence predicted the sex of sampled bees. We further asked whether the bee community differed between pollen-fertile and pollen-sterile cultivars. Females of most bee species were more abundant on sunflowers compared to males, and females were usually more abundant on pollen-fertile cultivars. In three bee species, pollen presence was predictive of a bee's sex, with females more abundant on pollen-fertile cultivars than males. Further, the bee community differed significantly between pollen-fertile and pollen-sterile cultivars, with two bee species functioning as indicators for pollen-fertile sunflowers. Our results demonstrate that a bee's sex shapes foraging associations on sunflowers and influences abundance between pollen-fertile and pollen-sterile cultivars, and that pollen-fertile and pollen-sterile cultivars are visited by different bee communities. Bee sexes and flower pollen presence may be under-appreciated factors shaping pollination services in both agricultural and natural ecosystems, and could be important considerations for pollination of crops with pollen-fertile and pollen-sterile flowers.

A bacteriocyte symbiont determines whitefly sex ratio by regulating mitochondrial function.

Nutritional symbionts influence host reproduction, but the underlying molecular mechanisms are largely unclear. We previously found that the bacteriocyte symbiont Hamiltonella impacts the sex ratio of the whitefly Bemisia tabaci. Hamiltonella synthesizes folate by cooperation with the whitefly. Folate deficiency by Hamiltonella elimination or whitefly gene silencing distorted whitefly sex ratio, and folate supplementation restored the sex ratio. Hamiltonella deficiency or gene silencing altered histone H3 lysine 9 trimethylation (H3K9me3) level, which was restored by folate supplementation. Genome-wide chromatin immunoprecipitation-seq analysis of H3K9me3 indicated mitochondrial dysfunction in symbiont-deficient whiteflies. Hamiltonella deficiency compromised mitochondrial quality of whitefly ovaries. Repressing ovary mitochondrial function led to distorted whitefly sex ratio. These findings indicate that the symbiont-derived folate regulates host histone methylation modifications, which thereby impacts ovary mitochondrial function, and finally determines host sex ratio. Our study suggests that a nutritional symbiont can regulate animal reproduction in a way that differs from reproductive manipulators.

Role of omega-6 and omega-3 fatty acids in fetal programming.

Maternal nutrition plays a critical role in fetal development and can influence adult onset of disease. Linoleic acid (LA) and alpha-linolenic acid (ALA) are major omega-6 (n-6) and n-3 polyunsaturated fatty acids (PUFA), respectively, that are essential in our diet. LA and ALA are critical for the development of the fetal neurological and immune systems. However, in recent years, the consumption of n-6 PUFA has increased gradually worldwide, and elevated n-6 PUFA consumption may be harmful to human health. Consumption of diets with high levels of n-6 PUFA before or during pregnancy may have detrimental effects on fetal development and may influence overall health of offspring in adulthood. This review discusses the role of n-6 PUFA in fetal programming, the importance of a balance between n-6 and n-3 PUFAs in the maternal diet, and the need of further animal models and human studies that critically evaluate both n-6 and n-3 PUFA contents in diets.

Pollen competition is the mechanism underlying a variety of evolutionary phenomena in dioecious plants.

It has long been known that more pollen grains often arrive on stigmas than there are ovules to fertilize, resulting in pollen competition. Moreover, this competition among pollen grains (gametophytes) depends, in part, on their extensive haploid gene expression. Here I review how this leads to a variety of phenomena in dioecious plants of interest to evolutionary biologists. For example, pollen competition can lead to extreme female-biased sex ratios. In addition, gene expression by individual pollen grains can slow mutation accumulation and degeneration of the Y chromosome. Lastly, I review work on how the haploid selection resulting from pollen competition has been proposed to influence which alleles are linked to the Y chromosome, and some recent empirical evidence in support of this theory.

Oxidative stress differentially impacts male and female bovine embryos depending on the culture medium and the stress condition.

Male and female embryos are known to differ for their metabolism and response to environmental factors very early in development. The present study aimed to evaluate the response to oxidative stress of male and female bovine embryos at the morula-blastocyst stages in terms of developmental rates, total cell number and apoptotic rates in two culture conditions. Embryos where cultured in a medium supplemented with either 5% fetal calf serum (FCS) or 4 mg/mL bovine serum albumin and a mixture of insulin, transferrin and selenium (BSA-ITS). Oxidative stress was applied at Day-5 post insemination (pi) by adding either AAPH or menadione to the culture medium, and blastocysts were analyzed at Day-7pi. The impact on development and blastocyst quality was dependent on the culture medium and the stress inducer but differed between male and female embryos. Male embryos resisted better to oxidative stress in FCS supplemented medium, no matter the stress inducer. Accordingly, the impact on blastocyst cell number tended to be higher in female blastocysts after stress induction with AAPH in FCS supplemented medium. On the other hand, in BSA-ITS supplemented medium, female embryos were more resistant to AAPH induced stress, while menadione had no impact on sex ratio. The weaker resistance of males to AAPH in this medium is in accordance with their trend to show a higher increase in apoptotic rates than females in this condition. In conclusion, this study shows that oxidative stress has differential impact on male and female bovine blastocysts depending on the culture condition and on the way oxidative stress is induced.

Sex ratio rather than population size affects genetic diversity in Antennaria dioica.

Habitat fragmentation and small population size can lead to genetic erosion in threatened plant populations. Classical theory implies that dioecy can counteract genetic erosion as it decreases the magnitude of inbreeding and genetic drift due to obligate outcrossing. However, in small populations, sex ratios may be strongly male- or female-biased, leading to substantial reductions in effective population size. This may theoretically result in a unimodal relationship between sex ratios and genetic diversity; yet, empirical studies on this relationship are scarce. Using AFLP markers, we studied genetic diversity, structure and differentiation in 14 highly fragmented Antennaria dioica populations from the Central European lowlands. Our analyses focused on the relationship between sex ratio, population size and genetic diversity. Although most populations were small (mean: 35.5 patches), genetic diversity was moderately high. We found evidence for isolation-by-distance, but overall differentiation of the populations was rather weak. Females dominated 11 populations, which overall resulted in a slightly female-biased sex ratio (61.5%). There was no significant relationship between population size and genetic diversity. The proportion of females was not unimodally but positively linearly related to genetic diversity. The high genetic diversity and low genetic differentiation suggest that A. dioica has been widely distributed in the Central European lowlands in the past, while fragmentation occurred only in the last decades. Sex ratio has more immediate consequences on genetic diversity than population size. An increasing proportion of females can increase genetic diversity in dioecious plants, probably due to a higher amount of sexual reproduction.

Polyunsaturated fatty acids influence offspring sex ratio in cows.

Dietary polyunsaturated fatty acids (PUFAs) can influence fertility in farm animals. Some evidence in mice and sheep have suggested that PUFAs may influence offspring sex ratio, which may have significant value for cattle production. To test this hypothesis, three groups of Holstein cows were supplemented with either 0%, 3% or 5% protected fat (PF) in the form of calcium salt of fatty acids (rich in omega-6) from 14-21 days pre-partum until conception. Proven-fertile frozen semen from the same ejaculate was used for insemination. Calf sex recorded at birth was 8/19 (42.1%) male offspring in the control group, increasing to 14/20 (70%, P > 0.05) and 17/20 (85%, P < 0.05) in 3% and 5% PF, respectively. To test if this effect was caused by a direct influence on the oocyte, we supplemented bovine cumulus oocyte complexes during in vitro maturation with either omega-3 alpha-linolenic acid (ALA), omega-6 linoleic acid (LA) or trans-10, cis-12 conjugated linoleic acid (CLA). Sex ratio of the produced transferable embryos was determined using PCR of SRY gene. Similar to the in vivo results, sex ratio was skewed to the male side in the embryos derived from LA- and CLA-treated oocytes (79% and 71%) compared to control and ALA-treated oocytes (44% and 54%, respectively). These results indicate that both dietary and in vitro supplementation of omega-6 PUFAs can skew the sex ratio towards the male side in cattle. Further experiments are required to confirm this effect on a larger scale and to study the mechanisms of action that might be involved.

The female advantage in natural populations of gynodioecious Plantago coronopus: seed quantity vs. offspring quality.

In gynodioecious plant species, females can only persist when they have a reproductive advantage in comparison with hermaphrodites. However, several studies have shown that females do not necessarily produce more seeds than hermaphrodites, since seed production can be affected by population characteristics, such as female frequency or population size. The aim of this study was to quantify the female advantage across a large number of natural populations, examine its relationship with population sex ratio and size, and to assess the role of competition on the magnitude of the female advantage. We sampled 27 populations of Plantago coronopus (nuclear-cytoplasmic gynodioecy) along the Belgian and Dutch coast. In each population, we estimated population sex ratio and size, and assessed seed production per flower and seed production per plant. Subsequently, germination, growth, and competition experiments were performed in the greenhouse to determine the female advantage regarding offspring quality. Females produced fewer seeds per plant than hermaphrodites (FA = 0.90), and seed production was negatively related to female frequency. Since both sex morphs were equally affected by pollen availability, the female advantage was not related to population sex ratio. On the other hand, offspring of females showed higher germination and growth rates, resulting in higher competitive abilities when seeds of a female and a hermaphrodite were grown together. Overall, these results indicate that differences in competitive abilities between the offspring of females and hermaphrodites may have contributed to the maintenance of females in relatively high frequencies in populations of this short-lived gynodioecious plant species.

Brain carnitine deficiency causes nonsyndromic autism with an extreme male bias: A hypothesis.

Could 10-20% of autism be prevented? We hypothesize that nonsyndromic or "essential" autism involves extreme male bias in infants who are genetically normal, but they develop deficiency of carnitine and perhaps other nutrients in the brain causing autism that may be amenable to early reversal and prevention. That brain carnitine deficiency might cause autism is suggested by reports of severe carnitine deficiency in autism and by evidence that TMLHE deficiency - a defect in carnitine biosynthesis - is a risk factor for autism. A gene on the X chromosome (SLC6A14) likely escapes random X-inactivation (a mixed epigenetic and genetic regulation) and could limit carnitine transport across the blood-brain barrier in boys compared to girls. A mixed, common gene variant-environment hypothesis is proposed with diet, minor illnesses, microbiome, and drugs as possible risk modifiers. The hypothesis can be tested using animal models and by a trial of carnitine supplementation in siblings of probands. Perhaps the lack of any Recommended Dietary Allowance for carnitine in infants should be reviewed. Also see the video abstract here: https://youtu.be/BuRH_jSjX5Y.

Concentrate supplementations of grazing pregnant Kalahari Red goats: Effects on pregnancy variables, reproductive performance, birth types and weight of kids.

A study was conducted for 22 weeks to determine the effects of concentrate supplementations at three crude protein (CP) levels, i.e. low protein diet-LPD 12.42% CP (124.93 g day-1); medium protein diet-MPD 14.18% CP (145.87 g day-1) and high protein diet-HPD 16.35% CP (168.19 g day-1) on some pregnancy variables, reproductive performance, birth types and weight of kids kidded by pregnant Kalahari Red goats grazed on Chloris gayana. Thirty-three matured Kalahari Red goats of first parity within age range of 2 to 21/2 years with an average body weight of 38.10 ± 1.13 kg were randomly allotted to the diets with 11 goats per treatment. Data obtained were subjected to analysis of variance in a completely randomised design. It was observed that goats fed HPD had the highest (p < 0.05) values for products of pregnancy (10.35 kg) taken within 24hour before kidding and foetal growth rate (88.37 g day-1) during pregnancy. Goats fed MPD had the lowest values (p < 0.05) of 9.28 kg and 80.07 g day-1 for each of the respective parameters. Afterbirth weight was also the highest and lowest (p < 0.05) for goats supplemented with HPD (3.38 kg) and MPD (3.04 kg), respectively. On the other hand, MPD-supplemented goats had the highest values (p < 0.05) for litter size (2.14) and litter weight (6.80 kg) at birth, the value which was the least for HPD supplementation. From the results obtained from this study, it could be concluded that concentrate diet supplementation with 14.18% CP improved litter size and weight at birth with resultant reduction in weights of pregnancy variables of pregnant Kalahari Red goats grazed on Rhodes grass.

Incidence of multiple sclerosis in the Republic of Ireland: A prospective population-based study.

BACKGROUND: Multiple sclerosis (MS) incidence and prevalence is increasing worldwide, with a disproportionally higher rate in women. Recent studies have questioned the presence of a latitudinal gradient in Europe. Ireland is a high prevalence country for MS with a previously reported North-South gradient making it ideal to further explore this concept. OBJECTIVES: In this study we prospectively determined the incidence rate of newly diagnosed MS in Ireland over a 12-month period and demonstrated the presence of a North-South gradient. METHODS: A national prospective population-based observational study was performed to ascertain all new cases of MS diagnosed from 1st March 2014 - 28th February 2015 in the Ireland. Within the main study there was a smaller nested cohort study to explore clinical outcomes with a view to future prospective follow-up of this cohort. Sources of case ascertainment included neurologists, MS nurse specialists and MS support services. The Irish census 2011 was used to obtain population statistics and the incidence rate was age-standardized to a European Standardised Population (ESP 2011). The North-South gradient was assessed, by comparing incidence rates between northern and southern counties. RESULTS: 292 patients fulfilled the inclusion criteria equating to an age-standardised incidence rate (A-SIR) of 6/100,000 (95% CI: 5.3-6.6); for women the rate was 8.7/100,000 (95% CI: 7.7-9.6) and for men 3.3/100,000 (95% CI: 3.0-3.7). The female to male sex ratio was 2.7:1. Mean age at diagnosis amongst the RRMS group was 37 years (SD: 9.6) and 55 years (SD: 7.7) in the PPMS group; there were no gender differences associated with age of diagnosis. Onset was progressive in 10% of cases. A significant difference was seen in incidence rates between the northern region (A-SIR: 9.6×105, CI: 6.9-12.3) and the southern region (A-SIR: 5.1×105, CI: 3.8-6.3) (Z-score =3.34, p<0.05). Amongst the nested cohort (n=113) mean age at symptom onset in the RRMS group (n=106) was 34 years (SD: 8.7) and 50 years (SD: 11.8) in the PPMS group (n=7). The female to male sex ratio was 3.5:1. Eighty percent had started or were due to start disease modifying therapy at time of review and 77% were taking supplemental vitamin D. Using the hospital depression and anxiety scale (HADS) mild to severe depressive symptoms were reported in 34% with no prior history of depression. Seventy-five percent were in full or part-time employment with 8% not working due to disability arising from their MS. CONCLUSIONS: This is the first study to prospectively assess the incidence rate of MS in Ireland and shows that Ireland has a high incidence rate, comparable with the rest of the British Isles, with a persistent North-South gradient. The age of onset of relapsing remitting multiple sclerosis appears to be increasing over the last 20 years. It will be of interest to re-assess this population over time to see if increasing incidence rates, as well as improved survival, are driving the reported increases in MS prevalence.

Physiological effects of temperature do not explain prevalence of females in populations of gynodioecious Lobelia siphilitica growing in warmer climates.

PREMISE OF THE STUDY: Gynodioecy is a sexual polymorphism whereby female and hermaphroditic plants co-occur within populations. In many gynodioecious species, stressful abiotic environments are associated with higher frequencies of females. This association suggests that abiotic stress affects the relative fitness of females and hermaphrodites and, thus, the maintenance of gynodioecy. METHODS: To test whether abiotic stress affects the fitness of females and hermaphrodites, we grew open-pollinated Lobelia siphilitica families in temperature regimes characteristic of the southern portion of the species' range (where females are common) and the northern portion of the range (where females are rare). We measured physiological and phenological traits that are indicative of heat stress, and fitness components of females and hermaphrodites that could affect the maintenance of gynodioecy. KEY RESULTS: Contrary to expectations if growth at high temperatures is stressful, we found that the hot treatment increased leaf chlorophyll content, decreased the percentage of plants that delayed flowering initiation, and did not affect the quantum efficiency of photosystem II. Growth at high temperatures did not affect the magnitude of the difference in rosette size (a correlate of flower number) between females and hermaphrodites, or the variance in pollen viability among hermaphrodites. CONCLUSIONS: We found that growing-season temperatures typical of high female L. siphilitica populations were not stressful and did not affect either the fitness of females compared to hermaphrodites or variation in fitness among hermaphrodites. Consequently, further research is necessary to explain correlations between abiotic environmental factors and the frequency of females in this and other gynodioecious species.

Effects of Diets Enriched in Omega-9 or Omega-6 Fatty Acids on Reproductive Process.

OBJECTIVE: Maternal type and amounts of dietary fatty acids affect on reproductive process in the mice. The present study investigated the effects of maternal supplementation with different amounts of omega-6 or omega-9 during pregnancy on the number of offspring, sex-ratio and duration of gestation. MATERIALS AND METHODS: Eight-week-old female C57BL/6 mice were randomly assigned into four dietary groups including low omega-6 (16%; LO6), low omega-9 (16%; LO9), high omega-6 (45%; HO6) and high omega-9 (45%; HO9) during gestation. Number of offspring, sex-ratio and duration of pregnancy were compared among four dietary groups. RESULTS: There was significant difference between LO6 and HO6 (p < 0.0001), LO9 and HO9 (p < 0.0001) groups in total number of pups. The number of female and male offspring were significantly different between LO6 and LO9 (p = 0.009 and p = 0.001, respectively), LO9 and HO9 (p = 0.01 and p = 0.025) groups. Duration of pregnancy was significantly higher in low fat diet than high fat diet groups (< 0.001). CONCLUSION: High fat diet reduced number of pups, gestation duration and lead to early labor. Omega-9 fatty acids shifted sex of offspring to females.

Responses of bovine early embryos to S-adenosyl methionine supplementation in culture.

AIM: There is a growing concern about the potential adverse effects of high dose folic acid (FA) supplementation before and during pregnancy. FA metabolism generates S-adenosyl methionine (SAM) which is an important cofactor of epigenetic programming. We sought to assess the impact of a large dose of SAM on early embryo development. MATERIALS & METHODS: In vitro cultured bovine embryos were treated with SAM from the eight-cell stage to the blastocyst stage. In addition to the phenotype, the genome-wide epigenetic and transcription profiles were analyzed. RESULTS: Treatment significantly improved embryo hatching and caused a shift in sex ratio in favor of males. SAM caused genome-wide hypermethylation mainly in exonic regions and in CpG islands. Although differentially expressed genes were associated with response to nutrients and developmental processes, no correspondence was found with the differentially methylated regions, suggesting that cellular responses to SAM treatment during early embryo development may not require DNA methylation-driven changes. CONCLUSION: Since bovine embryos were not indifferent to SAM, effects of large-dose FA supplements on early embryonic development in humans cannot be ruled out.

Male-biased hermaphrodites in a gynodioecious shrub, Daphne jezoensis.

Gynodioecy, a state where female and hermaphrodite plants coexist in populations, has been widely proposed an intermediate stage in the evolutionary pathway from hermaphroditism to dioecy. In the gynodioecy-dioecy pathway, hermaphrodites may gain most of their fitness through male function once females invade populations. To test this prediction, comprehensive studies on sex ratio variation across populations and reproductive characteristics of hermaphrodite and female phenotypes are necessary. This study examined the variation in sex ratio, sex expression, flower and fruit production and sexual dimorphism of morphological traits in a gynodioecious shrub, Daphne jezoensis, over multiple populations and years. Population sex ratio (hermaphrodite:female) was close to 1:1 or slightly hermaphrodite-biased. Sex type of individual plants was largely fixed, but 15% of plants changed their sex during a 6-year census. Hermaphrodite plants produced larger flowers and invested 2.5 times more resources in flower production than female plants, but they exhibited remarkably low fruit set (proportion of flowers setting fruits). Female plants produced six times more fruits than hermaphrodite plants. Low fruiting ability of hermaphrodite plants was retained even when hand-pollination was performed. Fruit production of female plants was restricted by pollen limitation under natural conditions, irrespective of high potential fecundity, and this minimised the difference in resources allocated to reproduction between the sexes. Negative effects of previous flower and fruit production on current reproduction were not apparent in both sexes. This study suggests that gynodioecy in this species is functionally close to a dioecious mating system: smaller flower production with larger fruiting ability in female plants, and larger flower production with little fruiting ability in hermaphrodite plants.

Impaired imprinted X chromosome inactivation is responsible for the skewed sex ratio following in vitro fertilization.

Dynamic epigenetic reprogramming occurs during normal embryonic development at the preimplantation stage. Erroneous epigenetic modifications due to environmental perturbations such as manipulation and culture of embryos during in vitro fertilization (IVF) are linked to various short- or long-term consequences. Among these, the skewed sex ratio, an indicator of reproductive hazards, was reported in bovine and porcine embryos and even human IVF newborns. However, since the first case of sex skewing reported in 1991, the underlying mechanisms remain unclear. We reported herein that sex ratio is skewed in mouse IVF offspring, and this was a result of female-biased peri-implantation developmental defects that were originated from impaired imprinted X chromosome inactivation (iXCI) through reduced ring finger protein 12 (Rnf12)/X-inactive specific transcript (Xist) expression. Compensation of impaired iXCI by overexpression of Rnf12 to up-regulate Xist significantly rescued female-biased developmental defects and corrected sex ratio in IVF offspring. Moreover, supplementation of an epigenetic modulator retinoic acid in embryo culture medium up-regulated Rnf12/Xist expression, improved iXCI, and successfully redeemed the skewed sex ratio to nearly 50% in mouse IVF offspring. Thus, our data show that iXCI is one of the major epigenetic barriers for the developmental competence of female embryos during preimplantation stage, and targeting erroneous epigenetic modifications may provide a potential approach for preventing IVF-associated complications.