Genome-wide scanning to identify and validate single nucleotide polymorphism markers associated with drought tolerance in spring wheat seedlings.

Unlike other growth stages of wheat, very few studies on drought tolerance have been done at the seedling stage, and this is due to the complexity and sensitivity of this stage to drought stress resulting from climate change. As a result, the drought tolerance of wheat seedlings is poorly understood and very few genes associated with drought tolerance at this stage were identified. To address this challenge, a set of 172 spring wheat genotypes representing 20 different countries was evaluated under drought stress at the seedling stage. Drought stress was applied on all tested genotypes by water withholding for 13 days. Two types of traits, namely morphological and physiological traits were scored on the leaves of all tested genotypes. Genome-wide association study (GWAS) is one of the effective genetic analysis methods that was used to identify target single nucleotide polymorphism (SNP) markers and candidate genes for later use in marker-assisted selection. The tested plant materials were genotyped using 25k Infinium iSelect array (25K) (herein after it will be identified as 25K) (for 172 genotypes) and genotyping-by-sequencing (GBS) (for 103 genotypes), respectively. The results of genotyping revealed 21,093 25K and 11,362 GBS-SNPs, which were used to perform GWAS analysis for all scored traits. The results of GWAS revealed that 131 and 55 significant SNPs were controlling morphological and physiological traits, respectively. Moreover, a total of eight and seven SNP markers were found to be associated with more than one morphological and physiological trait under drought stress, respectively. Remarkably, 10 significant SNPs found in this study were previously reported for their association with drought tolerance in wheat. Out of the 10 validated SNP markers, four SNPs were associated with drought at the seedling stage, while the remaining six SNPs were associated with drought stress at the reproductive stage. Moreover, the results of gene enrichment revealed 18 and six pathways as highly significant biological and molecular pathways, respectively. The selection based on drought-tolerant alleles revealed 15 genotypes with the highest number of different drought-tolerant alleles. These genotypes can be used as candidate parents in future breeding programs to produce highly drought-tolerant genotypes with high genetic diversity. Our findings in this study provide novel markers and useful information on the genetic basis of drought tolerance at early growth stages.

Regulation of uterus and placenta remodeling under high estradiol levels in gestational diabetes mellitus models†.

The present study aimed to investigate the regulation of placentas and uterus remodeling and involvement of estradiol in gestational diabetes mellitus. To achieve this, we established in vitro and in vivo models for gestational diabetes mellitus placentas by culturing human placental choriocarcinoma cells (BeWo) under hyperglycemic concentration and treating pregnant rats with streptozotocin. We evaluated the expression of angiogenesis-related proteins. The expression of the anti-angiogenic factor, excess placental soluble fms-like tyrosine kinase 1 was increased in our in vitro gestational diabetes mellitus model compared with the control. Moreover, the expressions of placental soluble fms-like tyrosine kinase 1 and the von Willebrand factor were also significantly elevated in the placenta of streptozotocin-treated rats. These data indicate the disruption of angiogenesis in the gestational diabetes mellitus placentas. The expression levels of connexin 43, a component of the gap junction and collagen type I alpha 2 chain, a component of the extracellular matrix, were decreased in the gestational diabetes mellitus uterus. These results suggest that uterus decidualization and placental angiogenesis are inhibited in gestational diabetes mellitus rats. Our results also showed upregulation of the expression of genes regulating estradiol synthesis as well as estrogen receptors in vivo models. Accordingly, the concentration of estradiol measured in the culture medium under hyperglycemic conditions, as well as in the serum and placenta of the streptozotocin-treated rats, was significantly elevated compared with the control groups. These results suggest that the dysregulated remodeling of the placenta and uterus may result in the elevation of estradiol and its signaling pathway in the gestational diabetes mellitus animal model to maintain pregnancy.

Screening Spring Wheat Genotypes for TaDreb-B1 and Fehw3 Genes under Severe Drought Stress at the Germination Stage Using KASP Technology.

Drought stress is a major yield-limiting factor throughout the world in wheat (Triticum aestivum L.), causing losses of up to 80% of the total yield. The identification of factors affecting drought stress tolerance in the seedling stage is especially important to increase adaptation and accelerate the grain yield potential. In the current study, 41 spring wheat genotypes were tested for their tolerance to drought at the germination stage under two different polyethylene glycol concentrations (PEG) of 25% and 30%. For this purpose, twenty seedlings from each genotype were evaluated in triplicate with a randomized complete block design (RCBD) in a controlled growth chamber. The following nine parameters were recorded: germination pace (GP), germination percentage (G%), number of roots (NR), shoot length (SL), root length (RL), shoot-root length ratio (SRR), fresh biomass weight (FBW), dry biomass weight (DBW), and water content (WC). An analysis of variance (ANOVA) revealed highly significant differences (p < 0.01) among the genotypes, treatments (PEG25%, PEG30%) and genotypes × treatment interaction, for all traits. The broad-sense heritability (H2) estimates were very high in both concentrations. They ranged from 89.4 to 98.9% under PEG25% and from 70.8 to 98.7% under PEG30%. Citr15314 (Afghanistan) was among the best performing genotypes under both concentrations for most of the germination traits. Two KASP markers for TaDreb-B1 and Fehw3 genes were used to screen all genotypes and to study the effect of these on drought tolerance at the germination stage. All genotypes with Fehw3 (only) showed a better performance for most traits under both concentrations compared to other genotypes having TaDreb-B1 or having both genes. To our knowledge, this work is the first report showing the effect of the two genes on germination traits under severe drought stress conditions.

Genetic and morpho-physiological analyses of the tolerance and recovery mechanisms in seedling stage spring wheat under drought stress.

Drought is one of the complex abiotic stresses that affect the growth and production of wheat in arid and semiarid countries. In this study, a set of 172 diverse spring wheat genotypes from 20 different countries were assessed under drought stress at the seedling stage. Besides seedling length, two types of traits were recorded, namely: tolerance traits (days to wilting, leaf wilting, and the sum of leaf wilting), and recovery traits (days to regrowth, regrowth biomass, and drought survival rate). In addition, tolerance index, recovery index, and drought tolerance index (DTI) were estimated to select the most drought tolerant genotypes. Moreover, leaf protein content (P), amino acid (AM), proline content (PRO), glucose (G), fructose (F), and total soluble carbohydrates (TSC) were measured under control and drought conditions to study the changes in each physiological trait due to drought stress. All genotypes showed a high significant genetic variation in all the physio-morphological traits scored under drought stress. High phenotypic and genotypic correlations were found among all seedling morphological traits. Among the studied indices, the drought tolerance index (DTI) had the highest phenotypic and genotypic correlations with all tolerance and recovery traits. The broad-sense heritability (H2) estimates were high for morphological traits (83.85-92.27), while the physiological traits ranged from 96.41 to 98.68 under the control conditions and from 97.13 to 99.99 under drought stress. The averages of the physiological traits (proteins, amino acids, proline, glucose, fructose, and total soluble carbohydrates) denoted under drought stress were higher than those recorded under well-watered conditions except for proteins. In this regard, amino acids, glucose, and total soluble carbohydrates had a significant correlation with all morphological traits. The selection for drought tolerance revealed 10 tolerant genotypes from different countries (8 genotypes from Egypt, one from Morocco, and one from the United States). These selected genotypes were screened for the presence of nine specific TaDREB1 alleles. Six primers were polymorphic among the selected genotypes. Genetic diversity among the selected genotypes was investigated using 21,450 SNP markers. The results of the study shed light on the different mechanisms for drought tolerance that wheat plants use to tolerate and survive under drought stress. The genetic analysis performed in this study suggested the most suitable genotypes for selective breeding at the seedling stage under water deficit.

Differential Expressions of Aquaporin Subtypes in the Adult Mouse Testis.

Many efforts have been made to study the expression of aquaporins (AQP) in the mammalian reproductive system, but there are not enough data available regarding their localized expression to fully understand their specific roles in male reproduction. The present study investigated the expression and localization patterns of different AQP subtypes in the adult mouse testes and testicular spermatozoa using an immunofluorescence assay. All the studied AQPs were expressed in the testes and revealed subtype-specific patterns in the intensity and localization depending on the cell types of the testes. AQP7 was the most abundant and intensive AQP subtype in the seminiferous tubules, expressing in Leydig cells and Sertoli cells as well as all stages of germ cells, especially the spermatids and testicular spermatozoa. The expression pattern of AQP3 was similar to that of AQP7, but with higher expression in the basal and lower adluminal compartments rather than the upper adluminalcompartment. AQP8 expression was limited to the spermatogonia and Leydig cells whereas AQP9 expression was exclusive to tails of the testicular spermatozoa and elongated spermatids. Taken together, the abundance and distribution of the AQPs across the different cell types in the testes indicating to their relavance in spermatogenesis, as well as in sperm maturation, transition, and function.

Inhibitory effects and underlying mechanisms of Artemisia capillaris essential oil on melanogenesis in the B16F10 cell line.

The present study investigated the anti­melanogenic activity of 10 essential oils using the B16F10 cell model. Initially, a wide range of concentrations of these essential oils were screened in order to determine their toxicity levels. The assigned non­toxic concentrations of the tested essential oils were then used to evaluate their effects on melanogenesis. The effects of the essential oils with potent anti­melanogenic activity on cell proliferation, protection against H2O2­induced cell death and the expression of certain melanogenesis­related genes, including MITF, tyrosinase, tyrosinase related protein (TRP)­1 and TRP­2 were also evaluated. The results revealed that the essential oils extracted from Citrus unshiu, Juniperus chinensis L., Zanthoxylum piperitum and Artemisia capillaris (A. capillaris) inhibited melanogenesis. However, among these four extracts, only A. capillaris extract enhanced cell proliferation, exhibited anti­H2O2 activities and decreased the expression level of TRP­1. It was demonstrated that A. capillaris extract inhibited melanin synthesis via the downregulation of the TRP­1 translational level. These essential oil extracts, particularly that of A. capillaris, may thus be used as natural anti­melanogenic agents for therapeutic purposes and in the cosmetic industry for skin whitening effects with beneficial proliferative properties. However, further studies using in vivo models are required to validate these findings and to examine the effects of these extracts on various molecular pathways.

Effect of endocrine disruptors on the ratio of X and Y chromosome-bearing live spermatozoa.

Although equal numbers of X and Y spermatozoa are produced during spermatogenesis, the sex chromosome ratio in ejaculated spermatozoa can be altered by exposure to endocrine-disrupting chemicals (EDCs), which can be reflected by altered sex ratios at birth. Here, we hypothesized EDCs affect sperm functions and viability of X and Y chromosome-bearing human spermatozoa. After exposure to genistein (Gen), bisphenol A (BPA), 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), dibromochloropropane (DBCP), and diazinon (Diaz), we evaluated motility, viability, capacitation, and differential viability of X and Y spermatozoa. All EDCs tested altered sperm viability, motility, and capacitation. Interestingly, the Y/X ratio of live spermatozoa was significantly lower in sperm treated with TCDD, DBCP, and Diaz than control spermatozoa. Our results suggest that some of EDCs have larger effects on the viability of Y spermatozoa than X spermatozoa, implicating that a reduction in Y sperm viability may result in a female-biased sex ratio of offspring at birth.

2,3,7,8-Tetrachlorodibenzo-p-dioxin can alter the sex ratio of embryos with decreased viability of Y spermatozoa in mice.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a reproductive and developmental toxicant that can alter the sex ratio of offspring (proportion of male offspring). We hypothesized that the alteration of sex ratio is associated with sex chromosome ratio of live spermatozoa affected by exposure to TCDD. After exposure to TCDD we analyzed simultaneously sperm sex chromosome constitution and viability, and evaluated sperm sex chromosome ratio association with embryo sex ratio in mice. Short-term exposure to TCDD affects the decreased sperm motility and viability, and the increased acrosome reaction. Interestingly, Y spermatozoa survived shorter than X spermatozoa at high concentrations of TCDD. Moreover, the decreased sex ratio of embryos was associated with the short lifespan of Y spermatozoa. Our results suggest that TCDD may affect the fertility of Y spermatozoa more than X spermatozoa. Further studies are needed to compare the difference of fertilizing capability between X and Y spermatozoa by the effect of TCDD.

Maternal and Fetal Outcomes in Pregnant Women with a Prosthetic Mechanical Heart Valve.

BACKGROUND: Pregnancy is associated with several cardiocirculatory changes that can significantly impact underlying cardiac disease. These changes include an increase in cardiac output, sodium, and water retention leading to blood volume expansion, and reductions in systemic vascular resistance and systemic blood pressure. In addition, pregnancy results in a hypercoagulable state that increases the risk of thromboembolic complications. OBJECTIVES: The aim of this study is to assess the maternal and fetal outcomes of pregnant women with mechanical prosthetic heart valves (PHVs). METHODS: This is a prospective observational study that included 100 pregnant patients with cardiac mechanical valve prostheses on anticoagulant therapy. The main maternal outcomes included thromboembolic or hemorrhagic complications, prosthetic valve thrombosis, and acute decompensated heart failure. Fetal outcomes included miscarriage, fetal death, live birth, small-for-gestational age, and warfarin embryopathy. The relationship between the following were observed: - Maternal and fetal complications and the site of the replaced valve (mitral, aortic, or double)- Maternal and fetal complications and warfarin dosage (≤5 mg, >5 mg)- Maternal and fetal complications and the type of anticoagulation administered during the first trimester. RESULTS: This study included 60 patients (60%) with mitral valve replacement (MVR), 22 patients (22%) with aortic valve replacement (AVR), and 18 patients (18%) with double valve replacement (DVR). A total of 65 patients (65%) received >5 mg of oral anticoagulant (warfarin), 33 patients (33%) received ≤5 mg of warfarin, and 2 patients (2%) received low-molecular-weight heparin (LMWH; enoxaparin sodium) throughout the pregnancy. A total of 17 patients (17%) received oral anticoagulant (warfarin) during the first trimester: 9 patients received a daily warfarin dose of >5 mg while the remaining 8 patients received a daily dose of ≤5 mg. Twenty-eight patients (28%) received subcutaneous (SC) heparin calcium and 53 patients (53%) received SC LMWH (enoxaparin sodium). Prosthetic valve thrombosis occurred more frequently in patients with MVR (P = 0.008). Postpartum hemorrhage was more common in patients with aortic valve prostheses than in patients with mitral valve prostheses (P = 0.005). The incidence of perinatal death was higher in patients with AVR (P = 0.014). The incidence of live birth was higher in patients with DVR (P = 0.012). The incidence of postpartum hemorrhage was higher in patients who received a daily dose of >5 mg of warfarin than in patients who received ≤5 mg of warfarin (P = 0.05). The incidence of spontaneous abortion was also higher in patients receiving >5 mg of warfarin (P ≤ 0.001), while the incidence of live births was higher in patients receiving ≤5 mg of warfarin (P = 0.008). There was a statistically significant difference between the anticoagulant received during the first trimester and cardiac outcomes. Specifically, patients on heparin developed more heart failure (P = 0.008), arrhythmias (P = 0.008), and endocarditis (P = 0.016). There was a statistically significant relationship between heparin shifts during the first trimester and spontaneous abortion (P = 0.003). CONCLUSION: Warfarin use during the first trimester is safer for the mother but is associated with more fetal loss, especially in doses that exceed 5 mg. The incidence of maternal complications is greater in women who receive LMWH or unfractionated heparin during the first trimester, especially prosthetic valve thrombosis, although the fetal outcome is better because heparin does not cross the placenta.

Significant association of TNFα and IL-6 gene with male infertility--an explorative study in Indian populations of Uttar Pradesh.

In this study were aimed to identify the association of SNPs candidate genes of TNF-α and IL-6 with hormones levels and sperm cells death in infertile subjects of Uttar Pradesh population in North India. The study population comprised, fertile donor (control group) and infertile group patients i.e. normozoospermic (idiopathic unexplained), oligozoospermic and asthenozoospermic groups, with 260 subjects in each group. Subjects were selected from the Departments of Urology, K.G's Medical University and Urology, SGPGIMS, Lucknow, India. The allele-specific polymerase chain reaction (PCR) and PCR-RFLP were used to investigate the substitution of the guanine (G)-to-adenosine (A) at position-308 and guanine (G)-to-cytosine (C) at position-174 in the promoter regions of the TNF-α and IL-6 genes, respectively. Further their relation to male fertility and sperm function were also investigated. It was found that the substitution levels from G to A and from G to C in the TNF-α and IL-6 genes, respectively, were significantly higher in the infertile subjects as compared to that of control group. The apoptosis and necrosis levels were also higher in oligozoospermic and asthenozoospermic infertile subjects. Further it was found to be associated with increased level of reactive oxygen species as observed in oligozoospermic and asthenozoospermic subjects. However, a significant decrease in testosterone and luteinizing hormone with increased prolactin and follicle stimulating hormones was observed in infertile subjects. The study populations indicating a strong association between TNF-α G-308A and IL-6 G-174C substitution with infertile men which is further supported by allele and genotype meta-analysis and thus established it as a risk factor.