Journal articles on the topic 'Fertility of gems'

Soybean is including a major protein source with a relatively higher consumption rate. Similarly, various efforts have been adopted towards boosting production, including selecting high-yield species and soil fertility improvement using fertilizers. This research aims to increase soybean production by employing the appropriate variety and phosphate fertilizers to improve soil fertility. This experiment was conducted in Takalar, South Sulawesi, between April-July 2019. The split-plot design method was applied with three replications. Consequently, the main plot encompasses the treatment of three varieties, termed Argomulyo, Anjasmoro and Gema, while the subplots relate to phosphate fertilization, comprising three levels, including fertilization with a dose of SP-36 at 50, 100 and 150 kg.ha<sup>-1</sup>. The results showed that the modifications in variety and phosphorus fertilization significantly influenced soybean production, as Argomulyo obtained the maximum rate of 2.77 t.ha<sup>-1</sup>, followed by Anjasmoro at 2.45 t.ha<sup>-1</sup>. Furthermore, phosphate fertilization with a dose of SP-36 at 100-150 kg.ha<sup>-1</sup> demonstrated the optimal soybean production as indicated by the maximum productive nodes and pods. Therefore, no interaction was observed between the various species and phosphate fertilization.

ABSTRACT The mammalian Grf1 and Grf2 proteins are Ras guanine nucleotide exchange factors (GEFs) sharing a high degree of structural homology, as well as an elevated expression level in central nervous system tissues. Such similarities raise questions concerning the specificity and/or redundancy at the functional level between the two Grf proteins. grf1-null mutant mice have been recently described which showed phenotypic growth reduction and long-term memory loss. To gain insight into the in vivo function of Grf2, we disrupted its catalytic CDC25-H domain by means of gene targeting. Breeding among grf2 +/− animals gave rise to viable grf2 −/− adult animals with a normal Mendelian pattern, suggesting that Grf2 is not essential for embryonic and adult mouse development. In contrast to Grf1-null mice, analysis of grf2 −/− litters showed similar size and weight as their heterozygous or wild-type grf2 counterparts. Furthermore, adult grf2 −/− animals reached sexual maturity at the same age as their wild-type littermates and showed similar fertility levels. No specific pathology was observed in adult Grf2-null animals, and histopathological studies showed no observable differences between null mutant and wild-type Grf2 mice. These results indicate that grf2 is dispensable for mouse growth, development, and fertility. Furthermore, analysis of double grf1/grf2 null animals did not show any observable phenotypic difference with single grf1 −/− animals, further indicating a lack of functional overlapping between the two otherwise highly homologous Grf1 and Grf2 proteins.

Although there is general recognition that the mammalian epididymis is unique among the vertebrates and is essential for male fertility, there is relatively little understanding of its role in the process of reproduction. It has been suggested that it plays an important role in the competition between males to achieve paternity (sperm competition) by accumulating and storing sperm.1 We now provide evidence that specific protein synthesized and secreted by the epididymis of the echidna plays a major role in sperm competition by binding spermatozoa into bundles of about a 100 sperm and consequently greatly enhancing their rate of motility, particularly in viscous medium. The epididymis of the echidna is structurally differentiated into a large caput epididymidis that is structurally similar to the ‘initial segment’ of other mammals, and a small cauda epididymidis. Using radiolabelling and 2-D electrophoresis we have shown that the caput epididymidis synthesises and secretes a similar pattern of proteins along most of its length where sperm are not associated with one another. The sperm form into bundles as they enter the cauda epididymidis where the pattern of protein secretion changes considerably, being characterised by a new band of about 80 kDa in reduced gels. Electron microscopy of the bundles shows electron dense material binding the sperm together in the bundles. The bundles persist for more than an hour after sperm are released into physiological media. (1)Jones, RC (1998). Evolution of the vertebrate epididymis. J. Reprod. Fertil. Suppl. 53, 163–182.

There are numerous benefit claims for the supplementation of l-arginine in animal production systems. Its positive effect on fertility has been widely characterised in females of various species; however, the effects on the male reproductive system remain debatable with still unfolding molecular mechanisms. Here we investigated the effects of a dietary l-arginine supplementation on boar semen production outcomes, sperm characteristics, and seminal plasma proteome. Nine mature (20 months of age) Nebraska Index Line boars were individually housed and randomly assigned to a soybean meal-based diet containing 0.77% standard l-arginine (control, n=4) or 1.77% high l-arginine (n=5). Boar semen was collected weekly during the 6-week dietary arginine trial. Semen production outcomes (volume and total spermatozoa) and sperm motion (total and progressive motility) and morphology characteristics were subsequently analysed using a computer-assisted sperm analyzer (CEROS II). On Week 6, the seminal plasma of each boar was obtained by centrifugation at 4°C for proteome analysis. The clarified seminal plasma was precipitated; the purified proteins were resuspended in an appropriate buffer and loaded (300μg) onto an immobilized pH gradient strip (isoelectric point 3-10) for the first-dimension electrophoresis. The second-dimension was subsequently performed (4-20% gradient gel), and all gels were stained with Coomassie R250 dye. The PDQuest software (Bio-Rad, Hercules, CA, USA) was used to detect all significantly differentially expressed protein spots (Student’s t-test with P<0.05). Two-way ANOVA repeated measurements (SPSS Statistics, Chicago, IL, USA) was used to analyse dependent (semen outputs and sperm characteristics) and independent (week) variables. A difference was significant at P<0.05. During the l-arginine-consumption trial, semen volumes and total spermatozoa were not significantly affected (P>0.05). Similarly, various sperm motility (total and progressive motility) and velocity characteristics were not affected (P>0.05). By the end of the feed trial, the average (mean±standard error of the mean) semen volume (389±40mL), the total spermatozoa per ejaculate (39±8×109), the total (82±2%) and progressive (62±3%) sperm motility, the sperm velocity (e.g. average path: 98±7μm s−1), and sperm morphology (e.g. distal droplet: 5±1%) parameters of the control group were not significantly different from the group supplemented with high l-arginine: 393±35mL, 47±4×109, 82±2%, 61±3%, 92±8μm s−1, and 5±0%, respectively (P>0.05). In contrast, the proteome profiles of seminal plasma harvested on the sixth week of diet revealed various changes, characterised by the significantly increased expression of 8 protein spots in seminal plasma samples derived from arginine-fed boars (P<0.05). These results indicate that dietary supplementation of l-arginine does not affect the semen outputs, neither the sperm motility characteristics nor their morphology. However, the proteome profile of the seminal plasma was changed by the presence of l-arginine. The findings may have significant implications for boar fertility, and the identification of these proteins is ongoing. This work was supported by USDA-Agricultural Research Service Biophotonics Initiative #58-6402-3-018.

The aims of this study were to: (i) evaluate the relationships between vegetation indices (VIs) derived from Sentinel-2 imagery and grain yield (GY) and the number of spikes per square meter (SN) of winter wheat and triticale; (ii) determine the dates and plant growth stages when the above relationships were the strongest at individual field scale, thus allowing for accurate yield prediction. Observations of GY and SN were performed at harvest on six fields (three locations in two seasons: 2017 and 2018) in three regions of Poland, i.e., northeastern (A—Brożówka), central (B—Zdziechów) and southeastern Poland (C—Kryłów). Vegetation indices (Normalized Difference Vegetation Index (NDVI), Soil-Adjusted Vegetation Index (SAVI), modified SAVI (mSAVI), modified SAVI 2 (mSAVI2), Infrared Percentage Vegetation Index (IPVI), Global Environmental Monitoring Index (GEMI), and Ratio Vegetation Index (RVI)) calculated for sampling points from mid-March until mid-July, covering within-field soil and topographical variability, were included in the analysis. Depending on the location, the highest correlation coefficients (of about 0.6–0.9) for most of VIs with GY and SN were obtained about 4–6 weeks before harvest (from the beginning of shooting to milk maturity). Therefore, satellite-derived VIs are useful for the prediction of within-field cereal GY as well as SN variability. Information on GY, predicted together with the results for soil nutrient availability, is the basis for the formulation of variable fertilize rates in precision agriculture. All examined VIs were similarly correlated with GY and SN via the commonly used NDVI. The increase in NDVI by 0.1 unit was related to an average increase in GY by about 2 t ha−1.

Seminal plasma is a complex of secretions of the male accessory reproductive organs and appears to exert important effects on sperm function (Shivaji et al. 1990 Proteins of Seminal Plasma, Wiley, New York, NY, USA). The protein quality of the seminal plasma may affect positively the bulls’fertility (Killiam et al. 1993 Biol. Reprod. 49, 1202-1207). Peptides of 55 and 66 kDa were present in bulls with excellent spermatic conditions for example motility and vigor. On the other hand, 16- and 36-kDa peptides were observed with unfavorable spermatic conditions (Chacur et al. 2009 Anim. Reprod. 6, 339). The objective was to determine the influence of season on seminal plasma proteins in Brown Swiss bulls. Semen from 33 Brown Swiss bulls 24 months of age was collected by electroejaculation during winter (from June to August) and summer (from December to February) in the southern hemisphere in 2008. Semen samples were collected with 14-day intervals totalizing 196 ejaculates. Samples of semen were centrifuged (1500g/15 min) and the seminal plasma was conditioned in cryotubes and stored at -20°C until further processing. Proteins were extracted from 200 μL of each sample in 2 mL of extraction buffer composed of 0.625 M Tris-HCl, at pH 6.8, in 2% SDS, 5% fi-mercaptoethanol, and 20% of glycerol. Percentages of different plasma proteins by season were statistically compared by the chi-square test with significance level at P < 0.05). Proteins were quantified according to Bradford (1976 Anal. Biochem. 72, 248-254) and electrophoresis was performed according to Laemmili (1970 Nature 227, 680-685). Gels were fixed with isopropanol: acetic acid: water (4:1:5 v/v) for 30 minutes and stained in the same solution with 2% of Coomassie Blue R250. In 26 bulls, the absence of high molecular weight (HMW; 55 kDa, 66 kDa, and 80 kDa) proteins was found in the summer. There was a significant increase (P < 0.05) in total spermatic defects, acrosome defects, and distal cytoplasmatic droplets in these bulls. The 40-kDa protein that reflected low fertility was observed in 10 bulls in the summer with semen quality decreases. The 11 bulls showed presence of HMW (55 kDa) in the winter. In 11 bulls, HMW (55 kDa, 66 kDa, or 80 kDa) proteins were present with a satisfactory semen condition according to Killiam et al. (1993 Biol. Reprod. 49, 1202-1207). In conclusion, the seasons of the year may influence the presence of proteins in seminal plasma. There was a direct relationship of the season with seminal plasma proteins. The presence of the proteins of 20 kDa, 55 kDa, 66 kDa, and 80 kDa suggested an increase of the semen quality during the winter. UNOESTE and Bartira Farm.

Green gram (Vigna radiata) is considered the chief legume in Pakistan. Thus, current study was conducted to examine the ameliorating effect of phytohormones pre-treatments under salt stress on proteome profile of green gram by sodium-dodecyl-sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The soluble green gram seedlings proteins were resolved on 4% stacking and 12% resolving gels. The SDS-PAGE resolved 24 polypeptide bands ranging from 200 to 17kDa. Among these, 12 out of 24 bands of proteins were essentials house-keeping or growth proteins of green grams. While, 120, 114.6, 51.8, 29.1, and 22.8 kDa bands were over-expressed under 50 to 350mM salt with phytohormones treatments. The others 104.5 kDa, 99.8 kDa, 95.3 kDa, 91.0 kDa, 55 kDa, 46 kDa, and 17kDa bands were related to the GAᴣ, IAA, and SA induced tolerance. Overall 120 kDa, 114.6 kDa, 104.5 kDa, 99.8, 95.3 kDa, 51.8 kDa, 29.1 kDa and 22.8kDa bands were first time identified in the current study. The information retrieved from NCBI protein database, the resolved peptides were principally belonging to 7S and 8S vicilin, 2S, 8S, 11S, and 16.5S globulins. It is determined that seed priming with SA enhanced tolerance in green gram by rapidly synthesizing stress alleviating peptides.Key word: Cluster analysis, dendrogram, mungbean, salt stress, SDS-PAGEINTRODUCTIONVarious world-wide health concerning organization recommended the use of high graded plant protein such as legumes to prevent the risk of metabolic disorder (Hou et al., 2019). Legumes are most important protein crop on the earth. Among the legumes, the green gram is the major pulses. Its seeds are rich in superior quality storage protein, which account 85% of the total protein while, another 15% have not been broadly studied (Yi-Shen et al., 2018). The soluble storage protein comprises of 60% globulins, 25% albumin and 15% prolamins. Globulins are further divided into 3.4% basic-type (7S), 7.6% legumin-type (11S), and 89% vicilin-type (8S) (Mendoza et al., 2001; Itoh et al., 2006). Other than proteins, the green gram seeds also contain starch, fiber, phenolic compound, saponins, vitamins, calcium zinc, potassium, folate, magnesium, manganese and very low in fat that made it meager man’s meat (Hou et al., 2019). It is also a good source of green manure and fodder (El-Kafafi et al., 2015). Its root has ability to fix 30 to 50 Kg/ha atmospheric nitrogen in the soil which is essential for maintaining soil fertility (Chadha, 2010). The green gram is the valuable and the major Rabi pulse crop of Pakistan. Its cultivation area in 2016-2017 was about 179,000 hectares with seed yield of 130,000 tones. In comparison during 2017-2018, it was cultivated on 161,800 hectares land with 118,800 tones seed yield (GOP, 2018). One of the reasons of this 9% decrease in both land and productivity is the shortage of irrigated land due to soil salinity. The salinity induce oxidative bust in the mungbean cells, caused by responsive oxygen species (ROS) such as hydrogen peroxide, singlet oxygen, hydroxyl radical and superoxide radical. The ROS create hindrance in various metabolic processes of plant via interacting with macromolecules like proteins (Alharby et al., 2016). However, phytohormones like gibberellic acid (GAᴣ), indole acetic acid (IAA), and salicylic acid (SA) take part in the biosynthesis of salt tolerance proteins under salinity. These salt tolerance proteins acclimate plants under salinity stress. Application of biotechnology plays a significant role in agriculture (Khan et al., 2017). Therefore, production of particular proteins under salt stress is a specific response of cell which can be analyzed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). SDS-PAGE is the simple, valid, and cost-effective biochemical marker (Mushtaq et al., 2018). This marker has been widely used to determine the extent of evolutionary variations in crops (El-Kafafi et al., 2015).OBJECTIVES The present study was directed first time with the aim to investigate the toxic effect of sodium chloride (0-350 mM) and stress acclimation by pre-treatment of GAᴣ, IAA, and SA on the proteome profile of NM-92 cultivar of a Pakistani green gram.MATERIALS AND METHODSThe present study was replicated thrice in the plant laboratory of Department of Genetics, Faculty of Science, and University of Karachi. The seeds of mung bean cultivar NM-92 were acquired from National Agricultural Research Centre (NARC), Islamabad. These freshly collected 15 seedsˉ1 treatment / replication were divided into two sets. The first was named as sodium chloride (SC) stress treatments were imbibed in sterile distilled water (DW) whereas, second set soaked in gibberellic acid (GAᴣ) (BDH Chemicals, England), indole acetic acid (IAA) (Fluka, Switzerland), and salicylic acid (SA) (J.T. Baker, Holland) in the separate beaker for 24 hours under dark condition. After 24 hours, given ample time to both the sets at room temperature. After recovery, all 20 treatments were sown in the 150 X 30 mm sized petri-dishes containing 0, 50, 150, 250, and 350 millimolar (mM) sodium chloride solution (Fisher Scientific, UK) for 72 hours.Protein extraction: Protein extraction was done by taking 0.3g of seedlings in an ice chilled mortar and crushed by adding 600µL 0.2 M Tris-HCl buffer having pH 7.5 contained 5% SDS (w/v) and 5% 2-mercaptoethanol (v/v). The homogenate was incubated at 0oC for 30 min., boiled in the water bath for 3 min. at 100oC. Samples were centrifuged in Heraeus Biofuge D-37520, Germany for 30 min. at 8000 rpm. The protein supernatant was saved at below 0°C for quantitative and qualitative determination with minor modifications. The total soluble protein content of the samples was estimated via “Bovine Serum Albumin (BSA) standard curve” and explicit in µg protein milligramˉ1 fresh weight of mung seedlings.Bovine serum albumin standard curve (2000 μg/mL): Total protein standard curve was made by dissolving 0.05g of Bovine Serum Albumin (BSA) in 25mL of distilled water. Ten serial dilutions were made from 0.1 mL to 1mL by BSA solution then performed Lowry. A standard curve of total proteins was plotted by taking BSA absorbance at Y-axis and 2000 μg BSA / mL at X-axisSample preparation for SDS-PAGE: For qualitative assessment of total proteins; the 35μL of saved protein supernatant was combined with 15μL of sample diluting buffer (SDB). The SDB was made up of 0.0625 M Tris-HCl pH 6.8 with 2% of SDS, 10% of glycerol, 0.003% of bromophenol blue dye and 5% of 2-mercaptoethanol. Boil the 50μL protein SDB supernatant at 100oC in water bath for 3 min., centrifuged at 6000 rpm for 4 min. The supernatant was loaded on SDS-PAGE gel with the given formulae. The SDS- PAGE: Total proteins were fractionated via SDS-PAGE with 4% stacking and 12% resolving gel. The resolving gel of 12% was made by taking 6mL solution A, 1.8 mL 3 M Tris 1 M HCl buffer pH 8.8, 144μL 10% SDS, 5.74 mL sterile distilled water, 720μL 1.5% ammonium persulphate (APS) in deionized water and 10μL TEMED. While, stacking was composed of 1.25mL of solution A, 2.5mL of 0.5M Tris 1M HCl buffer pH 6.8, 100μL 10% SDS, 1.8 mL of distilled water, 500μL 1.5% APS and 12μL TEMED. Solution A was prepared by conjoining 30% acrylamide and 0.8% N, N’-methylene-bisacrylamide in deionized water. To avoid polymerization in the beaker; the prepared solution was quickly poured into the 3 mm thick gel plates after adding TEMED. The stacking was lined over resolving gel, then combs were inserted between the gel plates of SCIE-PLAS TV-100 separation system, UK, and allowed to polymerize for ½ an hour. After polymerization gel was placed in the tank which were filled with Tris-Glycine buffer (electrode buffer) pH 8.4 then combs were removed. The electrode buffer contained 0.3% Tris, 1.41% Glycine and 0.1% SDS in 2000mL d/w. The gel was pre-run for 15 min. at 60 volts and 120 mA currents. The prepared SDS-PAGE samples were loaded in wells with BlueStepTM Broad Range Protein Marker, AMRESCO, USA as standard and run at 60 volts & 120 mA for about 45 min. When samples entered in resolving gel, and then gave 100 volts and 200 mA currents for around 2.5 hours. Furthermore, electrophoresis was carried out at a constant watt.The Gel was washed with 30% ethanol on Uni Thermo Shaker NTS-1300 EYELA, Japan at the constant shaking for 30 min. Then gels were placed in 10% glacial acetic acid in 50% methanol solution (Fixative) for 24 hours. SDS Gel was stained until protein bands were visible thereat placed as 5% of Methanol in 7.5% acetic acid glacial solution to destain the bands background. SDS-PAGE stain composed of 0.125% coomassie brilliant blue R-250 dissolved in 40% of Methanol and 7% acetic acid glacial solution. The stain was stirred on Magnetic stirrer & hot plate M6/1, Germany for 6-10 hours before used. Photographs were taken by Sanyo digital camera VPC-T1284BL and bands were scored through numbering pattern. Gels preserved in 10% acetic acid solution at 4°C.Interpretation of bands and data analysis: The total soluble protein bands relative mobility calculated by below formulae and Dendrogram was constructed via SPSS v. 20Where,F=(Migrated distance of protein band)/(Migrated distance of dye front)Slop=(Log MW of protein marker lower limit band–log〖MW of protein marker upper limit band )/(RF protein marker lower limit band –RF of protein marker upper limit band)RESULTS:The total soluble proteins extracted from green gram were perceived by SDS-PAGE Blue StepTm broad range biochemical markers. The protein-based marker was used to evaluate the toxic effect of sodium chloride along with pre-treatments of GAᴣ, IAA, and SA on proteome assay. In the current work, seedlings total soluble proteome resolved 24 polypeptide bands ranging from 200 to 17.1 kDa were recognized by using SDS-PAGE. The figure 1 showed Dendrogram assay, which classified the 20 treatments of SC, GAᴣ, IAA and SA into two major clusters where, the cluster I was the largest one (figure 1). Cluster I consisted of 15 treatments that further divided into I-A, and I-B. The pre-treatments of SC50+SA, SC150+SA, SC250+SA, and SC350+IAA were grouped together into C-1 of sub-cluster I-A. The C-2 of sub-cluster I-A, pre-treatment SC350+SA was most diverse among 20 treatments. The C-1 treatments showed 99% homology when compared with each other while, it was 97% similar with C-2. The sub-cluster I-B comprised another 10 treatments, SC0+GAᴣ, SC50+GAᴣ, SC150+GAᴣ, SC250+GAᴣ, SC350+GAᴣ, SC0+IAA, SC50+IAA, SC150+IAA, SC250+IAA, and SC0+SA that were also 99% similar for total proteins. Sub-cluster I-B pre-treatments was exhibiting 94% homology with the sub-cluster I-A. The second cluster was the smallest one that was divided into two sub-clusters, II-A and II-B. The II-A was comprised of SC50, SC150, and SC250 while, sub-cluster II-B consisted of SC0 and SC350. Within each sub-cluster, pre-treatments expressed 99% homology whereas, II-A was 97 different from II-B. Furthermore, cluster I showed 75% similarities with cluster II (figure 1). The seedlings storage proteome profile of green gram was shown in table 1.The results showed that 120kDa, 114.6 kDa, 51.8 kDa, 29.1 kDa and 22.8 kDa proteins bands were not induced at 0 mM SC, GAᴣ, IAA, and SA. The table 1 depicted the presence of 120 kDa and 114.6 kDa bands only at 350 mM SC level with all phytohormones treatments. Similarly, 51.8 kDa protein bands were appearing at 150SC, 250SC and 350SC stress with phytohormones. Based on the information collected from the NCBI protein database, this peptide was related to the 8S globulin alpha subunits. The two other, 7S globulins sub-units having 29.1kDa and 22.8 kDa molecular weights bands were synthesized under 50mM, 150mM, 250mM, 350mM SC stress with phytohormones. Concerning protein polypeptide of molecular weight 104.5 kDa, 99.8 kDa, 91.0 kDa, 55.0 kDa, and 46.0 kDa, those were induced by GAᴣ, IAA and SA at 0 to 350 mM SC. While, 17kDa protein band was appearing in SA, and IAA treated samples and 95.3kDa band was only present in SA treatment. Other 12 protein bands were present in all treatments proved as house-keeping proteins of green gram (table 1).DISCUSSIONThe SDS-PAGE profiling for proteome is the reliable and applied biochemical approach that has been used as biochemical marker in various crop differentiation, and characterization. In the current study, first time SDS-PAGE was utilized to investigate the impact of GAᴣ, IAA, and SA pre-soaking on green gram under salt toxicity. The salt toxicity adversely affects all seed, seedling, and plant metabolic process (Parveen et al., 2016). At salt toxicity, the endogenous GAᴣ, IAA, and SA levels markedly decrease (El-Khallal et al., 2009). In such condition, exogenous application of GAᴣ, IAA, and SA enhance seedlings survival rate by increasing synthesis of seed storage proteins. Likewise, our Dendrogram characterization based on 20 treatments showed significant diversity under 0 to 350 mM SC stress. The salicylic acid treatments were grouped together except SC0+SA treatment, exhibiting a close relationship, which proved its acclimating role under salt stress. These findings will help plant breeder toward enhancing food quality and quantity of green gram in future breeding programme on saline sodic land.The SDS-PAGE assay revealed 200. kDa, 109.4 kDa, 77 kDa, 68 kDa, 49 kDa, 38 kDa, 33 kDa, 26 kDa, 24 kDa, 22 kDa, 21 kDa and 19 kDa fractions as essential green gram proteins. Among these, 68 kDa, 49 kDa, 33 kDa, 26 kDa, 24 kDa and 21 kDa peptides were seed biotinylated isoform protein (Riascos et al., 2009), putative NADH-ubiquinone oxidoreductase subunit H (Gostinčar et al., 2019), heat shock protein 33 (Hamidian et al., 2015), globulin protein, seed coat / maturation protein (Dhaubhadel et al., 2005), and protein for dimerization. While, 22 kDa proteins belonged to the class of prolamin alpha zein Z1C1_2, Z1C1_4, and Z1C1_8 precursors, and 19kDa peptide was related with Z1A1_2, Z1A2_2, and Z1B_6 precursors (Miclaus et al., 2011). Further, the 91 kDa peptide is sucrose synthase SS1 protein, and 77kDa protein is the NADPH-cytochrome P450 reductase (Wang et al., 2004). Also, the phosphatase-associated two other proteins having 46 and 55 kDa molecular weight were reported earlier in Mucuna pruriens. Hameed et al. (2012) and Malviya et al. (2008) found 55 and 46kDa peptides as 7S vicilin small sub-units and 17kDa as 11S globulins sub-unit in the studied Vigna radiata. Some other molecular weight proteome such as 68 kDa and 49kDa are 7S vicilin, 33kDa is 8S vicilin, 38 and 26kDa 8S globulins, 24kDa 11S globulins, and 22kDa 16.5S globulins. These proteins required for germination and seed establishment of green gram plant (Hameed et al., 2012).The vast accumulation of 23kDa and 22kDa peptides under salt stress by salicylic acid, were reported previously in the mangrove Bruguiera parviffora and Zea mays (El-Khallal et al., 2009). Correspondingly, El-Kafafi et al. (2015) reported the presence of 115kDa, 23kDa, and 22kDa bands in the salt tolerant lines of green gram. These proteomes induced under salt stress may play a pivotal part in the stress acclimation and osmotic adjustment. Similarly, the induction of 104 kDa and 100kDa MW polypeptide by SC stress in the salt tolerant genotypes of green gram indicated the functional role of phytohormones in various metabolic and defense response El-Kafafi et al. (2015); Alharby et al. (2016), El-Khallal et al. (2009), Qados (2010). Ali et al. (2007), Alharby et al. (2016), and El-Kafafi et al. (2015) observed 17kDa, 26kDa, 33kDa and 77kDa bands involving in salt tolerance and can be considered as a positive biochemical marker for salt stress. Further, 26 kDa MW peptide also functions as osmotin under the salt stress that involved in enhancing the accumulation of glycine betaine and proline in the cells. Hence, proteome assay of green gram showed that GAᴣ, IAA, and SA could regulate the expression of salt stress proteins that are anticipated to play a crucial part in the salt tolerance mechanism. Likewise, the involvement of phytohormones in the induction of changes in the proteome profile pattern was attributed to their part in managing cell division by regulating some genes of apical meristems.CONCLUSIONFinally, the results revealed the presence of the ten new bands with MW of 200kDa, 120 kDa, 114.6 kDa, 109.4kDa, 104.5kDa, 99.8kDa, 95.3kDa, 51.8kDa, 29.1kDa and 22.8kDa have not reported previously under salt stress with phytohormones treatments in green gram. Furthermore, it was observed that phytohormones alleviate the negative impact of salt stress on green gram by enhancing synthesis of salt defense polypeptides. Hence, higher accumulation of proteins was observed in salicylic acid treated seedlings. Thus, present work recommended the pre-soaking of phytohormones to overcome the toxic impact of sodium chloride on green gram. Further research is needed on a biomolecular level to reveal the mechanism of signalling pathways under sever salt stress.CONFLICT OF INTERESTBoth authors have declared that no disagreement of interest regarding this research.REFERENCES Alharby, H. F., E. M. Metwali, M. P. Fuller and A. Y. Aldhebiani, 2016. The alteration of mRNA expression of sod and gpx genes, and proteins in tomato (Lycopersicon esculentum Mill) under stress of Nacl and/or ZnO nanoparticles. Saudi journal of biological sciences, 23(6): 773-781.Ali, A., M. Mageed, I. Ahmed and S. Mariey, 2007. Genetic and molecular studies on barley salt tolerance. In: African crop science conference proceedings. pp: 669-682.Chadha, M., 2010. Short duration mungbean: A new success in South Asia. Asia-Pacific association of agricultural research institutions.Dhaubhadel, S., K. Kuflu, M. C. Romero and M. Gijzen, 2005. A soybean seed protein with carboxylate-binding activity. Journal of experimental botany, 56(419): 2335-2344.El-Kafafi, E.-S. H., A. G. Helal, S. F. El Hafnawy and R. Flaah, 2015. Characterization and evaluation of some mungbean genotypes for salt tolerance. World applied science journal, 33(3): 360-370.El-Khallal, S. M., T. A. Hathout, A. Ahsour and A.-A. A. Kerrit, 2009. Brassinolide and salicylic acid induced antioxidant enzymes, hormonal balance and protein profile of maize plants grown under salt stress. Research journal of agriculture biological sciences, 5(4): 391-402.GOP, 2018. Pakistan economic survey from 2017 to 2018. Ministry of Finance. Islamabad. Government of Pakistan. Accessed 18-8-2019, http://www.finance.gov.pk/su rvey/chapters18/02-Agriculture.pdf.Gostinčar, C., M. Turk, J. Zajc and N. Gunde‐Cimerman, 2019. Fifty aureobasidium pullulans genomes reveal a recombining polyextremotolerant generalist. Environmental microbiology, 21(10): 3638-3652.Hameed, A., M. Qureshi, M. Nawaz and N. Iqbal, 2012. Comparative seed storage protein profiling of mung bean genotypes. Pakistan jouranl of botany, 44(6): 1993-1999.Hamidian, M., J. Hawkey, K. E. Holt and R. M. Hall, 2015. Genome sequence of Acinetobacter baumannii strain d36, an antibiotic-resistant isolate from lineage 2 of global clone 1. Genome announced, 3(6): e01478-01415.Hou, D., L. Yousaf, Y. Xue, J. Hu, J. Wu, X. Hu, N. Feng and Q. Shen, 2019. Mung bean (vigna radiata l.): Bioactive polyphenols, polysaccharides, peptides, and health benefits. Nutrients, 11(6): 1238.Itoh, T., R. N. Garcia, M. Adachi, Y. Maruyama, E. M. Tecson-Mendoza, B. Mikami and S. J. A. C. S. D. B. C. Utsumi, 2006. Structure of 8sα globulin, the major seed storage protein of mung bean. Acta crystallographica section D: Biological crystallography, 62(7): 824-832.Khan, F. F., K. Ahmad, A. Ahmed and S. Haider, 2017. Applications of biotechnology in agriculture-review article. World journal of biology biotechnology, 2(1): 139-142.Malviya, N., S. Nayak and D. Yadav, 2008. Characterization of total salt soluble seed storage proteins of grain legumes using sds-page. Bulletin de ressources phytogénétiques(156): 50.Mendoza, E. M. T., M. Adachi, A. E. N. Bernardo and S. Utsumi, 2001. Mungbean [Vigna radiata (L.) wilczek] globulins: Purification and characterization. Journal of agricultural food chemistry, 49(3): 1552-1558.Miclaus, M., J.-H. Xu and J. Messing, 2011. Differential gene expression and epiregulation of alpha zein gene copies in maize haplotypes. PLoS genetics, 7(6).Mushtaq, F., S. A. Jatoi, S. S. Aamir and S. U. Siddiqui, 2018. Genetic variability for morphological attributes and seed protein profiling in chili (Capsicum annuum L.). Pakistan jouranl of botany, 50(4): 1661-1668.Parveen, A.-u.-H. M., J. Akhtar and S. M. Basra, 2016. Interactive effect of salinity and potassium on growth, biochemical parameters, protein and oil quality of soybean genotypes. Pakistan journal of agricultural sciences, 53(01): 69-78.Qados, A., 2010. Effect of arginine on growth, nutrient composition, yield and nutritional value of mung bean plants grown under salinity stress. Nature, 8: 30-42.Riascos, J., W. Burks, L. Pons, A. Weissinger and S. Weissinger, 2009. Identification of a soybean seed biotinylated protein as a novel allergen. Journal of allergy cinical Immunology, 123(2): S24.Wang, S. Y., J. H. Wu, T. Ng, X. Y. Ye and P. F. Rao, 2004. A non-specific lipid transfer protein with antifungal and antibacterial activities from the mung bean. Peptides, 25(8): 1235-1242.Yi-Shen, Z., S. Shuai and R. FitzGerald, 2018. Mung bean proteins and peptides: Nutritional, functional and bioactive properties. Food nutrition research, 62.

Aim: This study evaluate the agronomic and quality characteristics of grape (Vitis vinifera L.) varieties in a non-traditional region of the Agreste of Pernambuco States.
 Study Design: The experiment was conducted in a randomized block design with five replications and eight plants per plot.
 Place and Duration of Study: Was carried out in the municipality of Brejão, PE, at the Experimental Station of the Agronomic Institute of the Pernambuco. The vines were implanted on September, 2013, whose pruning was performed on August and harvesting began on December, 2016 to January, 2017.
 Methodology: Ten treatments represented by the varieties of European vines: Cabernet Sauvignon, Malbec, Merlot Noir, Petit Verdot, Pinot Noir and Syrah for producing of red wines and Chardonnay, Muscat Petit Grain, Sauvignon Blanc and Viogner for producing of white wines, grafted on the Paulsen 1103 rootstock were evaluated. The vineyard was conducted in espalier vine-tying system in double short pruning type, with spacing 3 m x 1 m. The characterization of the phenological stages was made using as reference the phenological scale. The thermal requirement of the crop per period was estimated. Agronomic characteristics were also evaluated, such as: fertility of gems, budding (%), production, productivity, number of bunches per plant, length and width of bunch, bunch weight, soluble solids, titratable acidity, hydrogen ionic potential, SS / TA ratio, volume of 100 berries, yield of must, mass of the husks and seeds. The data were submitted to two selection indices: Classic Index and Distance Genotype-Ideotype Index.
 Results: Sprouting varied from 13.68% (Petit Verdot) to 81.6% (Sauvignon Blanc) and the fertility of gems from 0.1 bunch.bud-1 (Chardonnay) to 0.67 bunch.bud-1 (Sauvignon Blanc). The pruning cycle and Day Degrees (DD) cumulated ranged from 133 days and 1,684 DD (Muscat Petit Grain) to 167 days and 2,070 DD (Merlot Noir). The number of bunches ranged from five (Merlot Noir) to 29 bunches.plant-1 (Sauvignon Blanc). Muscat Petit Grain stood out for bunch weight, not differing from Syrah and Malbec. The varieties showed no difference in length and width of bunches. In the volume of 100 berries, Muscat Petit Grain (213.6 ml) and Malbec (216.0 ml) stood out. For the yield of must, Sauvignon Blanc (70.87%) stood out, not differing from Malbec (64.31%), Viognier (69.79%), Muscat Petit Grain (70.22%). Muscat Petit Grain, Sauvignon Blanc and Viognier (white wine), Cabernet Sauvignon, Malbec, Merlot Noir and Syrah obtained acceptable values for soluble solids (SS), titratable acidity (TA), SS/TA ratio and pH. From the selection index analyzes, the Muscat Petit Grain, Cabernet Sauvignon and Syrah varieties were indicated for the selection by the highest Mulamba and Mock index and by the Genotype-Ideotype distance index.
 Conclusion: The cycle of grapevine varieties evaluated in the Garanhuns, PE, Microregion is longer than that observed in the sub Medio of the São Francisco Valley, similar to those in the South Region of Brazil. In the evaluated cycle the varieties produced grapes with characteristics suitable for the production of quality fine wines, showing to be promising for this non-traditional microregion in the production of fine grapes. From the selection index analyzes, the Muscat Petit Grain, Syrah and Cabernet Sauvignon varieties were indicated for selection by the highest Mulamba and Mock index and Genotype-Ideotype distance index.

Abstract Study question Does culture in high relative humidity conditions (HC) improve pregnancy rates when using a time-lapse system (TLS) and single-step (SS) culture medium? Summary answer Using an integrated-TLS and SS medium, culture under HC increases the likelihood of embryos to achieve a pregnancy with respect to those cultured in DC. What is known already Many variables affect embryo development, and need to be precisely tuned in every IVF laboratory, especially inside the incubators. TLS provide stability during embryo culture, which is a well-known key factor for a proper embryo development. The humidity content of culture atmosphere is especially relevant in order to avoid oscillations in culture media osmolality. It has been previously reported that culture under HC has a significant effect on embryo quality and morphokinetics. However, studies assessing the effect of HC in clinical outcome are rare and inconclusive, mostly due to the variability in the incubator device used and insufficient sample size. Study design, size, duration The present is a retrospective study performed over 1624 ICSI treatments from 3 fertility clinics from December 2017 to October 2020. Zygote cohorts were randomly assigned to dry (N = 794) or humid conditions (N = 830). It includes autologous treatments with (N = 555) and without (N = 368) pre-implantation genetic testing (PGT) and egg donation treatments (N = 701). Following selection by combining morphological and morphokinetic criteria, 1611 mostly single embryo transfers (92%) were performed, 779 from DC and 832 from HC. Participants/materials, setting, methods Stimulation, oocyte pickup and fertilization were performed according to the standard procedures of the clinic. We used a GERI incubator (Genea Biomedx), with 6 separated chambers for individual patients, 3 of them configured to work in DC, and 3 in HC. Embryos were cultured in specific 16-well GERI trays with single-step Gems® culture medium (Genea Biomedx). The effect of HC in pregnancy rate was assessed by multivariate logistic regression and Pearson Chi Square Test. Main results and the role of chance Types of treatment and patient demographics were homogeneously distributed in the two study groups. Mean patient age was 39.88±4.47 years, BMI: 23.54±4.21 Kg/m2 and number of correctly fertilized oocytes: 7.86±3.87. A logistic regression was performed, including other possible affecting factors: ovum age and origin, transfer day, fresh or frozen-warmed embryo transfer, number of transferred embryos and the use of PGT. Said analysis revealed that embryos cultured in HC are more likely to achieve a pregnancy than those cultured in DC (OR = 1.30, 95% CI (1.05-1.59), p=0.014). Pregnancy rate was significantly higher in HC (66.7%) than in DC (60.9%) in the total embryo transfers (p = 0.017). Pregnancy rate was also higher in HC in fresh embryo transfers (68.6% in HC vs 63.2% in DC; p = 0.133) and frozen-thawed transfers (65.2% in HC vs 59.1% in DC; p = 0.062), although differences were not statistically significant due to the reduced sample size. Stratifying the results, the significant difference remained in transfers belonging to autologous cycles (68.4% HC vs 56.5% in DC; p = 0.030) and in treatments in which PGT was performed (67.1% HC vs 56.0% in DC; p = 0.023), but the difference in egg donation procedures was not statistically significant (66.4% in HC vs 64.7% in DC, p = 0.577). Limitations, reasons for caution This is a retrospective analysis performed over the clinics’ treatments, so it might be compromised by some bias, although multivariable analysis may overcome them. For further assessing the effect of HC in clinical results a prospective controlled study, with a larger sample size could be performed, also comparing life-birth rates. Wider implications of the findings These results, alongside our previous findings (Valera et al. 2020, Albert et al. 2020), support that HC contributes to optimize embryo development and clinical results in undisturbed culture in TLS with single-step medium. To our knowledge, this is the largest study on the matter and the first performing multivariable analysis. Trial registration number Not applicable

O presente trabalho objetivou verificar a prenhez de ovelhas inseminadas por via intrauterina ou cervical com sêmen refrigerado a 5ºC durante 8 horas nos diluentes citrato-gema e Cornell University Extender (CUE). Foram utilizados 4 carneiros da raça Suffolk. Em cada inseminação era colhido sêmen de 2 animais e após a avaliação seminal as amostras eram homogeneizadas, divididas em 2 alíquotas e diluídas na proporção de uma parte de sêmen para três de diluente em citrato-gema e CUE. A diluição ocorria a 30ºC e o abaixamento progressivo da temperatura se dava em 2 horas em caixa de isopor contendo gelo. As amostras permaneceram nesta temperatura durante 8 horas. Foram inseminadas 91 ovelhas mestiças da raça Suffolk, as quais tiveram o estro sincronizado com pessários vaginais impregnados com 50 mg de acetato de medroxiprogesterona, permanecendo por 14 dias, quando se administrou 500 UI de Gonadotrofina Coriônica (eCG) via intramuscular. Todas as ovelhas foram inseminadas independente do aparecimento do estro. A aplicação do sêmen foi realizada pela via cervical e intra-uterina por laparoscopia. A dose mínima inseminante foi de 250 milhões de espermatozóides, em volume de 0,4 ml, por ovelha. A utilização do diluente CUE resultou em 69,56% (n=23) e 8,33% (n=24) de prenhez para as vias intra-uterina e cervical, respectivamente contra 85,71% (n=21) e 21,74% (n=23) para o diluente citrato-gema. O sêmen diluído em citrato-gema forneceu índices de prenhez superiores, porém não significativos (P>0,05), nas duas vias estudadas em relação ao CUE. A fertilidade foi superior quando se utilizou a via intra-uterina (P0.05) has been found between values of pregnancy attained with suspensions of spermatozoa either in CUE or in citrate-york extender, in spite of the fact that the latter one gave higher pregnancy rates. Intrauterine insemination gave higher values of fertility (P

Abstract Study question Whether empty follicle syndrome (EFS) in a patient has a genetic basis. Summary answer Our findings would expand the mutational spectrum of LHCGR, in patients with GEFS What is known already The LHCGR gene (OMIM #52790) is located on chromosome 2p21 has 11 exons The LHCGRs present in gonadal cells; granulosa, theca and luteal cells in women and Leydig cells in menandplays a critical role in male sexual differentiation, female ovarian development and fertility (folliculogenesis, ovulation, corpus luteum formation and progesterone secretion) Inactivating mutations in males can lead to Leydig cell hypoplasia, which causes disorders in sexual development (MIM #238 320). Phenotype of women is less severe and variable and has no effect on the secondary sex characteristics, but it could cause amenorrhoea and infertility Study design, size, duration: In the context of clinical genetics, Next Generation Sequencing libraries were prepared in line with the manufacturer’s orders using QIASeq™ Targeted DNA Custom Panel (Qiagen) targeting exons and 20 bp exon-intron boundaries of selected genes (AR, BMP15, CATSPER1, CFTR, CYP21A2, FSHB, FSHR, HESX1, LHB, LHCGR, NR5A1, POU1F1, SRY, ZP1). The variant detected in NGS analysis was further confirmed using Sanger sequencing (MiSeq, Illumina, San Diego, CA). Participants/materials, setting, methods A 27 years old Turkish women with 6 year history of primary unexplained infertility underwent controlled ovarian hyperstimulation and IVF with an antagonist protocole in Ankara City Hospital. Although normal follicular development, E2 levels, and bioavailable β-hCG plasma levels, no oocytes or cumulus-corona complexes were retrieved by follicular aspiration. Main results and the role of chance A novel heterozygous mutation on Exon 5 of LHCGR (NM_000233.4):c.453C>G (p.Phe151Leu). This variant has not been reported in GnomAD database and is a novel variant as per controlled from ClinVar and HGMD mutation databases Also coagulation tests were done Patient was heterozygote for the prothrombin mutation (FXIII) and homozygote for MTHFR C677T mutation. The patient has normal pubertal development and female karyotype (46,XX). Gonadotropin and E2 levels were normal, nor any history of anosmia, primary amenorrhoea, polycystic ovaries, hyperandrogenism, systemic disorder, or neurologic defect. According to ACMG 2015 and HGMD detected variant was classified as unknown clinical significance (VUS) variant In 22nd World Congress COGI in 2015 we have presented another recurrent GEFS case with compound heterozygous frameshift mutations in exon 5 and 11 of LHCGRgene(c.1764_1765insT) and (c.430G>T, p.V144F) who developed premature ovarian failure at the end The different types of LHCGR mutations would lead to the different functional effects and clinical phenotypes in different ages; primary amenorrhea with high levels of LH, poor ovarian response (Poesidon group1), GEFS and secondary amenorrhea (premature ovarian failure) GEFS may be a gradual biological occurrence related to ovarian ageing. Long term follow up is important, since some clinical manifestations appear later in life. Limitations, reasons for caution LH resistance in females were always found due to their effected brothers and only 10 GEFS cases of 46, XX females with LHCGR gene defect reported in literature. Patient was heterozygous for indicated mutation; therefore segregation study should be done in family members and in vitro studies should be performed. Wider implications of the findings: Screening for mutations in the LHCGR gene in Poseidon group 1 and in patients GEFS especially the recurrent ones will provide valuable information and time for clinical management, corresponding treatment and give the opportunity of wise and cost effective counseling of patients about their future reproductive choices. Trial registration number it is a case report

Abstract STUDY QUESTIONS Does the application of anti-adhesion gel, compared to no gel, following operative hysteroscopy to treat intrauterine pathology in women wishing to conceive increase the chance of conception leading to live birth? WHAT IS KNOWN ALREADY Intrauterine adhesions (IUAs) following operative hysteroscopy may impair reproductive success in women of reproductive age. Anti-adhesion barrier gels may decrease the occurrence of IUAs, but the evidence on their effectiveness to improve reproductive outcomes is sparse and of low quality. STUDY DESIGN, SIZE, DURATION This multicentre, parallel group, superiority, blinded and pragmatic randomised controlled trial is being carried out in seven participating centres in Belgium. Recruitment started in April 2019. Women will be randomly allocated to treatment with anti-adhesion gel (intervention group) or no gel (control group). Sterile ultrasound gel will be applied into the vagina as a mock-procedure in both treatment arms. The patient, fertility physician and gynaecologist performing the second-look hysteroscopy are unaware of the allocated treatment. Power analysis, based on a target improvement of 15% in conception leading to live birth using anti-adhesion gel, a power of 85%, a significance level of 5%, and a drop-out rate of 10%, yielded a number of 444 patients to be randomised. The baseline rate of conception leading to live birth in the control group is expected to be 45%. PARTICIPANTS/MATERIALS, SETTING, METHODS Women of reproductive age (18–47 years), wishing to conceive (spontaneously or by fertility treatment) and scheduled for operative hysteroscopy to treat intrauterine pathology (endometrial polyps, myomas with uterine cavity deformation, uterine septa, IUAs or retained products of conception) are eligible for recruitment. Women may try to conceive from 3 to 6 weeks after receiving allocated treatment with follow-up ending at 30 weeks after treatment. If the woman fails to conceive within this timeframe, a second-look hysteroscopy will be scheduled within 2–6 weeks to check for IUAs. The primary endpoint is conception leading to live birth, measured at 30 weeks after randomisation. The secondary endpoints are time to conception, clinical pregnancy, miscarriage and ectopic pregnancy rates, measured at 30 weeks after receiving allocated treatment. The long-term follow-up starts when the patient is pregnant and she will be contacted every trimester. STUDY FUNDING/COMPETING INTEREST(S) This work is funded by the Belgian Healthcare Knowledge Centre (KCE). The anti-adhesion gel is supplied at no cost by Nordic Pharma and without conditions. Dr. Tomassetti reports grants and non-financial support from Merck SA, non-financial support from Ferring SA, personal fees and non-financial support from Gedeon-Richter, outside the submitted work. None of the other authors have a conflict of interest.

Abstract Study question Can we quantify the variation in epigenetic signature of constitutive heterochromatin (cHC) in human sperm using Western blot analysis and correlate this with pregnancy outcome after IVF? Summary answer Variation in the quantified H3K9me3 to H3 showed an association with embryo morphokinetics and a H3k9me3/H3 ratio <0,5 is negatively associated with pregnancy outcome. What is known already In human sperm, 5–15% of the DNA remains associated to histones. We previously showed that human mature spermatozoa still exhibit the canonical marks H3K9me3 and H4K20me3 on cHC regions and transmit these nucleosomes to the embryo after fertilization. Additionally, other groups reported an increase in the nucleosome/protamine ratio when sperm from male factor subfertility patients were compared with sperm from fertile men, indicating that incomplete chromatin remodelling during spermatid elongation may affect fertility. Thus, variations in sperm chromatin occur during human spermatogenesis that go undetected by current sperm quality tests and may have clinical significance for IVF treatment outcomes. Study design, size, duration An observational study of patient couples (n = 53) undergoing IVF treatment at the Erasmus MC, Rotterdam, between 2017 and 2018. Inclusion criteria were normospermia according to the WHO criteria, at least 4 oocytes harvested after ovum pickup and an embryo-transfer. After routine insemination, resulting embryos from 38 cycles were cultured in the EmbryoScopeTM and developmental timings up to the 8-cell stage were recorded. The primary outcome measure was ongoing pregnancy (ultrasound at 12 weeks of gestation). Participants/materials, setting, methods Surplus sperm samples from 53 IVF cycles were lysed at a concentration of 200x106 spermatozoa/ml in RIPA buffer. Lysates were loaded and separated on SDS-PAGE gels and transferred onto blotting membranes. After incubation with antibodies against H3 and H3K9me3 and fluorescent detection, signal intensity was quantified using Image Studio Lite (LI-COR Biotechnology)). Additionally, time-lapse developmental kinetics of 130 transferred and cryopreserved embryos of 38 patient couples were analyzed. Main results and the role of chance No significant differences were found in patient characteristics between the pregnant and non-pregnant group. However, fertilization rate was found to be a confounding factor; no pregnancies occurred below a fertilization rate of 40% and only 4 occurred below a fertilization rate of 60%. The median H3k9me3/H3 ratio in the pregnant group was 1.01 (interquartile range (IQR): 0.69–1.16) and in the non-pregnant group 0.896 (IQR: 0.54–1.06), this difference was not significant. When using logistic regression analysis, adjusted for the fertilization rate (dichotomized as ≤ 60% or > 60%), we observed a positive association between the H3k9me3/H3 ratio and ongoing pregnancy (odds ratio (OR) 6.43 (95% confidence interval (CI): 1.12–36.8; p-value 0.037). More importantly, none of the samples with a ratio of 0.5 or lower resulted in an ongoing pregnancy (n = 7). After classifying sperm samples into quartiles according to the H3K9me3/H3 ratio, we observed that resulting embryos reached the 4-cell stage faster with sperm from to the lowest quartile of H3K9me3/H3 ratio, compared to the highest quartile (beta –4.4 hours, p-value 0.05). Our results point to an association between the H3k9me3/H3 ratio, embryo development and pregnancy outcome. Limitations, reasons for caution This initial analysis is performed with a relatively low number of samples. The OR of 6.43 has a wide 95% CI, probably due to low statistical power. Our observations await confirmation in a larger data set. Wider implications of the findings: Semen analysis remains the cornerstone of male infertility diagnosis, despite its low prognostic value. Sperm epigenetic cHC inheritance likely has an important impact on early embryo development. Therefore, the H3k9me3/H3 ratio could serve as a parameter for sperm quality and aid in the prediction of pregnancy outcome. Trial registration number Not applicable