Variation of female pronucleus reveal oocyte or embryo abnormality: An expert experience deep learning of non-dark box analysis

Background: Pronuclear assessment appears to have the ability to distinguish good and bad embryos in the zygote stage,but paradoxical results were obtained in clinical studies.This situation might be caused by the robust qualitative detection of the development of dynamic pronuclei. Here,we aim to establish a quantitative pronuclear measurement method by applying expert experience deep learning from large annotated datasets. Methods: Convinced handle-annotated 2PN images(13419) were used for deep learning then corresponded errors were recorded through handle check for subsequent parameters adjusting. We used 790 embryos with 52479 PN images from 155 patients for analysis the area of pronuclei and the preimplantation genetic test results.Establishment of the exponential fitting equation and the key coefficient β1 was extracted from the model for quantitative analysis for pronuclear(PN) annotation and automatic recognition. Findings: Based on the female original PN coefficient β1,the chromosome normal rate in the blastocyst with biggest PN area is much higher than that of the blastocyst with smallest PN area(58.06% vs.45.16%, OR=1.68[1.07-2.64];P=0.031).After adjusting coefficient β1 by the first three frames which high variance of outlier PN areas was removed, coefficient β1 at 12 hours and at 14 hours post-insemination,similar but stronger evidence was obtained. All these discrepancies resulted from the female propositus in the PGT(SR) subgroup and smaller chromosomal errors. Conclusion(s): The results suggest that detailed analysis of the images of embryos could improve our understanding of developmental biology. Funding: None

Time-dependent expression of ryanodine receptors in sea urchin eggs, zygotes and early embryos

Summary In this work, the presence of calcium-dependent calcium channels and their receptors (RyR) has been investigated in Paracentrotus lividus eggs and early embryos, from unfertilized egg to four-blastomere stages. Electrophysiological recordings of RyR single-channel current fluctuations showed that RyRs are functional during the first developmental events with a maximum at zygote stage, c. 40 min after fertilization, corresponding to the first cleavage. The nature of vertebrate-like RyRs active at this stage was established by specific activation/blockade experiments.

LEAFY COTYLEDON1 expression in the endosperm enables embryo maturation in Arabidopsis

The endosperm provides nutrients and growth regulators to the embryo during seed development. LEAFY COTYLEDON1 (LEC1) has long been known to be essential for embryo maturation. LEC1 is expressed in both the embryo and the endosperm; however, the functional relevance of the endosperm-expressed LEC1 for seed development is unclear. Here, we provide genetic and transgenic evidence demonstrating that endosperm-expressed LEC1 is necessary and sufficient for embryo maturation. We show that endosperm-synthesized LEC1 is capable of orchestrating full seed maturation in the absence of embryo-expressed LEC1. Inversely, without LEC1 expression in the endosperm, embryo development arrests even in the presence of functional LEC1 alleles in the embryo. We further reveal that LEC1 expression in the endosperm begins at the zygote stage and the LEC1 protein is then trafficked to the embryo to activate processes of seed maturation. Our findings thus establish a key role for endosperm in regulating embryo development.

46 Use of time-lapse imaging technology to assess relationships of morphological and phototextural attributes of presumptive ovine zygotes and early embryos with their developmental competence invitro

The assessment of morphology and digital image opacity may provide valuable information on embryo viability because such traits are linked to embryonic gene expression, metabolism and ultrastructure. Time-lapse imaging has been used in research to monitor the dynamic nature of the developing pre-implantation embryo, which includes capturing alterations in various morphological parameters over time. The present study examined the effectiveness of time-lapse technology in assessing several morphometric and phototextural parameters for predicting the developmental potential of ovine embryos. The development of 37 long wool sheep embryos from IVF to the blastocyst stage was monitored and evaluated using Primo Vision time-lapse imaging technology. Image-Pro Plus software was then used to measure zona pellucida thickness, embryo diameter, cellular grey-scale pixel intensity and heterogeneity, and total area of the perivitelline space. A one-way analysis of variance (ANOVA) was done using SigmaPlot® 11.0 for all attributes at various time points during embryo development [i.e. presumptive zygote stage, t(0); first cleavage, t(2) or t(3); second cleavage, t(4) or t(6); and third cleavage, t(7) or t(8)]. Our results indicate that most parameters analysed did not differ among embryos varying in their developmental fate, with the exception of the perivitelline space area, which was greater (P<0.05) for non-dividing embryos than for future blastocysts at the presumptive zygote stage (4040±4137 vs. 857±642µm2, respectively; mean±s.d.). Consequently, the measurement of perivitelline space at t(0) could be used to predict developmental potential of invitro-produced ovine embryos, but further investigation is required.

Observations on the Embryonic Development of Trichuris sylvilagi (Nematoda, Trichuridae) under Laboratory Conditions

In the present study, we observed in vitro the rate and stages of exogenous development of the nematode Trichuris sylvilagi Tiner, 1950 isolated from the European hare, Lepus europaeus. The viability of eggs and embryos at different temperatures was investigated. It was found that the most favorable temperature for embryonic development of the species was 30 °C, with the formation of 88.67 % of viable eggs with a motile larva in 20 days at laboratory conditions. With a decrease in temperature to 20 °C, the embryogenesis occurred in 32 days. The viability of eggs cultured at 20 °C decreased: 68.0 % of eggs reached larval stage, and 32.0 % of eggs died. At 10 °C eggs did not develop to infective stage. On the 32nd day of cultivation at 10 °C, 27.33 % of eggs remained at the zygote stage, 52.0 % at the stage of blastomere cleavage and formation, and 20.67 % died. Embryogenesis was characterized by metric changes in egg parameters. At optimal temperature, the growth and development of eggs was accompanied by an increase in the egg and plugs' width with a simultaneous decrease in their length, as well as with thinning of the shell

Features of exogenous development of Trichuris globulosa (Nematoda, Trichuridae)

Parasitic nematodes of the genus Trichuris Röderer, 1761 are hematophagous helminths, capable of parasitizing many different hosts including humans. The domestic and wild ruminants are hosts of several Trichuris species, with Trichuris globulosa (Linstow, 1901) one of the most prevalent parasites found in cattle. The exogenous stages of the helminth’s life cycle develop in the outer environment, and their activity and survival depend on the abiotic factors. Thus the aim of the work was to determine the influence of temperature on the rate and success of development of infectious eggs of T. globulosa in laboratory culture considering their morphological and metric changes. The results of experimental studies showed that the embryogenesis of T. globulosa eggs, cultured in laboratory conditions and obtained from gonads of female nematodes, occurs in six stages regardless of the temperature regime. At the same time, the rate of transition from one stage to another, the term for the formation of infectious eggs and their viability directly depends on the temperature of the external environment. At the optimal temperature for the development of T. globulosa eggs, 25 °C, 76.3% of eggs reached the mobile larva stage. A decrease in temperature to 20 °C and an increase to 30° C led to an increase in the egg mortality of up to 26% and 32%, respectively, and the viability of eggs decreased. Under such temperature conditions, 74% and 68% of eggs of Trichuris reached the mobile larva stage. With an increase in the culture temperature, the time of embryogenesis decreased and amounted to 56 days at 20 °C, 48 days at 25 °C, and 32 days at 30 °C. Depending on the temperature regime, the zygote stage lasted from the release of eggs from the gonads of female nematodes to 12 days, the stage of blastomere formation from 4 to 12 days, the stage of the bean-shaped embryo from 8 to 24 days, the stage of the tadpole embryo from 16 to 36 days, the stage of larval formation lasted from 16 to 48 days, the stage of mobile larva from days 20 to 56. The formation of the infectious T. globulosa egg from a non-infectious one is shown in an experiment to include the metric changes confirmed by metric indicators. Mature eggs with a mobile larva are shorter and wider, with shorter and narrower plugs and a thinner shell than eggs at the zygote stage. The obtained research results on the influence of temperature on the exogenous development of Trichuris globulosa will make it possible to predict the epizootic situation of trichurosis in animal husbandry, as well as take timely measures to arrest the embryonic stages of nematode development in the external environment.

Extracellular Vesicles Mediated Early Embryo–Maternal Interactions

Embryo–maternal crosstalk is an important event that involves many biological processes, which must occur perfectly for pregnancy success. This complex communication starts from the zygote stage within the oviduct and continues in the uterus up to the end of pregnancy. Small extracellular vesicles (EVs) are part of this communication and carry bioactive molecules such as proteins, lipids, mRNA, and miRNA. Small EVs are present in the oviductal and uterine fluid and have important functions during fertilization and early embryonic development. Embryonic cells are able to uptake oviductal and endometrium-derived small EVs. Conversely, embryo-derived EVs might modulate oviductal and uterine function. In this review, our aim is to demonstrate the role of extracellular vesicles modulating embryo–maternal interactions during early pregnancy.

A reanalysis of Schaefer et al. does not indicate extensive CRISPR/Cas9 mediated off-target editing events

Recently Schaefer et al. (1) reported the presence of more than a thousand genomic differences between mice that had been edited with S. pyogenes Cas9 at the zygote stage, and a control mouse of the same strain. Given the overlap of genomic differences between the two edited mice that were not found in the control mouse, the authors concluded that these differences arose from a Cas9-dependent activity. We feel that this conclusion is inappropriate, for three key reasons: 1) there was incomplete analysis of the genetic variation; 2) there was no consideration that the variation is naturally arising in these animals; and 3) the inferred behavior of Cas9 lies outside of its understood mechanism of action. Attribution to Cas9 activity should require a burden of proof, that we believe has not been met.