Hatchability Of Fertile Eggs Research Articles

Leachate from plastic food packaging induced reproductive and neurobehavioral toxicity in zebrafish

The present study mimicked daily life exposure to plastic food package bags and evaluated its effects on the reproductive and neurobehavioral responses using zebrafish model. Gas chromatography-mass spectrometer (GC/MS) full scan analysis revealed that phthalic acid, isobutyl octyl ester (DEHP) and its metabolites were the main leachate from plastic bags. Our results demonstrated that during the eight weeks exposure, leaching from plastic bags treated with boiling water (P-high group) significantly affected the spawn egg production, embryo hatching and larval malformation rate. Cross-spawning trails between zebrafish collected from the controls and P-high group at the end of eight weeks showed that these adverse effects were more severe in the offspring derived from paternal exposure than those derived from the maternal exposure, suggesting leached chemicals may have a more pronounced effect in sperm than in eggs. In addition, P-high group male testis weight, sperm motility and sperm swimming velocities were decreased significantly. After eight weeks treatment, neurobehavioral tests demonstrated significant changes in the swimming speed during free swimming and light-dark stimulation in the adult zebrafish from P-high group, with the effects being more severe in the males than females. P-high group males also showed altered response in the light/dark explore and mirror attacks assays.

The regulative effect of Urtica dioica on sex hormones imbalance: elevated follicle-stimulating hormone/luteinizing hormone ratio ≥4.5 is associated with low performance in aged breeder quails

The age-related reproductive disorders are the main concerns in old birds. It was suggested that a drop in egg production and reproductive performance, towards the end of their laying period was caused partly by a decrease in the baseline concentration of plasma LH. Urtica dioica (nettle) is a plant with natural aromatase inhibitors. Steroid hormone levels are regulated by inhibition of the aromatase enzyme. Few studies have examined the effect of nettle on the egg production in adult hens. The aim of this study was to investigate the effects of diet supplemented with nettle powder (NP) in aged quails. One hundred and forty-52-week-old Japanese quails were randomly assigned to four treatments consisting of seven replicates (n = 5; four females and one male) and fed with diets containing NP at 0% (control group), 0.5, 1.0, and 1.5% (treatment groups). At 62 week of age, our results indicated the NP improved egg production, feed conversion ratio, eggshell thickness and Haugh unit (p < .05). Notably, fertility and hatchability of fertile eggs were significantly increased, while total embryonic mortality decreased significantly by supplementing diet with nettle powder (p < .05). Higher luteinizing hormone, lower oestrogen, malondialdehyde and total cholesterol and triglyceride concentrations were associated with percent of the nettle powder in diet (p ≤ .05). Elevated follicle-stimulating hormone/luteinizing hormone ratio ≥4 is associated with low egg production in control group and nettle supplementation can balance FSH/LH ratio to ≤2.7. It is concluded that nettle powder could be used as a worthwhile feed additive at the late laying period of aged quails. HIGHLIGHTS Nettle powder can be used as a food additive with aged quails at late laying period. Addition of 1 and 1.5% nettle powder improves egg production, FCR, egg shell thickness. Nettle enhances reproductive performances, such as fertility, hatchability, weight of ovary, and weight of follicles by balancing reproductive hormones at late laying period.

Surface functionalisation-dependent adverse effects of metal nanoparticles and nanoplastics in zebrafish embryos

High throughput imaging is used to assess hatching, lethality and malformations in zebrafish embryos and is suitable for hazard ranking of different nanomaterials.

An investigation of mechanisms underlying mouse blastocyst hatching: a ribonucleic acid sequencing study

To investigate the regulatory mechanisms and signaling molecules underlying hatching in mouse embryos. Experimental laboratory study using a mouse embryo model. University-based basic scientific research laboratory. A total of 40 B6C3F1 × B6D2F1 mouse embryos were used in this study. Frozen/thawed mouse embryos, at the 8-cell stage, were cultured invitro for 2 days. The resulting hatching and prehatching blastocysts were then used for complementary deoxyribonucleic acid (cDNA) library preparation and ribonucleic acid (RNA) sequencing analysis (n = 8 for each group). Differentially expressed genes were then used for downstream functional analysis. In addition, a list of genes related to developmental progression in humans was used to identify genes that were potentially related to the hatching of human embryos. Differentially expressed genes, enriched Gene Ontology terms and canonical pathways, clustered gene networks, activated upstream regulators, and common genes between a gene list of hatching-related genes in mice and a gene list associated with developmental progression in humans. A total 275 differentially expressed genes were identified between hatching and prehatching blastocysts: 230 up-regulated and 45 down-regulated genes. Functional enrichment analysis suggested that blastocyst hatching invitro is an adenosine triphosphate (ATP)-dependent process that involves protein biosynthesis and organization of the cytoskeleton. Furthermore, by regulating cell motility, the RhoA signaling pathway (including Arpc2, Cfl1, Gsn, Pfn1, Tpi1, Grb2, Tmsb10, Enah, and Rnd3 genes) may be a crucial signaling pathway during hatching. We also identified a cluster of genes (Krt8, Krt7, Cldn4, and Aqp3) that exerted functional roles in cell-cell junctions and water homeostasis during hatching. Moreover, some growth factors (angiotensinogen and fibroblast growth factor 2) and endocrine factors (estrogen receptor and prolactin) were predicted to be involved in the regulation of embryo hatching. In addition, we identified 81 potential genes that are potentially involved in the hatching process in human embryos. Our analysis identified potential genes and molecular regulatory pathways involved in the blastocyst hatching process in mice; we also identified genes that may potentially regulate hatching in human embryos. Our findings enhance our knowledge of embryo development and provide useful information for further exploring the mechanisms underlying embryo hatching.

Acute toxicity of methomyl commercial formulation induces morphological and behavioral changes in larval zebrafish (Danio rerio)

The use of pesticides has continue grown over recent years, leading to several environmental and health concerns, such as the contamination of surface and groundwater resources and associated biota, potentially affecting populations that are not primary targets of these complex chemical mixtures. In this work, we investigate lethal and sublethal effects of acute exposure of methomyl commercial formulation in zebrafish embryo and larvae. Methomyl is a broad-spectrum carbamate insecticide and acaricide that acts primarily in acetylcholinesterase inhibition (AChE). Methomyl formulation 96 h-LC50 was determined through the Fish Embryo Acute Toxicity Test (FET) and resulted in 1.2 g/L ± 0.04. Sublethal 6-day exposure was performed in six methomyl formulation concentrations (0.5; 1.0; 2.2; 4.8; 10.6; 23.3 mg/L) to evaluate developmental, physiological, morphological, behavioral, biochemical, and molecular endpoints of zebrafish early-development. Methomyl affected embryo hatching and larva morphology and behavior, especially in higher concentrations; resulting in smaller body and eyes size, failure in swimming bladder inflation, hypolocomotor activity, and concentration-dependent reduction of AChE activity; demonstrating methomyl strong acute toxicity and neurotoxic effect.

In Vivo Evaluation of Cannabis sativa Full Extract on Zebrafish Larvae Development, Locomotion Behavior and Gene Expression.

Historically, humans have been using Cannabis sativa for both recreational and medical purposes. Nowadays, cannabis-based products have gained scientific interest due to their beneficial effects on several syndromes and illnesses. The biological activity of cannabinoids is essentially due to the interaction with the endocannabinoid system, and zebrafish (Danio rerio) is a very well-known and powerful in vivo model for studying such specific interactions. The aim of the study was to investigate the effects of different doses of a Cannabis sativa whole extract [dissolved in dimethyl sulfoxide (DMSO)] on zebrafish eggs’ hatchability, embryo post-hatching survival, larvae locomotion behavior and mRNA gene expression. The results showed the absence of toxicity, and no significant differences were observed between treatments for both embryo hatching and survival rate. In addition, larvae exposed to the cannabis extract at the highest dose [containing 1.73 nM and 22.3 nM of ∆9-tetrahydrocannabinol (THC) and cannabidiol (CBD), respectively] showed an increased locomotion compared to the control and DMSO treated groups. Moreover, qRT-PCR analysis showed that the highest dosage of cannabis induced an over-expression of cnr1 and cnr2 cannabinoid receptors. In conclusion, the exposition of zebrafish larvae to the whole extract of Cannabis sativa showed no negative effects on embryo development and survival and enhanced the larvae’s locomotor performances. These findings may open up possible Cannabis sativa applications in human pharmacology as well as in other animal sectors.

Effects of incubator oxygen and carbon dioxide concentrations on hatchability of fertile eggs, some blood parameters, and histopathological changes of broilers with different parental stock ages in high altitude

The effects of incubator carbon dioxide (CO2) and oxygen (O2) concentrations with parental stock age (PSA) on embryonic deaths (ED), hatchability of fertile eggs (HFE), some blood parameters, and the tissue development of broilers were investigated. Four consecutive repetitions following the similar materials and methods were carried. From 3 different aged ROSS 308 broiler parental flocks 7,680 hatching eggs were obtained and classified as young (Y; 29 wk), middle (M; 37 wk) and old (O; 55 wk) as regards PSA, and randomly distributed. Four different incubator ventilation programs (IVP) as control (C; 0.67% CO2 and 20.33% O2), high CO2 (HC; 1.57% CO2 and 20.26% O2), high O2 (HO; 0.50% CO2 and 21.16% O2), and high CO2 + O2 (HCO; 1.17% CO2 21.03% O2) were applied with oxygen concentrator, and ED and HFE were investigated. Lung and heart tissues, hemoglobin value, packed cell volume, and red blood cell count, triiodothyronine, thyroxine, adrenocorticotropic hormone (ACTH) values of the chicks were analyzed. It was found that IVP affected ED and HFE. Higher rate of early ED (EED) was obtained from the HC than HCO, and higher middle+late stage+pipped but unhatched ED (MLPED) with a lower rate of HFE was observed in the C group than HO and HCO (P < 0.05). Association was found between PSA and IVP (P < 0.05), being more evident in EED for young PSA, in MLPED with HFE for Y and O PSA. From hematological values, no statistical difference in RBC, PCV, and Hb values were found among the treatment groups, ACTH concentration known as a response to stress was found to be higher than C in all groups, triiodothyronine concentration was higher in the HO group than C. In the histopathological examination, used IVPs were found to have negative effects on the lung and heart such as vacuolization, hemorrhage in all PSA groups except for C. Conclusively, PSA and IVP affected some hatching, blood and tissue development parameters of the broiler chicks.

Impact of Treating Hatching Eggs with Curcumin after Exposure to Thermal Stress on Embryonic Development, Hatchability, Physiological Body Reactions, and Hormonal Profiles of Dokki-4 Chickens.

Simple SummaryCurcumin has been used as a suitable feed supplement for poultry to improve several hematological and biochemical indicators, diminish heat stress, and increase antioxidant activity. This experiment evaluated the effects of incubation temperatures and spraying hatching eggs with curcumin during the incubation phase on chick embryo development, hatchability, physiological body reactions, and hormonal profiles of Dokki 4 chickens. The findings indicated that the relative water loss and dead after piping in the chronic incubation temperature group were significantly increased compared with the normal incubation temperature group. Post-hatch, the triiodothyronine level in the high incubation temperature group was significantly decreased than in the control group. Regarding curcumin treatments, relative water loss from eggs and the hatchability of fertile eggs in treated groups significantly increased, while body surface temperature significantly decreased compared with the control. This study evaluated the impact of incubation temperature and spraying hatching eggs with curcumin during the early embryogenesis phase on chick embryo developments, hatchability, physiological body reactions, and hormonal profiles of Dokki 4 chickens. A total of 720 fertile eggs were equally distributed into two groups. In the first group, the eggs were incubated at normal incubation temperature/NIT (37.8 °C and 55–60% RH) for up to 19 days of incubation, whereas those in the second group were incubated in the same conditions except from 6 to 8 day, in which they were daily exposed to chronic incubation temperature/CIT (39.0 °C) for 3 h. Each group was classified into four curcumin treatment doses; the 1st treatment (control) was sprayed with distilled water, while the 2nd, 3rd, and 4th treatments were sprayed with 250, 500, and 1000 mg curcumin/liter distilled water. The results indicated that the lowest hatchability of fertile eggs (%) was obtained in the CIT group (p = 0.02), whereas the highest body surface temperature/BST compared in the NIT group (p = 0.01). Regarding curcumin treatments, the percentages of heart, gizzard, spleen, and T3 hormone levels in the treated group were significantly increased, while the H/L ratio was significantly reduced (p = 0.001) compared with the control. At 8 weeks of age, the testes and ovary percentages in treated groups were significantly (p = 0.05) increased compared with the control. In conclusion, exposure of hatching eggs to high thermal stress (39 °C) during the incubation phase had deleterious effects on chick performance and T3 hormone level. Moreover, spraying hatching eggs had beneficial impacts on growth, reproductive organs, T3 hormone level, and reducing H/L ratio.

In vitro fertilization and hybridization potential of the Indian white shrimp (Penaeus indicus)

Managing the reproduction of shrimp farmed in captivity is essential for selective breeding and interspecies hybridization. Procedures have not been developed for conducting in vitro fertilization in penaeid shrimp. In this study, the feasibility of in vitro fertilization (IVF) of the pre-ovulatory oocyte of Penaeus indicus was examined. Additionally, the processes of fertilization and possibility of hybridizing P. indicus with P. monodon with utilization of IVF procedures was also evaluated. The IVF was conducted by dissecting ovaries with fully developed follicles, and mixing these (~0.1 million oocytes) with 1 mL of a sperm suspension. Evaluations for fertilization and embryonic development occurred every l5 min. The eggs collected from the ovaries had the capacity for activation and fertilization. The hatching rate was 5.5 ± 1.1% of the total number of eggs fertilized, and 8.2 ± 4.8% of the nauplii developed to the post-larval stage. Results from the scanning electron micrograph evaluations provided detailed information about the changes occurring as a result of IVF in P. indicus. Interspecific hybridization of P. indicus with P. monodon did not result in hatching of embryos from the eggs, although 1% of eggs were fertilized. A reliable and inexpensive IVF procedure was developed, therefore, IVF could be an efficacious procedure for facilitation of intraspecific cross production in a penaeid breeding program. Furthermore, it was determined that IVF could be a possible procedure for interspecies hybridization between closely related species to circumvent prezygotic barriers in reproductive processes.

Hatchability and growth performances of normal feathered local, Sasso-RIR and their F1-cross chickens managed under on-station condition in southern Ethiopia.

The Normal feathered local chicken (LL), Sasso-RIR (SRSR) and their F1-cross (LSR) chickens were hatched to evaluate for egg hatchability, body weight, feed efficiency, and survival rate. After 14days of brooding, 150 chicks of each genotype were randomly selected and further replicated into five pens in a deep litter grower house consisting of 30 chicks each in a completely randomized design, and evaluated for a period of 16weeks. Hatchability of fertile eggs was highest for LL (80.0%), intermediate for LSR (68.6%), and lowest for SRSR (55.9%) chickens. The body weight (BW) of chicks at 2weeks of age was 80.0, 76.3, and 61.5g/bird for SRSR, LSR, and LL, respectively, the latter being the lowest (p < 0.05). The respective BW at 8weeks of age was 732, 587, and 451g while at 18weeks it was 1877, 1379, and 1070g/bird and different from each other (p < 0.05). During 3- to 8-week and 9- to 18-week growth periods, the LL chickens were inferior (p < 0.05) in feed intake (29.7 and 66.9g/d/bird) whereas the SRSR chickens were superior (p < 0.05) in body weight gain (15.5 and 16.3g/d/bird) and feed conversion ratio (2.67 and 5.35g feed/ g gain), respectively. The mortality rate of chicken was not affected by genotypes. It can be concluded that Sasso-RIR chicken genotype had played a significant role in upgrading the growth rate and market weight of the local normal feathered chicken without adverse effect on hatchability, feed efficiency, and survival rate.

Empty Zona Pellucida Only Case: A Critical Review of the Literature.

The presence of empty zona pellucida (EZP) in oocytes following oocyte retrieval (OR) during an in vitro fertilization (IVF) cycle presents a major clinical and laboratory challenge in assisted reproduction. It has been attributed to several factors such as the ovarian stimulation protocol employed, the damaging of the follicles during oocyte retrieval (OR) mainly through the high aspiration pressure, during the denudation technique, and the degeneration of oolemma within the zona pellucida (ZP) through apoptosis. The role of ZP is pivotal from the early stages of follicular development up to the preimplantation embryo development and embryo hatching. Polymorphisms or alterations on the genes that encode ZP proteins may contribute to EZP. We present a critical review of the published literature hitherto on EZP and available options when encountered with the phenomenon of EZP. Concerning the former, we found that there is rare data on this phenomenon that merits documentation. The latter includes technical, genetic, and pathophysiological perspectives, along with specific treatment options. In conclusion, we identify the lack of a definitive management proposal for couples presenting with this phenomenon, we underline the need for an algorithm, and indicate the questions raised that point towards our goal for a strategy when addressing a previous finding of EZP.

Chloroperoxidase-catalyzed oxidative degradation of sulfur mustard

As a natural heme protein catalyzing the oxidation of sulfides to sulfoxides without sulfone formation, chloroperoxidase (CPO) is well suited for the degradation of sulfur mustard (HD), a persistent chemical warfare agent that has been widely disposed since World War II and continuously leaks into aquatic environments. Herein, we report the first systematic investigation of CPO-catalyzed degradation of HD and the potential application of CPO in destroying chemical weapons under mild conditions. The related Michaelis–Menten parameters (Km=0.17 mM, Vmax=0.06 mM s−1 (R2 =0.935), and kcat= 2717 s−1) indicated nearly a prominent enzymatic efficiency. Under optimal conditions, 80% of HD was transformed to bis(2-chloroethyl) sulfoxide as identified by mass spectroscopy and nuclear magnetic resonance (NMR) spectroscopy. Other metabolites were also generated during the decontamination process. A plausible oxidation mechanism was proposed based on the degradation products, NMR titration experiments, and molecular dynamics simulations. CPO also promoted the degradation of other chemical weapon agents, namely, Lewisite (L) and venomous agent X (VX), thereby exhibiting a broad substrate scope. The high potential of the developed system for the decontamination of aquatic environments was demonstrated by the successful hatching of zebrafish embryos after HD degradation and the survival of zebrafish (Danio rerio, AB strain) larvae after the degradation of Agent Yellow (L+HD).

Transformation of copper oxide nanoparticles as affected by ionic strength and its effects on the toxicity and bioaccumulation of copper in zebrafish embryo

This study aimed to investigate the transformation of copper oxide nanoparticles (CuO NPs) in aquatic environments under different ionic strength and further examine its effects on copper toxicity and bioaccumulation by monitoring the responses and uptake behaviours of zebrafish embryo. Ionic strength (IS) was simulated according to surface water (1.5 mM), groundwater (15 mM), and wastewater (54 mM), representing low-, mid-, and high-IS water, respectively. At the highest exposure of 10 mg CuO/L, zebrafish larvae mortality was increased from 21.3% to 33.3%, when IS decreased from 54 to 1.5 mM. Low-IS solution also caused the highest numbers of delayed hatching embryo (81.3%) and opaque yolk deformation (36.3%). Copper bioaccumulation markedly increased when larvae were exposed to low-IS water (35%) relative to high-IS water (15%). Exposing to low-IS particularly enhanced copper uptake (~15 ng Cu/g inside embryo), facilitating the copper accumulation in the heart of larvae, whereas aggregated CuO NPs (>500 nm) in mid- and high-IS water were blocked from the embryo and found abundantly in the body axis and tail. Results indicate that CuO NPs in low-IS solutions rapidly form the relatively small CuO NP aggregates with a high copper dissolution, which would pose great concern for aquatic organisms.

Toxicity of boron and vanadium nanoparticles on Danio rerio embryos - Phenotypical, biochemical, and behavioral alterations.

Engineered nanoparticles (NPs) are emerging contaminants of concern and it is important to understand their environmental behavior and ecological risks to exposed organisms. Despite their ubiquitous presence in the environment, there is little information about the hazards of certain NPs, such as boron (BNPs) and vanadium (VNPs). The aim of the present research was to investigate the effects of commercial BNPs and VNPs (80 to 100 nm) to zebrafish embryos, at different levels of biological organization. A range of nominal concentrations for both NPs (0, 0.01, 0.1, 1, and 10mg/L) was tested. Due to the presence of triton X-100 in the NPs' stock dispersions, an additional control group was included (0.001% triton X-100). Survival, hatching, and malformations of embryos were assessed for 96 hours (h) exposure. Locomotor behavior was evaluated at 120 h. Furthermore, embryos were exposed to 0, 1, and 10 mg/L of NPs to evaluate a set of biomarker responses after 96 h: cholinesterase (ChE) and glutathione S-transferase (GST) activities, total glutathione (TG) and energy budgets levels. VNPs induced malformations (10 mg/L), hyperactivity (10 mg/L), erratic swimming (0.01 mg/L), altered swimming pattern (>0.01 mg/L), delayed hatching (10 mg/L) and altered biochemical responses involved in antioxidant defense (GST and TG at >1 mg/L), neurotransmission (ChE at 10 mg/L) and energy metabolism (lipids at >1 mg/L and carbohydrates at 10 mg/L). BNPs caused malformations (10 mg/L), affected swimming pattern (>0.01 mg/L), induced erratic swimming (10 mg/L) and decreased TG content and GST activity (>1 mg/L). At the same concentrations, VNPs affected a greater number of endpoints than BNPs, demonstrating a greater toxicity to zebrafish embryos. The present study shows that BNPs and VNPs may affect aquatic organisms, albeit at relatively great non-environmentally relevant concentrations, reinforcing the importance of the risk assessment of different NPs.

CRISPR/Cas9-based functional characterization of the pigmentation gene ebony in Plutella xylostella.

Body pigmentation is an important character of insects in adapting to biotic and abiotic environmental challenges. Additionally, based on the relative ease of screening, several genes involved in insect melanization have been used in classic genetic studies or as visual markers in constructing transgenic insects. Here, a homologue of the Bombyx mori melanization-inhibiting gene ebony, associated with the conversion of dopamine to N-β-alanyl dopamine, was identified in a global pest, Plutella xylostella. The CRISPR/Cas9 system was applied to generate multiple Pxebony knockout alleles which were crossed to produce a Pxebony knockout strain, showing darker pigmentation in larvae, pupae and adults, compared with wildtype. Interestingly, we observed that Pxebony heterozygotes displayed an intermediate darkened phenotype, indicating partial dominance between the knockout and wildtype alleles. The fitness costs of Pxebony deficiency were also assessed in the mutant strain, indicating that embryo hatchability and larval survival were significantly reduced, while the eclosion rate was not obviously affected. Our work provides a potential target for exploring CRISPR-based genetics-control systems in this economically important pest lepidopteran.

Effects of ventilation programme and eggshell thickness on hatchability rate and hatching time of broiler eggs

The aim of the research was to determine whether enrichment of the atmosphere in an incubator with carbon dioxide (CO2) and oxygen (O2) and eggshell thickness (EST) affected embryonic death (ED), hatchability of fertile eggs (HFE) and hatching time (HT). A total of 320 Ross 308 eggs were used and the experiment was repeated twice. Eggshell thickness was classified as thin (&lt;31 μm), medium (31 - 32 μm) and thick (&gt;32 μm). The incubators were operated with their internal atmosphere enriched with CO2 (1.57% CO2; 20.23% O2) or O2 (0.50% CO2; 22.44% O2). Embryonic death, HFE and HT data were monitored at three periods, namely early (&lt;486 hours), middle (486 - 492 hours), and late (492 - 510 hours). Early ED, late ED and hatchability of fertile eggs were not affected by EST or by the incubator’s internal atmosphere (P &gt;0.05). Thus, O2 supplementation to the incubator was deemed unnecessary at 822 m altitude. There was a highly significant interaction between EST and HT. Eggs with shells 31 to 32 μm thick hatched at an appreciably greater rate between 486 and 492 hours of incubation (17%) than eggs with thicker (0.6%) or thinner (0.4%) shells in both the CO2 and O2 enriched atmospheres. The hatching rate was significantly higher in the eggs with an intermediate EST than in thick-shelled eggs. A greater proportion of eggs hatched at the late HT as opposed to earlier, regardless of EST.

Site specificity of blastocyst hatching significantly influences pregnancy outcomes in mice.

Blastocysts hatch from the zona pellucida (ZP) to enable implantation into the uterine endometrial epithelium, but little is known regarding the effect of hatching sites on pregnancy outcomes. Murine hatching embryos were categorized into five groups based on initial trophectoderm projection (TEP)/ZP position corresponding to the inner cell mass center. In blastocysts (3.5 dpc) post-12hours in vitro culture, TEP rates of A-site (44.4%) and B-site (38.6%) embryos were higher than those of C-site (12.5%) and D-site (3.1%) embryos, while the O-site (1.4%) was the lowest (P<.05). Post-ET A-site (55.6%) and B-site (65.6%) birth rates were higher than those of C-site embryos (21.3%) and controls (P<.05). Furthermore, live birth rate of B-site embryos remained higher than C-site embryos (68.8% vs 31.3%; P<.05) when both were transferred into the same recipients. Different TEP site blastocysts exhibited different implantation competences: the implantation rate of C-site embryos was lower than that of both A- and B-site groups (67.7% vs 84.3% and 83.2%, respectively; P<.05) at 2days post-ET. C-site embryos also had a distinctly higher ratio of developmental defects (47.5%) than A- and B-site embryos (22.5% and 14.6%, respectively), with implantation failure mainly associated with poor birth rate, a finding corroborated by differential gene expression analysis such as LIF, LIFR, and S100a9. Surprisingly, acidified Tyrode's solution (AAH)-treated B-site blastocysts had a significantly increased birth rate (77.1%) than C-site (55.3%) and controls (43.4%). Site specificity and differential gene expression during embryo hatching can be applied in ART screening. More importantly, assisted hatching by AAH is effective and feasible for improving pregnancy and term development, particularly at the B-site, for humans and in animal husbandry.

P–182 The mechanism of mouse embryo hatching and the impact of laser drilling the zona pellucida: an RNA sequencing study

Abstract Study question What is the mechanism of embryo hatching? Will laser-assisted zona pellucida (ZP) drilling alter the embryonic transcriptome? Summary answer Hatching is an ATP-dependent process. Hatching is also associated with Rho-mediated signaling. Laser-assisted ZP drilling might cause alternation in embryo metabolism. What is known already Embryo hatching is a vital process for early embryo development and implantation. Animal data suggests that hatching is the result of multiple factors, such as mechanical pressure, protease activation, and the regulation of maternal secretions. However, little is known about the regulatory signaling mechanisms and the molecules involved. In addition, despite the extensive use of laser-assisted ZP drilling in the clinic, the safety profile of this technique at molecular level is very sparse. The impact of this technique on the embryonic transcriptome has not been studied systematically. Study design, size, duration Eighty mouse embryos were randomly divided into a laser ZP drilling group (n = 40) and an untreated group (n = 40). After treatment, embryos were cultured in vitro for two days. Then, hatching blastocyst (n = 8) and pre-hatching blastocyst (n = 8) from the untreated group, and the hatching blastocyst from the treatment group (n = 8) were processed for RNA sequencing (RNA-seq). Participants/materials, setting, methods Cryopreserved 8-cell stage mouse embryos (B6C3F1 × B6D2F1) were thawed, and a laser was used to drill the embryo ZP in the treatment group. Next, the treated and untreated embryos were individually cultured in vitro to the E4.5 blastocyst stage. The resulting blastocysts were lysed individually and used for subsequent cDNA library preparation and RNA-seq. Following data quality control and alignment, the RNA-seq data were processed for differentially expressed gene analysis and downstream functional analysis. Main results and the role of chance According to the RNA-seq data, 275 differentially expressed genes (DEGs) (230 up-regulated and 45 down-regulated, adjusted P &amp;lt; 0.05) were identified when comparing hatching and pre-hatching blastocysts in the control groups. Analysis suggested that the trophectoderm is the primary cell type involved in hatching, and revealed the potential molecules causing increased blastocyst hydrostatic pressure (Aqp3 and Cldn4). Functional enrichment analysis suggested that ATP metabolism and protein synthesis were activated in hatching blastocysts. DEGs were found to be significantly enriched in several gene ontology terms, particularly in terms of the organization of the cytoskeleton and actin polymerisation (P &amp;lt; 0.0001). Furthermore, according to QIAGEN ingenuity pathway analysis results, Rho signaling was implicated in blastocyst hatching (Actb, Arpc2, Cfl1, Myl6, Pfn1, Rnd3, Septin9, z-score=2.65, P &amp;lt; 0.0001). Moreover, the potential role of hormones (estrogen (z-score=2.24) and prolactin (z-score=2.4)) and growth factors (AGT (z-score=2.41) and FGF2 (z-score=2.213)) were implicated in the hatching process as indicated by the upstream regulator analysis. By comparing the transcriptome between laser-treated and untreated hatching blastocysts, 47 DEGs were identified (adjusted P &amp;lt; 0.05) following laser-assisted ZP drilling. These genes were enriched in metabolism-related pathways (P &amp;lt; 0.05), including the lipid metabolism pathway (Mvd, Mvk, Aacs, Gsk3a, Pik3c2a, Aldh9a1) and the xenobiotic metabolism pathway (Aldh18a1, Aldh9a1, Keap1, and Pik3c2a). Limitations, reasons for caution Findings in mouse embryos may not be fully representative of human embryos. Furthermore, the mechanism of hatching revealed here might only reflect the hatching process of embryos in vitro. Further studies are now necessary to confirm these findings in different conditions and species to determine their clinical significance. Wider implications of the findings: Our study profiled the mouse embryo transcriptome during in vitro hatching, identified potential key genes and mechanisms for future study. In addition, for the first time, we revealed the impact of laser-assisted ZP drilling on the transcriptome, this may help us to assess and improve the existing technique. Trial registration number Not applicable

Visual system development and changes in hatching performance in hybrid grouper embryos under different light conditions

Light greatly impacts the performance of teleost embryos. In this study, the development of visual system and performance of Epinehelus moara × Epinephelus lanceolatus embryos under different light spectra (red, orange, green, blue and full spectrum) and dark conditions were assessed. The development of visual system was not completed during embryonic development, characterized by the absence of structural integrity in the retina, indicating the absence of visual photosensitivity. However, nonvisual system may play an important role in photosensitivity. Light conditions contributed significantly to embryo performance, including hatching rate, deformity rate and Wnt signaling pathway expression. There was no statistical difference in the oxygen consumption rate of embryos in different groups during heartbeat onset stage. Embryos in dark group had a significantly lower hatching rate than other groups. The hatching rate in full light spectrum group was significantly higher than red spectrum group, and no significant differences were observed among the other groups. The deformity rate of newly hatched larvae exposed to orange light was significantly lower than those under full spectrum and dark conditions. The mRNA expression level of frizzled2, dickkopf2, limb bud and heart homolog, and natural kermes dye inhibitor of Wnt signaling pathway 1 were detected. The results suggested that dark conditions could lead to the abnormal expression of the Wnt signaling pathway, characterized by significantly lower mRNA expression levels of these factors, which in turn may cause decreased hatchability. In order to increase the hatchability and performance of embryos, and reduce the deformity rate, orange light should be utilized and dark conditions should be avoided during the embryonic development of E. moara × E. lanceolatus.

Comparative study of hatchability rate and egg quality between different strains of Japanese Quail (Coturnix japonica)

The objectives of this study were to determine morphology of Japanese quail strains andhatchability rate and egg quality between commercial strain which is Poultry Progress Institute(IKTA) strain and White Texas strains of Japanese Quail (Coturnix japonica) rear in Malaysia. Atotal of 300 quails were randomly picked and the breastbone length, chest girth, body weight,body length, shank length and drum length were measured for the morphology. Then, 270 eggs ofCommercial and White Texas strains were used to test the hatchability rate (n=150) and eggquality (n=120). The ratio of the parents of the quails between male and female of each type ofstrain were 1:3. The cleanliness, shape, weight, and texture of the eggshell were observed,weighed and measured. The eggs were incubated for 17 days (37.6 °C, 65% humidity). The shapeindex, egg weight, egg volume and eggshell surface were weighed and measured for the exteriortrait of eggs. For interior trait, the albumen index, Haugh units, and internal quality units (IQU)were measured. Significant findings were observed in all of the features (P&lt;0.05), except for thewing length in morphology. For hatchability rate, there was a high significant in the percentage ofhatchability of incubated eggs, hatchability of fertile eggs and smoothness texture of the eggshell.In egg quality there was high significant in Haugh unit but low significant internal quality unit.The significant variability might be due from the genetic information that inherited from theparents. The IKTA and White strains were not the same in morphology, hatchability rate andinterior trait of eggs but same exterior trait of eggs. In conclusion, this research is important ingiving information about Japanese quail strain in Malaysia and more genetic study should be donein the future to determine good strain and increase understanding about quail strain.