二化性家蚕卵低温和常温催青的胚胎期蛋白质表达差异分析

家蚕二化性品种的滞育性受上代胚胎期环境条件调控,查找家蚕胚胎期滞育关联蛋白,可为最终阐明家蚕滞育的分子机制提供实验依据。以家蚕二化性品种秋丰的蚕卵为材料,分别在25℃常温和18℃低温条件下催青,提取胚胎不同发育时期蚕卵的易溶性和难溶性蛋白,采用蛋白质双向电泳(2-DE)和图像分析技术,研究蚕卵在胚胎不同发育时期的蛋白质差异表达谱。在易溶性和难溶性蛋白图谱中分别检测到5个和7个差异显著的蛋白点。对这些差异蛋白点进行质谱鉴定,有8个蛋白点得到可信的最佳匹配蛋白报告,共鉴定出卵特异蛋白、卵黄原蛋白、表皮蛋白和丙酮酸脱氢酶激酶4种蛋白。这些差异表达蛋白可能导致蚕卵胚胎中物质代谢和能量代谢的不同,据此推测低温和常温催青可能在蚕卵胚胎中诱导了不同的生理生化反应。     

Chromosome analysis of early embryonic mortality in layer and broiler chickens

1. Twenty-three lines of chickens, obtained from grandparent stocks of 4 Australian breeding companies, were analysed to determine the incidence of early embryonic mortality attributable to chromosome abnormalities. The lines included 10 layer strains, consisting of 6 White Leghorn, 2 New Hampshire and 2 Australorp lines, and 13 broiler lines. 2. A total of 10,730 eggs was examined after 3 d incubation; of these 9746 (90.8%) were fertile. Abortive embryonic development was observed in 1379 (14.1%) of the fertile eggs. This consisted of 952 (69.0%) dead and dying embryos, including 646 malformed and 427 (31.0%) membranes without embryos. 3. Early embryonic mortality was found to vary from 9.8 to 26.8% (average 16.4%) in broiler lines and from 8.0 to 27.9% (average 11.9%) in layer lines. 4. Among 898 abortive embryos analysed, 112 had abnormal chromosomes consisting of 27 haploids, 38 haploid-euploids, 24 triploids, 16 diploid-polyploids, 4 aneuploids, 2 tetraploids and 1 translocation. 5. In broilers and layers respectively, chromosome abnormalities were responsible for 4.4 to 28.1% (average 11.8%) and 7.4 to 25.0% (average 13.4%) of the early embryonic mortality. 6. The overall frequency of chromosome abnormalities in all fertile eggs varied from 0.7 to 3.7% for the broiler lines and 0.7 to 3.4% for the layer lines.     

Nondestructive sex-specific monitoring of early embryonic development rate in white layer chicken eggs using visible light transmission

ABSTRACT

1. Sex-specific variations in early embryonic development rates may pre-empt later variations in embryonic development through to pipping and hatching. Given that erythropoiesis (blood production) can be equated with early embryonic growth rate, it was hypothesised that blood pigment haemoglobin can act as a specific spectral fingerprint for changes in growth rate. Moreover, by measuring longitudinal, rather than lateral, spectral transmission through the egg, a more consistent spectrum with a higher signal-to-noise ratio could be captured.

2. Longitudinal visible transmission (T575/T598 ratio), which is sensitive to haemoglobin, was used to monitor sex-specific early embryonic development rate in white layer chicken eggs from d 0 to 8 of incubation. The sex of these eggs was subsequently confirmed two days after hatching.

3. Embryonic development was detectable from d 3 (72 h) of incubation, 36 h earlier than previously reported lateral spectral measurements, supporting the greater sensitivity of longitudinal measurements.

4. At d 3, the mean T575/T598 ratio for male embryos was significantly lower (P < 0.001) (i.e. higher absorbance of haemoglobin) than for female embryos, which was thought to be due to sex-differences in early embryogenesis. On the other hand, female embryos had a significantly lower (P < 0.05) mean T575/T598 ratio than male embryos at d 7 of incubation, presumably due to the combined effects of oestrogen synthesis receptors and enzymes on erythropoiesis in female embryos at this time.

5. In conclusion, the proposed methodology has the sensitivity to differentiate sex-specific embryonic development rates during early incubation and the potentiality to advance precision incubation management and poultry research.     

家蚕性连锁平衡致死系雌性胚胎的组织形态学初步研究

为了调查家蚕性连锁平衡致死系中Z染色体携带致死基因l2的雌性胚子(简称l2胚子)致死的时间和原因,收集家蚕性连锁平衡致死系品种平2不同胚胎发育时期的雌性黑卵,解剖获取胚胎后用光学显微镜对不同发育时期的l2胚子进行组织形态学观察。形态观察可见l2胚子在胚胎发育早期和能正常发育的雌性胚子没有区别,但l2胚子发育至晚期的己2期,约有一半胚子的体表未能正常着色,因此推断那些仍没有正常着色的胚子即为致死的l2胚子。通过胚胎的半薄切片观察发现,己2期l2胚子的组织器官结构完整,与正常发育的雌性胚子之间无明显的差异。观察结果提示:l2胚子死亡可能不是由于胚胎组织结构异常造成的,有可能是在己2期胚子停滞发育致死。     

Heart rate in developing ostrich embryos

1. A non-invasive condenser microphone was used to detect cardiogenic, acoustic pressure changes (acoustocardiogram, ACG) over the eggshell in order to determine embryonic heart rate (HR) of ostriches in a commercial hatchery. 2. HR measured for 36 eggs at 36.3C was maintained at about 185 bpm during the middle stage of development (days 19 to 23) and then decreased with embryonic development. 3. The daily changes in HR were not related to egg mass, but HR of high water vapour conductance (GspH2O) eggs was found to decrease less during the last stages of incubation relative to low and medium GspH2O groups. 4. The averaged HR at 80% of incubation period was close to the value predicted from the allometric equation determined previously for embryos of domesticated precocial birds.     

L-Leucine In Ovo Administration Causes Growth Retardation and Modifies Specific Amino Acid Metabolism in Broiler Embryos

L-Leucine (L-Leu) in ovo administration was demonstrated to afford thermotolerance and modified amino acids metabolism in post-hatched broiler chicks under heat stress. This study aimed to investigate the changes in embryonic growth and amino acid metabolism after in ovo injection of L-Leu. Fertilized broiler eggs were subjected to in ovo injection of sterile water or L-Leu on embryonic day (ED) 7. The weight of embryos and yolk sacs were measured on ED 12, 14, 16, and 18. Plasma and livers were collected on ED 14 and 18 for free amino acid analysis. The weight and relative weight of embryos were significantly lowered by in ovo administration of L-Leu, but those of yolk sacs were not altered. Moreover, L-Leu in ovo injection significantly reduced the plasma proline concentration during embryogenesis and increased the plasma concentrations of tyrosine (Tyr) and lysine (Lys) in ED 18. Hepatic Lys concentration was also significantly increased by L-Leu in ovo injection. Interestingly, Leu concentrations in the plasma and liver were not affected by L-Leu administration. These results indicated that in ovo administered L-Leu was metabolized before ED 14 and affected embryonic growth and amino acid metabolism during embryogenesis.     

ICSI受精卵胞质注射生产转基因猪胚胎的初步研究

试验旨在探讨单精子胞浆内注射（intracytoplasmic sperm injection,ICSI）法生产猪体外受精卵和应用猪ICSI受精卵进行胞质注射生产转基因胚胎的可行性。首先对比了猪体外受精（in vitro fertilization,IVF）受精卵与ICSI受精卵的胚胎发育效率;然后观察了猪ICSI受精卵的双原核形成时间及效率,对精子注射到胞质后6~18 h分6个时间段进行地衣红染色,对比精子进入卵胞质后的状态及原核形成;最后对猪IVF受精卵受精后8~10 h及ICSI受精卵受精后12~14 h进行EGFP-N1质粒（20 ng/μL）胞质注射,观察胚胎发育效率及转基因效率。结果表明,ICSI受精卵的胚胎发育率（卵裂率89.4%和67.9%、囊胚率36.5%和16.1%）显著优于IVF组（P<0.05）,适合用于猪的体外受精卵试验;猪ICSI受精卵双原核在精子注射到卵胞质后12~14 h形成,双原核形成率为54.90%,显著高于其余5个试验组（P<0.05）;ICSI受精卵胞质注射组胚胎卵裂率（86.2%和66.3%）、囊胚率（30.0%和13.6%）及转基因效率（18.5%和0）均显著高于IVF受精卵胞质注射试验组（P<0.05）。本试验结果为采用ICSI受精卵进行胞质注射生产转基因猪的研究奠定了基础。     

Time-dependent recovery of Mycoplasma lipofaciens (strain ML64) from incubated infertile chicken eggs and dead in shell chicken embryos

Mycoplasmas are pathogens of different avian species, and they are able to be vertically transmitted. Even detected, Mycoplasma prevalence in raptor eggs is very low. In contrast to poultry, raptor eggs submitted for investigations are usually incubated. To investigate the influence of incubation length on the recovery of mycoplasmas from eggs, infertile specific-pathogen-free chicken eggs and embryos were infected with Mycoplasma lipofaciens (strain ML64), which had previously been isolated from an egg of a northern goshawk (Accipiter gentilis), in two different dosages. The eggs were investigated up to 12 days after infection (infertile eggs) or embryonic death. Mycoplasmas were recovered over the entire period after embryonic death by isolation. It was possible to re-isolate M. lipofaciens (strain ML64) from infertile eggs infected with 10(6) colony-forming units (CFUs) up to 12 days, but only up to 7 days if infected with 10(2) CFUs, which may be closer to the situation after natural infection. This study demonstrates that incubation of infertile eggs does have an influence on the recovery rate of mycoplasmas. This influence must be considered if interpreting results of Mycoplasma investigations in eggs of nonpoultry species. Additionally, it is recommended to use dead in shell embryos rather than infertile eggs for Mycoplasma detection.     

Detection of early embryonic development in chicken eggs using visible light transmission

1. In two separate experiments, the possibility of detecting embryonic development in chicken eggs was assessed using the same spectrophotometric method used to detect blood in Table eggs, using a combination of two wavelengths (577 and 610 nm) of the transmission spectrum. 2. In the first experiment, during the first 10 d of incubation, transmission spectra of 30 Hisex White eggs and 30 Hybro eggs were measured daily. 3. In the second experiment, 292 Hisex White eggs were incubated. Seven groups were randomly assigned. Six received an injection of sodium azide (NaN3) at different times during incubation in order to stop embryonic development, and during the first 12 d of incubation the transmission spectrum was measured daily. The acoustic resonance analysis method was also used on a group of uninjected eggs. 4. In the first experiment, it was possible to detect embryonic development from 120 h of incubation onwards in fertile eggs. In the second experiment changes in light transmission due to embryonic development were detected from 108 h of incubation. Detection of embryonic development using the acoustic resonance analysis method in the second experiment was possible only from 120 h of incubation. 5. It was concluded that the detection of embryonic development using visible light transmission is not directly linked with the formation of blood, but with the formation of sub-embryonic fluid, which takes place from 72 h of incubation onwards. This fluid makes the yolk sac translucent so that absorption of light at 577 nm can be detected.     

Early development of artificially spawned southern mullet,Liza richardsonii (Smith, 1846)

The embryonic development of artificially spawned southern mullet, Liza richardsonii, eggs and the development of the larvae to 46 days are described and illustrated using drawings and photographs. The floating eggs hatched in sea water at 18–24°C after 46–60 h. Newly stripped eggs usually had more than one oil droplet (up to 16) which merged during development. Free embryos averaged 2,36 mm at hatching and had distinct yellow pigment.     

激光照射对肉鸡鸡胚孵化的影响

采用不同照射时间和位置在孵化前用氦氖激光照射鸡蛋，对早期胚胎发育和减少死亡率有促进作用。试验组死亡率为4.13％～7.32％，对照组死亡率为8.13％。激光处理的效应取决于照射的时间和位置。结果证明，20秒垂直照射的种蛋孵化率最高，比对照组高4个百分点。.     

Reduced egg shell permeability affects embryonic development and hatchling traits in Lycodon rufozonatum and Pelodiscus sinensis

The response of embryos to unpredictable hypoxia is critical for successful embryonic development, yet there remain significant gaps in our understanding of such responses in reptiles with different types of egg shell. We experimentally generated external regional hypoxia by sealing either the upper half or bottom half of the surface area of eggs in 2 species of reptiles (snake [Lycodon rufozonatum] with parchment egg shell and Chinese soft‐shelled turtle [Pelodiscus sinensis] with rigid egg shell), then monitored the growth pattern of the opaque white patch in turtle eggs (a membrane that attaches the embryo to the egg shell and plays an important role in gas exchange), the embryonic heart rate, the developmental rate and the hatchling traits in turtle and snake eggs in response to external regional hypoxia. The snake embryos from the hypoxia treatments facultatively increased their heart rate during incubation, and turtle embryos from the upper‐half hypoxia treatment enhanced their growth of the opaque white patch. Furthermore, the incubation period and hatching success of embryos were not affected by the hypoxia treatment in these 2 species. External regional hypoxia significantly affected embryonic yolk utilization and offspring size in the snake and turtle. Compared to sham controls, embryos from the upper‐half hypoxia treatment used less energy from yolk and, therefore, developed into smaller hatchlings, but embryos from the bottom‐half hypoxia treatment did not.     

家蚕的半活化胚现象初探

滞育是家蚕的生理现象 ,家蚕滞育卵的活化处理是养蚕生产上重要的技术操作。研究了家蚕盐酸处理卵、复式冷藏卵和长期保存卵的胚胎活化过程 ,认为家蚕胚胎除滞育态和活化态以外 ,还存在半活化状态。半活化胚胎的形态与萌动期 (乙 1)至反转期 (己 1)的活化胚相似 ,但胚胎整体发育不平衡 ,后部发育慢于前部 ,在 2 5℃、RH 75 %～ 85 %催青环境中的发育速度明显慢于活化胚 ,而越接近反转期发育速度越接近活化胚。同一群体内出现的半活化胚胎之间发育比较整齐 ,而不同群体的半活化胚胎之间发育有一定差异。活化刺激量越大 ,出现的半活化胚胎发育越快。探讨了半活化胚胎产生的机理以及对蚕种生产的影响     

Effect of lighted incubation on embryonic growth and hatchability performance of two strains of layer breeder eggs

1. Eggs from two layer-type breeder flocks (ISA-W vs Leghorn) between 30 and 45 weeks of age were used in 4 trials to study the effects of lighted incubation on embryonic growth from 5 to 18 d of age and hatchability per cent (HP). The physical dimensions, eggshell characteristics and conductance (EC) of eggs of the two strains were compared. 2. Eggs were set in an incubator on trays either in the dark-control or under two tubes of 20-watt white fluorescent light during the first 18 d of incubation. The light intensity ranged from 1230 to 1790 lux at the surface of the eggs. Eggs were transferred to dark hatching compartments at d 19 of incubation. 3. The genetic make-up of birds influenced the physical dimensions and eggshell characteristics of eggs. ISA-W eggs had higher weight, surface area (ESA), volume (EV), width, EC, shell volume and HP, and lower ESA:EV ratio, per cent shell, shell density and dead embryos than those of Leghorn eggs. 4. Lighted incubation increased daily embryonic growth (mg/d) and HP by 3.9 and 5.9%, respectively, when compared with the dark-control incubation. 5. The physical dimensions and eggshell characteristics of eggs influenced the effects of lighted incubation on embryonic growth and HP. Lighted incubation increased embryonic growth and HP of ISA-W eggs. The increase in embryonic growth and HP of the Leghorn eggs was not significant. 6. The genetic make-up of birds influenced the physical dimensions and eggshell characteristics of their eggs and these differences in the characteristics of hatching eggs influenced embryonic growth and HP when incubated under light.     

数种驱虫药和化学药物对体内外鸡蛔虫产卵和虫卵发育的影响

为鉴定男性不育药和有些驱虫药对寄生蠕虫的不育作用,对培养在试管内的鸡蛔虫和卵胚,以及人工感染蛔虫的鸡分别进行了体内和体外试验。棉酚和硝基呋喃类的呋喃唑酮、呋喃西林、S_(73031)和S_(72055)均无抑制产卵和卵胚发育的作用,苯并咪唑类的丙硫咪唑、甲苯咪唑和噻苯咪唑以及吩噻嗪均有抑制产卵和卵胚发育的作用,它们的EMIC_(50)分别为0.006,1.326,0.029和0.052ppm。抑制产卵所需浓度远大于抑制卵胚发育所需浓度。吡喹酮和硫双二氯酚也不具有抑制产卵和卵胚成熟的作用。放线菌酮仅对试管内蛔虫有抑制产卵作用,但对体内蛔虫却无影响。     

胚胎母源-合子转换期中的表观遗传调控

哺乳动物个体由终端分化的单倍体精子和卵子受精融合成的双倍体受精卵发育而来。在胚胎发育的初始阶段,合子基因组处于休眠状态,胚胎发育调控由卵母细胞内母源调控逐渐转换为合子基因组调控(Maternal-Zygotic Transition,MZT)。在此期间,随着母源物质的清除,合子基因组激活(Zygotic Genome Activation,ZGA),但调控MZT的具体分子机制还不清楚。最新研究表明,DNA甲基化、染色质重塑、组蛋白表观修饰、ncRNA在MZT和ZGA中发挥重要的作用。本文总结了上述4种表观修饰在动物植入前胚胎MZT中的生物学功能和特异的分子机制,对揭示动物胚胎MZT的调控机理有借鉴意义。     

Based serum metabolomics analysis reveals simultaneous interconnecting changes during chicken embryonic development

Metabolic disorder is a major health problem and is associated with a number of metabolic diseases. Due to native hyperglycaemia and resistance to exogenous insulin, chickens as a model had used in the studies of adipose tissue biology, metabolism and obesity. But no detailed information is available about the comprehensive changes of serum metabolites at different stages of chicken embryonic development. This study employed LC/MS‐QTOF to determine the changes of major functional metabolites at incubation day 14 (E14d), 19 (E19d) and hatching day 1 (H1d), and the associated pathways of differential metabolites during chicken embryonic development were analysed using Metabolite Set Enrichment Analysis method. Results showed that 39 metabolites were significantly changed from E14d to E19d and 68 metabolites were significantly altered from E19d to H1d in chicken embryos. Protein synthesis was promoted by increasing the concentrations of L‐glutamine and threonine, and gonadal development was promoted through increasing oestrone content from E14d to E19d in chicken embryos, which indicated that serum glutamine, threonine and oestrone contents may be considered as the candidate indicators for assessment of early embryonic development. 2‐oxoglutaric acid mainly contributed to enhancing the citric cycle, and it plays an important role in improving the growth of chicken embryos at the late development; the decreasing of L‐glutamine, L‐isoleucine and L‐leucine contents from E19d to H1d in chicken embryonic development implied their possible functions as the feed additive during early posthatch period of broiler chickens to satisfy the growth. These results provided insights into understand the roles of serum metabolites at different developmental stages of chicken embryos, it also provides available information for chicken as a model to study metabolic disease or human obesity.     

In ovo injection of branched‐chain amino acids: Embryonic development,hatchability and hatching quality of turkey poults

In this study, the influence of a branched‐chain amino acid blend (BCAA composed of 3 l ‐leucine:1 l ‐valine:2 l ‐isoleucine) injected into the amniotic fluid was evaluated for embryonic growth, yolk‐sac (YS) utilization and development of gastrointestinal tract (GIT) and skeletal muscles of turkey embryos from day 24 of incubation (24E) to hatching, together with hatchability, poult quality and liver L* (lightness), a* (redness) and b* (yellowness) values at hatch. At day 22 of incubation, embryonated eggs (n = 240) were assigned to three treatments, that is, eggs were not injected (control, NC) or injected with 1.5 ml sterile solution with 0.9% salt (SA) or 0.2% BCAA blend (BCAAb). These solutions were injected manually into the amniotic fluid of the embryonated eggs. To determine weights and lengths (where appropriate) of the studied organs and tissues, four embryonated eggs and poults per treatment were selected at 24E and at hatch. While the BCAAb decreased the YS and embryo weight, hatchability and the liver L* value, it increased the weight and quality of poults and the weights of breast and thigh muscles at hatch. In conclusion, the in ovo feeding of the BCAA blend negatively affected hatchability but positively affected hatching weight and poult quality by improving development of skeletal muscles and by regulating energy metabolism.     

Comparison of a modern broiler and layer strain during embryonic development and the hatching process

1. This research focused on the embryonic development of broiler and layer embryos. 2. Egg, embryo and yolk weights were measured and partial pressure of gases in the air cell and blood were analysed at several embryonic ages. The static stiffness of the eggshell was measured before the start of incubation and at embryonic day (ED) 18 to register the change in shell strength. Times of internal pipping (IP), external pipping (EP) and hatch were recorded. Plasma corticosterone, triiodothyronine and thyroxine concentrations were determined. 3. Relative egg weight loss was higher in layer eggs. Before ED16, layer embryos showed a slower development which was reflected in lower (relative) embryo weight, lower air cell and blood pCO(2) and higher air cell O(2). From ED16 onwards, relative growth rate accelerated in the layer strain; as a consequence the difference in relative yolk-free chick weight at hatch had disappeared between strains. 4. Differences in physiological events necessary for hatching (thyroid hormones, corticosterone, air cell pCO(2)) are most probably responsible for the observed differences in timing of pipping and hatching events between layer and broilers.     

Ultrasonographic examinations of fetal growth of sheep between day 20 and day 50 of gestation

Previous studies on embryonic and fetal growth in sheep were mostly transversal using animals killed at various stages of gestation. Until now it was difficult to monitor the development of individual embryos/foetuses during pregnancy, especially during the first and second pregnancy month. Real-time ultrasound as a non invasive method could be an appropriate method for examination of embryonic and early foetal development in sheep. The aim of this study was to determine the embryonic and foetal development of the crown-rump-length (CRL) in pregnant ewes in relation to the number of fetuses and/or the breed. Between the 20th and 50th day of pregnancy the embryos/foetuses showed an exponential growth which can be best described by the equation of the form CRL (mm) = W * exp (k * day of pregnancy). The individual variability in embryofetal growth is in part due to the number of embryos per sheep and the sheep breed.