Search results for: embryo

Authors: Diah Tri Widayati

Abstract:

Various assisted reproductive techniques have been developed and refined to obtain a large number of offspring from genetically superior animals or obtain offspring from infertile (or subfertile) animals. The embryo transfer is one assisted reproductive technique developed well, aimed at increased productivity of selected females, disease control, importation and exportation of livestock, rapid screening of AI sires for genetically recessive characteristics, treatment or circumvention of certain types of infertility. Embryo transfer also is a useful research tool for evaluating fetal and maternal interactions. This technique has been applied to nearly every species of domestic animal and many species of wildlife and exotic animals, including humans and non-human primates. The successful of embryo transfers have been limited to within-animal, homologous replacement of the embryos. There are several examples of interspecific and intergeneric embryo transfers in which embryos implanted but did not develop to term: sheep and goat, mouse and rat. An immunological rejections and placental incompatibility between the embryo and the surrogate mother appear to restrict interspecific embryo transfer/interspecific pregnancy. Recently, preimplantation embryo manipulation procedures have been applied, such as technique of inner cell mass transfer. This technique will possible to overcome the reproductive barrier interspecific embryo transfer/interspecific pregnancy, if there is a protective mechanism which prevents recognition of the foreign fetus by the mother of the other species

Keywords: Embryo Transfer, Assisted Reproductive Techology, Intraspesific-Interspesific Pregnancy, Inner cell mass.

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Authors: Münüre Tanur Erkoyuncu, Mustafa Yorgancılar

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Crop improvement through genetic engineering depends on effective and reproducible plant regeneration systems. Immature embryos are the most widely used explant source for in vitro regeneration in barley (Hordeum vulgare L.). However, immature embryos require the continuous growth of donor plants and the suitable stage for their culture is also certainly limited. On the other hand, mature embryos can be procured and stored easily; they can be studied throughout the year. In this study, an effective callus induction and plant regeneration were aimed to develop from mature embryos of different barley genotypes. The effect of medium (MS₁ and MS₂), auxin type (2,4-D, dicamba, picloram and 2,4,5-T) and concentrations (2, 4, 6 mg/l) on callus formation and effect of cytokinin type (TDZ, BAP) and concentrations (0.2, 0.5, 1.0 mg/l) on green plant regeneration were evaluated in mature embryo culture of barley. Callus and shoot formation was successful for all genotypes. By depending on genotype, MS₁ is the best medium, 4 mg/l dicamba is the best growth regulator in the callus induction and MS₁ is the best medium, 1 mg/l BAP is the best growth regulator in the shoot formation were determined.

Keywords: Barley, callus, embryo culture, mature embryo.

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Authors: André M. Ornelas, Lise P. Labéjof, Ligia V. Lage dos Santos, Jackson A. Santos

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The excessive use of agricultural pesticides and the resulting contamination of food and beds of rivers have been a recurring problem nowadays. Some of these substances can cause changes in endocrine balance and impair reproductive function of human and animal population. In the present study, we evaluated the possible effects of the fungicide cuprous copper oxide Sandoz® on pregnant Wistar rats. They received a daily oral administration of 103 or 3.103 mg/kg of the fungicide from the 6th to the 15th day of gestation. On day 21 of gestation, the maternal and fetal toxicity parameters and indices were determined. The administration of cuprous oxide (Copper Sandoz) in Wistar rats, the period of organogenesis, revealed no evidence of maternal toxicity or embryo at the studied doses.

Keywords: Reproductive toxicity, endocrine disrupter, cupper Sandoz®, rodent

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Authors: Samaneh Nazeri, Bagher Mojazi Amiri, Hamid Farahmand

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Production of high quality fish eggs with reasonable hatching rate makes a success in aquaculture industries. It is influenced by the environmental stimulators and inhibitors. Diazinon is a widely-used pesticide in Golestan province (Southern Caspian Sea, North of Iran) which is washed to the aquatic environment (3 mg/L in the river). It is little known about the effect of this pesticide on the embryogenesis of sturgeon fish, the valuable species of the Caspian Sea. Hormonal content of the egg is an important factor to guaranty the successful passes of embryonic stages. In this study, the fate of Persian sturgeon embryo to 24, 48, 72, and 96-hours exposure of diazinon (LC₅₀ dose) was tested. Also, the effect of thyroid hormones (T3 and T4) on these embryos was tested concurrently or separately with diazinon LC ₅₀ dose. Fertilized eggs are exposed to T3 (low dose: 1 ng/ml, high dose: 10 ng/ml), T4 (low dose: 1 ng/ml, high dose: 10 ng/ml). Six eggs were randomly selected from each treatment (with three replicates) in five developmental stages (two cell- division, neural, heart present, heart beaten, and hatched larvae). The possibility of changing T3, T4, and cortisol contents of the embryos were determined in all treated groups and in every mentioned embryonic stage. The hatching rate in treated groups was assayed at the end of the embryogenesis to clarify the effect of thyroid hormones and diazinon. The results indicated significant differences in thyroid hormone contents, but no significant differences were recognized in cortisol levels at various early life stages of embryos. There was also significant difference in thyroid hormones in (T3, T4) + diazinon treated embryos (P˂0.05), while no significant difference between control and treatments in cortisol levels was observed. The highest hatching rate was recorded in HT3 treatment, while the lowest hatching rate was recorded for diazinon LC₅₀ treatment. The result confirmed that Persian sturgeon embryo is less sensitive to diazinon compared to teleost embryos, and thyroid hormones may increase hatching rate even in the presence of diazinon.

Keywords: Persian sturgeon, diazinon, thyroid hormones, cortisol, embryo.

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Authors: Okafor C. Uche, Agbo P. Ejiofor, Okezie C. Eziuche

Abstract:

This study provides a regeneration protocol for Treculia africana Decne (an endangered plant) through embryo culture. Mature zygotic embryos of T. africana were excised from the seeds aseptically and cultured on varied strengths (full, half and quarter) of Murashige and Skoog (MS) basal medium supplemented. All treatments experienced 100±0.00 percent sprouting except for half and quarter strengths. Plantlets in MS full strength had the highest fresh weight, leaf area, and longest shoot length when compared to other treatments. All explants in full, half, quarter strengths and control had the same number of leaves and sprout rate. Between the treatments, there was a significant difference (P>0.05) in their effect on the length of shoot and root, number of adventitious root, leaf area, and fresh weight. Full strength had the highest mean value in all the above-mentioned parameters and differed significantly (P>0.05) from others except in shoot length, number of adventitious roots, and root length where it did not differ (P<0.05) from half strength. The result of this study indicates that full strength MS basal medium offers a better option for the optimum growth for Treculia africana regeneration in vitro.

Keywords: Medium strengths, Murashige and Skoog, Treculia africana, zygotic embryos.

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Authors: Sansani Rungrattawatchai

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The White Kwao (Pueraria mirifica), a potent phytoestrogenic medicinal plant, has long been use in Thailand as a traditional folkmedicine. However, no scientific information of the direct effect of White Kwao on the development of mammalian embryo was available. Therefore, the purpose of this study was to investigate the effect of White Kwao extract on the in vitro development and implantation rate of mouse embryos. This study was designed into two experiments. In the first experiment, the two-cell stage mouse embryos were collected from the oviduct of superovulated mature female mice, and randomly cultured in three different media, the M16, M16 supplemented with 0.52μg esthinylestradiol-17β, and M16 supplemented with 10 mg/ml White Kwao extract. The culture was incubated in CO2 incubator at 37 oC . After the embryos were cultivated, the developments of embryos were observed every 24 hours for 5 days. The development rate of embryos from the two-cell stage to blastocyst stage in the media was with White Kwao was significantly higher (p<0.05) than those of the control group (68.50% versus 43.50%) but did not differ from the positive control group (68.50% versus 57.66%). In the second experiment, hatched blastocysts, which obtained from three different media, were differently labeled the nuclei with two polynucleotide-specific fluorochromes, the propidium iodide (PI) and the bisbenzimide. The results showed that the number of trophectoderm cells in the blastocysts that cultivated in the media with White Kwao did not significantly differ from the control (80.00 versus 70 cells) and the positive control group (80.00 versus 112.50 cells). The average number of inner cell mass in the White Kwao treated group did not significantly differ from the control group (20.50 versus 16.00 cells) and the positive control group (20.50 versus 20.50 cells). The total cell number including the trophectoderm and the inner cell mass of the individual hatched blastocyst was evaluated. The cell number in the blastocysts obtained from the media with the White Kwao did not significantly differ from the control (94.25 + 9.50 versus 92.33 + 4.05) and the positive control group (94.25 + 9.50 versus 110.33 + 9.16). The results demonstrated that the White Kwao treatment group did have a stimulating effect on the in vitro development of mouse embryos. The exact mechanism that White Kwao stimulated mouse embryo development is not known. The suspect mechanism may in a manner similar to the mechanism that of estrogen stimulated the development of the mouse embryos. Futher studies are needed to transfer the blastocyst into the endometrium of pseudopreagnancy mice to evaluate the effect of White Kwao on implantation

Keywords: White Kwao (Pueraria mirifica), blastocyst.

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Authors: M. H. Rajikin, S. M. M. Syairah, A. R. Sharaniza

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Effects of nicotine on pre-partum body weight and preimplantation embryonic development has been reported previously. Present study was conducted to determine the effects of annatto (Bixa orellana)-derived delta-tocotrienol (TCT) (with presence of 10% gamma-TCT isomer) on the nicotine-induced reduction in body weight and 8-cell embryonic growth in mice. Twenty-four 6-8 weeks old (23-25g) female balb/c mice were randomly divided into four groups (G1-G4; n=6). Those groups were subjected to the following treatments for 7 consecutive days: G1 (control) were gavaged with 0.1 ml tocopherol stripped corn oil. G2 was subcutaneously (s.c.) injected with 3 mg/kg/day of nicotine. G3 received concurrent treatment of nicotine (3 mg/kg/day) and 60 mg/kg/day of δ-TCT mixture (contains 90% delta & 10% gamma isomers) and G4 was given 60 mg/kg/day of δ-TCT mixture alone. Body weights were recorded daily during the treatment. On Day 8, females were superovulated with 5 IU Pregnant Mare’s Serum Gonadotropin (PMSG) for 48 hours followed with 5 IU human Chorionic Gonadotropin (hCG) before mated with males at the ratio of 1:1. Females were sacrificed by cervical dislocation for embryo collection 48 hours post-coitum. Collected embryos were cultured in vitro. Results showed that throughout Day 1 to Day 7, the body weight of nicotine treated group (G2) was significantly lower (p<0.05) than that of G1, G3 and G4. Intervention with δ-TCT mixture (G3) managed to increase the body weight close to the control group. This is also observed in the group treated with δ-TCT mixture alone (G4). The development of 8-cell embryos following in vitro culture (IVC) was totally inhibited in G2. Intervention with δ- TCT mixture (G3) resulted in the production of 8-cell embryos, although it was not up to that of the control group. Treatment with δ- TCT mixture alone (G4) caused significant increase in the average number of produced 8-cell embryo compared to G1. Present data indicated that δ-TCT mixture was able to reverse the body weight loss in nicotine treated mice and the development of 8-cell embryos was also improved. Further analysis on the quality of embryos need to done to confirm the effects of δ-TCT mixture on preimplantation embryos.

Keywords: δ-tocotrienol, body weight, nicotine, preimplantation embryonic development.

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Authors: G. A. Oladosu, P. O. Ogbodogbo, C. I. Makinde, M. O. Tijani, O. A. Adegboyega

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Saprolegniasis is a major pathogenic infection that contributes significantly to poor hatching rates in incubated fish eggs in the African catfish hatchery in Nigeria. Malachite green, known to be very effective against this condition, has been banned because it is carcinogenic. There is therefore the need for other effective, yet safer method of controlling saprolegniasis in incubated fish eggs. A total of 50 ml crude, chloroform extract of pyocyanin from which solvent was removed to attain 30 ml, having a concentration of 12.16 ug/ml was produced from 700 ml broth culture of Pseudomonas aeruginosa isolated from a previous study. In vitro susceptibility of the fungus was investigated by exposing fungal infected eggs to two different time-concentration ratios of pyocyanin; 0.275 ug/ml and 2.75 ug/ml for 1 and 24 h, and 5 mg/L malachite green as positive control while normal saline was the control. Efficacy of pyocyanin was evaluated using the degree of mycelial growth inhibition in the different treatments. Fertilized Clarias gariepinus eggs (between 45 to 64 eggs) were then incubated in 20 ml of medium containing the similar concentrations of pyocyanin and malachite green, with freshwater as control for 24 hours. Hatching rates of the incubated eggs were observed. Three samples of un-hatched eggs were taken from each medium and observed for the presence of fungal pathogens using microscopy. Another batch of three samples of un-hatched eggs from each treatment was also inoculated on Sabourand dextrose agar (SDA) using Egg-Agar Transfer technique to observe for fungal growth. Mycelial growth was inhibited in fungal infected eggs treated with 2.75 ug/ml for 24 h and the 5 mg/L malachite green for both 1 h and 24 h. The mortality rate was 100% in fertilized C. gariepinus eggs exposed for 24 h to 0.275 and 2.75 ug/ml of pyocyanin. The mortality rate was least in the malachite green followed by the control treatment. Embryonic development was observed to be arrested in the eggs treated with the two pyocyanin concentrations as they maintain their color but showed no development beyond the gastrula stage, whereas viable eggs in the control and malachite green treatments developed fully into healthy hatchlings. Furthermore, microscopy of the un-hatched eggs revealed the presence of a protozoan ciliate; Colpidium sp. (Tetrahymenidae), as well as a pathogenic fungus; Saprolegnia sp. in the control, but not in the malachite green and pyocyanin treatments. Growth of Saprolegnia sp. was also observed in SDA culture of un-hatched eggs from the control, but not from pyocyanin and malachite green treated eggs. Pyocyanin treatment of incubated eggs of Clarias gariepinus effectively prevented fungal infection in the eggs, but also arrested the development of the embryo. Therefore, crude chloroform extract of pyocyanin from Pseudomonas aeruginosa cannot be used in the control of Saprolegniasis in incubated Clarias gariepinus eggs at the concentration and duration tested in this study.

Keywords: African catfish, bioprophylaxis, embryo, saprolegniasis.

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Authors: Masoume Amirkhani, Kambiz Mashayekhi, Maurizio Lambardi

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Agropyron cristatum L. Gaertn. is a native grass of semiarid region in Iran which is quit resistant to cool and drought climate and withstand heavy grazing. This species has close phylogenetic relationship with Triticum and Hordeum. In this research, the effect of seven different concentrations of growth regulator 2,4-D on callus production and somatic embryogenesis of A. cristatum was investigated on Murashige and Skoog medium. The results showed that the rate of callus, embryo and neomorph were highest in 1 mg L-1 2,4-D. Callus production was increased in 1 mg L-1 2,4-D but dramatically decreased at 5.5 and 9 mg L-1 2,4-D. The somatic embryos were observed at 1 and 4 mg L-1 2,4-D but matured embryos and plantlet were only occurred at 1 mg L-1 2,4-D. There were significant differences between 1 mg L-1 2,4-D and other treatments for producing globular and torpedo embryos, plantlet, rooted callus and number of roots (p<0.05) and there was not any callus production and embryogenesis in control treatment without growth regulator.

Keywords: 2, 4-D, callus production, somatic embryogenesis, Agropyron cristatum.

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Authors: Leila Amjad, Mahsa Shafighi

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The role of the pollen grain, with to the reproductive process of higher plants, is to deliver the spermatic cells to the embryo sac for egg fertilization. The aim of this project was study the effect of electromagnetic fields on structure and pollen grains development in Chenopodium album. Anthers of Chenopodium album L. were collected at different stages of development from control (without electromagnetic field) and plants grown at 10m from the field sources. Structure and development of pollen grains were studied and compared. The studying pollen structure by Light and Scanning electron microscopy showed that electromagnetic fields reduction of pollen grains number and male sterility, thus , in some anthers, pollen grains were attached together and deformed compared to control ones. The data presented suggest that prolonged exposures of plants to magnetic field may cause different biological effects at the cellular tissue and organ levels.

Keywords: Electromagnetic fields, pollen, Chenopodium albumL.

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Authors: Sunil Dehipawala, Todd Holden, E. Cheung, Robert Regan, P. Schneider, G. Tremberger Jr, D. Lieberman, T. Cheung

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The zinc and iron environments in different growth stages have been studied with EXAFS and XANES with Brookhaven Synchrotron Light Source. Tissue samples included meat, organ, vegetable, leaf, and yeast. The project studied the EXAFS and XANES of tissue samples using Zn and Fe K-edges. Duck embryo samples show that brain and intestine would contain shorter EXFAS determined Zn-N/O bond; as with the cases of fresh yeast versus reconstituted live yeast and green leaf versus yellow leaf. The XANES Fourier transform characteristic-length would be useful as a functionality index for selected types of tissue samples in various physical states. The extension to the development of functional synchrotron imaging for tissue engineering application based on spectroscopic technique is discussed.

Keywords: EXAFS, Fourier Transform, metalloproteins, XANES

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Authors: S. Moghaddaszadeh-Ahrabi, S. Farajnia, Gh. Rahimi-Mianji, A. Nejati-Javaremi

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In recent years, there has been an increasing interest toward the use of bovine genotyped embryos for commercial embryo transfer programs. Biopsy of a few cells in morulla stage is essential for preimplantation genetic diagnosis (PGD). Low amount of DNA have limited performing the several molecular analyses within PGD analyses. Whole genome amplification (WGA) promises to eliminate this problem. We evaluated the possibility and performance of an improved primer extension preamplification (I-PEP) method with a range of starting bovine genomic DNA from 1-8 cells into the WGA reaction. We optimized a short and simple I-PEP (ssI-PEP) procedure (~3h). This optimized WGA method was assessed by 6 loci specific polymerase chain reactions (PCRs), included restriction fragments length polymorphism (RFLP). Optimized WGA procedure possesses enough sensitivity for molecular genetic analyses through the few input cells. This is a new era for generating characterized bovine embryos in preimplantation stage.

Keywords: Whole genome amplification (WGA), Genotyping, Bovine, Preimplantation genetic diagnosis (PGD)

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Authors: Agata Kowalska, Radosław K. Kowalski, Zdzisław Zakęś

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Our results showed that treatment with both cyclooxygenase (COX1 or COX2) inhibitors impair reproduction parameters of the medaka. Resveratrol (COX1 inhibitor) caused an decrease in the number of spawning females at the first week of feeding fish with experimental diets. In the group treated with NS- 398 (COX2 inhibitor) we found the lowest sperm velocity parameters and decreased linearity of movement. The ovaries of the medaka fed feed supplemented with Resveratrol or NS-398 were confirmed to have a lower share of matured oocytes however during the experiment (four weeks) the number of eggs spawned by females was similar. Both inhibitors in fish diet (20 mg/kg body weight/day) caused a decrease in the embryo survival. Our results revealed that for the medaka female reproduction, activity of both COX enzymes might be necessary whereas males reproduction competence, as expressed by sperm motility parameters, might be related to COX2 activity.

Keywords: COX innibitors, medaka, reproduction parameters

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Authors: Yulia Irnidayanti, Win Darmanto, Agus Abadi

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research goal was to determine the expression levels cDNA of brain embrio at gestation days 10 (GD-10). The Electroforesis DNA results showed that GAPDH, Fibronectin1, Ncam1, Tenascin, Vimentin, Neurofilament heavy, Neurofilament medium and Neurofilament low were 447 bp, 462 bp, 293 bp. 416 bp, 327 bp, 301 bp, 398 bp and 289 bp. Result of real-time RT-PCR on brain Embryo at gestation days 10 showed that the expression of copy gen Fibronectin 36 copies, Ncam 21,708 copies; Tenascin 24,505 copies; Vimentin 538,554 copies; Neurofilament heavy 2,419 copies; Neurofilament medium 92,928 copies; Neurofilament low 125,809 copies. Vimentin expressed gene copies is very high compared with other gene copies. This condition are caused by Vimentin, that contribute to proliferate of brain development. The vimentin role to cell proliferation of brain.

Keywords: GAPDH, Fibronectin, Ncam, Tenascin, vimentin, Neurofilamen heavy, Neurofilament medium, Neurofilamen low.

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Authors: Salsabeel F. M. Alfalah, Jannat F. Falah, Nadia Muhaidat, Amjad Hudaib, Diana Koshebye, Sawsan AlHourani

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Educational technology is changing the way how students engage and interact with learning materials. This improved the learning process amongst various subjects. Virtual Reality (VR) applications are considered one of the evolving methods that have contributed to enhancing medical education. This paper utilizes VR to provide a solution to improve the delivery of the subject of Embryology to medical students, and facilitate the teaching process by providing a useful aid to lecturers, whilst proving the effectiveness of this new technology in this particular area. After evaluating the current teaching methods and identifying students ‘needs, a VR system was designed that demonstrates in an interactive fashion the development of the human embryo from fertilization to week ten of intrauterine development. This system aims to overcome some of the problems faced by the students’ in the current educational methods, and to increase the efficacy of the learning process.

Keywords: Virtual reality, student assessment, medical education, 3D, embryology.

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Authors: Yulia Irnidayanti

Abstract:

Retinoic acid is like a steroid hormone that plays a role in embryo formation, proliferation of spermatogonia cells, ephitelial cells differentiation and organogenesis. Retinoic acid can influences seminiferous tubule formation during embryonic testis development and also play a role in the regulation of ovarian function and female reproductive tract by suppressing the hormones FSH receptor expression. The excessive use of retinoic acid caused abnormalities in the fetus. The result showed that there is the influence of retinoic acid on the developmet of mice fetal testes, for examples disruption of the formation of seminiferous tubules and tubules seemed to be hollow, spermatogonia cells are relatively few in number and caused Leydig cells count relatively more. While in the female fetus does not caused the formation of primordial follicles and disrupted the development of germinal ephitelial cells of fetal ovaries of female mice (mus musculus) Swiss Webster.

Keywords: Retinoic acid, Leydig cell, Spermatogonia cells, Semin- ferous tubules, Primordial follicles

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Authors: Mithilesh Singh, Rakhi Chaturvedi

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Neem is a highly heterozygous and commercially important perennial plant. Conventionally, it is propagated by seeds which loose viability within two weeks. Strictly cross pollinating nature of the plant causes serious barrier to the genetic improvement by conventional methods. Alternative methods of tree improvement such as somatic hybridization, mutagenesis and genetic transformation require an efficient in vitro plant regeneration system. In this regard, somatic embryogenesis particularly secondary somatic embryogenesis may offer an effective system for large scale plant propagation without affecting the clonal fidelity of the regenerants. It can be used for synthetic seed production, which further bolsters conservation of this tree species which is otherwise very difficult The present report describes the culture conditions necessary to induce and maintain repetitive somatic embryogenesis, for the first time, in neem. Out of various treatments tested, the somatic embryos were induced directly from immature zygotic embryos of neem on MS + TDZ (0.1 μM) + ABA (4 μM), in more than 76 % cultures. Direct secondary somatic embryogenesis occurred from primary somatic embryos on MS + IAA (5 μM) + GA3 (5 μM) in 12.5 % cultures. Embryogenic competence of the explant as well as of the primary embryos was maintained for a long period by repeated subcultures at frequent intervals. A maximum of 10 % of these somatic embryos were converted into plantlets.

Keywords: Azadirachta indica A. Juss., Cytokinin, Somatic embryogenesis, zygotic embryo culture.

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Authors: S. M. M. Syairah, M. H. Rajikin, A-R. Sharaniza

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Several embryonic cellular mechanism including cell cycle, growth and apoptosis are regulated by phosphatidylinositol-3- kinase (PI3K)/Akt signaling pathway. The goal of present study is to determine the effects of annatto (Bixa orellana)-derived δ-tocotrienol (δ-TCT) on the regulations of PI3K/Akt genes in murine morula. Twenty four 6-8 week old (23-25g) female balb/c mice were randomly divided into four groups (G1-G4; n=6). Those groups were subjected to the following treatments for 7 consecutive days: G1 (control) received tocopherol stripped corn oil, G2 was given 60 mg/kg/day of δ-TCT mixture (contains 90% delta & 10% gamma isomers), G3 was given 60 mg/kg/day of pure δ-TCT (>98% purity) and G4 received 60 mg/kg/day α-TOC. On Day 8, females were superovulated with 5 IU Pregnant Mare’s Serum Gonadotropin (PMSG) for 48 hours followed with 5 IU human Chorionic Gonadotropin (hCG) before mated with males at the ratio of 1:1. Females were sacrificed by cervical dislocation for embryo collection 48 hours post-coitum. About fifty morulas from each group were used in the gene expression analyses using Affymetrix QuantiGene Plex 2.0 Assay. Present data showed a significant increase (p<0.05) in the average number (mean + SEM) of morula produced in G2 (27.32 + 0.23), G3 (25.42 + 0.21) and G4 (27.21 + 0.34) compared to control group (G1 – 14.61 + 0.25). This is parallel with the high expression of PDK1 gene with increase of 2.75-fold (G2), 3.07-fold (G3) and 3.59-fold (G4) compared to G1. From the present data, it can be concluded that supplementation with δ-TCT(s) and α-TOC induced high expression of PDK1 in G2-G4 which enhanced the PI3K/Akt signaling activity, resulting in the increased number of morula.

Keywords: Embryonic development, morula, nicotine, vitamin E.

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Authors: A. B. A. Ahmed, Amar, D. I., R. M. Taha

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This research aims to investigate callus induction, somatic embryogenesis and indirect plant regeneration of Crassula ovata (Mill.) Druce – the famous ornamental plant. Experiment no.1: Callus induction was obtained from leaf and stem explants on Murashige and Skoog (MS) medium supplemented with various plant growth regulators (PGRs). Effects of different PGRs, plant regeneration and subsequent plantlet conversion were also assessed. Indirect plant regeneration was achieved from the callus of stem explants by the addition of 1.5 mg/L Kinetin (KN) alone. Best shoot induction was achieved (6.5 shoots/per explant) after 60 days. For successful rooting, regenerated plantlets were sub-cultured on the same MS media supplemented with 1.5 mg/L KN alone. The rooted plantlets were acclimatized and the survival rate was 90%. Experiment no.2: Results revealed that 0.5 mg/L 2,4-D alone and in combination with 1.0 mg/L 6-Benzyladenine (BA) gave 89.8% callus from the stem explants as compared to leaf explants. Callus proliferation and somatic embryo formation were also evaluated by ‘Double Staining Method’ and different stages of somatic embryogenesis were revealed by scanning electron microscope. Full Strength MS medium produced the highest number (49.6%) of cotyledonary stage somatic embryos (SEs). Mature cotyledonary stage SEs developed into plantlets after 12 weeks of culture. Wellrooted plantlets were successfully acclimatized at the survival rate of 85%. Indirectly regenerated plants did not show any detectable variation in morphological and growth characteristics when compared with the donor plant.

Keywords: Callus induction, Crassula ovata, Double Staining, Indirect plant regeneration, Somatic embryogenesis.

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Authors: Mohammadjavad Mehrabanpour, Maryam Ranjbar Bushehri, Dorsa Mehrabanpour

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Respiratory diseases are the most important problems in the country’s poultry industry, particularly when it comes to broiler flocks. Ornithobacterium rhinotracheale (ORT) is a species that causes poor performance in growth rate, egg production, and mortality. This pathogen causes a respiratory infection including pulmonary alveolar inflammation, and pneumonia of birds throughout the world. Newcastle disease (ND) is a highly contagious disease in poultry, and also, it causes considerable losses to the poultry industry. The aim of this study was to evaluate the simultaneous occurrence of ORT and ND and NDV isolation by inoculation in embryonated eggs and confirmed by RT-PCR in broiler chicken flocks in Fars province. In this study, 318 blood and 85 tissue samples (brain, trachea, liver, and cecal tonsils) were collected from 15 broiler chicken farms. Survey serum antibody titers against ORT by using a commercial enzyme-linked immunosorbent assay (ELISA) kit performed. Evaluation of antibody titer against ND virus is performed by hemagglutination inhibition test. Virus isolation with chick embryo eggs 9-11 and RT-PCR method were carried out. A total of 318 serum samples, 135 samples (42.5%) were positive for antibodies to ORT and titer of HI antibodies against NDV in 122 serum samples (38/4%) were 7-10 (log2) and 61 serum samples (19/2%) had occurrence antibody titer against Newcastle virus and ORT. Results of the present study indicated that 20 tissue samples were positive in embryonated egg and in rapid hemagglutination (HA) test. HI test with specific ND positive serum confirmed that 6 of 20 samples. PCR confirmed that all six samples were positive and PCR products of samples indicated 535-base pair fragments in electrophrosis. Due to the great economic importance of these two diseases in the poultry industry, it is necessary to design and implement a comprehensive plan for prevention and control of these diseases.

Keywords: ELISA, Newcastle disease, Ornithobacterium rhinotracheale, seroprevalence.

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Authors: D. Biswas, Sang H. Hyun

Abstract:

In mammalian reproductive tract, the oviduct secretes huge number of growth factors and cytokines that create an optimal micro-environment for the initial stages of preimplantation embryos. Secretion of these growth factors is stage-specific. Among them, VEGF is a potent mitogen for vascular endothelium and stimulates vascular permeability. Apart from angiogenesis, VEGF in the oviduct may be involved in regulating the oocyte maturation and subsequent developmental process during embryo production in vitro. In experiment 1, to evaluate the effect of VEGF during IVM of porcine COC and subsequent developmental ability after PA and SCNT. The results from these experiments indicated that maturation rates among the different VEGF concentrations were not significant different. In experiment 2, total intracellular GSH concentrations of oocytes matured with VEGF (5-50 ng/ml) were increased significantly compared to a control and VEGF group (500 ng/ml). In experiment 3, the blastocyst formation rates and total cell number per blastocyst after parthenogenesis of oocytes matured with VEGF (5-50 ng/ml) were increased significantly compared to a control and VEGF group (500 ng/ml). Similarly, in experiment 4, the blastocyst formation rate and total cell number per blastocyst after SCNT and IVF of oocytes matured with VEGF (5 ng/ml) were significantly higher than that of oocytes matured without VEGF group. In experiment 5, at 10 hour after the onset of IVF, pronuclear formation rate was evaluated. Monospermy was significantly higher in VEGF-matured oocytes than in the control, and polyspermy and sperm penetration per oocyte were significantly higher in the control group than in the VEGFmatured oocytes. Supplementation with VEGF during IVM significantly improved male pronuclear formation as compared with the control. In experiment 6, type III cortical granule distribution in oocytes was more common in VEGF-matured oocytes than in the control. In conclusion, the present study suggested that supplementation of VEGF during IVM may enhance the developmental potential of porcine in vitro embryos through increase of the intracellular GSH level, higher MPN formation and increased fertilization rate as a consequence of an improved cytoplasmic maturation.

Keywords: angiogenesis, GSH, monospermy, VEGF

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