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Poultry Science Mar 2018In this study, we analyzed selected morphological traits of eggs, as well as structure, strength, and protein composition of the vitelline membrane (VM) of ostrich, emu,... (Comparative Study)
Comparative Study
In this study, we analyzed selected morphological traits of eggs, as well as structure, strength, and protein composition of the vitelline membrane (VM) of ostrich, emu, and greater rhea eggs. Ninety eggs (30 for species) were analyzed for the following parameters: egg weight, yolk weight, yolk ratio, and yolk index. In addition, pH value, water activity, consistency index, and flow behavior index were determined. The strength of VM was measured using the TA.HDPlus Texture Analyzer. Micrograph images were taken via a scanning electron microscope. Polyacrylamide gel electrophoresis was conducted under denaturing conditions. Ostrich eggs were characterized by the highest egg and yolk weight compared with those of emu and greater rhea eggs, whereas emu eggs had the highest yolk ratio compared with those of ostrich and greater rhea eggs (P > 0.05). Yolk content differed among the species in terms of water activity; it was found to be higher in emu eggs than in ostrich and greater rhea eggs (P > 0.05). Based on flow curves, yolks of the ratites were classified as pseudoplastic non-Newtonian fluids. The consistency index was significantly higher in yolks of ostrich and emu than that of greater rhea eggs, whereas the VM of yolks of greater rhea eggs was the most resistant (had the highest breaking force = 26.4 g). All species differed significantly regarding the structure of VM, the outer layer (OL) in particular, which was found to constitute fibers of various thicknesses that were differently arranged. Fibers of the OL of the VM of emu, whose fibers were the least differentiated but formed the most compact network, were the most diverse in characterization. An electropherogram of the VM of ostrich revealed 11 primary protein bands: 6 for the OL and 5 for the inner layer (IL), that of emu revealed 9 bands: 5 for the OL and 4 for the IL, and that of greater rhea revealed 10 bands: 6 for the OL and 4 for the IL.
Topics: Animals; Dromaiidae; Egg Yolk; Ovum; Rheiformes; Struthioniformes; Vitelline Membrane
PubMed: 29253213
DOI: 10.3382/ps/pex356 -
PloS One 2017Most vectors of arthropod-borne diseases produce large eggs with hard and opaque eggshells. In several species, it is still not possible to induce molecular...
Most vectors of arthropod-borne diseases produce large eggs with hard and opaque eggshells. In several species, it is still not possible to induce molecular perturbations to the embryo by delivery of molecules using microinjections or eggshell permeabilization without losing embryo viability, which impairs basic studies regarding development and population control. Here we tested the properties and permeability of the eggshell of R. prolixus, a Chagas disease vector, with the aim to deliver pharmacological inhibitors to the egg cytoplasm and allow controlled molecular changes to the embryo. Using field emission scanning and transmission electron microscopy we found that R. prolixus egg is coated by three main layers: exochorion, vitelline layer and the plasma membrane, and that the pores that allow gas exchange (aeropiles) have an average diameter of 10 μm and are found in the rim of the operculum at the anterior pole of the egg. We tested if different solvents could permeate through the aeropiles and reach the egg cytoplasm/embryo and found that immersions of the eggs in ethanol lead to its prompt penetration through the aeropiles. A single five minute-immersion of the eggs/embryos in pharmacological inhibitors, such as azide, cyanide and cycloheximide, solubilized in ethanol resulted in impairment of embryogenesis in a dose dependent manner and DAPI-ethanol solutions were also able to label the embryo cells, showing that ethanol penetration was able to deliver those molecules to the embryo cells. Multiple immersions of the embryo in the same solutions increased the effect and tests using bafilomycin A1 and Pepstatin A, known inhibitors of the yolk proteolysis, were also able to impair embryogenesis and the yolk protein degradation. Additionally, we found that ethanol pre-treatments of the egg make the aeropiles more permeable to aqueous solutions, so drugs diluted in water can be carried after the eggs are pre-treated with ethanol. Thus, we found that delivery of pharmacological inhibitors to the embryo of R. prolixus can be performed simply by submersing the fertilized eggs in ethanol with no need for additional methods such as microinjections or electroporation. We discuss the potential importance of this methodology to the study of this vector developmental biology and population control.
Topics: Animals; Arthropod Vectors; Egg Shell; Electrophoresis, Polyacrylamide Gel; Embryo, Nonmammalian; Ethanol; Microscopy, Electron, Scanning; Microscopy, Electron, Transmission; Permeability; Rhodnius
PubMed: 28961275
DOI: 10.1371/journal.pone.0185770 -
Frontiers in Microbiology 2017During a study on high mortality cases of goose embryo in Shandong Province, China (2014-2015), we isolated an H9N2 avian influenza virus (AIV) strain...
During a study on high mortality cases of goose embryo in Shandong Province, China (2014-2015), we isolated an H9N2 avian influenza virus (AIV) strain (A/goose/Shandong/DP01/2014, DP01), which was supposedly the causative agent for goose embryo death. Sequence analysis revealed that DP01 shared 99.9% homology in the HA gene with a classic immune suppression strain SD06. To study the potential vertical transmission ability of the DP01 strain in breeder goose, a total of 105 Taizhou breeder geese, which were 360 days old, were equally divided into five groups (A, B, C, D, and E) for experimental infection. H9N2 AIV (DP01) was used for inoculating through intravenous (group A), intranasal instillation (group B), and throat inoculation (group C) routes, respectively. The geese in group D were inoculated with phosphate buffer solution (PBS) and those in group E were the non-treated group. At 24 h post inoculation, H9N2 viral RNA could be detected at vitelline membrane, embryos, and allantoic fluid of goose embryos from H9N2 inoculated groups. Furthermore, the HA gene of H9N2 virus from vitelline membrane, embryo, allantoic fluid, and gosling shared almost 100% homology with an H9N2 virus isolated from the ovary of breeder goose, which laid these eggs, indicating that H9N2 AIV can be vertically transmitted in goose. The present research study provides evidence that vertical transmission of H9N2 AIV from breeding goose to goslings is possible.
PubMed: 28861069
DOI: 10.3389/fmicb.2017.01559 -
Poultry Science Sep 2017Vitelline membrane (VM) is a multilayered structure that surrounds the egg yolk serving to separate the yolk and the white. Due to its poor solubility in aqueous-based...
Vitelline membrane (VM) is a multilayered structure that surrounds the egg yolk serving to separate the yolk and the white. Due to its poor solubility in aqueous-based media, VM proteins and their biological properties have not been fully defined. In the current study, VM was hydrolyzed using different enzymes under the optimum hydrolysis conditions. Antioxidant and anti-inflammatory properties were evaluated in chemical and cellular models. Flavourzyme- and trypsin-treated samples showed the highest radical scavenging and ferric ion reducing effect (31% and 20 μM of Trolox equivalents/mg, respectively). In cellular studies, all VM hydrolysates were cyto-compatible and inhibited nitric oxide production by RAW264.7 macrophage cells significantly. Lipopolysaccharide-stimulated up-regulation of pro-inflammatory cytokines in RAW264.7 cells was suppressed by flavourzyme-treated VM. These results revealed that enzymatic hydrolysis of VM is a promising approach to produce peptides with several bioactivities (free radical scavenging, metal chelation, and anti-inflammatory) as valuable ingredients for cosmeceuticals and nutraceuticals.
Topics: Animals; Anti-Inflammatory Agents; Antioxidants; Chickens; Egg Proteins; Hydrolysis; Models, Chemical; Protein Hydrolysates; Vitelline Membrane
PubMed: 28854750
DOI: 10.3382/ps/pex125 -
PloS One 2017The starlet sea-anemone Nematostella vectensis has emerged as a model organism in developmental biology. Still, our understanding of various biological features,...
The starlet sea-anemone Nematostella vectensis has emerged as a model organism in developmental biology. Still, our understanding of various biological features, including reproductive biology of this model species are in its infancy. Consequently, through histological sections, we study here key stages of the oogenesis (oocyte maturation/fertilization), as the state of the gonad region immediately after natural spawning. Germ cells develop in a secluded mesenterial gastrodermal zone, where the developing oocytes are surrounded by mucoid glandular cells and trophocytes (accessory cells). During vitellogenesis, the germinal vesicle in oocytes migrates towards the animal pole and the large polarized oocytes begin to mature, characterized by karyosphere formation. Then, the karyosphere breaks down, the chromosomes form the metaphase plate I and the eggs are extruded from the animal enclosed in a sticky, jelly-like mucoid mass, along with numerous nematosomes. Fertilization occurs externally at metaphase II via swimming sperm extruded by males during natural spawning. The polar bodies are ejected from the eggs and are situated within a narrow space between the egg's vitelline membrane and the adjacent edge of the jelly coat. The cortical reaction occurs only at the polar bodies' ejection site. Several spermatozoa can penetrate the same egg. Fertilization is accompanied by a strong ooplasmatic segregation. Immediately after spawning, the gonad region holds many previtellogenic and vitellogenic oocytes, though no oocytes with karyosphere. Above are the first histological descriptions for egg maturation, meiotic chromosome's status at fertilization, fertilization and the gonadal region's state following spawning, also documenting for the first time the ejection of the polar body.
Topics: Animals; Female; Fertilization; Gonads; Male; Oocytes; Oogenesis; Reproduction; Sea Anemones; Sperm-Ovum Interactions
PubMed: 28796817
DOI: 10.1371/journal.pone.0182677 -
Scientific Reports Jul 2017The major components of vitelline membrane (VM) are ovomucin, VM outer (VMO) I and VMO II. At present, the distribution pattern of maternal cells on the VM has not been...
The major components of vitelline membrane (VM) are ovomucin, VM outer (VMO) I and VMO II. At present, the distribution pattern of maternal cells on the VM has not been described in detail. In this study, the existence and distribution characteristics of maternal cells on VM were observed. There were more than 3.2 × 10 somatic cells on VM, which were uneven distributed. The calcein AM/PI staining of the maternal cells on the VM showed that the cells' viability changed with the freshness of the eggs, and that the maternal cells gradually underwent apoptosis and became degraded. The results of morphology of different tissues indicated that the most of maternal cells on the VM were granulosa cells. Moreover, the karyotype of the cultured granulosa cells, which is the main source of cells on VM, were identified as the normal diploid karyotype of chicken. Furthermore, the VM DNA extracted from chickens and quails, which represent the eggs of different size, was adequate for further genetic analysis. The VM DNA was easily accessible and relatively constant, without cross-contamination. Therefore, the VM DNA could potentially be applied for the molecular traceability between eggs and chickens, and be beneficial in avian ecology research studies.
Topics: Animals; Apoptosis; Cell Survival; Chickens; DNA; Diploidy; Female; Granulosa Cells; Karyotyping; Vitelline Membrane
PubMed: 28747770
DOI: 10.1038/s41598-017-06996-1 -
International Journal of Molecular... Jun 2017Time-dependent expression of proteins in ovary is important to understand oogenesis in insects. Here, we profiled the proteomes of developing ovaries from (Hendel) to...
Comparative Proteomic Profiling Reveals Molecular Characteristics Associated with Oogenesis and Oocyte Maturation during Ovarian Development of Bactrocera dorsalis (Hendel).
Time-dependent expression of proteins in ovary is important to understand oogenesis in insects. Here, we profiled the proteomes of developing ovaries from (Hendel) to obtain information about ovarian development with particular emphasis on differentially expressed proteins (DEPs) involved in oogenesis. A total of 4838 proteins were identified with an average peptide number of 8.15 and sequence coverage of 20.79%. Quantitative proteomic analysis showed that a total of 612 and 196 proteins were differentially expressed in developing and mature ovaries, respectively. Furthermore, 153, 196 and 59 potential target proteins were highly expressed in early, vitellogenic and mature ovaries and most tested DEPs had the similar trends consistent with the respective transcriptional profiles. These proteins were abundantly expressed in pre-vitellogenic and vitellogenic stages, including tropomyosin, vitellogenin, eukaryotic translation initiation factor, heat shock protein, importin protein, vitelline membrane protein, and chorion protein. Several hormone and signal pathway related proteins were also identified during ovarian development including piRNA, notch, insulin, juvenile, and ecdysone hormone signal pathways. This is the first report of a global ovary proteome of a tephritid fruit fly, and may contribute to understanding the complicate processes of ovarian development and exploring the potentially novel pest control targets.
Topics: Animals; Chromatography, Liquid; Female; Gene Expression Profiling; Insect Proteins; Oogenesis; Ovary; Proteomics; Tandem Mass Spectrometry; Tephritidae
PubMed: 28665301
DOI: 10.3390/ijms18071379 -
Parasitology Research Jul 2017The origin, differentiation and functional ultrastructure of oncospheral or egg envelopes in Echinococcus multilocularis Leuckart, 1863 were studied by transmission...
The origin, differentiation and functional ultrastructure of oncospheral or egg envelopes in Echinococcus multilocularis Leuckart, 1863 were studied by transmission electron microscopy (TEM) and cytochemistry. The purpose of our study is to describe the formation of the four primary embryonic envelopes, namely vitelline capsule, outer envelope, inner envelope and oncospheral membrane, and their transformation into the oncospheral or egg envelopes surrounding the mature hexacanth. This transformation takes place in the preoncospheral phase of embryonic development. The vitelline capsule and oncospheral membrane are thin membranes, while the outer and inner envelopes are thick cytoplasmic layers formed by two specific types of blastomeres: the outer envelope by cytoplasmic fusion of two macromeres and the inner envelope by cytoplasmic fusion of three mesomeres. Both outer and inner envelopes are therefore cellular in origin and syncytial in nature. During the advanced phase of embryonic development, the outer and inner envelopes undergo great modifications. The outer envelope remains as a metabolically active layer involved in the storage of glycogen and lipids for the final stages of egg development and survival. The inner envelope is the most important protective layer because of its thick layer of embryophoric blocks that assures oncospheral protection and survival. This embryophore is the principal layer of mature eggs, affording physical and physiological protection for the differentiated embryo or oncosphere, since the outer envelope is stripped from the egg before it is liberated. The embryophore is very thick and impermeable, consisting of polygonal blocks of an inert keratin-like protein held together by a cementing substance. The embryophore therefore assures extreme resistance of eggs, enabling them to withstand a wide range of environmental temperatures and physicochemical conditions.
Topics: Animals; Cell Differentiation; Cytoplasm; Echinococcus multilocularis; Female; Microscopy, Electron, Transmission; Ovum
PubMed: 28593390
DOI: 10.1007/s00436-017-5479-x -
Parasites & Vectors Apr 2017Neorickettsia are a group of intracellular α proteobacteria transmitted by digeneans (Platyhelminthes, Trematoda). These endobacteria can also infect vertebrate hosts...
BACKGROUND
Neorickettsia are a group of intracellular α proteobacteria transmitted by digeneans (Platyhelminthes, Trematoda). These endobacteria can also infect vertebrate hosts of the helminths and cause serious diseases in animals and humans. Neorickettsia have been isolated from infected animals and maintained in cell cultures, and their morphology in mammalian cells has been described. However, limited information is available on the morphology and localization of Neorickettsia in the trematode host.
METHODS
We used a Neorickettsia-infected strain of the model trematode Plagiorchis elegans to infect Syrian Golden hamsters to produce adult worms. Ultrastructure of Neorickettsia was assessed by transmission electron microscopy of high pressure freezing/freeze substitution fixed specimens. A Neorickettsia surface protein from P. elegans (PeNsp-3) was cloned and antibodies against the recombinant protein were used to localize Neorickettsia by immunohistochemistry.
RESULTS
Ultrastructural analysis revealed moderate numbers of pleomorphic endobacteria with a median size of 600 × 400 nm and characteristic double membranes in various tissue types. Endobacteria showed tubular membrane invaginations and secretion of polymorphic vesicles. Endobacteria were unevenly localized as single cells, or less frequently as small morula-like clusters in the ovary, Mehlis' gland, vitelline follicles, uterus, intrauterine eggs, testis, cirrus-sac, tegument, intestine and the oral and ventral sucker. Examination of hamster small intestine infected with P. elegans showed many endobacteria at the host-parasite interface such as the oral and ventral sucker, the tegument and the excretory pore.
CONCLUSIONS
We conclude that adult P. elegans trematodes carry Neorickettsia endobacteria in varying numbers in many tissue types that support vertical transmission, trematode to trematode transmission via seminal fluid, and possibly horizontal transmission from trematodes to vertebrate hosts. These means appear to be novel mechanisms of pathogen transmission by endoparasitic worms.
Topics: Animal Structures; Animals; Female; Immunohistochemistry; Male; Mesocricetus; Microscopy, Electron, Transmission; Neorickettsia; Trematoda
PubMed: 28407790
DOI: 10.1186/s13071-017-2123-7 -
Genetics Apr 2017The eggshell is an extracellular matrix that confers protection to the egg and also plays a role in transferring positional information from the ovary to pattern the...
The eggshell is an extracellular matrix that confers protection to the egg and also plays a role in transferring positional information from the ovary to pattern the embryo. Among the constituents of the eggshell, Nasrat, Polehole, and Closca form a group of proteins related by sequence, secreted by the oocyte, and mutually required for their incorporation into the eggshell. Besides their role in eggshell integrity, Nasrat, Polehole, and Closca are also required for embryonic terminal patterning by anchoring or stabilizing Torso-like at the eggshell. Here, we show that they are also required for dorsoventral patterning, thereby unveiling that the dorsoventral and terminal systems, hitherto considered independent, share a common extracellular step. Furthermore, we show that Nasrat, Polehole, and Closca are required for proper Nudel activity, a protease acting both in embryonic dorsoventral patterning and eggshell integrity, thus providing a means to account for the role of Nasrat, Polehole, and Closca. We propose that a Nasrat/Polehole/Closca complex acts as a multifunctional hub to anchor various proteins synthesized at oogenesis, ensuring their spatial and temporal restricted function.
Topics: Animals; Body Patterning; Drosophila; Drosophila Proteins; Egg Proteins; Embryo, Nonmammalian; Female; Membrane Proteins; Ovary; Proto-Oncogene Proteins c-raf; Serine Endopeptidases
PubMed: 28179368
DOI: 10.1534/genetics.116.197574