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Structure, function, and evolution of the oviducts of squamate reptiles, with special reference to viviparity and placentation.

In lizards and snakes, the oviducts function in fertilization, sperm storage, egg transport, eggshell deposition, maintenance of the early embryo, and expulsion of the egg or fetus. In viviparous forms they also contribute to placentae responsible for gas exchange and nutrient provision to the fetus. Dissections of species of 30 genera coupled with data from the literature indicate that squamate oviducts vary interspecifically in seven macroscopic features, including the extent and nature of regional differentiation, vascular supply, topographic asymmetry, number of oviducts, vaginal pouches, and relationship to the cloaca. The uterus, infundibulum, and vagina differ histologically in their epithelia, glands, and myometrial layers. Season cyclicity occurs in all three oviductal regions, most prominently in the uterus, and is under endocrinological control. Regional and cytological specializations reflect the diverse functions performed by the oviduct. Definitive evidence for oviductal albumen production and egg resorption is lacking. In viviparous squamates, three uterine specializations may facilitate maternal-fetal gas exchange: an attenuated epithelium, reduced uterine glands (and a reduced shell membrane), and increased vascularization. Contrary to previous reports, pregnant uteri show no epithelial erosion or capillary exposure. Specializations for nutrient provision to the fetus include mucosal hypertrophy, enlarged glandular epithelia, and multicellular glands whose secretions are absorbed by the chorioallantois. Comparisons with other amniotes indicate that squamates inherited the oviduct as an organ with capabilities for egg uptake and transport, fertilization, eggshell deposition, and oviposition. Other features have evolved convergently among squamates: infundibular sperm receptacles, unilateral oviduct loss, uterine gestation, placentation, and specializations for placentotrophy. Cladistic analysis indicates that oviductal features associated with deposition of tertiary egg investments in reptiles reflect evolutionary convergence as well as secondary simplification, rather than a unidirectional trend towards increased specialization.

Animals↗

Voltage associated with spontaneous embryonic motility in the developing chicken: an automated characterization during mid-late embryogenesis.

Movement of developing chicken embryos and their associated membranes generates voltage detectable with electrodes inserted just beneath the eggshell. Use of such voltages as a motility indicator offers an embryonic behavioral assessment method less subjective and invasive than observational methods using windows that disrupt substantial portions of the eggshell. We used a computerized signal recording and processing procedure to compare voltages from embryonic Day 12 (E12), E15, and E18 chicken eggs with embryos, assessed on the same day. Larger voltages were recorded from E18 subjects than from E12 or E15 subjects. Because this could have been due to embryonic size (mass) and/or proximity to the electrodes, making age comparisons uninterpretable, we used standard deviation-normalized and Z-score-based data transformations, comparing groups for relative deviations from basal voltages. E18 subjects still appeared more active than E12 subjects, with E15 a transitional age, in contrast to results from earlier window-based studies. The automated assessment method we used could enhance behavioral teratology studies of avian species.

Age Factors↗

Spectral Transforms as a Tool to Optimize Digital Phenotyping in Biological Images.

Modern livestock breeding has mastered genotyping. Genome-wide association studies, genomic selection, and SNP arrays enable genetic merit prediction at lower cost. However, phenotyping remains the bottleneck, as manual measurement is slow, expensive, subjective, and unable to capture spatial or temporal trait organization. Digital phenotyping via artificial intelligence could resolve this, but deep learning requires thousands of labelled examples, impractical when phenotyping cost itself limits datasets to hundreds of individuals. This creates a paradox: AI could accelerate phenotyping but requires large numbers of samples to train the models. Here, we demonstrate that integrating computer vision with machine learning offers sample-efficient digital phenotyping using eggshell colour as a model system. Rather than learning features from scratch (deep learning), we engineer physically motivated features via Wavelet transforms that decompose images into multi-scale spatial components. Wavelet features captured 14.2 percentage points more variance (R2&#x2009;=&#x2009;0.976 vs. 0.834, p&#x2009;<&#x2009;0.001) than standard colorimetry, with 50% better sample efficiency (achieving at n&#x2009;=&#x2009;60 what colorimetry required n&#x2009;=&#x2009;120). Variance decomposition revealed 77% of discriminative capacity derives from spatial patterns (bands, spots, gradients) invisible to scalar averages. Additionally, we identified "cryptic phenotypes" (3.3%) where spatial patterns contradicted average colour, cases where colorimeters failed but Wavelets succeeded. The underlying principle-that spatial decomposition can recover organizational information lost by scalar averaging-may be applicable to other traits with spatial or temporal structure, such as marbling, dermatitis, or pigmentation rhythms, although whether comparable performance gains would be observed remains to be tested empirically. Hence, for breeding programs implementing genomic selection, computer vision-based digital phenotyping captures complex trait variation without massive training datasets, addressing the bottleneck that increasingly limits genetic progress as genotyping becomes trivial.

Wavelet transform↗

Life history specializations of monogenean flatworms: a review of experimental and microscopical studies.

This review illustrates the use of experimental approaches combined with microscopy to study the biology of monogenean parasites. Studies of feeding, development, reproduction, and systematics have been based on gyrodactylids, flatworms infecting teleost fishes. In a contrasting system involving an amphibian host in a desert environment, analysis of adaptations to extreme conditions has focused on Pseudodiplorchis americanus. The unusual reproductive strategies, particularly the interactions between mother and offspring, are highlighted for both monogeneans. Species of Gyrodactylus are viviparous, maintaining up to three generations of embryos simultaneously in utero, and many of their reproductive specializations are related to progenesis. Embryo nutrition takes place via a metabolically-active syncytial uterine lining that has close association with the parental gut. Microscopy has also proved an essential adjunct to molecular studies of speciation and host specificity. P. americanus is ovoviviparous and the adaptations for embryo maintenance are unique. The primary keratin-type eggshell is replaced by a flexible secondary elastin capsule produced by the uterus; parental nutrients are transferred through cytoplasmic connections to the developing embryo. TEM has demonstrated unique adaptations of P. americanus to its micro-environments, including secretion of tegumental vesicles that provide protection from digestive enzymes during migration through the host gut. This paper highlights the potential of monogeneans for studies of fundamental biological principles.

Animals↗

Differential sensitivity of zonagenesis and vitellogenesis in Atlantic salmon (Salmo salar L) to DDT pesticides.

In Atlantic salmon (Salmo salar L) female sexual maturation entails both zonagenesis and vitellogenesis, both of which are controlled by increasing levels of estradiol-17 beta (E2). Antibodies against salmon zona radiata proteins (eggshell zr-proteins) and vitellogenin were used to monitor induction of oogenesis in juvenile salmon. Molecular weights of zr-monomers were estimated to about 66, 61, and 55 kDa, and to about 180 kDa for vitellogenin. Xenobiotics such as the pesticide DDT impair biological reproduction. The o,p'-DDT (1,1,1-trichloro-2[2-chlorophenyl]-2-[4-chlorophenyl]ethane) isomer seems to be a xenoestrogen. Serum levels of zr-proteins and vitellogenin, and hepatocytic biosynthesis of these components, were determined after in vivo treatment of salmon with DDT (technical, p,p'-(1,1,1-trichloro-2,2-bis[4-chlorophenyl]ethane) or o,p'-DDT) or E2. Exposing fish to frequent doses of o,p'-DDT (25 mg/kg b.w. (body weight) twice a week, six times totally) resulted in induction of all three zr-protein monomers, but not of vitellogenin. In contrast, three weekly injection of 10 mg/kg b.w. of either of the three DDT preparations did not induce typical zr-proteins or vitellogenin in serum. In vivo studies with combined DDT + E2 injections showed that none of the DDT preparations influenced E2-induced biosynthesis of zr-proteins or vitellogenin. E2 induction of these oogenetic processes was not blocked even by a high concentration (125 mg/kg b.w.) of o,p'-DDT. Furthermore, pretreatment of salmon with o,p'-DDT for 2 weeks, followed by one injection of E2, did not antagonize biosynthesis of zr-proteins, but serum concentration of vitellogenin was decreased. The data indicate that in juvenile salmon o,p'-DDT may be xenoestrogenic with regard to zonagenesis, but weakly anti-(xeno)estrogenic with regard to vitellogenesis. These findings suggest new complexities in fish reproductive toxicology of xenoestrogens. Compared to vitellogenesis, zonagenesis is a more sensitive parameter for monitoring reproductive effects of xenoestrogens.

Animals↗

Ontogeny of the extraembryonic membranes of the oviparous lizard, Eumeces fasciatus (Squamata: scincidae).

Oviposited eggs of Eumeces fasciatus contain embryos in the limb bud stage. Amniogenesis is complete and two yolk sac membranes, vascular trilaminar omphalopleure (choriovitelline membrane) and bilaminar omphalopleure, enclose the yolk vesicle. A small allantoic vesicle contacts the chorion. The choriovitelline membrane is the primary vascular system. Blood islands, sites of hematopoiesis, are associated with omphalomesenteric vessels of the choriovitelline membrane. The bilaminar omphalopleure, which contacts the eggshell over the abembryonic hemisphere of the egg, lies external to an isolated yolk mass and yolk cleft and is not vascularized. The definitive yolk sac (splanchnopleure) is formed when the extraembryonic coelom and allantoic vesicle intrude into the choriovitelline membrane. Omphalomesenteric vessels are retained with the yolk sac splanchnopleure and the associated hematopoietic sites are present throughout incubation. The chorioallantoic membrane reaches the equator of the egg, entirely supplanting the choriovitelline membrane, after 25% of incubation is completed. Further growth of the allantois is stalled until 65% of incubation is completed when rapid expansion of the allantoic vesicle, in conjunction with resorption of the isolated yolk mass, supplants the bilaminar omphalopleure. As a result, the chorioallantoic membrane completely envelopes the egg for the final 35% of incubation. This developmental event is coincident with published reports for the timing of increased growth and metabolism of embryos. As the isolated yolk mass regresses, intravitelline cells associated with the yolk cleft invade and resorb the yolk to form a large cavity. The wall of this cavity is a germinal epithelium that produces cells that fill the cavity. This structure appears to be a site of hematopoiesis previously undescribed in vertebrates.

Allantois↗

An unusual intraosseous lesion with fibroblastic, osteoclastic, osteoblastic, aneurysmal and fibromyxoid elements. "Solid" variant of aneurysmal bone cyst.

Four examples are reported of an unusual noncystic intraosseous lesion which does not conform to any hitherto recognized entity and which can be mistaken, not only by the general histopathologist but by the osteoarticular pathologist, for a variety of other conditions, including sarcoma and giant-cell tumor. They were in patients aged 5 to 13 years; three in the spine, one in the ethmoid. Local excision, supplemented by low-dose radiotherapy in cases with cord compression, produced a satisfactory outcome in all cases. At presentation the radiologic findings were nonspecific but, following treatment, an eggshell rim of bone developed in those lesions which had been incompletely excised. Histologically, they are characterised by florid fibroblastic or fibrohistiocytic proliferation, osteoblastic differentiation with osteoid production, areas rich in osteoclast-type giant cells, aneurysmal sinusoids, and occasional foci of degenerate calcifying fibromyxoid tissue. Because this combination of histologic features can be found in the solid parts of aneurysmal bone cyst and in no other condition, at this centre we have regarded this lesion as a variant of aneurysmal bone cyst devoid of any cystic component.

Adolescent↗

Chondrogenic potential of chick embryonic calvaria: II. Matrix calcium may repress cartilage differentiation.

Chick embryos cultured in the absence of their eggshell are rendered severely calcium-deficient, and develop a cartilage-like phenotype in the calvarium, a normally osteogenic tissue. In the preceding paper (Jacenko and Tuan [1995] Dev. Dyn. 202:13-26), experiments using organ cultured calvaria from day-12 normal and shell-less embryos showed that depletion of calcium alone may be responsible in promoting chondrogenic differentiation in calvaria. Here these findings were extended using an in vivo calvarial grafting technique, such that the extent of calvarial matrix calcification was a function of the calcium status of both the graft and the host. In these calvarial grafts, undermineralized regions again were shown to support chondrogenesis. To identify possible mechanisms which promote chondrogenesis in the calvaria, cells were enzymatically dissociated from the calvaria and cultured in media with varied levels of soluble calcium, under conditions which should modulate cell-to-cell interactions, including monolayer, micromass, agarose gels, and suspension cultures. Soluble calcium had no effect on calvarial cell differentiation, whereas conditions which enhanced cell-cell interactions, e.g., suspension culture, elicited cartilage expression. Based on these findings, we propose that the calcified matrix of the calvarium is repressive to chondrogenesis during normal development, but that the lack of mineral in a calcium-deficient calvarium creates a microenvironment permissive for cell-to-cell interactions which lead to chondrogenic differentiation.

Animals↗

Effects of varying chamber construction and embryo pre-incubation age on survival and growth of chick embryos in shell-less culture.

Shell-less culture involves culturing chick embryos with associated yolk and albumen outside of the eggshell and shell membranes. The technique allows direct access to and continuous observation of cultured embryos almost to the time of hatching. The plastic wrap/culture tripod technique described in this paper allows normal embryonic growth and differentiation from 48 hours (in ovo pre-incubation age) through at least 10 days of total incubation. As the duration of in ovo pre-incubation is decreased below 36 hours, there is a concomitant increase in the percentage of grossly abnormal embryos associated with decreased survival and retarded growth and differentiation. Survival of embryos pre-incubated for 72 hours through 13 and 18 days of total incubation is greater than 80% and 40%, respectively. The wrap/tripod technique allows substantially better survival and more normal development of cultured embryos than does the petri dish technique of Auerbach et al. (1974). Embryonic growth in 7.8-cm-diameter chambers is significantly greater than in either larger (10.6 cm) or smaller (5.2 cm) diameter chambers. Some gas exchange through the culture chamber walls appears necessary for optimal embryonic survival and growth. Suspending egg contents in either Safeway or Handi-Wrap plastic wrap (both of intermediate permeability) in 7.8-cm tripods resulted in superior growth and/or survival compared to suspension in Silastic sheeting (high permeability), Saran Wrap (low permeability), glass dishes (nonpermeable), or glass dishes lined with an inner layer of Safeway wrap(nonpermeable).

Animals↗

FT-Raman spectroscopy as diagnostic tool of Congo red binding to amyloids.

Chorion is the major component of silkmoth eggshell. More than 95% of its dry mass consists of the A and B families of low molecular weight structural proteins, which have remarkable mechanical and chemical properties protecting the oocyte and developing embryo from environmental hazards. We present data from FT-Raman spectroscopy of silkmoth chorion and amyloid-like fibrils formed from peptide analogues of chorion proteins, both unstained and stained by Congo red. The results show that FT-Raman spectroscopy is not a straightforward diagnostic tool for the specific interactions of Congo red with amyloids: a dilute aqueous solution of the Congo red dye at pH 5.5 and a thin solid film of the dye cast from this solution exhibit the same "diagnostic" Raman shifts relative to the neat Congo red dry powder as do amyloid fibrils formed from peptide analogues of chorion proteins stained by Congo red. An important consequence of this finding is that these shifts of the Raman active modes of Congo red are probably due to the formation of supramolecular dye aggregates in the presence of water. Therefore, this is not an appropriate diagnostic test for Congo red binding to amyloids.

Animals↗

Asymmetrical hatching behaviors: the development of postnatal motor laterality in three precocial bird species.

The effects of asymmetrical hatching behaviors on the development of turning bias and footedness in domestic chicks, bobwhite quail, and Japanese quail chicks were examined. Control tests with incubator reared domestic chicks and bobwhite quail revealed significant individual and population left-side turning bias and right footedness. When late stage hatching behaviors were disrupted, population laterality was not evident and individual laterality was reduced. By contrast, Japanese quail chicks demonstrated no population turning bias or footedness and only weak individual biases. Disruption of hatch behaviors further decreased laterality. Examination of discarded eggshells showed significant differences in the degree of rotation made to cut out of the egg by Japanese quail versus domestic chicks and bobwhite quail. Taken together these findings suggest that the counterclockwise hatching behaviors that are characteristic of many precocial bird species serve to facilitate the development of motor laterality at both the individual and population level.

Animals↗

Effects of prenatal visual stimulation on growth and heart rate in bobwhite quail (Colinus virginianus).

This study examined the effects of prenatal visual stimulation on bobwhite quail embryos' growth and heart rate. No differences in growth rate were found between embryos exposed to visual stimulation during the late prenatal period and control embryos. Embryos exposed to visual stimulation throughout incubation maintained lower heart rates in response to visual stimulation than did naïve embryos. In a subsequent experiment, naïve embryos that underwent an egg-opening procedure exhibited heart rates that were lower than embryos measured in intact eggshells. Embryos in opened eggs maintained lower heart rates than comparison embryos across time; however, a less invasive egg-opening procedure led to a quicker heart rate recovery than did a more invasive egg-opening procedure. These findings indicate that prenatal heart rate responses may be mediated by multiple features of the organism's developmental context, including intensity and duration of sensory stimulation.

Animals↗

Epicardial-like cells on the distal arterial end of the cardiac outflow tract do not derive from the proepicardium but are derivatives of the cephalic pericardium.

A series of recent studies strongly suggests that the myocardium of the cardiac distal outflow tract (d-OFT) does not derive from the original precardiac mesoderm but, instead, differentiates from a so-called anterior heart field. Similar findings were also reported for the endocardium of the d-OFT. However, very little information is available on the origin of the epicardium of the OFT. To address this issue, we have performed a study in which we have combined experimental in vivo and in vitro techniques (construction of proepicardial chimeras, proepicardial ablation, OFT insertion of eggshell membrane pieces, and culture on collagen gels) with molecular characterization techniques to determine this origin and define the properties of d-OFT epicardium compared with proepicardially derived epicardium. Our results demonstrate that the coelomic/pericardial epithelium in the vicinity of the aortic sac (and not the proepicardium) is the origin of d-OFT epicardium. This "pericardially" derived epicardium and the proepicardially derived epicardial tissues differ in their morphologic appearance, gene-expression profile, and in their ability to undergo epithelial-to-mesenchymal transformation. We conclude that the heterogeneity in the epicardial cell population of the OFT could be a factor in the complex developmental remodeling events at the arterial pole of the heart.

Animals↗

Genetic analysis of chorion formation in Drosophila melanogaster: I. The effects of one somatic-specific and seven germ-line-specific mutations.

Eight X-linked recessive female sterile mutations, derived from a hybrid dysgenic screen of Drosophila melanogaster and representing eight distinct loci, have been characterized by genetic and ultrastructural analysis. Four have abnormal respiratory appendages, three have essentially normal appendages but show moderate defects in the endochorion, and one mutant, fs(1)ne1a, exhibits major defects in both the endochorion and the respiratory appendages. Germ line clones of all eight mutants were generated using the dominant female sterile technique. Seven of the eight mutations are germ line specific, indicating that, although the eggshell is produced by the follicular cells, germ line functions play a significant role in its elaboration. The mutant that shows major defects, fs(1)ne1a, is somatic line specific, and exerts its effect in the ovary.

Animals↗

Localized Bicaudal-C RNA encodes a protein containing a KH domain, the RNA binding motif of FMR1.

The Bicaudal-C (Bic-C) gene of Drosophila melanogaster is required for correct targeting of the migrating anterior follicle cells and for specifying anterior position. Females lacking any wild type copies of Bic-C produce only eggshells open at the anterior end, because of the failure of the columnar follicle cells to migrate in the correct position at the nurse cell--oocyte boundary. Embryos which develop from eggs produced in females with only one wild type copy of Bic-C show defects in anterior patterning and an abnormal persistence of oskar RNA in anterior regions. We cloned Bic-C and found that, in ovaries, Bic-C RNA is expressed only in germline cells. Bic-C RNA is localized to the oocyte in early oogenesis, and later concentrates at its anterior cortex. The Bic-C protein includes five KH domains similar to those found in the human fragile-X protein FMR1. Alteration of a highly conserved KH domain codon by mutation abrogates in vivo Bic-C function. These results suggest roles for the Bic-C protein in localizing RNAs and in intercellular signaling.

Amino Acid Sequence↗

Zeolite A increases proliferation, differentiation, and transforming growth factor beta production in normal adult human osteoblast-like cells in vitro.

Silicon in trace amounts enhances bone formation, and the silicon-containing compound zeolite A (ZA) increases eggshell thickness in hens. In the studies reported here, treatment of nearly homogeneous strains of normal human osteoblast-like cells for 48 h with ZA at 0.1-100 micrograms/ml induced a dose-dependent increase (r = 0.35, P < 0.001) in DNA synthesis (n = 31) to 162 +/- 16% (mean +/- SEM) of control and in the proportion of cells in mitosis (n = 4) from 9.1 +/- 1.8 to 27.0 +/- 4.5% (r = 0.69, P < 0.005). ZA treatment also increased alkaline phosphatase activity (P < 0.05) and osteocalcin release (P < 0.05) but did not significantly affect collagen production per individual cell. The mitogenic action of ZA was dependent on cell seeding density over the range of 1250-40,000 cells per cm2, which is consistent with induction of an autocrine factor(s). TGF-beta is a potent mitogen for osteoblasts. ZA treatment increased the steady-state mRNA levels of transforming growth factor beta 1 (TGF-beta 1) and induced the release of the latent form of TGF-beta protein into the conditioned medium within 6 h. We conclude that ZA induces the proliferation and differentiation of cells of the osteoblast lineage.

Alkaline Phosphatase↗

From egg to pole cells: ultrastructural aspects of early cleavage and germ cell determination in insects.

Insect eggs are giant and very complex cells covered by an extremely resistant shell. Both the egg cell and surrounding eggshell express anteroposterior and ventrodorsal polarity. The molecular and cytoplasmic organization of both axes originates during oogenesis and leads to the production of an ooplasmic system which consists of euplasm and deutoplasm (yolk) and contains a nucleus as well as extranuclear determinants of maternal origin. Both are part of the store of information for early embryogenesis. In addition, the deutoplasm serves as raw material and early nutrient supply for building the embryo. The insect egg cell, which is arrested in the first maturation division when released from the ovary during oviposition, will be activated by different stimuli among different species to complete meiosis and start embryogenesis. The zygote nucleus undergoes a number of synchronous mitotic divisions leading to cleavage energids which initially form a syncytial blastoderm and subsequently the cellular blastoderm. In many insects, prior to blastoderm formation, polar granules (or oosome material) are incorporated in a single cell or a small number of cells which bud off at the posterior pole. These so called pole cells give rise to the primordial germ cells. Therefore, polar granules or the oosome material mark the germ line, and while structural counterparts of determinants of body pattern formation have so far not been found, the polar granules or oosome serve as an autonomous ooplasmic determinant for the pole or germ cells. Anteroposterior body polarity can arise independent of the germ plasm.

Animals↗