Ovalbumin is a component of the chicken eggshell matrix

The protein components of biomineralized structures (matrix proteins) are believed to modulate crystal nucleation and growth, and thereby influence the shape and strength of the final structure. The chicken eggshell contains a complex array of distinct matrix proteins. One of these was found to have similar molecular weight and chromatographic properties as purified egg ovalbumin. A commercially available antibody to ovalbumin was utilized for western blotting to demonstrate that ovalbumin is one of the matrix proteins this is extracted from decalcified eggshell. Immunohistochemistry revealed that ovalbumin is found only in the mammillary bodies of decalcified shell, and is not distributed throughout the shell matrix. These results indicate that ovalbumin is present during the initial phase of shell formation and becomes incorporated into the protein matrix of the mammillary bodies. However, it is not yet clear whether ovalbumin at this site plays a specific role in shell mineralization.     

Purification of ovocalyxin-32, a novel chicken eggshell matrix protein

The eggshell is a highly ordered structure resulting from the deposition of calcium carbonate and an organic matrix from the acellular uterine fluid. Characterization of the individual matrix components is necessary to determine their influence upon calcite crystal shape, size, and orientation during eggshell calcification. We have purified and sequenced a novel 32-kDa protein, ovocalyxin-32 (OCX-32), which is present at high levels in the uterine fluid during the terminal phase of eggshell formation, and is localized predominantly in the outer eggshell. Database searches identified expressed sequence tags (ESTs) whose alignment yielded the complete cDNA. OCX-32 protein possesses limited identity (32%) to two unrelated proteins: latexin, a carboxypeptidase inhibitor expressed in rat cerebral cortex and mast cells, and to a skin protein that is encoded by a retinoic acid receptor-responsive gene, TIG1. The timing of OCX-32 secretion into the uterine fluid suggests that it may play a role in the termination of mineral deposition.     

Taste bud development in chickens (Gallus gallus domesticus)

Oral epithelium in the anterior mandibular glands region was examined in embryonic, hatchling, and mature chickens to establish the timing of morphologic events during taste bud ontogeny. Hematoxylin-and-eosin-stained sections (10 microns) from 27 Anak (broiler breed) chickens were examined serially, and buds were quantified at 16-20 days of incubation (E) and, posthatch days 1 and 50-60. Taste buds were first recognized at the beginning of E17 as small clusters of cells in the basal epithelium. Only spherical-shaped buds were observed on E17 and E18, and these spherical clusters never penetrated to the surface of the stratified epithelial layer. E19 marked a transitional stage when mature bud features began to emerge: the buds assumed a more elongate shape, several kinds of cells comprising the bud were distinguishable and the first taste pores were observed. During the ensuing embryonic days, buds continued to elongate commensurate with the deepening oral epithelium and by hatching virtually all buds opened to the oral cavity. No marked morphological changes in taste bud structure were observed on the day of hatching and at 50-60 days posthatching. Taste bud numbers increased dramatically during E17 and E18, peaked on E19, and remained relatively constant thereafter. It is concluded that the morphological sequence of taste bud development in chickens is similar to that in mammals. The timing of bud ontogeny, though initiated only during the third trimester in ovo, essentially is completed by hatching, thus providing the precocial hatchling with the sensory apparatus essential for gustatory experience.     

Scanning electron microscope study of the spatial organization of the air and blood conducting components of the avian lung (Gallus gallus variant domesticus)

The lungs of the domestic fowl were prepared for scanning electron microscopy after vascular and airway latex rubber casting to demonstrate the spatial organization of the various structural components that are involved in the gas exchange that takes place in the parabronchial tissue mantle. The bulk of the intrapulmonary air flows through the parabronchial lumen and then centrifugally diffuses into the exchange tissue through the atria, the infundibula, and the air capillaries. The blood flows centripetally from the interparabronchial arteries, then into the intraparabronchial arterioles, and finally into the blood capillaries, which together with the air capillaries constitute the functional terminal gas exchange units. The relationship between the air flow in the parabronchial lumen and the incoming blood (into the exchange tissue) has been shown to be crosscurrent, where the directions of the flow of these two gas exchange media are essentially perpendicularly disposed to each other; whereas the relationship between the blood capillaries and the air capillaries is countercurrent, the blood flowing towards the parabronchial lumen and the air in the opposite direction, i.e., towards its periphery. Both these spatial structural relationships between the air and blood are significant factors that contribute to the remarkable efficiency of the avian lung in gas exchange.(ABSTRACT TRUNCATED AT 250 WORDS)     

Differential expression of extracellular matrix metalloproteinase inducer (EMMPRIN/CD147) in avian tibial dyschondroplasia

Tibial dyschondroplasia (TD) is an avian bone disorder of different aetiologies that may be associated with lameness. The disorder is characterized by focal disruption of endochondral bone formation, with a lack of matrix proteolysis and an accumulation of non-mineralized avascular cartilage. The aim of this study was to determine the expression of extracellular matrix metalloproteinase inducer (EMMPRIN/CD147) in normal, thiram-induced TD lesions and in the process of recovery from TD in broiler chickens. An extracellular matrix (ECM) degrading enzyme, matrix metalloproteinase-9 (MMP-9), was selected to investigate the effects of CD147 in the degradation of ECM. Gene expression was analysed by quantitative real-time polymerase chain reaction and protein levels by immunohistochemistry and western blotting. The birds were divided into three groups: thiram fed; recovery; and controls. Genes encoding CD147 and MMP-9 were down-regulated during the development of the disease, and were up-regulated during recovery. Western blotting also showed lower protein levels of CD147 in TD, which increased during the recovery phase associated with ECM degradation and growth plate repair. The findings of this study suggest that ECM has a crucial role in the occurrence of TD and that CD147 appears to play a pivotal role in matrix proteolysis in the chicken, similar to that in other species.     

Development of the supraoptic decussation in the chick (Gallus gallus)

Summary The developing supraoptic decussation (SOD), a major interhemispheric tract in birds, has been implicated in both transfer of visual information and in the modulation of brain asymmetry. Moreover little is known of its morphology during development. We have examined the development of the chick SOD, which consists of three subregions; dorsal, ventral and subventral SOD. In the dorsal SOD the total number of fibres reach 968000 on the 19th day of incubation (E-19), falling to 570000 by the 8th day after hatching (P-8). In the ventral SOD, the number of fibres at E-19 reach 660000, followed by a gradual reduction in their number to about 490000 at P-22. In the subventral SOD the number of fibres estimated was 87000 at E-15 falling to about 36000 at P-1. Compared with adult levels, there is, respectively, a drop in the number of fibres of 44%, 25% and 69% in the dorsal, ventral and subventral SOD during development. At E-19 in both the dorsal and ventral SOD there is qualitative evidence of axonal loss; disrupted axonal profiles, increased extracellular space and cells containing lysosomal cytoplasmic inclusions indicative of macrophages. Differences were also observed in the pattern of myelination, the dorsal, ventral and subventral SOD were shown to myelinate at different rates. Thus, in a single named tract, the SOD, there are major differences in the onset, rate and extent of fibre loss and myelogenesis within its three subregions. The functional implications of these differences are considered.     

Archistriatal lesions enhance tonic immobility in the chicken (Gallus gallus)

Chickens sustaining lesions in the archistriatum intermedium showed an 8-fold increase in the duration of tonic immobility as compared with control birds. Septal lesions had no significant effect when compared with sham operates. It was assumed that the tractus occipitomesencephalicus, a possible avian variant of the primate pyramidal tract was damaged. The behavioral effect found here supports the notion that the avian archistriatum is “conjunctive” cortex and that the archistriatum cannot be described as a homolog of the mammalian amygdala without qualifications.     

Congenital cerebellar dysplasia in White Leghorn chickens (Gallus gallus domesticus)

Congenital cerebellar anomalies have been rarely reported in birds. We examined cerebellums with disorganized folia from seven specific-pathogen-free White Leghorn chickens (Gallus gallus domesticus). Islands of heterotopic cortex were distributed from the deeper cortices to the medulla in the cerebellum. The characteristic lesions were composed of randomly admixed components of the cerebellar cortex, including Purkinje cells, a molecular layer and granular cells. Immunofluorescent analysis revealed Purkinje cells with haphazardly extended dendrites and a lack of Bergmann's glial fibres in the foci. Chicken parvovirus, Aino virus and avian retrovirus were not detected in the affected birds by polymerase chain reaction. This is the first report of cerebellar dysplasia in chickens possibly caused by a genetic abnormality.     

Ultramorphology of eggshell of flesh fly Liosarcophaga dux (Diptera: Sarcophagidae)

Eggs of Liosarcophaga dux (Thomson) (Diptera: Sarcophagidae) are elongated and slightly bean-shaped, measuring approximately 1.5 mm in length. Each is covered externally by an eggshell comprised of polygonal patterns. In this study, scanning and transmission electron microscopy were used to examine the eggshell of this species of flesh fly for the first time. No plastron region or median area was detected. Ultrathin sectioning of the eggshell revealed multiple layers in the shell that could be observed using transmission electron microscopy.     

Developmental changes in tonic immobility in young chicks (Gallus gallus)

Tonic immobility (TI) was induced by means of ventral restraint in White Leghorn chicks within 12-hr posthatch (Day 0) and, in independent groups, on succeeding days through Day 10. Durations of immobility and eye closure increased and incidence of immobility termination and vocalization decreased significantly with age. Marked qualitative changes in immobility reactions were also apparent over the first few posthatch days.     

Structure and origin of the eggshell ofQuinqueserialis quinqueserialis (Trematoda: Notocotylidae)

Ultrastructural studies were conducted on the eggshell and portions of the female reproductive system of the notocotylid trematodeQuinqueserialis quinqueserialis (Barker and Laughlin 1911). The eggshell consists of long bipolar filaments extending from an ovoid egg capsule. The egg capsule, formed within the ootype, is composed of an inner electron-dense layer of vitelline origin and an outer membranous layer containing many shallow pits derived from membranous secretions of the Mehlis' gland. Filaments are formed at the level of the fourth uterine loop from electron-dense secretions produced within the uterine epithelium. Filament elongation continues as eggs pass through the remaining uterine loops.     

Expression of fibroblast growth factor-2 (FGF-2) in early stages (days 3-11) of the development of the avian lung, Gallus gallus variant domesticus: an immunocytochemical study

In the avian lung, the bronchial system forms from epithelial (endodermal) cells. The intrapulmonary primary bronchus is the focal point of airway development. It originates secondary bronchi (SB) along its proximal-distal extent and parabronchi (tertiary bronchi) arise from and connect the SB. From as early as day 3.5, fibroblast growth factor-2 (FGF-2) is diffusely expressed in the epithelial and mesenchymal cells. Up-regulation of FGF-2 in discrete areas of the developing lung seem to set the growth rate, trajectories followed, areas appropriated, three-dimensional symmetry and coupling of the airways. Expressed early in development and persisting into the incubation period, FGF-2 may be involved in the formation of the avian lung. Morphogenetic differences between the avian and the mammalian lungs may explain the existing structural contrarieties.     

Ovotransferrin is a matrix protein of the hen eggshell membranes and basal calcified layer

The eggshell is an highly ordered structure deposited in the distal oviduct and composed of calcium carbonate and an organic matrix which is believed to influence its fabric. We have identified ovotransferrin as an 80 kDa matrix protein observed at high concentration in the uterine fluid at the initial stage of shell mineralization, by N-terminal sequencing and western blotting using monoclonal and polyclonal antibodies. It is present in extracts from demineralized eggshell and was localized by immunofluorescence in the eggshell membranes and mammillae, which are the sites of calcite nucleation. Northern blotting and RT-PCR demonstrated that ovotransferrin message was expressed in the proximal oviduct (magnum and white isthmus), and at a lower magnitude in the distal oviduct (red isthmus and uterus). Ovotransferrin was revealed by immunofluorescence in the tubular gland cells of the uterus. Calcium carbonate crystals grown in vitro in the presence of purified ovotransferrin showed large modifications of the calcite morphology. These observations and its presence in eggshell and membranes suggest a dual role for ovotransferrin, as a protein influencing nucleation and growth of calcite crystals and as a bacteriostatic filter to reinforce its inhibition of Salmonella growth in egg albumen.     

Neuronal morphology in subdivisions of the inferior colliculus of chicken (Gallus gallus)

The avian inferior colliculus (IC), also referred to as the nucleus mesencephalicus lateralis pars dorsalis (MLd), is an auditory midbrain nucleus that converges auditory cues from tonotopically organized brainstem nuclei. This information is relayed onto the optic tectum on the one hand and to nucleus ovoidalis on the other hand. Morphologically, there has been considerable debate about the number and nomenclature of the subnuclei within the IC. Here, we provide morphological characteristics of single cells in five IC subnuclei in chicken. The cellular structure within the IC was studied by whole-cell patch technique and biocytin iontophoresis. In addition, histological staining was performed, to delineate the borders between subnuclei of the IC. We were able to discriminate between 5 subnuclei: the core of the central nucleus (ICCc), the medial and lateral shell of the central nucleus (ICCms and ICCls), the external nucleus (ICX) and the superficial nucleus (ICS) of the IC. Our findings suggest the existence of at least two different morphologies of neurons with two subtypes each. The IC in chicken is a largely homogenous nucleus in terms of neuronal anatomy on a cellular level. However, its compartmentation into diversified subnuclei with different neurophysiological characteristics suggests a complex system to process auditory information. The auditory system in chicken is not as hypertrophied as in specialists such as the barn owl, but appears to have comparable connectivity and cellular morphology.     

Behavioral reactions to gustatory stimuli in young chicks (Gallus gallus domesticus)

Freely-moving, posthatch chicks were individually presented 2 concentrations each of quinine, citric acid, fructose, sucrose, sodium saccharin, and distilled water and their behavioral reactions were videotaped and analyzed. Already during the first posthatch day distinct rejection responses to quinine and citric acid could be recognized. Prolonged head shaking and beak clapping episodes were the most dominant features of these reactions. While responses to water and sweet stimuli could be interpreted as acceptance behaviors, the resolution was generally not fine enough to discriminate between reactions to the 2 different sweet concentrations of these stimuli or between them and water. When only water or sugar solutions were presented to other hatchlings in a single session, there was a suggestion of more definite acceptance behavior to some sweet stimuli as compared to water. It is concluded that the systems mediating aversive gustatory responses are present and functioning in posthatching chicks while acceptance responses, though present, are less discriminative among stimuli.     

Haematological changes in domestic fowl (Gallus gallus) and cranes (Gruiformes) with Mycobacterium avium infection

Haematological findings in 26 domestic fowl and 17 cranes with confirmed Mycobacterium avium infection were compared with normal values from the species concerned. Heterophils and monocytosis occurred frequently in the infected birds; in many cases very large numbers of these cells were present. Haematological findings were related, where possible, to the results of the intradermal tuberculin test and the haemagglutination test. It was concluded that haematological testing has a useful role in both diagnosis and screening for avian TB infections in the species studied.     

Vitellogenesis and eggshell formation in Caryophyllaeus laticeps (Pallas) and Caryophyllaeides fennica (Schneider) (Cestoidea: Caryophyllaeidea)

Summary Eggshell formation in Caryophyllaeus laticeps and Caryophyllaeides fennica (Cestoidea: Caryophyllaeidea), as shown by histochemical tests for proteins, phenols, and polyphenol oxidase, is similar to that of D. latum, Ligula sp., and Schistocephalus sp.Maturation of vitelline cells is characterized by (1) an increase in size of two or three times, (2) vacuolation of the nucleus with the nucleolus and a small amount of nucleoplasm and dispersed chromatin displaced to one side, and in the cytoplasm, (3) synthesis of shell globules that are later released in the uterus to form the shell.Tests for glycogen (PAS) and nucleic acids show that maturing and mature cells have glycogen in the nuclear vacuole and cytoplasm and that there is a decrease in DNA in the nucleus and an increase in RNA in the cytoplasm as protein synthesis is initiated.Glycogenesis and protein synthesis appear to be largely completed before the cell is released from the follicle.

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Zusammenfassung Die Eischalenbildung bei Caryophyllaeus laticeps und Caryophyllaeides fennica (Cestoidea: Caryophyllaeidea) ist — wie die histochemischen Untersuchungen auf Proteine, Phenol und Polyphenoloxidase zeigen — ähnlich der bei D. latum, Ligula spec. und Schistocephalus spec. Die Reifung der Dotterzellen ist charakterisiert 1. durch eine Vergrößerung auf das Zwei- bis Dreifache, 2. eine Vakuolisation des Zellkerns mit Nukleolus und einer kleinen Menge von Nukleoplasma und fein verteiltem Chromatin, das sich auf eine Seite verlagert, und im Cytoplasma, 3. Synthese der Schalenkügelchen, die später in den Uterus zur Schalenbildung freigesetzt werden.Die Teste auf Glykogen (PAS) und Nukleinsäuren zeigen, daß die reifenden und die reifen Zellen in den Kernvakuolen und Zytoplasma Glykogen enthalten, und daß eine Verminderung der DNS im Nukleus und eine Vermehrung der RNS im Zytoplasma auftritt, wenn die Proteinsynthese beginnt.Die Glykogenese und die Proteinsynthese erscheint im wesentlichen abgeschlossen zu sein, bevor die Zelle aus dem Follikel entlassen wird.

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This investigation was started at Neuchâtel and finished at Albany.

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NIH Postdoctoral fellow.     

Tissue tropism of a Thailand strain of high-pathogenicity avian influenza virus (H5N1) in tissues of naturally infected native chickens (Gallus gallus), Japanese quail (Coturnix coturnix japonica) and ducks (Anas spp.).

The tropism of a Thailand strain of highly pathogenic avian influenza H5N1 virus was demonstrated on tissues (lung, trachea, heart, liver, spleen, pancreas, rectum, kidney, brain, skeletal muscle, duodenum, and oviduct) from naturally infected native chickens (Gallus gallus), Japanese quail (Coturnix coturnix japonica) and ducks (Anas spp.) by indirect immunofluorescence assay. In chickens and quail, the distribution and localization of nucleoprotein viral antigen was similar and detected at the highest level in cardiac myocytes, at 88% (chickens) and 89% (quail), and respiratory, digestive and urinary systems all showed high levels of antigen. Antigen in duck tissues were detected at significantly lower levels (P < 0.05) with the exception of brain and skeletal muscle samples. In most cases, antigen in duck tissue was absent in the digestive organs but present in respiratory organs, which supports the hypothesis that aerosol and oral-oral transmission are the main method of highly pathogenic avian influenza virus transmission from this species.     

Ultrastructure of eggshell and embryological development of Salvator merianae (Squamata: Teiidae)

The objective of this study was to characterize the external morphology of Salvator meriane embryos in different stages of embryonic development and establish a relationship with the ultrastructure of the shell from oviductal transit to hatching. A total of 120 embryos were analyzed to describe their external morphology, and 78 eggs were used for the analysis of the shell. For embryonic development, the series was established according to the total length of the body. We established 40 embryonic stages from the primitive streak. In the early stages, the external morphological features are the C-shaped body, the maxillary, and mandibular fusion processes with the frontal process and the fusion of the forelimb with the digital plate. In the middle stages, the eyelid appears, and there are claws on the toes, cornification of fingers, and the onset of pigmentation. The last stage of embryonic development is characterized by the beginning of the formation of the scales, appear the toenails, and finalize the entire pigmentation. Regarding the relationship that exists with the ultrastructure of the egg during development, it was possible to observe a marked change in the composition of the shell and well-marked compaction during embryonic development, which may be related to the transport of calcium during embryonic ossification. Our results allowed us to show the complete sequence of embryonic development, determining the laying stage for this species. It was possible to establish a relationship with the ultrastructure of the eggshell from the oviductal transit to the moment of hatching.