Mammillary Cores Research Articles

New zonal structure and transition of the membrane to mammillae in the eggshell of chicken Gallus domesticus

Avian eggshell is a typical bio-engineered ceramics characterized by layer structures. These layers are categorized mainly by the form of crystalline calcite. Whether there exist other layer structures, how the membrane layer is transformed to the carbonate one, what form the carbonate takes after the transition. These questions remain to be clarified. Here we examine the eggshell of chicken Gallus domesticus by optical microscope, scanning electron microscope and transmission electron microscope. We find that there exists another layer structure defined by variation of organic matrices. The transition from the membrane to the mammillary cones is implemented through the calcium reserve assemblies or the mammillary cores. The integrity of the transitional structure was weakens as the reserved calcium is displaced, and loses completely in about 10 days of incubation. As the first deposited carbonate layer after the transition, the mammillary cones comprise amorphous calcium carbonate and clusters/assemblies of calcite crystallites the size about a nanometer, plus bubble pores extending preferentially in the lateral direction. Our results provide new insights into the structure and component of the avian eggshell, and may help decipher the constitution of the bio-ceramics in the perspective of material science.

Amorphous calcium carbonate controls avian eggshell mineralization: A new paradigm for understanding rapid eggshell calcification.

Avian eggshell mineralization is the fastest biogenic calcification process known in nature. How this is achieved while producing a highly crystalline material composed of large calcite columnar single crystals remains largely unknown. Here we report that eggshell mineral originates from the accumulation of flat disk-shaped amorphous calcium carbonate (ACC) particles on specific organic sites on the eggshell membrane, which are rich in proteins and sulfated proteoglycans. These structures known as mammillary cores promote the nucleation and stabilization of a amorphous calcium carbonate with calcitic short range order which predetermine the calcite composition of the mature eggshell. The amorphous nature of the precursor phase was confirmed by the diffuse scattering of X-rays and electrons. The nascent calcitic short-range order of this transient mineral phase was revealed by infrared spectroscopy and HRTEM. The ACC mineral deposited around the mammillary core sites progressively transforms directly into calcite crystals without the occurrence of any intermediate phase. Ionic speciation data suggest that the uterine fluid is equilibrated with amorphous calcium carbonate, throughout the duration of eggshell mineralization process, supporting that this mineral phase is constantly forming at the shell mineralization front. On the other hand, the transient amorphous calcium carbonate mineral deposits, as well as the calcite crystals into which they are converted, form by the ordered aggregation of nanoparticles that support the rapid mineralization of the eggshell. The results of this study alter our current understanding of avian eggshell calcification and provide new insights into the genesis and formation of calcium carbonate biominerals in vertebrates.

Changes in eggshell ultrastructure of Falco naumanni and Tyto alba exposed to pesticides and polychlorinated biphenyls (PCBs)

Changes in the quality of eggs of birds exposed to environmental contaminants have been described, but few reports concern eggshell ultrastructure. In this study, infertile or addled Lesser Kestrels (Falco naumanni) and Barn owls (Tyto alba) eggs were collected from the polluted area of Gela plain (Sicily) during 2007, and compared in terms of organophosphate and organochlorine pesticides, and PCBs levels, and eggshell ultrastructure as determined by scanning electron microscopy. Pesticide and PCB residues in eggs were determined by Gas chromatography/ mass spectrometry (GC/MS) [GC Agilent 7890A/MS Agilent 5975C (Agilent technologies) using a DB-5 capillary column in the selected ion monitoring mode]. The GC/MS analysis revealed that eggs contained measurable amounts of some pesticides and PCBs. There was a low detection of organophosphate pesticides while the most abundant organochlorine residues detected were p,p’ DDT, p,p’ DDE, and Hexachlorobenzene. While, the most abundant PCBs detected congeners were PCB 138, 153, 170, 180, and 187. Although the general structure of the eggshell layers was maintained, the results showed ultrastructural differences in mammillary and palisade eggshell layers between high level and low level contaminated eggs in Tyto alba. Furthermore, mammillary cores of the eggshell had an increased distance between themselves with respect to well organized structures present in uncontaminated egg. In this paper we verify the presence of environmental contaminants in the eggs and document structural changes in bird of prey eggshells. The data could suggest that some contaminants can contribute to reduced reproductive performance (infertile or addled egg) by structural changes in the eggshell. The alteration in morphological disposition of mammillary cores could also suggest an impairment of gas exchange.

The Effect of Manganese Deficiency upon the Ultrastructure of the Eggshell

Manganese deficiency in the laying hen results in decreased egg production and eggshell thickness. Eggshells from deficient hens show specific changes in appearance characterized by translucent areas. Examination of these translucent areas with the scanning electron microscope revealed changes in the ultrastructure of the shell. The most prominent feature was large irregular mammillary knobs probably due to the fusion of several mammillary cores during the early phases of eggshell formation.Chemical analysis of the organic matrix of the eggshell showed a decrease in hexosamine and hexuronic acid content consistent with the known role of manganese in polysaccharide synthesis. Studies on the characteristics of the polysaccharide chains isolated from normal eggshells suggested that these polysaccharides were considerably different from those isolated from cartilage proteoglycans.

Biochemistry of the Organic Matrix of the Eggshell

INTRODUCTIONAlthough the organic material accounts for a very small portion of the eggshell, recent evidence suggests the perterbations in synthesis and secretion of this organic material can severely impair eggshell formation. The organic matter of the eggshell can be divided into at least four distinct fractions based upon chemical, histochemical, and morphological characteristics. These are shell membranes, mammillary cores, shell matrix, and cuticle. This discussion will emphasize the biochemistry of the first three organic constituents of the eggshell.SHELL MEMBRANESThe shell membranes are characterized by having two layers, which have been designated as inner and outer membranes. Figure 1 shows a scanning electron micrograph of the eggshell membranes. This meshwork-like structure is characterized by having fibers arranged in a random fashion. On these fibers are small fiber protuberances, the function of which is unknown at the present time. When the fibers are viewed with the transmission electron microscope, .

Structure of the Eggshell

The ultrastructure of the eggshell of the domestic fowl is reviewed with regard to the influence that the various shell layers have on determining eggshell quality. Emphasis is placed on the importance of normal mammillary knob formation. Eggshells having good eggshell quality have a regular distribution of mammillary cores. These initiate the production of many small mammillary cap and cone formations. Early fusion of the forming mammillary knobs is a key to eggshell strength through a reduction in the penetration of intermammillary clefts into the calcified eggshell. The formation of narrow palisade columns is implicated in this process. The impact of the membranes, vertical crystal layer, and cuticle is also discussed.

Use of Low Viscosity Expoxy Plastic Casts to Study Mammillary Core and Mammillary Knob Organization of Avian Eggshells

A technique has been developed to facilitate scanning electron microscopic analysis of eggshell mammillary formations with minimal physical disturbance to the shell. The method employs a low viscosity epoxy resin to make a negative replica of the mammillary surface. The cast can be viewed after decalcification using the scanning electron microscope without further physical manipulation which may cause structural artifacts.

P,p′-DDE-induced changes in the organic and inorganic structure of eggshells of the Mallard, Anas platyrhynchos

Scanning electron microscopic examination of decalcified and inorganic preparations of Mallard ( Anas platyrhynchos) eggshells showed that dietary p,p′-DDE induced significant and consistent changes in both the organic and inorganic constituents of the eggshell. The numbers of mammillary cores, structures composed of organic knobs which serve as crystallization sites on the outer surface of eggshell membranes, as well as the numbers of basal caps, were reduced by dietary p,p′-DDE in eggshells which also showed a reduction in shell thickness. Polychlorinated biphenyls induced a lesser but significant reduction in the numbers of mammillary cores; a comparable decrease in numbers of basal caps was not found to be statistically significant with the sample sizes used. Effects of a diet containing both DDE and PCB were equivalent to those of DDE. Thickness of eggshells from mallards maintained on the PCB diet was equivalent to that of the controls. The reduction in the number of mammillary cores may therefore occur independently of the DDE-induced eggshell thinning. The density of mammillary cores on the eggshell membranes was found to be linearly related to the density of basal caps.

Ultrastructure of The Mammillary Region of Low Puncture Strength Avian Eggshells

The mammillary region of poor quality avian egg shells displays a number of aberrant structural alterations not commonly observed in high quality egg shells. Three general categories of mammillary alterations were observed in radial sections of extremely low quality shells: 1) Shells with a proliferation of discrete rounded calcified bodies situated upon the shell membrane fibers; 2) shells with an extremely disorganized, multinucleate mammillary region without numerous calcified bodies; and 3) mammillary knobs possessing cavernous tips without mammillary core formations.Adhesion discs (fracture surfaces between adjacent calcified structures) were observed and classified into two types, small-rounded (Type I) adhesion discs between adjacent calcified bodies, or between calcified bodies and larger mammillary formations; and large oval (Type II) adhesion discs between adjacent mammillary knob formations. The nucleation zone or external surface of the outer shell membrane fibers, upon which calcium salts initially precipitate, was also observed to vary in low-quality shells. Waves or bulges in the nucleation zone were observed directly beneath the multinucleate unorganized mammillary knob formations.These findings suggest that the initial process of shell mineralization (mammillary knob formation) is responsible for the base upon which the palisade (main shell) layer is deposited. Disorganization of the basal calcified region may give rise to poorly indigitating calcite (palisade) columns and decreased intrinsic shell strength.

Structure and relationship of the mammillary gore to membrane fibres and initial calcification of the avian Egg Shell1

1. The interior of the mammillary knob is shown to be composed of: (1) outer shell membrane fibres that penetrate the basal portion of the knob and become associated with (2) The mammillary core, and (3) the bulk calcification of the main knob. 2. The mammillary knob consists of radial crystals which are arranged in a hemisphere on the outer shell membrane fibres, and are composed of a central zone which is a homogeneous solid, crystalline in nature, and an outer heterogeneous vesiculated zone. 3. The walls of the mammillary knob are continuous with the base and sides of the mammillary core radial crystals; the dorso‐central zone of the core crystals are exposed to a cavity that communicates with the upper zones of the mammillary knob and palisade layer via the central channel. The dorsal surface of the core cavity is formed by a layer of unorganised crystals of the mammillary knob proper. 4. Initial calcification of the knob begins at the base and sides of the mammillary core and expands over the organic...

Studies of membrane, mammilary cores and cuticle of the hen egg shell

Synopsis Mammillary cores and outer shell membrane have been studied by comparing the weight and chemical composition of outer membrane with cores still attached and of outer membrane free from cores, from the same shells. Inner membrane and cuticle have also been investigated. The weights of inner and outer membrane are significantly correlated (P < 0.05). Both membranes contain galactosamine, glucosamine, sialic acid, glucose, mannose and fucose but in different proportions. Uronic acid is probably absent. An average of 3.3 mg of organic mammillary core material per shell (52 μg/cm2) was present in the shells investigated, but there was a trend suggesting that eggs laid by more mature hens may contain rather more. Cores are rich in hexo‐samine, sialic acid and hexose, but the probable absence of uronic acid suggests that chondroitin sulphates play no part in shell initiation. There were no major differences in the amino acid spectrum of membrane samples with and without cores. Galactosamine, glucosamine...

The structure of the organic mammillary cores in some weak egg shells

Synopsis The microscopic structure of the outermost layer of the outer shell membrane containing decalcified mammillary cores of some of the egg shells laid by a large flock of Shaver 288 Starcross hybrids from November 1967 to September 1968 has been examined. It has been found that a high percentage of the weak and thin egg shells contain abnormal organic mammillary cores. The results are discussed with special reference to the recent findings of other workers who have studied the crystalline structure of shell concretions and egg shells laid by hens under climatic stress.

Salmonella Penetration of Fertile and Infertile Chicken Eggs at Progressive Stages of Incubation

The shell and shell membrane systems of fertile chicken eggs undergo marked changes during incubation and embryonic development. Tyler and Simkiss (2) described these changes and confirmed that the membranes separate from the shell of fertile eggs (except for that portion over the air cell) by about the 15th or 16th day of incubation. The outer membrane does not come away cleanly from the shell but carries with it the ends of the mammillary knobs, including the mammillary cores. In earlier studies (4) of the penetration patterns of salmonella organisms through the outer structures of chicken eggs, the incubation periods used most frequently have been short periods up to 24 hrs at 990F, or eggs have been exposed to salmonella and stored for extended periods before examination. The work reported herein was undertaken as a systematic investigation of salmonella penetration into fertile hatching eggs at various stages of embryonic development, in comparison with penetration of infertile eggs maintained under identical conditions of incubation.

The occurrence of neutral and acidic mucins in the reproductive tract of the laying hen.

The tissues of the domestic hen's reproductive tract have been shown to contain both acidic and neutral mucins. The presence of an egg in the isthmus was always accompanied with an increase in the concentration in the isthmus secretory epithelium of PAS-positive mucins without free acidic groups. Staining of uterine tissue for strongly acidic groups with basic dyes was greater when an egg had been present in the shell gland at the time of death and the same was true for tissues of the upper oviduct. The results are discussed with special reference to the periodic secretion of organic mammillary core and egg shell mucins.

Studies on egg shells. XII.—Some changes in the shell during incubation

AbstractThe changes taking place in egg shells during the course of incubation have been studied, using histochemical tests and plastic embedding techniques. It has been confirmed that the membrane comes off all the shell, apart from that over the air space, by about the 15th or 16th day. the membrane does not come away cleanly from the shell but carries with it the ends of the mammillary knobs, including the mammillary cores. At the end of incubation there is still a great deal of calcium carbonate present on the membrane. These results are discussed in relation to those of previous workers on the subject.

Studies on egg shells. VIII.—The determination of membrane‐, pore‐ and matrix‐protein—a reassessment

AbstractThe first paper in this series suggested methods for determining membrane‐, pore‐ and matrix‐protein in egg shells. Further studies by means of histochemical methods and by the etching of plastic‐embedded specimens have shown that the method for membrane protein is sound. On the other hand, although the true shell contains nitrogen fractions, one of which is soluble in 10% sodium hydroxide and the other not, the allocation of the names ‘pore’ and ‘matrix’ protein respectively to these fractions is wrong. The alkali treatment removes the mammillary cores, the cuticle and part of the matrix of the spongy layer. Further, the organic matter of the shell contains acid mucopolysaccharide as well as protein. It is, therefore, recommended that the terms ‘pore’ and ‘matrix’ protein should be replaced respectively by the terms ‘soluble’ and ‘insoluble’ nitrogen fraction.