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Organic matrix morphology and distribution in the palisade layer of eggshells sampled at selected periods during lay.

1. 1 cm2 pieces of eggshells from a commercial battery flock were plasma etched to remove the outer shell membranes. 2. They were decalcified using EDTA (200 g/l, pH 6.9 to 7.0) in paraformaldehyde (20 g/l) and 25% gluteraldehyde (20 ml in 0.98 l) in phosphate buffer, then prepared for light and transmission electron microscopy. 3. Light microscopy revealed a differential distribution of matrix material within all 3 regions of the palisade layer at the beginning of lay. 4. Transmission electron microscopy revealed a more even distribution of matrix at the beginning of lay, although morphological differences were observed. At the end of lay all 3 regions showed an increase in % matrix and vesicles/10 cm2 of micrograph compared to the middle and beginning of lay periods. 5. It is hypothesised that matrix vesicles are involved in the regulation of the physiochemical environment within the forming eggshell and that the decline in shell quality associated with the end of lay is related to a concomitant change in matrix quality.

Animals↗

Features of eggshell formation in guinea fowl: kinetics of shell deposition, uterine protein secretion and uterine histology.

1. Rate of calcium carbonate deposition, duration of eggshell formation, organic composition of the uterine fluid, morphology of the egg shells and histochemistry of the uterus were studied in guinea fowl to analyse the origin of such thick, strong egg shells. 2. The egg shell was linearly deposited from 6.4 h to 21.8 h after the oviposition of the previous egg. The rate of egg shell deposition was similar to that in laying hens. However, the duration of linear shell deposition was increased by 2.1 h relative to that in hens. This explained the increased egg shell weight observed in the guinea fowl. 3. Intervals between oviposition of intra-clutch eggs were 24 h throughout the laying period. Ovulation occurred just after oviposition of the previous egg in the guinea fowl, as previously observed in hens but the duration of egg white protein deposition, of plumping and of initiation of shell mineralisation were all 1.5 h shorter than in domestic hen. 4. Uterine fluid can only be collected during the growth and terminal phase of shell formation. The electrophoretic profiles of the uterine fluid differed between phases and were somewhat different from those previously observed in the hen. Ovalbumin and ovocleidin-17 were both present in the uterine fluid and also in egg shell extract. Ovocleidin-17 was predominant during the growth phase. 5. The histology of the uterus differed slightly in guinea fowl compared to hens. Ovocleidin and ovalbumin are both secreted by the tubular glands. 6. Examination of radial ultrathin sections of eggshell showed, above the mammillary layer, intricate interlacing of adjacent exospherite in guinea fowl in contrast to the continuous columnar microstructure in hens. 7. The kinetics of egg shell deposition largely explains the increased egg shell weight of guinea fowl. The organic matrix proteins may be associated with the contrast between the structural organisation of the guinea fowl egg shell and that of the hen egg shell.

Animals↗

Improving eggshell quality at high temperatures with dietary sodium bicarbonate.

Two experiments were conducted that confirmed the hypothesis that a dietary bicarbonate supplement will improve eggshell quality in hens at high temperatures as long as feed is consumed during the period of eggshell formation. End-of-lay hens were maintained on continuous light at temperatures of 30 and 35 C. Individual egg weights and shell quality measures for each hen were calculated as a proportion of the initial values determined during an acclimatization period at 25 C. Improvements in shell breaking strength in both experiments were observed as a result of supplementing control diets with 1% sodium bicarbonate (NaHCO3). This response to NaHCO3 was not a reflection of a reduced rate of lay or egg mass output, as these were similar or inferior on the control diets. Similar feed intakes on the control and NaHCO3 diets indicated that the response was not related to differences in calcium intakes. Supplements of zinc methionine and ascorbic acid proved to be inferior to NaHCO3. Improvements in egg weight were associated with the introduction of continuous lighting.

Acclimatization↗

Effects of dietary fat and eggshell cuticle removal on egg water loss and embryo growth in broiler hatching eggs.

The effects of dietary fat and eggshell cuticle removal on egg water loss, embryo growth, and hatchability were determined in eggs from broiler breeder hens at different ages. Hens were fed isocaloric diets containing one of three different types and levels of added fat. In addition, eggs were either left intact or washed to remove the eggshell cuticle prior to set. Cuticle removal increased egg water loss between 43 and 62 wk. Cuticle removal increased relative wet embryo weight at Week 52 and relative dry embryo weight at 52 and 62 wk. Furthermore, at 62 wk, diet and day of incubation interacted to affect wet embryo weight, and diet variably affected dry embryo weight. No treatment differences were observed for cumulative hatchability, rate of hatch, and relative yolk sac weight at Day 19 of incubation. It was concluded that cuticle removal and the addition of fat to breeder diets may influence embryonic growth without having any subsequent effects on hatchability.

Animal Feed↗

Paternal influences on turkey embryonic growth in the absence of changes in egg weight and eggshell conductance.

The hypothesis was proposed that embryonic growth could be influenced paternally in the absence of changes in egg weight or eggshell conductance. Sire families were established by selecting 11 sires based on blood sampling of approximately 50 commercial turkey males. Southern blot analysis was used to identify the most distantly related individuals in the subpopulation. Five hen siblings or half-siblings of the males were randomly assigned to each male to randomize effects of egg size and eggshell conductance. Artificial inseminations were performed weekly for 17 wk, using each sire with the assigned hens. Fertilized eggs were collected and set in incubators at biweekly intervals. Offspring were observed at the time of hatching for sex, body weight, and blood glucose concentration. Significant differences among sires were observed at hatching for poult weights and blood glucose concentration, and a significant correlation between the BW and glucose concentration was noted. Progeny of sires with the highest and lowest BW and blood glucose concentrations were subsequently compared. Mean BW of poults from different sires differed by as much as 10 g, although they were hatched from eggs of the same weight. Elevated blood glucose was associated with heavier BW; heavier heart, liver, and muscle weights; a rapid utilization of glycogen at pipping; and increased gluconeogenesis as measured by plasma organic acids and glucose-6-phosphatase activity. We conclude that poult embryonic growth differs even when not mediated by egg size and functional characteristics.

Animals↗

Mineral, amino acid, and hormonal composition of chicken eggshell powder and the evaluation of its use in human nutrition.

Chicken eggshell powder (ESP) might be an attractive source of Ca for human nutrition. To study its nutritional value, we analyzed minerals, amino acids, and hormones in commercially available Slovakian ESP. The mineral composition was compared with three Dutch ESP samples that differed in feed and housing, a Japanese ESP, refined CaCO3, and an oyster shell supplement. Chicken eggshell powder contains high levels of Ca (mean +/- SD/g EPS: 401+/-7.2 mg) and Sr (372+/-161 microg) when compared with recommended or estimated daily intakes for humans 51 to 70 yr of age. Levels of potentially toxic Pb, Al, Cd, and Hg were very low as were levels of V, B, Fe, Zn, P, Mg, N, F, Se, Cu, and Cr. Large differences in the levels of F, Se, Cu, Cr, and Sr in the Dutch and Slovakian ESP indicated a strong influence of feed and environment. The small protein fraction of ESP contains high levels of Gly and Arg. Furthermore, small amounts of transforming growth factor-beta1 (0.75 to 7.28 ng/g ESP), calcitonin (10 to 25 ng/g ESP), and progesterone (0.30 to 0.33 ng/g ESP) were detected. Estradiol-17beta and calcitriol were below the detection limit of the methods used. Compared with ESP, refined CaCO3 was found to contain increased levels of Cd, and the oyster shell supplement showed increased levels of Al and Cd. Therefore, ESP seems to have a beneficial composition with about 39% of elemental Ca, relevant amounts of Sr, and low levels of Al, Pb, Cd and Hg. It may be used as a Ca source in human nutrition.

Aged↗

Pleistocene geochronology and palaeothermometry from protein diagenesis in ostrich eggshells: implications for the evolution of modern humans.

Proteinaceous residues incorporated within the crystal structure of ostrich eggshells (OES) are retained without loss over geological time exceeding 10 million years. Degradation of the polypeptides, including hydrolysis to smaller peptide fragments and eventual release of free amino acids, decomposition, and racemization and epimerization occur at regular, predictable rates dependent on ambient temperature. The extent of isoleucine epimerization (aIle/Ile ratio) in OES follows linear first-order reversible kinetics in controlled-temperature laboratory simulations of time up to an aIle/Ile ratio in excess of 1.0. The hydrolysis of leucine also follows a predictable pattern, but deviates from first-order kinetics. A nonlinear mathematical model has been developed that adequately describes the pattern of leucine hydrolysis through a wide temperature range. Arrhenius parameters were derived from laboratory experiments combined with rate constant values found for 14C-dated OES from stratified caves in southern Africa. These parameters for isoleucine epimerization and leucine hydrolysis differ by ca. 10%, allowing the simultaneous solution of the two equations for temperature, independent of sample age. Although the uncertainty of the simultaneous temperature is relatively high (+/- 10 degrees C), it provides an effective means of identifying burned samples. If sample age is known, palaeotemperatures (the integrated thermal history experienced by an eggshell as opposed to an 'instantaneous' temperature) can be calculated with a precision of better than +/- 1 degrees C. The ages of levels at Border Cave, South Africa, from which anatomically modern human skeletal remains have been recovered, are dated by the extent of isoleucine epimerization in associated OES.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Cloning and analysis of the dec-1 female-sterile locus, a gene required for proper assembly of the Drosophila eggshell.

Female-sterile mutations at the dec-1 (defective chorion-1) locus of Drosophila severely disrupt the organization of the eggshell late in oogenesis. Previous characterization of dec-1 mutations has correlated the defects with failure to accumulate an early eggshell protein that undergoes proteolytic cleavage during choriogenesis. To enable further study of the regulation and processing of dec-1 products, we have molecularly cloned the locus and characterized its transcripts. Chromosome jumping was used to isolate a deficiency breakpoint within the locus. Overlapping genomic clones from a wild-type library were then obtained, and a region including the dec-1 locus was identified by hybridization to cDNA probes complementary to RNA from stage 9-10 egg chambers. Analysis of genomic rearrangements associated with the locus verified its identity. Two transcripts from the locus have been identified and characterized using cDNA clones, RNase protection, and primer extension analyses. A 4.0-kb transcript accumulates maximally in stages 9-10, when the primary follicle cell protein associated with dec-1 mutations is synthesized. A second transcript of 5.8 kb, generated by alternative splicing, accumulates during stages 11-12. These results are discussed in light of previous analysis of dec-1 mutations.

Animals↗

Isolation of messenger RNA coding for eggshell protein of the DNA-eliminating nematode Ascaris lumbricoides.

Poly(A)-containing RNA from polyploid uterine epithelial cells of Ascaris lumbricoides has been isolated by poly(U)-Sepharose chromatography. The bulk of poly(A)-containing RNA migrates as 18-S RNA in formamide/polyacrylamide gels. In a cell-free wheat germ system, this RNA directs the synthesis of a polypeptide with identical migration behavior in dodecylsulphate/urea/polyacrylamide gels as the polypeptide isolated from the proteinaceous eggshell. The two proteins reveal almost identical peptide patterns in fingerprint analysis. The authentic eggshell protein has been identified as a glycoprotein with a molecular weight of about 10000, as determined by dodecylsulphate/polyacrylamide gel electrophoresis. The apparent discrepancy between mRNA length and the required coding length for the protein is discussed.

Animals↗

The inhibitory effect of some chlorinated hydrocarbon pesticides on the ATP-dependent Ca2+ binding of the particulate fraction of the eggshell gland mucosa cells.

The pesticide p-p'-DDT and its persistent metabolite p-p'-DDE cause thinning of the eggshells in several species of birds. In earlier investigations on ducks this thinning was found to be associated with a reduction of the ATP-dependent Ca2+ binding to a homogenate of the shell gland mucosal cells by DDE. The activity of a Ca2+-Mg2+-activated ATPase in the homogenate was also decreased on administration of DDE in vivo. We have therefore investigated the in vitro effects of some other chlorinated hydrocarbon pesticides of ecotoxicological interest on the ATP-dependent Ca2+ binding and the Ca2+-Mg2+-activated ATPase activity in a homogenate of the eggshell gland mucosa of the hen and determined the molar concentrations that produced 50% inhibition (=IC50). Several of the investigated compounds, namely toxaphene, chlordane, p-p'-DDD, o-p'-DDE, p-p'-DDT, methoxychlor and PCB (Arochlor 1242), had a similar IC50 to inhibit the Ca2+ binding as p-p'-DDE. Lindane, p-p'-DDA and biphenyl had an IC50 3.3-4 times higher and that of 2.4 D was 13.5 times higher than that of p-p'-DDE. When the IC50 of some of the compounds (p-p'-DDE, PCB, toxaphene, Lindane) was determined that decreased the Ca2+-Mg2+-activated ATPase of the homogenate it was found to be only 18 to 29 per cent of that needed to inhibit the Ca2+ binding by the homogenate. It is therefore probable that some other effect than inhibition of this enzyme is also involved in the Ca2+-binding process and affected by the compounds.

Adenosine Triphosphatases↗

Effect of some metal compounds on the Ca2+ binding and Ca2+-Mg2+-ATPase activity of eggshell gland mucosa homogenate from the domestic fowl.

Some metal ions, e.g. Hg2+, Cd2+ and Al3+, can have the effects as ecotoxicological agents, of causing eggshell thinning and breakage in birds. In a homogenate of the Ca2+-secreting part of the eggshell gland mucosa, a study was made of the influence of Hg2+, Cd2+, Cu2+, Pb2+, methyl-Hg+, Zn2+, V3+, Al3+ and Ni2+ in different concentrations on the rate of ATP-dependent 10(-4) M Ca2+ binding. All compounds had an inhibitory action. The most potent metal (Hg2+) produced 50% inhibition (IC50) at 1.1 X 10(-6) M, whereas this value for the least potent compound (Ni2+) was 9 X 10(-4) M. The specific Ca2+-Mg2+-ATPase activity was also inhibited by the tested metal ions. In all cases except methyl-Hg+ the IC50 for this activity was lower than that for Ca2+ binding. The most potent ion in this respect was Cd2+, with an IC50 of 8 X 10(-8) M, and the least potent was methyl-Hg+, with an IC50 of 1.4 X 10(-3) M. Pb2+ and Cd2+ in a concentration range of 10(-5)-10(-4) stimulated the Mg2+-ATPase activity, however, to almost the same extent as 10(-4) M Ca2+. A possible explanation for this effect is that these ions may have an affinity for sites of Ca2+ binding of the polypeptide calmodulin and thereby influence the Ca2+ metabolism of the shell gland mucosa.

Animals↗

DDE residues and eggshell changes in Alaskan falcons and hawks.

Eggshell thickness after exposure to DDT was reduced by 21.7 percent in Alaskan tundra peregrines, by 16.8 percent in taiga peregrines, by 7.5 percent in Aleutian peregrines, by 3.3 percent in rough-legged hawks, and not at all in gyrfalcons. Tundra peregrine eggs contain an average of 889 parts of DDE per million (lipid basis); taiga peregrine eggs contain 673 parts per million; Aleutian peregrine eggs contain 167 parts per million; rough-legged hawk eggs contain 22.5 parts per million; and gyrfalcon eggs contain 3.88 parts per million. These changes in eggshell thickness and the pesticide residues reflect different degrees of exposure to contamination. There is a highly significant negative correlation between shell thickness and DDE content in peregrine eggs. Tundra and taiga peregrines have fledged progressively fewer young each year since 1966.

Alaska↗

Changes in eggshell conductance after transfer of hens from an altitude of 3,800 to 1,200 m.

Hens acclimated to an altitude of 3,800 m (PB 480 Torr) were transferred to 1,200 m (PB 657 Torr). Eggs were collected before departure and daily after the transfer so that changes in eggshell conductance could be studied. Over the next 2 mo eggshell conductance increased 30%, presumably to compensate for the 37% reduction (from 657 to 480 Torr) in gas diffusivity at the lower altitude. Measurements of shell thickness and number of pores in the shell allow one to calculate that most of the change in total pore area occurred by an increase in cross-sectional area of individual pores.

Adaptation, Physiological↗

Two signalling pathways specify localised expression of the Broad-Complex in Drosophila eggshell patterning and morphogenesis.

The Drosophila eggshell, which has a pair of chorionic appendages (dorsal appendages) located asymmetrically along both the anterior/posterior and dorsal/ventral axes, provides a good model to study signal instructed morphogenesis. We show that the Broad-Complex, a gene encoding zinc-finger transcription factors, is essential for the morphogenesis of dorsal appendages and is expressed in a bilaterally symmetrical pattern in the lateral-dorsal-anterior follicle cells during late oogenesis. This is induced and specified along the dorsoventral axis by an epidermal growth factor receptor signalling pathway, which includes a localised transforming growth factor-alpha like molecule, Gurken, in the oocyte and the Drosophila EGF receptor homologue, Torpedo, in the surrounding somatic follicle cells. Furthermore, the precisely localised expression of BR-C along the AP axis requires a separate signalling pathway, initiated by a transforming growth factor-beta homologue, Decapentaplegic, in nearby follicle cells. These two signalling pathways, one from the oocyte and the other from the follicle cells, co-ordinately specify patches of follicle cells to express the Broad-Complex in a unique position in respect to both major axes, which in turn directs the differentiation of the dorsal appendages in the correct position on the eggshell.

Animals↗

Effect of concentration of mixed-function oxidase on concentration of estrogen, rate of egg lay, eggshell thickness, and plasma calcium in laying hens.

The objective of this study was to investigate the effect of induction or depression of mixed-function oxidases (MFO) by xenobiotics on the peripheral concentration of estrogens, plasma Ca, rate of egg lay, and eggshell thickness in laying hens. In Exp. 1, 100 mg of phenobarbital (PB) administered orally each day for 3 or 7 d reduced concentrations of estrone and estradiol (E2) in serum. In Exp. 2, 25, 50, or 100 mg of PB was administered orally to laying hens for 3 or 7 d. Treatment with 100 mg of PB for 3 d or with 25, 50, or 100 mg for 7 d significantly increased liver:body weight ratios. Microsomal protein increased after 7 d of 50 or 100 mg of PB administration. Feeding PB decreased concentration of E2 and total plasma Ca in a dose- and period-dependent manner. Concentration of E2 was reduced to 10% of control, whereas hepatic cytochrome P-450 increased significantly with dose. The correlation between concentration of E2 and P-450 was negative and significant. Total Ca in the plasma was highly, positively correlated with concentration of E2. Eggshells were thinner from hens treated with 100 mg of PB for both 3 or 7 d than those from control hens. Rate of egg lay was reduced by 100 mg of PB for 7 d. In Exp. 3, .5 mL of CCl4 given orally for 1 d decreased P-450 and increased E2.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Influence of large doses of ascorbic acid on performance, plasma calcium, bone characteristics, and eggshell quality in broilers and Leghorn hens.

Four experiments were conducted using broilers (Experiments 1 and 2) or White Leghorn hens (Experiments 3 and 4) to determine the effects of large doses of dietary ascorbic acid on performance, plasma concentration of total and ionic calcium, bone characteristics, and eggshell quality. A total of 564 male broilers were fed diets containing ascorbic acid (ranging from 0 to 3,000 ppm) from 3 to 7 wk of age. Weight gain and feed conversion were measured, and blood plasma was analyzed for total and ionic calcium. Leg bones (femur, tibia, and metatarsus) were analyzed for bone mineral content, density, and breaking strength. In Experiments 3 and 4, a total of 484 Leghorn hens were fed diets containing ascorbic acid (ranging from 0 to 3,000 ppm) for 4 wk. Egg weight and specific gravity were determined, and plasma and tibiae were analyzed as in Experiments 1 and 2. Results of the broiler experiment (Experiment 2) indicated that plasma ionic calcium was significantly increased (P < .05) in ascorbic-acid-treated birds. Among leg bones examined, femur strength was improved by 16% in birds fed 2,000 ppm of ascorbic acid (Experiment 2). Other bone characteristics were not affected. In the layer experiments (Experiments 3 and 4), egg weight increased up to 5% and egg specific gravity was improved in hens fed 2,000 or 3,000 ppm of ascorbic acid, which also had increased calcium in the blood. Results suggest that large doses of ascorbic acid in the diet influence calcium metabolism, affecting bone and eggshell mineralization in chickens.

Animals↗

Effects of dietary thiouracil on thyroid activity, egg production, and eggshell quality in commercial layers.

The effects of thyroid suppression induced during the rearing period by providing various dietary thiouracil (TU) regimens on plasma thyroxine (T4) concentrations, growth, and subsequent egg production (EP) and eggshell quality were determined in Single Comb White Leghorn chickens. Thiouracil was provided in the feed at levels of 0, .1, and .2% from 0 to 6 wk of age in Experiment 1, and at levels of 0, .05, and .1% from 6 to 16 wk of age in Experiment 2. In both experiments, T4 concentrations were reduced during TU treatment. However, T4 later became elevated at 12 and 20 wk in both dosage level groups in Experiment 1. Additionally, BW and egg weights were suppressed by both TU treatments, and EP was reduced up to Week 23 in the .1% TU-treated birds and through Week 25 in the .2% TU-treated birds. No effects on EP were noted in Experiment 2, but feed consumption (FC) was reduced during Week 6 in birds fed .05% TU and during Weeks 6, 10, and 19 in birds fed .1% TU. Both liver and thyroid weights were increased in .1% TU-treated birds relative to controls at Week 16. Eggshell quality was affected only in Experiment 2, in which birds given .05% TU had a higher relative conductance, or maximum rate of water loss, at Week 38 than 0 and .1% TU dosage levels, and .1% TU-treated birds had a higher breaking strength than 0 and .05% TU-treated birds at Week 22.(ABSTRACT TRUNCATED AT 250 WORDS)

Animal Feed↗

The effect of four levels of ascorbic acid and two levels of calcium on eggshell quality of forced-molted White Leghorn hens.

The present experiment evaluated the effect of four levels of ascorbic acid (0, 100, 250, and 500 ppm) and two levels of calcium (3.0 and 3.5%) on the productivity and eggshell quality of molted Single Comb White Leghorn hens, housed four per 30.5 cm wide x 45.7 cm deep cage. The 4 x 2 factorial treatment combinations were randomly assigned to eight blocks of cages. Egg production increased (P < .06) an average of 5% for the levels of 250 and 500 ppm ascorbic acid. No differences were observed for feed consumption, egg weight, and shell thickness for the different levels of ascorbic acid or calcium in the diet. An increase (P < .01) was observed for specific gravity, as the levels of ascorbic acid increased. Increasing the level of calcium from 3.0 to 3.5% increased (P < .01) specific gravity from 1.073 to 1.075. Shell weight increased (P < .01) with the higher levels of ascorbic acid (250 and 500 ppm). No interactions were observed between the levels of ascorbic acid and calcium. Results suggest that supplementing ascorbic acid to molted laying hens can be beneficial to egg production and eggshell quality.

Animal Feed↗