Mammillary Layer Research Articles

Dietary sodium sulfate supplementation improves eggshell quality, uterine ion transportation and glycosaminoglycan synthesis in laying hens.

This study evaluated the effects of dietary sodium sulfate (Na2SO4) supplementation on eggshell quality, uterine ion transportation, and glycosaminoglycan (GAG) synthesis. A total of 432 48-wk-old Hy-line Brown laying hens were randomly divided into 6 dietary treatments with 8 replicates of 9 birds each. The experimental laying hens were fed the corn-soybean meal diets (containing 0.15% NaCl) supplemented with 0.22%, 0.37%, 0.52%, 0.68%, 0.83%, or 0.99% Na2SO4 for 12 weeks. Results showed that the eggshell breaking strength and eggshell ratio significantly increased in the 0.68% Na2SO4 group at the end of wk 56 and wk 60 (P < 0.05). In addition, eggshell thickness and weight significantly increased in the 0.68% Na2SO4 group at the end of wk 60 (P < 0.05). Eggshell calcium (Ca) content in the 0.68% Na2SO4 group was higher than that of 0.22% and 0.99% groups (P < 0.001). The concentrations of K+ and Ca2+ in the uterine fluid were significantly greater in the 0.68% group than in the other groups (P < 0.05). Dietary Na2SO4 increased the gene expression of SLC8A1, SCNN1A, ATP1B1, and KCNMA1 quadratically in the uterus (P < 0.05), and higher values were observed in 0.68% group. Additionally, the GAG contents of the eggshell, and ATP-S, SULT, CS, and DS contents of the isthmus increased linearly with the increment of dietary Na2SO4 (P < 0.05). There was a remarkable reduction in mammillary knob width, mammillary thickness, and the percentage of the mammillary layer (P < 0.05), and an increment in mammillary knob density, effective thickness, and total thickness in the 0.68% group compared with the 0.22% and 0.99% groups (P < 0.05). Overall, there was no dose-related difference with the increment of dietary Na2SO4 levels. The addition of 0.68% Na2SO4 in the corn-soybean basal diet (0.15% Cl) regulated uterine ion transport, increased GAG contents of eggshell, and improved eggshell ultrastructure and quality.

Organic Trace Elements Improve the Eggshell Quality via Eggshell Formation Regulation during the Late Phase of the Laying Cycle.

The quality of eggshells is critical to the egg production industry. The addition of trace elements has been shown to be involved in eggshell formation. Organic trace elements have been found to have higher biological availability than inorganic trace elements. However, the effects of organic trace elements additive doses on eggshell quality during the laying period of commercial laying hens required further investigation. This experiment aims to explore the potential mechanisms of different doses of organic trace elements replacing inorganic elements to remodel the eggshell quality of egg-laying hens during the laying period. A total of 360 healthy hens (Lohmann Pink, 45-week-old) were randomly divided into four treatments, with six replications per treatment and 15 birds per replication. The dietary treatments included a basal diet supplemented with inorganic iron, copper, zinc and manganese at commercial levels (CON), a basal diet supplemented with organic iron, copper, zinc and manganese at 20% commercial levels (LOT), a basal diet supplemented with organic iron, copper, zinc and manganese at 30% commercial levels (MOT), and a basal diet supplemented with organic iron, copper, zinc and manganese at 40% commercial levels (HOT). The trial lasted for 8 weeks. The results of the experiment showed that the replacement of organic trace elements did not significantly affect the production performance of laying hens (p > 0.05). Compared with inorganic trace elements, the MOT and HOT groups improved the structure of the eggshells, enhanced the hardness and thickness of the eggshells, increased the Haugh unit of the eggs, reduced the proportion of the mammillary layer in the eggshell, and increased the proportion of the palisade layer (p < 0.05). In addition, the MOT and HOT groups also increased the enzyme activity related to carbonate transport in the blood, the expression of uterine shell gland-related genes (CA2, OC116, and OCX32), and the calcium and phosphorus content in the eggshells (p < 0.05). We also found that the MOT group effectively reduced element discharge in the feces and enhanced the transportation of iron (p < 0.05). In conclusion, dietary supplementation with 30-40% organic micronutrients were able to improve eggshell quality in aged laying hens by modulating the activity of serum carbonate transport-related enzymes and the expression of eggshell deposition-related genes.

Comparative Analysis of the Ultrastructure, Bubble Pores, and Composition of Eggshells of Dwarf Layer-White and Guinea Fowl.

The decrease in eggshell quality seriously affects production efficiency. Guinea fowl (GF) eggs possess strong eggshells because of their unique crystal structure, and few systematic studies have compared laying hen and GF eggs. Sixty eggs were collected from both 40-week-old Dwarf Layer-White (DWL-White) laying hens and GF, and the eggshell quality, ultrastructure, bubble pores, and composition were measured. The results showed that the DWL-White eggs had a higher egg weight and a lower eggshell strength, strength per unit weight, thickness, and ratio than the GF eggs (p < 0.01). There were differences in the mammillary layer thickness ratio, the effective layer thickness ratio, the quantity of bubble pores (QBPs), the ratio of the sum of the area of bubble pores to the area of the eggshell in each image (ARBE), and the average area of bubble pores (AABPs) between the DWL-White and GF eggs (p < 0.01). The composition analysis demonstrated that there were differences in the organic matter, inorganic matter, calcium, and phosphorus between the DWL-White and GF eggs (p < 0.01). There were positive associations between the mammillary knob number in the image and the QBPs and ARBE and a negative correlation with the AABPs in the DWL-White eggs (p < 0.01). This study observed distinctions that offer new insights into enhancing eggshell quality.

Effects of manganese glycine on eggshell quality, eggshell ultrastructure, and elemental deposition in aged laying hens

Poor eggshell quality of eggs laid by aged laying hens is the major problem affecting the length of the rearing period in the laying hen industry. Trace elements are required and play vital roles in the eggshell quality of laying hens. Appropriate dose of organic microelements is environmentally friendly and sufficient to satisfy the needs of hens because of their greater bioavailability and lower excretion than inorganic forms. The aim of this experiment was to investigate the effects of manganese (Mn) glycine (MG) on eggshell quality, elemental deposition, and eggshell ultrastructure in aged laying hens. A total of 720 Hy-Line Brown hens 70 weeks old were assigned equally to four groups with six replicates of 30 birds each. The hens were fed basal diets (without Mn supplementation) supplemented with 120 mg/kg of Mn from manganese sulfate monohydrate (MSM), or 40, 80, or 120 mg/kg Mn from MG for 12 weeks. Dietary supplementation with 80 mg/kg Mn from MG resulted in the greatest eggshell strength after 6 weeks of treatment (P = 0.047), and in greater eggshell strength than observed in the MSM control after 12 weeks of treatment (P = 0.025). After 12 weeks of treatment, the eggs of hens in the MG groups showed lower mammillary layer thickness in the blunt end, equator, and acute end than observed in the MSM control group (P < 0.001). With the exception of the blunt ends of eggs from hens in the 120 mg/kg MG group, the eggs of hens in the MG groups, compared with the MSM control group, exhibited a lower mammillary layer ratio, and greater palisade layer ratio and effective layer ratio in the blunt end, equator, and acute end (P < 0.001). Dietary supplementation with 80 mg/kg Mn from MG, compared with the MSM control and 40 and 120 mg/kg MG, resulted in the greatest palisade layer thickness and effective layer thickness, and the lowest mammillary layer thickness in the equator (P < 0.001, P = 0.001, P < 0.001, respectively). Furthermore, supplementation with 80 mg/kg Mn from MG exhibited the greatest ratio of the palisade layer and effective layer, and the lowest mammillary layer ratio in the blunt end and equator (all P < 0.001). The Mn content of eggshells in hens-fed diets supplemented with 80 and 120 mg/kg Mn from MG was greater than that in the MSM control and 40 mg/kg MG groups (P = 0.035). Dietary supplementation with 80 or 120 mg/kg Mn from MG resulted in greater tibia Mn content than observed in the 40 mg/kg MG group (P = 0.019), and greater yolk Mn content than observed in the 40 mg/kg MG and MSM control groups (P = 0.018). In conclusion, dietary supplementation with 80 mg/kg Mn from MG, compared with the MSM control (120 mg/kg Mn), may increase the deposition efficiency of Mn, alter eggshell elemental composition, improve eggshell ultrastructure, and enhance eggshell strength in aged laying hens.

Eggshell microstructure, shell quality indices, mineralogy, and UV–Vis absorbance of domestic eggs of Iran

The structure and quality of eggshells (ES) vary around a mean value depending on different items, and these variations are important in their many industrial and technological applications as will be mentioned (vide infra). Five commercial egg brands of different laying hens are examined and their quality indices, morphological microstructure, elemental composition, and light absorbance at 200–700 nm UV/Vis spectra were compared. The ES layers include a limiting membrane, inner and outer shell membranes, a mammillary layer, a palisade layer, a surface vertical crystal layer, and a bilayer cuticle. The elemental composition of each layer reflects the proteinous or calcified nature of the layers, and various elements, C, O, N, Ca, Fe, Mg, Mn, Pb, Sr, Sc, Hf, Co, and La, were found. The weight difference between egg brands was significant, as well as the difference in ES percentage between white and colored eggs. ES weight possessed a positive relationship with the thickness of the calcified layer. So, the ES is heavier and the egg is larger and heavier when the calcified layer is thicker. The calculated average ES index was 7.99% ± 0.16 SE. In all ES samples, the absorbance in UV wavelength spectra (300–350 nm) was slightly higher than in Vis spectra (400 - 700 nm) and their difference was significant. The difference in absorbance of various treatments was significant and mean absorbance was the highest in the after-furnace (ash) samples and was the lowest for acid-treated ones. It seems that turning the ES into ash can improve the absorbance capability, especially in white ESs.

Comparison of the eggshell mammillary microstructure of Galliformes and Anseriformes

AbstractIn free‐living species, the comparison of eggshell microstructure focus mainly on species representing extremely different reproductive strategies or with phylogenetically divergent taxa. The purpose of this study was to compare the microstructure of the mammillary layer in two bird orders (Galliformes and Anseriformes) representing phylogenetically close precocials, and to relate eggshell microstructure to female body weight and egg characteristics. The results showed significant differences between the studied orders. The mammillary layer in Galliformes had more knobs, and consequently, the percent coverage with them was higher than in Anseriformes. The size of individual knobs did not differ significantly between the orders. The obtained results are consistent with the reports that the ossification at the time of hatching is more advanced in Galliformes, so representatives of this order must resorb more calcium from the eggshell. Egg parameters and female body weight were correlated significantly negatively only with the number of knobs per mm2 for Anseriformes and, to a lesser extent, for Galliformes, but not with the coverage per cent of knobs or the size of individual knobs. Only for the three largest species, which lay proportionally larger eggs than the other species studied, the number of knobs per mm2 was significantly lower, but no difference was found in the per cent coverage of knobs. Our results showed that order was a stronger predictor of microstructure than species. Different ossification patterns of the embryonic skeletal system described in these two orders could be the explanation for the observed differences in eggshell microstructure; however, more research is needed.

Effects of dietary Lactobacillus rhamnosus GG supplementation on the production performance, egg quality, eggshell ultrastructure, and lipid metabolism of late-phase laying hens

BackgroundToward the late phase of laying, the production performance of laying hens decreases, egg quality deteriorates, lipid metabolism weakens, and hepatic lipid accumulation is exacerbated. Probiotics as an alternative to antimicrobials have been employed in poultry-related industries. Lactobacillus rhamnosus GG (LGG) is currently the most researched and clinically validated probiotic, showing promising effects in multiple application areas. However, few studies have been conducted on livestock (including poultry) production.ResultsCompared with the CON group, the feed conversion ratio (P < 0.01) declined significantly in the LGG group. Eggshell strength (P < 0.001) and eggshell thickness (P < 0.001) were significantly increased by supplementation with LGG in the diet. The height (P < 0.001) and proportion (P < 0.05) of the effective layer and the mammillary knob density (P < 0.01) in the eggshell ultrastructure of the LGG group increased significantly, while the mammillary layer (P < 0.05) and knob width (P < 0.01) decreased significantly. The LGG-treated hens had significantly lower serum concentrations of low-density lipoprotein (P < 0.05), free fatty acids (P < 0.01), and liver triglyceride (P < 0.05) levels than those in the CON group.ConclusionsLGG supplementation significantly decreases the feed conversion ratio, improves eggshell quality by altering the ultrastructure, and improves lipid metabolism in the late laying period.

Grape Pomace: Agrifood By-Product with Potential to Enhance Performance, Yolk Quality, Antioxidant Capacity, and Eggshell Ultrastructure in Laying Hens.

Grape pomace (GP) is an industrial by-product of grape juice making and is commonly discarded as a waste product, even with its large quantity of phytochemicals. Thus, the objectives of this trial were to examine the effects of graded dietary GP on laying rate, egg quality, yolk lipid profile, oxidative stability, shell quality and ultrastructure, and serum biochemistry. Two hundred 35-week-old laying hens were allocated to four dietary treatments with ten replicates each. Four diets were formulated by mixing a standard basal diet with GP at 0 g/kg (control), 30 g/kg (GP3%), 60 g/kg (GP6%), and 90 g/kg (GP9%). Egg production percent, egg weight, and egg mass were linearly improved (p < 0.01) due to adding GP to the diets of laying hens. Eggs obtained from laying hens fed with GP diets had (p < 0.01; linear, p < 0.01) greater Haugh units, yolk color, albumen index, and yolk index than those of the control. The GP9% group had the greatest values (p < 0.05) for shell weight, thickness, and breaking strength. Electron microscopy scanning of eggshells indicated that the incremental dietary level of GP linearly augmented the thickness of the palisade layer but reduced both the mammillary layer and mammillary knob width (p < 0.01). Improved tibia-breaking strength and ash content were shown (p < 0.05) in the GP-fed laying hens. The dietary addition of GP by up to 90 g/kg linearly (p < 0.01) mitigated lipid oxidation and improved the antioxidant capacity in both the serum and stored eggs. A reduction in the percentages of saturated fatty acids was observed, while the contents of monounsaturated fatty acids, polyunsaturated fatty acids, and n-3 fatty acids were augmented because of increasing dietary GP levels (p < 0.001). Additionally, the eggs obtained from laying hens fed on the GP6% and GP9% had lower yolk cholesterol content (p < 0.001); this effect was confirmed by linear and quadratic responses (p < 0.001). Laying hens on GP diets had lower (p < 0.01) serum hepatic enzymes, cholesterol, triglycerides, and low-density lipoprotein but greater high-density lipoprotein compared to the control. To sum up, the addition of GP in the layers' diets by up to 90 g/kg increased laying performance, enriched the yolk with beneficial fatty acids, enhanced antioxidant potential in yolk lipids, and improved shell quality and ultrastructure.

Effect of pearl guinea fowl eggshell ultrastructure and microstructure on keets hatchability

Variability in shell structure is an evolutionary mechanism in birds that enables them to adapt to specific environmental conditions. This variability may also occur within the same species under the influence of individual indicators, such as the age or health status of females. While interspecies variation is quite obvious and easy to interpret, the reasons for intraspecies variation remain unclear. In this study, we examined the ultra- and microstructure of guinea fowl eggshells to identify the association between variations in shell structure and hatchability outcomes. We analyzed the visual differences between shells with low (L), intermediate (I), and high (H) external porosity using scale invariant feature transform analysis with NaturePatternMatch software. We found that the external pore image was closely related to the overall porosity of the shell before incubation. The total pore area, total porosity, and diffusion index (GH2O) were highest in group H shells (P < 0.001). Posthatching shells were characterized by an increased diameter and total surface area, decreased pore number (P < 0.001), as well as shortened mammillary layer (P < 0.001) and decreased total consumption of mammillary knobs (P < 0.001). The porosity indices of posthatching H shells had intermediate values between L and I. Although the effect of shell structure parameters on hatching was not confirmed, we assumed that all categories (L, I, and H) of shells were ideal for incubation. This suggests that the shell structure adapts to the metabolic rate of developing embryos; however, differences in shell structure affect the duration of incubation and synchronization of hatching. Both L and H shells showed delayed and prolonged hatching. Therefore, we recommended that guinea fowl eggs with different external porosity parameters should be incubated separately for better hatching synchronization. Differences in GH2O between L, I, and H eggs suggest that the shell porosity characteristics of guinea fowl eggs may be a key determinant of the rate of water loss during storage before incubation.

The structure of the eggshell and eggshell membranes of Crocodylus niloticus.

The macro- and microstructure, elemental composition, and crystallographic characteristics of the eggshell and eggshell membranes of the Crocodylus niloticus egg was investigated using optical and electron microscopy, energy-dispersive X-ray spectroscopy (EDS), electron backscatter diffraction (EBSD) and computerised tomography. The translucent ellipsoid egg is composed of two basic layers, the outer calcified layer referred to as the shell and an inner organic fibre layer, referred to as the shell membrane. The outer inorganic calcite shell is further divided into an external, palisade and mammillary layers with pore channels traversing the shell. The external layer is a thin layer of amorphous calcium and phosphorus, the underlying palisade layer consist of irregular wedge-shaped crystals composed calcite with traces of magnesium, sodium, sulphur and phosphorus. The crystals are mostly elongated, orientated perpendicular to the shell surface ending in cone-shaped knobs, which forms the inner mammillary layer. The elemental composition of the mammillae is like that of the palisade layer, but the crystal structure is much smaller and orientated randomly. The highest number of mammillae and shell pores are found at the equator of the egg, becoming fewer towards the egg poles. The shell thickness follows the same pattern, with the thickest area located at the equator. The eggshell membrane located right beneath and embedded in the mammillary layer of the shell; it is made up of unorganised fibre sheets roughly orientated at right angles to one another. Individual fibres consist of numerous smaller fibrils forming open channels that run longitudinally through the fibre.

산란후기 사료 내 가수분해 효모의 첨가 급여가 생산성과 계란 품질에 미치는 영향

An experiment was conducted to investigate the effects of varying levels of hydrolyzed yeast on egg production and egg quality in aged laying randomly allotted to three dietary treatments such that egg production was similar in each treatment (6 replicates of 10 birds each). The layers were fed diets containing 0, 0.1, or 0.2% hydrolyzed yeast for eight weeks. No significant difference was observed in egg production during the first half of the experiment. Egg production and daily egg mass in groups fed diets containing hydrolyzed yeast were significantly higher (<i>P</i>&lt;0.05) than those of the control groups during the second half of the experiment. Egg weight was not affected by the dietary treatment. Eggshell strength and thickness in groups fed diets containing hydrolyzed yeast were significantly higher than those of the control groups during the overall experimental period (<i>P</i>&lt;0.05). Although no significant differences were observed in the Haugh units, yolk color in the group fed diets containing 0.1% hydrolyzed yeast was significantly higher than that in the control group (<i>P</i>&lt;0.05). The mammillary layer thickness increased in a linear manner and significantly following treatment with dietary hydrolyzed yeast (<i>P</i>&lt;0.05). Antibody titer against avian influenza virus in the group fed diets containing 0.2% hydrolyzed yeast was significantly higher (<i>P</i>&lt;0.05) than that in the control group. In conclusion, dietary hydrolyzed yeast improved egg production and eggshell quality of laying hens in the late stages of production.

Genetic variations for the eggshell crystal structure revealed by genome-wide association study in chickens

BackgroundEggshell is a bio-ceramic material comprising columnar calcite (CaCO3) crystals and organic proteinaceous matrix. The size, shape and orientation of the CaCO3 crystals influence the microstructural properties of chicken eggshells. However, the genetic architecture underlying eggshell crystal polymorphism remains to be elucidated.ResultsThe integral intensity of the nine major diffraction peaks, total integral intensity and degree of orientation of the crystals were measured followed by a genome-wide association study in 839 F2 hens. The results showed that the total integral intensity was positively correlated with the eggshell strength, eggshell thickness, eggshell weight, mammillary layer thickness and effective layer thickness. The SNP-based heritabilities of total integral intensity and degree of orientation were 0.23 and 0.06, respectively. The 621 SNPs located in the range from 55.6 to 69.1 Mb in GGA1 were significantly associated with TA. PLCZ1, ABCC9, ITPR2, KCNJ8, CACNA1C and IAPP, which are involved in the biological process of regulating cytosolic calcium ion concentration, can be suggested as key genes regulating the total integral intensity.ConclusionsThe findings greatly advance the understanding of the genetic basis underlying the crystal ultrastructure of eggshell quality and thus will have practical significance in breeding programs for improving eggshell quality.

Hydroxychloride trace elements improved eggshell quality partly by modulating uterus histological structure and inflammatory cytokines expression in aged laying hens

The objectives of this study were to investigate the effectiveness of dietary zinc, copper, and manganese hydroxychloride (HC) supplementation on performance, minerals deposition, serum parameters, eggshell ultrastructure, uterus histological structure, and inflammatory cytokines in aged hens. A total of 560 Hyline Brown layers at 62 wk of age were randomly allotted into 3 groups (CON, basal diet without extra minerals supplemented; Sulphate and HC, basal diet with sulphate or hydroxychloride zinc, copper, and manganese supplementation at levels of 80, 15, and 80 mg/kg, respectively). The trial lasted for 16 wk consisting of 4 wk depletion period and 12 wk testing period. The results indicated that dietary hydroxychloride trace elements increased egg weight (P < 0.05) when compared with CON group and improved average Haugh unit and albumen height (P < 0.05) when compared with Sulphate group from 70 to 73 wk. Trace element supplementation significantly increased eggshell strength, ceruloplasmin content in serum, and modified crystallographic structure of eggshell (P < 0.05) that included effective layer height, palisade height, mammillary layer width, and mammillary internal area ratio, but the results did not differ regarding the trace mineral sources used. Furthermore, hens fed with hydroxychloride trace element showed the highest mucosal fold height (P < 0.05) and epithelial height (P = 0.053) in eggshell gland, as well as mRNA expression of TNF-α (P < 0.05) and IL-22 (P = 0.094). It is concluded that supplementation of Zn, Cu, and Mn mixture modified eggshell quality partly through enhancing histological structure and immune responses of uterus. Hydroxychloride source of Zn, Cu, and Mn excelled sulphate in its beneficial effects for birds.

Tiny, ornamented eggs and eggshell from the Upper Cretaceous of Utah represent a new ootaxon with theropod affinities

A new Cretaceous ootaxon (eggshell type) from the Kaiparowits Formation of Grand Staircase-Escalante National Monument is among a growing number of very small eggs described from the Mesozoic. Analyses of two partial eggs (~ 17.7 mm in diameter) and 29 eggshell fragments reveal that this new ootaxon exhibits nodose ornamentation with distinctive branching pore canals that open atop the nodes. Its two-layered microstructure consists of a mammillary layer and a continuous layer with rugged grain boundaries between calcite grains. Although the exact identity of the egg producer is unknown, the eggshell microstructure and small size is consistent with a small-bodied avian or non-avian theropod. The specific combination of small egg size, branching pores, two-layered microstructure, and dispersituberculate ornamentation preserved in this new ootaxon is unique among theropod eggs. This underscores that both eggshell and skeletal fossils of Cretaceous theropods can display a mosaic of transitional morphological and behavioural features characteristic of both avian and non-avian taxa. As such, this new ootaxon increases the diversity of Cretaceous eggs and informs our understanding of the evolution of theropod eggshell microstructure and morphology.

The partial replacement of sodium chloride with sodium bicarbonate or sodium sulfate in laying hen diets improved laying performance, and eggshell quality and ultrastructure

This study investigated the effects of dietary chloride (Cl) reduction on laying performance and eggshell quality by substitution of sodium bicarbonate (NaHCO3) or sodium sulfate (Na2SO4) for part of dietary sodium chloride (NaCl), and further explored its mechanism for improving eggshell quality. A total of 360 29-wk-old Hy-line Brown laying hens were randomly allocated to 5 dietary treatments, including a basal diet contained 0.33% NaCl (control group, 0.27% dietary Cl), and 4 experimental diets that contained 0.21% and 0.15% dietary Cl by substituting Na2SO4 or NaHCO3 for part of NaCl in the basal diet. No significant differences were observed in blood Na+, Cl−, K+ and Ca2+ levels and pH value as well as serum creatinine and uric acid contents among 5 treatments (P > 0.05). Dietary Cl reduction increased egg production and ADFI during wk 33 to 36, 37 to 40 and 29 to 40 of age and decreased feed conversion ratio during wk 37 to 40 of age (P < 0.05). The hens fed with diets containing 0.15% Cl increased eggshell breaking strength, thickness and weight ratio in wk 40 of age (P < 0.05). Birds fed with dietary 0.21% and 0.15% Cl exhibited higher effective layer thickness and lower mammillary layer thickness of eggshell than those fed with dietary 0.27% Cl (P < 0.05). Apparent Ca metabolizability of hens was increased with dietary Cl reduction (P < 0.05). Total Ca of eggshell of dietary 0.15% Cl group was higher than that of dietary 0.27% Cl group (P < 0.05). No significant differences in laying performance, eggshell quality and Ca metabolism of layers were observed between Na2SO4 and NaHCO3 replacement groups (P > 0.05). Overall, dietary Cl reductions could improve laying performance and eggshell quality by substitution of NaHCO3 or Na2SO4 for part of NaCl, and there were no differences in the improvements between these two substitutes. The improved eggshell quality may be attributed to improved eggshell ultrastructure and increased supply of eggshell Ca2CO3.

Dietary supplementation with sodium bicarbonate or sodium sulfate affects eggshell quality by altering ultrastructure and components in laying hens

Dietary sodium (Na) levels were related to the content of the eggshell matrix. We therefore speculated that dietary Na supplementation as sodium bicarbonate (NaHCO3) or sodium sulfate (Na2SO4) may improve eggshell quality. Additionally, dietary NaHCO3 or Na2SO4 supplementation may further affect eggshell quality in different ways due to differences in anions. This study investigated and compared the effects of dietary Na supplementation in either NaHCO3 or Na2SO4 form on laying performance, eggshell quality, ultrastructure and components in laying hens. A total of 576 29-week-old Hy-Line Brown laying hens were randomly allocated to 8 dietary treatments that were fed a Na-deficient basal diet (0.07% Na, 0.15% Cl) supplemented with Na2SO4 or NaHCO3 at 0.08, 0.18, 0.23 or 0.33% Na for 12 weeks. No differences were observed in laying production performance with dietary Na supplementation. Dietary Na supplementation resulted in quadratic increases of eggshell breaking strength in both Na2SO4 and NaHCO3 added groups (P < 0.05), and Na2SO4-fed groups had a quadratic increase in the eggshell ratio at week 12 (P < 0.05). Compared with supplementing 0.08% Na, dietary supplementation of 0.23% Na increased the effective thickness (P < 0.05) in both Na2SO4 and NaHCO3 added groups, but decreased the thickness and knob width of the mammillary layer (P < 0.05). A linear increase on the calcium content of the shell was only observed with Na supplementation from NaHCO3 (P < 0.05). No differences were observed in Na contents of the shell with dietary Na supplemented by both sources. Dietary Na addition had a quadratic increase on uronic acid contents of shell membrane in NaHCO3-fed groups (P < 0.05). Moreover, the sulfated glycosaminoglycan (GAG) contents of shell membranes increased linearly with dietary Na supplementation (P < 0.05). Dietary supplementation of 0.23% Na from Na2SO4 increased the sulfated GAG contents of calcified eggshell (P < 0.05). Additionally, compared with NaHCO3-fed groups, Na2SO4-fed groups had higher eggshell breaking strength, thickness, eggshell weight ratio, effective thickness and the sulfated GAG contents of calcified eggshell at week 12. Overall, dietary supplementation of NaHCO3 or Na2SO4 could increase eggshell breaking strength, which may be related to increased sulfated GAG contents in eggshell membranes and improved ultrastructure. Higher eggshell breaking strength, thickness and eggshell ratio could be obtained when the diet was supplemented with 0.23% Na from Na2SO4.

Comparative morphotaxonomical affinities, palaeoecological implications and cladistic analyses of ornithoid eggshells (oofamily Laevisoolithidae) from the Late Cretaceous intertrappean beds of Anjar, District Kachchh, Gujarat, India

ABSTRACT We re-evaluate fragmentary eggshells of avian affinities (ratite morphotype) from the Late Cretaceous Intertrappean beds of Anjar (District Kachchh, Gujarat), India in view of additional material and cladistic analysis. Previously, Anjar eggshells were encompassed in the oofamily Subtiliolithidae; however, now the oofamily Subtiliolithidae has been re-examined as a junior synonym of the oofamily Laevisoolithidae. External surface of the eggshells is smooth to rugose, with unevenly distributed, sub-rounded tubercles; average eggshell thickness (467–491 µm) characterised by two layers: (1) a well-differentiated mammillary layer (196.1 µm), comprising conical, crystalline aggregates with petaloid wedges, and (2) a continuous spongy layer (274.58 µm, CL:ML, 1.3:1) divulges a squamatic zone. The eggshells have been compared with three oofamilies (Elongatoolithidae, Prismatoolithidae and Laevisoolithidae) and their palaeoecological implications have been discussed in detail. Shell microstructure is comparable with oofamily Laevisoolithidae, an ootaxon usually related with enantiornithid birds. Cladistic analysis performed on the eggshells are consistent with the avian kinships recognised based on macro, micro and ultrastructural features.

Uterine transcriptome analysis reveals mRNA expression changes associated with the ultrastructure differences of eggshell in young and aged laying hens

BackgroundLower eggshell quality in the late laying period leads to economic loss. It is a major threat to the quality and safety of egg products. Age-related variations in ultrastructure were thought to induce this deterioration. Eggshell formation is a highly complex process under precise regulation of genes and biological pathways in uterus of laying hens. Herein, we evaluated the physical, mechanical and ultrastructure properties of eggshell and conducted RNA sequencing to learn the transcriptomic differences in uterus between laying hens in the peak (young hens) and late phase (aged hens) of production.ResultsThe declined breaking strength and fracture toughness of eggshell were observed in aged hen group compared to those in young hen group, accompanied with ultrastructure variations including the increased thickness of mammillary layer and the decreased incidence of early fusion. During the initial stage of eggshell formation, a total of 183 differentially expressed genes (DEGs; 125 upregulated and 58 downregulated) were identified in uterus of laying hens in the late phase in relative to those at peak production. The DEGs annotated to Gene Ontology terms related to antigen processing and presentation were downregulated in aged hens compared to young hens. The contents of proinflammatory cytokine IL-1β in uterus were higher in aged hens relative to those in young hens. Besides, the genes of some matrix proteins potentially involved in eggshell mineralization, such as ovalbumin, versican and glypican 3, were also differentially expressed between two groups.ConclusionsAltered gene expression of matrix proteins along with the compromised immune function in uterus of laying hens in the late phase of production may conduce to age-related impairments of eggshell ultrastructure and mechanical properties. The current study enhances our understanding of the age-related deteriorations in eggshell ultrastructure and provides potential targets for improvement of eggshell quality in the late laying period.

Effects of ecofriendly synthesized calcium nanoparticles with biocompatible Sargassum latifolium algae extract supplementation on egg quality and scanning electron microscopy images of the eggshell of aged laying hens

In the present study, 200 Brown commercial egg-type layers (60 wk old) were used to study the effects of different levels of ecofriendly synthesis of calcium (Ca) nanoparticles (0.0, 0.50, 1.0, and 1.5 g/kg diet) with biocompatible Sargassum latifolium algae extract (SL-CaNps) on exterior egg quality traits, electronic microscopic view of eggshells, Ca and phosphorus (P) retention, serum Ca and P concentrations, and the histology of the uterus. Hens fed with dietary SL-CaNps powder had higher egg weight and shell weight % values than those of the control group. All SL-CaNps treatment groups had the greatest values of shell weight per unit surface area and shell thickness. Dietary supplementation of SL-CaNps at graded levels up to 1.5 g/kg diet had higher serum Ca and inorganic P levels than that of the control. Laying hens fed with SL-CaNps-added diets had beneficial effects on shell ultrastructure in terms of well-developed palisade and mammillary layers. The numbers of apical cells along the branched tubular gland were greater in SL-CaNps-treated groups than those of control. Conclusively, supplementing SL-CaNps powder up to 1.5 g/kg to the diet of laying hens improved eggshell thickness, shell weight% and shell weight per unit surface and has no adverse effect on their eggshell quality or electronic microscopic view of their eggshell.

Dietary cadmium chloride impairs shell biomineralization by disrupting the metabolism of the eggshell gland in laying hens.

In this study, we identified cadmium (Cd) as a potential endocrine disruptor that impairs laying performance, egg quality, and eggshell deposition and induces oxidative stress and inflammation in the eggshell glands of laying hens. A total of 480 38-wk-old laying hens were randomly assigned into 5 groups that were fed a basal diet (control) or a basal diet supplemented with Cd (provided as CdCl2·2.5 H2O) at 7.5, 15, 30, and 60 mg Cd per kg feed for 9 wk. The results showed that, when compared with the control group, a low dose of dietary Cd (7.5 mg/kg) had positive effects on egg quality by improving albumen height, Haugh unit, yolk color, and shell thickness at the third or ninth week. However, with the increase in the dose and duration of Cd exposure, the laying performance, egg quality, and activities of eggshell gland antioxidant enzymes (catalase [CAT], glutathione peroxide [GSH-Px]), and ATPase (Na+/K+-ATPase, Ca2+-ATPase, and Mg2+-ATPase) deteriorated, and the activity of total nitric oxide synthase (T-NOS) and the level of malondialdehyde (MDA) increased significantly (P < 0.05). The histopathology and real-time quantitative PCR results showed that Cd induced endometrial epithelial cell proliferation accompanied by upregulation of the mRNA levels of progesterone receptor (PgR) and epidermal growth factor receptor (EGFR), downregulation of the mRNA levels of estrogen receptor α (ERα) and interleukin 6 (IL6), and inflammation of the eggshell gland accompanied by significantly increased expression of complement C3 and pro-inflammatory cytokine tumor necrosis factor α (TNFα) (P < 0.05). In addition, the ultrastructure of the eggshell showed that dietary supplementation with 7.5 mg/kg Cd increased the palisade layer and total thickness of the shell, but with the increase in dietary Cd supplementation (30 and 60 mg/kg) the thickness of the palisade layer and mammillary layer decreased significantly (P < 0.05), and the outer surface of the eggshell became rougher. Correspondingly, the expression of calbindin 1 (CALB1), ovocalyxin-32 (OCX-32), ovocalyxin-36 (OCX-36), osteopontin (SPP1), and ovocledidin-17 (OC-17) decreased significantly (P < 0.05) with increasing dietary Cd supplementation. Conclusively, the present study demonstrates that dietary supplementation with Cd negatively affects laying performance, egg quality, and eggshell deposition by disturbing the metabolism of eggshell glands in laying hens but has a positive effect on egg quality at low doses.