Guinea fowl eggshell structural analysis at different scales reveals how organic matrix induces microstructural shifts that enhance its mechanical properties.

Guinea fowl eggshells have an unusual structural arrangement that is different from that of most birds, consisting of two distinct layers with different microstructures. This bilayered organization, and distinct microstructural characteristics, provides it with exceptional mechanical properties. The inner layer, constituting about one third of the eggshell thickness, contains columnar calcite crystal units arranged vertically as in most bird shells. However, the thicker outer layer has a more complex microstructural arrangement formed by a switch to smaller calcite domains with diffuse/interlocking boundaries, partly resembling the interfaces seen in mollusk shell nacre. The switching process that leads to this remarkable second-layer microstructure is unknown. Our results indicate that the microstructural switching is triggered by changes in the inter- and intracrystalline organic matrix. During production of the outer microcrystalline layer in the later stages of eggshell formation, the interactions of organic matter with mineral induce an accumulation of defects that increase crystal mosaicity, instill anisotropic lattice distortions in the calcite structure, interrupt epitaxial growth, reduce crystallite size, and induce nucleation events which increase crystal misorientation. These structural changes, together with the transition between the layers and each layer having different microstructures, enhance the overall mechanical strength of the Guinea fowl eggshell. Additionally, our findings provide new insights into how biogenic calcite growth may be regulated to impart unique functional properties. STATEMENT OF SIGNIFICANCE: Avian eggshells are mineralized to protect the embryo and to provide calcium for embryonic chick skeletal development. Their thickness, structure and mechanical properties have evolved to resist external forces throughout brooding, yet ultimately allow them to crack open during chick hatching. One particular eggshell, that of the Guinea fowl, has structural features very different from other galliform birds - it is bilayered, with an inner columnar mineral structure (like in most birds), but it also has an outer layer with a complex microstructure which contributes to its superior mechanical properties. This work provides novel and new fundamental information about the processes and mechanisms that control and change crystal growth during the switch to microcrystalline domains when the second outer layer forms.

Comparative analysis of the morphology, chemistry and structure of the tibiotarsus, humerus and keel bones in laying hens.

1. Bone properties are adapted to their specific functions in the animal, so various types of bones develop different characteristics depending on their location in the skeleton.2. The aim of this research was to compare the chemical composition, mineral characteristics and structural organisation in tibiotarsus, humerus and keel bones as representatives of hen skeletal mineralisation. Complementary analytical techniques, such as X-ray radiography, optical and electron microscopy, thermogravimetry and 2D X-ray diffraction, were used for characterisation.3. The humerus had a thinner cortex and cortical bone mineral had higher crystallinity and a greater degree of crystal orientation than the tibiotarsus. The humerus generally lacks medullary bone although, when present, it has a higher mineral content than seen in the tibiotarsus. These differences were attributed to the different forces that stimulate bone formation and remodelling.4. The keel cortical bone had a lower degree of mineralisation than the tibiotarsus or humerus. Its degree of mineralisation decreased from the cranial to the distal end of the bone. This gradient may affect keel mechanical properties, making it more prone to deformation and fractures.5. Data from studying different bones in laying hens can help to understand mineralisation as well as finding solutions to prevent osteoporosis-related fractures.

Influence of physical activity on tibial bone material properties in laying hens.

Laying hens develop a type of osteoporosis that arises from a loss of structural bone, resulting in high incidence of fractures. In this study, a comparison of bone material properties was made for lines of hens created by divergent selection to have high and low bone strength and housed in either individual cages, with restricted mobility, or in an aviary system, with opportunity for increased mobility. Improvement of bone biomechanics in the high line hens and in aviary housing was mainly due to increased bone mass, thicker cortical bone and more medullary bone. However, bone material properties such as cortical and medullary bone mineral composition and crystallinity as well as collagen maturity did not differ between lines. However, bone material properties of birds from the different type of housing were markedly different. The cortical bone in aviary birds had a lower degree of mineralization and bone mineral was less mature and less organized than in caged birds. These differences can be explained by increased bone turnover rates due to the higher physical activity of aviary birds that stimulates bone formation and bone remodeling. Multivariate statistical analyses shows that both cortical and medullary bone contribute to breaking strengthThe cortical thickness was the single most important contributor while its degree of mineralization and porosity had a smaller contribution. Bone properties had poorer correlations with mechanical properties in cage birds than in aviary birds presumably due to the greater number of structural defects of cortical bone in cage birds.

Quality assessment of chicken eggshell cuticle by infrared spectroscopy and staining techniques: a comparative study.

1. The cuticle is a very thin organic layer that coats the eggshell surface and plugs the eggshell pores preventing bacterial penetration. It also reduces eggshell permeability which is important to maintain internal quality of the egg. Thus, the eggshell cuticle quality is crucial to ensure the food safety and quality of eggs. 2. A new methodology to assess eggshell cuticle quality, based on attenuated total reflection-Fourier transform infrared spectroscopy (ATR-FTIR), is compared with a more traditional method based on cuticle staining techniques. 3. Both techniques are useful to determine the amount of cuticle; however, the ATR-FTIR technique is independent of egg colour, more sensitive and provides complete information about the cuticle and its composition. Additionally, it provides information about eggshell permeability. 4. The methodology for cuticle quality assessment described in this work can be very useful for genetic selection programmes aimed to improve the safety and quality of eggs.