Guinea fowl eggshell quantitative proteomics yield new findings related to its unique structural characteristics and superior mechanical properties.

The Guinea fowl eggshell is a bioceramic material with the remarkable mechanical property of being twice as strong as the chicken eggshell. Both eggshells are composed of 95% calcite and 3.5% organic matrix, which control its structural organization. Chicken eggshell is made of columnar calcite crystals arranged vertically. In the Guinea fowl, the same structure is observed in its inner half, followed by a dramatic change in crystal size and orientation in the outer region. Guinea fowl eggshell is thicker than chicken eggshell. Both structure and shell thickness confer a superior resistance to breakage compared to eggshells of other bird species. To understand the underlying mechanisms controlling the structural organization of this highly resistant material, we used quantitative proteomics to analyze the protein composition of the Guinea fowl eggshell organic matrix at key stages of the biomineralization process. We identified 149 proteins, which were compared to other bird eggshell proteomes and analyzed their potential functions. Among the 149 proteins, 9 are unique to Guinea fowl, some are involved in the control of the calcite precipitation (Lysozyme, Ovocleidin-17-like, Ovocleidin-116 and Ovalbumin), 61 are only found in the zone of microstructure shift and 17 are more abundant in this zone. SIGNIFICANCE: The avian eggshell is a critical physical barrier to protect the contents of this autonomous reproductive enclosure from physical and microbial assault. The Guinea fowl (Numida meleagris) eggshell exhibits a unique microstructure (texture), which confers exceptional mechanical properties compared to eggshells of other species. In order to understand the mechanisms that regulate formation of this texture in the Guinea fowl eggshell, we performed comparative quantitative proteomics at key stages of shell mineralization and particularly during the dramatic shift in shell microstructure. We demonstrate that the Guinea fowl eggshell proteome comprises 149 proteins, of which 61 were specifically associated with the change in size and orientation of calcite crystals. Comparative proteomics analysis with eggshell of other bird species leads to new insights into the biomineralization process. Moreover, our data represents a list of organic compounds as potential additives to regulate material design for industrial fabrication of ceramics. This information also provides molecular markers for efficient genomic selection of chicken strains to lay eggs with improved shell mechanical properties for enhanced food safety.