Elongated shape and unusual eggshell microstructure enable first confirmed hatching of avian twins†

The aim of the study was to elucidate the causes that led to the hatching of goose twins for the first time. Analyses included the reconstruction of the egg's dimensions based on preserved fragments of shell and characterization of its microstructure. Sequencing of the genome of the twins was performed. Based on the results, the dimensions of the egg were recreated, while the structure and porosity of the shell were characterized. The main factors that allowed the twins to survive to the end of incubation were, first, the highly elongated shape of the egg, which "forced" the embryos to adopt a parallel position in line with the long axis of the egg, and second, the shell's altered porosity indices. The pores in the twins' post-hatching shell had a smaller surface area, but there were significantly more of them than in the control shells. As a result, for the twins' egg, the total pore area of the shell and the ratio of total pore area to shell thickness was low. These changes contributed to changes in the dynamics of water diffusion from the egg, adapting it to the needs of the two embryos. Analyses, including whole-genome sequencing, indicate that most of the single nucleotide polymorphism (SNP) variants, and insertions, and deletions in both twins' cases were located within introns and in the intergenic parts of the genome. A comparison of the type and frequency of SNP and InDel variations showed that the twins are characterized by high level of genetic similarity.