Structural and proteomic analyses of vitelline membrane proteins of blackbird (Turdus merula) and song thrush (Turdus philomelos)

Abstract

In this study, we aimed to perform structural and proteomic analysis of the vitelline membrane (VM) of two species birds belonging to the family Turdidae: blackbird (Turdus merula) and song thrush (Turdus philomelos). We performed structural analyses using scanning electron microscopy. The VM proteins were identified and compared to the best-known chicken VM proteins. According to our results, VM of both species has a typical three-layered structure: the outer layer, inner layer, and the continuous membrane between them. An unusual observation was the finding of “convexity” formed by the inner layer in blackbird. The role of these convex structures is not known, but they can be typical for the species and can be used in their identification. In addition, we identified two proteins in the VM of both species of birds, of which U3KEZ1 FICAL was not previously identified in any other bird species, and the U3JXV8 FICAL protein was confirmed only once in cockatiel parrot VM. The function of these proteins is not exactly known, but their structure shows similarities to the SERPIN proteins that are involved in microbiological defense, i.e., they are immune proteins. This study contributes to the current knowledge about the structure and composition of proteins of VM, especially because similar analyses have never been performed for Turdidae family. Knowledge of the structure and specific proteins of blackbird and song thrush VM can be beneficial in research on ecology and bird biology and also helpful in developing noninvasive and nongenetic identification methods.

Analysis of ultrastructure and microstructure of blackbird (Turdus merula) and song thrush (Turdus philomelos) eggshell by scanning electron microscopy and X-ray computed microtomography

“…However, according to the SEM analysis of chicken eggshells in earlier studies 4,20 and analyses of eggshells of other species whose eggs weigh greater than chicken eggs [21][22][23][24] , Mk are hidden within Ma without forming such structures or are markedly smaller. In contrast, the eggshell micrographs of zebra finch 22 or American restart 25 , whose eggs weigh significantly lesser than blackbird and song thrush eggs, have shown similar structures 26 . Apart from these two recent studies on www.nature.com/scientificreports/ the eggshell morphology of small-sized eggs, most studies have only examined the shells of eggs with a high or very high weight, and the systematic impact [21][22][23][24] .…”

“…3 ). The eggs of two species classified among Phasianidae, which are distinct from Turdidae, and Chinese quail have similar weight (5.6–7.1 g 27 ) to blackbird and song thrush eggs (7.01 ± 0.17 and 6.06 ± 0.16, respectively 26 ), while domestic chicken eggs are significantly heavier (about 50–60 g). This suggests that Mk of Chinese quail eggshell are more elongated and similar to those in blackbird and song thrush eggs, but not in the larger chicken eggs, although both species belong to the same family.…”

“…The eggshell surface area A s (cm 2 ) was calculated using the formula of Paganelli et al 45 : A s = 4.835·M 0.666 , where M denotes egg weight. Egg weight measured by Damaziak et al 26 was used for the calculations.…”

“…After collection eggs from nests and transport to the Institute of Animal Science laboratory (Warsaw, PL), which lasted for about 40 min, the eggs were placed in a cold store at 8 °C for 24 h. Then all eggs were broken, and the eggshells were washed in deionized water, dried at room temperature (about 23 °C) for another 24 h and analyzed successively. The specific procedures used for collecting eggs from nests, their transport, and storage were previously described in detail by Damaziak et al 26 www.nature.com/scientificreports/ SEM analysis. Eggs were cut in the middle portion of the long axis, and approximately 3 × 3 mm samples were taken toward the blunt end.…”

Sperm morphology and performance in relation to postmating prezygotic isolation in two recently diverged passerine species

“…The divergence in the acrosome size among species might thus indicate differences in the interaction between the sperm and the egg’s glycoprotein surroundings prior to the fertilization. It is for example possible that the nightingale species differ in thickness of the glycoprotein layer around the eggs and differences in acrosome sizes between the species reflect the ease with which sperm can penetrate it 78 . Further research is needed to test these possibilities.…”