Do the differences in egg contamination, penetration, and resistance against microorganisms among the hen genotypes exist?

The aim of this study was to evaluate and compare the impact of genotype and storage conditions (temperature and time) on microbiological contamination and eggshell quality. There were four genotypes of laying hens used, Czech golden spotted (CGS), Greenleg Partridge (GP), White Leghorn (WL) and commercial hybrid (CH) hens were included. After collection, the eggs were divided equally into five groups regarding the storage time (0, 14, 28 days) and temperature (5 and 20 °C). The microbiological analysis included counting of colonies forming units (CFU) of Escherichia coli (EC), Enterococcus (ENT) and total number of microorganisms (TNM) on eggshell surface, eggshell membranes and in thin albumen. The analysis of eggshell quality included the determination of eggshell proportion (SP), thickness (ST), strength (SST), index (SI) and surface (SS). Moreover, egg weight (EW) and egg weight loss (EWL) were determined. The significant effect of genotype was found in contamination of eggshell by EC, ENT and TNM, eggshell membranes by TNM and albumen by EC (all P ≤ 0.05). The significantly lowest contamination of eggshell from EC was in eggs from the WL hens (4.42 log CFU/eggshell), while from ENT was in eggs from the CGS hens (1.22 log CFU/eggshell) and from the WL hens (1.40 log CFU/eggshell). The lowest incidence of TNM was also detected in eggs from the WL hens (5.03 log CFU/eggshell). Statistically the lowest contamination of eggshell membranes by TNM was found in eggs from the WL (0.12 log CFU/eggshell membranes) and CH hens (0.15 log CFU/eggshell membranes). Regarding the effect of genotype, the GP (not detected) and WL (not detected) hens was in eggs with statistically the lowest occurrence of EC bacteria in albumen. Regarding the EW and eggshell quality, all the parameters were significantly affected by the genotype (P ≤ 0.0001). Also EWL was significantly (P ≤ 0.05) affected by genotype (after 14, 21 and 28 days of storage). There were found to be significant differences of microbial contamination of egg surface among observed hen genotypes. The penetration of selected microorganisms was also significant in contamination of eggshell membranes by TNM and in contamination of albumen by EC.

Eggshell membrane for DNA sexing of the endangered Maleo (Macrocephalon maleo)

Background: Noninvasive DNA sampling has been applied across many avian genetic studies for a variety of purposes including conservation and management of endangered birds. However, its application in megapodes is still lacking. The previous genetic studies on megapodes used either blood or fresh tissue. Here we present the first demonstration of the use of eggshell membrane for research on endangered Maleo (Macrocephalon maleo). Methods: We used 24 post-hatched eggshell membranes collected from two different sites, Tambun and Tanjung Binerean, in North Sulawesi, 12 samples in each. Two different DNA extraction methods: alkaline lysis method and gSYNCTM DNA Extraction Kit were applied. To determine the sex of Maleo, we utilized PCR-based DNA sexing using CHD genes, with the primer set 2550F/2718R. Results: We successfully extracted all samples; the mean sample concentration was 267.5 ng/µl (range 47–510.5 ng/µl) and samples were of high purity (A260/280 ratio 1.85±0.03). All samples were used to successfully identified sexes, 9 females and 15 males. Conclusions: Our research clearly illustrates that eggshell membranes can be used for DNA sexing and open the possibility to build noninvasive DNA collections over large spatial scales for population study of endangered birds.

Eggshell membrane for DNA sexing of the endangered Maleo (Macrocephalon maleo)

Background: Noninvasive DNA sampling has been applied across many avian genetic studies for a variety of purposes including conservation and management of endangered birds. However, its application in megapodes is still lacking. The previous genetic studies on megapodes used either blood or fresh tissue. Here we present the first demonstration of the use of eggshell membrane for research on endangered Maleo (Macrocephalon maleo). Methods: We used 24 post-hatched eggshell membranes collected from two different sites, Tambun and Tanjung Binerean, in North Sulawesi, 12 samples in each. Two different DNA extraction methods: alkaline lysis method and gSYNCTM DNA Extraction Kit were applied. To determine the sex of Maleo, we utilized PCR-based DNA sexing using CHD genes, with the primer set 2550F/2718R. Results: We successfully extracted all samples; the mean sample concentration was 267.5 ng/µl (range 47–510.5 ng/µl) and samples were of high purity (A260/280 ratio 1.85±0.03). All samples were used to successfully identified sexes, 9 females and 15 males. Conclusions: Our research clearly illustrates that eggshell membranes can be used for DNA sexing and open the possibility to build noninvasive DNA collections over large spatial scales for population study of endangered birds.

Eggshell membrane for DNA sexing of the endangered Maleo (Macrocephalon maleo)

Background: Noninvasive DNA sampling has been applied across many avian genetic studies for a variety of purposes including conservation and management of endangered birds. However, its application in megapodes is still lacking. The previous genetic studies on megapodes used either blood or fresh tissue. Here we present the first demonstration of the use of eggshell membrane for research on endangered Maleo (Macrocephalon maleo). Methods: We used 24 post-hatched eggshell membranes collected from two different sites, Tambun and Tanjung Binerean, in North Sulawesi, 12 samples in each. Two different DNA extraction methods: alkaline lysis method and gSYNCTM DNA Extraction Kit were applied. To determine the sex of Maleo, we utilized PCR-based DNA sexing using CHD genes, with the primer set 2550F/2718R. Results: We successfully extracted all samples; the mean sample concentration was 267.5 ng/µl (range 47–510.5 ng/µl) and samples were of high purity (A260/280 ratio 1.85±0.03). All samples were used to successfully identified sexes, 9 females and 15 males. Conclusions: Our research clearly illustrates that eggshell membranes can be used for DNA sexing and open the possibility to build noninvasive DNA collections over large spatial scales for population study of endangered birds.

Low genetic diversity and no genetic differentiation between maleo hatched at coastal and inland nesting grounds in North Sulawesi, Indonesia

Saputra AW, Yuda P. 2020. Low genetic diversity and no genetic differentiation between maleo hatched at coastal and inland nesting grounds in North Sulawesi, Indonesia. Biodiversitas 21: 4772-4777. Maleo Senkawor (Macrocephalon maleo), an endemic and endangered bird of Sulawesi (Indonesia), is burrow-nesting megapodes that incubate its eggs in communal nesting sites in soils heated by sun on beaches and by volcanic activity in inland. The aims of this study were to assess genetic diversity of the Maleo and examine whether those which have different nesting sites have become genetically differentiated. In total, 24 eggshell membranes of Maleo were collected from Tanjung Binerean (coastal nesting ground) and Tambun (inland nesting ground), and the DNA was extracted using silica spin-column kit. PCR was applied to amplify the hypervariable region 1 (HV1) and partial mtDNA control region of HV2 using a specific primer set designed for Maleo. The PCR products were sequenced, resulted in 612 bp, and showed 9 polymorphic sites and 9 haplotypes (H). Further sequences analysis suggested that there was no genetic differentiation between coastal nesting population and inland nesting population (Fst = 0.0009; P = 0.431). As expected, the genetic diversity of Maleo was relatively low (coastal nesting population, Hd: 0.727270, ℼ: 0.002377 and inland nesting population Hd: 0.848480, ℼ: 0.002203).

Polycarboxylated Eggshell Membrane Scaffold as Template for Calcium Carbonate Mineralization

Biomineralization is a process in which specialized cells secrete and deliver inorganic ions into confined spaces limited by organic matrices or scaffolds. Chicken eggshell is the fastest biomineralization system on earth, and therefore, it is a good experimental model for the study of biomineralization. Eggshell mineralization starts on specialized dispersed sites of the soft fibrillar eggshell membranes referred to as negatively charged keratan sulfate mammillae. However, the rest of the fibrillar eggshell membranes never mineralizes, although 21% of their amino acids are acidic. We hypothesized that, relative to the mammillae, the negatively charged amino acids of the fibrillar eggshell membranes are not competitive enough to promote calcite nucleation and growth. To test this hypothesis, we experimentally increased the number of negatively charged carboxylate groups on the eggshell membrane fibers and compared it with in vitro calcite deposition of isolated intact eggshell membranes. We conclude that the addition of poly-carboxylated groups onto eggshell membranes increases the number of surface nucleation sites but not the crystal size.

Natural Bioactive Compound Loaded Eggshell Membrane from Waste Food Industry: Preparation and Characterization

Objective: Eggshell membrane (ESM), the protein and collagen-rich membrane between the eggshell and egg white, has usually been regarded as waste and overlooked. However, its potential is now being highlighted in many industrial and technological applications. In the literature natural bioactive compounds (NBCs) have been used together with various proteins. Methods: Preparation and characterization methods of the eggshell and eggshell membrane is summarised. Results: Various studies showed how to utilize eggshell membranes as an adsorbent, scaffold, wound dressing, and vascular graft. Due to the chemical composition of the eggshell membrane, its superior binding and adsorptive properties increase the stability of the bioactive compounds. Conclusion: This review focuses on the potential utilization of eggshell membranes as functional food and nutraceutical.