Yacon (Smallanthus sonchifolius) root extracts affect laying performance, egg quality, serum biochemical parameters and intestinal microbiota in hens.

Objective: The objective of this study was to investigate the influence of yacon root extracts (YREs) on productive performance and health of laying hens. Methods: Six hundred 30-week-old Xiaoshan Chicken layers were divided into 5 groups, control group, antibiotic positive control group, and 3 YREs treatment groups. In a 9-wk feeding experiment, at the end of wk 3, 6 and 9, twenty eggs were collected from each replicate to measure egg qualities. At the end of wk 9, three hen serum samples, and 5 hen cecal content samples were collected from each replicate. Results: Compared to the control group, 0.8%, 1.6% and 2.4% YREs treatments could increase hens' daily feed intake, and YREs supplementation affected daily feed intake in linear manner. YREs did not change egg size, but 0.8% and 2.4% YREs changed egg shape by decreasing the egg shape index and sphericity, and 0.8% YREs tended to improve the eggshell breaking strength. 1.6% YREs might decrease yolk color grade but optimize the pH of thick egg white in fresh egg; moreover, 1.6% and 2.4% YREs might be helpful for eggs to inhibit water loss during storage, and YREs supplementation affected water loss rate in linear manner. 2.4% YREs could decrease the serum lactate dehydrogenases (LDH) level, and YREs supplemental levels linearly affected serum LDH content. Finally, YREs could enrich the diversity of intestinal microbiota of hens fed with 0.8% and be beneficial for the relative abundance of phylum Bacteroidota and Halobacterota; 2.4% YREs might increase the abundance of phylum Actinobacteriota and genus Bifidobacterium, while decrease genus Bacteroides; YREs supplemental levels affected the abundance of phylum Actinobacteriota, and genera Bifidobacterium and Bacteroides in linear manner. Conclusion: Dietary supplementation with YREs could affect egg quality, protect the health of organs and exhibit prebiotic activity.

Evaluation of fertilization capability of frozen-thawed completely immotile spermatozoa collected from a white bengal tiger after interspecific ICSI with bovine oocytes.

The objective of this study was to evaluate the fertilization capability of White Bengal Tiger frozen-thawed completely immotile spermatozoa after interspecific intracytoplasmic sperm injection (ICSI) with bovine oocytes. The fertilization status of presumptive zygotes was assessed 18 h after ICSI by immunofluorescence staining and confocal microscopy. The fertilization rate was 34.8% (8/23), as confirmed by the extrusion of two polar bodies, or male and female pronuclei formation. For unfertilized oocytes (65.2%, 15/23), one activated oocyte had an activated spermatozoon but most were unactivated oocytes with unactivated spermatozoa (1/15, 6.7% vs 10/15, 66.7%, respectively, p < 0.05). These results showed that White Bengal Tiger frozen-thawed completely immotile spermatozoa retained the capacity to fertilize bovine oocytes after interspecific ICSI. This is the first report of in vitro produced zygotes using tiger immotile sperm with bovine oocytes by interspecific ICSI technique, which provides an efficient and feasible method for preservation and utilization of endangered feline animals.

Comparative proteomics of matrix fractions between pimpled and normal chicken eggshells.

Eggshell matrix can be dissociated into three matrix fractions: acid-insoluble matrix (M1), water-insoluble matrix (M2) and acid-water facultative-soluble matrix (M3). Matrix fractions from pimpled and normal eggshells were compared using label-free proteomic method to understand the differences among three matrix fractions and the proteins involved with eggshell quality. A total of 738 and 600 proteins were identified in the pimpled and normal calcified eggshells, respectively. Both eggshells showed a combined proteomic inventory of 769 proteins. In the same type of eggshell, a high similarity was present in the proteomes of three matrix fractions. These triply overlapped common proteins formed the predominant contributor to proteomic abundance in the matrix fractions. In each matrix fraction and between both eggshell models, normal and pimpled eggshells, a majority of the proteomes of the fractions were commonly observed. Forty-two common major proteins (iBAQ-derived abundance ≥0.095% of proteomic abundance) were identified throughout the three matrix fractions and these proteins might act as backbone constituents in chicken eggshell matrix. Finally, using 1.75-fold as up-regulated and using 0.57-fold as down-regulated cutoff values, twenty-five differential major proteins were screened and they all negatively influence and none showed any effect on eggshell quality. Overall, we uncovered the characteristics of proteomics of three eggshell matrix fractions and identified candidate proteins influencing eggshell quality. The next research on differential proteins will uncover the potential mechanisms underlying how proteins affect eggshell quality. BIOLOGICAL SIGNIFICANCE: It was reported that the proteins in an eggshell can be divided into insoluble and soluble proteins. The insoluble proteins are thought to be an inter-mineral matrix and acts as a structural framework, while the soluble proteins are thought as intra-mineral matrix that are embedded within the crystal during calcification. However, the difference between matrix fractions is unknown. Cross-analysis of proteomic data of three matrix fractions from the same type of eggshell, uncovered triply overlapped common proteins formed the predominant contributor to proteomic abundance of any matrix fraction, and we suggested that abundance variance of some common proteins between the three matrix fractions might be an important cause of their solubility differences. Moreover, eggshell is formed in hen's uterus, and uterus tend to be considered as unique organ determining eggshell quality. By cross-analysis on proteomic data of three matrix fractions between two eggshell models, normal and pimpled eggshells, the differential proteins were screened as candidates influencing eggshell quality. And we suggested that the liver and spleen or lymphocytes might be the major organs influencing eggshell quality, because the most promising candidates are almost blood and non-collagenous proteins, and originated from above organs.

Keratan sulfate glycosaminoglycan from chicken egg white.

Keratan sulfate (KS) was isolated from chicken egg white in amounts corresponding to ∼0.06 wt% (dry weight). This KS had a weight-average molecular weight of ∼36-41 kDa with a polydispersity of ∼1.3. The primary repeating unit present in chicken egg white KS was →4) ß-N-acetyl-6-O-sulfo-d-glucosamine (1 → 3) ß-d-galactose (1→ with some 6-O-sulfo galactose residues present. This KS was somewhat resistant to depolymerization using keratanase 1 but could be depolymerized efficiently through the use of reactive oxygen species generated using copper (II) and hydrogen peroxide. Of particular interest was the presence of substantial amounts of 2,8- and 2,9-linked N-acetylneuraminic acid residues in the form of oligosialic acid terminating the non-reducing ends of the KS chains. Most of the KS appears to be N-linked to a protein core as evidenced by its sensitivity to PNGase F.

Compositional analysis and structural elucidation of glycosaminoglycans in chicken eggs.

Glycosaminoglycans (GAGs) have numerous applications in the fields of pharmaceuticals, cosmetics, nutraceuticals, and foods. GAGs are also critically important in the developmental biology of all multicellular animals. GAGs were isolated from chicken egg components including yolk, thick egg white, thin egg white, membrane, calcified shell matrix supernatant, and shell matrix deposit. Disaccharide compositional analysis was performed using ultra high-performance liquid chromatography-mass spectrometry. The results of these analyses showed that all four families of GAGs were detected in all egg components. Keratan sulfate was found in egg whites (thick and thin) and shell matrix (calcified shell matrix supernatant and deposit) with high level. Chondroitin sulfates were much more plentiful in both shell matrix components and membrane. Hyaluronan was plentiful in both shell matrix components and membrane, but was only present in a trace of quantities in the yolk. Heparan sulfate was plentiful in the shell matrix deposit but was present in a trace of quantities in the egg content components (yolk, thick and thin egg whites). Most of the chondroitin and heparan sulfate disaccharides were present in the GAGs found in chicken eggs with the exception of chondroitin and heparan sulfate 2,6-disulfated disaccharides. Both CS and HS in the shell matrix deposit contained the most diverse chondroitin and heparan sulfate disaccharide compositions. Eggs might provide a potential new source of GAGs.

Spatiotemporal expression profile of a putative ß propeller WDR72 in laying hens.

The purpose of this study is to characterize the expression profile of a novel gene WDR72 in laying hens. Sixty-week old Hy-line Brown layers with similar laying sequence, egg weight, and shell strength, were selected and divided into 5 groups. The oviduct segments, such as magnum, white isthmus, and uterus, were sampled from each group of hens which were killed at 3 h post-oviposition (3 h P.O.), 4.15-4.5 h P.O., 8.5-9 h P.O., 12 h P.O. and 18 h P.O., respectively. To the 8.5-9 h P.O. hens, additional organs were also sampled besides oviduct tissues. Moreover, another group of hens with weak shell strength were selected and their oviduct segments were sampled at 12 h P.O. Then the expression profile of WDR72 was analyzed using real-time quantitative RT-PCR. The results showed as follows. (1) WDR72 transcripts specifically distributed in parts of organs investigated. At 8.5-9 h P.O., WDR72 appeared to be much more abundantly expressed in hens' oviduct sections, then followed in turn by brain, kidney, lung, glandular stomach and spleen. However, there were almost no WDR72 transcripts expressed in pectoral muscle, liver, heart and jejunum. (2) During the process of an "egg" passing through an oviduct, the expression of WDR72 in the magnum was greatly superior to that in the other two oviduct segments at 3 h P.O., 8.5-9 h P.O., and 12 h P.O.; while it was white isthmus in which WDR72 transcript levels were the highest at 4.15-4.5 h P.O. and 18 h P.O. (3) To any oviduct segment, not only uterus but also magnum and white isthmus, the expression of WDR72 in which was significantly up-regulated at the stages of active calcification. (4) WDR72 transcript levels in any oviduct segments of strong-shell hens were significantly higher than that of weak-shell layers (P < 0.01), which arose the possibility that WDR72 was positively associated with chicken eggshell strength. In conclusion, the expression profile of WDR72 gene in laying hens has been characterized, which would facilitate to further probe into its functions.

Microarray analysis of genes involved with shell strength in layer shell gland at the early stage of active calcification.

The objective of this study was to get a comprehensive understanding of how genes in chicken shell gland modulate eggshell strength at the early stage of active calcification. Four 32-week old of purebred Xianju hens with consistent high or low shell breakage strength were grouped into two pairs. Using Affymetrix Chicken Array, a whole-transcriptome analysis was performed on hen's shell gland at 9 h post oviposition. Gene ontology enrichment analysis for differentially expressed (DE) transcripts was performed using the web-based GOEAST, and the validation of DE-transcripts was tested by qRT-PCR. 1,195 DE-transcripts, corresponding to 941 unique genes were identified in hens with strong eggshell compared to weak shell hens. According to gene ontology annotations, there are 77 DE-transcripts encoding ion transporters and secreted extracellular matrix proteins, and at least 26 DE-transcripts related to carbohydrate metabolism or post-translation glycosylation modification; furthermore, there are 88 signaling DE-transcripts. GO term enrichment analysis suggests that some DE-transcripts mediate reproductive hormones or neurotransmitters to affect eggshell quality through a complex suite of biophysical processes. These results reveal some candidate genes involved with eggshell strength at the early stage of active calcification which may facilitate our understanding of regulating mechanisms of eggshell quality.

A putative transcriptional elongation factor hIws1 is essential for mammalian cell proliferation.

Iws1 has been implicated in transcriptional elongation by interaction with RNA polymerase II (RNAP II) and elongation factor Spt6 in budding yeast Saccharomyces cerevisiae, and association with transcription factor TFIIS in mammalian cells, but its role in controlling cell growth and proliferation remains unknown. Here we report that the human homolog of Iws1, hIws1, physically interacts with protein arginine methyltransferases PRMT5 which methylates elongation factor Spt5 and regulates its interaction with RNA polymerase II. Gene-specific silencing of hIws1 by RNA interference reveals that hIws1 is essential for cell viability. GFP fusion protein expression approaches demonstrate that the hIws1 protein is located in the nucleus, subsequently, two regions harbored within the hIws1 protein are demonstrated to contain nuclear localization signals (NLSs). In addition, mouse homolog of hiws1 is found to express ubiquitously in various tissues.