Wen Ming Zhao

Effects of dietary probiotic supplementation on LXRα and CYP7α1 gene expression, liver enzyme activities and fat metabolism in ducks

Abstract 1. The objective of this study was to investigate the effects of dietary probiotic supplementation on liver X receptor alpha (LXRα) and cholesterol 7α-hydroxylase (CYP7α1) mRNA levels, protein enzymatic activities and fat metabolism in Cherry Valley Pekin ducks. 2. A total of 750 one-day-old Cherry Valley Pekin ducks were randomly divided into 5 groups with three replicates of 50 ducks each in a completely randomised experiment. Each group was fed on a basal diet supplemented with 0, 500, 1000, 1500 or 2000 mg probiotics/kg. 3. Body rate and feed conversion ratio were highest and abdominal subcutaneous fat % was lowest at 1000 mg probiotic/kg. 4. The mRNA levels of LXRα and CYP7α1 in liver tissue was estimated by RT-PCR; serum triglyceride (TG) and total cholesterol (TC) concentrations were measured by ELISA. 5. The expression levels and enzyme activity of LXRα and CYP7α1 increased in conjunction with decreases in TG and TC concentrations following probiotic supplementation to a maximum at 1000 mg probiotics/kg and decreased thereafter. 6. It is concluded that dietary probiotics can enhance LXRα and CYP7α1 enzyme activities in the liver and reduce lipid concentrations and fat deposition in ducks.

DNA Methylation and Regulation of the CD8A after Duck Hepatitis Virus Type 1 Infection

Background Cluster of differentiation 8 (CD8) is expressed in cytotoxic T cells, where it functions as a co-receptor for the T-cell receptor by binding to major histocompatibility complex class I (MHCI) proteins, which present peptides on the cell surface. CD8A is critical for cell-mediated immune defense and T-cell development. CD8A transcription is controlled by several cis-acting elements and trans-acting elements and is also regulated by DNA methylation. However, the epigenetic regulation of CD8A in the duck and its relationship with virus infection are still unclear. Results We investigated the epigenetic transcriptional regulatory mechanisms, such as DNA methylation, for the expression of the CD8A and further evaluated the contribution of such epigenetic regulatory mechanisms to DHV-I infection in the duck. Real-time quantitative polymerase chain reaction (RT-qPCR) revealed the highest level of CD8A expression to be in the thymus, followed by the lungs, spleen, and liver, and the levels of CD8A expression were very low in the kidney, cerebrum, cerebellum, and muscle in the duck. RT-qPCR also demonstrated that the CD8A mRNA was down-regulated significantly in morbid ducklings treated with DHV-1 and up-regulated significantly in non-morbid ducklings in all the tissues tested. In addition, hypermethylation of CD8A was detected in the morbid ducklings, whereas relatively low methylation of CD8A was evident in the non-morbid ducklings. The CD8A mRNA level was negatively associated with the CpG methylation level of CD8A and global methylation status. Conclusions We concluded that the mRNA level of the CD8A was negatively associated with the CpG methylation level of CD8A and global methylation status in the duck, suggesting that the hypermethylation of CD8A may be associated with DHV-1 infection. The first two CpG sites of the CD8A promoter region could be considered as epigenetic biomarkers for resistance breeding against duckling hepatitis disease in the duck.

Polymorphism in Intron-1 of ApoVLDL-II Gene and its Association with Production Traits and Biochemical Levels in White Muscovy (Cairina moschata)

Abstract Zhang, Y.Y., Chen, G.H., Xu, Q., Zhang, H.B., Duan, X.J., Zhao, W.M. and Sun, G.B. 2010. Polymorphism in intron-1 of ApoVLDL-II gene and its association with production traits and biochemical levels in white muscovy (Cairina moschata). J. Appl. Anim. Res., 38: 23–28. The current study was designed to investigate the possible association of apoVLDL-II gene intron-1 polymorphism with White Muscovy production traits and biochemical levels. There were two alleles (C and D) and three genotypes (CC, CD and DD) in the experimental population. CC genotype and C allele frequency were the highest. One SNP (G803A) and one 13bp insertion deletion AAAATCTTGTTTA after position 770bp were found by sequencing. The apoVLDL-II gene intron-1 polymorphisms were associated (P<0.05) with body slope length, gizzard weight, pH, percentage of chest muscle water, intramuscular fat, serum triglyceride and serum cholinesterase in the 120 individuals. The CC may be an advantage genotype for growth and lipid transportation in White Muscovy. The apoVLDL-II gene intron-1 polymorphisms could be used in marker assisted selection (MAS) as a genetic marker for the birds growth and meat quality.

The CD8α gene in duck (Anatidae): cloning, characterization, and expression during viral infection

Cluster of differentiation 8 alpha (CD8α) is critical for cell-mediated immune defense and T-cell development. Although CD8α sequences have been reported for several species, very little is known about CD8α in ducks. To elucidate the mechanisms involved in the innate and adaptive immune responses of ducks, we cloned CD8α coding sequences from domestic, Muscovy, Mallard, and Spotbill ducks using reverse transcription polymerase chain reaction (RT-PCR). Each sequence consisted of 714 nucleotides and encoded a signal peptide, an IgV-like domain, a stalk region, a transmembrane region, and a cytoplasmic tail. We identified 58 nucleotide differences and 37 amino acid differences among the four types of duck; of these, 53 nucleotide and 33 amino acid differences were between Muscovy ducks and the other duck species. The CD8α cDNA sequence from domestic duck consisted of a 61-nucleotide 5′ untranslated region (UTR), a 714-nucleotide open reading frame, and an 849-nucleotide 3′ UTR. Multiple sequence alignments showed that the amino acid sequence of CD8α is conserved in vertebrates. RT-PCR revealed that expression of CD8α mRNA of domestic ducks was highest in the thymus and very low in the kidney, cerebrum, cerebellum, and muscle. Immunohistochemical analyses detected CD8α on the splenic corpuscle and periarterial lymphatic sheath of the spleen. CD8α mRNA in domestic ducklings was initially up-regulated, and then down-regulated, in the thymus, spleen, and liver after treatment with duck hepatitis virus type I (DHV-1) or the immunostimulant polyriboinosinic polyribocytidylic acid (poly I:C).

Transcriptome profiling to identify key mediators of granulosa cell proliferation upon FSH stimulation in the goose (Anser cygnoides).

ABSTRACT 1. The low reproductive performance of geese has seriously hampered the development of the industry. Reproductive performance, particularly the egg laying rate mainly depends on the development of the follicle. Previous studies have shown that follicle-stimulating hormone (FSH) plays an important role in the process of follicular development, but the exact underlying mechanism remains unclear. 2. This study showed that FSH stimulated granulosa cell proliferation in a dose-dependent manner. The effect of FSH treatment on granulosa cell proliferation was greatest at a dose of 100 mIU/ml FSH for 24 h. 3. Secondly, the effect of different concentrations of FSH on goose granulosa cell proliferation was investigated, and de novo transcriptome assembly and gene expression analysis performed using short-read sequencing technology (Illumina). High-throughput sequencing results yielded 62.61 M reads and 7.8 G base pairs from granulosa cells treated with 100 mIU/ml FSH. These reads were assembled into 65,757 unigenes (mean length: 705 bp) with an N50 of 903 bp. A total of 110 upregulated and 510 downregulated differentially expressed genes (DEGs) were identified by RNA-seq. 4. Functional analysis by gene ontology (GO) and KEGG pathway annotation indicated that hormone biosynthesis (GO:0042446), positive regulation of hormone secretion (GO:0046887), steroid biosynthesis, oxidative phosphorylation and carbon metabolism pathways were involved in FSH-mediated proliferation of goose granulosa cells. 5. After screening, a group of key responsive genes including superoxide dismutase 1, fatty acyl-CoA reductase 1, transforming growth factor-beta receptor-associated protein 1 and follistatin were tested by real-time reverse transcription PCR to confirm differential expression in granulosa cells stimulated by FSH. 6. FSH-stimulated goose granulosa cells and DEG profiling data provided comprehensive gene expression information at the transcriptional level that could promote better understanding of the molecular mechanisms underlying follicle development in response to FSH stimulation.

Effects of forage feeding versus grain feeding on the growth performance and meat quality of Yangzhou geese

ABSTRACT 1. Highly palatable and nutritious meat products can be produced through dietary interventions. Previous studies have shown that forage feeding has a significant impact on the growth performance and nutrition of cattle in various regions, but whether the same effects can be induced in geese remains unclear. 2. Three hundred and sixty Yangzhou goslings were divided according to body weight at 29 d old, assigned to one of 4 treatments and raised in separate pens. The treatments applied were (A) grazing, (B) grazing, grain supplemental diet (64 to 70 d), (C) grazing, grain supplemental diet and (D) confined. 3. Eviscerated carcass yield was lower in the grazing treatment. Protein content and muscle collagen in both the breast and thigh muscles were significantly higher in the grazing treatment than the confined, while fat content exhibited the opposite tendency. Those fed on grass and supplementary grain had a higher Mg and Cu content in breast muscle. 4. Geese will grow to their full potential when they are allowed to consume grass from pasture supplemented with grain, protein, collagen, Mg and Cu content was greater to a degree, which suggests this feeding regime is an ideal model for goose production.

Variations in endothelin receptor B subtype 2 (EDNRB2) coding sequences and mRNA expression levels in 4 Muscovy duck plumage colour phenotypes

ABSTRACT 1. Endothelin receptor B subtype 2 (EDNRB2) is a paralog of EDNRB, which encodes a 7-transmembrane G-protein coupled receptor. Previous studies reported that EDNRB was essential for melanoblast migration in mammals and ducks. 2. Muscovy ducks have different plumage colour phenotypes. Variations in EDNRB2 coding sequences (CDSs) and mRNA expression levels were investigated in 4 different Muscovy duck plumage colour phenotypes, including black, black mutant, silver and white head. 3. The EDNRB2 gene from Muscovy duck was cloned; it had a length of 6435 bp and encoded 437 amino acids. The coding region was screened and potential single nucleotide polymorphisms were identified. Eight mutations were obtained, including one missense variant (c.64C > T) and 7 synonymous substitutions. The substitutions were associated with plumage colour phenotypes. 4. The EDNRB2 mRNA expression levels were compared between feather pulp from black birds and black mutant birds. The results indicated that EDNRB2 transcripts in feather pulp were significantly higher in black feathers than in white feathers. 5. The results determined the variation of EDNRB2 CDS and mRNA expression in Muscovy ducks of various plumage colours.

Molecular cloning of the SMAD4 gene and its mRNA expression analysis in ovarian follicles of the Yangzhou goose (Anser cygnoides).

Abstract Mothers against decapentaplegic homolog 4 (SMAD4) is an important protein in animal reproduction. It plays pivotal roles in cellular pathways, including apoptosis. The expression profile of the SMAD4 gene in goose ovarian follicles has not been reported. In this study, the SMAD4 coding sequence was cloned from the Yangzhou goose. A phylogenetic analysis was performed and mRNA expression was examined in various tissues using quantitative real-time PCR. An alternative splice form of SMAD4, SMAD4-b having 1656 bp, was identified. SMAD4-a mRNA was widely expressed in various healthy tissues, whereas SMAD4-b was very weakly expressed. SMAD4 mRNA in the ovary and oviduct was significantly higher than that in the pituitary and hypothalamus. SMAD4 mRNA expression analysis in hierarchical follicles showed that the level of SMAD4 mRNA was higher in large white follicles and post-ovulatory follicles than in the other follicles. The results indicate that SMAD4 might be involved in the recruitment of hierarchical follicles.