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Meng X.,Yangzhou University | Zhu C.,Jiangsu Institute of Poultry Science | Wang H.,Yangzhou University | Wang J.,Beijing Agricultural Vocational College | And 3 more authors.
FEMS Microbiology Letters | Year: 2013

Salmonella Enteritidis is an intracellular pathogen that causes enteritis and systemic disease in humans and other animals. The RNA chaperone protein Hfq mediates the binding of small noncoding RNAs to target mRNA and assists in post-transcriptional gene regulation in bacteria. In this study, we constructed an hfq deletion mutant in S. Enteritidis SE50336 and analyzed the expression of major fimbrial subunits sefA, bcfA, fimA, safA, stbA, sthA, csgA, csgD, and pegA using quantitative real-time PCR. The gene expression of sefA increased about 14-fold in the hfq mutant, as compared with its expression in the wild-type strain. The expression of fimA and pegA did not change significantly, while the expression of the other fimbrial genes was significantly down-regulated in the hfq mutant. The ability of SE50336Δhfq adhering to Caco-2 cells was also reduced as compared with wild-type adherence. The virulence of the hfq mutant was significantly reduced in a 1-day-old chicken model of S. Enteritidis disease, as determined by quantifying the lethal dose 50% of the bacterial strains. We conclude that Hfq critically contributes to S. Enteritidis virulence, likely partially affected by regulating fimbrial gene expression. © 2013 Federation of European Microbiological Societies.

Li Y.,Changshu Institute of Technology | Wang X.,Changshu Institute of Technology | Wang X.,Jiangsu Institute of Poultry Science | Yu J.,Changshu Institute of Technology | And 5 more authors.
Poultry Science | Year: 2016

As the most abundant microRNA (miRNA) in the liver, miR-122 plays important roles in the growth and development of liver, lipid metabolism, and liver diseases. Vanin 1 (VNN1) plays an important role in hepatic lipid metabolism, and VNN1 may serve as a potential therapeutic target for the treatment of metabolic diseases caused by overactivated gluconeogenesis. In our previous RNA-seq study, we found the expression of VNN1 increased significantly when the expression of miR-122 (gga-miR-122-5p) was knocked down in primary chicken hepatocytes. In this study, we verified this result by real-time qRT-PCR, and we also found that the chicken VNN1 was highly expressed in the liver. By bioinformatics analyses, we found the 3′UTR of VNN1 contained sequences completely complementary to the nucleotides 1 to 8 of miR-122. Co-transfection and dual-luciferase reporter assays showed that overexpression of miR-122 decreased the expression of luciferase reporter gene linked to the 3′UTR of chicken VNN1 in the Chinese hamster ovary cells (P < 0.01), and the decrease was further demonstrated to be dependent on the predicted miR-122 binding sites by site mutation analyses. These results further support miR-122 as a negative regulator of VNN1 expression in chicken hepatocytes. Overall, this study suggests that miR-122 might play an important role in lipid metabolism in the chicken liver by negatively regulating the expression of the VNN1 gene. © 2016 Poultry Science Association Inc.

Sun C.,China Agricultural University | Lu J.,Jiangsu Institute of Poultry Science | Yi G.,China Agricultural University | Yuan J.,China Agricultural University | And 5 more authors.
PLoS ONE | Year: 2015

Because it serves as the cytoplasm of the oocyte and provides a large amount of reserves, the egg yolk has biological significance for developing embryos. The ovary and its hierarchy of follicles are the main reproductive organs responsible for yolk deposition in chickens. However, the genetic architecture underlying the yolk and ovarian follicle weights remains elusive. Here, we measured the yolk weight (YW) at 11 age points from onset of egg laying to 72 weeks of age and measured the follicle weight (FW) and ovary weight (OW) at 73 weeks as part of a comprehensive genome-wide association study (GWAS) in 1,534 F2 hens derived from reciprocal crosses between White Leghorn (WL) and Dongxiang chickens (DX). For all ages, YWs exhibited moderate single nucleotide polymorphism (SNP)- based heritability estimates (0.25-0.38), while the estimates for FW (0.16) and OW (0.20) were relatively low. Independent univariate genome-wide screens for each trait identified 12, 3, and 31 novel significant associations with YW, FW, and OW, respectively. A list of candidate genes such as ZAR1, STARD13, ACER1b, ACSBG2, and DHRS12 were identified for having a plausible function in yolk and follicle development. These genes are important to the initiation of embryogenesis, lipid transport, lipoprotein synthesis, lipid droplet promotion, and steroid hormone metabolism, respectively. Our study provides for the first time a genome-wide association (GWA) analysis for follicle and ovary weight. Identification of the promising loci as well as potential candidate genes will greatly advance our understanding of the genetic basis underlying dynamic yolk weight and ovarian follicle development and has practical significance in breeding programs for the alteration of yolk weight at different age points. © 2015 Sun et al.

Musa H.-H.,Yangzhou University | Musa H.-H.,University of Nyala | Zhang W.-J.,Yangzhou University | Lv J.,Yangzhou University | And 8 more authors.
Journal of Microbiology, Immunology and Infection | Year: 2014

Background: The fimbriae of Salmonella enterica serovar Enteritidis are used for colonization and invasion into host cells, and have drawn considerable interest because fimbriae can serve as potential immunogens against many pathogenic bacteria that colonize on epithelial surfaces. The purpose of the study is to use a molecular adjuvant, C3d, to enhance the immunogenicity of FimA proteins against Salmonella enterica serovar Enteritidis. Methods: FimA of type I fimbriae from Salmonella enteritidis and FimA with one copy of mC3d, two copies of mC3d2 and three copies of mC3d3 were cloned into the expression vector pCold-TF. Soluble fusion proteins of FimA with different copy of mC3d were induced by IPTG and expressed into Escherichia coli BL21 (DE3). Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) showed that the recombinant proteins from pCold-TF-fimA, TF-fimA-mC3d, TF-fimA-mC3d2, TF-fimA-mC3d3 were 70kDa, 100kDa, 130kDa and 160kDa, respectively. The fusion protein was recognized by rabbit anti-fimbriae polyclonal antibodies, and then visualized by goat anti-rabbit polyclonal antibodies with a chrome appearance by enzyme-subtract interaction. The recombinant proteins were purified by Ni-TED (tris-carboxymethyl ethylene diamine), immobilized metal ion affinity chromatography (IMAC). Balb/c mice were subcutaneously immunized with the purified proteins and the immune response was monitored by an enzyme-linked immunosorbent assay (ELISA) for FimA-specific antibody. The immunized mice were challenged with a 10-fold LD50 dose (i.e., 100 CFU) of Salmonella enterica serovar Enteritidis standard strain (SD-2) 1 week after the second immunization. Results: The immunized mice with the fusion proteins FimA-mC3d2 and FimA-mC3d3 had increased levels of ELISA titer of antibody that were 2 and 4 logs, respectively, more immunogenic than the TF-FimA protein alone. The challenge results showed that immune protection rate in the mice immunized with 10μg of FimA, FimA-mC3d2, and FimA-mC3d3 were 50%, 75% and 100%, respectively. Conclusion: We conclude that mC3d can be expressed in a prokaryotic vector and enhance the immune response of the recombinant protein. FimA-mC3d3 is potentially a subunit vaccine against S. enterica serovar Enteritidis infection. © 2012.

Wang Y.,China Agricultural University | Gao Y.,China Agricultural University | Imsland F.,Uppsala University | Gu X.,China Agricultural University | And 13 more authors.
PLoS ONE | Year: 2012

The Crest phenotype is characterised by a tuft of elongated feathers atop the head. A similar phenotype is also seen in several wild bird species. Crest shows an autosomal incompletely dominant mode of inheritance and is associated with cerebral hernia. Here we show, using linkage analysis and genome-wide association, that Crest is located on the E22C19W28 linkage group and that it shows complete association to the HOXC-cluster on this chromosome. Expression analysis of tissues from Crested and non-crested chickens, representing 26 different breeds, revealed that HOXC8, but not HOXC12 or HOXC13, showed ectopic expression in cranial skin during embryonic development. We propose that Crest is caused by a cis-acting regulatory mutation underlying the ectopic expression of HOXC8. However, the identification of the causative mutation(s) has to await until a method becomes available for assembling this chromosomal region. Crest is unfortunately located in a genomic region that has so far defied all attempts to establish a contiguous sequence. © 2012 Wang et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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