title: Signature-tagged mutagenesis screening revealed a novel smooth-to-rough transition determinant of Salmonella enterica serovar Enteritidis.
abstract: Salmonella enterica serovar Enteritidis (S. Enteritidis) has emerged as one of the most important food-borne pathogens for humans. Lipopolysaccharide (LPS), as a component of the outer membrane, is responsible for the virulence and smooth-to-rough transition in S. Enteritidis. In this study, we screened S. Enteritidis signature-tagged transposon mutant library using monoclonal antibody against somatic O9antigen (O9MAb) and O9factor rabbit antiserum to identify novel genes that are involved in smooth-to-rough transition.
  
  
  
  A total of 480 mutants were screened and one mutant with transposon insertion in rfbG gene had smooth-to-rough transition phenotype. In order to verify the role of rfbG gene, an rfbG insertion or deletion mutant was constructed using λ-Red recombination system. Phenotypic and biological analysis revealed that rfbG insertion or deletion mutants were similar to the wild-type strain in growth rate and biochemical properties, but the swimming motility was reduced. SE Slide Agglutination test and ELISA test showed that rfbG mutants do not stimulate animals to produce agglutinating antibody. In addition, the half-lethal dose (LD50) of the rfbG deletion mutant strain was 106.6-fold higher than that of the parent strain in a mouse model when injected intraperitoneally.
  
  
  
  These data indicate that the rfbG gene is involved in smooth-to-rough transition, swimming motility and virulence of S. Enteritidis. Furthermore, somatic O-antigen antibody-based approach to screen signature-tagged transposon mutants is feasible to clarify LPS biosynthesis and to find suitable markers in DIVA-vaccine research.
authors.affiliation: Jiangsu Key Laboratory of Zoonosis, Yangzhou University, 48 East Wenhui Road, Yangzhou, Jiangsu, 225009, China.|Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Jiao
authors.affiliation: Jiangsu Key Laboratory of Zoonosis, Yangzhou University, 48 East Wenhui Road, Yangzhou, Jiangsu, 225009, China.|Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China.
authors.forename: Rongxian
authors.initials: R
authors.lastname: Guo
authors.affiliation: Jiangsu Key Laboratory of Zoonosis, Yangzhou University, 48 East Wenhui Road, Yangzhou, Jiangsu, 225009, China.|Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China.
authors.forename: Peipei
authors.initials: P
authors.lastname: Tang
authors.affiliation: Jiangsu Key Laboratory of Zoonosis, Yangzhou University, 48 East Wenhui Road, Yangzhou, Jiangsu, 225009, China.|Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China.
authors.forename: Xilong
authors.initials: X
authors.lastname: Kang
authors.affiliation: Jiangsu Key Laboratory of Zoonosis, Yangzhou University, 48 East Wenhui Road, Yangzhou, Jiangsu, 225009, China.|Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China.
authors.forename: Junlei
authors.initials: J
authors.lastname: Yin
authors.affiliation: Jiangsu Key Laboratory of Zoonosis, Yangzhou University, 48 East Wenhui Road, Yangzhou, Jiangsu, 225009, China.|Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China.
authors.forename: Kaiyue
authors.initials: K
authors.lastname: Wu
authors.affiliation: Jiangsu Key Laboratory of Zoonosis, Yangzhou University, 48 East Wenhui Road, Yangzhou, Jiangsu, 225009, China.|Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China.
authors.forename: Shizhong
authors.initials: S
authors.lastname: Geng
authors.affiliation: Jiangsu Key Laboratory of Zoonosis, Yangzhou University, 48 East Wenhui Road, Yangzhou, Jiangsu, 225009, China.|Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China.
authors.forename: Qiuchun
authors.initials: Q
authors.lastname: Li
authors.affiliation: Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China.|Division of Gastroenterology and Hepatology Department of Medicine, University of Illinois at Chicago, 840 S Wood Street, Room 704 CSB, 60612, Chicago, IL, USA.
authors.forename: Jun
authors.initials: J
authors.lastname: Sun
authors.affiliation: Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China.
authors.forename: Xiulong
authors.initials: X
authors.lastname: Xu
authors.affiliation: Pathobiology and Veterinary Science, College of Agriculture, Health and Natural Resources, University of Connecticut, 61 North Eagleville Road, Unit-3089, Mansfield, CT, USA.
authors.forename: Xiaohui
authors.initials: X
authors.lastname: Zhou
authors.affiliation: Jiangsu Key Laboratory of Zoonosis, Yangzhou University, 48 East Wenhui Road, Yangzhou, Jiangsu, 225009, China.|Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China.
authors.forename: Junji
authors.initials: J
authors.lastname: Gan
authors.affiliation: Jiangsu Key Laboratory of Zoonosis, Yangzhou University, 48 East Wenhui Road, Yangzhou, Jiangsu, 225009, China.|Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China.
authors.forename: Xinan
authors.initials: X
authors.lastname: Jiao
authors.affiliation: Jiangsu Key Laboratory of Zoonosis, Yangzhou University, 48 East Wenhui Road, Yangzhou, Jiangsu, 225009, China. xfliu@yzu.edu.cn.|Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China. xfliu@yzu.edu.cn.
authors.forename: Xiufan
authors.initials: X
authors.lastname: Liu
authors.affiliation: Jiangsu Key Laboratory of Zoonosis, Yangzhou University, 48 East Wenhui Road, Yangzhou, Jiangsu, 225009, China. zmpan@yzu.edu.cn.|Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009, China. zmpan@yzu.edu.cn.
authors.forename: Zhiming
authors.initials: Z
authors.lastname: Pan
mesh_terms: D000372:Agglutination Tests / Q000379:methods; D000818:Animals; D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins / Q000235:genetics*; D004251:DNA Transposable Elements / Q000235:genetics; D004195:Disease Models, Animal; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005798:Genes, Bacterial; D007928:Lethal Dose 50; D008070:Lipopolysaccharides / Q000096:biosynthesis; D016296:Mutagenesis; D016254:Mutagenesis, Insertional; D019081:O Antigens / Q000235:genetics / Q000276:immunology; D010641:Phenotype; D011817:Rabbits; D012481:Salmonella Infections, Animal / Q000517:prevention & control; D012477:Salmonella enteritidis / Q000235:genetics* / Q000254:growth & development / Q000302:isolation & purification* / Q000472:pathogenicity; D014774:Virulence / Q000235:genetics*
chemical_list: D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D008070:Lipopolysaccharides; D019081:O Antigens; C079397:rfb protein, Bacteria
keywords: O9 MAb; S. Enteritidis; Signature-tagged mutagenesis (STM); Smooth-to-rough transition; rfbG gene