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Background    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 O 9  antigen (O 9  MAb) and O 9  factor rabbit antiserum to identify novel genes that are involved in smooth-to-rough transition.    Results  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 (LD 50 ) of the  rfbG  deletion mutant strain was 10 6.6  -fold higher than that of the parent strain in a mouse model when injected intraperitoneally.    Conclusions  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.

Signature-tagged mutagenesis screening revealed a novel smooth-to-rough transition determinant of Salmonella enterica serovar Enteritidis