Su Liangke1, Yan Yaxian, Lu Chengping. 1. Shanghai Key Laboratory of Veterinary Biotechnology, School of Agriculture and Biology, Shanghai Jiaotong University, Shanghai 200240, China. suliangke@yahoo.com.cn
Abstract
OBJECTIVE: To construct an stx2 gene mutant phage phi Min27(delta stx::cat) and to observe its infectiousness of various serotypes Escherichia coli strains. METHODS: With the help of Red recombinant system, the stx2 gene of the E. coli O157:H7 Min27 strain isolated from intestinal feces of piglet with diarrhea at a swine farm of Shanghai, was replaced by the chloramphenicol acetyltrasferase (cat) gene from plasmid pLacI. Phage phi Min27(delta stx::cat) was isolated after induction of E. coli Min27(delta stx::cat) strain with mitomycin C. Twenty-one E. coli strains with various serotypes were infected with phi Min27(delta stx::cat), and plaque formation and lysogenic conversion of them were investigated. RESULTS: Of the 21 E. coli isolates, 2 with the serotypes of O60 and O138 integrated the phi Min27(delta stx::cat) in their chromosomes and expressed resistance to chloramphenicol. With the exception of one laboratory E. coli strain MG1655, none of the tested E. coli strains supported the formation of plaques and lysogenization when used as indicators for phi Min27(delta stx::cat). Following induction with mitomycin C, these lysogenic strains released infectious particles of phi Min27(delta stx::cat) that formed plaques on a lawn of E.coli laboratory strain MC1061. CONCLUSION: These results demonstrated that phi Min27(delta stx::cat) was able to infect and lysogenize particular E. coli strains and that the lysogens could produce infectious phage progeny. It could be inferred that Stx bacteriophages were able to spread exogenous genes among E. coli strains. The work provided a basis for further study on mechanisms of Stx phages infection and control of Stx expression.
OBJECTIVE: To construct an stx2 gene mutant phage phi Min27(delta stx::cat) and to observe its infectiousness of various serotypes Escherichia coli strains. METHODS: With the help of Red recombinant system, the stx2 gene of the E. coli O157:H7 Min27 strain isolated from intestinal feces of piglet with diarrhea at a swine farm of Shanghai, was replaced by the chloramphenicol acetyltrasferase (cat) gene from plasmid pLacI. Phage phi Min27(delta stx::cat) was isolated after induction of E. coli Min27(delta stx::cat) strain with mitomycin C. Twenty-one E. coli strains with various serotypes were infected with phi Min27(delta stx::cat), and plaque formation and lysogenic conversion of them were investigated. RESULTS: Of the 21 E. coli isolates, 2 with the serotypes of O60 and O138 integrated the phi Min27(delta stx::cat) in their chromosomes and expressed resistance to chloramphenicol. With the exception of one laboratory E. coli strainMG1655, none of the tested E. coli strains supported the formation of plaques and lysogenization when used as indicators for phi Min27(delta stx::cat). Following induction with mitomycin C, these lysogenic strains released infectious particles of phi Min27(delta stx::cat) that formed plaques on a lawn of E.coli laboratory strain MC1061. CONCLUSION: These results demonstrated that phi Min27(delta stx::cat) was able to infect and lysogenize particular E. coli strains and that the lysogens could produce infectious phage progeny. It could be inferred that Stx bacteriophages were able to spread exogenous genes among E. coli strains. The work provided a basis for further study on mechanisms of Stx phages infection and control of Stx expression.