Transduction of porcine enteropathogenic Escherichia coli with a derivative of a shiga toxin 2-encoding bacteriophage in a porcine ligated ileal loop system

Abstract

In this study, we have investigated the ability of detoxified Shiga toxin (Stx)-converting bacteriophages Phi3538 (Deltastx(2)::cat) (H. Schmidt et al., Appl. Environ. Microbiol. 65:3855-3861, 1999) and H-19B::Tn10d-bla (D. W. Acheson et al., Infect. Immun. 66:4496-4498, 1998) to lysogenize enteropathogenic Escherichia coli (EPEC) strains in vivo. We were able to transduce the porcine EPEC strain 1390 (O45) with Phi3538 (Deltastx(2)::cat) in porcine ligated ileal loops but not the human EPEC prototype strain E2348/69 (O127). Neither strain 1390 nor strain E2348/69 was lysogenized under these in vivo conditions when E. coli K-12 containing H-19B::Tn10d-bla was used as the stx1 phage donor. The repeated success in the in vivo transduction of an Stx2-encoding phage to a porcine EPEC strain in pig loops was in contrast to failures in the in vitro trials with these and other EPEC strains. These results indicate that in vivo conditions are more effective for transduction of Stx2-encoding phages than in vitro conditions.

FIG. 1.

Agarose gel electrophoresis of stx₂::cat-specific (A) and eae-specific (B) PCR products. Samples: M, λ phage DNA digested with HindIII; 1, stx₂::cat donor 3538, eae⁺; 2, EPEC recipient strain 1390, eae⁺; 3, transductant E-12 (stx₂::cat), eae⁺; 4, transductant DJ-04 (stx₂::cat), eae⁺; 5, transductant ME-08 (stx₂::cat), eae⁺; 6, E. coli C600 (negative control).

FIG. 2.

RFLP of stx₂::cat-specific PCR products. The undigested (A) and PvuII-digested (B) stx₂::cat amplicons were electrophoresed in a 1% agarose gel. Samples: M, λ phage DNA digested with HindIII; lane 1, strain 3538; 2, strain 1390; 3, strain E12; 4, strain DJ-04; 5, strain ME-08. The PvuII-digested stx₂::cat-specific PCR uniformly yielded two fragments of 257 and 1,310 bp, respectively.