A chromosomally lux-marked (Tn5 luxCDABE) strain of nontoxigenic Escherichia coli O157:H7 was constructed by transposon mutagenesis and shown to have retained the O157, H7, and intimin phenotypes. The survival characteristics of this strain in the experiments performed (soil at ؊5, ؊100, and ؊1,500 kPa matric potential and artificial groundwater) were indistinguishable from the wild-type strain. Evaluation of potential luminescence was found to be a rapid, cheap, and quantitative measure of viable E. coli O157:H7 Tn5 luxCDABE populations in environmental samples. In the survival studies, bioluminescence of the starved populations of E. coli O157:H7 Tn5 luxCDABE could be reactivated to the original levels of light emission, suggesting that these populations remain viable and potentially infective to humans. The attributes of the construct offer a cheap and low-risk substitute to the use of verocytotoxin-producing E. coli O157:H7 in long-term survival studies.

Aims: To evaluate the suitability of a multiplex PCR‐based assay for sensitive and rapid detection of Escherichia coli O157:H7 in soil and water. Methods and Results: Soil and water samples were spiked with E. coli O157:H7 and subjected to two stages of enrichment prior to multiplex PCR. Detection sensitivities were as high as 1 cfu ml–1 drinking water and 2 cfu g–1 soil. Starvation of E. coli O157:H7 for 35 d prior to addition to soil did not affect the ability of the assay to detect initial cell numbers as low as 10 cfu g–1 soil. Use of an 8‐h primary enrichment enabled detection of as few as 6 cfu g–1 soil, and 104 cfu g–1 soil with a 6‐h primary enrichment. When soil was inoculated with 105 cfu g–1, the PCR assay indicated persistence of E. coli O157:H7 during a 35 d incubation. However, when soil was inoculated with lower numbers of pathogen, PCR amplification signals indicated survival to be dependent on cell concentration. Conclusions: A multiplex PCR‐based assay, in combination with an enrichment strategy enabled sensitive and rapid detection of E. coli O157:H7 in soil and water. Significance and Impact of the Study: The ability to sensitively detect E.coli O157:H7 in environmental material within one working day represents a considerable advancement over alternative more time‐consuming methods for detection of this pathogen.