Detection of Enterotoxigenic Escherichia coli by DNA Colony Hybridization

Abstract

A method fo detecting large numbers of isolates of enterotoxigenic Escherichia coli is described in which the genes encoding th enterotoxins are detected, rather than the toxins themselves. Radiolabeled fragments of DNA encoding the heat-labile (LT) or heat-stable (ST) toxins were used as hybridization probes for homologous DNA sequences in E. coli colonies grown and lysed in situ on nitrocellulose filters. The LT probe detected all of 31 E. coli strains producing ST and LT or only LT, while the ST probe detected 12 of 17 strains producing only ST and three of 26 strains producing ST and LT. These results suggest that the LTs produced by different isolates of E. coli are homologous and that human isolates of E. coli produce at least two heterologous STs detectable in the infant mouse assay. The hybridization method also detected the presence of enterotoxigenic E. coli in bacterial growth in directly spotted stools from patients with acute diarrhea.

Prevalence and Genetic Profiling of Virulence Determinants of Non-O157 Shiga Toxin-ProducingEscherichia coliIsolated from Cattle, Beef, and Humans, Calcutta, India

“…A colony hybridization test for detecting pathogenic factors was carried out as described previously (26). DNA probes for espP, katP, eae, and hlyA were prepared by PCR.…”

Escherichia colidiarrhoea

“…The method is complicated by the existence of more than one kind of ST (see above). Nevertheless, surveys have been successfully carried out using ST probes and LT probes (Moseley et al, 1980(Moseley et al, , 1982Echeverria et al 1982). …”

Plasmid profiles and phage types ofSalmonella typhimuriumisolated from successive flocks of chickens on three parent stock farms

“…Agarplates were incubated at 37°C for 4 to 6 h, and colonies were lysed by the procedure described by Moseley et al (1980). Hybridization with a digoxigenin-dUTP labeled probe and posthybridization washes were carried out at 68°C according to the digoxigenin application manual (Anon., 1989), using 25 to 30 ng probe/ml hybridization solution, but with the modification that filters were gently wiped with a piece of paper cloth after one hour of prehybridization in order to decrease background signals resulting from cellular debris.…”