Immunomagnetic separation and DNA hybridization for detection of enterotoxigenic Escherichia coli in a piglet model.

摘要

Enterotoxigenic Escherichia coli (ETEC) strains were detected by stool blot hybridization assays using different oligonucleotide probes for the colonization fimbrial antigen F4, heat-stable enterotoxin I (ST I), and heat-labile enterotoxin (LT I) genes. Forty-eight fecal samples and seven samples of intestinal content from ETEC-challenged newborn piglets were processed in two ways: (i) by direct inoculation of bacterial suspension onto nylon membranes overlaying blood agar and (ii) by immunomagnetic enrichment of F4+ ETEC using magnetic beads coated with F4 monoclonal antibodies before inoculation onto nylon membranes. In samples obtained from nondiarrheic piglets pre- and postchallenge, E. coli genes for F4, ST I, and LT I could be detected only after immunomagnetic enrichment. No difference between the two methods in detection of these E. coli genes was observed when stool blots from diarrheic piglets were examined. By using magnetic separation, it was easy to decrease background bacterial flora, intestinal cells, and fecal debris and thus produce purer specimens. The method evaluated in this animal model appeared simple and quick and increased the sensitivity of detection of ETEC strains 100-fold compared with the direct stool blot hybridization assays. Prior bacterial isolation and identification were not necessary.