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Infection and Immunity, October 2000, p. 5710-5715, Vol. 68, No. 10
0019-9567/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

IS1414, an Escherichia coli Insertion Sequence with a Heat-Stable Enterotoxin Gene Embedded in a Transposase-Like Gene

Annette McVeigh,1,2 Alessio Fasano,3 Daniel A. Scott,1 Sandra Jelacic,4 Steve L. Moseley,4 Donald C. Robertson,5 and Stephen J. Savarino1,*

Enteric Diseases Department, Naval Medical Research Center, Silver Spring,1 Henry M. Jackson Foundation for the Advancement of Military Medicine, Rockville,2 and Center for Vaccine Development and Department of Pediatrics, University of Maryland School of Medicine, Baltimore,3 Maryland; Department of Microbiology, University of Washington, Seattle, Washington4; and Department of Molecular Biology, Biochemistry and Microbiology, University of Idaho, Moscow, Idaho5

Received 20 December 1999/Returned for modification 2 March 2000/Accepted 6 July 2000

Enteroaggregative Escherichia coli (EAEC) heat-stable enterotoxin 1 (EAST1) was originally discovered in EAEC but has also been associated with enterotoxigenic E. coli (ETEC). Multiple genomic restriction fragments from each of three ETEC strains of human origin showed homology with an EAST1 gene probe. A single hybridizing fragment was detected on the plasmid of ETEC strain 27D that also encodes heat-stable enterotoxin Ib and colonization factor antigen I. We isolated and characterized this fragment, showing that it (i) carries an allele of astA nearly identical to that originally reported from EAEC 17-2 and (ii) expressed enterotoxic activity. Sequence analysis of the toxin coding region revealed that astA is completely embedded within a 1,209-bp open reading frame (ORF1), whose coding sequence is on the same strand but in the -1 reading frame in reference to the toxin gene. In vitro expression of the predicted Mr-~46,000 protein product of ORF1 was demonstrated. ORF1 is highly similar to transposase genes of IS285 from Yersinia pestis, IS1356 from Burkholderia cepacia, and ISRm3 from Rhizobium meliloti. It is bounded by 30-bp imperfect inverted repeat sequences and flanked by 8-bp direct repeats. Based on these structural features, pathognomonic of a regular insertion sequence, this element was designated IS1414. Preliminary experiments to show IS1414 translocation were unsuccessful. Overlapping genes of the type suggested by the IS1414 core region have heretofore not been described in bacteria. It seems to offer a most efficient mechanism for intragenomic and horizontal dissemination of EAST1.

* Corresponding author. Mailing address: Naval Medical Research Center, 503 Robert Grant Ave., Silver Spring, MD 20910-7500. Phone: (301) 319-7663. Fax: (301) 319-7679. E-mail: savarinos{at}nmrc.navy.mil.

Infection and Immunity, October 2000, p. 5710-5715, Vol. 68, No. 10
0019-9567/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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