Typing of Intimin Genes in Human and Animal Enterohemorrhagic and Enteropathogenic Escherichia coli: Characterization of a New Intimin Variant

doi:10.1128/IAI.68.1.64-71.2000

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Infection and Immunity, January 2000, p. 64-71, Vol. 68, No. 1
0019-9567/0/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Typing of Intimin Genes in Human and Animal Enterohemorrhagic and Enteropathogenic Escherichia coli: Characterization of a New Intimin Variant

E. Oswald,¹^,^* H. Schmidt,² S. Morabito,³ H. Karch,² O. Marchès,¹ and A. Caprioli³

Unité INRA-ENVT de Microbiologie Moléculaire, Ecole Vétérinaire de Toulouse, 31076 Toulouse Cedex, France¹; Institut für Hygiene und Mikrobiologie, University of Wurzburg, Wurzburg 97080, Germany²; and Laboratorio di Medicina Veterinaria, Istituto Superiore di Sanità, 00161 Rome, Italy³

Received 26 July 1999/Returned for modification 8 September 1999/Accepted 30 September 1999

Enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli (EHEC) produce the characteristic "attaching and effacing"(A/E) lesion of the brush border. Intimin, an outer membrane proteinencoded by eae, is responsible for the tight association of bothpathogens with the host cell. Several eae have been cloned fromdifferent EPEC and EHEC strains isolated from humans and animals.These sequences are conserved in the N-terminal region but highlyvariable in the last C-terminal 280 amino acids (aa), where thecell binding activity is localized. Based on these considerations,we developed a panel of specific primers to investigate the eaeheterogeneity of the variable 3' region by using PCR amplification.We then investigated the distribution of the known intimin typesin a large collection of EPEC and EHEC strains isolated from humansand different animal species. The existence of a yet-unknown familyof intimin was suspected because several EHEC strains, isolatedfrom human and cattle, did not react with any of the specificprimer pairs, although these strains were eae positive when primersamplifying the conserved 5' end were used. We then cloned andsequenced the eae present in one of these strains (EHEC of serotypeO103:H2) and subsequently designed a PCR primer that recognizesin a specific manner the variable 3' region of this new intimintype. This intimin, referred to as " $varepsilon$ ," was present in human andbovine EHEC strains of serogroups O8, O11, O45, O103, O121, andO165. Intimin $varepsilon$ is the largest intimin cloned to date (948 aa)and shares the greatest overall sequence identity with intimin $beta$ , although analysis of the last C-terminal 280 aa suggests agreater similarity with intimins $alpha$ and $gamma$ .

^* Corresponding author. Mailing address: Unité mixte INRA-ENVT de Microbiologie Moléculaire, 23 chemin des Capelles, 31076Toulouse Cedex, France. Phone: (33) 561-19-39-91. Fax: (33) 561-19-39-75.E-mail: e.oswald{at}envt.fr.

Infection and Immunity, January 2000, p. 64-71, Vol. 68, No. 1
0019-9567/0/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

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