Enterohemolysin production is associated with a temperate bacteriophage in Escherichia coli serogroup O26 strains

Abstract

A temperate bacteriophage that determines the expression of enterohemolysin was isolated from Escherichia coli 026 strain C3888. The genetic determinant associated with enterohemolysin production (E-Hly determinant) was cloned from EcoRI-digested bacteriophage DNA in vector plasmid pUC8. pUC8 recombinant plasmid pEOl9 carries a 3.7-kb EcoRI insert of phage DNA, and enterohemolysin was expressed in E. coli K-12 after transformation. Hemolysin-negative derivatives of pEOl9 were generated by transposon mutagenesis with Tnl725. By subcloning, the phage E-Hly determinant was assigned to a 2,150-bp piece of DNA which is flanked by EcoRI and AccI restriction sites. The enterohemolysin-producing recombinant strains and wild-type strain C3888 express a 60-kDa protein which was detected in the bacterial outer membrane by Western immunoblotting. Biologically active enterohemolysin was detected only in bacteria grown to the stationary phase, and the hemolysin was not released into the culture medium. Lysis of erythrocytes was inhibited by 30 mM dextran 4, which functions as an osmotic protectant without destroying the enterohemolysin itself.

The ability of haemolysins expressed by atypical enteropathogenic Escherichia coli to bind to extracellular matrix components

“…2). This observation differs from previous reports, which describe the lack of haemolytic activity in the culture supernatant of strain C3888 (Beutin et al 1990, Figueiredo et al 2003. However, these authors detected such activity in disrupted bacterial cells collected by centrifugation.…”

Prevalence of Hemolysin Genes and Comparison of ehxA Subtype Patterns in Shiga Toxin-Producing Escherichia coli (STEC) and Non-STEC Strains from Clinical, Food, and Animal Sources

“…The bacteriophage-associated enterohemolysin (e-hlyA) is genetically unrelated to the plasmid-carried ehxA (38)(39)(40). It was suggested that the e-hlyA determinant does not encode a hemolysin, per se, as reported in earlier studies (38,39), but instead causes the release of the silent hemolysin by lysis of the bacterial cells (40).…”

“…It was suggested that the e-hlyA determinant does not encode a hemolysin, per se, as reported in earlier studies (38,39), but instead causes the release of the silent hemolysin by lysis of the bacterial cells (40). The silent hemolysin (sheA), also called cytolysin A, encodes a pore-forming toxin (41,42) not related to the RTX family (43,44).…”

Thiol-independent activity of a cholesterol-binding enterohemolysin produced by enteropathogenic Escherichia coli

“…The gene structure of this hemolysin is still unknown. A 60-kDa outer membrane protein, present only in EHly-positive bacteria (5), may be the EHly itself, but no further information on the purification of this protein is available. In the present paper, we describe the purification of EHly and show that it shares several properties with thiolactivated hemolysins having the following general characteristics, in addition to being inactivated by cholesterol: inactivation by oxidation and reactivation with reducing agents, lethality to mice, and lytic activity towards eukaryotic cells through the formation of pores in the cell membrane (6).…”