Cloning and characterization of the genes encoding the haemolysin of Haemophilus ducreyi

Abstract

We previously identified a heat- and protease-labile haemolytic activity expressed by Haemophilus ducreyi. In order to characterize the haemolysin at the molecular level, genomic DNA from H. ducreyi was probed with haemolysin genes from other Gram-negative organisms. The haemolysin genes of Proteus mirabilis hybridized to H. ducreyi DNA suggesting that the haemolysin of H. ducreyi is related to the Proteus/Serratia pore-forming family of haemolysins. Tn916 mutagenesis was employed to isolate haemolysin-deficient mutants. Approximately 5000 Tn916 transposon mutants were screened for the loss of haemolytic activity and two mutants were identified. One mutant, designated 35,000-1, was further characterized. Sequences flanking the Tn916 element in strain 35,000-1 were employed to identify clones from a lambda DASHII library of H. ducreyi strain 35,000 DNA. A 13 kb insert from one lambda clone was selected for further study. This 13 kb fragment was able to both confer haemolytic activity to Escherichia coli and complement the haemolysin deficiency in strain 35,000-1. The haemolysin gene cluster was cloned from this 13 kb insert and two genes, designated hhdA and hhdB, were identified. The derived amino acid sequence of these genes demonstrated homology to the haemolysin and activation/secretion proteins of P. mirabilis and Serratia marcescens.

The PhlA Hemolysin from the Entomopathogenic Bacterium Photorhabdus luminescens Belongs to the Two-Partner Secretion Family of Hemolysins

“…In Serratia marcescens, ShlB that belongs to the TpsB protein family has been shown to be essential for secretion and activation of the ShlA hemolysin (47). HpmB and HpmA in Proteus mirabilis (52), EthB and EthA in E. tarda (29), and HhdB and HhdA of H. ducreyi (39), which all belong to the TpsB and TpsA protein families, have the same functions (31). Furthermore, putative hemolysin genes were also identified in the Y. pestis genome.…”

Channel Formation by FhaC, the Outer Membrane Protein Involved in the Secretion of the Bordetella pertussis Filamentous Hemagglutinin

“…The mechanism of FHA secretion is of particular interest, as despite its high efficiency it requires only one specific accessory protein, FhaC. It appears to be a distinct terminal branch of Sec-dependent secretion widely distributed among Gram-negative bacteria, as a growing number of FhaC homologs are being identified that are involved in the secretion of hemolysins or adhesins (5)(6)(7)(8).…”

A CdtA-CdtC Complex Can Block Killing of HeLa Cells by Haemophilus ducreyi Cytolethal Distending Toxin

“…While the mechanism(s) by which this gram-negative bacterium causes genital ulceration remains to be determined (31), it has been established that this pathogen produces at least two toxins. The first of these is a cytotoxin with hemolytic activity (20,21); killing of human cells in vitro by this hemolysin requires direct contact between the bacterium and the eukaryotic cells (2,19). The other toxin can be detected by its activity in H. ducreyi culture supernatant fluid (26) and is a member of the cytolethal distending toxin (CDT) family (4,24).…”