Direct and indirect bacterial killing functions of neutrophil defensins in lung explants

Studies of the antimicrobial activity of neutrophil defensins have mostly been carried out in microbiological media, and their effects on the host defense in physiological conditions are unclear. We examined 1) the antibacterial activity of defensins in physiological media with and without lung tissue present, 2) the effect of defensins on hydrogen peroxide (H(2)O(2)) production by lung tissue that had been exposed to bacteria, and 3) the effect of diphenyleneiodonium (DPI), an inhibitor of reactive oxygen species formation, on the antibacterial activity of defensins in the presence of lung tissue. Defensins were incubated with Escherichia coli or Pseudomonas aeruginosa in the absence or presence of primary cultured mouse lung explants. Defensins reduced bacterial counts by approximately 65-fold and approximately 25-fold, respectively, at 48 h; bacterial counts were further decreased by approximately 600-fold and approximately 12,000-fold, respectively, in the presence of lung tissue. Defensins induced H(2)O(2) production by lung tissue, and the rate of killing of E. coli by defensins was reduced by approximately 2,500-fold in the presence of 10 microM DPI. We conclude that defensins exert a significant antimicrobial effect under physiological conditions and that this effect is enhanced in the presence of lung tissue by a mechanism that involves the production of reactive oxygen species.     

Induction of tumour necrosis factor by staphylococcal toxic shock toxin 1

Abstract Staphylococcal toxic shock syndrome toxin 1 (TSST1) induced the release of tumour necrosis factor (TNF) from human and rabbit monocytes in vitro. Nanogram amounts of TSST1 were sufficient to induce TNF release. There was considerable variation in response between cells from different rabbits and different donors. Rabbit monocytes were slightly more sensitive to TSST1 than were human monocytes. Release of TNF in vivo could explain many of the symptoms of toxic shock syndrome.     

Cloning and nucleotide sequence of the eae gene homologue from enterohemorrhagic Escherichia coli serotype O157:H7

The eae gene has recently been shown to be necessary for the attaching and effacing (AE) activity of enteropathogenic Escherichia coli (EPEC) on intestinal epithelial cells. In this paper we report the cloning and nucleotide sequence of a similar gene from a strain of enterohemorrhagic E. coli (EHEC) serotype O157:H7. An EHEC eae sequence was identified which was 97% homologous to the EPEC eae gene for the first 2200 bp and 59% homologous over the last 800 bp. Both eae sequences show 50% homology to the central region of the Yersinia pseudotuberculosis inv gene. The receptor-binding domain of the inv gene product lies near the carboxyl terminus. This suggests that the predicted amino acid sequence divergence in the carboxyl termini of the eae gene products might result in different antigenic and receptor specificity of these putative adhesins.     

The two-component system sivS/R regulates virulence in Streptococcus iniae

Streptococcus iniae causes invasive disease and death in fish, and to a lesser extent, sporadic cases of soft-tissue infections in humans. A two-component system termed sivS/R, which regulates capsule expression, was previously identified and characterized. In this study, it is shown that a sivS/R deletion-insertion mutant, termed 9117Deltasiv, causes transient bacteremia and reduced virulence compared with the parent strain when tested in a murine model of bacteremic infection. Furthermore, real-time PCR studies indicated that SivS/R regulates the expression levels of the streptolysin S structural gene, sagA, as well as the CAMP factor gene, cfi. Sodium dodecyl sulphate polyacrylamide gel electrophoresis of S. iniae spheroplasts revealed downregulation of three surface proteins in the mutant strain compared with the parent strain. These proteins were identified by MS to be a putative lipoprotein, a hyaluronate-associated protein and a pyruvate kinase. This study demonstrates that SivS/R regulates virulence in vivo, and controls the expression of a number of genes in S. iniae.     

Induction of tumour necrosis factor by staphylococcal toxic shock toxin 1

Staphylococcal toxic shock syndrome toxin 1 (TSST1) induced the release of tumour necrosis factor (TNF) from human and rabbit monocytes in vitro. Nanogram amounts of TSST1 were sufficient to induce TNF release. There was considerable variation in response between cells from different rabbits and different donors. Rabbit monocytes were slightly more sensitive to TSST1 than were human monocytes. Release of TNF in vivo could explain many of the symptoms of toxic shock syndrome.     

A toxic shock syndrome toxin mutant of Staphylococcus aureus isolated by allelic replacement lacks virulence in a rabbit uterine model

The gene coding for toxic shock syndrome toxin-1 in S. aureus was inactivated by allelic replacement in two TSS-associated strains. One mutant derived from FRI1169 (a non-enterotoxigenic strain) lacked virulence in the rabbit uterine chamber infection model. This suggests that TSST-1 is the only determinant produced by this strain that can induce the symptoms of shock in rabbits. A novel method for allelic replacement involving transduction of plasmid integrants is described.