The effect of UVB radiation on skin microbiota in patients with atopic dermatitis and healthy controls.

OBJECTIVES: To investigate Staphylococcus aureus and Staphylococcus epidermidis quantitatively in adult patients with atopic dermatitis and in healthy controls treated with UVB radiation. STUDY DESIGN: Twenty-three adult patients (of these, 3 were excluded) with flexural atopic dermatitis and 20 healthy controls were randomly selected at the outpatient clinic of the Dermatological Department, University Hospital, North Norway. METHODS: Adult patients with atopic dermatitis (n = 20) and healthy controls (n = 20) were given 20 UVB treatments. Bacterial samples were collected before treatment, after 4 weeks of treatment, and finally after 2 weeks follow-up. RESULTS: The main bacteria found were Staphylococcus aureus and Staphylococcus epidermidis. 16 of the 20 patients with atopic dermatitis had Staphylococcus aureus in lesional skin and 12 in non-lesional skin. None of the healthy controls had Staphylococcus aureus in the sample from the flexural elbow. The Staphylococcus aureus counts decreased (not significant) in lesional skin after 4 weeks of treatment and Staphylococcus aureus counts were slightly higher after 2 weeks follow up. The same figures were also seen in non-lesional skin and forehead. CONCLUSIONS: Staphylococcus aureus is widely colonised in the skin of atopic dermatitis patients, but is rare in healthy adults. UVB treatment decreases the Staphylococcus aureus count in patients with atopic dermatitis.

No detectable Chlamydia pneumoniae and cytomegalovirus DNA in leukocytes in subjects with echolucent and echogenic carotid artery plaques.

BACKGROUND: Controversy exists whether persistent Chlamydia pneumoniae or cytomegalovirus infections cause initiation or progression of atherosclerosis. C. pneumoniae DNA in peripheral blood mononuclear cells (PBMC) has been proposed to be a more reliable marker of cardiovascular risk than are C. pneumoniae antibodies. Reported prevalences of C. pneumoniae DNA among cardiovascular patients vary greatly, indicating methodological limitations. There is an increasing concern that published results may have been biased by extensive use of less specific polymerase chain reaction (PCR) technology. METHODS: C. pneumoniae DNA and cytomegalovirus DNA were determined by probe-based real-time PCR technology in PBMCs among subjects with echolucent (n=29) or echogenic (n=28) carotid artery plaques, and in controls without carotid plaques (n=38), all recruited from a population-based study. Samples were examined in multiple repeats with PCR assays targeting two different sequences of the genome for both microorganisms. RESULTS AND CONCLUSION: IgG seropositivity was frequent in all three groups, confirming previous exposure, but C. pneumoniae DNA or cytomegalovirus DNA was not detected in a single PBMC sample by means of probe-based, highly sensitive, and specific real-time PCR assays. Our results indicate that persistent C. pneumoniae or CMV infection is not a common phenomenon in subjects with carotid atherosclerosis.

Staphylococcus aureus small colony variants are resistant to the antimicrobial peptide lactoferricin B.

OBJECTIVES: To determine whether Staphylococcus aureus small colony variants (SCVs) are resistant to the antimicrobial peptide lactoferricin B. To assess if deficiency in transmembrane potential, a common characteristic of SCVs that are haemin- or menadione-auxotrophs, affects the uptake of the peptide into the bacterial cytoplasm. METHODS: A broth microdilution technique was used for susceptibility testing to determine the MIC of lactoferricin B for SCVs with three different auxotrophisms (haemin, menadione or thymidine) and their isogenic parent strains. Both clinical isolates and genetically defined mutants were used. The internalization of lactoferricin B in a hemB mutant and the respective parent strain was studied using transmission electron microscopy and immunogold labelling. RESULTS: All SCVs showed reduced susceptibility to lactoferricin B irrespective of their auxotrophy compared with their isogenic parent strains. The MIC for all SCVs was >256 mg/L, whereas the MICs for the parent strains ranged from 16-256 mg/L. Surprisingly, the hemB mutant contained significantly more lactoferricin B intracellularly than the respective parent strain. CONCLUSIONS: The resistance mechanism of SCVs towards the antimicrobial peptide lactoferricin B is presumably caused by the metabolic changes present in SCVs rather than by a changed transmembrane potential of SCVs or reduced uptake of the peptide.

Induced resistance to the antimicrobial peptide lactoferricin B in Staphylococcus aureus.

This study was designed to investigate inducible intrinsic resistance against lactoferricin B in Staphylococcus aureus. Serial passage of seven S. aureus strains in medium with increasing concentrations of peptide resulted in an induced resistance at various levels in all strains. The induced resistance was unstable and decreased relatively rapidly during passages in peptide free medium but the minimum inhibitory concentration remained elevated after thirty passages. Cross-resistance to penicillin G and low-level cross-resistance to the antimicrobial peptides indolicidin and Ala(8,13,18)-magainin-II amide [corrected] was observed. No cross-resistance was observed to the human cathelicidin LL-37. In conclusion, this study shows that S. aureus has intrinsic resistance mechanisms against antimicrobial peptides that can be induced upon exposure, and that this may confer low-level cross-resistance to other antimicrobial peptides.

Lactoferricin B inhibits bacterial macromolecular synthesis in Escherichia coli and Bacillus subtilis.

Most antimicrobial peptides have an amphipathic, cationic structure, and an effect on the cytoplasmic membrane of susceptible bacteria has been postulated as the main mode of action. Other mechanisms have been reported, including inhibition of cellular functions by binding to DNA, RNA and proteins, and the inhibition of DNA and/or protein synthesis. Lactoferricin B (Lfcin B), a cationic peptide derived from bovine lactoferrin, exerts slow inhibitory and bactericidal activity and does not lyse susceptible bacteria, indicating a possible intracellular target. In the present study incorporation of radioactive precursors into DNA, RNA and proteins was used to demonstrate effects of Lfcin B on macromolecular synthesis in bacteria. In Escherichia coli UC 6782, Lfcin B induces an initial increase in protein and RNA synthesis and a decrease in DNA synthesis. After 10 min, the DNA-synthesis increases while protein and RNA-synthesis decreases significantly. In Bacillus subtilis, however, all synthesis of macromolecules is inhibited for at least 20 min. After 20 min RNA-synthesis increases. The results presented here show that Lfcin B at concentrations not sufficient to kill bacterial cells inhibits incorporation of radioactive precursors into macromolecules in both Gram-positive and Gram-negative bacteria.

Anti-complement effects of lactoferrin-derived peptides.

Lactoferrin is an important biological molecule with many functions such as modulation of the inflammatory response, iron metabolism and antimicrobial defense. One effect of lactoferrin is the inhibition of the classical complement pathway. This study reports that antimicrobial peptides derived from the N-terminal region from both human and bovine lactoferrin, lactoferricin H and lactoferricin B, respectively, inhibit the classical complement pathway. No inhibitory effect of these peptides was observed on the alternative complement pathway in an AP50 assay. However, lactoferricin B reduced the inhibitory properties of serum against Escherichia coli in a concentration dependent manner. These results suggest that the N-terminal region of lactoferrin is the important part in the inhibition of complement activation and that these peptides possess other important properties than their antimicrobial effect.

Proteases in Escherichia coli and Staphylococcus aureus confer reduced susceptibility to lactoferricin B.

Lactoferricin B is a cationic antimicrobial peptide derived from the N-terminal part of bovine lactoferrin. The effect of bacterial proteases on the antibacterial activity of lactoferricin B towards Escherichia coli and Staphylococcus aureus was investigated using various protease inhibitors and protease-deficient E. coli mutants. Sodium-EDTA, a metalloprotease inhibitor, was the most efficient inhibitors in both species, but combinations of sodium-EDTA with other types of protease inhibitor gave a synergic effect. The results indicate that several groups of proteases are involved in resistance to lactoferricin B in both E. coli and S. aureus. We also report that genetic inactivation of the heat shock-induced serine protease DegP increased the susceptibility to lactoferricin B in E. coli, suggesting that this protease, at least, is involved in reduced susceptibility to lactoferricin B.

Evidence for a direct antitumor mechanism of action of bovine lactoferricin.

BACKGROUND: Bovine lactoferrin (LFB) and its pepsin-generated peptide lactoferricin (LfcinB) possess antitumor activities. The mechanism underlying the antitumor activities of LfcinB in vivo has not been elucidated. In this study the antitumor activities exerted by LFB, LfcinB and murine lactoferricin (LfcinM) on murine tumor cell lines and experimental tumors were investigated. MATEIALS AND METHODS: The protein and peptides were tested against Meth A fibrosarcoma, B16F10 melanoma and C26 colon carcinoma cells in vitro and their derived tumors in vivo, exploring the mechanisms of antitumor activity by way of histological and scanning electron microscopical studies. RESULTS: LfcinB exerted significant cytotoxic activity against the three tumor cell lines in vitro and significantly reduced the size of solid Meth A tumors. Scanning electron micrographs revealed tumor cell membrane disruption and eventually cell lysis, while extensive hemorrhagic necrosis was apparent in tumor sections one day after LfcinB treatment. No species-specific antitumor effect of LfcinM was observed. CONCLUSION: Our study demonstrated that LfcinB elicits an antitumor effect mediated through a direct mechanism of action not observed with LFB or LfcinM.