The importance of skin manifestations, serology and nailfold (video)capillaroscopy in morphea and systemic sclerosis: current understanding and new insights.

Since the field around morphea and systemic sclerosis (SSc) is evolving rapidly, this review approaches conventional as well as more recent clinical developments from a dermatological point of view. Skin manifestations are critical in sub-classifying these diseases ensuring a correct prognosis for these patients. They can be discretely present, and therefore, diagnosis can be challenging sometimes, implicating a thorough dermatological examination is mandatory. Furthermore, a growing amount of dermatologists perform nailfold videocapillaroscopy (NVC), a more recent reliable non-invasive imaging technique used for in vivo assessment of the microcirculation at the nailfold. After all, specific NVC-changes are present in a majority of patients with SSc. This way, dermatologists not only take part in the diagnosis process through clinical investigation but also through the use of a modern state of the art imaging technique that is becoming the golden standard in SSc multidisciplinary workup. In this review, current understandings for NVC in morphea and SSc are revised. So far, the role of NVC in the diagnosis/prognosis/classification of morphea patients has not been thoroughly investigated to make proper conclusions. As for SSc, it is well known that NVC contributes to the diagnosis and can make a fundamental difference especially when obvious clinical SSc signs are absent. This review emphasizes the (somewhat underestimated) role of dermatologists in the process of diagnosis and follow-up, and thus, the difference we can make for our patients and fellow colleagues in the multidisciplinary workup of SSc and morphea.

Inhibition by EGTA of the formation of a biofilm by clinical strains of Staphylococcus aureus.

The effect of EGTA on the adhesion and on the formation of a biofilm by two reference and eight clinical strains of Staphylococcus aureus was studied. All the clinical strains were isolated from patients from Kinshasa. Spa typing confirmed that these clinical strains were distinct. The Biofilm Ring Test (BFRT®) showed that EGTA (100 µM-10 mM) inhibited the adhesion of the four clinical methicillin-resistant (MRSA) strains and the crystal violet staining method that it inhibited the formation of a biofilm by all the strains. Divalent cations abolished the effect of EGTA on the formation of a biofilm, specially in the clinical MRSA strains. EGTA had no effect on established biofilms. Only concentrations of EGTA higher than 10 mM were toxic to eukaryotic cells. Our results establish the effectiveness and the safety of lock solutions with EGTA to prevent the formation in vitro of biofilms by S. aureus.

Contribution of the production of quormones to some phenotypic characteristics of Pseudomonas aeruginosa clinical strains.

The contribution of quorum sensing in some phenotypic and pathogenic characteristics of Pseudomonas aeruginosa was studied. The production of acylhomoserine lactones (AHL) by planktonic cultures of eight clinical and reference strains of P. aeruginosa was evaluated using two biosensors. The adhesion of the bacteria on a surface (Biofilm Ring Test ®, BFRT), their capacity to develop a biofilm (crystal violet staining method, CVSM), their sensitivity to tobramycin and their secretion of proteases or of rhamnolipids were also measured. The production and the release of AHL widely varied among the eight strains. An analysis of the extracts by TLC showed that 3-oxo-C8-HSL, 3-oxo-C10-HSL and 3-oxo-C12-HSL were released by the five strains producing the highest amount of Cn≥6-HSL. The genes lasI and lasR involved in the synthesis and response to 3-oxo-C12-HSL were detected in the genomes of all strains. Two clinical strains had deletions in the lasR gene leading to truncation of the protein. One subpopulation of the PAO1 strain had a major deletion (98 bp) of the lasR gene. Strains with significant mutations of lasR secreted the lowest amount of AHL, probably due to deficiencies in the self-induction and amplification of the synthesis of the lactone. These strains formed a biofilm with low biomass. C4-HSL production also differed among the strains and was correlated with rhamnolipid production and biofilm formation. Whereas the production of AHL varied among P. aeruginosa strains, few correlations were observed with their phenotypic properties except with their ability to form a biofilm.

Identification of peptides derived from the human antimicrobial peptide LL-37 active against biofilms formed by Pseudomonas aeruginosa using a library of truncated fragments.

Persistent Pseudomonas aeruginosa infections are a major cause of morbidity and mortality in cystic fibrosis (CF) patients and are linked to the formation of a biofilm. The development of new biofilm inhibition strategies is thus a major challenge. LL-37 is the only human antimicrobial peptide derived from cathelicidin. The effects on the P. aeruginosa PAO1 strain of synthetic truncated fragments of this peptide were compared with the effects of the original peptide. Fragments of LL-37 composed of 19 residues (LL-19, LL13-31, and LL7-25) inhibited biofilm formation. The strongest antibiofilm activity was observed with the peptides LL7-37 and LL-31, which decreased the percentage of biomass formation at a very low concentration. Some peptides were also active on the bacteria within an established biofilm. LL7-31, LL-31, and LL7-37 increased the uptake of propidium iodide (PI) by sessile bacteria. The peptide LL7-37 decreased the height of the biofilm and partly disrupted it. The peptides active within the biofilm had an infrared spectrum compatible with an α-helix. LL-37, but not the peptides LL7-31 and LL7-37, showed cellular toxicity by permeabilizing the eukaryotic plasma membrane (uptake of ethidium bromide and release of lactate dehydrogenase [LDH]). None of the tested peptides affected mitochondrial activity in eukaryotic cells. In conclusion, a 25-amino-acid peptide (LL7-31) displayed both strong antimicrobial and antibiofilm activities. The peptide was even active on cells within a preformed biofilm and had reduced toxicity toward eukaryotic cells. Our results also suggest the contribution of secondary structures (α-helix) to the activity of the peptides on biofilms.

Effect of pluronic acid F-127 on the toxicity towards eukaryotic cells of CSA-13, a cationic steroid analogue of antimicrobial peptides.

AIMS: CSA-13 is an antimicrobial cationic steroid with some toxicity against eukaryotic cells. The purpose of this work was to test whether pluronic acid F-127 could interfere with the toxicity of CSA-13 on human umbilical vein endothelial (HUVEC) without modifying its bactericidal activity against Pseudomonas aeruginosa. METHODS AND RESULTS: The addition of pluronic acid F-127 slightly decreased the number of dead cells after exposure to CSA-13. Pluronic acid F-127 blocked the permeabilizing effect of CSA-13 on the plasma membrane of HUVEC (uptake of ethidium bromide, release of lactate dehydrogenase) without modifying its toxic effect on their mitochondrial function (MTT test, uptake of tetramethyl rhodamine ethyl ester). CONCLUSION: Pluronic acid F-127 decreased the toxicity of CSA-13 against eukaryotic cells without completely protecting them from mitochondrial damage at high concentrations of the drug. SIGNIFICANCE AND IMPACT OF THE STUDY: This work establishes that studies on the toxic effects of synthetic antimicrobials on eukaryotic cells should not only focus on the permeability of the plasma membrane but also on the integrity of the mitochondria.

Effect of a low concentration of a cationic steroid antibiotic (CSA-13) on the formation of a biofilm by Pseudomonas aeruginosa.

AIMS: Cationic steroids like CSA-13 have been designed by analogy with antimicrobial cationic peptides and have bactericidal properties. The purpose of this work was to evaluate the effect of a low concentration (1 mg l(-1)) of CSA-13 on the formation of a biofilm by eight strains of Pseudomonas aeruginosa (four mucoid and four nonmucoid strains) on an inert surface. METHOD AND RESULTS: The biofilm formation was measured with the Crystal Violet method. CSA-13 inhibited the formation of a biofilm by three strains. The zeta potential varied among the strains. The inhibition by the cationic steroid analogue affected the populations of bacteria with the lowest zeta potential. P. aeruginosa bound a fluorescent, more hydrophobic analogue of CSA-13 but there was no correlation between this binding and the inhibition by CSA-13 of biofilm formation. The interaction of CSA-13 with bacteria did not modify their ability to produce rhamnolipids. CONCLUSIONS: A low concentration of CSA-13 inhibits the formation of a biofilm by P. aeruginosa through electrostatic interactions and without affecting the production of rhamnolipids. SIGNIFICANCE AND IMPACT OF THE STUDY: A low, nontoxic concentration of CSA-13 might be beneficial for the prevention of biofilm formation.

Study of the initial phase of biofilm formation using a biofomic approach.

AIMS: The purpose of this work was to study the initial steps of formation of a biofilm using the BioFilm Ring Test and the Crystal violet staining technique. METHODS AND RESULTS: Eight strains of Pseudomonas aeruginosa were studied. The two methods revealed that four strains formed a rapid biofilm. The biofilm formed by these strains was detected after only 45 min with the BioFilm Ring Test and after 6h with the Crystal violet method. The enumeration of bacteria of the PA01 strain confirmed that, after 30 min, a significant amount of bacteria had attached on the bottom of the culture wells. After 48 h the Crystal violet method detected a biofilm with all strains. The four strains which rapidly formed a biofilm did not differ from the slow-forming strains by their mucoid character or their swarming motility or their synthesis of rhamnose. They showed higher swimming mobility. CONCLUSIONS: Our results show that the BioFilm Ring Test is a method specially suited for the study of the initial phase of the formation of a biofilm. SIGNIFICANCE AND IMPACT OF STUDY: The BioFilm Ring Test is an easy and rapid alternative to the Crystal violet staining and the enumeration methods.

Klinefelter's syndrome and lupus erythematosus: report of a case.

We report a case of SLE related polyarthritis in a 28 year-old man with Klinefelter's syndrome treated with testosterone for 13 years. Clinical outcome was favorable with low doses of corticosteroids. The pathogenesis of that strange association is discussed.