Search

The aim of this study was to estimate the durability of tetanus toxoid specific seroprotection in a cohort of people with HIV (PWH).A cross-sectional study.PWH with a last date of tetanus toxoid booster available were identified. Tetanus toxoid specific IgG were detected using commercial ELISA kit. Durability of seroprotection was estimated using a linear regression model and analyzed according to the country of birth. The impact of baseline parameters at the time of vaccination (CD4 + T cell count, viral load, and antiretroviral therapy) was also assessed.One hundred three individuals were included. The median duration between last tetanus toxoid booster and sampling was 5.6years (IQR 2.6-8.9). Using a linear regression model, half-life of tetanus toxoid specific antibody was estimated at 9.9 years [95% confidence interval (95% CI: 5.5-50)] in the whole cohort. Half-life was reduced in individuals born outside Europe: 4.4 years (95% CI: 2.9-8.5). PWH born outside Europe had lower CD4 + T cell count at the time of immunization and more frequently a CD4 + T cell count nadir less than 200 cells/μl before vaccination.PWH born outside Europe have lower half-life of tetanus toxoid specific antibody as compared to previous study performed in the general population. Possible causes include lower nadir or current CD4 + T cell count or under-immunization status in country of origin before migration. Longer interval of booster vaccination, as recommended in the general population, might not be appropriate in this subgroup of PWH. Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.

Highlights • Early cytokine measurement after hospitalization predicts disease progression • A score combining the results of IL-6, IL-8 and IL-10 allows to predict with a very high performance: The severity of the disease (IL-6 * IL-8 * IL-10 score : threshold value of 2068 pg/mL) The need for intensive care (IL-6 * IL-10 score: cut-off value of 178 pg/mL), Objectives We aimed to explore cytokine profile in patients related to Coronavirus Disease 2019 (COVID-19) severity and to establish a predictive cytokine score to discriminate severe from non-severe cases and provide a prognosis parameter for patients that will require intensive care unit (ICU) transfer. Methods Serum samples of 63 patients diagnosed with SARS-CoV-2 infection were collected early after hospital admission (day 0-3). Patients were categorized in five groups based on the clinical presentation, the PaO2/FiO2 ratio and the requirement of mechanical ventilation. Results Three cytokines, IL-6, IL-8 and IL-10 were markedly higher in severe forms (n = 44) than in non-severe forms (n = 19) (p

Early evidence during the coronavirus disease 2019 (COVID-19) pandemic indicated high levels of interleukin (IL)-6 in patients with severe COVID-19. This led to the off-label use of tocilizumab (TCZ) during the first wave of the pandemic. While the monoclonal antibody blocks IL-6 pathway, its effect on other inflammatory cytokines remains poorly described. To better understand the effect of TCZ on the biological inflammatory profile, we monitored a large panel of inflammatory cytokines in critically ill COVID-19 patients receiving off-label TCZ. Twenty-three patients with polymerase chain reaction-confirmed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection were included in the study, among which 15 patients received TCZ and 8 patients did not. Serum samples were collected for 8 days, before and following TCZ administration or hospital admission for the control group. Serum profile of 12 cytokines (IL-1

Infection with the novel coronavirus SARS-CoV-2 triggers severe illness with high mortality in a subgroup of patients. Such a critical course of COVID-19 is thought to be associated with the development of cytokine storm, a condition seen in macrophage activation syndrome (MAS) and secondary hemophagocytic lymphohistiocytosis (HLH). However, specific data demonstrating a clear association of cytokine storm with severe COVID-19 are still lacking. The aim of this study was to directly address whether immune activation in COVID-19 does indeed mimic the conditions found in these classic cytokine storm syndromes.Levels of 22 biomarkers were quantified in serum samples from patients with COVID-19 (n = 30 patients, n = 83 longitudinal samples in total), patients with secondary HLH/MAS (n = 50), and healthy controls (n = 9). Measurements were performed using bead array assays and single-marker enzyme-linked immunosorbent assay. Serum biomarker levels were assessed for correlations with disease outcome.In patients with secondary HLH/MAS, we observed pronounced activation of the interleukin-18 (IL-18)-interferon-γ axis, increased serum levels of IL-1 receptor antagonist, intercellular adhesion molecule 1, and IL-8, and strongly reduced levels of soluble Fas ligand in the course of SARS-CoV-2 infection. These observations appeared to discriminate immune dysregulation in critical COVID-19 from the well-recognized characteristics of other cytokine storm syndromes.Serum biomarker profiles clearly separate COVID-19 from MAS or secondary HLH in terms of distinguishing the severe systemic hyperinflammation that occurs following SARS-CoV-2 infection. These findings could be useful in determining the efficacy of drugs targeting key molecules and pathways specifically associated with systemic cytokine storm conditions in the treatment of COVID-19.

Objectives Tetanus is a vaccine-preventable disease. Booster immunization is required in order to induce long-lived tetanus-toxoid (TT) specific antibody response. We investigated the prevalence and risk factors of TT seronegativity in a cohort of people living with HIV (PWH) in Belgium along with the respective performance of vaccine history and a rapid dipstick test (Tetanus Quick Stick ® or TQS) compared to ELISA testing. Methods PWH were prospectively enrolled and answered a questionnaire. ELISA was performed on serum or plasma using a commercial kit. A TT antibody level ≥ 0.15 IU / mL was considered protective. The TQS test was performed on a limited number of subjects. Results Three-hundred forty-four subjects were included. The prevalence of tetanus seroprotection was 84,9%. Median age was 46.7 and 68% were born outside Belgium. Antiretroviral therapy coverage was almost universal (98.5%). After multivariable analysis, two risk factors were independently associated with TT seronegativity: an education level equivalent or below than secondary school and being born outside Europe. Vaccine history was shown to be unreliable (sensitivity: 43.8%; specificity: 76.5%; positive predictive value: 91.4% and negative predictive value :19.3%). The correlation between vaccine history and tetanus seroprotection was low (kappa coefficient = 0.09). The TQS performances were good (sensitivity 86.4%, specificity 96.0%, positive predictive value 99.3%, negative predictive value 52.17%). The correlation between TQS and tetanus seroprotection was substantial (kappa coefficient = 0.61). Conclusions In this cohort of PWH with a high proportion of migrants, socio-demographic and educational factors were associated with TT seronegativity while HIV-related factors were not, indicating that vaccine information should be tailored to cultural and educational background. As vaccine history is not reliable, TQS could represent an efficient tool for screening of TT-seronegativity.

Early evidence during the COVID-19 pandemic indicated high levels of IL-6 in patients with severe COVID-19. This led to the off-label use of tocilizumab (TCZ) during the first wave of the pandemic.We aimed to monitor IL-6 and several inflammatory cytokines in critically ill COVID-19 patients receiving off-label TCZ. Fifteen critically ill SARS-CoV-2 PCR confirmed cases were enrolled and serum samples were collected during 8 days, before and following administration of a single dose of TCZ. In parallel, a control group consisting of 8 non-treated COVID-19 patients not receiving TCZ was established. Serum profile of 12 cytokines (IL-1β, -2, -4, -6, -8, -10, -12, -13, -17, -18, TNF-α and INF-γ) and of IL-6R were assessed in these two groups. Although the increased IL-6 concentrations after TCZ infusion were expected, we observed an unexpected increase in IL-1β, -2, -4, -10, -12p70, -18 and IL-6R levels in the treated patients with maximal values reached 2 to 4 days after TCZ. In contrast, no change in cytokine levels was observed in the control group. There was no significant difference in cytokine levels between survivors (TCZ/S) or non-survivors (TCZ/D). This observation suggests that some inflammatory pathways escape IL-6R blockade leading to an increase in several pro-inflammatory cytokines. Our findings could highlight an anti-inflammatory role of IL-6 and may explain why TCZ has failed to improve survival in critically ill COVID-19 patients when given alone.

Introduction Coronavirus disease 2019 (COVID-19) appeared in China in December 2019 and has spread around the world. High Interleukin-6 (IL-6) levels in COVID-19 patients suggest that a cytokine storm may play a major role in the pathophysiology and are considered as a relevant parameter in predicting most severe course of disease. The aim of this study was to assess repeated IL-6 levels in critically ill COVID-19 patients admitted to our Intensive Care Unit (ICU) and to evaluate their relationship with patient’s severity and outcome. Methods We conducted a retrospective study on patients admitted to the ICU with a diagnosis of COVID-19 between March 10 (i.e. the date of the first admitted patients) and April 30, 2020. Demographic, clinical and laboratory data were collected at admission. On the day of IL-6 blood concentration measurement, we also collected results of D-Dimers, C-Reactive Protein, white blood cells and lymphocytes count, lactate dehydrogenase (LDH) and ferritin as well as microbiological samples, whenever present. Results Of a total of 65 patients with COVID-19 admitted to our ICU we included 41 patients with repeated measure of IL-6. There was a significant difference in IL-6 levels between survivors and non-survivors over time (p = 0.001); moreover, non survivors had a significantly higher IL-6 maximal value when compared to survivors (720 [349–2116] vs. 336 [195–646] pg/mL, p = 0.01). The IL-6 maximal value had a significant predictive value of ICU mortality (AUROC 0.73 [95% CI 0.57–0.89]; p = 0.01). Conclusions Repeated measurements of IL-6 can help clinicians in identifying critically ill COVID-19 patients with the highest risk of poor prognosis., info:eu-repo/semantics/published

Many biomarkers have been proposed for the diagnosis of secondary hemophagocytic lymphohistiocytosis (HLH) in adults, but comparative studies are lacking. We analyzed ferritin, glycosylated ferritin, soluble CD25, CD163 and CD14, IL-6, IFN-γ, IL-18, IL-10, IL-1s, IL-12p70, IL-17α, IP-10, and CXCL9 levels to differentiate HLH from sepsis in critically ill patients. Of 120 patients, HLH was confirmed for 14 patients. Among the biomarkers tested, ferritin, IL-18, and glycosylated ferritin were the most efficient parameters for early diagnosis of HLH. With a sensitivity set at 85%, ferritin, IL-18, and glycosylated ferritin were the biomarkers with the highest specificity: 84, 79, and 71% respectively. Combining IL-18 with the HScore provided a new score with an increased specificity compared to the HScore alone, 86% compared to 70% with a sensitivity set at 100%. A distinct cytokine pattern was highlighted in patients with malignancy-triggered HLH, with highly increased levels of INF-ɣ and CXCL9, compared to HLH secondary to infection. This is the largest study available to date, comparing diagnostic biomarkers for HLH on a cohort of critically ill adult patients. Serum ferritin was the most discriminating parameter for early diagnosis of secondary HLH. IL18*HScore was identified as a highly potential score.

INTRODUCTION The discrimination of leukemia lymphoblasts (LB) in diagnosis and follow-up of B-cell progenitor acute lymphoblastic leukemia (BCP-ALL) by multiparameter flow cytometry (MFC) may be difficult due to the presence of hematogones (HG). The aim of this study was to compare lymphoblasts of BCP-ALL and HG for the expression of the most discriminating antigens. METHODS A total of 82 bone marrow samples (39 BCP-ALL and 43 patients with HG) were analyzed using MFC. Mean fluorescence intensity (MFI) was measured for ten markers commonly used in hematology laboratories: CD45, CD19, CD10, CD34, CD38, CD20, CD22, CD58, CD81, and CD123. Statistical comparison of the MFI between LB and HG was performed. The presence on LB of aberrant expression of myeloid and/or T-cell markers was also investigated. RESULTS Qualitative pattern expression of antigens showed overexpression on LB of CD58, CD22, CD34, CD10 and underexpression of CD81, CD45, CD38 when compared to HG. Expression of CD123 was positive in 34% of BCP-ALL LB and always absent on HG. Aberrant antigen expression (myeloid and/or T-cell marker) including CD123 was observed in 58% of BCP-ALL patients. The use of a MFI antigen ratio of the most discriminating markers (CD81/CD58) (analysis of variance, P

Von Willebrand disease (MW) is the most common constitutional bleeding disorders. It is caused by a quantitative or qualitative abnormality of the von Willebrand factor (VWF). The laboratory assessment of the disease combines a FVIII assay, and a determination of the antigen and activity of VWF. The analytical validation of VWF:Ag, VWF:Act, vWF:CB is reported in this work and demonstrates good test performance of all three assays, with a coefficient of variation lower than 10% for both the repeatability and reproducibility, stability with a deviation of less than 5% from the target value after six hours at room temperature. The dosages are linear through the following ranges: 2% to 346% for VWF:Ag, 3% to 170% for VWF:Act, and 0.07% to 259% for VWF:CB. The usual values determined on 32 control subjects are in the range of reference values published in the literature. However as the number of control samples tested is small, we will adopt the reference values of the literature of 50 to 200% in routine. The other functional test VWF:CB will be used in our daily practice to differentiate the type of 2M 2A type. However, given that the type 2M is extremely rare, we think it is rather aimed at specialized laboratories in which a large number of patients referred.

BACKGROUND Preanalytical issues are a major part of routine coagulation laboratory errors. Automation in detection of preanalytical problems, including hemolysis, icterus and lipemia (HIL), improper tube fill volume, and undue clotting, has recently been implemented on specific hemostasis instruments. The aim of this study was to assess the added value of a new preanalytical module integrated into hemostasis analyzers compared to visual inspection of samples. METHODS The detection of preanalytical issues was performed by visual or manual inspection of the samples and by the new preanalytical module integrated on the ACL TOP 550. Additional tests were done to evaluate the interference of hemoglobin (Hb), bilirubin or triglycerides (TG). Plasma pools containing the interference substance were tested for routine coagulation assays on the STA-R Evolution, CS-5100, and ACL TOP 550. RESULTS Visual or manual inspection detected statistically less samples with preanalytical issues than the new preanalytical module integrated on the ACL TOP 550 (3.5% vs. 6.6%, p < 0.001). The majority of the samples were rejected for poor filling. HIL interferences appeared on assays when the concentration of Hb, bilirubin or TG exceeded a certain threshold that was analyzer and reagent dependent. CONCLUSIONS Automatic and standardized check of routine coagulation samples by ACL TOP 550 increased the accuracy and consistency in detection of preanalytical issues as compared to visual inspection only. The main advantages were the detection of insufficient filled tubes and icteric samples that are not detected visually.

Three chromogenic media, chromID MRSA SMART (SMART), chromID MRSA first generation (chromID), and Brilliance MRSA (OX2), were evaluated for methicillin-resistant Staphylococcus aureus (MRSA) screening using 1,220 samples. The sensitivity at 24 h was significantly better with the SMART agar (66.4%) than that with chromID agar (50.5%). Enrichment and incubation until 48 h are still needed for an optimal yield.

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 (100μM-10mM) 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 10mM 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. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The formation of a Pseudomonas aeruginosa biofilm, a complex structure enclosing bacterial cells in an extracellular polymeric matrix, is responsible for persistent infections in cystic fibrosis patients leading to a high rate of morbidity and mortality. The protective environment created by the tridimensional structure reduces the susceptibility of the bacteria to conventional antibiotherapy. Cationic steroid antibiotics (CSA)-13, a nonpeptide mimic of antimicrobial peptides with antibacterial activity on planktonic cultures, was evaluated for its ability to interact with sessile cells. Using confocal laser scanning microscopy, we demonstrated that the drug damaged bacteria within an established biofilm showing that penetration did not limit the activity of this antimicrobial agent against a biofilm. When biofilms were grown during exposure to shear forces and to a continuous medium flow allowing the development of robust structures with a complex architecture, CSA-13 reached the bacteria entrapped in the biofilm within 30 min. The permeabilizing effect of CSA-13 could be associated with the death of the bacteria. In static conditions, the compound did not perturb the architecture of the biofilm. This study confirms the potential of CSA-13 as a new strategy to combat persistent infections involving biofilms formed by P. aeruginosa. © 2013 The Authors., SCOPUS: ar.j, info:eu-repo/semantics/published

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.

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.

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.

Cationic antimicrobial peptides are major components of innate immunity and help control the initial steps of the infectious process. They are expressed not only by immunocytes, but also by epithelial cells. They share an amphipathic secondary structure with a polar cationic site, which explains their tropism for prokaryote membranes and their hydrophobic site contributing to the destructuration of these membranes. LL-37 is the only cationic antimicrobial peptide derived from human cathelicidin. LL-37 can also cross the plasma membrane of eukaryotic cells, probably through special domains of this membrane called lipid rafts. This transfer could be beneficial in the context of vaccination: the activation of intracellular toll-like receptors by a complex formed between CpG oligonucleotides and LL-37 could conceivably play a major role in the building of a cellular immunity involving NK cells., SCOPUS: re.j, info:eu-repo/semantics/published

article i nfo Article history: 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 6 h 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.

The bactericidal activity of a cholic acid antimicrobial derivative, CSA-13, was tested against eight strains of Pseudomonas aeruginosa (both reference and clinical strains) and compared with the response to tobramycin. In planktonic cultures, the minimal inhibitory and minimal bactericidal concentrations of CSA-13 and tobramycin were in the 1-25 mg/L range except for one mucoid clinical strain which was much less sensitive to tobramycin (minimal bactericidal concentration, 65-125 mg/L). In young (24 h) biofilms, the sensitivity to CSA-13 was reduced (half-maximal concentration CSA-13 averaged 88 mg/L) and varied among the eight strains. The sensitivity to tobramycin was also very variable among the strains and some were fully resistant to the aminoglycoside. The combination of tobramycin with CSA-13 was synergistic in five strains. Only one strain showed antagonism between the two drugs at low concentrations of CSA-13. One reference and five clinical strains were tested in mature (12 days) biofilms. The effect of CSA-13 was delayed, some strains requiring 9 days exposure to the drug to observe a bactericidal effect. All the strains were tolerant to tobramycin but the addition of CSA-13 with tobramycin was synergistic in three strains. CSA-13 permeabilized the outer membrane of the bacteria (half-maximal concentration, 4.4 mg/L). At concentrations higher than 20 mg/L, it also permeabilized the plasma membrane of human umbilical vein endothelial cells. In conclusion, CSA-13 has bactericidal activity against P. aeruginosa even in mature biofilms and cationic steroid antibiotics can thus be considered as potential candidates for the treatment of chronic pulmonary infections of patients with cystic fibrosis. Considering its interaction with the plasma membrane of eukaryotic cells, less toxic derivatives of CSA-13 should be developed.

Peritoneal macrophages were isolated from wild type (WT) mice and from mice invalidated for the P2X(7) receptor (KO) which had been pretreated with thioglycolate. In cells from WT mice, 1 mM ATP increased the intracellular concentration of calcium ([Ca(2+)](i)), the uptake of ethidium bromide, the production of reactive oxygen species (ROS), the secretion of IL-1beta, the release of oleic acid and of lactate dehydrogenase; it decreased the intracellular concentration of potassium ([K(+)](i)). In KO mice, ATP transiently increased the [Ca(2+)](i) confirming that the P2X(7) receptor is a major receptor of peritoneal macrophages. WKYMVm, an agonist of receptors for formylated peptides (FPR) also increased the [Ca(2+)](i) in murine macrophages. The slight increase of the [Ca(2+)](i) was strongly potentiated by ivermectin confirming the expression of functional P2X(4) receptors by murine peritoneal macrophages. CRAMP, the unique antimicrobial peptide derived from cathelin in mouse inhibited all the responses coupled to P2X(7) receptors in macrophages from WT mice. Agonists for FPR had no effect on the increase of the [Ca(2+)](i) in response to ATP. CRAMP had no effect on the increase of the [Ca(2+)](i) evoked by a combination of ATP and ivermectin in macrophages from P2X(7)-KO mice. In summary CRAMP inhibits the responses secondary to the activation of the murine P2X(7) receptors expressed by peritoneal macrophages. This inhibition is not mediated by FPR receptors and is specific since CRAMP has no effect on the response coupled to P2X(4) receptors. It can thus be concluded that the interaction between P2X(7) receptors and cathelin-derived antimicrobial peptides is species-specific, in some cases (man) positive in others (mouse) negative.

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 C≥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.

Respiratory disease is the main cause of morbidity and mortality in patients with cystic fibrosis (CF). In particular, patients suffer from chronic infection due to biofilm formation by opportunistic Pseudomonas aeruginosa (32). Therefore, there is an urgent need to develop alternative ways to treat biofilm-associated clinical infections. The aim of this study was to compare the antimicrobial effects in vitro of the combinations tobramycin-clarithromycin and tobramycin-azithromycin against five P. aeruginosa biofilms and to establish the most effective combination. We performed a kinetic study over a period of 28 days of a twice-daily coadministration of the combinations tobramycin-clarithromycin and tobramycin-azithromycin on 12-day-old, mature P. aeruginosa biofilms formed on microplate pegs for 4 clinical isolates and one laboratory strain (PAO1) to simulate the treatment of CF patients with tobramycin inhalation solution (TOBI) through aerosolization. A synergy between tobramycin and clarithromycin was recorded for 3/5 biofilms, with a bacterial decrease of more than 5 log. Conversely, we found an antagonistic activity when 4 μg/ml tobramycin was administered with azithromycin at 2 μg/ml for P. aeruginosa PAO1 and with azithromycin at 2, 20, 50, 100, and 200 μg/ml for P. aeruginosa PYO1. Treatment with tobramycin at 4 μg/ml combined with clarithromycin at 200 μg/ml eradicated all five biofilms, while tobramycin-azithromycin at the same concentrations eradicated only three biofilms. Results of this study suggest that local administration of tobramycin and clarithromycin into the respiratory tract represents a better strategy than the combination tobramycin-azithromycin for the treatment of P. aeruginosa -associated pulmonary infections.

Background: Staphylococcus epidermidis is a pathogen that is frequently encountered in the hospital environment. Healthcare workers (HCWs) can serve as a reservoir for the transmission of S. epidermidis to patients.Methods: The aim of this study was to compare and identify differences between S. epidermidis isolated from 20 patients with catheter-related bloodstream infections (CRBSIs) and from the hands of 42 HCWs in the same hospital in terms of antimicrobial resistance, biofilm production, presence of the intercellular adhesion (ica) operon and genetic diversity (pulsed field gel electrophoresis (PFGE), multilocus sequence typing (MLST) and staphylococcal cassette chromosome (SCC) mec typing).Results: S. epidermidis isolates that caused CRBSI were resistant to significantly more non-betalactam drugs than were isolates collected from HCWs. Among the 43 mecA positive isolates (26 from HCWs), the most frequent SCCmec type was type IV (44%). The ica operon was significantly more prevalent in CRBSI isolates than in HCWs (P < 0.05). Weak in vitro biofilm production seemed to correlate with the absence of the ica operon regardless of the commensal or pathogenic origin of the isolate. The 62 isolates showed high diversity in their PFGE patterns divided into 37 different types: 19 harbored only by the CRBSI isolates and 6 shared by the clinical and HCW isolates. MLST revealed a total of ten different sequence types (ST). ST2 was limited to CRBSI-specific PFGE types while the " mixed" PFGE types were ST5, ST16, ST88 and ST153.Conclusion: One third of CRBSI episodes were due to isolates belonging to PFGE types that were also found on the hands of HCWs, suggesting that HCW serve as a reservoir for oxacillin resistance and transmission to patients. However, S. epidermidis ST2, mecA-positive and icaA-positive isolates, which caused the majority of clinically severe CRBSI, were not recovered from the HCW's hands. © 2014 Cherifi et al. licensee BioMed Central Ltd., SCOPUS: ar.j, info:eu-repo/semantics/published