Pseudomonas Aeruginosa Colonization Research Articles

Elexacaftor – Tezacaftor – Ivacaftor treatment improves systemic infection parameters and Pseudomonas aeruginosa colonization rate in patients with cystic fibrosis a monocentric observational study

Background & aimsThe CFTR-modulating therapy Elexaftor – Tezacaftor - Ivacaftor (ETI) has been widely prescribed since its approval in 2020 in the European Union. The aim of this study was to methodically evaluate the effects of an ETI treatment on clinical, biochemical data and Pseudomonas colonization in order to demonstrate its efficacy. MethodsThis prospective monocentric study comprised 69 patients diagnosed with cystic fibrosis aged at least 12 years and treated with ETI between September 2020 and November 2021. Clinical and laboratory data of each patient and study visit were collected before and after 24 weeks of ETI treatment. Follow-up status of Pseudomonas aeruginosa (PsA) colonization was assessed after one year of therapy by regularly determined sputum or throat swab samples. ResultsMarked improvements biochemical markers of systemic inflammation as white blood cell count, levels of immunoglobulins A, G and M and albumin within 24 weeks of therapy were observed. ETI treatment proved to be effective as seen by amelioration of lung function and sweat chloride concentration. Assessment of PsA colonization status revealed a conversion from a positive to negative detection in 36% of the cases after one year of therapy. ConclusionsETI treatment effectively improves systemic inflammation parameters and shows promising results in PsA status conversion.

Gradual increase in sweat chloride concentration is associated with a higher risk of CRMS/CFSPID to CF reclassification.

Universal implementation of cystic fibrosis (CF) newborn screening (NBS) has led to the diagnostic dilemma of infants with CF screen-positive, inconclusive diagnosis (CFSPID), with limited guidance regarding prognosis and standardized care. Rates of reclassification from CFSPID to CF vary and risk factors for reclassification are not well established. We investigated whether clinical characteristics are associated with the risk of reclassification from CFSPID to a CF diagnosis. Children with a positive CF NBS were recruited from two sites in California. Retrospective, longitudinal, and cross-sectional data were collected. A subset of subjects had nasal epithelial cells collected for CF transmembrane conductance regulator(CFTR) functional assessment. Multivariate logistic regression was used to assess the risk of reclassification. A total of 112 children completed the study (CF = 53, CFSPID = 59). Phenotypic characteristics between groups showed differences in pancreatic insufficiency prevalence, immunoreactive trypsinogen (IRT) levels, and Pseudomonas aeruginosa (PSA) colonization. Spirometry measures were not different between groups. Nasal epithelial cells from 10 subjects showed 7%-30% of wild-type (WT)-CFTR (wtCFTR) function in those who reclassified and 27%-67% of wtCFTR function in those who retained the CFSPID designation. Modeling revealed that increasing sweat chloride concentration (sw[Cl- ]) and PSA colonization were independent risk factors for reclassification to CF. Increasing sw[Cl- ] and a history of PSA colonization are associated with the risk of reclassification from CFSPID to CF in a population with high IRT and two CFTR variants. A close follow-up to monitor phenotypic changes remains critical in this population. The role of CFTR functional assays in this population requires further exploration.

The Pseudomonas aeruginosa Secreted Protein PA3611 Promotes Bronchial Epithelial Cell Epithelial-Mesenchymal Transition via Integrin αvβ6-Mediated TGF-β1-Induced p38/NF-κB Pathway Activation.

Pseudomonas aeruginosa (PA) is an important pathogen that has been proven to colonize and cause infection in the respiratory tract of patients with structural lung diseases and to lead to bronchial fibrosis. The development of pulmonary fibrosis is a complication of PA colonization of the airway, resulting from repeated infection, damage and repair of the epithelium. Bronchial epithelial cell epithelial-mesenchymal transition (EMT) plays a vital role in bronchial fibrosis. To date, research on bronchial epithelial cell EMT caused by PA-secreted virulence factors has not been reported. Here, we found that PA3611 protein stimulation induced bronchial epithelial cell EMT with mesenchymal cell marker upregulation and epithelial cell marker downregulation. Moreover, integrin αvβ6 expression and TGF-β1 secretion were markedly increased, and p38 MAPK phosphorylation and NF-κB p65 subunit phosphorylation were markedly enhanced. Further research revealed that PA3611 promoted EMT via integrin αvβ6-mediated TGF-β1-induced p38/NF-κB pathway activation. The function of PA3611 was also verified in PA-infected rats, and the results showed that ΔPA3611 reduced lung inflammation and EMT. Overall, our results revealed that PA3611 promoted EMT via integrin αvβ6-mediated TGF-β1-induced p38/NF-κB pathway activation, suggesting that PA3611 acts as a crucial virulence factor in bronchial epithelial cell EMT and is a potential target for the clinical treatment of bronchial EMT and fibrosis caused by chronic PA infection.

Combined Resistance to Ceftolozane-Tazobactam and Ceftazidime-Avibactam in Extensively Drug-Resistant (XDR) and Multidrug-Resistant (MDR) Pseudomonas aeruginosa: Resistance Predictors and Impact on Clinical Outcomes Besides Implications for Antimicrobial Stewardship Programs.

A retrospective case-control study was conducted at Modena University Hospital from December 2017 to January 2019 to identify risk factors and predictors of MDR/XDR Pseudomonas aeruginosa (PA) isolation with resistance to ceftazidime/avibactam (CZA) and ceftolozane/tazobactam (C/T), and of mortality among patients infected/colonized. Among 111 PA isolates from clinical/surveillance samples, 60 (54.1%) were susceptible to both drugs (S-CZA-C/T), while 27 (24.3%) were resistant to both (R-CZA-C/T). Compared to patients colonized/infected with S-CZA-C/T, those with R-C/T + CZA PA had a statistically significantly higher Charlson comorbidity score, greater rate of previous PA colonization, longer time before PA isolation, more frequent presence of CVC, higher exposure to C/T and cephalosporins, longer hospital stay, and higher overall and attributable mortality. In the multivariable analysis, age, prior PA colonization, longer time from admission to PA isolation, diagnosis of urinary tract infection, and exposure to carbapenems were associated with the isolation of a R-C/T + CZA PA strain, while PA-related BSI, a comorbidity score > 7, and ICU stay were significantly associated with attributable mortality. C/T and CZA are important therapeutic resources for hard-to-treat PA-related infections, thus specific antimicrobial stewardship interventions should be prompted in order to avoid the development of this combined resistance, which would jeopardize the chance to treat these infections.

Sputum and Plasma Neutrophil Elastase in Stable Adult Patients With Cystic Fibrosis in Relation to Chronic Pseudomonas Aeruginosa Colonization.

Background and purposeNeutrophil elastase (NE) has been implicated in the pathogenesis of airway inflammation in cystic fibrosis (CF) patients and it impairs defenses against Pseudomonas aeruginosa (PA) infection or colonization. Sputum NE may act as a biomarker of neutrophilic inflammation in CF patients. This study aimed to determine sputum and plasma total NE levels in clinically stable adult CF patients and control subjects, and their correlation to PA colonization and lung functions.MethodsThis is a cross-sectional study. Total NE was measured on spontaneously expectorated sputum and plasma obtained from 21 CF patients, aged 18-40 years, during routine visits to the adult CF clinic. This was compared to plasma obtained from 22 matching healthy controls. The levels of NE were measured by the magnetic bead-based multiplex assay.ResultsSputum and plasma NE levels had a significant positive correlation (Pearson r=0.533, P=0.013) with PA colonization. Sixteen CF patients (76.2%) were chronically colonized with PA. Both median sputum and plasma NE were found to be higher in CF patients with PA as compared with non-PA patients, even though this difference was statistically insignificant. Sputum and plasma NE levels did not correlate with the percentage predicted forced expiratory volume in one second (FEV1), the forced vital capacity (FVC), and FEV1/FVC and no association with PA.ConclusionThe findings suggest that clinically stable adult CF patients colonized with PA may have higher NE levels in both plasma and sputum as compared to non-PA CF patients and probably total NE does not influence lung functions.

Exacerbations and Pseudomonas aeruginosa colonization are associated with altered lung structure and function in primary ciliary dyskinesia

BackgroundRecurrent bacterial infections of the respiratory tract are one of the major clinical features of the primary ciliary dyskinesia (PCD), a rare genetic disease due to malfunctioning of motile cilia. Chronic infections and persistent inflammation of the respiratory system result in progressive lung disease.Aim of the study was to highlight the main factors associated with clinical, functional and anatomical deterioration in PCD patients.MethodsWe retrospectively analyzed data from 58 patients with PCD, 37 adults and 21 children. The demographic and clinical data, forced expiratory volume at 1 s (FEV1) and forced vital capacity (FVC), sputum microbiology and imaging results (chest CT scores-modified Bhalla) were recorded. Patients were stratified according to the number of exacerbations (< 2/year vs ≥ 2/year) and chronic Pseudomonas aeruginosa (PA) colonization. The possible correlations between lung function and chest CT scores were assessed; we also evaluated the correlation between these parameters and the severity scores for bronchiectasis (BSI, FACED and e-FACED).ResultsChest CT scores showed a significant correlation with FEV1 (p = 0.0002), age (p < 0.0001), BMI (p = 0.0002) and number of lung lobes involved (p < 0.0001). PA colonization had an overall prevalence of 32.6%: no significant difference in FEV1 between PA colonized and non-colonized patients was found (p = 0.70), while chest CT score was significantly worse in chronic PA colonized patients (p = 0.009). Patients with a high number of exacerbation (≥ 2/year) were older (p = 0.01), had lower FEV1 (p = 0.03), greater number of lobes involved (p < 0.001) and worse CT score than patients with low number of exacerbations (p = 0.001); they also had higher prevalence of PA chronic bronchial infection (33.3% versus 13.6%, p = 0.10). Multivariable linear regression analyses adjusted for gender, age and BMI showed positive associations between PA colonization and number of exacerbations with severity of disease (number of lobes involved, CT score, BSI, FACED, and e-FACED).ConclusionsIn our PCD population the number of exacerbations (≥ 2/year) and PA colonization were the two most relevant factors associated with severity of disease.

Sputum Exosomal microRNAs Profiling Reveals Critical Pathways Modulated By Pseudomonas aeruginosa Colonization In Bronchiectasis.

BackgroundPseudomonas aeruginosa (PA) colonization confers poor prognosis in bronchiectasis. However, the biomarkers and biological pathways underlying these associations are unclear.ObjectiveTo identify the roles of PA colonization in bronchiectasis by exploring for sputum exosomal microRNA profiles.MethodsWe enrolled 98 patients with clinically stable bronchiectasis and 17 healthy subjects. Sputum was split for bacterial culture and exosomal microRNA sequencing, followed by validation with quantitative polymerase chain reaction. Bronchiectasis patients were stratified into PA and non-PA colonization groups based on sputum culture findings. We applied Gene Ontology and Kyoto Encyclopedia of Genes and Genome pathway enrichment analysis to explore biological pathways corresponding to the differentially expressed microRNAs (DEMs) associated with PA colonization.ResultsEighty-two bronchiectasis patients and 9 healthy subjects yielded sufficient sputum that passed quality control. We identified 10 overlap DEMs for the comparison between bronchiectasis patients and healthy subjects, and between PA and non-PA colonization group. Both miR-92b-5p and miR-223-3p could discriminate PA colonization (C-statistic >0.60) and independently correlated with PA colonization in multiple linear regression analysis. The differential expression of miR-92b-5p was validated by quantitative polymerase chain reaction (P<0.05), whereas the differential expression of miR-223 trended towards statistical significance (P=0.06). These DEMs, whose expression levels correlated significantly with sputum inflammatory biomarkers (interleukin-1β and interleukin-8) level, were implicated in the modulation of the nuclear factor-κB, phosphatidylinositol and longevity regulation pathways.ConclusionSputum exosomal microRNAs are implicated in PA colonization in bronchiectasis, highlighting candidate targets for therapeutic interventions to mitigate the adverse impacts conferred by PA colonization.

MULTI-DRUG RESISTANT PATHOGEN SCORES PROMOTE THE OVERUSE OF ANTI-PSEUDOMONAS ANTIBIOTICS

SESSION TITLE: Tuesday Electronic Posters 1 SESSION TYPE: Original Inv Poster Discussion PRESENTED ON: 10/22/2019 01:00 PM - 02:00 PM PURPOSE: Appropriate Pseudomonas aeruginosa (PA) antibiotic empiric selection represents a clinical and practical challenge for healthcare providers. We aimed to assess the appropriate, over- and under-utilization of anti-PA antibiotics when using the recommended regimen based on specific multi-drug resistant (MDR) scores for hospitalized adult patients with community acquire pneumonia (CAP). METHODS: We used the GLIMP study platform (international point-prevalence study) of adult patients hospitalized for CAP (n=3193)[1]. Anti-PA empiric utilization was tested according to the MDR scores: HCAP, Shorr, Shindo, DRIP, Park, Aliberti and GLIMP-PA derived (included: prior PA infection or colonization (pPA) +/- at least one lung disease (i.e. bronchiectasis, very severe COPD [FEV1 <30%], tracheostomy present on admission), respectively [1-7]. Scores were adapted accordingly, with the use of prior hospitalization and antibiotic therapies within one year prior to the index pneumonia hospitalization. The primary outcomes were whether appropriate, over- and under-utilization of anti-PA therapy was recommended. Appropriate use was defined when the score recommended anti-PA therapy when PA-CAP was diagnosed. Over-utilization refers to a score that recommends anti-PA coverage without PA-CAP confirmation. Underutilization was defined when the score did not recommend the use of anti-PA antibiotics when PA-CAP is confirmed. RESULTS: The different scores suggested anti-PA coverage for 22.2-49.0% of hospitalized CAP patients (except for GLIMP-PA [10.7%]). PA-CAP was diagnosed in 4.2% of patients (n=133). Appropriate suggested anti-PA coverage was 6.4%-Shorr, 7.1%-Shindo, 10.0%-DRIP, 7.3-%Park, 6.3%-Aliberti, 6.8%-HCAP and 20.5%-GLIMP-PA, respectively. Over-utilization of anti-PA therapy was 93.6%-Shorr, 92.9%-Shindo, 90.0%-DRIP, 92.7%-Park, 93.7%-Aliberti, 93.2%-HCAP and 79.5%-GLIMP-PA, respectively. Underutilization of anti-PA therapies was 2.0%-Shorr, 3.0%-Shindo, 2.5%-DRIP, 2.5%-HCAP, 2.6%-Park, 2.7%-Aliberti and 2.2%-GLIMP-PA, respectively. CONCLUSIONS: Specific PA scoring systems over recommend anti-PA antibiotics for adult patients with CAP and despite the strong concern for PA there is a very low underutilization risk. CLINICAL IMPLICATIONS: Implementation programs may prevent over-utilization of antibiotics in patients admitted with CAP at risk for MDR pathogens, in particular PA. DISCLOSURES: No relevant relationships by Stefano Aliberti, source=Web Response No relevant relationships by Marcos Restrepo, source=Web Response No relevant relationships by Alejandro Rodriguez Oviedo, source=Web Response No relevant relationships by Alexander Shaffer, source=Web Response No relevant relationships by Nilam Soni, source=Web Response

Simultaneous analysis of gene expression and T cell repertoire in peripheral blood mononuclear cells from patients with cystic fibrosis at the single cell level

Abstract Highly sensitive RNA sequencing (RNA-Seq) methods enables gene expression analysis of low-input samples and even single cells. Single-Cell RNA sequencing (scRNA-seq) is a powerful tool for studying the transcriptome and enables discovery of cellular differences usually masked by bulk sampling methods. The Chromium Single Cell Immune Profiling Solution developed by 10X genomics makes it possible to examine gene expression and T cell receptors simultaneously on a cell-by-cell basis. Patients with cystic fibrosis (CF) often suffer inflammatory lung diseases caused by chronic Pseudomonas aeruginosa (PA) colonization. This colonization could also skew the lymphocytes repertoire in peripheral blood mononuclear cells from CF patients. We investigated the use of scRNA-seq to assess the lymphocyte repertoire in CF patients with PA colonization and whether the single-cell transcriptomic changes correlate with preferential T cell receptor (TCR) usage. Our analysis did not find evidence for clonal expansion of T cells in peripheral blood of these CF patients, suggesting that PA colonization mainly elicits local adaptive immune responses in the lungs. However, this dataset did reveal a striking depletion of the population of T cells known as mucosal associated invariant T cells (MAIT cells) in CF patients. Within the MAIT cell population, we found an overrepresentation of certain genes that was observable in healthy but not CF samples. In this study we show the feasibility of simultaneous analysis of gene expression and lymphocytes repertoire in PBMCs. We recapitulate previous findings that implicate a protective role for MAIT cells against PA colonization, and also provide evidence for the involvement of particular genes in this effect.

Does screening neonates in the neonatal intensive care unit for Pseudomonas aeruginosa colonization help prevent infection?

Pseudomonas aeruginosa (PA) is a Gram-negative environmental organism that can cause severe infection in immunosuppressed patients, including preterm neonates. In recent years, it has become common practice to screen neonates for PA colonization. To assess the value of screening neonates for PA in (1) predicting the risk of developing severe PA infection and (2) directing infection control practice. Between August 2012 and September 2015, babies admitted to the neonatal intensive care unit (NICU) at North Bristol NHS Trust were screened routinely for PA colonization on admission and weekly thereafter. Data were also collected on babies who developed PA infection. Environmental samples from the NICU were tested for the presence of PA. Variable number tandem repeat (VNTR) typing was performed on all strains of PA from babies and the environment. No babies with positive screens subsequently developed PA infection. There was no VNTR strain evidence supporting cross-infection from the environment or other babies. Screening neonates for PA did not identify babies who subsequently developed PA infection. Following cessation of screening in September 2015, there was no increase in the number of babies identified with PA infection.

Increased cytokines in cystic fibrosis patients' upper airways during a new P. aeruginosa colonization.

Previously, we found linkages of inflammatory mediator levels in CF upper airways (UAW) sampled by nasal lavage (NL) to disease severity and to chronic pathogen colonization such as Pseudomonas aeruginosa (PsA). Here, we assess UAW cytokine dynamics in CF patients with a new PsA colonization. We measured cytokines in 149 longitudinally obtained NL samples from 34 CF patients. Cytokine concentrations determined prior to, at the time of de novo PsA detection in either UAW or lower airways (LAW), and in a subsequent PsA free period in newly colonized patients (PsA-new/n = 7) were compared to levels of not- (PsA-free/n = 13) and chronically colonized patients (PsA-chron/n = 14). Moreover, serological and clinical data were compiled. Concentrations of IL-1ß, IL-6, and IL-8 in samples taken prior to new PsA detection were comparable with PsA-free patients. At the time of PsA detection and, most interestingly, irrespective of whether PsA occurred in the UAW or LAW, IL-8 increased (P = 0.009) and IL-6 tended to increase (P = 0.081). In these patients, detection of PsA was not related to elevated PsA antibody-titers. In comparison, NL of PsA-chron patients revealed generally lower IL-8 and IL-1β concentrations as in PsA-free patients, most likely due to a consequent antibiotic and anti-inflammatory therapy (eg, with azithromycin). Monitoring cytokine dynamics in the UAW by serial NL sampling may be valuable in the early phase of PsA acquisition and, thus, increase the chance to adjust treatment options early and more specifically.

Clinical impact of Pseudomonas aeruginosa colonization in patients with Primary Ciliary Dyskinesia.

Airway infections in Primary Ciliary Dyskinesia (PCD) are caused by different microorganisms, including pseudomonas aeruginosa (PA). The aim of this study was to investigate the association of PA colonization and the progression of lung disease in PCD. Data from 11PCD centers were retrospectively collected from 2008 to 2013. Patients were considered colonized if PA grew on at least two separate sputum cultures; otherwise, they were classified as non-colonized. These two groups were compared on the lung function computed tomography (CT) Brody score and other clinical parameters. Data were available from 217 patients; 60 (27.6%) of whom were assigned to the colonized group. Patients colonized with PA were older and were diagnosed at a later age. Baseline forced expiratory volume at 1s (FEV1) was lower in the colonized group (72.4±22.0 vs. 80.1±18.9, % predicted, p=0.015), but FEV1 declined throughout the study period was similar in both groups. The colonized group had significantly worse CT-Brody scores (36.07±24.38 vs. 25.56±24.2, p=0.034). A subgroup analysis with more stringent definitions of colonization revealed similar results. Lung PA colonization in PCD is associated with more severe disease as shown by the FEV1 and CT score. However, the magnitude of decline in pulmonary function was similar in colonized and non-colonized PCD patients.

Relative resistance index (RRI) – a scoring system for antibiotic resistance in Pseudomonas aeruginosa

Background: There is a need to measure antibiotic resistance of Pseudomonas aeruginosa (PA) in cystic fibrosis (CF), either qualitatively or quantitatively, to inform patient management. The aim of this study was to develop a simple method by which resistance can be quantified by calculating a relative resistance index (RRI), and to assess correlation of RRIs with clinical variables.Methods: In our model, RRIs were calculated based on resistance to aztreonam, ceftazidime, ciprofloxacin, colistin, meropenem, tazocin, temicillin and tobramycin. Eighty-five adults with CF and chronic PA colonisation were identified. For each, all PA cultures were allocated a score of 0 for susceptible, 0.5 for intermediate resistance or 1 for resistance for each antibiotic listed above, and the RRI calculated by dividing the sum of these by the number of antibiotics, giving a maximum score of 1. The mean RRIs for all cultures were correlated with key clinical variables monitored in CF patients (including age, FEV1, IV antibiotic days and BMI).Results: RRIs for non-mucoid PA exhibited moderate positive correlation with total number of IV days (r = 0.405; p < 0.001) and moderate negative correlation with FEV1 % predicted (r = −0.437; p < 0.001). RRIs were not significantly correlated with duration of colonisation, typing (clonal vs other strain) or BMI. Median RRIs were significantly higher for females (0.26, IQR 0.13–0.54) than males (0.18, IQR 0.07–0.37) for non-mucoid PA only (p = 0.03).Conclusions: RRI is an easily calculated measure that correlates with other clinical variables in CF patients and enables quantitative monitoring of resistance.

Low Bone Mineral Density and Associated Factors in Patients with Cystic Fibrosis: A Cross-Sectional Study

Background: Failure to maintain bone mass density is a major complication in patients with cystic fibrosis (CF). This study was conducted to evaluate the prevalence of low bone mineral density (BMD) and also identifying associated risk factors in CF patients. Materials and Methods: Present study conducted on 59 CF patients aged 5-35 years referred to respiratory clinic of Masih Daneshvari Hospital, Tehran-Iran. BMD was measured using dual energy X-ray absorptiometry (DXA) scan. Patients were divided in two groups: cases aged 5-18 years as group A and cases over 18 years as group B. Anthropometric variables, corticosteroid usage, pulmonary function test, serum calcium, phosphate and 25-OH vitamin D were assessed and correlation of them with BMD was investigated.Results: Low BMD (Z score < -2 standard deviation) was found in 72.8% (44) of patients. There was a positive correlation between malnutrition, Forced Expiratory Volume 1 (FEV1) and BMD (r=59 and 0.47, P<0.01, respectively). Steroid therapy and Pseudomonas aeruginosa colonization correlated significantly inversely with BMD (r = -0.34 and -0.32, P<0.05). Vitamin D deficiency was found in 36.7% (18) CF patients. No significant correlation was found between 25-OH vitamin D levels and BMD (r = 0.17; P=0. 23).Conclusion: In present study, the prevalence of low BMD was about 72.8% with significant correlation with low weight, BMI (poor nutritional status), FEV1,Pseudomonas aeruginosacolonization and the use of glucocorticoids.

Co-colonisation with Aspergillus fumigatus and Pseudomonas aeruginosa is associated with poorer health in cystic fibrosis patients: an Irish registry analysis

BackgroundPulmonary infection is the main cause of death in cystic fibrosis (CF). Aspergillus fumigatus (AF) and Pseudomonas aeruginosa (PA) are the most prevalent fungal and bacterial pathogens isolated from the CF airway, respectively. Our aim was to determine the effect of different colonisation profiles of AF and PA on the clinical status of patients with CF.MethodsA retrospective analysis of data from the Cystic Fibrosis Registry of Ireland from 2013 was performed to determine the effect of intermittent and persistent colonisation with AF or PA or co-colonisation with both microorganisms on clinical outcome measures in patients with CF. Key outcomes measured included forced expiratory volume in one second (FEV1), number of hospitalisations, respiratory exacerbations and antimicrobials prescribed, and complications of CF, including CF related diabetes (CFRD) and allergic bronchopulmonary aspergillosis (ABPA).ResultsThe prevalence of AF and PA colonisation were 11% (5% persistent, 6% intermittent) and 31% (19% persistent, 12% intermittent) in the Irish CF population, respectively. Co-colonisation with both pathogens was associated with a 13.8% reduction in FEV1 (p = 0.016), higher levels of exacerbations (p = 0.042), hospitalisations (p = 0.023) and antimicrobial usage (p = 0.014) compared to non-colonised patients and these clinical outcomes were comparable to those persistently colonised with PA. Intermittent and persistent AF colonisation were not associated with poorer clinical outcomes or ABPA. Patients with persistent PA had a higher prevalence of CFRD diagnosis (p = 0.012).ConclusionsCF patients co-colonised with AF and PA had poor clinical outcomes comparable to patients persistently colonised with PA, emphasising the clinical significance of co-colonisation with these microorganisms.

Pseudomonas aeruginosa Ventilator-Associated Pneumonia Induces Lung Injury through TNF-α/c-Jun NH2-Terminal Kinase Pathways.

Ventilator-associated pneumonia (VAP) is a common nosocomial infection among intensive care unit (ICU) patients. Pseudomonas aeruginosa (PA) is the most common multidrug-resistant Gram-negative pathogen and VAP caused by PA carries a high rate of morbidity and mortality. This study examined the molecular mechanism of PA VAP-induced lung injury. C57BL/6 wild-type (WT) mice and JNK1 knockout (JNK1-/-) mice received mechanical ventilation (MV) for 3 h at 2 days after receiving nasal instillation of PA. The WT and JNK1-/- mice also received MV after the induction of lung injury by instillation of supernatants from PA-stimulated alveolar macrophages (AMs). AMs isolated from WT, IκB-kinase (IKK)βΔMye (IKKβ was selectively deleted in macrophages), and JNK1-/- mice were ex vivo stimulated with live PA and supernatants were collected for cytokine assay. Intranasal instillation of 106 PA enhanced MV-induced NF-κB DNA binding activity in the lungs and nitrite levels in BALF. MV after PA instillation significantly increased the expression of ICAM and VCAM in the lungs and TNF-α, IL-1β, and IL-6 levels in bronchoalveolar lavage fluid (BALF) of WT mice, but not in JNK1-/- mice. MV after supernatant instillation induced more total protein concentration in BALF and neutrophil sequestration in the lungs in WT mice than JNK1-/- mice and cytokine assay of supernatants indicated that TNF-α is a critical regulator of PA VAP-induced lung injury. Ex vivo PA stimulation induced TNF-α production by AMs from WT as well as JNK1-/- mice but not IKKβΔMye mice. In summary, PA colonization plays an important role in PA VAP-induced lung injury through the induction of JNK1-mediated inflammation. These results suggest that the pathogenesis mechanism of PA VAP involves production of TNF-α through activation of IKK/NF-κB pathways in AMs and JNK signaling pathway in the lungs.

Predictors of deterioration of lung function in Polish children with cystic fibrosis

IntroductionSeverity of lung disease varies in patients with the same CFTR genotype. It suggests that other factors affect the severity of cystic fibrosis (CF). The aim of the study was to identify risk factors that determine lung function decline in Polish cystic fibrosis children.Material and methodsThe follow-up time was no less than 5 years of respiratory status observation based on the forced expiratory volume in 1 s value (FEV1). The socio-economic data, perinatal interview, presence of meconium ileus (MI), time of CF diagnosis, initiation of tobramycin inhalation solution (TIS), pancreatic function, sensitization to Aspergillus fumigatus, presence of impaired glucose tolerance (IGT) or diabetes mellitus, chronic bacterial colonization and number of exacerbations and hospitalizations were assessed.ResultsThe mean age of 61 included children was 13.3 ±7.6 years. Delta F508 homozygosity was detected in 45.9%, 44.3% were delta F508 heterozygous, and 9.8% had other genotypes. FEV1 decline was observed among 20% of patients; the rest of the patients presented stable values of FEV1 during at least 5 years of observation. The most significant predictors related to the decline of FEV1 were presentation of MI (p = 0.0344), IGT (p = 0.0227), number of exacerbations (p = 0.0288), and early Pseudomonas aeruginosa (PA) chronic colonization (p = 0.0165) followed by late TIS initiation after the first detection of PA (p=0.0071). Neither time of diagnosis nor type of CFTR mutation was statistically significant as a predictor of lung deterioration.ConclusionsThe presence of MI, IGT, chronic PA colonization, and number of exacerbations are risk factors for lung function deterioration.

Prospective observational study of prior rectal colonization status as a predictor for subsequent development of Pseudomonas aeruginosa clinical infections.

The potential role of Pseudomonas aeruginosa (PA) intestinal colonization in the subsequent development of infections has not been thoroughly investigated. The aims of this study were to assess the role of PA intestinal colonization as a predictor of subsequent infections and to investigate the risk factors associated with the development of PA infection in patients in the intensive care unit (ICU). For this purpose, a prospective study was conducted that included (i) active surveillance of PA rectal colonization at ICU admission and weekly until ICU discharge, (ii) detection of PA clinical infections, and (iii) genotypic analysis by pulsed-field gel electrophoresis (PFGE). A total of 414 patients were included, of whom 179 (43%) were colonized with PA. Among the 77 patients who developed PA infection, 69 (90%) had prior PA colonization, and 60 (87%) of these showed genotyping concordance between rectal and clinical isolates. The probability of PA infection 14 days after ICU admission was 26% for carriers versus 5% for noncarriers (P < 0.001). Cox regression analysis identified prior PA rectal colonization as the main predictor of PA infection (hazard ratio [HR], 15.23; 95% confidence interval [CI], 6.9 to 33.7; P < 0.001). Prior use of nonantipseudomonal penicillins was also identified as an independent variable associated with PA infection (HR, 2.15; 95% CI, 1.3 to 3.55; P < 0.003). Our study demonstrated that prior PA rectal colonization is a key factor for developing PA infection.

107 Sputum flora among teenage cystic fibrosis patients using inhaled TOBI therapy for 5 and more years from Sofia CF center

Objectives To assess the diversity of the sputum flora among teenaged cystic fibrosis (CF) patients with chronic Pseudomonas aeruginosa (PA) colonization treated for 5 years and more with inhaled TOBI (TIS), as well as the safety of inhaled drug. Methods Microbiological investigations of sputum flora from 22 CF patients (13 boys and 9 girls), aged 12–18 y (mean age 15.7 y) receiving 5 years and more of inhaled TOBI were done together with a main blood biochemical assessment of liver condition (ALAT, ASAT, LDH), renal function (creatinine, urea, ureic acid), lung functional tests (FEV1) and ototoxicity (audiometric testing). Results PA in sputum was permanently established in 6 patients (27.27%), in two of them in combination with other pathogens; Candida sp. was found in 3 (13.63%), methicillin resistant Staphylococcus aureus in 6 patients, Burkholderia cepacia in one. Stenotrophomonas maltophilia , Achromobacter xylosoxidans, Aspergillus sp. and non-tuberculous mycobacteria were not found. 7 patients (31.81%) were free of sputum pathogens. FEV1 less than 40% of predicted showed 5 patients (22.73%). Out of a slight elevation of ureic acid level in 1 patient none of the others revealed liver and renal dysfunction, neither signs of ototoxicity. Conclusion 5 years and more of inhaled TOBI therapy is safe, shows effective results in more than 75% of chronically PA infected teenaged CF patients and leads to permanent full sputum pathogen eradication in at least 1/3 of them. New drug inhaled formulae are needed for each fourth teenaged CF patients with chronic PA colonization.

S100 Novel T helper cell responses against Pseudomonas aeruginosa in healthy individuals and patients with cystic fibrosis

Introduction and Objectives Pseudomonas aeruginosa (PA) colonisation is a hallmark of cystic fibrosis (CF) resulting in damaging neutrophilic inflammation. Patients with CF produce anti-pseudomonal antibodies but the role of CD4 + T cell responses to PA remains unclear. Novel T helper cell subsets, Th17 and Th22 cells, have important roles in host defense but may also enhance tissue damage. We aimed to define the antigen-specific memory T helper cell responses to Pseudomonas aeruginosa in healthy humans and patients with cystic fibrosis. Methods CD14 + monocytes and memory CD4 + CD45RO + T cells were isolated from peripheral blood of CF patients with PA colonisation (n = 8) and healthy controls (n = 10). Monocyte-derived dendritic cells (DCs) were stimulated with live Pseudomonas strain PA103 and PA isolates derived from CF patients. Autologous T cells were co-cultured with activated DCs. The resultant T helper response was determined by measuring proliferation, immunoassay of cytokine output, and immunostaining of intracellular cytokines. Lavage samples from explanted CF lungs were assayed for IL-22 secretion. Results Healthy individuals andpatients with cystic fibrosis had robust antigen-specific memory CD4 + T cell responses to Pseudomonas aeruginosa that not only contained a Th1 and Th17 component but also Th22 cells. In contrast to previous descriptions of human Th22 cells, these Pseudomonal-specific Th22 cells lacked the skin homing markers CCR4 or CCR10, although they did express the chemokine receptor CCR6 that would direct migration to damaged epithelial surfaces. Furthermore, IL-22 production was evident in the lungs of CF patients colonised with PA. Healthy individuals and patients with cystic fibrosis had similar levels of Th22 and Th1 cells, but the patient group had significantly fewer Th17 cells in peripheral blood. Conclusions MemoryTh22 cells specific to Pseudomonas aeruginosa are induced in both healthy individuals and patients with CF, with IL-22 secretion being demonstrated in the CF lung. These Th22 cells do not express tissue specificity for gut or skin sites and we thus hypothesise may have a role in respiratory defense. Along with Th17 cells, they may play an important role in the pathogenesis of pulmonary infection with this microbe in patients with cystic fibrosis.