Effect of shear stress on Pseudomonas aeruginosa isolated from the cystic fibrosis lung

Chronic colonization of the lungs by Pseudomonas aeruginosa is one of the major causes of morbidity and mortality in cystic fibrosis (CF) patients. To gain insights into the characteristic biofilm phenotype of P. aeruginosa in the CF lungs, mimicking the CF lung environment is critical. We previously showed that growth of the non-CF-adapted P. aeruginosa PAO1 strain in a rotating wall vessel, a device that simulates the low fluid shear (LS) conditions present in the CF lung, leads to the formation of in-suspension, self-aggregating biofilms. In the present study, we determined the phenotypic and transcriptomic changes associated with the growth of a highly adapted, transmissible P. aeruginosa CF strain in artificial sputum medium under LS conditions. Robust self-aggregating biofilms were observed only under LS conditions. Growth under LS conditions resulted in the upregulation of genes involved in stress response, alginate biosynthesis, denitrification, glycine betaine biosynthesis, glycerol metabolism, and cell shape maintenance, while genes involved in phenazine biosynthesis, type VI secretion, and multidrug efflux were downregulated. In addition, a number of small RNAs appeared to be involved in the response to shear stress. Finally, quorum sensing was found to be slightly but significantly affected by shear stress, resulting in higher production of autoinducer molecules during growth under high fluid shear (HS) conditions. In summary, our study revealed a way to modulate the behavior of a highly adapted P. aeruginosa CF strain by means of introducing shear stress, driving it from a biofilm lifestyle to a more planktonic lifestyle. IMPORTANCE: Biofilm formation by Pseudomonas aeruginosa is one of the hallmarks of chronic cystic fibrosis (CF) lung infections. The biofilm matrix protects this bacterium from antibiotics as well as from the immune system. Hence, the prevention or reversion of biofilm formation is believed to have a great impact on treatment of chronic P. aeruginosa CF lung infections. In the present study, we showed that it is possible to modulate the behavior of a highly adapted transmissible P. aeruginosa CF isolate at both the transcriptomic and phenotypic levels by introducing shear stress in a CF-like environment, driving it from a biofilm to a planktonic lifestyle. Consequently, the results obtained in this study are of great importance with regard to therapeutic applications that introduce shear stress in the lungs of CF patients

Ferrous Iron Is a Significant Component of Bioavailable Iron in Cystic Fibrosis Airways

Chronic, biofilm-like infections by the opportunistic pathogen Pseudomonas aeruginosa are a major cause of mortality in cystic fibrosis (CF) patients. While much is known about P. aeruginosa from laboratory studies, far less is understood about what it experiences in vivo. Iron is an important environmental parameter thought to play a central role in the development and maintenance of P. aeruginosa infections, for both anabolic and signaling purposes. Previous studies have focused on ferric iron [Fe(III)] as a target for antimicrobial therapies; however, here we show that ferrous iron [Fe(II)] is abundant in the CF lung (~39 µM on average for severely sick patients) and significantly correlates with disease severity (ρ = −0.56, P = 0.004), whereas ferric iron does not (ρ = −0.28, P = 0.179). Expression of the P. aeruginosa genes bqsRS, whose transcription is upregulated in response to Fe(II), was high in the majority of patients tested, suggesting that increased Fe(II) is bioavailable to the infectious bacterial population. Because limiting Fe(III) acquisition inhibits biofilm formation by P. aeruginosa in various oxic in vitro systems, we also tested whether interfering with Fe(II) acquisition would improve biofilm control under anoxic conditions; concurrent sequestration of both iron oxidation states resulted in a 58% reduction in biofilm accumulation and 28% increase in biofilm dissolution, a significant improvement over Fe(III) chelation treatment alone. This study demonstrates that the chemistry of infected host environments coevolves with the microbial community as infections progress, which should be considered in the design of effective treatment strategies at different stages of disease. IMPORTANCE: Iron is an important environmental parameter that helps pathogens thrive in sites of infection, including those of cystic fibrosis (CF) patients. Ferric iron chelation therapy has been proposed as a novel therapeutic strategy for CF lung infections, yet until now, the iron oxidation state has not been measured in the host. In studying mucus from the infected lungs of multiple CF patients from Europe and the United States, we found that ferric and ferrous iron change in concentration and relative proportion as infections progress; over time, ferrous iron comes to dominate the iron pool. This information is relevant to the design of novel CF therapeutics and, more broadly, to developing accurate models of chronic CF infections

Respiratory Bacterial Culture Sampling in Expectorating and Non-expectorating Patients With Cystic Fibrosis

Purpose: Different respiratory sampling methods exist to identify lower airway pathogens in patients with cystic fibrosis (CF), of which bronchoalveolar lavage (BAL), and expectorated sputum are considered the “gold standard.” Because BAL cannot be repeated limitless, the diagnosis of lower respiratory tract infections in non-expectorating patients is challenging. Other sampling techniques are nasal swab, cough swab, and induced sputum. The purpose of this study (NCT02363764) was to compare concordance between the microbiological yield of nasal swab, cough swab, and expectorated sputum in expectorating patients; nasal swab, cough swab, and induced sputum in non-expectorating patients; nasal swab, cough swab, induced sputum, and BAL in patients requiring bronchoscopy (“BAL-group”); and to determine the clinical value of cough swab in non-expectorating patients with CF.Methods: Microbiological yield detected by these different sampling techniques was compared between and within 105 expectorating patients, 30 non-expectorating patients and BAL-group (n = 39) in a single CF clinic. Specificity, sensitivity, positive (PPV), and negative (NPV) predictive values were calculated.Results: Overall low sensitivity (6.3–58.0%) and wide-ranging predictive values (0.0–100.0%) indicated that nasal swab was not appropriate to detect lower airway pathogens [Pseudomonas aeruginosa (Pa), Staphylococcus aureus (Sa), and Haemophilus influenzae (Hi)] in all three patient groups. Microbiological yield, specificity, sensitivity, PPV, and NPV of cough swab and induced sputum were largely similar in non-expectorating patients and in BAL-group (except sensitivity (0.0%) of induced sputum for Hi in BAL-group). Calculations for Pa and Hi could not be performed for non-expectorating patients because of low prevalence (n = 2 and n = 3, respectively). In expectorating patients, concordance was found between cough swab and expectorated sputum, except for Hi (sensitivity of 40.0%).Conclusion: Our findings suggest that cough swab might be helpful in detecting the presence of some typical CF pathogens in the lower airways of clinically stable patients with CF. However, in symptomatic patients, who are unable to expectorate and who have a negative cough swab and induced sample, BAL should be performed as it currently remains the “gold standard.

Ferrous Iron Is a Significant Component of Bioavailable Iron in Cystic Fibrosis Airways

Chronic, biofilm-like infections by the opportunistic pathogen Pseudomonas aeruginosa are a major cause of mortality in cystic fibrosis (CF) patients. While much is known about P. aeruginosa from laboratory studies, far less is understood about what it experiences in vivo. Iron is an important environmental parameter thought to play a central role in the development and maintenance of P. aeruginosa infections, for both anabolic and signaling purposes. Previous studies have focused on ferric iron [Fe(III)] as a target for antimicrobial therapies; however, here we show that ferrous iron [Fe(II)] is abundant in the CF lung (~39 µM on average for severely sick patients) and significantly correlates with disease severity (ρ = −0.56, P = 0.004), whereas ferric iron does not (ρ = −0.28, P = 0.179). Expression of the P. aeruginosa genes bqsRS, whose transcription is upregulated in response to Fe(II), was high in the majority of patients tested, suggesting that increased Fe(II) is bioavailable to the infectious bacterial population. Because limiting Fe(III) acquisition inhibits biofilm formation by P. aeruginosa in various oxic in vitro systems, we also tested whether interfering with Fe(II) acquisition would improve biofilm control under anoxic conditions; concurrent sequestration of both iron oxidation states resulted in a 58% reduction in biofilm accumulation and 28% increase in biofilm dissolution, a significant improvement over Fe(III) chelation treatment alone. This study demonstrates that the chemistry of infected host environments coevolves with the microbial community as infections progress, which should be considered in the design of effective treatment strategies at different stages of disease. IMPORTANCE: Iron is an important environmental parameter that helps pathogens thrive in sites of infection, including those of cystic fibrosis (CF) patients. Ferric iron chelation therapy has been proposed as a novel therapeutic strategy for CF lung infections, yet until now, the iron oxidation state has not been measured in the host. In studying mucus from the infected lungs of multiple CF patients from Europe and the United States, we found that ferric and ferrous iron change in concentration and relative proportion as infections progress; over time, ferrous iron comes to dominate the iron pool. This information is relevant to the design of novel CF therapeutics and, more broadly, to developing accurate models of chronic CF infections

Comparison of culture and qPCR for the detection of Pseudomonas aeruginosa in not chronically infected cystic fibrosis patients

Pseudomonas aeruginosa is the major respiratory pathogen causing severe lung infections among CF patients, leading to high morbidity and mortality. Once infection is established, early antibiotic treatment is able to postpone the transition to chronic lung infection. In order to optimize the early detection, we compared the sensitivity of microbiological culture and quantitative PCR (qPCR) for the detection of P. aeruginosa in respiratory samples of not chronically infected CF patients.Comparative StudyJournal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe

Immunodeficiency in a Child with Rapadilino Syndrome: A Case Report and Review of the Literature

Rapadilino syndrome is a genetic disease characterized by a characteristic clinical tableau. It is caused by mutations in RECQL4 gene. Immunodeficiency is not described as a classical feature of the disease. We present a 2-year-old girl with Rapadilino syndrome with important lymphadenopathies and pneumonia due to disseminated Mycobacterium lentiflavum infection. An immunological work-up showed several unexpected abnormalities. Repeated blood samples showed severe lymphopenia. Immunophenotyping showed low T, B, and NK cells. No Treg cells were seen. T cell responses to stimulations were insufficient. The IL12/IL23 interferon gamma pathway was normal. Gamma globulin levels and vaccination responses were low. With this report, we aim to stress the importance of screening immunodeficiency in patients with RECQL4 mutations for immunodeficiency and the need to further research into its physiopathology

Pandemic A/H1N1v influenza 2009 in hospitalized children: a multicenter Belgian survey

<p>Abstract</p> <p>Background</p> <p>During the 2009 influenza A/H1N1v pandemic, children were identified as a specific "at risk" group. We conducted a multicentric study to describe pattern of influenza A/H1N1v infection among hospitalized children in Brussels, Belgium.</p> <p>Methods</p> <p>From July 1, 2009, to January 31, 2010, we collected epidemiological and clinical data of all proven (positive H1N1v PCR) and probable (positive influenza A antigen or culture) pediatric cases of influenza A/H1N1v infections, hospitalized in four tertiary centers.</p> <p>Results</p> <p>During the epidemic period, an excess of 18% of pediatric outpatients and emergency department visits was registered. 215 children were hospitalized with proven/probable influenza A/H1N1v infection. Median age was 31 months. 47% had ≥ 1 comorbid conditions. Febrile respiratory illness was the most common presentation. 36% presented with initial gastrointestinal symptoms and 10% with neurological manifestations. 34% had pneumonia. Only 24% of the patients received oseltamivir but 57% received antibiotics. 10% of children were admitted to PICU, seven of whom with ARDS. Case fatality-rate was 5/215 (2%), concerning only children suffering from chronic neurological disorders. Children over 2 years of age showed a higher propensity to be admitted to PICU (16% vs 1%, p = 0.002) and a higher mortality rate (4% vs 0%, p = 0.06). Infants less than 3 months old showed a milder course of infection, with few respiratory and neurological complications.</p> <p>Conclusion</p> <p>Although influenza A/H1N1v infections were generally self-limited, pediatric burden of disease was significant. Compared to other countries experiencing different health care systems, our Belgian cohort was younger and received less frequently antiviral therapy; disease course and mortality were however similar.</p