Chlamydia muridarum lung infection in infants alters hematopoietic cells to promote allergic airway disease in mice

Background Viral and bacterial respiratory tract infections in early-life are linked to the development of allergic airway inflammation and asthma. However, the mechanisms involved are not well understood. We have previously shown that neonatal and infant, but not adult, chlamydial lung infections in mice permanently alter inflammatory phenotype and physiology to increase the severity of allergic airway disease by increasing lung interleukin (IL)-13 expression, mucus hyper-secretion and airway hyper-responsiveness. This occurred through different mechanisms with infection at different ages. Neonatal infection suppressed inflammatory responses but enhanced systemic dendritic cell:T-cell IL-13 release and induced permanent alterations in lung structure (i.e., increased the size of alveoli). Infant infection enhanced inflammatory responses but had no effect on lung structure. Here we investigated the role of hematopoietic cells in these processes using bone marrow chimera studies. Methodology/Principal Findings Neonatal (<24-hours-old), infant (3-weeks-old) and adult (6-weeks-old) mice were infected with C. muridarum. Nine weeks after infection bone marrow was collected and transferred into recipient age-matched irradiated naïve mice. Allergic airway disease was induced (8 weeks after adoptive transfer) by sensitization and challenge with ovalbumin. Reconstitution of irradiated naïve mice with bone marrow from mice infected as neonates resulted in the suppression of the hallmark features of allergic airway disease including mucus hyper-secretion and airway hyper-responsiveness, which was associated with decreased IL-13 levels in the lung. In stark contrast, reconstitution with bone marrow from mice infected as infants increased the severity of allergic airway disease by increasing T helper type-2 cell cytokine release (IL-5 and IL-13), mucus hyper-secretion, airway hyper-responsiveness and IL-13 levels in the lung. Reconstitution with bone marrow from infected adult mice had no effects. Conclusions These results suggest that an infant chlamydial lung infection results in long lasting alterations in hematopoietic cells that increases the severity of allergic airway disease in later-life.

Chronic lung infection by Pseudomonas aeruginosa biofilm is cured by L-Methionine in combination with antibiotic therapy

Bacterial biofilms are associated with 80-90% of infections. Within the biofilm, bacteria are refractile to antibiotics, requiring concentrations >1,000 times the minimum inhibitory concentration. Proteins, carbohydrates and DNA are the major components of biofilm matrix. Pseudomonas aeruginosa (PA) biofilms, which are majorly associated with chronic lung infection, contain extracellular DNA (eDNA) as a major component. Herein, we report for the first time that L-Methionine (L-Met) at 0.5 mu M inhibits Pseudomonas aeruginosa (PA) biofilm formation and disassembles established PA biofilm by inducing DNase expression. Four DNase genes (sbcB, endA, eddB and recJ) were highly up-regulated upon L-Met treatment along with increased DNase activity in the culture supernatant. Since eDNA plays a major role in establishing and maintaining the PA biofilm, DNase activity is effective in disrupting the biofilm. Upon treatment with L-Met, the otherwise recalcitrant PA biofilm now shows susceptibility to ciprofloxacin. This was reflected in vivo, in the murine chronic PA lung infection model. Mice treated with L-Met responded better to antibiotic treatment, leading to enhanced survival as compared to mice treated with ciprofloxacin alone. These results clearly demonstrate that L-Met can be used along with antibiotic as an effective therapeutic against chronic PA biofilm infection.

Treatment of lung infection in patients with cystic fibrosis:Current and future strategies

In patients with cystic fibrosis (CF) lung damage secondary to chronic infection is the main cause of death. Treatment of lung disease to reduce the impact of infection, inflammation and subsequent lung injury is therefore of major importance. Here we discuss the present status of antibiotic therapy for the major pathogens in CF airways, including prophylaxis against infection, eradication of early infection, suppression of chronic infection, and the treatment of infective exacerbations. We outline measures to optimize maintenance treatment for infection in the light of novel antibiotic drug formulations. We discuss new developments in culture-independent microbiological diagnostic techniques and the use of tools for monitoring the success of antibiotic treatment courses. Finally, cost-effectiveness analyses for antibiotic treatment in CF patients are discussed.

A Secretory Leukocyte Protease Inhibitor Variant with Improved Activity against Lung Infection

Secretory leukocyte protease inhibitor (SLPI) is an important respiratory tract host defense protein, which is proteolytically inactivated by excessive neutrophil elastase (NE) during chronic Pseudomonas infection in the cystic fibrosis (CF) lung. We generated two putative NE-resistant variants of SLPI by site-directed mutagenesis, SLPI-A16G and SLPI-S15G-A16G, with a view to improving SLPI’s proteolytic stability. Both variants showed enhanced resistance to degradation in the presence of excess NE as well as CF patient sputum compared with SLPI-wild type (SLPI-WT). The ability of both variants to bind bacterial lipopolysaccharides and interact with nuclear factor-κB DNA binding sites was also preserved. Finally, we demonstrate increased anti-inflammatory activity of the SLPI-A16G protein compared with SLPI-WT in a murine model of pulmonary Pseudomonas infection. This study demonstrates the increased stability of these SLPI variants compared with SLPI-WT and their therapeutic potential as a putative anti-inflammatory treatment for CF lung disease.

Lung infection rates in two sympatric Tropiduridae lizard species by pentastomids and nematodes in northeastern Brazil

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Role of Toll interleukin-1 receptor (IL-1R) 8, a negative regulator of IL-1R/Toll-like receptor signaling, in resistance to acute Pseudomonas aeruginosa lung infection

Toll interleukin-1 receptor (IL-1R) 8 (TIR8), also known as single Ig IL-1 receptor (IL-R)-related molecule, or SIGIRR, is a member of the IL-1R-like family, primarily expressed by epithelial cells. Current evidence suggests that TIR8 plays a nonredundant role as a negative regulator in vivo under different inflammatory conditions that are dependent on IL-R and Toll-like receptor (TLR) activation. In the present study, we examined the role of TIR8 in innate resistance to acute lung infections caused by Pseudomonas aeruginosa, a Gram-negative pathogen responsible for life-threatening infections in immunocompromised individuals and cystic fibrosis patients. We show that Tir8 deficiency in mice was associated with increased susceptibility to acute P. aeruginosa infection, in terms of mortality and bacterial load, and to exacerbated local and systemic production of proinflammatory cytokines (gamma interferon [IFN-γ], tumor necrosis factor alpha [TNF-α], IL-1β, and IL-6) and chemokines (CXCL1, CXCL2, and CCL2). It has been reported that host defense against P. aeruginosa acute lung infection can be improved by blocking IL-1 since exaggerated IL-1β production may be harmful for the host in this infection. In agreement with these data, IL-1RI deficiency rescues the phenotype observed in Tir8-deficient mice: in Tir8-/- IL-1RI-/- double knockout mice we observed higher survival rates, enhanced bacterial clearance, and reduced levels of local and systemic cytokine and chemokine levels than in Tir8-deficient mice. These results suggest that TIR8 has a nonredundant effect in modulating the inflammation caused by P. aeruginosa, in particular, by negatively regulating IL-1RI signaling, which plays a major role in the pathogenesis of this infectious disease.

The therapeutic potential of the humoral pattern recognition molecule PTX3 in chronic lung infection caused by Pseudomonas aeruginosa

Chronic lung infections by Pseudomonas aeruginosa strains are a major cause of morbidity and mortality in cystic fibrosis (CF) patients. Although there is no clear evidence for a primary defect in the immune system of CF patients, the host is generally unable to clear P. aeruginosa from the airways. PTX3 is a soluble pattern recognition receptor that plays nonredundant roles in the innate immune response to fungi, bacteria, and viruses. In particular, PTX3 deficiency is associated with increased susceptibility to P. aeruginosa lung infection. To address the potential therapeutic effect of PTX3 in P. aeruginosa lung infection, we established persistent and progressive infections in mice with the RP73 clinical strain RP73 isolated from a CF patient and treated them with recombinant human PTX3. The results indicated that PTX3 has a potential therapeutic effect in P. aeruginosa chronic lung infection by reducing lung colonization, proinflammatory cytokine levels (CXCL1, CXCL2, CCL2, and IL-1β), and leukocyte recruitment in the airways. In models of acute infections and in in vitro assays, the prophagocytic effect of PTX3 was maintained in C1q-deficient mice and was lost in C3- and Fc common γ-chain-deficient mice, suggesting that facilitated recognition and phagocytosis of pathogens through the interplay between complement and FcγRs are involved in the therapeutic effect mediated by PTX3. These data suggested that PTX3 is a potential therapeutic tool in chronic P. aeruginosa lung infections, such as those seen in CF patients.

The surface of Pseudomonas aeruginosa in cystic fibrosis lung infection

Chronic experimental lung infection in rats was induced by intratracheal inoculation of agar beads containing Pseudomonas aeruginosa. Bacteria were recovered directly without subculture from the lungs of rats at 14 days post-infection and the outer membrane (OM) antigens were studied. The results indicated that bacteria grew under iron-restricted conditions as revealed by the expression of several iron-regulated membrane proteins (IRMPs) which could also be observed when the isolate was grown under iron-depleted conditions in laboratory media. The antibody response to P. aeruginosa OM protein antigens was investigated by immunoblotting with serum and lung fluid from infected rats. These fluids contained antibodies to all the major OM proteins, including the IRMPs, and protein H1. Results obtained using immunoblotting and enzyme-linked immunosorbent assay indicated that lipopolysaccharide (LPS) was the major antigen recognised by antibodies in sera from infected rats. The animal model was used to follow the development of the immune response to P. aeruginosa protein and LPS antigens. Immunoblotting was used to investigate the antigens recognised by antibodies in sequential serum samples. An antibody response to the IRMPs and OM proteins D, E, G and H1 and alao to rough LPS was detected as early as 4 days post-infection. Results obtained using immunoblotting and crossed immunoelectrophoresis techniques indicated that there was a progressive increase in the number of P. aeruginosa antigens recognised by antibodies in these sera. Both iron and magnesium depletion influenced protein H1 production. Antibodies in sera from patients with infections due to P. aeruginosa reacted with this antigen. Results obtained using quantitative gas-liquid chromatographic analysis indicated that growth phase and magnesium and iron depletion also affected the amount of LPS fatty acids, produced by P. aeruginosa. The silver stained SDS-polyacrylamide gels of proteinase K digested whole cell lysates of P. aeruginosa indicated that the O-antigen and core LPS were both affected by growth phase and specific nutrient depletion.

Comparison of intranasal and transcutaneous immunization for induction of protective immunity against chlamydia muridarum respiratory tract infection

Chlamydia pneumoniae causes a range of respiratory infections including bronchitis, pharyngitis and pneumonia. Infection has also been implicated in exacerbation/initiation of asthma and chronic obstructive pulmonary disease (COPD) and may play a role in atherosclerosis and Alzheimer's disease. We have used a mouse model of Chlamydia respiratory infection to determine the effectiveness of intranasal (IN) and transcutaneous immunization (TCI) to prevent Chlamydia lung infection. Female BALB/c mice were immunized with chlamydial major outer membrane protein (MOMP) mixed with cholera toxin and CpG oligodeoxynucleotide adjuvants by either the IN or TCI routes. Serum and bronchoalveolar lavage (BAL) were collected for antibody analysis. Mononuclear cells from lung-draining lymph nodes were stimulated in vitro with MOMP and cytokine mRNA production determined by real time PCR. Animals were challenged with live Chlamydia and weighed daily following challenge. At day 10 (the peak of infection) animals were sacrificed and the numbers of recoverable Chlamydia in lungs determined by real time PCR. MOMP-specific antibody-secreting cells in lung tissues were also determined at day 10 post-infection. Both IN and TCI protected animals against weight loss compared to non-immunized controls with both immunized groups gaining weight by day 10-post challenge while controls had lost 6% of body weight. Both immunization protocols induced MOMP-specific IgG in serum and BAL while only IN immunization induced MOMP-specific IgA in BAL. Both immunization routes resulted in high numbers of MOMP-specific antibody-secreting cells in lung tissues (IN > TCI). Following in vitro re-stimulation of lung-draining lymph node cells with MOMP; IFNγ mRNA increased 20-fold in cells from IN immunized animals (compared to non-immunized controls) while IFNγ levels increased 6- to 7-fold in TCI animals. Ten days post challenge non-immunized animals had >7000 IFU in their lungs, IN immunized animals <50 IFU and TCI immunized animals <1500 IFU. Thus, both intranasal and transcutaneous immunization protected mice against respiratory challenge with Chlamydia. The best protection was obtained following IN immunization and correlated with IFNγ production by mononuclear cells in lung-draining LN and MOMP-specific IgA in BAL.

Interleukin-13 promotes susceptibility to chlamydial infection of the respiratory and genital tracts

Chlamydiae are intracellular bacteria that commonly cause infections of the respiratory and genital tracts, which are major clinical problems. Infections are also linked to the aetiology of diseases such as asthma, emphysema and heart disease. The clinical management of infection is problematic and antibiotic resistance is emerging. Increased understanding of immune processes that are involved in both clearance and immunopathology of chlamydial infection is critical for the development of improved treatment strategies. Here, we show that IL-13 was produced in the lungs of mice rapidly after Chlamydia muridarum (Cmu) infection and promoted susceptibility to infection. Wild-type (WT) mice had increased disease severity, bacterial load and associated inflammation compared to IL-13 deficient (−/−) mice as early as 3 days post infection (p.i.). Intratracheal instillation of IL-13 enhanced bacterial load in IL-13−/− mice. There were no differences in early IFN-g and IL-10 expression between WT and IL-13−/− mice and depletion of CD4+ T cells did not affect infection in IL-13−/− mice. Collectively, these data demonstrate a lack of CD4+ T cell involvement and a novel role for IL-13 in innate responses to infection. We also showed that IL-13 deficiency increased macrophage uptake of Cmu in vitro and in vivo. Moreover, the depletion of IL-13 during infection of lung epithelial cells in vitro decreased the percentage of infected cells and reduced bacterial growth. Our results suggest that enhanced IL-13 responses in the airways, such as that found in asthmatics, may promote susceptibility to chlamydial lung infection. Importantly the role of IL-13 in regulating infection was not limited to the lung as we showed that IL-13 also promoted susceptibility to Cmu genital tract infection. Collectively our findings demonstrate that innate IL-13 release promotes infection that results in enhanced inflammation and have broad implications for the treatment of chlamydial infections and IL-13-associated diseases.