The lipopolysaccharide core of Brucella abortus acts as a shield against innate immunity recognition

Innate immunity recognizes bacterial molecules bearing pathogen-associated molecular patterns to launch inflammatory responses leading to the activation of adaptive immunity. However, the lipopolysaccharide (LPS) of the gram-negative bacterium Brucella lacks a marked pathogen-associated molecular pattern, and it has been postulated that this delays the development of immunity, creating a gap that is critical for the bacterium to reach the intracellular replicative niche. We found that a B. abortus mutant in the wadC gene displayed a disrupted LPS core while keeping both the LPS O-polysaccharide and lipid A. In mice, the wadC mutant induced proinflammatory responses and was attenuated. In addition, it was sensitive to killing by non-immune serum and bactericidal peptides and did not multiply in dendritic cells being targeted to lysosomal compartments. In contrast to wild type B. abortus, the wadC mutant induced dendritic cell maturation and secretion of pro-inflammatory cytokines. All these properties were reproduced by the wadC mutant purified LPS in a TLR4-dependent manner. Moreover, the core-mutated LPS displayed an increased binding to MD-2, the TLR4 co-receptor leading to subsequent increase in intracellular signaling. Here we show that Brucella escapes recognition in early stages of infection by expressing a shield against recognition by innate immunity in its LPS core and identify a novel virulence mechanism in intracellular pathogenic gram-negative bacteria. These results also encourage for an improvement in the generation of novel bacterial vaccines.

Relationship between Azithromycin Susceptibility and Administration Efficacy for Nontypeable Haemophilus influenzae Respiratory Infection

Nontypeable Haemophilus influenzae (NTHI) is an opportunistic pathogen that is an important cause of acute exacerbations of chronic obstructive pulmonary disease (AECOPD). COPD is an inflammatory disease of the airways, and exacerbations are acute inflammatory events superimposed on this background of chronic inflammation. Azithromycin (AZM) is a macrolide antibiotic with antibacterial and anti-inflammatory properties and a clinically proven potential for AECOPD prevention and management. Relationships between AZM efficacy and resistance by NTHI and between bactericidal and immunomodulatory effects on NTHI respiratory infection have not been addressed. In this study, we employed two pathogenic NTHI strains with different AZM susceptibilities (NTHI 375 [AZM susceptible] and NTHI 353 [AZM resistant]) to evaluate the prophylactic and therapeutic effects of AZM on the NTHI-host interplay. At the cellular level, AZM was bactericidal toward intracellular NTHI inside alveolar and bronchial epithelia and alveolar macrophages, and it enhanced NTHI phagocytosis by the latter cell type. These effects correlated with the strain MIC of AZM and the antibiotic dose. Additionally, the effect of AZM on NTHI infection was assessed in a mouse model of pulmonary infection. AZM showed both preventive and therapeutic efficacies by lowering NTHI 375 bacterial counts in lungs and bronchoalveolar lavage fluid (BALF) and by reducing histopathological inflammatory lesions in the upper and lower airways of mice. Conversely, AZM did not reduce bacterial loads in animals infected with NTHI 353, in which case a milder anti-inflammatory effect was also observed. Together, the results of this work link the bactericidal and anti-inflammatory effects of AZM and frame the efficacy of this antibiotic against NTHI respiratory infection.

Relative contributions of lipooligosaccharide inner and outer core modifications to nontypeable Haemophilus influenzae pathogenesis

Nontypeable Haemophilus influenzae (NTHi) is a frequent commensal of the human nasopharynx that causes opportunistic infection in immunocompromised individuals. Existing evidence associates lipooligosaccharide (LOS) with disease, but the specific and relative contributions of NTHi LOS modifications to virulence properties of the bacterium have not been comprehensively addressed. Using NTHi strain 375, an isolate for which the detailed LOS structure has been determined, we compared systematically a set of isogenic mutant strains expressing sequentially truncated LOS. The relative contributions of 2-keto-3-deoxyoctulosonic acid, the triheptose inner core, oligosaccharide extensions on heptoses I and III, phosphorylcholine, digalactose, and sialic acid to NTHi resistance to antimicrobial peptides (AMP), self-aggregation, biofilm formation, cultured human respiratory epithelial infection, and murine pulmonary infection were assessed. We show that opsX, lgtF, lpsA, lic1, and lic2A contribute to bacterial resistance to AMP; lic1 is related to NTHi self-aggregation; lgtF, lic1, and siaB are involved in biofilm growth; opsX and lgtF participate in epithelial infection; and opsX, lgtF, and lpsA contribute to lung infection. Depending on the phenotype, the involvement of these LOS modifications occurs at different extents, independently or having an additive effect in combination. We discuss the relative contribution of LOS epitopes to NTHi virulence and frame a range of pathogenic traits in the context of infection.

Development and testing of an intermediate temperature glass sealant for use in mixed ionic and electronic conducting membrane reactors

High temperature ceramic membranes have interesting possibilities for application in areas of new and developing technologies such as hydrocarbon combustion with carbon dioxide capture and electrochemical promotion of catalysis (EPOC). However, membrane module sealing remains a significant technical challenge. In this work a borosilicate glass sealant (50SiO₂·25B₂O₃·25Na₂O, mol%) was developed to fit the requirements of sealing an air separation membrane system at intermediate temperatures (300-600 °C). The seal was assessed by testing the leak rates under a range of conditions. The parameters tested included the effect of flowrate on the leak rate, the heating and cooling rates of the reactor and the range of temperatures under which the system could operate. Tests for durability and reliability were also performed. It was found that the most favourable reactor configuration employed a reactor with the ceramic pellet placed underneath the inner chamber alumina tube (inverted configuration), using a quartz wool support to keep the membrane in place prior to sealing. Using this configuration the new glass-based seal was found to be a more suitable sealant than traditional alternatives; it produced lower leak rates at all desirable flowrates, with the potential for rapid heating and cooling and multiple cycling, allowing for prolonged usage. © 2010 Elsevier B.V. All rights reserved.

Hydrogen-permeation characteristics of a SrCeO3-based ceramic separation membrane:Thermal, ageing and surface-modification effects

Dense ceramics with mixed protonic-electronic conductivity are of considerable interest for the separation and purification of hydrogen and as electrochemical reactors. In this work, the hydrogen permeability of a Sr_0.97Ce_0.9Yb_0.1O_{3 - δ} (SCYb) membrane with a porous Pt catalytic layer on the hydrogen feed-exposed side has been studied over the temperature range 500-804 °C employing Ar as the permeate sweep gas. A SiO₂-B₂O₃-BaO-MgO-ZnO-based glass-ceramic sealant was successfully employed to seal the membrane to the dual-chamber reactor. After 14 h of exposure to 10% H₂:90% N₂ at 804 °C, the H₂ flux reached a maximum of 33 nmol cm^{- 2} s^{- 1}, over an order of magnitude higher than that obtained on membranes of similar thickness without surface modification. The permeation rate then decreased slowly and moderately on annealing at 804 °C over a further 130 h. Thereafter, the flux was both reproducible and stable on thermal cycling in the range 600-804 °C. The results indicate an important role of superficial activation processes in the flux rate and suggest that hydrogen fluxes can be further optimised in cerate-based perovskites. © 2009 Elsevier B.V. All rights reserved.