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Moraxella catarrhalis, a major nasopharyngeal pathogen of the human respiratory tract, is exposed to rapid and prolonged downshifts of environmental temperature when humans breathe cold air. In the present study, we show that a 26°C cold shock up-regulates the expression of UspA1, a major adhesin and putative virulence factor of M. catarrhalis, by prolonging messenger RNA half-life. Cold shock promotes M. catarrhalis adherence to upper respiratory tract cells via enhanced binding to fibronectin, an extracellular matrix component that mediates bacterial attachment. Exposure of M. catarrhalis to 26°C increases the outer membrane protein-mediated release of the proinflammatory cytokine interleukin 8 in pharyngeal epithelial cells. Furthermore, cold shock at 26°C enhances the binding of salivary immunoglobulin A on the surface of M. catarrhalis. These data indicate that cold shock at a physiologically relevant temperature of 26°C affects the nasopharyngeal host-pathogen interaction and may contribute to M. catarrhalis virulence

Background. Early exposure of infants and long-term immunity suggest that colonization with Moraxella catarrhalis is more frequent than is determined by routine culture. We characterized a reservoir of M. catarrhalis in pharyngeal lymphoid tissue. Methods. Tissue from 40 patients (median age, 7.1 years) undergoing elective tonsillectomy and/or adenoidectomy was analyzed for the presence of M. catarrhalis by culture, real-time DNA and RNA polymerase chain reaction (PCR), immunohistochemical analysis (IHC), and fluorescent in situ hybridization (FISH). Histologic sections were double stained for M. catarrhalis and immune cell markers, to characterize the tissue distribution of the organism. Intracellular bacteria were identified using confocal laser scanning microscopy (CLSM). Results. Twenty-nine (91%) of 32 adenoids and 17 (85%) of 20 tonsils were colonized with M. catarrhalis. Detection rates for culture, DNA PCR, RNA PCR, IHC, and FISH were 7 (13%) of 52, 10 (19%) of 52, 21 (41%) of 51, 30 (61%) of 49, and 42 (88%) of 48, respectively (P < .001). Histologic analysis identified M. catarrhalis in crypts, intraepithelially, subepithelially, and (using CLSM) intracellularly. M. catarrhalis colocalized with macrophages and B cells in lymphoid follicles. Conclusions. Colonization by M. catarrhalis is more frequent than is determined by surface culture, because the organism resides both within and beneath the epithelium and invades host cells

Objectives The outer membrane protein M35 of Moraxella catarrhalis is an antigenically conserved porin. Knocking out M35 significantly increases the MICs of aminopenicillins. The aim of this study was to determine the biological mechanism of this potentially new antimicrobial resistance mechanism of M. catarrhalis and the behaviour of M35 in general stress situations. Methods PCR using m35-specific primers was used to detect the m35 gene in clinical isolates. The m35 mRNA expression of strains 300, O35E and 415 after exposure to amoxicillin and different stress conditions was measured by real-time PCR and normalized in relation to their 16S rRNA expression. The expression of M35 protein was analysed by SDS-PAGE and western blotting. Results Screening of 52 middle ear isolates resulted in positive PCR products for all tested strains. The analysis of m35 mRNA expression after amoxicillin treatment showed 24%-85% down-regulation compared with the respective amoxicillin-free controls in all three strains tested. Also, analysis of protein concentrations revealed lower M35 expression after growth with amoxicillin. Investigation of M35 during general stress responses showed down-regulation of the porin with growth at 26°C and 42°C, under hyperosmolar stress and under iron restriction. Conclusions The reduced expression of M35 after aminopenicillin exposure indicates a novel resistance mechanism against aminopenicillins in M. catarrhalis, which may be relevant in vivo. The differences in expression after different stress treatments demonstrate that M35 is involved in general stress responses

BACKGROUND: Moraxella catarrhalis, a major nasopharyngeal pathogen of the human respiratory tract, is exposed to rapid downshifts of environmental temperature when humans breathe cold air. The prevalence of pharyngeal colonization and respiratory tract infections caused by M. catarrhalis is greatest in winter. We investigated how M. catarrhalis uses the physiologic exposure to cold air to regulate pivotal survival systems that may contribute to M. catarrhalis virulence. RESULTS: In this study we used the RNA-seq techniques to quantitatively catalogue the transcriptome of M. catarrhalis exposed to a 26 °C cold shock or to continuous growth at 37 °C. Validation of RNA-seq data using quantitative RT-PCR analysis demonstrated the RNA-seq results to be highly reliable. We observed that a 26 °C cold shock induces the expression of genes that in other bacteria have been related to virulence a strong induction was observed for genes involved in high affinity phosphate transport and iron acquisition, indicating that M. catarrhalis makes a better use of both phosphate and iron resources after exposure to cold shock. We detected the induction of genes involved in nitrogen metabolism, as well as several outer membrane proteins, including ompA, m35-like porin and multidrug efflux pump (acrAB) indicating that M. catarrhalis remodels its membrane components in response to downshift of temperature. Furthermore, we demonstrate that a 26 °C cold shock enhances the induction of genes encoding the type IV pili that are essential for natural transformation, and increases the genetic competence of M. catarrhalis, which may facilitate the rapid spread and acquisition of novel virulence-associated genes. CONCLUSION: Cold shock at a physiologically relevant temperature of 26 °C induces in M. catarrhalis a complex of adaptive mechanisms that could convey novel pathogenic functions and may contribute to enhanced colonization and virulence.

Purpose: Curcumin exerts its anti-inflammatory activity via inhibition of nuclear factor κB. Oropharyngeal epithelia and residing bacteria closely interact in inflammation and infection. This in vitro model investigated the effects of curcumin on bacterial survival, adherence to, and invasion of upper respiratory tract epithelia, and studied its anti-inflammatory effect. We aimed to establish a model, which could offer insights into the host-pathogen interaction in cancer therapy induced mucositis. Methods: Moraxella catarrhalis (Mcat) and the oropharyngeal epithelial cell line Detroit 562 were used. Time-kill curves assessed the inhibition of bacterial growth and adherence assays and gentamicin protection assays determined the effect of curcumin-preincubated cells on bacterial adherence and invasion. Curcumin-mediated inhibition of pro-inflammatory activation by Mcat was determined via interleukin-8 concentrations in the supernatants. The synergistic role of secretory IgA (sIgA) on adherence was investigated. Results: Curcumin was bactericidal at concentrations >50µM. Preincubation of Detroit cells for 60min demonstrated that concentrations >100µM inhibited bacterial adherence. Together with sIgA, curcumin inhibited adherence at concentrations ≥50µM. Both 100 and 200µM curcumin significantly inhibited Mcat cell invasion. Finally, curcumin inhibited Mcat-induced pro-inflammatory activation by strongly suppressing IL-8 release. At a concentration of 200µM, 10min of curcumin exposure inhibited IL-8 release significantly, and complete suppression required a pre-exposure time of ≥45min. Conclusion: Curcumin, in clinically relevant concentrations for topical use, displayed strong antibacterial effect against a facultative upper respiratory tract pathogen by inhibiting bacterial growth, adherence, invasion, and pro-inflammatory activation of upper respiratory tract epithelial cells in vitro

Moraxella catarrhalis, a major nasopharyngeal pathogen of the human respiratory tract, is exposed to rapid and prolonged downshifts of environmental temperature when humans breathe cold air. In the present study, we show that a 26°C cold shock up-regulates the expression of UspA1, a major adhesin and putative virulence factor of M. catarrhalis, by prolonging messenger RNA half-life. Cold shock promotes M. catarrhalis adherence to upper respiratory tract cells via enhanced binding to fibronectin, an extracellular matrix component that mediates bacterial attachment. Exposure of M. catarrhalis to 26°C increases the outer membrane protein-mediated release of the proinflammatory cytokine interleukin 8 in pharyngeal epithelial cells. Furthermore, cold shock at 26°C enhances the binding of salivary immunoglobulin A on the surface of M. catarrhalis. These data indicate that cold shock at a physiologically relevant temperature of 26°C affects the nasopharyngeal host-pathogen interaction and may contribute to M. catarrhalis virulence

Background. Early exposure of infants and long-term immunity suggest that colonization with Moraxella catarrhalis is more frequent than is determined by routine culture. We characterized a reservoir of M. catarrhalis in pharyngeal lymphoid tissue. Methods. Tissue from 40 patients (median age, 7.1 years) undergoing elective tonsillectomy and/or adenoidectomy was analyzed for the presence of M. catarrhalis by culture, real-time DNA and RNA polymerase chain reaction (PCR), immunohistochemical analysis (IHC), and fluorescent in situ hybridization (FISH). Histologic sections were double stained for M. catarrhalis and immune cell markers, to characterize the tissue distribution of the organism. Intracellular bacteria were identified using confocal laser scanning microscopy (CLSM). Results. Twenty-nine (91%) of 32 adenoids and 17 (85%) of 20 tonsils were colonized with M. catarrhalis. Detection rates for culture, DNA PCR, RNA PCR, IHC, and FISH were 7 (13%) of 52, 10 (19%) of 52, 21 (41%) of 51, 30 (61%) of 49, and 42 (88%) of 48, respectively (P < .001). Histologic analysis identified M. catarrhalis in crypts, intraepithelially, subepithelially, and (using CLSM) intracellularly. M. catarrhalis colocalized with macrophages and B cells in lymphoid follicles. Conclusions. Colonization by M. catarrhalis is more frequent than is determined by surface culture, because the organism resides both within and beneath the epithelium and invades host cells

Objectives The outer membrane protein M35 of Moraxella catarrhalis is an antigenically conserved porin. Knocking out M35 significantly increases the MICs of aminopenicillins. The aim of this study was to determine the biological mechanism of this potentially new antimicrobial resistance mechanism of M. catarrhalis and the behaviour of M35 in general stress situations. Methods PCR using m35-specific primers was used to detect the m35 gene in clinical isolates. The m35 mRNA expression of strains 300, O35E and 415 after exposure to amoxicillin and different stress conditions was measured by real-time PCR and normalized in relation to their 16S rRNA expression. The expression of M35 protein was analysed by SDS-PAGE and western blotting. Results Screening of 52 middle ear isolates resulted in positive PCR products for all tested strains. The analysis of m35 mRNA expression after amoxicillin treatment showed 24%-85% down-regulation compared with the respective amoxicillin-free controls in all three strains tested. Also, analysis of protein concentrations revealed lower M35 expression after growth with amoxicillin. Investigation of M35 during general stress responses showed down-regulation of the porin with growth at 26°C and 42°C, under hyperosmolar stress and under iron restriction. Conclusions The reduced expression of M35 after aminopenicillin exposure indicates a novel resistance mechanism against aminopenicillins in M. catarrhalis, which may be relevant in vivo. The differences in expression after different stress treatments demonstrate that M35 is involved in general stress responses