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PubMed2003

Morphostructural damage and the inhibition of bacterial adhesiveness of Staphylococcus aureus and Moraxella catarrhalis induced by moxifloxacin.

The aim of this study was to investigate the ability of moxifloxacin to interfere with the mechanism of bacterial adhesion and disrupt the morphological and structural integrity of bacteria. Three Staphylococcus aureus and three Moraxella catarrhalis strains were grown in the presence of 1/2-1/128 minimum inhibitory concentration (MIC) serial dilutions and incubated with human epithelial cells. A significant decrease in adhesion was observed from 1/2 MIC to 1/64 MIC for S. aureus, and from 1/2 MIC to 1/16 MIC for M. catarrhalis. The use of atomic force microscopy, a new technique capable of revealing surface structures in three-dimensional detail and at very high resolution, showed the rapid onset and time course of the sequence of disruptive morphostructural events following the incubation of both S. aureus and M. catarrhalis with sub-MICs of moxifloxacin. Our findings suggest that less than conventional MIC moxifloxacin concentrations may be effective in reducing bacterial adhesiveness and structural integrity on which the maintenance of bacterial activity depends.

Aza Compounds↗
PubMed2004

Antimicrobial activity of innate immune molecules against Streptococcus pneumoniae, Moraxella catarrhalis and nontypeable Haemophilus influenzae.

BACKGROUND: Despite its direct connection to the nasopharynx which harbors otitis media pathogens as part of its normal flora, the middle ear cavity is kept free of these bacteria by as yet unknown mechanisms. Respiratory mucosal epithelia, including those of the middle ear and eustachian tube, secrete antimicrobial effectors including lysozyme, lactoferrin and beta defensins-1 and -2. To elucidate the role of these innate immune molecules in the normal defense and maintenance of sterility of respiratory mucosa such as that of the middle ear, we assessed their effect on the respiratory pathogens nontypeable Haemophilus influenzae (NTHi) 12, Moraxella catarrhalis 035E, and Streptococcus pneumoniae 3, and 6B. METHODS: Two assay methods, the radial assay and the liquid broth assay, were employed for testing the antimicrobial activity of the molecules. This was done in order to minimize the possibility that the observed effects were artifacts of any single assay system employed. Also, transmission electron microscopy (TEM) was employed to evaluate the effect of antimicrobial innate immune molecules on OM pathogens. For the statistical analysis of the data, Student's t-test was performed. RESULTS: Results of the radial diffusion assay showed that beta defensin-2 was active against all four OM pathogens tested, while treatment with beta defensin-1 appeared to only affect M. catarrhalis. The radial assay results also showed that lysozyme was quite effective against S. pneumoniae 3 and 6B and was partially bacteriostatic/bactericidal against M. catarrhalis. Lysozyme however, appeared not to affect the growth of NTHi. Thus, lysozyme seems to have a more pronounced impact on the growth of the Gram-positive S. pneumoniae as compared to that of Gram-negative pathogens. Lactoferrin on the other hand, enhanced the growth of the bacteria tested. The results of the radial assays were confirmed using liquid broth assays for antimicrobial activity, and showed that lysozyme and beta defensin-2 could act synergistically against S. pneumoniae 6B. Moreover, in the liquid broth assay, beta defensin-1 showed a modest inhibitory effect on the growth of S. pneumoniae 6B. As assessed by ultrastructural analysis, lysozyme and beta defensin-2, and to a much lesser extent, beta defensin-1, appeared to be able to cause damage to the bacterial membranes. CONCLUSIONS: Here we report that lysozyme and the beta defensins can inhibit the growth of clinical isolates of otitis media pathogens - namely NTHi strain 12, S. pneumoniae strains 3 and 6B and M. catarrhalis strain 035E - and cause ultrastructural damage to these pathogens. Moreover, we demonstrate that lysozyme and beta defensin-2 can act synergistically against S. pneumoniae. These findings are consistent with the concept that secreted antimicrobial peptides and other components of innate immunity constitute the first line of defense protecting host mucosal surfaces, including the tubotympanal (eustachian tube and middle ear cavity) mucosa, against pathogens.

Anti-Infective Agents↗
PubMed1999

Acute laryngitis in the rat induced by Moraxella catarrhalis and Bordetella pertussis: number of neutrophils, dendritic cells, and T and B lymphocytes accumulating during infection in the laryngeal mucosa strongly differs in adjacent locations.

Infectious laryngotracheitis results in fulminant respiratory distress. During the disease, the subglottic mucosa is selectively infected and swollen, the reason for this preference being unknown. Therefore, in the present study the immunoreaction of the laryngeal mucosa was studied in the rat after inhalation of either heat-killed Moraxella catarrhalis (PVG rats) or application of viable Bordetella pertussis (BN rats). The number of neutrophils, macrophages, dendritic cells, and T and B lymphocytes was determined in the mucosa of the supraglottic, glottic, and subglottic area of the larynx as well as in the trachea. After application of the pathogens, the mucosa of the subglottic area was significantly more affected than the glottic mucosa. Already 1 h after application of M. catarrhalis, not only neutrophils but also dendritic cells and T and B lymphocytes were found both subepithelially and within the epithelium. They showed a similar kinetic progression, although at a different level. Two hours after application of M. catarrhalis, at the peak of inflammation, dendritic cells (173 +/- 10 cells/0.1 mm2) outnumbered neutrophils (54 +/- 9 cells/0.1 mm2), T lymphocytes (25 +/- 2 cells/0.1 mm2), and B lymphocytes (4.3 cells/0.1 mm2). The subglottic area (and the trachea) contained about three to five times more cells than the glottic area. In contrast, the number of local macrophages was lower in the subglottic area (24 +/- 5 cells/0.1 mm2) compared with that of the glottic area (38 +/- 6 cells/0.1 mm2), and did not change after application of both M. catarrhalis and B. pertussis. Thus, infectious laryngotracheitis in the rat closely resembles the clinical picture in children. In addition, the present results show a major difference in cellular influx in the mucosa of the glottic and subglottic area. This demonstrates that even in two closely adjacent locations, inflammatory responses of different magnitudes can occur, and it underlines the importance of regulatory mechanisms specific for the respective microenvironment.

Animals↗
PubMed1994

Prolonged occurrence of Moraxella bovis infection on a restricted heifer farm.

An epizootiologic study was conducted for infectious bovine keratoconjunctivitis (IBK) on a large grassland farm rearing a total of 1,300-1,400 heifers during May to October in both 1991 and 1992. Heifers were examined for infection with Moraxella bovis, and isolation of M. bovis from the affected eyes and from Asian face flies (Musca bezzii Patton et Cragg) swarming on the heifer face was carried out. During the observation period, 10.7% of the heifers in 1991 and 5.3% of the heifers in 1992 were affected with IBK. Eight M. bovis isolates were isolated from the ocular swabs of affected heifers with IBK for 2 years and identified by biochemical examinations and Southern DNA-DNA hybridization, but no M. bovis was obtained from 331 Asian face flies tested. Plasmid profile analysis showed that two plasmids (35-kb and 4.0-kb) were commonly found in all 8 M. bovis isolates. This indicates that M. bovis isolates with the same plasmid profile were widely disseminated on this farm for 2 years.

Animals↗
PubMed2004

Moraxella osloensis blood and catheter infections during anticancer chemotherapy: clinical and microbiologic studies of 10 cases.

Moraxella osloensis, a gram-negative bacterium that is saprophytic on skin and mucosa, rarely causes infections. Moreover, infections in patients with cancer have not been reported. We describe 10 cases of M. osloensis blood or catheter infections that occurred during anticancer chemotherapy with or without preexisting neutropenia. The organism was identified definitively by sequencing analysis of the 16S ribosomal RNA gene. Fever (up to 39.7 degrees C) with substantial neutrophilia characterized these infections. The infections were monomicrobic for 3 patients and polymicrobic for 7 patients. Nine patients acquired the infection through central venous catheter colonization. The likely sources of the organism were sinusitis (3 cases), bronchitis (1 case), presumed subclinical mucositis from anticancer therapy (4 cases), and cutaneous graft-vs-host disease (2 cases). The infections resolved, without catheter removal, after antibiotic therapy with cell wall-active agents, to which all strains were shown to be susceptible. The M. osloensis strains exhibited significant morphologic variations on gram stain, and sheep blood agar was the preferred culture medium for 9 strains.

Adult↗
PubMed1976

Pneumonitis and pulmonary abscess associated with Moraxella nonliquefaciens.

A 78-year-old man with pneumonitis and pulmonary abscess associated with Moraxella nonliquefaciens is presented. This organism was found by culture of both transtracheal aspirate and sputum. No previous reports have associated M nonliquefaciens with infection of the lower respiratory tract, although sinusitis and bronchitis have been reported. Possible predisposing factors in our patient included carcinoma of the larynx, as well as alcohol ingestion and cigarette smoking.

Aged↗
PubMed2005

Vaccine development for non-typeable Haemophilus influenzae and Moraxella catarrhalis: progress and challenges.

An urgent need exists for vaccines to prevent infections caused by nontypeable Haemophilus influenzae and Moraxella catarrhalis. These bacteria cause otitis media in children, a clinical problem associated with enormous morbidity and cost. H. influenzae and M. catarrhalis also cause lower respiratory tract infections in adults with chronic lung disease. Infections in this clinical setting are associated with disability and death. Recent progress in identifying potential vaccine antigens in both bacteria raises great promise in developing effective vaccines. This paper reviews the key issues in vaccine development for H. influenzae and M. catarrhalis, including areas where progress has been stalled, and proposes areas that deserve investigation in the next 5 years.

Animals↗
PubMed2006

Structure and immunological action of the human pathogen Moraxella catarrhalis IgD-binding protein.

Several pathogens have acquired the capacity to bind immunoglobulins in a nonimmune manner, that is, the binding does not involve the normal antigen-binding sites of the antibodies. In contrast to gram-positive bacteria, for example Staphylococus aureus, nonimmune binding to gram-negative bacteria is rare. Moraxella catarrhalis outer membrane protein MID is the first to date known IgD-binding protein. MID is a 200-kDa autotransporter protein that exists as an oligomer and is governed at the transcriptional level. The majority of M. catarrhalis clinical isolates expresses MID. Two functional domains have been attributed to MID. MID764-913 functions as an adhesin and promotes the bacteria to attach to epithelial cells. The IgD-binding domain is located within MID962-1200 and the IgD-binding is related to the secondary and tertiary structure, that is, an oligomer is required for an optimal interaction. In parallel, M. catarrhalis activates B lymphocytes through the IgD B-cell receptor. This stimulatory capacity can be blocked by anti-IgD polyclonal antibodies, and M. catarrhalis mutants devoid of MID do not stimulate B cells. Moreover, MID and MID962-1200 activates B lymphocytes in the presence of T-helper 2 cytokines or soluble CD40L. Thus, available data suggest that MID is a T-cell-independent antigen.

Adhesins, Bacterial↗
PubMed1996

The incidence of respiratory tract pathogens and antimicrobial susceptibilities of Streptococcus pneumoniae, Haemophilus influenzae and Moraxella (Branhamella) catarrhalis isolated between 1990 and 1993.

Using a quantitative culture of sputum, the incidence of pathogenic bacteria in respiratory infection in our laboratory between 1990 and 1993 were investigated. While Haemophilus influenzae, Streptococcus pneumoniae and Moraxella (Branhamella) catarrhalis were isolated at high rates (67-78%) from the specimens of outpatients throughout the study period, the incidence of S pneumoniae has increased gradually. The antimicrobial susceptibilities of these three pathogens were examined with the agar dilution method. A marked increase of penicillin (PC) resistant S. pneumoniae (MIC > or = 0.1 microgram/ml) was observed with a resistance rate of 2.1% in 1990 and 25% in 1993. Resistance to erythromycin (EM, MIC > or = 1.56 micrograms/ml) was 8.5% in 1990 but then increased to 34% in 1992. Most of the PC resistant isolates were resistant to multidrugs such as EM, minocycline and clindamycin. The MICs of all beta-lactams examined for S. pneumoniae increased along with the MICs of PC, though the level varied between drugs. The rates of beta-lactamase positive H. influenzae gradually decreased, being 14.3% in 1990 and 7.4% in 1993, whereas those of M. (B) catarrhalis were consistently high (> 90%) every year. In addition to beta-lactamase production, the emergence of strains of H. influenzae and M. (B) catarrhalis resistant to new quinolone drugs should be noted.

Drug Resistance, Microbial↗
PubMed2001

Cloning and characterization of a Moraxella bovis cytotoxin gene.

OBJECTIVE: To identify the Moraxella bovis cytotoxin gene. PROCEDURE: Hemolytic and nonhemolytic strains of M. bovis were compared by use of western blotting to identify proteins unique to hemolytic strains. Oligonucleotide primers, designed on the basis of amino acid sequences of 2 tryptic peptides derived from 1 such protein and conserved regions of the C and B genes from members of the repeats in the structural toxin (RTX) family of bacterial toxins, were used to amplify cytotoxin-specific genes from M. bovis genomic DNA. Recombinant proteins were expressed, and antisera against these proteins were produced in rabbits. RESULTS: Several proteins ranging in molecular mass from 55 to 75 kd were unique to the hemolytic strain. An open reading frame encoding a 927-amino acid protein with a predicted molecular mass of 98.8 kd was amplified from M. bovis genomic DNA. The deduced amino acid sequence encoded by this open reading frame was homologous to RTX toxins. Antisera against the recombinant carboxy terminus encoded by this open reading frame neutralized hemolytic and cytolytic activities of native M. bovis cytotoxin. CONCLUSIONS AND CLINICAL RELEVANCE: A gene was identified in M bovis that encodes a protein with sequence homology to other RTX toxins. Results of cytotoxin neutralization assays support the hypothesis that M. bovis cytotoxin is encoded by this gene and belongs in the RTX family of bacterial exoproteins. Identification of this gene and expression of recombinant cytotoxin could facilitate the development of improved vaccines against infectious bovine keratoconjunctivitis.

Amino Acid Sequence↗
PubMed2001

Detection of shared magnetic antigenic determinants on whole Moraxella bovis pili by use of antisera to cyanogen bromide-cleaved M. bovis pilus protein.

OBJECTIVE: To determine the ability of antisera against cyanogen bromide-cleaved pili from 4 strains of Moraxella bovis to react with whole or nondenatured pili. SAMPLE POPULATION: Antisera to 4 strains of M. bovis produced by New Zealand White rabbits. PROCEDURE: Pili from 4 strains of M. bovis were collected and purified. Pilus proteins (pilin) were cleaved, using cyanogen bromide. Whole pilus and cyanogen bromide-cleaved pilin were injected into rabbits. Antisera were serially diluted, reacted with 4 strains of M. bovis, and examined by immunoelectron microscopy and indirect immunofluorescence. RESULTS: Antisera to whole pili aggregated and distorted pili from homologous strains, but pili from heterologous strains were unaffected. Antisera to cleaved pilin fragments resulted in partial aggregation and thickening of homologous and heterologous pili, suggestive of heterospecific antibodies. Attachment of antibodies to pili was detected by indirect immunofluorescence, indicating a strong reaction of antisera to whole pili with homologous pili. Weak cross-reactions were evident with certain heterologous strains. In contrast, antisera to cleaved pilin fragments reacted strongly with pili from homologous and heterologous strains. CONCLUSIONS AND CLINICAL RELEVANCE: We detected shared antigenic determinants on pili from various strains of M. bovis that were not immunogenic in intact pili. These sites were immunogenic after cleavage of pilus protein with cyanogen bromide, and antisera produced to protein fragments reacted with whole pili from heterologous strains of the organism. Vaccines produced from cyanogen bromide-treated pili may induce broader immunity against infectious bovine keratoconjuctivitis than that provided by currently available vaccines.

Animals↗
PubMed2004

Stability, antigenicity, and aggregation of Moraxella bovis cytolysin after purification and storage.

OBJECTIVES: To compare stability, antigenicity, and aggregation characteristics of Moraxella bovis cytolysins among isolates from geographically diverse areas. STUDY POPULATION: 8 isolates of M. bovis. PROCEDURE: Filter-sterilized broth culture supernatants of M. bovis were concentrated, diafiltered, and chromatographed. The endotoxin and cytolysin activities in samples were measured. Chromatographed cytolysins of M. bovis were examined by immunoblotting. Hemolytic and leukotoxic activities were measured from samples collected at each step of purification and before and after storage. Hemolysis was measured directly by use of washed bovine erythrocyte targets. Leukotoxicity was measured by use of a 51Cr release assay. RESULTS: Cytolysin was retained by a filter with 100-kd nominal molecular weight limit. Hemolytic activity, leukotoxic activity, and endotoxin were eluted together in void volume of a gel-filtration column (molecular mass exclusion limit = 4 X 10(7) d). Gel-column chromatographed diafiltered retentate had the greatest specific cytolytic activity and the highest endotoxin-to-protein ratio. Frozen diafiltered retentate(-80 degrees C, 4 months) was cytolytic after thawing. Immunoblots of gel-column chromatographed cytolysin contained 4 proteins with molecular masses between 90 and 68 kd. Fractions with high lytic activities also had additional protein bands with molecular masses of 98 and 63 kd. Immunoblots of gel-column chromatographed diafiltered retentate revealed proteins with molecular masses between 90 and 68 kd. CONCLUSIONS AND CLINICAL RELEVANCE: Diafiltered M. bovis cytolysin is aggregated with endotoxin. Antigenicity and cytolytic activities in diafiltered retentate are conserved among M. bovis isolates. Diafiltration could be useful for bulk semipurification of M. bovis cytolysin. Cytolysin-enriched vaccines of M. bovis could be contaminated by endotoxin.

Analysis of Variance↗
PubMed2003

Moraxella catarrhalis induces mast cell activation and nuclear factor kappa B-dependent cytokine synthesis.

Human mast cells are often found perivascularly and at mucosal sites and may play crucial roles in the inflammatory response. Recent studies have suggested a prominent role for mast cells in host defense. In this study, we analyzed the effects of a common airway pathogen, Moraxella catarrhalis and a commensal bacterium, Neiserria cinerea, on activation of human mast cells. Human mast cell leukemia cells (HMC-1) were activated with either phorbol myristate acetate (PMA) and calcium ionophore or with varying concentrations of heat-killed suspensions of bacteria. Supernatants were assayed for the cytokines interleukin-4 (IL-4), granulocyte macrophage colony stimulating factor (GM-CSF), IL-6, IL-8, IL-13 and monocyte chemotactic protein-1 (MCP-1). Nuclear proteins were isolated and assayed by electrophoretic mobility shift assay (EMSA) for nuclear factor kappaB (NF-kappaB) nuclear binding activity. In some experiments, NF-kappaB inhibitor, Bay-11 was added to determine functional significance. Both M. catarrhalis and N. cinerea induced mast cell activation and selective secretion of two key inflammatory cytokines, IL-6 and MCP-1. This was accompanied by NF-kappaB activation. Neither spun bacterial supernatants nor bacterial lipopolysaccharide induced cytokine secretion, suggesting need for direct bacterial contact with mast cells. Scanning electron microscopy revealed active aggregation of bacteria over mast cell surfaces. The NF-kappaB inhibitor, Bay-11, inhibited expression of MCP-1. These findings suggest the possibility of direct interactions between human mast cells and common bacteria and provide evidence for a novel role for human mast cells in innate immunity.

Cell Line, Tumor↗
PubMed1994

Moraxella (Branhamella) catarrhalis-induced experimental otitis media in the chinchilla.

Otitis media was introduced in chinchillas by direct transbullar inoculation with either viable or formalin-inactivated Moraxella (Branhamella) catarrhalis. Both groups of animals developed middle ear fluids (MEF) and severe inflammatory changes in the tubotympanum. Pure cultures of M. catarrhalis were recovered for up to 5 days after inoculation from those animals inoculated with viable bacteria. Significantly elevated anti-M. catarrhalis antibody titers were detected in post-inoculation sera and in MEF or bullar lavages, and an increased number of IgG-bearing cells was also observed in the tubotympanum of these animals. Control chinchillas inoculated with pyrogen-free sterile saline failed to show any signs of otitis media. Our data indicate that viable M. catarrhalis induce acute otitis media in the chinchilla but that the bacteria are rapidly eliminated from the middle ear, precluding its usefulness as a model to study acute otitis media. Moreover, the same concentration of nonviable organisms also induces severe inflammatory changes in the middle ear. These data indicate that the chinchilla may be a useful model for studying the role of nonviable M. catarrhalis or its cell wall components on the induction and persistence of inflammation in the middle ear.

Animals↗
PubMed1991

Mechanism of adherence of Moraxella (Branhamella) catarrhalis.

We examined the mechanisms of adherence of Moraxella catarrhalis to nasopharyngeal epithelial cells. Fimbriae were detected by electron microscopy on most of the strains studied. A role of fimbriae in adherence was supported by the reduction in adherence by treatments denaturing the fimbriae or by antifimbrial antibodies. There was, however, no significant difference in adhesive capacity or hemagglutination between fimbriated and non-fimbriated strains. Furthermore, there was no correlation between hemagglutination and adherence. The possibility that receptor epitopes were provided by cell surface glycolipids was examined by thin-layer chromatography. Glycolipids from various sources, including nasopharyngeal cells were separated by thin layer chromatography plates and overlayed with bacteria. No binding was detected. The results suggest that lectin-glycolipid interactions do not explain the attachment of M. catarrhalis to epithelial cells.

Adult↗
PubMed1993

Erythromycin treatment is beneficial for longstanding Moraxella catarrhalis associated cough in children.

The benefits of antibiotic treatment and a nasopharyngeal culture in children with longstanding cough were analysed in a prospective randomized open study. Clinically suspected pertussis was excluded. Of 40 children given erythromycin for 7 days, 35 (88%) recovered in one week, compared with 17/47 (36%) untreated (p < 0.0001). Erythromycin eliminated Moraxella catarrhalis from the nasopharynx in 21/31 children (68%), compared with spontaneous disappearance in 7/35 (20%) untreated controls (p < 0.001). Purulent bronchitis or otitis media occurred in 2 children (5%) in the treatment group and in 21 (45%) in the control group (p < 0.01). To evaluate the clinical role of isolated pathogens, the 47 untreated subjects were studied. Seven of 35 children harbouring M. catarrhalis recovered, compared with 8/12 in whom this bacterium was absent (p < 0.01). No correlation was found between the isolation of Haemophilus influenzae or Streptococcus pneumoniae and the clinical outcome. Children with persistent cough > 10 days may benefit from erythromycin treatment. M. catarrhalis in the nasopharynx indicates prolonged symptoms and increased risk of bacterial complications.

Child↗
PubMed1994

Moraxella catarrhalis--an uncommon cause of community-acquired pneumonia in Swedish children.

In 284 Swedish children with community-acquired, roentgenologically verified pneumonia, antibodies to Moraxella (Branhamella) catarrhalis were determined in paired serum samples with an enzyme immunoassay using a whole-cell antigen preparation from 10 strains of M. catarrhalis. Only 9 children (3%) had significant increases in antibodies to M. catarrhalis. Among these 9 children, 11-39 months of age, 6 had serologic evidence of concurrent infection with other respiratory pathogens such as S. pneumoniae, non-capsulated H. influenza, RS virus and adenovirus. In 6 (67%) of the 9 children with antibody response and in 74 (27%) of the 275 children without antibody response to M. catarrhalis, nasopharyngeal cultures yielded growth of this bacterium. M. catarrhalis seems to be a common commensal in the upper respiratory tract, but a rare cause of pneumonia in children.

Antibodies, Bacterial↗
PubMed1994

Significance of isolation of Moraxella catarrhalis in routine cultures from the respiratory tract in adults: antibody response studied in a whole cell EIA.

The significance of the isolation of Moraxella catarrhalis from sputum or nasopharynx was studied in patients treated at an infectious disease clinic. A whole-cell enzyme immunoassay was used to detect a specific antibody response to M. catarrhalis during infection. In all, 27 patients with respiratory tract infections and 4 with other infections were studied. Titre rises were recorded in 11/23 patients with lower respiratory tract infections, whereas patients with common cold or infections elsewhere all had negative serology. In patients with acute bronchitis, 7/10 patients responded with a significant titre rise. Patients with a low titre in their acute serum sample were those who responded with a titre increase during infection. The findings indicate that isolation of M. catarrhalis from sputa and nasopharyngeal samples in adults is of value for the etiological diagnosis of acute bronchitis and other lower respiratory tract infections, and is therefore important for the choice of drug for treatment, as many isolates are resistant to beta-lactam antibiotics.

Adolescent↗