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PubMed2005

Susceptibility patterns for amoxicillin/clavulanate tests mimicking the licensed formulations and pharmacokinetic relationships: do the MIC obtained with 2:1 ratio testing accurately reflect activity against beta-lactamase-producing strains of Haemophilus influenzae and Moraxella catarrhalis?

Amoxicillin/clavulanate has recently undergone formulation changes (XR and ES-600) that represent 14:1 and 16:1 ratios of amoxicillin/clavulanate. These ratios greatly differ from the 2:1 ratio used in initial formulations and in vitro susceptibility testing. The objective of this study was to determine if the reference method using a 2:1 ratio accurately reflects the susceptibility to the various clinically used amoxicillin/clavulanate formulations and their respective serum concentration ratios. A collection of 330 Haemophilus influenzae strains (300 beta-lactamase-positive and 30 beta-lactamase-negative) and 40 Moraxella catarrhalis strains (30 beta-lactamase-positive and 10 beta-lactamase-negative) were tested by the broth microdilution method against eight amoxicillin/clavulanate combinations (4:1, 5:1, 7:1, 9:1, 14:1, and 16:1 ratios; 0.5 and 2 microg/mL fixed clavulanate concentrations) and the minimum inhibitory concentration (MIC) results were compared with those obtained with the reference 2:1 ratio testing. For the beta-lactamase-negative strains of both genera, there was no demonstrable change in the MIC values obtained for all ratios analyzed (2:1 to 16:1). For the beta-lactamase-positive strains of H. influenzae and M. catarrhalis, at ratios >or=4:1 there was a shift in the central tendency of the MIC scatterplot compared with the results of testing 2:1 ratio. As a result, there was a 2-fold dilution increase in the MIC(50) and MIC(90) values, most evident for H. influenzae and BRO-1-producing M. catarrhalis strains. For beta-lactamase-positive strains of H. influenzae, the shift resulted in a change in the interpretive result for 3 isolates (1.0%) from susceptible using the reference method (2:1 ratio) to resistant (8/4 microg/mL; very major error) at the 16:1 ratio. In addition, the number of isolates with MIC values at or 1 dilution lower than the breakpoint (4/2 microg/mL) increased from 5% at 2:1 ratio to 32-33% for ratios 14:1 and 16:1. Our results indicate that, for the beta-lactamase-positive strains of H. influenzae and M. catarrhalis, the results of the amoxicillin/clavulanate reference 2:1 ratio testing do not accurately represent all the currently licensed formulations. Pharmacokinetic/pharmacodynamic (PK/PD) target attainment might be compromised when higher amoxicillin/clavulanate ratios are used clinically. With a better understanding of PK/PD parameters, reevaluation of the amoxicillin/clavulanate in vitro susceptibility testing should be considered by the standardizing authorities to reflect the licensed formulations and accurately predict clinical outcomes.

Amoxicillin-Potassium Clavulanate Combination↗
PubMed2006

Quantitative detection of Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis in lower respiratory tract samples by real-time PCR.

The limitation of polymerase chain reaction (PCR) in diagnosis of lower respiratory tract infections (LRTIs) caused by Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis has been a distinguishing colonization from infection. We assess here the usefulness of real-time quantitative PCR (RQ-PCR) performed on lower respiratory tract samples to overcome this problem. Consecutive respiratory tract samples from patients with and without signs of infection (n = 203) were subjected to RQ-PCR, targeting the genes pneumolysin (S. pneumoniae), fumarate reductase (H. influenzae), and outer membrane protein B (M . catarrhalis). DNA from positive controls with predefined colony forming units (CFUs) per milliliter were included to allow estimation of CFU per milliliter for the test samples. In parallel, assessment of quantitative cultures from all samples was performed. In the group of patients with LRTI, significant pathogens (>/=10(5) CFU/mL) were found in 32/135 samples (23.7%) with culture, in 51/135 (37.7%) with RQ-PCR, and in 59/135 (43.7%) when combining the methods.

Bronchoalveolar Lavage Fluid↗
PubMed2004

Exploration of Moraxella catarrhalis outer membrane proteins, CD and UspA, as new carriers for lipooligosaccharide-based conjugates.

Moraxella catarrhalis outer membrane proteins, CD and ubiquitous surface protein A (UspA), were used as carriers for M. catarrhalis detoxified lipooligosaccharide (dLOS)-based conjugates. Our study was designed to investigate the feasibility of CD and UspA as protein carriers for dLOS-based conjugates and their possible synergic effects on protection from both anti-LOS and anti-CD or anti-UspA antibody responses. Female Balb/c mice were immunized subcutaneously three times with dLOS-CD or dLOS-UspA conjugate in Ribi adjuvant. Antisera elicited by the conjugates showed high titers of specific anti-LOS antibodies with complement-dependent bactericidal activity towards M. catarrhalis strain 25238. In a mouse aerosol challenge model, mice immunized with both conjugates showed a significant enhancement of the clearance of strain 25238 from lungs as compared with the control mice. Although both conjugates elicited reduced (relative to unconjugated CD or UspA) but significant levels of anti-CD or UspA antibodies, they did not show synergetic effects with anti-LOS antibodies on the bactericidal activity or the pulmonary bacterial clearance. Nevertheless, CD and UspA are safe and effective new carriers for dLOS-based or other potential carbohydrate-based conjugate vaccines to help thymus-independent carbohydrate antigens for production of anti-carbohydrate antibodies against target pathogens.

Animals↗
PubMed2005

Moraxella catarrhalis is only a weak activator of the mannose-binding lectin (MBL) pathway of complement activation.

A hemolytic bystander assay was used to assess the functional serum mannose-binding lectin (MBL) activating capacity of five isolates of Moraxella catarrhalis obtained from children who suffered recurrent acute otitis media episodes. Results showed that this organism is only a poor activator of the lectin pathway of complement activation, with subsequent consequences for the etiology of otitis media by this organism.

Child↗
PubMed2004

Antimicrobial susceptibility of Haemophilus influenzae, Haemophilus parainfluenzae and Moraxella catarrhalis isolated from adult patients with respiratory tract infections in four southern European countries. The ARISE project.

Over a 7-month period in 2000-2001, 1213 Haemophilus influenzae, 112 Haemophilus parainfluenzae and 142 Moraxella catarrhalis isolates were recovered from adult patients with respiratory tract infections. Patients were from four southern European countries (Spain, Italy, Portugal and Greece). The antimicrobial susceptibility of the isolates to 11 antibiotics was determined in a central laboratory. The most active drugs on the basis of MICs were levofloxacin, cefditoren, cefotaxime, cefpodoxime and amoxicillin/clavulanate. MICs > or = 2 mg/l for amoxicillin were found in 19.5, 28.6, and 75.4% of H. influenzae, H. parainfluenzae and M. catarrhalis isolates, respectively. Isolates of H. influenzae and H. parainfluenzae with reduced susceptibility or that were fully resistant to amoxicillin/clavulanate, cefuroxime and clarithromycin were detected (0.2-1.8%) as well as M. catarrhalis resistant to clarithromycin (0.7%). Regular surveys of resistance patterns for antimicrobial agents are necessary.

Adult↗
PubMed2005

Antimicrobial resistance in Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis and group A beta-haemolytic streptococci in 2002-2003. Results of the multinational GRASP Surveillance Program.

A multinational surveillance study, GRASP, was conducted between November 2002 and April 2003 with the aim of assessing rates of antimicrobial resistance among 2656 isolates of Streptococcus pneumoniae, 2486 isolates of group A beta-haemolytic streptococci, 1358 isolates of Haemophilus influenzae and 1047 of Moraxella catarrhalis from 20 countries in Europe, eastern Asia and southern Africa. Conspicuous differences between various countries were noted in the S. pneumoniae resistance rates observed for penicillin (0-79.2%) and erythromycin (4-66%), along with other antimicrobials. The percentage of MDR strains was above 25% in 8 of the 20 countries studied. Group A streptococcal macrolide resistance rates ranged from 0% to 35% by country, while rates of beta-lactamase production ranged from 0% to 39% for H. influenzae and 80-100% for M. catarrhalis. Antibiotic resistance in S. pneumoniae remains a significant problem world wide.

Adult↗
PubMed2005

Molecular analysis of intrafamiliar transmission of Moraxella catarrhalis.

The possible intrafamiliar transmission of Moraxella catarrhalis was evaluated in 3 pairs between children and their parents, and 8 pairs between siblings from 11 families. Of the 22 isolates, all were found producing beta-lactamase. Molecular typing by pulsed-field gel electrophoresis (PFGE) with Not I and Spe I showed that the PFGE patterns in 2 of 3 pairs between children and their parents, and 4 of 8 pairs between siblings were indistinguishable and those of the remaining pairs were different. These data indicate a possible high rate of intrafamiliar transmission of M. catarrhalis.

Adult↗
PubMed2004

A novel plasmid (pEMCJH03) isolated from moraxella catarrhalis possibly useful as a cloning and expression vector within this species.

A preliminary screening study of six Moraxella catarrhalis isolates from primary school children in the Netherlands identified a small 3.5 kb plasmid (pEMCJH03), containing four open reading frames, which encoded three mobilizing and one replicase protein. Insertion of a kanamycin containing transposon (yielding pEMCJH04) allowed selection and isolation of the plasmid in Escherichia coli. Natural transformation of pEMCJH04 into M. catarrhalis was successful for 25% (3/12) of non-isogenic isolates, with no link between (lack of) transformability and genetic lineage or (lack of) transformability and complement phenotype, though the transformation efficiency was found to be rather low at approximately 615CFU/microg (range=60-1040CFU/microg ). This is only the second publication detailing a plasmid isolated from this important respiratory pathogen, and the ability to clone and express foreign proteins in M. catarrhalis using pEMCJH04 could help in the development of a vaccine expression vector, as well as providing a useful tool for studying promoter activity and in complementation studies of gene knockout mutants.

Bacterial Proteins↗
PubMed2005

Development of a shuttle vector for Moraxella catarrhalis.

Efforts to perform genetic analysis in Moraxella catarrhalis have been hampered by the lack of a cloning vector. M. catarrhalis strain E22 was previously shown to contain plasmid pLQ510 which lacked a selectable antibiotic resistance marker. Several methods were used to eliminate unnecessary DNA from pLQ510. Then, a 1.2 kb spectinomycin resistance cartridge, a multiple cloning site, and the origin of replication from pACYC184 were cloned into this plasmid backbone to obtain the 7.2 kb plasmid pWW102B. This new plasmid could replicate in M. catarrhalis as well as in both Escherichia coli and Haemophilus influenzae. This shuttle vector was used to clone and express two different M. catarrhalis genes, respectively, encoding an adhesin and a protein involved in serum resistance. When these two plasmids were introduced into appropriate M. catarrhalis mutants, they complemented the phenotypic deficiency of each mutant. This is the first report of functional complementation in trans in this pathogen.

Adhesins, Bacterial↗
PubMed2006

Plasmid pWW115, a cloning vector for use with Moraxella catarrhalis.

The plasmid shuttle vector pWW102B is able to replicate in only a modest number of Moraxella catarrhalis strains. Plasmid pWW115, a spontaneous deletion mutant of pWW102B, was shown to lack both the pACYC184-derived origin of replication and the associated chloramphenicol-resistance gene but was able to replicate in every M. catarrhalis strain tested in this study, including one strain that had been previously refractory to all types of genetic manipulations. To test the utility of this plasmid, a M. catarrhalis gene encoding the UspA2 serum-resistance factor was cloned into pWW115 and the resultant recombinant plasmid was shown to confer serum-resistance on a serum-sensitive M. catarrhalis uspA2 mutant.

Bacterial Outer Membrane Proteins↗
PubMed2004

Neisseria meningitidis, Neisseria lactamica and Moraxella catarrhalis share cross-reactive carbohydrate antigens.

Carriage of commensal bacteria species is associated with the development of natural immunity to meningococcal disease, with lipo-oligosaccharides (LOS) of meningococci being one of the main virulence factors associated with severity of meningococcal disease. Meningococcal reference strains and isolates from the commensal species Neisseria lactamica and Moraxella catarrhalis were assessed for the presence of cross-reactive glycoconjugate antigens. Binding of human blood group antibodies of the P and Ii system to meningococcal immunotype reference strains were in accordance with the presence of known LOS carbohydrate structures. Binding studies with meningococcal immunotyping antibodies and blood group phenotyping antibodies to N. lactamica strains from different European countries showed, that a greater number of isolates obtained from native Greek and Scottish adults and children bound anti-meningococcal L(3, 7, 9) immunotyping (P < 0.001), pK (P = 0.035) and paragloboside (P < 0.001) blood group typing antibodies compared to isolates obtained from children of Russian immigrants in Greece. A greater number of M. catarrhalis strains isolated from children in Scotland bound anti-L(3, 7, 9) antibodies (38.2%) compared to strains isolated from adults (22.2%) (P = 0.017). These findings provide evidence that blood group like glycoconjugate antigens found on the commensal species N. lactamica and M. catarrhalis might be involved in the development of natural immunity to meningococcal endotoxins during childhood, and might be exploited as anti-meningococcal vaccine candidates.

Adolescent↗
PubMed2004

Identification and characterization of outer membrane proteins G1a and G1b of Moraxella catarrhalis.

Moraxella catarrhalis is an important cause of otitis media, sinusitis, and lower respiratory tract infections in patients with chronic obstructive pulmonary disease. The purified outer membrane of M. catarrhalis contains a 29 kDa band, previously named outer membrane protein G1 (OMP G1). Polyclonal antiserum to the OMP G1 band was used to screen a genomic lambda phage library and the gene for OMP G1a was cloned and sequenced. Analysis of outer membrane by isoelectric focusing and amino-terminal protein sequence of the 29 kDa band revealed that the band is actually two individual proteins designated OMP G1a and OMP G1b. OMP G1a is a lipoprotein with an isoelectric point of 4. OMP G1b contains an unblocked amino-terminus and has an isoelectric point of 9. Analysis of the sequence of OMP G1a and OMP G1b from 25 clinical isolates revealed a high degree of conservation among strains. The sequence conservation of OMP G1a and OMP G1b among strains, combined with previous observations that OMP G1a and OMP G1b contain epitopes on the bacterial surface, indicate that OMP G1a and OMP G1b are potential vaccine antigens for M. catarrhalis.

Amino Acid Sequence↗
PubMed2004

Prevention of naturally occurring infectious bovine keratoconjunctivitis with a recombinant Moraxella bovis cytotoxin-ISCOM matrix adjuvanted vaccine.

The efficacy of a recombinant Moraxella bovis cytotoxin subunit vaccine to prevent naturally occurring infectious bovine keratoconjunctivitis (IBK) was evaluated in a randomized, blinded, controlled field trial. Ninety-three cross bred beef calves were vaccinated with either saline, ISCOM matrix (adjuvant control), or a recombinant M. bovis cytotoxin carboxy terminus peptide plus ISCOM matrix and boostered 21 days later. Ocular examinations were performed once weekly for 20 weeks. At week 12, the cumulative proportion of calves with ulcerated eyes in the recombinant vaccine group was significantly lower than in the saline control group. Throughout the 20 week trial, the cumulative proportion of ulcerated calves remained lowest in the recombinant vaccine group. By week 7, nonulcerated calves in the recombinant vaccine group had significantly higher changes in serum neutralizing titers and cytotoxin specific to total IgG ratios in serum and tears as compared to calves in the control groups. The trend for a reduced cumulative proportion of IBK in the vaccinated calves over the 20 week trial suggests that a recombinant M. bovis cytotoxin vaccine may be beneficial in helping to prevent naturally occurring IBK.

Animals↗
PubMed2005

Moraxella catarrhalis strains with reduced expression of the UspA outer membrane proteins belong to a distinct subpopulation.

The outer membrane proteins UspA1 and UspA2 are candidate antigens for a Moraxella catarrhalis vaccine. We previously reported that 103 of 108 isolates (95%) from young children expressed UspA1 detected by reactivity with the monoclonal antibody mAb24B5. The aim of the present study was to investigate mechanisms controlling UspA1 expression by analysis of five mAb24B5 non-reactive isolates. Four of these strains were characterized by (i) decreased or absent transcription of uspA1 and uspA2 and (ii) clustered mutations and deletions in the promoter region of both uspA1 and uspA2. Antigenic or phase variation were not responsible for reduced levels of UspA1 expression. While mAb24B5-positive isolates expressing normal levels of uspA1 and uspA2 mRNA belonged to the previously described 16S rRNA type 1 phylogenetic group, these four mAb24B5-negative isolates were found to belong to the 16S rRNA gene types 2 or 3. The remaining mAb24B5-negative isolate (#610) belonged to 16S rRNA type 1 and exhibited a posttranscriptional defect of UspA1 expression defined by normal levels of uspA1 mRNA and both recombinant and in vitro expression of mAb24B5-reactive UspA1. In conclusion, M. catarrhalis clinical isolates exhibiting reduced expression of UspA1 and UspA2 belonged to a distinct phylogenetic subpopulation. A UspA-based vaccine is unlikely to be effective against such isolates.

Amino Acid Sequence↗
PubMed2006

Filamentous-haemagglutinin-like protein genes encoded on a plasmid of Moraxella bovis.

The complete nucleotide sequence of a plasmid, pMBO-1, from Moraxella bovis strain Epp63 was determined. We identified 30 open reading frames (ORFs) encoded by the 44,215bp molecule. Two large ORFs, flpA and flpB, encoding proteins with similarity to Bordetella pertussis filamentous haemagglutinin (FHA), were identified on the same plasmid. The gene for a specific accessory protein (Fap), which may play a role in the secretion of Flp protein, was also identified. Reverse transcriptase PCR analysis of total RNA isolated from M. bovis Epp63 indicated that the flpA, flpB, and fap genes are all transcribed. Southern blot analysis indicated that the flp and fap genes are present in other clinical isolates of geographically diverse M. bovis.

Animals↗
PubMed1996

The structures of oligosaccharides isolated from the lipopolysaccharide of Moraxella catarrhalis serotype B, strain CCUG 3292.

The oligosaccharides from the lipopolysaccharides of Moraxella catarrhalis serotype B, strain CCUG 3292, were isolated after mild acid hydrolysis and separated by high-performance anion-exchange chromatography. The structures of the oligosaccharides were established by fast atom bombardment mass spectrometry and nuclear magnetic resonance spectroscopy. It is concluded that the oligosaccharides comprise a mixture of mainly a nona- and a deca-saccharide. [formula: see text] Smaller amounts of undeca-saccharides and of truncated forms, namely, hexa-, hepta-, and octa-saccharides, were also detected.

Carbohydrate Conformation↗
PubMed2002

beta-Lactamase-producing Moraxella catarrhalis may prevent the emergence of penicillin-resistant Streptococcus pneumoniae in children with recurrent acute otitis media.

We studied the effect of concomitant nasopharyngeal carriage of beta-lactamase producing Moraxella catarrhalis and Haemophilus influenzae on the occurrence of penicillin resistance of Streptococcus pneumoniae. We took nasopharyngeal samples from 306 children with recurrent otitis media and a history of several antibiotic treatments. We could isolate at least one of the pathogens in 89 subjects. Of these children 13% carried more than one pathogen. Of the isolated M. catarrhalis and H. influenzae strains 93% and 43% produced beta-lactamase, respectively. Of the S. pneumoniae strains 25% were non-susceptible (I/R) to penicillin. However, in patients carrying beta-lactamase-producing M. catarrhalis together with pneumococci all strains were susceptible to penicillin (P=0.0353). This finding suggests that beta-lactamase producing M. catarrhalis may hinder the emergence of penicillin resistance of S. pneumoniae in children with recurrent acute otitis media.

Acute Disease↗
PubMed2003

Acute mastoiditis caused by Moraxella catarrhalis.

Acute mastoiditis is the most frequent intratemporal complication of otitis media. The bacteriology of acute otitis media is changing continuously and it differs markedly from the bacteriology of acute mastoiditis. Moraxella catarrhalis (M. catarrhalis) is the third most common bacteria found in acute otitis media, and in recent years its importance as an etiological factor of acute otitis media has markedly increased in certain geographic areas. However, there are no reports of acute mastoiditis caused solely by M. catarrhalis. This report describes a case of a 2-year-old girl with acute mastoiditis and M. catarrhalis in the bacterial culture of middle ear effusion.

Acute Disease↗