Functional Characteristics of a Protective Monoclonal Antibody against Serotype A and C Lipooligosaccharides from
            <i>Moraxella catarrhalis</i>

Hu, Weigang; Chen, Jing; McMichael, John; Gu, Xin-Xing

doi:10.1128/iai.69.3.1358-1363.2001

Cited by 39 publications

(35 citation statements)

References 37 publications

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“…Fitzgerald et al showed that the bacterial adherence of M. catarrhalis might be mediated by carbohydrate moieties (12). Our previous study revealed that a specific anti-LOS monoclonal antibody significantly inhibits the adherence of M. catarrhalis to human epithelial cells (28). In this report, attachment to human epithelial cells by the kdtA mutant showed more than a 50% reduction in attachment to Chang or cervix epithelial cells while there was no reduction in attachment to lung epithelial cells.…”

Section: Discussionmentioning

confidence: 66%

“…The OMV preparation was also applied since there was no detectable LOS in O35EkdtA whole-cell lysates. Western blot using a rabbit anti-LOS antibody was performed (28). For the composition analysis, 30 to 35 g of wet cells from O35E and O35EkdtA were prepared for LOS purification by phenol-water extraction (21) and then the phenol-chloroform-petroleum ether (PCP) method (17), due to the lack of LOS extracts from O35EkdtA.…”

Section: Methodsmentioning

confidence: 99%

“…Bacteria were suspended on a slide in a small amount of water (28). Formvar-coated 200 mesh grids (Electron Microscopy Sciences, Fort Washington, PA) were floated on the bacterial suspension for 1 min, blotted dry, and then floated on a mixture of equal volumes of 2% ammonium acetate (Mallinckrodt Chemical, Inc., Paris, KY) and 2% ammonium molybdate (Sigma).…”

Section: Methodsmentioning

confidence: 99%

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Roles of 3-Deoxy- d - manno -2-Octulosonic Acid Transferase from Moraxella catarrhalis in Lipooligosaccharide Biosynthesis and Virulence

et al. 2005

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Lipooligosaccharide (LOS), a major outer membrane component of Moraxella catarrhalis, is a possible virulence factor in the pathogenesis of human infections caused by the organism. However, information about the roles of the oligosaccharide chain from LOS in bacterial infection remains limited. Here, a kdtA gene encoding 3-deoxy-D-manno-2-octulosonic acid (Kdo) transferase, which is responsible for adding Kdo residues to the lipid A portion of the LOS, was identified by transposon mutagenesis and construction of an isogenic kdtA mutant in strain O35E. The resulting O35EkdtA mutant produced only lipid A without any core oligosaccharide, and it was viable. Physicochemical and biological analysis revealed that the mutant was susceptible to hydrophobic reagents and a hydrophilic glycopeptide and was sensitive to bactericidal activity of normal human serum. Importantly, the mutant showed decreased toxicity by the Limulus amebocyte lysate assay, reduced adherence to human epithelial cells, and enhanced clearance in lungs and nasopharynx in a mouse aerosol challenge model. These data suggest that the oligosaccharide moiety of the LOS is important for the biological activity of the LOS and the virulence capability of the bacteria in vitro and in vivo. This study may bring new insights into novel vaccines or therapeutic interventions against M. catarrhalis infections.Moraxella catarrhalis, a gram-negative diplococcus, is now considered to be an important human respiratory tract pathogen in children and adults (6,29). In children, it causes otitis media, the most common childhood infection and the leading cause of conductive hearing loss, while in adults it exacerbates chronic obstructive pulmonary diseases, the fourth leading cause of death in the United States. Sporadic cases of conjunctivitis, meningitis, endocarditis, ophthalmia neonatorum, keratitis, urethritis, peritonitis, and septicemia have also been reported in individuals with reduced immune defense (36). In addition, the number of antibiotic-resistant strains of M. catarrhalis has significantly increased over the past decades (3, 23). Currently, the molecular pathogenesis of M. catarrhalis infection is not fully understood. Based on limited information about this organism and other respiratory tract pathogens, it is generally believed that it has several virulence factors involved in colonization, adherence, or complement resistance that enable the organism to evade the normal host defense and to establish the infection (47).Previous studies suggested that complement resistance involving multifactorial processes might be an important mechanism of M. catarrhalis virulence. Several M. catarrhalis isogenic mutants of UspA2 (1, 30), Fur (15), CopB (22), and outer membrane protein (OMP) E (37) showed their susceptibility to the bactericidal activity of normal human serum when compared with their parental strains. This indicates that these bacterial components may play critical roles for the bacterial resistance to killing caused by normal human serum. In addition, ...

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Section: Discussionmentioning

confidence: 66%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Roles of 3-Deoxy- d - manno -2-Octulosonic Acid Transferase from Moraxella catarrhalis in Lipooligosaccharide Biosynthesis and Virulence

et al. 2005

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“…Previous studies have suggested that LOS is important for the pathogenesis of other respiratory pathogens, such as Neisseria meningitidis and Haemophilus influenzae, aiding in adherence of the bacterium to the mucosal epithelium and in the destruction of the mucosal barrier (16,30,51,52). Some studies have demonstrated that the LOS of M. catarrhalis may also aid in adherence of this bacterium to epithelial cells, while other studies have indicated that LOS may elicit antibody production, suggesting that this major glycolipid has potential as a vaccine candidate (2,23,24,27).Previous studies using polyclonal antisera and structural analyses have identified three major LOS serotypes for M. catarrhalis (7-9, 21, 55). Clinical isolates were grouped into LOS serotypes A (60%), B (30%), and C (5%), with 5% of the strains unidentified (21,26,55).…”

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confidence: 99%

Characterization of a Cluster of Three Glycosyltransferase Enzymes Essential for Moraxella catarrhalis Lipooligosaccharide Assembly

Edwards¹,

Allen

Gibson

et al. 2005

J Bacteriol

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Moraxella catarrhalis isolates express lipooligosaccharide (LOS) molecules on their surface, which share epitopes similar to that of the Neisseria and Haemophilus species. These common LOS epitopes have been implicated in various steps of pathogenesis for the different organisms. In this study, a cluster of three LOS glycosyltransferase genes (lgt) were identified in M. catarrhalis 7169, a strain that produces a serotype B LOS. Mutants in these glycosyltransferase genes were constructed, and the resulting LOS phenotypes were consistent with varying degrees of truncation compared to wild-type LOS. The LOS structures of each lgt mutant were no longer detected by a monoclonal antibody (MAb 4G5) specific to a highly conserved terminal epitope nor by a monoclonal antibody (MAb 3F7) specific to the serotype B LOS side chain. Mass spectrometry of the LOS glycoforms assembled by two of these lgt mutants indicated that lgt1 encodes an ␣(1-2) glucosyltransferase and the lgt2 encodes a ␤(1-4) galactosyltransferase. However, these structural studies could not delineate the function for lgt3. Therefore, M. catarrhalis lgt3 was introduced into a defined ␤(1-4) glucosyltransferase Haemophilus ducreyi 35000glu؊ mutant in trans, and monoclonal antibody analysis confirmed that Lgt3 complemented the LOS defect. These data suggest that lgt3 encodes a glucosyltransferase involved in the addition of a ␤(1-4)-linked glucose to the inner core. Furthermore, we conclude that this enzymatic step is essential for the assembly of the complete LOS glycoform expressed by M. catarrhalis 7169.Moraxella catarrhalis is a gram-negative human respiratory pathogen that causes 15 to 20% of acute otitis media in children (56). In addition, this bacterium is responsible for 10 to 35% of lower respiratory infections in adults with chronic obstructive pulmonary disease, the fourth leading cause of death in the United States (20). The emergence of M. catarrhalis as an important human pathogen has occurred in the past 15 years due to the increasing prevalence of ␤-lactamase-positive strains, the high incidence of recurrent infections despite successful antibiotic treatment, and the lack of an effective vaccine (10,39,50,56). Another factor that likely contributes to the persistence of M. catarrhalis disease is the lack of understanding of the basic bacterial factors and mechanisms that promote colonization and survival in the host.Although there have been a number of putative virulence factors described for M. catarrhalis, the actual role of these components in pathogenesis remains largely undefined (13,19,20,31,53). One of the most prominent surface components expressed in the outer membrane of this bacterium is the lipooligosaccharide (LOS) (12). The LOS of M. catarrhalis is similar to the lipopolysaccharide (LPS) of other gram-negative organisms, but it lacks a repeating O antigen (21). Previous studies have suggested that LOS is important for the pathogenesis of other respiratory pathogens, such as Neisseria meningitidis and Haemophilus influenzae, aid...

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“…For many bacteria, adherence to epithelial surfaces contributes to the evasion of host clearance mechanisms (4,25,59). Previous studies with M. catarrhalis have identified UspA1 (32, 36), UspA2 (2, 8, 36), Hag (14,15,17,19,24,45,47; M. M. Holm and E. R. Lafontaine, unpublished data), lipooligosaccharide (24), OMPCD (49), and UspA2H (32) as potential adhesins. Of these, only UspA1 and UspA2H have been directly shown to mediate adherence to human cells (32,36).…”

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confidence: 99%

Identification of aMoraxella catarrhalisOuter Membrane Protein Exhibiting Both Adhesin and Lipolytic Activities

Timpe¹,

Holm²,

Vanlerberg³

et al. 2003

Infect Immun

116

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The UspA1 and Hag proteins have previously been shown to be involved in the ability of the Moraxella catarrhalis wild-type strain O35E to bind to human Chang and A549 cells, respectively. In an effort to identify novel adhesins, we generated a plasmid library of M. catarrhalis DNA fragments, which was introduced into a nonadherent Escherichia coli strain. Recombinant E. coli bacteria were subsequently enriched for clones that gained the ability to bind to Chang and A549 cells, yielding the plasmid pELFOS190. The unencapsulated, gram-negative bacterium Moraxella catarrhalis is one of the leading causes of otitis media in young children and of lower respiratory tract infections in adults with chronic obstructive pulmonary disease (COPD). In developed countries, more than 80% of children under the age of 3 years will be diagnosed at least once with otitis media, and M. catarrhalis is responsible for 15 to 25% of all of these cases (6, 7, 10, 12, 26-29, 41, 51, 60). Lower respiratory tract infections (exacerbations) have been shown to contribute to the progression of COPD, and of the approximately 20 million cases of exacerbations documented each year in the United States, up to 35% result from M. catarrhalis infections (3,41,43,54,55).A vaccine that provides protection against M. catarrhalis is highly desirable due to this substantial morbidity as well as the growing concern over antibiotic resistance observed in clinical isolates (30). Toward this end, current research is focused on the identification of potential antigens with emphasis on the outer membrane proteins (OMPs) (26,34,35). While M. catarrhalis OMPs with a wide variety of functions have been identified, adhesins are particularly attractive vaccine candidates because they are surface-exposed antigens. Furthermore, adhesins generally play a crucial role in colonization and pathogenesis. For many bacteria, adherence to epithelial surfaces contributes to the evasion of host clearance mechanisms (4,25,59). Previous studies with M. catarrhalis have identified UspA1 (32, 36), UspA2 (2, 8, 36), Hag (14,15,17,19,24,45,47; M. M. Holm and E. R. Lafontaine, unpublished data), lipooligosaccharide (24), OMPCD (49), and UspA2H (32) as potential adhesins. Of these, only UspA1 and UspA2H have been directly shown to mediate adherence to human cells (32,36). However, while UspA1 is expressed by virtually all strains tested to date, only ϳ20% of clinical isolates contain an uspA2H gene (32, 37).The present study describes the identification of the novel M. catarrhalis OMP McaP. While this protein was identified based on its ability to function as an adhesin, we report that it also displays the enzymatic activity of an esterase and a phospholipase B (PLB). MATERIALS AND METHODSStrains, plasmids, tissue culture (TC) cell lines, and growth conditions. The bacterial strains and plasmids described in this study are listed in Table 1. M. catarrhalis was routinely cultured at 37°C on Todd-Hewitt medium (Difco). When cultured on agar plates, M. catarrhalis was incubated in an at...

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Functional Characteristics of a Protective Monoclonal Antibody against Serotype A and C Lipooligosaccharides from Moraxella catarrhalis

Cited by 39 publications

References 37 publications

Roles of 3-Deoxy- d - manno -2-Octulosonic Acid Transferase from Moraxella catarrhalis in Lipooligosaccharide Biosynthesis and Virulence

Roles of 3-Deoxy- d - manno -2-Octulosonic Acid Transferase from Moraxella catarrhalis in Lipooligosaccharide Biosynthesis and Virulence

Characterization of a Cluster of Three Glycosyltransferase Enzymes Essential for Moraxella catarrhalis Lipooligosaccharide Assembly

Identification of aMoraxella catarrhalisOuter Membrane Protein Exhibiting Both Adhesin and Lipolytic Activities

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