Lipooligosaccharide P
            <sup>k</sup>
            (Galα1-4Galβ1-4Glc) Epitope of
            <i>Moraxella catarrhalis</i>
            Is a Factor in Resistance to Bactericidal Activity Mediated by Normal Human Serum

Zaleski, Anthony; Scheffler, N. Karoline; Densen, Peter; Lee, Frank K. N.; Campagnari, Anthony A.; Gibson, Bradford W.; Apicella, Michael A.

doi:10.1128/iai.68.9.5261-5268.2000

Cited by 49 publications

(47 citation statements)

References 58 publications

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“…Previous studies have shown that MAb 4G5 reacts to a conserved LOS epitope, containing the terminal portion of the main branch, expressed by all M. catarrhalis isolates tested (33,59). Western blot analysis demonstrated that MAb 3F7 reacts to an epitope that is specific to M. catarrhalis strains expressing serotype B LOS, suggesting that the side chain is important for antibody reactivity (data not shown).…”

Section: Vol 187 2005 Glycosyltransferase Enzymes Essential For Losmentioning

confidence: 97%

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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Section: Vol 187 2005 Glycosyltransferase Enzymes Essential For Losmentioning

confidence: 97%

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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“…This information is critical for elucidating the virulence functions of the LOS molecule. Previously, Zaleski et al identified a UDP-glucose-4-epimerase and showed that a corresponding knockout mutant resulted in a truncated LOS structure lacking two terminal galactosyl residues (50). Recently, Luke et al identified a kdsA gene coding for Kdo-8-phosphate synthase and found a kdsAdeficient mutant consisting only of lipid A in its LOS structure (32).…”

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confidence: 99%

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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“…A number of putative virulence factors have been described for M. catarrhalis (8,11,12,17,20), including a surface-exposed lipooligosaccharide (LOS) (6,18,23,32). Structural and serological studies with M. catarrhalis have described only three different LOS serotypes (termed A, B, and C), which vary in length and content of the oligosaccharide branches (3-5, 13, 15, 28).…”

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confidence: 99%

Multiplex PCR Assay That Identifies the Major Lipooligosaccharide Serotype Expressed by Moraxella catarrhalis Clinical Isolates

Edwards¹,

Schwingel²,

Datta

et al. 2005

J Clin Microbiol

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A heterologous cluster of glycosyltransferase genes was identified in the three Moraxella catarrhalis LOS serotype strains. Multiple PCR primers designed to this region amplified products that differentiate between the serotypes more rapidly and efficiently than previously described serological analyses. This assay will be valuable for clinical and research-based studies.Moraxella catarrhalis, a gram-negative diplococcus, is considered a significant cause of acute otitis media in children and lower respiratory infections in adults with chronic obstructive pulmonary disease (COPD) (12,21,29). A number of putative virulence factors have been described for M. catarrhalis (8,11,12,17,20), including a surface-exposed lipooligosaccharide (LOS) (6,18,23,32). Structural and serological studies with M. catarrhalis have described only three different LOS serotypes (termed A, B, and C), which vary in length and content of the oligosaccharide branches (3-5, 13, 15, 28). One serological study by Vaneechoutte et al. grouped clinical isolates into serotypes A (60%), B (30%), and C (5%), with 5% of the strains unidentified (28). That has been the only study to investigate the prevalence of specific M. catarrhalis LOS serotypes in the population. The difficulties with serological determinations of M. catarrhalis LOS expression are the limited quantities of antibodies, the absolute requirement for purified sample, and the potential for cross-reactivity between serotypes A and C (13,24,25).Recently, a cluster of three glycosyltransferase (lgt) genes were identified and characterized in a strain of M. catarrhalis 7169 expressing serotype B LOS (6). Primers 406 and 408 (Table 1) designed to this region were subsequently used in PCRs with chromosomal DNA from M. catarrhalis 25238, the previously defined LOS serotype A strain, and M. catarrhalis RS10, the previously defined LOS serotype C strain (Table 2) (3, 4). PCR was performed in 50-l reaction mixtures containing PCR SuperMix (Invitrogen, Carlsbad, CA), 20 pmol/l of each primer, and 1 l chromosomal DNA prepared as previously described (26). Amplifications were performed in a GeneAmp PCR system 9700 (Applied Biosystems, Foster City, CA) according to the manufacturer's protocol for 25 cycles with an annealing temperature of 53.1°C and extension time of 4 min. Primers 406 and 408 (Table 1) produced an amplicon of 4.3 kb in serotype A and C strains, which was 1 kb larger than the product amplified in the serotype B strain, 7169 (data not shown).Sequence analyses (MacVector 7.2 software; Accelrys, San Diego, CA) of the entire region amplified by primer 406 and flanking primer 704 (Table 1) identified an additional open reading frame upstream of the original lgt cluster described in 7169 and in the same orientation as lgt1, as illustrated in Fig. 1A and C. A ClustalW alignment with the translated sequence of this open reading frame (Lgt4) in both strains revealed 46% identity and 60% similarity to Lgt1, an ␣(1-2) glucosyltransferase in M. catarrhalis 7169 (6). The 5Ј end of this cluster...

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Lipooligosaccharide P ^k (Galα1-4Galβ1-4Glc) Epitope of Moraxella catarrhalis Is a Factor in Resistance to Bactericidal Activity Mediated by Normal Human Serum

Cited by 49 publications

References 58 publications

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

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

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

Multiplex PCR Assay That Identifies the Major Lipooligosaccharide Serotype Expressed by Moraxella catarrhalis Clinical Isolates

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Lipooligosaccharide P k (Galα1-4Galβ1-4Glc) Epitope of Moraxella catarrhalis Is a Factor in Resistance to Bactericidal Activity Mediated by Normal Human Serum

Cited by 49 publications

References 58 publications

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

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

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

Multiplex PCR Assay That Identifies the Major Lipooligosaccharide Serotype Expressed by Moraxella catarrhalis Clinical Isolates

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Resources

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Lipooligosaccharide P ^k (Galα1-4Galβ1-4Glc) Epitope of Moraxella catarrhalis Is a Factor in Resistance to Bactericidal Activity Mediated by Normal Human Serum