The Experts below are selected from a list of 2598 Experts worldwide ranked by ideXlab platform
M A Apicella - One of the best experts on this subject based on the ideXlab platform.
-
Nontypeable Haemophilus influenzae Lipooligosaccharide Expresses a Terminal Ketodeoxyoctanoate In Vivo, Which Can Be Used as a Target for Bactericidal Antibody
American Society for Microbiology, 2018Co-Authors: M A Apicella, Christopher J. Day, Jeremy Coffin, Margaret Ketterer, Deborah M. B. Post, Freda E.-c. Jen, Michael P. JenningsAbstract:Nontypeable Haemophilus influenzae (NTHi) is an important pathogen in individuals of all ages. The Lipooligosaccharide (LOS) of NTHi has evolved a complex structure that can be attributed to a multiplicity of glycosyltransferases, the random switching of glycosyltransferase gene expression via phase variation, and the complex structure of its core region with multiple glycoform branch points. This article adds to that complexity by describing a multifunctional enzyme (LsgB) which optimally functions when the species is grown on a solid surface and which can add either a ketodeoxyoctanoate (KDO) or an N-acetylneuramic acid (Neu5Ac) moiety to a terminal N-acetyllactosamine structure of LOS. Our studies show that expression of lsgB is reduced four- to sixfold when NTHi is grown in broth. The substrate that the enzyme utilizes is dependent upon the concentration of free Neu5Ac (between 1 and 10 µg/ml) in the environment. In environments in which Neu5Ac is below that level, the enzyme utilizes endogenous CMP-KDO as the substrate. Our studies show that during in vivo growth in an NTHi biofilm, the KDO moiety is expressed by the organism. Monoclonal antibody 6E4, which binds KDO, is bactericidal for NTHi strains that express the KDO epitope at high levels. In a survey of 33 NTHi strains isolated from healthy and diseased individuals, the antibody was bactericidal (>90% kill) for 12 strains (36%). These studies open up the possibility of using a KDO-based glycoconjugate vaccine as part of a multicomponent vaccine against NTHi.Nontypeable Haemophilus influenzae is an important pathogen in middle ear infections in children, sinusitis in adults, and acute bronchitis in individuals with chronic obstructive lung disease. The organism is very well adapted to the human host environment, and this has hindered successful development of an effective vaccine. In this article, we describe a mechanism by which the bacteria decorates its surface Lipooligosaccharide with a sugar unique to Gram-negative bacteria, ketodeoxyoctanoate (KDO). This sugar decoration is present during active infection and we have shown that an antibody directed against this sugar can result in killing of the organism. These data demonstrate that the Lipooligosaccharide ketodeoxyoctanoate epitope may be a novel NTHi-specific candidate vaccine antigen
-
REVIEW ARTICLES Nontypable Haemophilus influenzae: A Review of Clinical Aspects, Surface Antigens, and the Human Immune Response to Infection
2016Co-Authors: Timothy F Murphy, M A ApicellaAbstract:Nontypable Haemophilus influenzae has now become well established as an important pathogen in both adults and children. Recent work has identified clear distinctions be-tween nontypable and type b strains of H. influenzae. These organisms affect different patient populations, cause different infections, present different surface antigens to the host, and are genetically different. The commonest clinical manifestation of infection due to nontypable H. influenzae in adults is lower respiratory tract infection, particularly in the elderly and in those with chronic bronchitis. The bacterium is a frequent cause of acute otitis media in children. The surface of nontypable H. influenzae is composed of outer-membrane proteins and Lipooligosaccharide, and both ofthese demonstrate sub-stantial antigenic heterogeneity, which can be used to serotype isolates. Some respiratory tract isolates are fimbriated, but the role of fimbriae in pathogenesis is unclear. Antibod-ies to outer-membrane proteins and Lipooligosaccharide are present in human serum. In-vestigation of human immunity to infection is focusing on identification of those anti-gens to which protective antibody is directed. In the past several years, nontypable (nonencapsu
-
nontypeable haemophilus influenzae strain 2019 produces a biofilm containing n acetylneuraminic acid that may mimic sialylated o linked glycans
Infection and Immunity, 2004Co-Authors: L Greiner, Bradford W. Gibson, Nancy J. Phillips, Anthony Zaleski, Hiroshi Watanabe, Jianqiang Shao, A Morgan, M A ApicellaAbstract:Previous studies suggested that nontypeable Haemophilus influenzae (NTHI) can form biofilms during human and chinchilla middle ear infections. Microscopic analysis of a 5-day biofilm of NTHI strain 2019 grown in a continuous-flow chamber revealed that the biofilm had a diffuse matrix interlaced with multiple water channels. Our studies showed that biofilm production was significantly decreased when a chemically defined medium lacking N-acetylneuraminic acid (sialic acid) was used. Based on these observations, we examined mutations in seven NTHI strain 2019 genes involved in carbohydrate and Lipooligosaccharide biosynthesis. NTHI strain 2019 with mutations in the genes encoding CMP-N-acetylneuraminic acid synthetase (siaB), one of the three NTHI sialyltransferases (siaA), and the undecaprenyl-phosphate α-N-acetylglucosaminyltransferase homolog (wecA) produced significantly smaller amounts of biofilm. NTHI strain 2019 with mutations in genes encoding phosphoglucomutase (pgm), UDP-galactose-4-epimerase, and two other NTHI sialyltransferases (lic3A and lsgB) produced biofilms that were equivalent to or larger than the biofilms produced by the parent strain. The biofilm formed by the NTHI strain 2019pgm mutant was studied with Maackia amurensis fluorescein isothiocyanate (FITC)-conjugated and Sambucus nigra tetramethyl rhodamine isocyanate (TRITC)-conjugated lectins. S. nigra TRITC-conjugated lectin bound to this biofilm, while M. amurensis FITC-conjugated lectin did not. S. nigra TRITC-conjugated lectin binding was inhibited by incubation with α2,6-neuraminyllactose and by pretreatment of the biofilm with Vibrio cholerae neuraminidase. Matrix-assisted laser desorption ionization—time of flight mass spectometry analysis of Lipooligosaccharides isolated from a biofilm, the planktonic phase, and plate-grown organisms showed that the levels of most sialylated glycoforms were two- to fourfold greater when the Lipooligosaccharide was derived from planktonic or biofilm organisms. Our data indicate that NTHI strain 2019 produces a biofilm containing α2,6-linked sialic acid and that the sialic acid content of the Lipooligosaccharides increases concomitant with the transition of organisms to a biofilm form.
-
construction of acetate auxotrophs of neisseria meningitidis to study host meningococcal endotoxin interactions
Journal of Biological Chemistry, 2001Co-Authors: Peter C Giardina, M A Apicella, Theresa L Gioannini, Benjamin A Buscher, Anthony Zaleski, Deshang Zheng, Lynn L Stoll, Athmane Teghanemt, Jerrold WeissAbstract:Abstract To facilitate studies of the molecular determinants of host-meningococcal Lipooligosaccharide (endotoxin) interactions at patho-physiologically relevant endotoxin concentrations (i.e. ≤10 ng/ml), we have generated acetate auxotrophs NMBACE1 from encapsulated Neisseria meningitidis (serogroup B, strain NMB) and NMBACE2 from an isogenic bacterial mutant lacking the polysialic acid capsule. Growth of the auxotrophs in medium containing [14C]acetate yielded14C-Lipooligosaccharides containing ∼600 cpm/ng. Gel sieving resolved 14C-Lipooligosaccharide-containing aggregates with an estimated molecular mass of ≥20 × 106 Da (peak A) and ∼1 × 106 Da (peak B) from both strains. Lipooligosaccharides in peaks A and B had the same fatty acid composition and SDS-polyacrylamide gel electrophoresis profile. 14C-Labeled capsule copurified with 14C-Lipooligosaccharides in peak B from NMBACE1, whereas the other aggregates contained only14C-Lipooligosaccharide. For all aggregates, lipopolysaccharide-binding protein and soluble CD14-induced delivery of Lipooligosaccharides to endothelial cells and cell activation correlated with disaggregation of Lipooligosaccharides. These processes were inhibited by the presence of capsule but unaffected by the size of the aggregates. In contrast, endotoxin activation of cells containing membrane CD14 was unaffected by capsule but diminished when endotoxin was presented in larger aggregates. These findings demonstrate that the physical presentation of Lipooligosaccharide, including possible interactions with capsule, affect the ability of meningococcal endotoxin to interact with and activate specific host targets.
-
Gonococcal Lipooligosaccharide: an adhesin for bacterial dissemination? Response.
Trends in microbiology, 2000Co-Authors: M A ApicellaAbstract:Dr Nassif discusses gonococcal Lipooligosaccharide (LOS) and its role as an adhesin in bacterial dissemination. Our laboratory has shown that, in HepG2 cells and sperm cells, the terminal lactosamine present on lacto-N-neotetraose-terminal gonococcal LOS is a ligand for the human asialoglycoprotein receptor (ASGP-R) 1.xNeisseria gonorrhoeae utilizes and enhances the biosynthesis of the asialoglycoprotein receptor expressed on the surface of the hepatic HepG2 cell line. Porat, N. et al. Infect. Immun. 1995; 63: 1498–1506PubMedSee all References, 2.xGonococcal Lipooligosaccharide is a ligand for the asialoglycoprotein receptor on human sperm. Harvey, H.A. et al. Mol. Microbiol. 2000; 36: 1059–1070Crossref | PubMed | Scopus (56)See all References. We agree with Dr Nassif's assessment of the role of LOS in the transmission of gonorrhoea. In addition, we would like to expand the hypothesis that LOS plays a role in gonococcal pathogenesis to include adherence and invasion of host epithelial cells. Previous work from our laboratory with a primary human urethral epithelial cell tissue culture system showed that the ASGP-R is present on these cells 3xUltrastructural analysis of primary human urethral epithelial cell cultures infected with Neisseria gonorrhoeae. Harvey, H.A. et al. Infect. Immun. 1997; 65: 2420–2427PubMedSee all References3. Recent studies with primary urethral cells have pro-vided evidence that clathrin-dependent receptor-mediated endocytosis via interaction between gonoococcal LOS and the ASGP-R is a major route of gonococcal entry into these cells (M.A. Apicella, unpublished). Taken together, our findings — those discussed by Dr Nassif and our recent work — support the hypothesis that a specific interaction between the ASGP-R on human cells and the terminal galactose on lacto-N-neotetraose-terminal Neisseria gonorrhoeae LOS contributes to the pathogenesis and transmission of the disease gonorrhoea.
Bradford W. Gibson - One of the best experts on this subject based on the ideXlab platform.
-
nontypeable haemophilus influenzae strain 2019 produces a biofilm containing n acetylneuraminic acid that may mimic sialylated o linked glycans
Infection and Immunity, 2004Co-Authors: L Greiner, Bradford W. Gibson, Nancy J. Phillips, Anthony Zaleski, Hiroshi Watanabe, Jianqiang Shao, A Morgan, M A ApicellaAbstract:Previous studies suggested that nontypeable Haemophilus influenzae (NTHI) can form biofilms during human and chinchilla middle ear infections. Microscopic analysis of a 5-day biofilm of NTHI strain 2019 grown in a continuous-flow chamber revealed that the biofilm had a diffuse matrix interlaced with multiple water channels. Our studies showed that biofilm production was significantly decreased when a chemically defined medium lacking N-acetylneuraminic acid (sialic acid) was used. Based on these observations, we examined mutations in seven NTHI strain 2019 genes involved in carbohydrate and Lipooligosaccharide biosynthesis. NTHI strain 2019 with mutations in the genes encoding CMP-N-acetylneuraminic acid synthetase (siaB), one of the three NTHI sialyltransferases (siaA), and the undecaprenyl-phosphate α-N-acetylglucosaminyltransferase homolog (wecA) produced significantly smaller amounts of biofilm. NTHI strain 2019 with mutations in genes encoding phosphoglucomutase (pgm), UDP-galactose-4-epimerase, and two other NTHI sialyltransferases (lic3A and lsgB) produced biofilms that were equivalent to or larger than the biofilms produced by the parent strain. The biofilm formed by the NTHI strain 2019pgm mutant was studied with Maackia amurensis fluorescein isothiocyanate (FITC)-conjugated and Sambucus nigra tetramethyl rhodamine isocyanate (TRITC)-conjugated lectins. S. nigra TRITC-conjugated lectin bound to this biofilm, while M. amurensis FITC-conjugated lectin did not. S. nigra TRITC-conjugated lectin binding was inhibited by incubation with α2,6-neuraminyllactose and by pretreatment of the biofilm with Vibrio cholerae neuraminidase. Matrix-assisted laser desorption ionization—time of flight mass spectometry analysis of Lipooligosaccharides isolated from a biofilm, the planktonic phase, and plate-grown organisms showed that the levels of most sialylated glycoforms were two- to fourfold greater when the Lipooligosaccharide was derived from planktonic or biofilm organisms. Our data indicate that NTHI strain 2019 produces a biofilm containing α2,6-linked sialic acid and that the sialic acid content of the Lipooligosaccharides increases concomitant with the transition of organisms to a biofilm form.
-
metabolic incorporation of unnatural sialic acids into haemophilus ducreyi Lipooligosaccharides
Proceedings of the National Academy of Sciences of the United States of America, 2003Co-Authors: Scarlett Goon, Bradford W. Gibson, Birgit Schilling, Michael V Tullius, Carolyn R BertozziAbstract:The Lipooligosaccharides (LOS) of Haemophilus ducreyi are highly sialylated, a modification that has been implicated in resistance to host defense and in virulence. In previous work, we demonstrated that H. ducreyi scavenges sialic acid from the extracellular milieu and incorporates those residues into LOS. Here we report that H. ducreyi can use unnatural sialic acids bearing elongated N-acyl groups from three to seven carbon atoms in length, resulting in outer membrane presentation of unnatural sialyl-LOS. The unnatural variant comprises ≈90% of cell surface sialosides when exogenous substrates are added to the media at micromolar concentrations, despite the availability of natural sialic acid in the growth media. Although they represent the majority of cell surface sialosides, analogs with longer N-acyl groups diminish the overall level of LOS sialylation, culminating in complete inhibition of LOS sialylation by N-octanoyl sialic acid. Thus, sialylation of H. ducreyi LOS can be modulated with respect to the structure of the terminal sialic acid residue and the extent to which the LOS acceptor is modified by supplying the bacteria with various sialic acid analogs.
-
characterization of Lipooligosaccharides from haemophilus ducreyi containing polylactosamine repeats
Journal of the American Society for Mass Spectrometry, 2002Co-Authors: Birgit Schilling, Bradford W. Gibson, Melanie J Filiatrault, Anthony A. CampagnariAbstract:Haemophilus ducreyi, a gram-negative human mucosal pathogen, is one of the principal causes of genital ulcer disease. The Lipooligosaccharides (LOS) of these bacteria are considered to be a major virulence factor and have been implicated in the adherence and invasion of H. ducreyi to several human cell types. An isogenic heptosyltransferase-III knockout strain (waaQ) was recently constructed from H. ducreyi 35000 wild-type strain and immunochemical and molecular weight data of the isolated LOS suggested the presence of poly-N-acetyllactosamine (LacNAc) (Filiatrault et al., Infect. Immun.2000, 68, 3352–3361). In this present study, the structures of these novel LOS-glycoforms were characterized by matrix-assisted laser desorption/ionization (MALDI) and electrospray ionization (ESI) mass spectrometry in combination with exoglycosidase digestion. Detailed structural information was obtained for the oligosaccharide (OS) portions of these LOS showing between one to five linear LacNAc repeats on the non-reducing terminus of the main oligosaccharide branch. When grown on solid media, the organism produced LacNAc repeats that were further modified by the addition of sialic acid. Enzymatic digestion with β-galactosidase, β-N-acetylhexosaminidase, and neuraminidase type VI-A yielded truncated glycoforms consistent with a polyLacNAc structure capped at various end points with sialic acid. ESI-MS/MS mass spectrometry on a quadrupole time-of-flight instrument was particularly effective in obtaining detailed structural information on the least abundant, high-mass glycoforms. Although LOS containing terminal di-LacNAc have been reported, this is the first time to our knowledge that a linear polyLacNAc structure has been characterized in bacteria.
-
Gonococcal Lipooligosaccharide is a ligand for the asialoglycoprotein receptor on human sperm
Molecular microbiology, 2000Co-Authors: H. A. Harvey, Bradford W. Gibson, Nancy J. Phillips, Michael P. Jennings, Nurith Porat, Colleen A. Campbell, M A Apicella, Blake Milan SAbstract:In the present study, we show that Neisseria gonorrhoeae Lipooligosaccharide (LOS) can bind to the asialoglycoprotein receptor (ASGP-R) on human sperm. This work demonstrates the presence of ASGP-R on human sperm. Binding of purified ASGP-R ligand decreased in the presence of gonococci. Binding of purified iodinated gonococcal LOS identified a protein of molecular weight corresponding to that of human ASGP-R. The presence of excess unlabelled LOS blocked binding of iodinated gonococcal LOS. Binding of wild-type gonococcal LOS to sperm was higher than that of mutant LOS lacking the galactose ligand for ASGP-R. These data suggest that the ASGP-R on human sperm cells recognizes and binds wild-type gonococcal LOS. This interaction may contribute to the transmission of gonorrhea from infected males to their sexual partners.
-
investigation of the kinetic mechanism of cytidine 5 monophosphate n acetylneuraminic acid synthetase from haemophilus ducreyi with new insights on rate limiting steps from product inhibition analysis
Biochemistry, 1999Co-Authors: Nicole M Samuels, Bradford W. Gibson, Susan M MillerAbstract:The presence of sialic acid as a component of cell surface Lipooligosaccharides or capsular polysaccharides has been shown to be correlated with the virulence of a number of Gram-negative mucosal pathogens, including several Haemophilus and Neisseria spp. As part of our efforts to evaluate the role of sialic acid in the pathobiology of these organisms, we have initiated studies of the enzymes from Haemophilus ducreyi (the infectious agent of chancroid) responsible for the activation and attachment of sialic acid to the Lipooligosaccharide. In this report, we describe results of an investigation of the steady-state kinetic mechanism of the activating enzyme, cytidine 5‘-monophosphate N-acetylneuraminic acid (CMP-NeuAc) synthetase. Using a combination of initial velocity, product inhibition, and dead-end inhibition studies, the reaction is shown to be freely reversible and to proceed through an ordered bi-bi kinetic mechanism in which CTP binds first and CMP-NeuAc dissociates last. In addition, a detailed a...
Timothy F Murphy - One of the best experts on this subject based on the ideXlab platform.
-
REVIEW ARTICLES Nontypable Haemophilus influenzae: A Review of Clinical Aspects, Surface Antigens, and the Human Immune Response to Infection
2016Co-Authors: Timothy F Murphy, M A ApicellaAbstract:Nontypable Haemophilus influenzae has now become well established as an important pathogen in both adults and children. Recent work has identified clear distinctions be-tween nontypable and type b strains of H. influenzae. These organisms affect different patient populations, cause different infections, present different surface antigens to the host, and are genetically different. The commonest clinical manifestation of infection due to nontypable H. influenzae in adults is lower respiratory tract infection, particularly in the elderly and in those with chronic bronchitis. The bacterium is a frequent cause of acute otitis media in children. The surface of nontypable H. influenzae is composed of outer-membrane proteins and Lipooligosaccharide, and both ofthese demonstrate sub-stantial antigenic heterogeneity, which can be used to serotype isolates. Some respiratory tract isolates are fimbriated, but the role of fimbriae in pathogenesis is unclear. Antibod-ies to outer-membrane proteins and Lipooligosaccharide are present in human serum. In-vestigation of human immunity to infection is focusing on identification of those anti-gens to which protective antibody is directed. In the past several years, nontypable (nonencapsu
-
biofilm formation by nontypeable haemophilus influenzae strain variability outer membrane antigen expression and role of pili
BMC Microbiology, 2002Co-Authors: Timothy F Murphy, Charmaine KirkhamAbstract:Nontypeable Haemophilus influenzae is an important cause of otitis media in children and lower respiratory tract infection in adults with chronic obstructive pulmonary disease (COPD). Several lines of evidence suggest that the bacterium grows as a biofilm in the human respiratory tract. Fifteen clinical isolates from middle ear fluid of children with otitis media and 15 isolates from sputum of adults with COPD were studied in an in vitro assay of biofilm formation. Striking variability among isolates was observed in their ability to form biofilms. Analysis of cell envelopes revealed minimal differences in banding patterns in polyacrylamide gels, alteration of expression of an epitope on Lipooligosaccharide, and preservation of expression of selected epitopes on outer membrane proteins P2, P5 and P6 in biofilms compared to planktonically grown cells. A pilus-deficient variant showed a marked impairment in biofilm formation compared to its isogenic parent. Nontypeable H. influenzae forms biofilms in vitro. Clinical isolates show substantial variability in their ability to grow as biofilms. Three major outer membrane proteins (P2, P5 and P6) are expressed during growth as a biofilm. Expression of Lipooligosaccharide is altered during growth as a biofilm compared to planktonic growth. Pili are important in biofilm formation. As the role of biofilms in human infection becomes better defined, characterization of biofilms may be important in understanding the pathogenesis of infection and immune response to nontypeable H. influenzae in children with otitis media and adults with COPD.
Charmaine Kirkham - One of the best experts on this subject based on the ideXlab platform.
-
biofilm formation by nontypeable haemophilus influenzae strain variability outer membrane antigen expression and role of pili
BMC Microbiology, 2002Co-Authors: Timothy F Murphy, Charmaine KirkhamAbstract:Nontypeable Haemophilus influenzae is an important cause of otitis media in children and lower respiratory tract infection in adults with chronic obstructive pulmonary disease (COPD). Several lines of evidence suggest that the bacterium grows as a biofilm in the human respiratory tract. Fifteen clinical isolates from middle ear fluid of children with otitis media and 15 isolates from sputum of adults with COPD were studied in an in vitro assay of biofilm formation. Striking variability among isolates was observed in their ability to form biofilms. Analysis of cell envelopes revealed minimal differences in banding patterns in polyacrylamide gels, alteration of expression of an epitope on Lipooligosaccharide, and preservation of expression of selected epitopes on outer membrane proteins P2, P5 and P6 in biofilms compared to planktonically grown cells. A pilus-deficient variant showed a marked impairment in biofilm formation compared to its isogenic parent. Nontypeable H. influenzae forms biofilms in vitro. Clinical isolates show substantial variability in their ability to grow as biofilms. Three major outer membrane proteins (P2, P5 and P6) are expressed during growth as a biofilm. Expression of Lipooligosaccharide is altered during growth as a biofilm compared to planktonic growth. Pili are important in biofilm formation. As the role of biofilms in human infection becomes better defined, characterization of biofilms may be important in understanding the pathogenesis of infection and immune response to nontypeable H. influenzae in children with otitis media and adults with COPD.
Edward W Swords - One of the best experts on this subject based on the ideXlab platform.
-
luxs promotes biofilm maturation and persistence of nontypeable haemophilus influenzae in vivo via modulation of Lipooligosaccharides on the bacterial surface
Infection and Immunity, 2009Co-Authors: Chelsie E Armbruster, Wenzhou Hong, Bing Pang, Kristin E Dew, Richard A Juneau, Matthew S Byrd, Cheraton F Love, Nancy D Kock, Edward W SwordsAbstract:Nontypeable Haemophilus influenzae (NTHI) is an extremely common airway commensal which can cause opportunistic infections that are usually localized to airway mucosal surfaces. During many of these infections, NTHI forms biofilm communities that promote persistence in vivo. For many bacterial species, density-dependent quorum-signaling networks can affect biofilm formation and/or maturation. Mutation of luxS, a determinant of the autoinducer 2 (AI-2) quorum signal pathway, increases NTHI virulence in the chinchilla model for otitis media infections. For example, bacterial counts in middle-ear fluids and the severity of the host inflammatory response were increased in luxS mutants compared with parental strains. As these phenotypes are consistent with those that we have observed for biofilm-defective NTHI mutants, we hypothesized that luxS may affect NTHI biofilms. A luxS mutant was generated using the well-characterized NTHI 86-028NP strain and tested to determine the effects of the mutation on biofilm phenotypes in vitro and bacterial persistence and disease severity during experimental otitis media. Quantitation of the biofilm structure by confocal microscopy and COMSTAT analysis revealed significantly reduced biomass for NTHI 86-028NP luxS biofilms, which was restored by a soluble mediator in NTHI 86-028NP supernatants. Analysis of Lipooligosaccharide moieties using an enzyme-linked immunosorbent assay and immunoblotting showed decreased levels of biofilm-associated glycoforms in the NTHI 86-028NP luxS strain. Infection studies showed that NTHI 86-028NP luxS had a significant persistence defect in vivo during chronic otitis media infection. Based on these data, we concluded that a luxS-dependent soluble mediator modulates the composition of the NTHI Lipooligosaccharides, resulting in effects on biofilm maturation and bacterial persistence in vivo.
-
Lipooligosaccharides containing phosphorylcholine delay pulmonary clearance of nontypeable haemophilus influenzae
Infection and Immunity, 2008Co-Authors: Bing Pang, Wenzhou Hong, Nancy D Kock, Dana Winn, Ryan C Johnson, Shayla Westbarnette, Edward W SwordsAbstract:Nontypeable Haemophilus influenzae (NTHi) causes pulmonary infections in patients with chronic obstructive pulmonary disease and other mucociliary clearance defects. Like many bacteria inhabiting mucosal surfaces, NTHi produces Lipooligosaccharide (LOS) endotoxins that lack the O side chain. Persistent NTHi populations express a discrete subset of LOS glycoforms, including those containing phosphorylcholine (PCho). In this study, we compared two NTHi strains with isogenic mutants lacking PCho for clearance from mice following pulmonary infection. Consistent with data from other model systems, populations of the strains NTHi 2019 and NTHi 86-028NP recovered from mouse lung contained an increased proportion of PCho+ variants compared to that in the inocula. PCho− mutants were more rapidly cleared. Serial passage of NTHi increased both PCho content and bacterial resistance to clearance, and no such increases were observed for PCho− mutants. Increased PCho content was also observed in NTHi populations within non-endotoxin-responsive C3H/HeJ and Toll-like receptor 4 null (TLR4−/−) mice, albeit at later times postinfection. Changes in bacterial subpopulations and clearance were unaffected in TLR2−/− mice compared to the subpopulations in and clearance from mice of the parental strain. The clearance of PCho− mutants occurred at earlier time points in both strain backgrounds and in all types of mice. Comparison of bacterial populations in lung tissue cryosections by immunofluorescent staining showed sparse bacteria within the air spaces of C57BL/6 mice and large bacterial aggregates within the lungs of MyD88−/− mice. These results indicate that PCho promotes bacterial resistance to pulmonary clearance early in infection in a manner that is at least partially independent of the TLR4 pathway.
-
sialylation of Lipooligosaccharides promotes biofilm formation by nontypeable haemophilus influenzae
Infection and Immunity, 2004Co-Authors: Edward W Swords, Miranda L Moore, Luciana Godzicki, Gail Bukofzer, Michael J Mitten, Jessica L VoncannonAbstract:Nontypeable Haemophilus influenzae (NTHi) is a major cause of opportunistic respiratory tract infections, including otitis media and bronchitis. The persistence of NTHi in vivo is thought to involve bacterial persistence in a biofilm community. Therefore, there is a need for further definition of bacterial factors contributing to biofilm formation by NTHi. Like other bacteria inhabiting host mucosal surfaces, NTHi has on its surface a diverse array of Lipooligosaccharides (LOS) that influence host-bacterial interactions. In this study, we show that LOS containing sialic (N-acetyl-neuraminic) acid promotes biofilm formation by NTHi in vitro and bacterial persistence within the middle ear or lung in vivo. LOS from NTHi in biofilms was sialylated, as determined by comparison of electrophoretic mobilities and immunochemical reactivities before and after neuraminidase treatment. Biofilm formation was significantly reduced in media lacking sialic acid, and a siaB (CMP-sialic acid synthetase) mutant was deficient in biofilm formation in three different in vitro model systems. The persistence of an asialylated siaB mutant was attenuated in a gerbil middle ear infection model system, as well as in a rat pulmonary challenge model system. These data show that sialylated LOS glycoforms promote biofilm formation by NTHi and persistence in vivo.