Workshop on In Vitro Neutralization of Chlamydia trachomatis: Summary of Proceedings

Byrne, Gerald I.; Stephens, Richard S.; Ada, G. L.; Caldwell, Harlan D.; Su, Hua; Morrison, Robert; Pol, Barbara Van Der; Bavoil, Patrik M.; Bobo, Linda; Everson, S; Ho, Yu Min; Hsia, Ru-ching; Kennedy, Kathleen A.; Kuo, Cho‐Chou; Montgomery, P. C.; Peterson, Ellena M.; Swanson, Albertina F.; Whitaker, Charles W.; Hudson, J. Whittum; Yang, Chunlin; Zhang, Y.-X.; Zhong, Guangming

doi:10.1093/infdis/168.2.415

Cited by 57 publications

(44 citation statements)

References 17 publications

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“…Two methods of assaying inhibition of cell culture infectivity by ovalbumin glycopeptides or defined oligosaccharides were performed using HeLa 229 cell monolayers grown in 96-well microtiter plates (16). One was the pretreatment of organisms with sugars, then inoculation onto HeLa cells.…”

Section: Methodsmentioning

confidence: 99%

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An N-linked high-mannose type oligosaccharide, expressed at the major outer membrane protein of Chlamydia trachomatis, mediates attachment and infectivity of the microorganism to HeLa cells.

Kuo

Takahashi²,

Swanson³

et al. 1996

J. Clin. Invest.

118

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The structure of the carbohydrate of the 40-kD major outer membrane component of Chlamydia trachomatis and its role in defining infectivity of the organism were investigated. The oligosaccharides were released from the glycoprotein by N-glycanase digestion, coupled to a 2-aminopyridyl residue, and subjected to two-dimensional sugar mapping technique. The major fractions consisted of "highmannose type" oligosaccharides containing 8-9 mannose residues. Bi-and tri-antennary "complex type" oligosaccharides having terminal galactose were detected as minor components. These oligosaccharides were N-linked and contained no sialic acid. This structural profile is consistent with our previous characterization based on lectin-binding and glycosidase digestion. Functional specificity of identified chlamydial oligosaccharides was analyzed using glycopeptides fractionated from ovalbumin and structurally defined oligosaccharides from other sources. The glycopeptide fraction having high-mannose type oligosaccharide, as compared to those having complex or hybrid-type, showed a stronger inhibitory effect on attachment and infectivity of chlamydial organisms to HeLa cells. Among high-mannose type oligosaccharides, the strongest inhibition was observed with mannose 8 as compared with mannose 6, 7, or 9. These results indicate that a specific high-mannose type oligosaccharide linked to the major outer membrane protein of C. trachomatis mediates attachment and infectivity of the organism to HeLa cells. ( J. Clin. Invest. 1996. 98:2813-2818.)

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Section: Methodsmentioning

confidence: 99%

“…These monoclonal antibodies have been described previously (5,10,17). A reduction of more than 50% of inclusion counts is regarded as indicating positive neutralization for chlamydia (16).…”

Section: Methodsmentioning

confidence: 99%

An N-linked high-mannose type oligosaccharide, expressed at the major outer membrane protein of Chlamydia trachomatis, mediates attachment and infectivity of the microorganism to HeLa cells.

Kuo

Takahashi²,

Swanson³

et al. 1996

J. Clin. Invest.

118

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“…Consequently, many immunological studies of human genital tract infections have been focused on the major outer membrane protein (MOMP) because C. trachomatis serovars are defined by serological variation in this protein. In humans, MOMP elicits C. trachomatisneutralizing antibodies that recognize its surface-exposed variable segments (VSs), where amino acid sequences vary substantially among serovars (4). Therefore, some have speculated that B-cell responses directed toward serovar-specific epitopes located in VSs of MOMP are a critical component of protective immunity (16)(17)(18).…”

mentioning

confidence: 99%

T-Cell Epitopes in Variable Segments ofChlamydia trachomatisMajor Outer Membrane Protein Elicit Serovar-Specific Immune Responses in Infected Humans

Ortiz¹,

Angevine²,

Kim

et al. 2000

Infect Immun

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We previously identified 18 stimulatory Chlamydia trachomatis major outer membrane protein (MOMP) peptides containing at least 23 epitopes presented with various HLA class II allotypes. Only one peptide contained an epitope localized in a variable segment (VS2). Continued studies reported here identified a total of five VS peptides containing T-cell epitopes that are distributed among MOMPs VS1, VS2, and VS4. Only MOMP-primed T-cell cultures from subjects infected with serovar E responded to the serovar E VS peptides, while the response of such cultures to constant-segment peptides was independent of the infecting serovar. Furthermore, MOMP-primed T cells proliferated in response only to the VS peptides encoded in serovar E but not to the corresponding peptides derived from serovar F, I, or J, confirming that these responses were serovar specific.Chlamydia trachomatis causes ocular and genital tract infections worldwide. In the United States, the annual cost of treating sexually transmitted diseases caused by C. trachomatis is billions of dollars (9). Effective preventive measures against C. trachomatis infections are not yet available, due in part to insufficient understanding of human immune responses to C. trachomatis. Evidence for protective immunity against C. trachomatis is scant and inconclusive. However, earlier studies have suggested that short-lived immunity develops after an episode of chlamydial infection (1,10,12) and that protection may be serovar specific (reviewed in reference 8); while reinfection with C. trachomatis is common, it is usually not with serovars of preceding infections. Consequently, many immunological studies of human genital tract infections have been focused on the major outer membrane protein (MOMP) because C. trachomatis serovars are defined by serological variation in this protein. In humans, MOMP elicits C. trachomatisneutralizing antibodies that recognize its surface-exposed variable segments (VSs), where amino acid sequences vary substantially among serovars (4). Therefore, some have speculated that B-cell responses directed toward serovar-specific epitopes located in VSs of MOMP are a critical component of protective immunity (16)(17)(18).Because of the role of T helper (Th) cells in priming and maintaining B-cell responses, we have investigated HLA class II-restricted Th cell responses to MOMP in humans genitally infected with C. trachomatis, focusing on commonly involved serovar E. The main results of our first study (14) were that (i) in vitro priming of peripheral blood mononuclear cells (PBMC) with MOMP yielded MOMP-specific Th cell cultures from 83% of serovar E-infected subjects; (ii) proliferative responses of T-MOMPЈ cultures to synthetic peptides localized at least 23 Th cell epitopes, and T-MOMPЈ cultures from most subjects responded to multiple epitopes; (iii) multiple HLA-DR and other HLA class II molecules were used to present the MOMP epitopes to the Th cells; and (iv) remarkably, 22 of the 23 epitopes were distributed among the five MOMP constant segments ...

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“…In vitro neutralization assays using HaK (Syrian hamster kidney) cells were performed as previously described (23). Serial dilutions of monospecific PorB peptide antisera were prepared in SPG.…”

Section: In Vitro Neutralizationmentioning

confidence: 99%

“…In vitro neutralization assays are a central component in evaluating functionality of chlamydial immune responses and provide a correlate of protective immune responses (23). The neutralization data for the peptide antisera revealed four regions of neutralizing activity on the PorB protein, with 50% reciprocal neutralization endpoint titers ranging from ϳ1:32 to ϳ1:2048 (Fig.…”

Section: In Vitro Neutralization Of Chlamydial Infectivitymentioning

confidence: 99%

Antigenic Topology of Chlamydial PorB Protein and Identification of Targets for Immune Neutralization of Infectivity

Kawa

Stephens

2002

The Journal of Immunology

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The outer membrane protein PorB is a conserved chlamydial protein that functions as a porin and is capable of eliciting neutralizing Abs. A topological antigenic map was developed using overlapping synthetic peptides representing the Chlamydia trachomatis PorB sequence and polyclonal immune sera. To identify which antigenic determinants were surface accessible, monospecific antisera were raised to the PorB peptides and were used in dot-blot and ELISA-based absorption studies with viable chlamydial elementary bodies (EBs). The ability of the surface-accessible antigenic determinants to direct neutralizing Ab responses was investigated using standardized in vitro neutralization assays. Four major antigenic clusters corresponding to Phe34-Leu59 (B1-2 and B1-3), Asp112 -Glu145 (B2-3 and B2-4), Gly179-Ala225 (B3-2 to B3-4), and Val261-Asn305 (B4-4 to B5-2) were identified. Collectively, the EB absorption and dot-blot assays established that the immunoreactive PorB Ags were exposed on the surface of chlamydial EBs. Peptide-specific antisera raised to the surface-accessible Ags neutralized chlamydial infectivity and demonstrated cross-reactivity to synthetic peptides representing analogous C. pneumoniae PorB sequences. Furthermore, neutralization of chlamydial infectivity by C. trachomatis PorB antisera was inhibited by synthetic peptides representing the surface-exposed PorB antigenic determinants. These findings demonstrate that PorB Ags may be useful for development of chlamydial vaccines.

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Workshop on In Vitro Neutralization of Chlamydia trachomatis: Summary of Proceedings

Cited by 57 publications

References 17 publications

An N-linked high-mannose type oligosaccharide, expressed at the major outer membrane protein of Chlamydia trachomatis, mediates attachment and infectivity of the microorganism to HeLa cells.

An N-linked high-mannose type oligosaccharide, expressed at the major outer membrane protein of Chlamydia trachomatis, mediates attachment and infectivity of the microorganism to HeLa cells.

T-Cell Epitopes in Variable Segments ofChlamydia trachomatisMajor Outer Membrane Protein Elicit Serovar-Specific Immune Responses in Infected Humans

Antigenic Topology of Chlamydial PorB Protein and Identification of Targets for Immune Neutralization of Infectivity

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