Expression of Opacity Proteins Interferes with the Transmigration of Neisseria gonorrhoeae across Polarized Epithelial Cells

Stein, Daniel C.; Adriana, LeVan; Britney, Hardy; Wang, Liangchun; Zimmerman, Lindsey I.; Song, Wenxia

doi:10.1371/journal.pone.0134342

Cited by 22 publications

(40 citation statements)

References 57 publications

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“…To address the potential role of mAb 2C7-dependent complement activation in facilitating opsonophagocytosis of the 3-Hex/G+ mutant, we recreated the 3-Hex/G+ LOS structure in the background of an Opa-negative MS11 strain (42). The 3-Hex/G+ Opa-negative MS11 mutant strain bound similar (low) levels of mAb 2C7 and was fully resistant (>100% survival) to killing by 4 μg/ml of mAb 2C7 plus 20% complement (Fig.…”

Section: Resultsmentioning

confidence: 99%

Phase-Variable Heptose I Glycan Extensions Modulate Efficacy of 2C7 Vaccine Antibody Directed against Neisseria gonorrhoeae Lipooligosaccharide

Chakraborti

Lewis

Cox

et al. 2016

The Journal of Immunology

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Neisseria gonorrhoeae , the causative agent of the sexually transmitted infection, gonorrhea, has developed resistance to most conventional antibiotics. Safe and effective vaccines against gonorrhea are needed urgently. A candidate vaccine that targets a lipooligosaccharide (LOS) epitope recognized by monoclonal (mAb) 2C7 attenuates gonococcal burden in the mouse vaginal colonization model. Glycan extensions from the LOS core heptoses (HepI and HepII) are controlled by phase-variable LOS glycosyltransferase (lgt) genes; we sought to define how HepI glycan extensions affect mAb 2C7 function. Isogenic gonococcal mutants in which the lgt required for mAb 2C7 reactivity (lgtG) was genetically locked ‘ON’ and the lgt loci required for HepI variation (lgtA, lgtC and lgtD) were genetically locked ‘ON’ or ‘OFF’ in different combinations were created. We observed 100% complement-dependent killing by mAb 2C7 of a mutant that expressed lactose (Gal-Glc) from HepI, while a mutant that expressed Gal-Gal-Glc-HepI fully resisted killing (>100% survival). Mutants that elaborated 4- (Gal-GlcNAc-Gal-Glc-HepI) and 5-glycan (GalNAc-Gal-GlcNAc-Gal-Glc-HepI) structures displayed ‘intermediate’ phenotypes (<50% killing with 2 μg/ml and >95% killing with 4 μg/ml of mAb 2C7). The contrasting phenotypes of the lactose-HepI and the Gal-Gal-Glc-HepI LOS structures were recapitulated with phase-variants of a recently isolated clinical strain. Despite lack of killing of the Gal-Gal-Glc-HepI mutants, mAb 2C7 deposited sufficient C3 on these bacteria for opsonophagocytic killing by human neutrophils. In conclusion, mAb 2C7 showed functional activity against all gonococcal HepI LOS structures defined by various lgtA/C/D ‘ON/OFF’ combinations, thereby providing further impetus for use of the 2C7 epitope in a gonococcal vaccine.

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Section: Resultsmentioning

confidence: 99%

Phase-Variable Heptose I Glycan Extensions Modulate Efficacy of 2C7 Vaccine Antibody Directed against Neisseria gonorrhoeae Lipooligosaccharide

Chakraborti

Lewis

Cox

et al. 2016

The Journal of Immunology

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“…GC aggregation and adherence to host cells through these molecules may influence each other, therefore changing both the infectivity and antibiotic resistance of GC in the reproductive tract of women. Indeed, we have previously shown that Opa expression facilitates GC adherence to host cells and GC aggregation, but reduces GC penetration into both polarized epithelial cell monolayers and the endocervical epithelium in the tissue explant model [ 16 , 20 ]. The functions of Opa in both GC-GC and GC-epithelial interactions enable Opa-expressing GC to survive better in the presence of antibiotic agents and dominate the colonization of the epithelial surface.…”

Section: Discussionmentioning

confidence: 99%

“…The GC biofilm progresses from an intimately associated aggregation of microcolonies to an organized biofilm. Pili, opacity-associated protein (Opa), and lipooligosaccharides (LOS), three well-characterized phase variable surface molecules of GC, have been shown to mediate inter-bacterium interactions to form aggregates [ 15 , 16 , 17 ]. How phase variations in their expression contribute to different GC-GC interactions and whether these interactions affect GC susceptibility to antibiotics is unknown.…”

Section: Introductionmentioning

confidence: 99%

Neisseria gonorrhoeae Aggregation Reduces Its Ceftriaxone Susceptibility

et al. 2018

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Antibiotic resistance in Neisseria gonorrhoeae (GC) has become an emerging threat worldwide and heightens the need for monitoring treatment failures. N. gonorrhoeae, a gram-negative bacterium responsible for gonorrhea, infects humans exclusively and can form aggregates during infection. While minimal inhibitory concentration (MIC) tests are often used for determining antibiotic resistance development and treatment, the knowledge of the true MIC in individual patients and how it relates to this laboratory measure is not known. We examined the effect of aggregation on GC antibiotic susceptibility and the relationship between bacterial aggregate size and their antibiotic susceptibility. Aggregated GC have a higher survival rate when treated with ceftriaxone than non-aggregated GC, with bacteria in the core of the aggregates surviving the treatment. GC lacking opacity-associated protein or pili, or expressing a truncated lipooligosaccharide, three surface molecules that mediate GC-GC interactions, reduce both aggregation and ceftriaxone survival. This study demonstrates that the aggregation of N. gonorrhoeae can reduce the susceptibility to antibiotics, and suggests that antibiotic utilization can select for GC surface molecules that promote aggregation which in turn drive pathogen evolution. Inhibiting aggregation may be a potential way of increasing the efficacy of ceftriaxone treatment, consequently reducing treatment failure.

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“…In addition, we created a mutant in the parent (or wild-type [WT]) strain MS11 4/3/1, where only lgtG was locked on to create WT/Gϩ; the ability of HepI glycans to phase vary was unaffected in this mutant. As previously described (15), a 3-Hex/Gϩ LOS mutant was made in the background of MS11 ΔOpa for opsonic killing studies by PMNs, where all 11 opa genes were deleted (16).…”

Section: Methodsmentioning

confidence: 99%

“…where all 11 Opa proteins had been deleted (MS11 ΔOpa) (15,16). Elimination of Opa(s) avoids interactions with CEACAM3 on human PMNs that would result in nonopsonic killing (17,18) and thereby confound interpretation of results.…”

Section: -Hex/g؉ Opsonized With 2c7-ximab(s) Is Killed By Pmns In a mentioning

confidence: 99%

Bypassing Phase Variation of Lipooligosaccharide (LOS): Using Heptose 1 Glycan Mutants To Establish Widespread Efficacy of Gonococcal Anti-LOS Monoclonal Antibody 2C7

et al. 2020

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The sialylatable lacto-N-neotetraose (LNnT; Gal-GlcNAc-Gal-Glc) moiety from heptose I (HepI) of the lipooligosaccharide (LOS) of Neisseria gonorrhoeae undergoes positive selection during human infection. Lactose (Gal-Glc) from HepII, although phase variable, is commonly expressed in humans; loss of HepII lactose compromises gonococcal fitness in mice. Anti-LOS monoclonal antibody (MAb) 2C7, a promising antigonococcal immunotherapeutic that elicits complement-dependent bactericidal activity and attenuates gonococcal colonization in mice, recognizes an epitope comprised of lactoses expressed simultaneously from HepI and HepII. Glycan extensions beyond lactose on HepI modulate binding and function of MAb 2C7 in vitro. Here, four gonococcal LOS mutants, each with lactose from HepII but fixed (unable to phase-vary) LOS HepI glycans extended beyond the lactose substitution of HepI (lactose alone, Gal-lactose, LNnT, or GalNAc-LNnT), were used to define how HepI glycan extensions affect (i) mouse vaginal colonization and (ii) efficacy in vitro and in vivo of a human IgG1 chimeric derivative of MAb 2C7 (2C7-Ximab) with a complement-enhancing E-to-G Fc mutation at position 430 (2C7-Ximab-E430G). About 10-fold lower 2C7-Ximab-E430G concentrations achieved similar complement-dependent killing of three gonococcal mutants with glycan extensions beyond lactose-substituted HepI (lactose alone, LNnT, or GalNAc-LNnT) as 2C7-Ximab (unmodified Fc). The fourth mutant (Gal-lactose) resisted direct complement-dependent killing but was killed approximately 70% by 2C7-Ximab-E430G in the presence of polymorphonuclear leukocytes and complement. Only mutants with (sialylatable) LNnT from HepI colonized mice for >3 days, reiterating the importance of LNnT sialylation for infection. 2C7-Ximab-E430G significantly attenuated colonization caused by the virulent mutants.

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Expression of Opacity Proteins Interferes with the Transmigration of Neisseria gonorrhoeae across Polarized Epithelial Cells

Cited by 22 publications

References 57 publications

Phase-Variable Heptose I Glycan Extensions Modulate Efficacy of 2C7 Vaccine Antibody Directed against Neisseria gonorrhoeae Lipooligosaccharide

Phase-Variable Heptose I Glycan Extensions Modulate Efficacy of 2C7 Vaccine Antibody Directed against Neisseria gonorrhoeae Lipooligosaccharide

Neisseria gonorrhoeae Aggregation Reduces Its Ceftriaxone Susceptibility

Bypassing Phase Variation of Lipooligosaccharide (LOS): Using Heptose 1 Glycan Mutants To Establish Widespread Efficacy of Gonococcal Anti-LOS Monoclonal Antibody 2C7

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