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. 2024 Feb 2:14:1342621.
doi: 10.3389/fcimb.2024.1342621. eCollection 2024.

Antibody responses to Chlamydia trachomatis vaccine candidate antigens in Chlamydia-infected women and correlation with antibody-mediated phagocytosis of elementary bodies

Hong Yu  1 , William M Geisler  2 , Chuanbin Dai  1 , Kanupriya Gupta  2 , Gary Cutter  3 , Robert C Brunham  1
Affiliations

Antibody responses to Chlamydia trachomatis vaccine candidate antigens in Chlamydia-infected women and correlation with antibody-mediated phagocytosis of elementary bodies

Hong Yu et al. Front Cell Infect Microbiol. .

Abstract

Murine research has revealed a significant role for antibody responses in protection against Chlamydia reinfection. To explore potential humoral immune markers of protection elicited by Chlamydia trachomatis (CT) antigens in humans in the context of presumed clinical correlates of protection, we used both an IgG1-based ELISA and a conventional total IgG ELISA to evaluate antibody responses. We evaluated responses to five CT outer membrane proteins (PmpE, PmpF, PmpG, PmpH, and MOMP), along with other promising CT antigens (Pgp3 and HSP60), negative control antigens (RecO and AtpE), and CT elementary bodies (EBs) in sera from a well-characterized cohort of 60 women with different CT infection outcomes, including two outcomes that are likely clinical correlates of protective immunity: spontaneous resolution of infection and absence of reinfection after treatment. Furthermore, we used a flow cytometry-based assay to measure antibody-mediated phagocytosis by neutrophils in these sera. Results demonstrated that IgG1 ELISA displayed higher sensitivity than conventional total IgG ELISA in assessing antibody responses to CT EBs and antigens. Pgp3 IgG1 ELISA exhibited the highest sensitivity compared to IgG1 ELISA incorporating CT EBs or other antigens, confirming Pgp3 IgG1 ELISA as an ideal assay for CT antibody detection. Most (95%) sera from women with CT infection outcomes exhibited antibody-mediated phagocytosis of CT EBs, which was significantly correlated with IgG1 antibody responses to MOMP, Pgp3, HSP60, and PmpF. However, neither IgG1 responses to CT antigens and EBs nor antibody-mediated phagocytosis were associated with clinical correlates of protection. These findings suggest that neither CT IgG1 antibody detection nor antibody-mediated phagocytosis will be useful as immune correlates of protection against CT infection in humans.

Keywords: Chlamydia; IgG1 ELISA; Pgp3; antibody; antibody-mediated phagocytosis; antigen; serology.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Enhanced sensitivity of IgG1 ELISA for detecting antibody responses to Chlamydia trachomatis (CT) antigens compared to total IgG ELISA. Five serum samples from five women with CT infection outcomes (A) and serum samples from five CT naïve women (B) were utilized for both IgG1 ELISA and total IgG ELISA.
Figure 2
Figure 2
Pgp3 IgG1 ELISA is more sensitive for Chlamydia trachomatis (CT) antibody detection compared to HSP60 IgG1, MOMP IgG1, and CT EB IgG1 ELISAs. (A–D) Antibody responses to Pgp3, MOMP, CT EB or HSP60 determined by IgG1 ELISA in 120 serum samples (enrollment and follow-up) from 60 women with CT infection outcomes (CT+) and 20 serum samples from CT-naïve women (CT-). (E–H) Antibody responses to Pgp3, MOMP, CT EB or HSP60, at enrollment visit versus follow-up visit (paired t test).
Figure 3
Figure 3
Pgp3 IgG1 ELISA is highly sensitive, with the strongest Pgp3 IgG1 responses during current Chlamydia trachomatis (CT) infection. IgG1 antibody responses to Pgp3 (A), HSP60 (B), MOMP (C), and CT EBs (D) in the combined persisting infection groups at the enrollment visit (Persisting_V1), the spontaneous resolution group at the enrollment visit (Resolved_V1), the persisting infection with reinfection group at the follow-up visit (Persisting with reinf._V2), the persisting infection without reinfection group at the follow-up visit (Persisting w/o reinf._V2) and the spontaneous resolution group at the follow-up visit (Resolved_V2). (Unpaired t test between groups with different outcomes).
Figure 4
Figure 4
Assessment of Antibody-Mediated Phagocytosis of Chlamydia trachomatis (CT) EBs Using a Flow Cytometry-Based Assay. CFSE-labeled CT EBs were preincubated with human serum samples, followed by incubation with DMF-stimulated PLB-985 cells (neutrophil-like cells). The CFSE signal was quantified using flow cytometry. Cells were gated based on CFSE-positive (indicating phagocytosis) events. (A) Flow cytometry plots depict phagocytosing CFSE+ cells from PLB cells alone, PLB cells with CT EBs, and PLB cells with CT EBs plus serum samples from representative women with CT infection outcomes (CT+) or who are CT-naïve (CT-). (B) Percentage of phagocytosing CFSE+ cells in serum from 120 women with CT infection outcomes and 20 CT naïve women. (C) Percentage of phagocytosing CFSE+ cells at enrollment visit versus follow-up visit. (paired t test). (D) Percentage of phagocytosing CFSE+ cells in groups with different outcomes of CT infection (Unpaired t test).
Figure 5
Figure 5
Strong correlations between phagocytosis of Chlamydia trachomatis (CT) EBs and IgG1 antibodies against specific CT antigens and EBs in serum samples from 60 women with CT infection outcomes measured by IgG1 ELISA at follow-up. (A) The percentage of cells with phagocytosed CT EBs versus the OD values of IgG1 antibodies specific to CT antigens (B) Correlation analysis between phagocytosis and IgG1 responses against 9 CT antigens and CT EBs (Pearson’s correlation test).
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