Interferon-Gamma Responses to Chlamydia trachomatis Vaccine Candidate Proteins in Chlamydia-Infected Women with Different Chlamydia Outcomes.

BACKGROUND: Chlamydia trachomatis (CT) testing and treatment strategies have not decreased infection rates, justifying need for a CT vaccine. A murine study showed that a vaccine consisting of MOMP and 4 polymorphic membrane proteins (Pmps E, F, G, H) elicited protective immunity; studies on human cellular immune responses to Pmps are sparse. METHODS: Interferon gamma (IFN-γ) responses to these 5 CT proteins were measured by ELISPOT in PBMCs from women returning for treatment of a positive CT screening test. Responses were compared in those with spontaneous CT clearance vs. persisting infection at baseline and no reinfection vs. reinfection at a 3-month follow-up visit. RESULTS: IFN-γ response to one or more proteins was detected in 39% at baseline and 51.5% at follow-up; PmpE and MOMP most often elicited positive responses. IFN-γ responses to MOMP were detected less often at follow-up vs. baseline in women with reinfection, but were maintained in those without reinfection. Women with spontaneous clearance had a higher magnitude of IFN-γ response to PmpE and MOMP. CONCLUSIONS: IFN-γ responses to these 5 CT vaccine candidate proteins were heterogenous and primarily directed against MOMP and PmpE. Spontaneous clearance of infection and absence of reinfection may be clinical correlates of protection.

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

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.

Identification of an Optimized Receptor-Binding Domain Subunit Vaccine against SARS-CoV-2.

Current vaccine efforts to combat SARS-CoV-2 are focused on the whole spike protein administered as mRNA, viral vector, or protein subunit. However, the SARS-CoV-2 receptor-binding domain (RBD) is the immunodominant portion of the spike protein, accounting for 90% of serum neutralizing activity. In this study, we constructed several versions of RBD and together with aluminum hydroxide or DDA (dimethyldioctadecylammonium bromide)/TDB (d-(+)-trehalose 6,6'-dibehenate) adjuvant evaluated immunogenicity in mice. We generated human angiotensin-converting enzyme 2 knock-in mice to evaluate vaccine efficacy in vivo following viral challenge. We found that 1) subdomain (SD)1 was essential for the RBD to elicit maximal immunogenicity; 2) RBDSD1 produced in mammalian HEK cells elicited better immunogenicity than did protein produced in insect or yeast cells; 3) RBDSD1 combined with the CD4 Th1 adjuvant DDA/TDB produced higher neutralizing Ab responses and stronger CD4 T cell responses than did aluminum hydroxide; 4) addition of monomeric human Fc receptor to RBDSD1 (RBDSD1Fc) significantly enhanced immunogenicity and neutralizing Ab titers; 5) the Beta version of RBDSD1Fc provided a broad range of cross-neutralization to multiple antigenic variants of concern, including Omicron; and 6) the Beta version of RBDSD1Fc with DDA/TDB provided complete protection against virus challenge in the knock-in mouse model. Thus, we have identified an optimized RBD-based subunit vaccine suitable for clinical trials.

Diagnosis and Management of Uncomplicated Chlamydia trachomatis Infections in Adolescents and Adults: Summary of Evidence Reviewed for the 2021 Centers for Disease Control and Prevention Sexually Transmitted Infections Treatment Guidelines.

To prepare for the development of the 2021 Centers for Disease Control and Prevention (CDC) sexually transmitted infections treatment guidelines, the CDC convened a committee of expert consultants in June 2019 to discuss recent abstracts and published literature on the epidemiology, diagnosis, and management of sexually transmitted infections.This paper summarizes the key questions, evidence, and recommendations for the diagnosis and management of uncomplicated Chlamydia trachomatis (CT) infections in adolescents and adults that were reviewed and discussed for consideration in developing the guidelines. The evidence reviewed mostly focused on efficacy of doxycycline and azithromycin for urogenital, rectal, and oropharyngeal CT infection, CT risk factors in women, performance of CT nucleic acid amplification tests on self-collected meatal specimens in men, and performance of newer CT point-of-care tests.

Problems With Understanding Chlamydia trachomatis Immunology.

The payoff for understanding Chlamydia trachomatis immunology is the development of a vaccine. Two lines of research have contributed to our current understanding of C. trachomatis immunology. The first is the Grayston model of type-specific immunity and genus-specific pathology, which was elaborated by Caldwell and Morrison as the major outer membrane protein and heat shock protein 60 paradigm. The second is the murine model of Chlamydia muridarum infection, which established the essential role of major histocompatibility complex class II and CD4 T cells in immunity. However, neither approach has yielded a vaccine. I review these 2 lines of research and conclude with 6 problem areas in human C. trachomatis immunology whose resolution may result in a vaccine.

Using Epidemiology, Immunology, and Genomics to Study the Biology of Chlamydia trachomatis.

ABSTRACT: The traditional framework in which to study the biology of human infectious diseases involves characterizing interactions and features of the host, pathogen, and environment. Using the tools of epidemiology, immunology, and genomics allows one to study the biology of infectious disease within this framework. The study of Chlamydia trachomatis biology vividly illustrates the usefulness for the approach. I note key findings from my own studies on C. trachomatis epidemiology, immunology, and genomics to show how important light has been shed on its biology and how this has impacted the Chlamydia field generally. In particular, the epidemiology of C. trachomatis diseases in women shows its impact on reproduction and how public health programs to detect and treat infection has reduced that impact but at the cost of arresting the development of protective immunity and increasing the risk of infection and reinfection. Immunological studies demonstrate the importance of CD4 Th1 cells in protection and that antibiotic treatment interferes with the development of protective immunity when given early in the course of infection. Evaluating the T-cell antigen landscape for C. trachomatis and Chlamydia muridarum demonstrates the role of surface proteins such as the major outer-membrane protein and the polymorphic membrane proteins as major protective CD4 T-cell antigens. Genomic studies reveal that the genome of organism has 3 loci of immunological interest. The antigen loci of the major outer-membrane protein and polymorphic membrane proteins are hotspots for both mutation and recombination, and the plasticity zone contains immune evasion genes that are highly variable from species to species. Interestingly, these 3 loci seem to have entered the Chlamydia phylum at the time of the evolution of the Chlamydiaceae when they became pathogens of vertebrates and encountered the adaptive immune system. In aggregate, these 3 approaches have shed light on human C. trachomatis infections and suggest paths for vaccine development. These approaches are likely to remain useful for the further study of C. trachomatis and for other human pathogens.

Reproductive system infections in women: upper genital tract, fetal, neonatal and infant syndromes.

Lower genital tract infection and bloodborne spread of infection are the two principal modes for infection of the upper genital tract or for infection of the fetus, neonate or infant. Treponema pallidum and human immunodeficiency virus (HIV) are the two most common bloodborne pathogens that infect the fetus, neonate or infant. Most infections of the upper genital tract, however, spread along epithelial surfaces from the vagina or cervix to the upper genital tract or chorioamnion, fetus, neonate or infant. These infections are caused by either pathogens associated with a dysbiotic vaginal microbiome or those that are sexually transmitted. The clinical syndromes that these pathogens produce in the lower genital tract were discussed in part one of this review. We now discuss the syndromes and pathogens that affect the upper genital tract of both non-pregnant and pregnant women as well as fetus, neonate and infant.

Reproductive system infections in women: lower genital tract syndromes.

Gynecological and obstetrical infectious diseases are an important component of women's health. A system approach to gynecological and obstetrical infection helps unify and classify microbial etiology and pathogenesis within a clinical anatomical framework of lower and upper genital tract syndromes. The reproductive system of women includes the vulva, vagina, cervix, uterus, fallopian tubes and ovaries. During pregnancy, additional tissues include the chorioamnion and placenta together with the fetus and amniotic fluid. We review in two parts reproductive system infection syndromes in women using selected research results to illustrate the clinical utility of the system approach in terms of diagnosis, treatment and prevention. We conclude that a reproductive system perspective will lead to improvements in understanding, management and prevention of these diseases.

Comparison of Chlamydia outer membrane complex to recombinant outer membrane proteins as vaccine.

The Chlamydial outer membrane complex (COMC) from the elementary body (EB) is a protein rich insoluble outer membrane shell from which cytosolic proteins have been extracted with detergent. In this study we conducted mass spectrometry experiments to detect proteins in the COMC prepared from C. muridarum EB. Proteomic analysis showed that the COMC contained 75 proteins that included 10 outer membrane proteins (OMPs) such as the major outer membrane protein (MOMP) and polymorphic membrane proteins (Pmps) that were previously identified as CD4 T cell vaccine candidates. We tested the vaccine efficacy of COMC in comparison to individual or combination of recombinant OMPs formulated with Th1 polarizing adjuvant DDA/MPL in two murine genital tract models (C. muridarum and C. trachomatis) by measuring organismal shedding, tubal pathology and immune responses including neutralizing antibodies. In the C. muridarum model, the COMC vaccine generated broadly reactive immune responses against multiple outer membrane proteins, high levels of EB binding and neutralizing antibody and exhibited superior protection against genital infection and pathology when compared to the recombinant PmpG vaccine. Denaturing the COMC by boiling significantly reduced protection. In the C. trachomatis model, the COMC vaccine also conferred greater protection compared to individual or multiple recombinant outer membrane proteins. Immunization with multiple COMCs from C. trachomatis serovars D, F and J generated neutralizing antibodies against multiple C. trachomatis serovars. We conclude that broader immunogenicity and generation of neutralizing antibody may explain the superior efficacy of COMC vaccine. The study suggests that conformationally intact proteins will be necessary for a successful recombinant OMP vaccine.

Discordance in the Epithelial Cell-Dendritic Cell Major Histocompatibility Complex Class II Immunoproteome: Implications for Chlamydia Vaccine Development.

BACKGROUND: Chlamydia trachomatis and Chlamydia muridarum are intracellular bacterial pathogens of mucosal epithelial cells. CD4 T cells and major histocompatibility complex (MHC) class II molecules are essential for protective immunity against them. Antigens presented by dendritic cells (DCs) expand naive pathogen-specific T cells (inductive phase), whereas antigens presented by epithelial cells identify infected epithelial cells as targets during the effector phase. We previously showed that DCs infected by C trachomatis or C muridarum present epitopes from a limited spectrum of chlamydial proteins recognized by Chlamydia-specific CD4 T cells from immune mice. METHODS: We hypothesized that Chlamydia-infected DCs and epithelial cells present overlapping sets of Chlamydia-MHC class II epitopes to link inductive and effector phases to generate protective immunity. We tested that hypothesis by infecting an oviductal epithelial cell line with C muridarum, followed by immunoaffinity isolation and sequencing of MHC class I- and II-bound peptides. RESULTS: We identified 26 class I-bound and 4 class II-bound Chlamydia-derived peptides from infected epithelial cells. We were surprised to find that none of the epithelial cell class I- and class II-bound chlamydial peptides overlapped with peptides presented by DCs. CONCLUSIONS: We suggest the discordance between the DC and epithelial cell immunoproteomes has implications for delayed clearance of Chlamydia and design of a Chlamydia vaccine.

National Institute of Allergy and Infectious Diseases workshop report: "Chlamydia vaccines: The way forward".

Chlamydia trachomatis (Ct), an intracellular pathogen, is the most common bacterial sexually transmitted infection. In addition to acute cervicitis and urethritis, Ct can lead to serious sequelae of significant public health burden including pelvic inflammatory disease (PID) and infertility. Ct control efforts have not resulted in desired outcomes such as reduced incidence and reinfection, and this highlights the need for the development of an effective Ct vaccine. To this end, NIAID organized a workshop to consider the current status of Ct vaccine research and address critical questions in Ct vaccine design and clinical testing. Topics included the goal(s) of a vaccine and the feasibility of achieving these goals, animal models of infection including mouse and nonhuman primate (NHP) models, and correlates of protection to guide vaccine design. Decades of research have provided both whole cell-based and subunit vaccine candidates for development. At least one is currently in clinical development and efforts now need to be directed toward further development of the most attractive candidates. Overall, the discussions and presentations from the workshop highlighted optimism about the current status of Ct vaccine research and detailed the remaining gaps and questions needed to move vaccines forward.

B Cell Presentation of Chlamydia Antigen Selects Out Protective CD4γ13 T Cells: Implications for Genital Tract Tissue-Resident Memory Lymphocyte Clusters.

Surveillance and defense of the enormous mucosal interface with the nonsterile world are critical to protecting the host from a wide range of pathogens. Chlamydia trachomatis is an intracellular bacterial pathogen that replicates almost exclusively in the epithelium of the reproductive tract. The fallopian tubes and vagina are poorly suited to surveillance and defense, with limited immune infrastructure positioned near the epithelium. However, a dynamic process during clearing primary infections leaves behind new lymphoid clusters immediately beneath the epithelium. These memory lymphocyte clusters (MLCs) harboring tissue-resident memory (Trm) T cells are presumed to play an important role in protection from subsequent infections. Histologically, human Chlamydia MLCs have prominent B cell populations. We investigated the status of genital tract B cells during C. muridarum infections and the nature of T cells recovered from immune mice using immune B cells as antigen-presenting cells (APCs). These studies revealed a genital tract plasma B cell population and a novel genital tract CD4 T cell subset producing both gamma interferon (IFN-γ) and interleukin-13 (IL-13). A panel of CD4 T cell clones and microarray analysis showed that the molecular fingerprint of CD4γ13 T cells includes a Trm-like transcriptome. Adoptive transfer of a Chlamydia-specific CD4γ13 T cell clone completely prevented oviduct immunopathology without accelerating bacterial clearance. Existence of a CD4γ13 T cell subset provides a plausible explanation for the observation that human peripheral blood mononuclear cell (PBMC) Chlamydia-specific IFN-γ and IL-13 responses predict resistance to reinfection.

Perspective: my 37 year journey through Chlamydia research: Chlamydia antigen analysis using monoclonal antibodies and major histocompatibility complex molecules.

Chlamydia antigen analysis enables understanding of disease pathogenesis, facilitates development of diagnostic immunoassays and is essential to the design of a subunit Chlamydia trachomatis vaccine. Using an autobiographical narrative, I review over three decades of antigen analysis research findings coming from my research laboratory and provide an outlook to the broader field of Chlamydia seroepidemiology and vaccinology. Based on the experiences of my scientific career I conclude with thoughts for young scientists newly entering the Chlamydia research field.

Update on Chlamydia trachomatis Vaccinology.

Attempts to produce a vaccine to protect against Chlamydia trachomatis-induced trachoma were initiated more than 100 years ago and continued for several decades. Using whole organisms, protective responses were obtained. However, upon exposure to C. trachomatis, disease exacerbation developed in some immunized individuals, precluding the implementation of the vaccine. Evidence of the role of C. trachomatis as a sexually transmitted pathogen started to emerge in the 1960s, and it soon became evident that it can cause acute infections and long-term sequelae in women, men, and newborns. The main focus of this minireview is to summarize recent findings and discuss formulations, including antigens, adjuvants, routes, and delivery systems for immunization, primarily explored in the female mouse model, with the goal of implementing a vaccine against C. trachomatis genital infections.

A double-edged sword: does highly active antiretroviral therapy contribute to syphilis incidence by impairing immunity to Treponema pallidum?

BACKGROUND AND HYPOTHESIS: Recently, the world has experienced a rapidly escalating outbreak of infectious syphilis primarily affecting men who have sex with men (MSM); many are taking highly active antiretroviral therapy (HAART) for HIV-1 infection. The prevailing hypothesis is that HAART availability and effectiveness have led to the perception among both individuals who are HIV-1 infected and those who are uninfected that HIV-1 transmission has become much less likely, and the effects of HIV-1 infection less deadly. This is expected to result in increased sexual risk-taking, especially unprotected anal intercourse, leading to more non-HIV-1 STDs, including gonorrhoea, chlamydia and syphilis. However, syphilis incidence has increased more rapidly than other STDs. We hypothesise that HAART downregulates the innate and acquired immune responses to Treponema pallidum and that this biological explanation plays an important role in the syphilis epidemic. METHODS: We performed a literature search and developed a mathematical model of HIV-1 and T. pallidum confection in a population with two risk groups with assortative mixing to explore the consequence on syphilis prevalence of HAART-induced changes in behaviour versus HAART-induced biological effects. CONCLUSIONS AND IMPLICATIONS: Since rising syphilis incidence appears to have outpaced gonorrhoea and chlamydia, predominantly affecting HIV-1 positive MSM, behavioural factors alone may be insufficient to explain the unique, sharp increase in syphilis incidence. HAART agents have the potential to alter the innate and acquired immune responses in ways that may enhance susceptibility to T. pallidum. This raises the possibility that therapeutic and preventative HAART may inadvertently increase the incidence of syphilis, a situation that would have significant and global public health implications. We propose that additional studies investigating the interplay between HAART and enhanced T. pallidum susceptibility are needed. If our hypothesis is correct, HAART should be combined with enhanced patient management including frequent monitoring for pathogens such as T. pallidum.