Chlamydial Antigen Research Articles

CT584 Is Not a Protective Vaccine Antigen against Respiratory Chlamydial Challenge in Mice.

Background:Chlamydia trachomatis is the most prevalent bacterial sexually transmitted pathogen in humans worldwide. Since chlamydial infection is largely asymptomatic with the potential for serious complications, a preventative vaccine is likely the most viable long-term answer to this public health threat. Cell-free protein synthesis (CFPS) utilizes the cellular protein manufacturing machinery decoupled from the requirement for maintaining cellular viability, offering the potential for flexible, rapid, and decentralized production of recombinant protein vaccine antigens. Methods: Here, we use CFPS to produce the full-length putative chlamydial type three secretion system (T3SS) needle-tip protein, CT584, for evaluation as a vaccine antigen in mouse models. High-speed atomic force microscopy (HS-AFM) (RIBM, Tsukuba, Japan) imaging and computer simulations confirm that CFPS-produced CT584 retains a native-like structure prior to immunization. Female mice were primed with CT584 adjuvanted with CpG-1826 intranasally (i.n.) or CpG-1826 + Montanide ISA 720 intramuscularly (i.m.), followed four weeks later by an i.m. boost before respiratory challenge with 104 inclusion forming units (IFU) of Chlamydia muridarum. Results: Immunization with CT584 generated robust antibody responses but weak cell-mediated immunity and failed to protect against i.n. challenge as demonstrated by body weight loss, increased lung weights, and the presence of high numbers of IFUs in the lungs. Conclusion: While CT584 was not a protective vaccine candidate, the speed and flexibility with which CFPS can be used to produce other potential chlamydial antigens make it an attractive technique for antigen production.

Evaluation in mice of cell-free produced CT584 as a Chlamydia vaccine antigen.

Chlamydia trachomatis is the most prevalent bacterial sexually transmitted pathogen worldwide. Since chlamydial infection is largely asymptomatic with the potential for serious complications, a preventative vaccine is likely the most viable long-term answer to this public health threat. Cell-free protein synthesis (CFPS) utilizes the cellular protein manufacturing machinery decoupled from the requirement for maintaining cellular viability, offering the potential for flexible, rapid, and de-centralized production of recombinant protein vaccine antigens. Here, we use CFPS to produce the putative chlamydial type three secretion system (T3SS) needle-tip protein, CT584, for use as a vaccine antigen in mouse models. High-speed atomic force microscopy (HS-AFM) imaging and computer simulations confirm that CFPS-produced CT584 retains a native-like structure prior to immunization. Female mice were primed with CT584 adjuvanted with CpG-1826 intranasally (i.n.) or CpG-1826 + Montanide ISA 720 intramuscularly (i.m.), followed four-weeks later by an i.m. boost before respiratory challenge with 104 inclusion forming units (IFU) of Chlamydia muridarum. Immunization with CT584 generated robust antibody responses but weak cell mediated immunity and failed to protect against i.n. challenge as demonstrated by body weight loss, increased lungs' weights and the presence of high numbers of IFUs in the lungs. While CT584 alone may not be the ideal vaccine candidate, the speed and flexibility with which CFPS can be used to produce other potential chlamydial antigens makes it an attractive technique for antigen production.

Stability and antigenicity of Chlamydia muridarum major outer membrane protein antigen at body temperature

Chlamydia is an obligate intracellular bacterial pathogen responsible for disease and infertility across multiple species. Currently vaccines are being studied to help reduce the prevalence of this disease. The main advantage of protein subunit vaccines is their high degree of safety although this is traded off with the requirement for multiple booster doses to achieve complete protection. Although in certain populations the booster dose can be difficult and costly to administer, development of delayed vaccine delivery techniques, such as a vaccine capsule, could be the solution to this problem. One of the main drawbacks in this technology is that the antigen must remain stable at body temperature (37 °C) until release is achieved. Here we elucidate the stability of a recombinant chlamydial major outer membrane protein (MOMP) antigen and assess its antigenic and immunogenic properties after subjecting the antigen to 37 °C for four to six weeks. Through in vitro and in vivo assessment we found that the aged chlamydial MOMP was able to produce equivalent humoral and cell-mediated immune responses when compared with the unaged vaccine. It was also found that vaccines formulated with the aged antigen conferred equivalent protection against a live infection challenge as the unaged antigen. Thus ageing chlamydial MOMP antigens at 37 °C for four to six weeks did not cause any significant structural or antigenic/immunogenic degradation and recombinant C. muridarum MOMP is suitable for use in a delayed vaccine delivery system.

Preclinical screen for protection efficacy of chlamydial antigens that are immunogenic in humans.

To search for subunit vaccine candidates, immunogenic chlamydial antigens identified in humans were evaluated for protection against both infection and pathology in a mouse genital tract infection model under three different immunization regimens. The intramuscular immunization regimen was first used to evaluate 106 chlamydial antigens, which revealed that two antigens significantly reduced while 11 increased genital chlamydial burden. The two infection-reducing antigens failed to prevent pathology and 23 additional antigens even exacerbated pathology. Thus, intranasal mucosal immunization was tested next since intranasal inoculation with live Chlamydia muridarum prevented both genital infection and pathology. Two of the 29 chlamydial antigens evaluated were found to prevent genital infection but not pathology and three exacerbate pathology. To further improve protection efficacy, a combinational regimen (intranasal priming + intramuscular boosting + a third intraperitoneal/subcutaneous boost) was tested. This regimen identified four infection-reducing antigens, but only one of them prevented pathology. Unfortunately, this protective antigen was not advanced further due to its amino acid sequence homology with several human molecules. Two pathology-exacerbating antigens were also found. Nevertheless, intranasal mucosal priming with viable C. muridarum in control groups consistently prevented both genital infection and pathology regardless of the subsequent boosters. Thus, screening 140 different chlamydial antigens with 21 repeated multiple times in 17 experiments failed to identify a subunit vaccine candidate but demonstrated the superiority of viable chlamydial organisms in inducing immunity against both genital infection and pathology, laying the foundation for developing a live-attenuated Chlamydia vaccine.

Designing a Histological Analyzer for Diagnosing Pathomorphological Changes in Tissues as an Example of Chlamydial Infection

The article is devoted to the development of a device to study tissue destruction caused by damage to the histo-hematic barriers of the body, under the influence of chlamydial infection. Cell pathology refers to changes in its components and ultrastructures with causal relationships. Chlamydiacea is a spectrum of diseases that, because of their polymorphism, cannot be united by a specific symptom complex, and sometimes affect all systems and organs. Due to the lack of organotropy and host specificity of the different representatives of chlamydiae, the clinic of chlamydiae is extremely diverse. The pathological process in chlamydial infections may localize in many organs, thus causing pathomorphologic changes in various body structures. The complex of adequate and modern methods of investigation makes it possible to evaluate the changes occurring in the macroorganism at the cellular and ultrastructural level. Emerging dystrophic, dyscirculatory, inflammatory processes in general, while not specific for chlamydia, are complemented by signs pathognomonic for this infection (presence of chlamydial antigens in cells in immunohistochemical method of study, detection of chlamydial structures in cells in electron microscopy). Currently, automation and robotization of research are penetrating all areas of medicine and veterinary medicine, including histological analysis. Currently, in the preparation of histological preparation, the technological process is automated in a fragmented way. The development of a histology robot will help to solve the problem of the shortage of highly qualified histology lab technicians and pathologists and reduce the burden on medical personnel in general. Processes of automation and modeling of technological flows and resources in the preparation of histological images and acceptance of the diagnosis in medicine and veterinary medicine is an urgent tasks. In order to identify pathological processes at the cellular level, as well as to reduce the error in the performance of histological manipulations, approaches to the design of a histological robot were developed. The model of the express analyzer structurally consists of two modules: a histological image preparation module and a pathology recognition module. Laboratory experiments were carried out to identify indicators of pathologies. Software for programmed OMRON controllers has been developed. Analysis of the simulation of the circuit operation showed positive results. The probability of pathology recognition was 0.8-0.9.

Effect of chlamydia trachomatis on infertile women

Objective: This study was carried out to determine the seroprevalence of Chlamydia trachomatis in hospitalized women has been made. Materials and methods: Chlamydial antigen and anti-chlamydial antibodies were investigated in 145 patients who applied to Al Khanasaa teaching hospitals in City of Mosul-Iraq and were diagnosed as infertile, and in 81 women who gave birth. Results: Endocervical chlamydial antigen in one (0.7%) patient in the infertile group; While anti-chlamydial antibody was positive in 7 (4.8%) patients, endocervical chlamydial antigen and anti-chlamydial antibody positivity were not detected in fertile cases. Conclusion: Seropositivity of 5.5% (seven antibody positive and one antigen positive case) was obtained in 145 infertile women; In infertile cases, antibody positivity intensifies between the ages of 25-35; It was determined that a case with positive chlamydial antigen was 20 years old.

Effect of chlamydia trachomatis on infertile women

Objective: This study was carried out to determine the seroprevalence of Chlamydia trachomatis in hospitalized women has been made. Materials and methods: Chlamydial antigen and anti-chlamydial antibodies were investigated in 145 patients who applied to Al Khanasaa teaching hospitals in City of Mosul-Iraq and were diagnosed as infertile, and in 81 women who gave birth. Results: Endocervical chlamydial antigen in one (0.7%) patient in the infertile group; While anti-chlamydial antibody was positive in 7 (4.8%) patients, endocervical chlamydial antigen and anti-chlamydial antibody positivity were not detected in fertile cases. Conclusion: Seropositivity of 5.5% (seven antibody positive and one antigen positive case) was obtained in 145 infertile women; In infertile cases, antibody positivity intensifies between the ages of 25-35; It was determined that a case with positive chlamydial antigen was 20 years old.

Immunoassay to categorize chlamydial inflammatory disease in women to develop biomarker from peptide derived stimulant and its response to cytokine

Chlamydia trachomatis infection in women can result in serious sequelae such as pelvic inflammatory disease and infertility. Previous research has suggested that peptide epitopes from HtrA and HSP60 are potential biomarkers for immune cell reactions. Thus, in this study we tested the hypothesis that these peptides could be used for future development of an immunoassay to detect at-risk women of pathology by a profile of the cytokine response. Our findings indicate that some chlamydial peptide antigens stimulated cervical mononuclear cell production of IL-6 and IFN-y resulting from cervical cell proliferation from women with pelvic inflammatory disease (PID) and/or cervicitis to a greater extent than that observed in negative control cohorts. Proliferative response of cervical cells induced by peptide antigens strongly reduces IL-10 concentration. We evaluated the suitability of different pairs of peptide antigens for the predictive value of chlamydial infection based on the cytokine induction. Whilst the cytokine response was not statistically significant, the data suggests that an immunoassay may be possible to develop in future for early detection of serious chlamydial infection sequelae.

New Possibilities of Antibiotic Therapy for Blepharoconjunctivitis in Children

The problem of drug therapy for bacterial eye infections in children has remained relevant for many years. The greatest interest of ophthalmologists in recent years is associated with the use of fluoroquinolones in the treatment of inflammatory eye diseases of the bacterial etiology. At the same time, new ophthalmic dosage forms of fluoroquinolones that have appeared in recent years naturally require additional research on their effectiveness.Objective: to study the clinical efficacy of the antibacterial drug Oftocypro (0.3 % cyprofloxacin in ophthalmic ointment) in the treatment of chronic blepharoconjunctivitis in children.Materials. The study involved 38 children aged 3 to 18 years (mean age 10.3 ± 2.7 years) with clinical manifestations of blepharoconjunctivitis. All patients were divided into 2 groups of equal size: 18 children (36 eyes) — with bacterial blepharoconjunctivitis and 20 (40) — with chlamydial blepharoconjunctivitis. The diagnosis was verified based on the clinical picture of blepharoconjunctivitis and laboratory data: detection of pathogenic microflora in the conjunctival cavity of patients of the first group and chlamydia antigen — in epithelial cells in scraping material from the conjunctiva by immunohistochemical analysis.Results. The analysis of the data obtained during the examination and treatment of children with blepharoconjunctivitis of bacterial etiology (group I), a reliable dynamics of controlled clinical and laboratory parameters was established. There was a significant positive dynamics of all controlled parameters of the clinical course of chronic bacterial blepharoconjunctivitis against the background of the drug Oftocipro ophthalmic ointment 0.3 % use. All children with chronic chlamydial blepharoconjunctivitis with the background of treatment with Oftocypro, ophthalmic ointment 0.3 %, showed a steady tendency towards relief of the estimated clinical signs of the disease. According to the results of laboratory studies, it was found that on the 28th day of treatment with Oftocypro chlamydia in the cells of the epithelium of the conjunctiva was re-detected in 4 out of 20 patients (20 %).Conclusion. The high efficacy of the drug Oftocypro ophthalmic ointment 0.3 %, in combination with the absence of pronounced side effects, makes it possible to recommend this drug for wider practical use.

Chlamydia trachomatis Cross-Serovar Protection during Experimental Lung Reinfection in Mice.

Chlamydia trachomatis causes most bacterial sexually transmitted diseases worldwide. Different major outer membrane proteins (MOMPs) define various serovars of this intracellular pathogen: In women, D to L3 can cause urethritis, cervicitis, salpingitis, and oophoritis, and, thus, infertility. Protective immunity might be serovar-specific since chlamydial infection does not appear to induce an effective acquired immunity and reinfections occur. A better understanding of induced cross-serovar protection is essential for the selection of suitable antigens in vaccine development. In our mouse lung infection screening model, we evaluated the urogenital serovars D, E, and L2 in this regard. Seven weeks after primary infection or mock-infection, respectively, mice were infected a second time with the identical or one of the other serovars. Body weight and clinical score were monitored for 7 days. Near the peak of the second lung infection, bacterial load, myeloperoxidase, IFN-γ, and TNF-α in lung homogenate, as well as chlamydia-specific IgG levels in blood were determined. Surprisingly, compared with mice that were infected then for the first time, almost independent of the serovar combination used, all acquired parameters of disease were similarly diminished. Our reinfection study suggests that efficient cross-serovar protection could be achieved by a vaccine combining chlamydial antigens that do not include nonconserved MOMP regions.

Retrospective Analysis of Drug Sensitivity of Neisseria gonorrhoeae in Teaching Hospitals of South China.

The aim of this study was retrospective analysis of drug sensitivity of Neisseria gonorrhoeae in two teaching hospitals of South China. A total of 304 Neisseria gonorrhoeae isolates obtained from patients in South China from 2016 to 2020 were evaluated. The MICs of penicillin, cefuroxime, ceftriaxone (CRO), cefepime, ciprofloxacin, ceftazidime and azithromycin (AZM) against the isolates were determined by the agar dilution method. Then, Neisseria gonorrhoeae isolates were categorized into sensitive, moderately sensitive and resistant according to MICs. Also, β-lactamases were detected by enzyme linked immunosorbent assay (ELISA). Ureaplasma urealyticum and Mycoplasma hominis were determined by culture in liquid medium, and Chlamydia was detected by rapid antigen test. The result showed there was 50.99%, 20.72%, 9.87%, 14.47%, 86.84%, 7.57%, 6.91%, 11.18% resistance to penicillin, cefuroxime, ceftriaxone, cefepime, ciprofloxacin, ceftazidime and azithromycin, respectively. Also, β-lactamase positivity was 53.29% and Chlamydia antigen positivity was 20.07%. Ureaplasma urealyticum and Mycoplasma hominis positivity was 11.84% and 6.25%, respectively. From 2016 to 2020, the resistant rate of ceftriaxone and azithromycin gradually increased. In conclusion, Southern China is among the area reporting gonococci with high-level resistance to AZM and CRO, so N. gonorrhoeae culture and drug sensitivity test will be vital for monitoring trends in antimicrobial resistance.

Intranasal immunization with inactivated chlamydial elementary bodies formulated in VCG-chitosan nanoparticles induces robust immunity against intranasal Chlamydia psittaci challenge

Vaccines based on live attenuated Chlamydia elementary bodies (EBs) can cause disease in vaccinated animals and the comparably safer inactivated whole EBs are only marginally protective. Recent studies show that a vaccine formulation comprising UV-inactivated EBs (EB) and appropriate mucosal delivery systems and/or adjuvants induced significant protective immunity. We tested the hypothesis that intranasal delivery of UV-inactivated C. psittaci EB formulated in Vibrio cholerae ghosts (VCG)-chitosan nanoparticles will induce protective immunity against intranasal challenge in SPF chickens. We first compared the impact of VCG and CpG adjuvants on protective immunity following IN mucosal and IM systemic delivery of EB formulated in chitosan hydrogel/microspheres. Immunologic analysis revealed that IN immunization in the presence of VCG induced higher levels of IFN-γ response than IM delivery or the CpG adjuvanted groups. Also, vaccine efficacy evaluation showed enhanced pharyngeal bacterial clearance and protection against lung lesions with the VCG adjuvanted vaccine formulation, thereby establishing the superior adjuvanticity of VCG over CpG. We next evaluated the impact of different concentrations of VCG on protective immunity following IN mucosal immunization. Interestingly, the adjuvanticity of VCG was concentration-dependent, since protective immunity induced following IN mucosal immunization showed dose-dependent immune responses and protection. These studies reveal that formulation of inactivated chlamydial antigens with adjuvants, such as VCG and chitosan increases their ability to induce protective immune responses against challenge.

A comparative study of prevalence of Chlamydia trachomatis infection among infertile and fertile women at a tertiary care center

Background and Aim: Chlamydia trachomatis is an obligate intracellular Gram-negative bacterium causing sexually transmitted infection leading to urogenital infections which are asymptomatic. If untreated, leads to complications such as chronic pelvic pain, inflammation, and occlusion of the fallopian tubes, resulting in infertility and ectopic pregnancy. The association is found between C. trachomatis infection and female infertility with a prevalence rate of 15%–30%. The aim was to determine the prevalence of C. trachomatis infection in women with infertility. Materials and Methods: This was a hospital-based cross-sectional study of patients presenting with or without infertility. Group A (75 cases) cases with infertility with or without symptoms of pelvic inflammatory disease. Group B (75 cases) with no infertility matching age, symptoms with Group A. After history, physical examination cervical swab for chlamydial antigen and serology sample was taken and processed. Positive cases were followed by hysterosalpingography (HSG) and laparoscopy. Results: Out of 150 patients, 14 were positive for chlamydia in cases and 4 in the control group. Chlamydial prevalence was four times more in cases as compared to the control group, which is significant. The mean age was 27.97 + 4.520 years. Out of 14 positive cases, 8 (57.14%) had symptoms, whereas 6 (42.85%) were asymptomatic; in controls, 2 were symptomatic, whereas 2 were asymptomatic. HSG and laparoscopy were positive in chlamydia positive cases. Conclusion: It can be presumed that there is a significant role of C. trachomatis in infertility and also there is an association between chlamydia antigen detection and tubal factor infertility.

MICROBIOLOGICAL PROFILE OF VAGINOSIS AMONG WOMEN OF THE REPRODUCTIVE AGE GROUP, WHO ATTENDED AT DMC, LAHERIASARAI, BIHAR

Background: An abnormal vaginal discharge is a common complaint in women and it can be due to vaginal infections like bacterial vaginosis, candidiasis and trichomoniasis. Aim: A descriptive cross sectional study was done at Microbiology Department, DMC, Laheriasarai, Bihar to find out the common pathogens which caused vaginosis in patients of the reproductive age group, who attended the Obstetrics and Gynaecology Department at the Darbhanga Medical College and Hospital, Laheriasarai, Bihar and referred to Microbiology Department of DMC, from 1st April to 30th September, 2020. Materials and Methods: All the 140 vaginal discharge samples were subjected to gram staining to view the morphological nature of the bacteria which caused bacterial vaginosis and gram positive, budding yeast cells, wet film for the motility of Trichomonas and Chlamydial antigen detection by ELISA. Results: The overall positivity was 44%, which included bacterial vaginosis (36.4%). The positivity of candidiasis was 4.2%, that of trichomoniasis was 2.1% and that of chlamydiasis was 1.4%. Conclusion: Bacterial vaginosis was found to be the commonest cause of the abnormal vaginal discharge in women of the reproductive age group.

Intranasal vaccination with a Chimeric Chlamydial Antigen BD584 confers protection against Chlamydia trachomatis genital tract infection

<em>Chlamydia</em> is the most common sexually transmitted infection of bacterial origin, and a broadly protective vaccine is urgently needed given the largely asymptomatic nature of the infection and the severe reproductive sequelae in women with untreated infections.

NK Cells Contribute to Protective Memory T Cell Mediated Immunity to Chlamydia muridarum Infection.

We previously reported that NK cells can promote type 1 T cell immune response that is essential for protection to a primary infection of Chlamydia muridarum. In this study, we have investigated the contribution of NK cells to memory T cells associated immunity during chlamydial infection. We have found that NK cell depletion led to impaired production of IFN-γ by memory T cells upon re-stimulation with chlamydial antigens in vitro. Mice with depleted NK cells also exhibited reduced type 1 T cell recall responses, with increased production of IL-4 from CD4+ T cells and a lower level of Chlamydia-specific IgG2a titers compared to control mice. In addition, Tregs response was significantly increased in mice with NK cell depletion. Moreover, NK cell-depleted mice showed an increased bacterial loads and more severe inflammatory pathological changes than control mice. These findings indicate that NK cells contribute to protective memory T cell associated immunity to chlamydial re-infection through modulating the cytokine pattern of T cell and inhibition of Tregs response.

MON-LB123 Diagnosis and Management of Euglycemic DKA in the Setting of SGLT2 Inhibitor Use and Prostate Abscess

Introduction: SGLT2i has been associated with euglycemic DKA by increasing lipid oxidation and glucagon synthesis. In the event of an underlying infection, increased ketogenesis could lead to the presentation of DKA. Clinical case: A 35 year old male patient who was recently diagnosed with type 2 Diabetes mellitus after he presented to his primary care physician with increased urinary frequency, frothy urine. He was found to have a hemoglobin A1c of 12.5% and blood glucose 408 mg/dl. Urinalysis was negative for nitrites and leukocyte esterase. Patient endorsed to have one sexual partner and safe sexual practices. Urinary gonorrhea and chlamydia antigens were negative. He was diagnosed with diabetes mellitus and was started on ertugliflozin, exenatide and bactrim for possible UTI. Three days later, the patient presented to the ED with dysuria, polyuria, rectal pain, nausea and fatigue. Patient appeared to be in distress with tachypnea and tachycardia. Physical exam showed signs of dehydration. Laboratory evaluation revealed blood glucose at 185 mg/dL. Given the history of recent diagnosis of diabetes mellitus and SGLT2 inhibitor use, euglycemic DKA was suspected. ABG revealed metabolic acidosis (arterial blood gas pH 7.32, pCo2 20, pO2 130) with elevated anion gap of 24, HCO3 level of 10 mmol/L (reference range 22 - 28 mmol/L) and beta-hydroxybutyrate of 5.7 mmol/L (reference range 0 - 0.3 mmol/L). The lactate levels were normal. Wbc count 20,100. Urinalysis showed glucose 4+, ketones 4+, WBC 4, negative for infection. Computed tomography scan of his abdomen and pelvis with contrast showed 3.7 x 2.7 cm prostate abscess and acute cystitis. Patient was treated for euglycemic DKA with infusion of insulin and dextrose 10% with 0.45% NS. Careful monitoring of his blood sugar was required as it dropped with slight increase in infusion rate. He was started on ciprofloxacin for prostate abscess. Patient’s metabolic acidosis resolved and he showed clinical improvement. Endocrinologist was on consult. Patient was switched to insulin regimen for glycemic control. Conclusion: SGLT2i class of drugs has been increasingly used in the treatment of diabetes due to improved cardiovascular outcomes and renal protective effects. However, diagnosing DKA can be a concern while treating patients with this drug. While the SGLT2i is responsible for his euglycemic presentation, DKA was likely triggered by his prostate abscess. They can be used safely with extensive education to the physicians about euglycemic DKA presentation. Management requires starting D10 infusion along with insulin drip to avoid the risk of hypoglycemia.

Proteome array of antibody responses to Chlamydia trachomatis infection in nonhuman primates

AimsNonhuman primates have been used to investigate pathogenic mechanisms and evaluate immune responses following Chlamydia trachomatis inoculation. This study aimed to systemically profile antibody responses to C. trachomatis infection in nonhuman primates. Materials and methodsSera were obtained from 4 pig-tailed and 8 long-tailed macaques which were intravaginally or ocularly infected with live C. trachomatis organisms, and analyzed by C. trachomatis proteome array of antigens. Key findingsThe sera from 12 macaques recognized total 172 C. trachomatis antigens. While 84 antigens were recognized by pig-tailed macaques intravaginally infected with serovar D strain, 125 antigens were recognized by long-tailed macaques ocularly infected with serovar A, and 37 antigens were recognized by both. Ocular inoculation with virulent A2497 strain induced antibodies to more antigens. Among the antigens uniquely recognized by A2497 strain infected macaques, outer membrane complex B antigen (OmcB) induced robust antibody response. Although macaques infected by less virulent A/HAR-13 strain failed to develop antibodies to OmcB, reinfection by A2497 strain induced high levels of antibodies to OmcB. SignificanceProteome array has revealed a correlation of chlamydial infection invasiveness with chlamydial antigen immunogenicity, and identified antibody responses to OmcB potentially as biomarkers for invasive infection with C. trachomatis.

A Novel LNP-Based &amp;lt;i&amp;gt;Chlamydia&amp;lt;/i&amp;gt; Subunit Vaccine Formulation That Induces Th1 Responses without Upregulating IL-17 Provides Equivalent Protection in Mice as Formulations That Induced IL-17 and Th1 Cytokines

We evaluated novel Chlamydial vaccines, consisting of major outer membrane protein (MOMP) alone or in combination with polymorphic membrane proteins D (PmpD) and G (PmpG) using a C57BL/6 mouse model. Native MOMP (nMOMP) isolated from C. muridarum elementary bodies (EBs) and recombinant PmpD and PmpG proteins were adjuvanted with Monophosphoryl lipid A (MPLA), with either lipid nanoparticles (LNPs) or the cationic lipid dimethyldioctadecylammonium bromide (DDA). Antibody titers to C. muridarum nMOMP, and EBs were evaluated by ELISA, and T-cell responses were analyzed by intracellular cytokine staining (ICS). Protection from challenge was determined by qPCR. Vaccine immunized mice showed significantly higher antibody titers to nMOMP (P C. muridarum EBs (P C. muridarum EBs and PmpG. ICS analysis showed more robust CD4 + T-cell responses (IFN-γ/IL-2/TNF-a) in the DDA and LNP groups compared to the adjuvant alone group. The DDA + MPLA gave robust Th17 responses in comparison to MPLA and LNP group. Mice immunized with Chlamydia antigens also showed protection from C. muridarum challenge, by reduction in bacterial shedding for all groups (P < 0.003) compared to shedding from the adjuvant control. Both vaccine formulations generated robust immunological responses, and both were protective by reducing bacterial shedding after challenge. This data indicates equal protection can be achieved without the induction of Th17 responses.

Multi-peptide ELISAs overcome cross-reactivity and inadequate sensitivity of conventional Chlamydia pneumoniae serology

Cross-reactivity of classical chlamydial antigens compromises Chlamydia (C.) pneumoniae serology. By testing with 185 human antisera, we expanded 18 previously discovered C. pneumoniae-specific B-cell epitopes to 48 peptide antigens from 12 C. pneumoniae immunodominant proteins. For specific detection of antibodies against C. pneumoniae, we developed novel ELISAs with strongly reactive individual peptide antigens and mixtures of these peptides. By comparison to a composite reference standard (CRS) for anti-C. pneumoniae antibody status of human sera, the top-performing CpnMixF12 peptide assay showed 91% sensitivity at 95% specificity, significantly higher than 4 commercial anti-C. pneumoniae IgG ELISAs (36-12% sensitivity at 95% specificity). Human C. pneumoniae (Cpn) and C. trachomatis (Ctr) seroreactivity was 54% biased towards co-positivity in commercial Cpn and Ctr ELISAs, but unbiased in Cpn and Ctr peptide antibody assays, suggesting severe cross-reactivity of commercial ELISAs. Using hyperimmune mouse sera against each of 11 Chlamydia spp., we confirm that commercial Cpn and Ctr ELISA antigens are cross-reactive among all Chlamydia spp., but Cpn and Ctr peptide antigens react only with antisera against the cognate chlamydial species. With simultaneously high specificity and sensitivity, and convenient use for non-specialized laboratories, these ELISAs have the potential to improve serodiagnosis of C. pneumoniae infection.