Update on Chlamydia trachomatis Vaccinology de la Maza, Luis M; Zhong, Guangming; Brunham, Robert C
Clinical and vaccine immunology,
04/2017, Letnik:
24, Številka:
4
Journal Article
Recenzirano
Odprti dostop
Attempts to produce a vaccine to protect against
-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
, disease exacerbation developed in some immunized individuals, precluding the implementation of the vaccine. Evidence of the role of
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
genital infections.
Chlamydia trachomatis remains one of the few major human pathogens for which there is no transformation system. C. trachomatis has a unique obligate intracellular developmental cycle. The ...extracellular infectious elementary body (EB) is an infectious, electron-dense structure that, following host cell infection, differentiates into a non-infectious replicative form known as a reticulate body (RB). Host cells infected by C. trachomatis that are treated with penicillin are not lysed because this antibiotic prevents the maturation of RBs into EBs. Instead the RBs fail to divide although DNA replication continues. We have exploited these observations to develop a transformation protocol based on expression of β-lactamase that utilizes rescue from the penicillin-induced phenotype. We constructed a vector which carries both the chlamydial endogenous plasmid and an E.coli plasmid origin of replication so that it can shuttle between these two bacterial recipients. The vector, when introduced into C. trachomatis L2 under selection conditions, cures the endogenous chlamydial plasmid. We have shown that foreign promoters operate in vivo in C. trachomatis and that active β-lactamase and chloramphenicol acetyl transferase are expressed. To demonstrate the technology we have isolated chlamydial transformants that express the green fluorescent protein (GFP). As proof of principle, we have shown that manipulation of chlamydial biochemistry is possible by transformation of a plasmid-free C. trachomatis recipient strain. The acquisition of the plasmid restores the ability of the plasmid-free C. trachomatis to synthesise and accumulate glycogen within inclusions. These findings pave the way for a comprehensive genetic study on chlamydial gene function that has hitherto not been possible. Application of this technology avoids the use of therapeutic antibiotics and therefore the procedures do not require high level containment and will allow the analysis of genome function by complementation.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The transmission of the urogenital serovars of
can be significantly influenced by vaginal gels. Hydroxyethyl cellulose is a commonly used gelling agent that can be found in vaginal gels. Hydroxyethyl ...cellulose showed a concentration-dependent growth-enhancing effect on
serovars D and E, with a 26.1-fold maximal increase
and a 2.57-fold increase
.
Spontaneous clearance of
(CT) infections can occur between diagnosis and treatment. We followed CT patients to assess clearance using a conventional definition (no total CT-DNA, assessed by routine ...quantitative PCR methods) and a definition accounting for viability, assessed by viability PCR testing.
Three outpatient STI clinics included CT-diagnosed women (The Netherlands, 2016-2017, FemCure study); participants had vaginal CT (vCT) and rectal CT (rCT) (group A: n=155), vCT and were rectally untested (group B: n=351), single vCT (group C: n=25) or single rCT (group D: n=29). Follow-up (median interval 9 days) vaginal and rectal samples underwent quantitative PCR testing (detecting total CT-DNA). When PCR positive, samples underwent V-PCR testing to detect 'viable CT' (CT-DNA from intact CT organisms; V-PCR positive). 'Clearance' was the proportion PCR-negative patients and 'clearance of viable CT' was the proportion of patients testing PCR negative or PCR positive but V-PCR negative. We used multivariable logistic regression analyses to assess diagnosis group (A-D), age, days since initial CT test (diagnosis) and study site (STI clinic) in relation to clearance and clearance of viable CT.
Clearance and clearance of viable CT at both anatomic sites were for (A) 0.6% and 3.9%; (B) 5.4% and 9.4%; (C) 32.0% and 52.0% and (D) 27.6% and 41.4%, respectively. In multivariate analyses, women with single infections (groups C and D) had higher likelihood of clearance than women concurrently infected with vCT and rCT (p<0.001).Of rectally untested women (group B), 76.9% had total CT-DNA and 46.7% had viable CT (V-PCR positive) at the rectal site.
Of untreated female vCT patients who had CT also at the rectal site, or who were rectally untested, only a small proportion cleared CT (in fact many had viable CT) at their follow-up visit (median 9 days). Among single site infected women clearance was much higher.
NCT02694497.
Chlamydia spp. utilize multiple secretion systems, including the type III secretion system (T3SS), to deploy host-interactive effector proteins into infected host cells. Elucidation of secreted ...proteins has traditionally required ectopic expression in a surrogate T3SS followed by immunolocalization of endogenous candidate effectors to confirm secretion by chlamydiae. The ability to transform Chlamydia and achieve stable expression of recombinant gene products has enabled a more direct assessment of secretion. We adapted TEM-1 β-lactamase as a reporter system for assessment of chlamydial protein secretion. We provide evidence that this system facilitates visualization of secretion in the context of infection. Specifically, our findings provide definitive evidence that C. trachomatis CT695 is secreted during infection. Follow-up indirect immunofluorescence studies confirmed CT695 secretion and indicate that this effector can be secreted at multiple points during the chlamydial developmental cycle. Our results indicate that the BlaM-fusion reporter assay will allow efficacious identification of novel secreted proteins. Moreover, this approach can easily be adapted to enable more sophisticated studies of the secretion process in Chlamydia.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
IFN-β has been implicated as an effector of oviduct pathology resulting from genital chlamydial infection in the mouse model. In this study, we investigated the role of cytosolic DNA and engagement ...of DNA sensors in IFN-β expression during chlamydial infection. We determined that three-prime repair exonuclease-1, a host 3' to 5' exonuclease, reduced IFN-β expression significantly during chlamydial infection using small interfering RNA and gene knockout fibroblasts, implicating cytosolic DNA as a ligand for this response. The DNA sensor cyclic GMP-AMP synthase (cGAS) has been shown to bind cytosolic DNA to generate cyclic GMP-AMP, which binds to the signaling adaptor stimulator of IFN genes (STING) to induce IFN-β expression. We determined that cGAS is required for IFN-β expression during chlamydial infection in multiple cell types. Interestingly, although infected cells deficient for STING or cGAS alone failed to induce IFN-β, coculture of cells depleted for either STING or cGAS rescued IFN-β expression. These data demonstrate that cyclic GMP-AMP produced in infected cGAS(+)STING(-) cells can migrate into adjacent cells via gap junctions to function in trans in cGAS(-)STING(+) cells. Furthermore, we observed cGAS localized in punctate regions on the cytosolic side of the chlamydial inclusion membrane in association with STING, indicating that chlamydial DNA is most likely recognized outside the inclusion as infection progresses. These novel findings provide evidence that cGAS-mediated DNA sensing directs IFN-β expression during Chlamydia trachomatis infection and suggest that effectors from infected cells can directly upregulate IFN-β expression in adjacent uninfected cells during in vivo infection, contributing to pathogenesis.
Timely detection and treatment are important for the control of Chlamydia trachomatis, Neisseria gonorrhoeae, and Trichomonas vaginalis. The objective of this study was to measure the performance of ...the Visby Medical Sexual Health Test, a single-use, point-of-care PCR device.
Women aged 14 years and older who presented consecutively to ten clinical sites across seven US states were enrolled for a cross-sectional, single-visit study. Patients who consented to participate, and who had not used any exclusionary products in the genital area in the previous 48 h, provided self-collected vaginal swabs for testing with the investigational device. Untrained operators received the specimens and ran the device using the guide provided. Specimens had to be run within 2 h of collection to be considered valid. For comparison, patient-infected status was derived by testing clinician-collected vaginal specimens with the Hologic Aptima Combo 2 Assay and Aptima Trichomonas vaginalis Assay, as well as the BD ProbeTec CT/GC Qx Amplified DNA Assay and BD ProbeTec Trichomonas vaginalis Qx Assay. If the results of those assays did not match, the BD MAX CT/GC/TV was used as a tiebreaker. The primary outcomes were the sensitivity and specificity of the investigational device for the detection of C trachomatis, N gonorrhoeae, and T vaginalis compared with patient-infected status.
Between Feb 25, 2019, and Jan 6, 2020, 1585 participants aged between 14 years and 80 years (mean 34·8 SD 14·2) were enrolled. 1555 participants had tests run with the investigational device, of whom 1532 (98·5%) had a valid result on either the first or repeat test. Among the patients with evaluable results (including a determinate patient-infected status), the device had a sensitivity of 97·6% (95% CI 93·2–99·2) and specificity of 98·3% (97·5–98·9) for C trachomatis (n=1457), sensitivity of 97·4% (86·5–99·5) and specificity of 99·4% (98·9–99·7) for N gonorrhoeae (n=1468), and sensitivity of 99·2% (95·5–99·9) and specificity of 96·9% (95·8–97·7) for T vaginalis (n=1449).
This innovative, rapid, easy-to-use, single-use, point-of-care device to detect C trachomatis, N gonorrhoeae, and T vaginalis infections showed excellent sensitivity and specificity, and could represent an important advance in the development of rapid diagnostics for sexually transmitted infections and other infectious diseases.
Division of Microbiology and Infectious Diseases, National Institute of Allergy and Infectious Diseases.
The VD4 region from the Chlamydia trachomatis major outer membrane protein contains important neutralizing B-cell epitopes of relevance for antibody-mediated protection against genital tract ...infection. We developed a multivalent vaccine construct based on VD4s and their surrounding constant segments from serovars D, E, and F. Adjuvanted with cationic liposomes, this construct promoted strong immune responses to serovar-specific epitopes, the conserved LNPTIAG epitope and neutralized serovars D, E, and F. Vaccinated mice were protected against challenge, with protection defined as reduced bacterial numbers in vagina and prevention of pathological changes in the upper genital tract. Adoptive transfer of serum and T-cell depletion experiments demonstrated a dominant role for antibodies and CD4⁺ T cells in the protective immune response. Integrating a multivalent VD4 construct into the sequence of the major outer membrane protein resulted in a protective and broadly neutralizing vaccine. Our findings emphasize the important role of antibodies in protection against Chlamydia trachomatis.
Background. The biology of recurrent or long-term infections of humans by Chlamydia trachomatis is poorly understood. Because repeated or persistent infections are correlated with serious ...complications in humans, understanding these processes may improve clinical management and public health disease control. Methods. We conducted whole-genome sequence analysis on C. trachomatis isolates collected from a previously described patient set in which individuals were shown to be infected with a single serovar over a lengthy period. Results. Data from 5 of 7 patients showed compelling evidence for the ability of these patients to harbor the same strain for 3–5 years. Mutations in these strains were cumulative, very uncommon, and not linked to any single protein or pathway. Serovar J strains isolated from 1 patient 3 years apart did not accumulate a single base change across the genome. In contrast, the sequence results of 2 patients, each infected only with serovar Ia strains, revealed multiple same-serovar infections over 1–5 years. Conclusions. These data demonstrate examples of long-term persistence in patients in the face of repeated antibiotic therapy and show that pathogen mutational strategies are not important in persistence of this pathogen in patients.