Carolyn M. Black

Multiple Drug-Resistant Chlamydia trachomatis Associated with Clinical Treatment Failure

In vitro susceptibility testing and genotyping were done on urogenital isolates of Chlamydia trachomatis from 3 patients, 2 of whom showed evidence of clinical treatment failure with azithromycin and one of whom was the wife of a patient. All 3 isolates demonstrated multidrug resistance to doxycycline, azithromycin, and ofloxacin at concentrations >4.0 microg/mL. Recurrent disease due to relapsing infection with the same resistant isolate was documented on the basis of identical genotypes of both organisms. This first report of clinically significant multidrug-resistant C. trachomatis causing relapsing or persistent infection may portend an emerging problem to clinicians and public health officials.

Suppression of Endogenous IL-10 Gene Expression in Dendritic Cells Enhances Antigen Presentation for Specific Th1 Induction: Potential for Cellular Vaccine Development

A new paradigm for designing vaccines against certain microbial pathogens, including Chlamydia trachomatis, is based on the induction of local mucosal Th1 response. IL-10 is an anti-inflammatory cytokine that exerts negative immunoregulatory influence on Th1 response. This study investigated whether biochemical modulation of endogenous IL-10 expression at the level of APCs is a practical strategy for enhancing the specific Th1 response against pathogens controlled by Th1 immunity. The results revealed that the high resistance of genetically engineered IL-10−/− (IL-10KO) mice to genital chlamydial infection is a function of the predilection of their APCs to rapidly and preferentially activate a high Th1 response. Thus, in microbiological analysis, IL-10KO mice suffered a shorter duration of infection, less microbial burden, and limited ascending infection than immunocompetent wild-type mice. Also, IL-10KO were resistant to reinfection after 8 wk of the primary infection. Cellular and molecular immunologic evaluation indicated that IL-10KO mice induced greater frequency of chlamydial-specific Th1 response following C. trachomatis infection. Moreover, IL-10KO APCs or antisense IL-10 oligonucleotide-treated wild-type APCs were potent activators of Th1 response from naive or immune T cells. Furthermore, both Ag-pulsed dendritic cells from IL-10KO mice and IL-10 antisense-treated dendritic cells from wild-type mice were efficient cellular vaccines in adoptive immunotherapeutic vaccination against genital chlamydial infection. These findings may furnish a novel immunotherapeutic strategy for boosting the Th1 response against T cell-controlled pathogens and tumors, using IL-10-deficient APCs as vaccine delivery agents.

Epidemiology of Human Papillomavirus Infection and Abnormal Cytologic Test Results in an Urban Adolescent Population

We determined the prevalence of and the risk factors for human papillomavirus (HPV) infection and abnormal cytologic test results in 312 adolescent girls (mean age, 16.1 years). Subjects had a median of 2 years of sexual activity and 4 lifetime sex partners. Cervical HPV was detected by use of L1-consensus polymerase chain reaction in 64% of subjects; half of those with HPV had >1 type, and 77% had >/=1 high-risk type. Independent risk factors for HPV were lifetime number of sex partners, age of partner, and douching. Cytologic abnormalities were common (20.9% of subjects had atypical squamous cells of uncertain significance, and 17.0% had high- or low-grade squamous intraepithelial lesions) and were significantly associated with detection of HPV (P=.0001); however, most (51.6%) subjects with HPV had normal cytologic test results.

Antibody regulation of Tcell immunity: implications for vaccine strategies against intracellular pathogens.

Intracellular microbial pathogens cause a plethora of diseases that pose a huge public health challenge. Efficacious prophylactic vaccines are needed to protect the population from this myriad of infectious diseases. Contemporary approaches to vaccine design are guided by the immunobiological paradigm that extracellular pathogens are controlled principally by humoral immunity, involving specific antibodies, whereas host protection against intracellular pathogens requires effectors of cell-mediated immunity. However, this distinct T-helper (Th) type 1 and 2 paradigm of host defense has encountered a major challenge due to the reality that most antigens or vaccines induce mixed immune responses comprising of both humoral and CMI effectors. Besides, the true functional independence of antibodies and T-cells under in vivo physiologic conditions is uncertain. Recent findings have revealed that antibodies exert a significant immunoregulatory effect on T-cell immunity. Thus, a robust and protective T-cell memory response against microbial pathogens such as Chlamydia and Mycobacteria require an effective primary humoral immune response characterized by specific antibody isotypes whose role is to modulate Th1 activation via Fc receptors (FcR) by facilitating a rapid uptake, processing and presentation of pathogen-derived antigens for an enhanced T-cell response. These findings have crystallized into a paradigm shift in host defense wherein different components of the apparently disparate mixed immune responses elicited against a microbial pathogen function concertedly to maximize the principal effector mechanism. This review focuses on the essential role of both arms of the immune system in controlling intracellular microbial pathogens, especially the regulatory role of FcR-mediated antibody function in optimizing the induction of a protective Th1 response. The immunobiological implications are discussed in the context of vaccine design, delivery and evaluation against intracellular microbial pathogens of bacteria, fungi and parasitic origin.

Fc Receptor–Mediated Antibody Regulation of T Cell Immunity against Intracellular Pathogens

Immunity to intracellular microbial pathogens, including Chlamydia species, is controlled primarily by cell-mediated effector mechanisms, yet, the absence of antibodies results in inefficient microbial clearance. We investigated the hypothesis that certain Fc receptor functions promote the rapid induction of elevated T helper type 1 (Th1) response, which effectively clears chlamydiae. FcR(-/-) mice exhibited a delayed and reduced frequency of Chlamydia-specific Th1 cells, compared to FcR(+/+) mice. In vitro, antichlamydial antibodies increased the rate of Th1 activation by FcR(+/+) but not FcR(-/-) antigen-presenting cells. FcR(-/-) dendritic cells and the T cell-associated IgG2A and IgA mediate enhanced Th1 activation by antibodies. Immunization with chlamydia-antibody complexes induced elevated and protective Th1 response. These results provide a mechanistic basis for requiring both T cell and humoral immune responses in protective immunity and vaccine evaluation. Findings offer a paradigm in host defense wherein different effector components function indirectly to maximize the principal effector mechanism.

The natural history of untreated Chlamydia trachomatis infection in the interval between screening and returning for treatment.

Background: Studies of the natural history of genital chlamydial infections in humans are sparse and have had study design limitations. An improved understanding of chlamydial natural history may influence recommendations for elements of control efforts such as chlamydia screening frequency or time parameters for partner notification. Methods: Addressing limitations of prior studies in part, we are prospectively studying chlamydial natural history in sexually transmitted diseases clinic patients in the interval between screening and returning for treatment of positive chlamydial tests. Results of repeat chlamydial testing and clinical outcomes and their associated predictors are being evaluated. Results: In the initial 129 subjects, 89% were female, 88% were black, median age was 21 years, and the median interval between screening and treatment was 13 days. Based on nucleic acid amplification testing at treatment, spontaneous resolution of chlamydia occurred in 18%. Resolution was somewhat more common in subjects with longer intervals between screening and treatment. Persisting infections more often progressed to develop clinical signs at the time of treatment (e.g., urethritis or cervicitis). Two women and one man developed chlamydial complications between screening and treatment. Conclusions: Our findings demonstrate that although spontaneous resolution of chlamydia is common, many persons with persisting chlamydia progress to develop signs of infection and some develop complications.

Head-to-Head Multicenter Comparison of DNA Probe and Nucleic Acid Amplification Tests for Chlamydia trachomatis Infection in Women Performed with an Improved Reference Standard

ABSTRACT Few evaluations of tests for Chlamydia trachomatis have compared nucleic acid amplification tests (NAATs) with diagnostic tests other than those by culture. In a five-city study of 3,551 women, we compared the results of commercial ligase chain reaction (LCR) and PCR tests performed on cervical swabs and urine with the results of PACE 2 tests performed on cervical swabs, using independent reference standards that included both cervical swabs and urethral swab-urine specimens. Using cervical culture as a standard, the sensitivities of PACE 2, LCR, and PCR tests with cervical specimens were 78.1, 96.9, and 89.9%, respectively, and the specificities were 99.3, 97.5, and 98.2%, respectively. Using either cervical swab or urine LCR-positive tests as the standard decreased sensitivities to 60.8% for PACE 2 and to 75.8 and 74.9% for PCR with cervical swabs and urine, respectively. Specificities increased to 99.7% for PACE 2 and to 99.7 and 99.4% for PCR with cervical swabs and urine, respectively. Sensitivities with a cervical swab-urine PCR standard were 61.9% for PACE 2 and 85.5 and 80.8% for LCR with cervical swabs and urine, respectively. Specificities were 99.6% for PACE 2 and 99.0 and 98.9% for LCR with cervical swabs and urine, respectively. Cervical swab versus urine differences were significant only for PCR specificities (P = 0.034). Overall, LCR sensitivity exceeded that of PCR, and sensitivities obtained with cervical swabs exceeded those obtained with urine specimens by small amounts. These data have substantiated, using a large multicenter sample and a patient standard, that LCR and PCR tests performed on endocervical swabs and urine are superior to PACE 2 tests for screening C. trachomatis infections in women. In our study, NAATs improved the detection of infected women by 17 to 38% compared to PACE 2.

Incidence and prevalence of chlamydia, herpes, and viral hepatitis in a homeless adolescent population.

Background High rates of unprotected intercourse and illegal drug use have been reported among homeless adolescents. As a transient population with the potential to act as disease vectors from one location to another, incidence and prevalence of sexually transmitted infections in this population are of particular concern. Goal To assess a homeless adolescent population for incidence and prevalence of Chlamydia trachomatis, herpes simplex virus type 2, hepatitis B virus, hepatitis C virus, HIV, and psychosocial correlates of the acquisition of sexually transmitted infections. Study Design Longitudinal with assessments at baseline, 3 months, and 6 months (n = 536; 319 males and 217 females). Results Baseline prevalence of C trachomatis was 4.17% for males and 6.30% for females. Prevalence of herpes simplex virus type 2 was 5.73% for males and 12.50% for females. Hepatitis B virus and hepatitis C virus prevalences were 3.60% and 5.0%, respectively. HIV seroprevalence was 0.3%. The incidence of sexually transmitted infections was significantly higher among females than among males (16.7% versus 9.8%) and was associated with inconsistent condom use and, for females, number of partners and sex with older partners. Incident hepatitis B virus and hepatitis C virus infection rates were 3.44% and 6.61%, respectively; both were associated with injection drug use. Conclusions Among females, the incidence of herpes simplex virus type 2 (> 25%) and C trachomatis (12%) was relatively high. Inconsistent condom use was the primary factor associated with a significantly greater risk of incident sexually transmitted infections. This was especially true for females with multiple partners. Homeless adolescents also are at high risk for hepatitis B virus and hepatitis C virus infection, primarily associated with self-reported injection drug use.

A Novel Recombinant Multisubunit Vaccine against Chlamydia

The administration of an efficacious vaccine is the most effective long-term measure to control the oculogenital infections caused by Chlamydia trachomatis in humans. Chlamydia genome sequencing has identified a number of potential vaccine candidates, and the current challenge is to develop an effective delivery vehicle for induction of a high level of mucosal T and complementary B cell responses. Vibrio cholerae ghosts (VCG) are nontoxic, effective delivery vehicles with potent adjuvant properties, and are capable of inducing both T cell and Ab responses in mucosal tissues. We investigated the hypothesis that rVCG could serve as effective delivery vehicles for single or multiple subunit chlamydial vaccines to induce a high level of protective immunity. rVCG-expressing chlamydial outer membrane proteins were produced by a two-step genetic process, involving cloning of Omp genes in V. cholerae, followed by gene E-mediated lysis of the cells. The immunogenicity and vaccine efficacy of rVCG-expressing single and multiple subunits were compared. Immunologic analysis indicated that i.m. immunization of mice with either vaccine construct induced a strong mucosal and systemic specific Th1 response against the whole chlamydial organism. However, there was an immunogenic advantage associated with the multiple subunit vaccine that induced a higher frequency of Th1 cells and a relatively greater ability to confer protective immunity, compared with the single subunit construct. These results support the operational theory that the ability of a vaccine to confer protective immunity against Chlamydia is a function of the level of Th1 response elicited.

Population-Based Genetic and Evolutionary Analysis of Chlamydia trachomatis Urogenital Strain Variation in the United States

Chlamydia trachomatis is a major cause of ocular and sexually transmitted diseases worldwide. While much of our knowledge about its genetic diversity comes from serotyping or ompA genotyping, no quantitative assessment of genetic diversity within serotypes has been performed. To accomplish this, 507 urogenital samples from a multicenter U.S. study were analyzed by phylogenetic and statistical modeling. No B, Da, or I serotypes were represented. Based on our analyses, all but one previous urogenital B serotype was identified as Ba. This, coupled with the lack of B serotypes in our population, suggests that B has specific tropism for ocular mucosa. We identified a Ba/D recombinant (putative crossover nucleotide 477; P < 0.0001) similar to a B/D mosaic we described previously from an African trachoma patient. Computational analyses of the Ba/D recombinant indicated that upstream changes were less important for tissue tropism than downstream incorporation of the D sequence. Since most serotypes had nonsynonymous/synonymous ratios of <1.0, the major outer membrane protein, encoded by ompA, has many functional constraints and is under purifying selection. Surprisingly, all serotype groups except for J had a unimodal population structure indicating rapid clonal expansion. Of the groups with a unimodal structure, E and Ia and, to a lesser extent, G and K were prevalent, had infrequent incorporation of mutations, and, compared to other groups, had a relatively greater degree of diversifying selection, consistent with a selective sweep of mutations within these groups. Collectively, these data suggest a diverse evolutionary strategy for different serogroups of the organism.