THE NATURAL HISTORY of genital infections with Chlamydia trachomatis is poorly defined. In particular, the duration of untreated infections remains unknown. In a widely cited study, McCormick1 reported that 50% of 14 women with untreated genital chlamydial infections were infected when retested after 16 to 17 months. Based on this finding, some authors have used 15 months as the median duration of infection2,3; however, no consensus exists on this subject.
Better defining the duration of genital chlamydial infections has important implications. From a clinical perspective, knowing how long people remain infected would aid in patient counseling and partner notification. A better estimate of the duration of infection would also be essential in the assessment of the burden of chlamydial disease and the potential impact of different public health interventions. The duration of infectivity is one of the critical factors that determine the extent to which an infectious disease spreads in a population.4 Recent mathematical modeling studies have estimated the incidence of genital chlamydial infections and the comparative impact of partner-notification and screening programs. These studies used estimates of duration that varied from 100 to 450 days in men and from 185 to 450 days in women.3,5,6 Given the uncertainty of how long infections persist, findings derived from such models must be considered tenuous.
We reviewed the scientific literature to summarize what is known about the duration of and the factors that affect untreated genital chlamydial infections. Our interest in performing this review was to define the duration of infection as it relates to disease control. As a result, we briefly review cell-culture findings as they relate to the duration of infection, and concentrate on human and animal studies that shed light on how long people remain infectious.
We conducted a MEDLINE search combining the subject headings chlamydia, C trachomatis infection, and the key word chlamydia. The results were sequentially combined with results of searches using the subject headings spontaneous remission, follow-up studies, time, time factors, and randomized trial to identify articles with both subject headings. We also searched the key words resolution and follow-up, and combined them with articles with chlamydial subject headings. All subject-heading searches used exploded terms. Both human and animal studies were reviewed.
Article titles and abstracts were reviewed for studies in which untreated or inadequately treated subjects underwent serial chlamydial tests. Relevant articles were then reviewed for information on the duration of chlamydial infection. References from articles were reviewed by hand for additional sources.
Leading chlamydial researchers were also contacted regarding unpublished data on the duration of chlamydial infection and the availability of data for a pooled analysis.
Studies in Cell-Culture Systems
Numerous studies have documented persistent chlamydial infections in cell-culture systems.7 Exposure to nutrient-deficient media, antimicrobials, and τ interferon can each interfere with the normal maturation of reticulate bodies to elementary bodies in culture. The result is the “perpetuation of chlamydiae within the host cell without overt growth or replication,” which has been termed a microbiologically inapparent state.7 Removal of these factors is associated with the resumption of normal chlamydial proliferation. This observation suggests that chlamydiae can be persistent, nonreplicating, and uncultivatable, but have the potential for reactivation. Whether this cell-culture phenomena occurs in human infections is unknown. If chlamydiae do enter such a quiescent state, the duration of potentially transmissible infection may be much longer than previously estimated.
Studies using various animal models and strains of chlamydiae have serially tested animals to measure the duration of infection, have used immunosuppression and nonculture techniques to look for evidence of persistent infection beyond the period of culture positivity, and have investigated the effects of previous chlamydial exposure on duration of infection. The results of studies in which animals were serially tested for the presence of chlamydiae following experimental infection are summarized in Table 1.8–22 With the exceptions of the study by Beale10 and several animals subjects in studies by Gale20 and Johnson,16 these studies repeatedly tested animals at fixed intervals. Thirteen of 16 studies used either culture or culture and immunofluorescent staining to assess the presence of infection. Seven studies required more than one sequential negative diagnostic test to define clearance of infection. In the remaining nine reports, the number of negative tests used to define the absence of infection was not stated or the study ended when the last animal had a single negative culture. Eight studies quantitated the intensity of infections using inclusion-forming units.
From existing animal data, it appears that primates experience more prolonged infections than either guinea pigs or mice. In mice and guinea pigs, studies demonstrate a fairly orderly decline in inclusion-forming units leading to negative cultures.9,10,15 Conversely, small studies of primates showed intermittent culture positivity, and sporadically elevated inclusion counts were common.18,21 Within the limits defined by the sensitivity of the tests used, most animals seemed to resolve infections spontaneously. However, because of the often sporadic nature of culture positivity in primates it is difficult to determine if infections cleared, even when measured by culture. Data in male primates are particularly limited, with no study including more than three animals.
Several investigators have immunosuppressed animals to determine whether viable, potentially transmissible chlamydiae persist beyond the period of culture positivity. In an experiment by Beale,10 75% of mice exposed to the mouse pneumonitis biovar of C trachomatis remained positive by polymerase chain reaction (PCR) 42 days after infection despite being culture negative. To assess whether this PCR positivity reflected the existence of viable organisms, Beale immunosuppressed animals but was not able to reinduce culture positivity. Johnson17 also was unable to reactivate culture-positive infections by subjecting marmosets to cyclophosphamide 76 days after infection. However, Cotter23 demonstrated that immunosuppression can reactivate vaginal chlamydial shedding in mice when they are immunosuppressed 2 to 3 weeks after a first-negative culture. In his experiment, mice exposed to immunosuppression at longer intervals after infection did not generally become culture positive. Reactivation of culture-positive infection after immunosuppression has also been demonstrated in guinea pigs.24 These findings suggest that the period of potential infectivity may exceed that of culture positivity. The fact that investigators were only able to reinduce culture positivity for a short period after the first negative chlamydial culture suggests that immunosuppression allowed organisms replicating below the threshold of detection by culture to proliferate above that threshold.
Studies using nonculture techniques have also investigated whether chlamydiae persist beyond the period of culture positivity. Using a pocket model of chlamydial infection in monkeys, Cappuccio25 demonstrated evidence of chlamydiae by in situ hybridization 28 days after experimental inoculation. All animals in the experiment were persistently culture negative starting 3 days after infection. Studying macaques, Patton26 used in situ hybridization and immunocytochemistry to look for evidence of infection in cervical, endometrial, and fallopian tube specimens taken 12 to 16 weeks after infection. More than half of the animals had evidence of chlamydiae by in situ hybridization or immunocytochemistry despite negative cultures, and, in most cases, negative testing by ligase chain reaction. Of note, half of the animals treated with doxycycline also had evidence of persistent organisms. Whether the presence of chlamydial DNA in the upper genital tract observed in these studies is indicative of the presence of transmissible infection is uncertain.
Numerous investigators have taken previously infected animals and have reexposed them to chlamydial species to assess the host's susceptibility to reinfection and the effect of previous exposure on the duration of infection. In mice,8 guinea pigs,14,27,28 female marmosets,17 pig-tail macaques,18 and male baboons,21 the reported duration of culture-positive infection is shorter when previously infected animals are reexposed to chlamydial species.
Human studies have limitations that are distinct from those that affect animal studies. Subjects' onset of infection is generally unknown, reexposure to infections is common, and reported results generally reflect the findings of a single follow-up culture. As a result, duration of infection before testing is unknown, clearance of infection in most studies is even less certain than in animal studies, and reported findings reflect the natural history of infections as they may be modified by reexposure to chlamydiae. To the extent that many studies are based on asymptomatic cases detected through screening, findings are also subject to length-time bias, the tendency of cross-sectional studies to disproportionately identify cases of long duration.
We identified seven studies that report the duration of untreated genital chlamydial infection in women.1,29–35 The results of these studies are presented in Table 2. The two largest studies of untreated women were reported by Rahm32,33 and Parks.35 Rahm et al prospectively followed 109 young asymptomatic women with genital C trachomatis diagnosed by screening culture. During a 10-12 week follow-up period, 16 women (15%) developed symptoms that required treatment. Of 85 asymptomatic untreated women available for repeat culture, 68 (80%) remained culture positive. Ten of 17 culture-negative women were recultured within 6 months, and all remained culture negative.
More recently, Parks et al35 reviewed all cases of chlamydial infection diagnosed by culture at the `Jefferson County STD clinic in Alabama. They identified instances in which patients were untreated and retested within 45 days. Most study subjects (86%) were recultured within 20 days. The accuracy of all negative follow-up cultures was verified using chlamydial PCR of frozen specimens. At time of repeat testing, 28% of women were both culture and PCR negative, suggesting that the infections were spontaneously cleared.
It is surprising that Parks et al found a slightly higher rate of spontaneous clearance than Rahm et al, despite a much shorter follow-up period and the use of more sensitive testing to detect persistent infection. In part, this disparity may reflect differences in the populations studied. Rahm studied asymptomatic women, 94% of whom were younger than 20 years. In contrast, the majority of Park's subjects were symptomatic, and only 21% were younger than 20 years. Higher clearance rates were associated with older age, a finding that is consistent with animal and human data suggesting that partial immunity may diminish the duration of infection in subjects previously exposed to C trachomatis.8,17,18,21,27,28,36,37 Alternatively, because Rahm et al primarily studied women in whom infections were diagnosed as a result of screening, these findings may reflect length-time bias.
Studies of women treated with regimens with poor activity against chlamydiae provide further evidence that most infections persist if inadequately treated.38,39 In a study of 129 women with gonorrhea and chlamydial coinfection, Rees38 serially cultured women at 1- to 3-week intervals for up to 12 weeks after treatment with penicillin-probenicid or 1 g ampicillin. Seventy-two percent of women were followed up for 1 month or longer and 84% remained infected at the time of last culture.
Several case series and case reports document that many women remain infected for long periods. Schachter29 reported that 25 of 33 untreated women (76%) retested 2 to 28 weeks after an initial positive culture remained culture positive when retested. In a group of 14 untreated asymptomatic women who were retested by McCormick1 16 to 17 months after an initially positive culture or test for genital antichlamydial antibody, 50% had evidence of infection. Three women denied sexual activity in the preceding 5 to 11 months. Stenberg reported a case of chlamydial conjunctivitis that persisted in a 6-year-old boy from birth.40 At the time the infection was diagnosed, the child's mother had a cervical chlamydial infection that, potentially, was persistent from the time of the his birth. Hammerschlag41 found culture-positive vaginal chlamydial infections in two victims (aged 6 and 10 years) of sexual assault who were believed to have been infected for 3 years.
Based primarily on nonculture laboratory findings, studies in women seeking evaluation for infertility also suggest that infections can be prolonged. Investigators using in situ hybridization and immunoperoxidase staining have found evidence of chlamydiae in 26% to 56% of patients with tubal infertility.42,43 However, findings from studies reporting culture results have been variable, with 0% to 33% of infertile women testing positive by culture from one or more anatomical sites.43–55 In one study, upper genital tract specimens were positive more often than endocervical specimens, suggesting that infection may be isolated to the upper tract.46 To the extent that the women tested in these studies were infertile as a result of chlamydial infections, their infections were likely long-standing. However, as in animal studies, whether nonculture evidence of chlamydiae in the upper genital tract is indicative of viable, replicating, or transmissible infection is uncertain. Several studies have documented that PCR performed on lower genital tract specimens can remain positive for several weeks after cultures become negative, though persistent lower-tract PCR positivity appears to be rare.56,57 Recently, Gerard et al58 used reverse transcriptase PCR (RT-PCR) to demonstrate the presence of primary transcripts of chlamydial rRNA in the fallopian tubes of six women with ectopic pregnancy who were PCR positive for C trachomatis. No primary rRNA transcripts were detected in fallopian-tube samples from four women who were PCR negative. RT-PCR has also been used to demonstrate the persistence of chlamydial RNA in patients with reactive arthritides secondary to chlamydial infection.59 Because RNA is produced in metabolically active cells and is rapidly degraded, these studies provide the best evidence to date that PCR positivity is indicative of the presence of viable microorganisms. However, it remains unclear whether evidence of upper genital tract infection is indicative of transmissible disease, and results of RT-PCR need to be replicated in larger populations.
Seven studies have reported the results of serial chlamydial cultures in untreated men (Table 3).31,35,60–64 In the largest study, Johannisson recultured 52 men with chlamydial urethritis 1 to 4 weeks after an initial positive chlamydial culture. After 1 or 2 weeks, 85% of subjects were recultured, and 60% remained culture positive.63 In the next-largest study of untreated men, Paavonen31 observed somewhat higher rates of persistent infection, with 76% of 21 men remaining culture positive after 4 weeks.
Ten additional studies have recultured men after treatment with medications that were subsequently found to be inadequate to consistently eradicate infection with C trachomatis65–74 (Table 4). Because some subjects were probably cured with suboptimal therapy, it is uncertain what proportion of negative cultures in these studies reflect spontaneous clearance of infection. However, subjects with persistently positive cultures define a low estimate of the proportion of men who would remain infected if untreated. In a study with serial follow-up cultures, Stamm68 observed 10 initially asymptomatic men for 14 to 35 days (mean, 24 days). Participants were cultured three or four times before treatment, and the mean duration of infection after positive screening was 22 days. Six of 10 men remained infected at time of treatment. One patient had spontaneous clearing of infection documented by three negative follow-up cultures without previous treatment. One patient developed symptoms, suggesting that symptomatic urethritis can occur in an initially asymptomatic man identified through screening.
Taken together, studies in untreated and inadequately treated subjects suggest that most symptomatic men remain infected for more than 3 weeks and that spontaneous clearance can occur. Chlamydial urethritis has been reported to persist for as long as 1 year29; however, no study has consistently evaluated men for more than 4 weeks. As a result, the frequency of prolonged infections is unknown. Existing data provide some evidence to support the common belief that infections spontaneously resolve more quickly in men than in women.5 However, culture in men may be less sensitive than in women,75 making it difficult to draw firm conclusions.
The extent to which prolonged infections reflect the natural history of a single, persistent chlamydial infection as opposed to a result of serial exposures and recurrent infections is uncertain. To date, studies of untreated patients have not controlled for the possibility of reexposure. However, studies of treated patients provide some data on this subject. A subset of women treated for chlamydial infections retest positive even if they have had a negative test-of-cure culture.76 Dean77 genotyped chlamydial isolates from seven women with at least five positive chlamydial cultures with the same serotype over a 2-year period. All women had identical or near-identical genotypes with each infection, leading the author to conclude that these infections were persistent despite treatment. Further supporting the idea that women with repeated positive chlamydial cultures after treatment have a persistent as opposed to a new or recurrent infection, high rates of infection have been reported among adolescents who report sexual abstinence after single-dose azithromycin treatment for C trachomatis.78 Other studies do not support the idea that infections persistent after conventional therapy. Despite evaluating patients for as long as 20 weeks, studies using DNA amplification techniques to sample the lower genital tract have not detected evidence of persistent infection after treatment.56,57,79
Studies of untreated nongenital chlamydial infections also support the idea that longstanding chlamydial infections can result from persistent as opposed to recurrent infection. Following up children with perinatal chlamydial infections, Bell80 documented a median duration of culture-positive infection of 189 days in untreated subjects and 226 days in unsuccessfully treated subjects. One child was infected for as long as 866 days. Most infections were nasopharyngeal. Lymphogranuloma venereum (LGV) has been reported to persist for as long as 20 years,81 and clinically active trachoma has been reported in a humans subjected to topical opthalmic corticosteroids after living for years outside of a trachoma endemic area.82
We systematically reviewed the literature on C trachomatis to better define the duration of transmissible genital infections. We conclude that existing data do not allow one to determine the median duration that untreated infections remain contagious or to construct a curve that describes the natural history of the disease. Definitively determining the duration of potentially transmissible genital chlamydial infections would require the serial testing of untreated patients whose time of infection was known using tests for which the relationship to transmissibility was defined. Such natural history studies could not be performed ethically and, at present, the relation of various laboratory tests to risk of transmission is unknown.
Although animal infections are imperfect models for human infections, they have allowed investigators to serially culture animals with untreated chlamydial infections until they become culture negative. No similar studies have been performed in humans. These studies suggest that animals, particularly primates, may experience prolonged culture-positive infections and that, at least as measured by culture, these infections spontaneously clear. Studies in which animals were immunosuppressed document that viable, potentially transmissible organisms persist for a finite time beyond the period of culture positivity, whereas reports using nonculture techniques demonstrate more persistent evidence of chlamydiae of uncertain viability and transmissibility. Animal studies also suggest that the duration of infection is affected by a previous history of chlamydial infection. The usefulness of existing animal data is limited by potential dissimilarities between humans and the animals studied, by the fact that many studies used chlamydial species not implicated in human disease, and because of the theoretical impact that serially culturing animals may have on the natural history of infection. In addition, little data exists on infections in male animals. Studies of primates often have shown intermittent culture positivity, making it uncertain whether negative cultures documented in studies are indicative of clearance of transmissible infection. As a result, infections in these animals may be more prolonged than reported, with the quantity of organisms shed periodically dropping below the threshold of detection by culture.
Although potentially limited by length-time bias, human studies in which patients were serially tested provide evidence that most infections in women persist for more than 60 days. Small case series and case reports suggest that infections may persist for years, whereas studies of women seeking care for infertility provide evidence of presumably long-standing chlamydial persistence in the upper genital tract. Recent evidence using RT-PCR suggests that this persistence reflects viable infection; however, more studies are needed to clearly establish the extent to which chlamydiae establish longstanding upper genital tract infections and whether these infections are indicative of transmissible infection. Studies in men are less definitive still. Although untreated infections in men can be protracted, inconclusive data suggest that the duration of infection may be shorter than in women. Because of variable length of follow-up evaluation among studies and, in some instances, among subjects in individual studies, drawing firm conclusions regarding the rate at which genital chlamydial infections resolve is not possible, and one cannot determine the proportion of infections that persist for long periods. Furthermore, it is not possible to determine the extent to which reinfection may influence the observed duration of infection.
Despite the difficulties inherent in better defining the duration of chlamydial infections, it may be possible to improve existing knowledge. First, animal studies could be performed in primates using DNA-amplification testing and serial cultures to assess the natural history of untreated infections. Such studies might include quantitative measures of infection to define the relation of infectious burden to test results. Second, as screening programs expand and include men and difficult-to-reach populations, it will be possible to retest persons who inadvertently delay treatment. Many persons who test positive for chlamydial infections cannot be immediately located,83,84 and when eventually found can be retested to assess whether they are still infected. This has been done in Alabama, and similar studies elsewhere could make a contribution to our ability to estimate the duration of infection. In cases reevaluated 3 or more weeks after an initial test, DNA amplification testing should be done in addition to culture to maximize the sensitivity of repeat testing and to assess for the existence of viable organisms. Data on symptoms, recent sexual history, history of chlamydial infections, partnership duration and change, and partner infection status might allow investigators to evaluate factors thought to affect the duration of infection and to define populations in which reinfection is an unlikely cause for repeatedly positive test results. Third, collection of standardized information could allow for the pooling of data to better define the natural history of chlamydial infections and the factors affecting infection duration. We attempted to gather existing data for such an analysis. Unfortunately, because most studies were conducted many years ago, not enough data were available to merit such an effort. Fourth, further work using RT-PCR is warranted to assess whether nonculture evidence of upper genital tract disease in women is indicative of active and transmissible infection.
Because of the inherent difficulties of attempting to study the issue of duration of infection, better defining this important determinant of transmission of genital chlamydial infection will be challenging. However, we believe that an effort to do so is justified. Knowing how long people remain infected will improve patient counseling and will provide essential information for modeling disease transmission, estimating disease burden, and designing prevention programs.
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