CHLAMYDIA TRACHOMATIS UROGENITAL INFECTIONS are a major global public health concern and remain the most commonly reported bacterial sexually transmitted diseases (STDs) in the United States with an estimated 3 million new cases yearly.1 Young age and black race are risk markers for chlamydial infection.2,3 The major outer membrane protein (MOMP) of C. trachomatis is an antigenically diverse and abundant surface protein of the organism, and chlamydial strains may be differentiated into individual serovars by typing with anti-MOMP monoclonal antibodies using microimmunofluorescence. At least 18 C. trachomatis serovars have been identified.4–6 The serovars primarily isolated from the urogenital tract include D, Da, E, F, G, H, I, Ia, J, and K, with the most prevalent serovars reported in the United States being serovars D, E, and F.7,8
Individual chlamydial serovars have been reported to differ among specific racial groups.9 Of particular interest in this regard is serovar Ia, which was observed to be increasing in prevalence in Seattle and was associated with black race.7 Chlamydial serovar and racial distributions have also been reported to influence the quantitative yield of chlamydial organisms, measured as inclusion-forming units (IFUs) in chlamydial culture.10 Higher IFU counts have been associated with clinical and microbiologic evidence of acute inflammation11 and may represent a surrogate for infectivity or transmissibility. Hence, differences in chlamydial serovars by racial distribution could impact the epidemiology of chlamydial infections in a given population, especially in terms of transmission dynamics. The objective of this study was to evaluate the C. trachomatis serovar distribution in a predominantly black population in Birmingham, Alabama, focusing particularly on serovar Ia.
Study Population and Data Collection
The study population consisted of heterosexual men and women with culture-documented C. trachomatis urogenital infections detected at routine care visits at the Jefferson County Department of Health (JCDH) STD Clinic in Birmingham, Alabama. Chlamydial isolates were initially collected from patients seen from February 2003 through April 2003. To more meaningfully study the relationship of chlamydial serovar to patient demographics, we then collected more isolates for an additional 3-month time period (from December 2003 through February 2004). Chlamydial isolates were selected consecutively during each study period, and all patients whose chlamydial isolate could be harvested for serotyping were included. Each isolate was from a different patient, and there were no individuals with repeat infections in either time period. Gender, age, race, and culture site were recorded for every patient on a laboratory requisition form accompanying the urogenital swab specimen submitted for chlamydial culture. Race was self-reported by patients at the time of clinic registration and then entered into a computerized clinic database, which was the source of the demographic data for the laboratory requisition forms. The Institutional Review Boards of the JCDH and University of Alabama at Birmingham (UAB) approved the study.
Specimen Collection and Isolation of C. trachomatis
The methods used for specimen collection and isolation of C. trachomatis have been previously described in detail.12 Briefly, Dacron-tipped swabs were inserted 1 to 2 cm into the urethra in men and the endocervix in women and were immediately placed in vials containing 1.5 mL of 0.2 mol/L sucrose-phosphate buffer transport media containing 2% fetal bovine serum and antibiotics. Transport media were then stored at 4°C for up to 18 hours before transportation to the UAB STD Research Laboratory, where they may be stored up to 72 hours at −70°C. Specimens were thawed and inoculated onto DEAE–dextran-treated McCoy cells in 96-well microtiter plates. After centrifugation, plates were incubated in 5% CO2 for 30 minutes and media was aspirated and replaced with cycloheximide medium. After incubation in 5% CO2 at 37°C for 48 to 72 hours, cells were fixed with methanol and stained with monoclonal antibody reagents to MOMP and lipopolysaccharide. Chlamydial inclusions were visualized with a fluorescence microscope. Transport media containing specimens with chlamydial inclusions were stored at −70°C until the time of serotyping.
Transport media specimens were sent to the University of Washington Chlamydia Laboratory for serotyping. A low-passage microtiter plate culture method was used for rapid immunotyping with a panel of 18 subspecies- and serovar-specific monoclonal antibodies on mature fixed inclusions, as previously described in detail.13 Monoclonal antibodies were run individually or in noncrossreactive pools to prevent ambiguous crossreactivity. Wells were then reacted with a fluorescein isothiocyanate conjugate and read with an epifluorescence microscope. Final reaction patterns were compared with prototype strain reaction patterns to determine serotype.
Statistical analyses were conducted using Stata (Release 6.0; Stata Corp., College Station, TX). Relationship of the overall serovar distribution, the major serovar distribution (including only serovars occurring in >5%), and individual serovar infections to race, gender, and age were assessed by the Fisher exact test. Mixed serovar infections were not included in the analyses. Because of the small sample size of the nonblack race group in our study, associations of chlamydial serovar and race were considered significant if 2-sided P values were less than 0.1.
Ninety-eight men and 109 women with chlamydial infection had C. trachomatis isolates consecutively collected that could be serotyped. The median patient age was 23 years (range, 13–61 years), and the racial distribution consisted of 189 (91%) black individuals and 18 (9%) persons of other races (predominantly white). Ninety-five percent of men versus 88% of women were black. Age and race did not significantly differ by gender.
C. trachomatis serovar E (60; 29%) was the most frequent serovar isolated followed by serovars D (40 [19%]), F (40 [19%]), Ia (32 [16%]), J (19 [9%]), G (4 [2%]), H (4 [2%]), K (4 [2%]), and mixed (4 [2%]). Serovar B was not isolated. The serovar distribution by gender and by race are displayed in Figure 1A and Figure 1B, respectively. Serovar E was the most common in both women and men (31% and 27%, respectively) and in both black and other racial groups (28% and 39%, respectively).
The overall chlamydial serovar distribution in those with single chlamydial serovar infections (n = 203; serovars D through K) did not significantly differ by age, gender, or race. However, limiting to the analyses to only the major chlamydial serovars (those >5% [D, E, F, Ia, J]; n = 191) revealed differences by race (P = 0.07), with serovar Ia infections only occurring in blacks (17% vs. 0%), and serovar E (30% vs. 41%) and J (9% vs. 24%) infections occurring less often in blacks. Analyses of individual serovars confirmed the associations between black race and serovars Ia and J (P = 0.08 and 0.06, respectively). Furthermore, serovar Ia infections were significantly associated with older age (21% in those age ≥23 years vs. 10% in those age <23 years; P = 0.03).
The primary objective of our study was to determine the distribution of C. trachomatis serovars in a predominantly black STD clinic patient population in Birmingham, Alabama, focusing particularly on serovar Ia, which has been found to be associated with the black race in other studies.7,9 We found serovar E was most frequently isolated, followed by serovars D and F. In general, these 3 chlamydial serovars have been reported to be the most prevalent cause of urogenital infection in heterosexuals,7,8,14,15 and as seen in other studies,7,9 their prevalence did not vary by race. However, we found frequency of serovar Ia and J infections did vary by race. Serovar J infections occurred less frequently in blacks and interestingly, serovar Ia infections, which were the fourth most frequent overall, were only seen in blacks (occurring in 17%).
Studies assessing serovar distribution in a predominantly black population in the United States are sparse, particular in the Southeast. Batteiger et al. reported 12% of chlamydial infections in persons attending a STD clinic in Indianapolis in 1986 were the result of serovar I.8 In their study, serovar I represented a pooled serovar group consisting of serovar I and other I variants (possibly including Ia, which was identified and reported later by Wang and Grayston in 1991).5,8 In an abstract presented at the International Congress of Sexually Transmitted Infections (Berlin, June 2001) by Martin et al., 18% of chlamydial isolates obtained from patients, mostly pregnant women, in New Orleans were genotype Ia.16 In a study of chlamydial serovar distribution in a STD clinic population in Seattle, in which blacks comprised only 40% of infected individuals, black men and women were significantly more likely to be infected with serovar Ia (14% and 5.5%, respectively).9 Reports of serovar Ia infections outside the United States are limited, and when reported, serovar Ia usually accounts for only a small proportion of infections. Selected clinical studies reporting chlamydial serovar Ia infections are summarized in Table 1.
It has been suggested that choice of sexual partners (e.g., less frequent mixing with partners of a different race) could influence the distribution of chlamydial serovars.9 We did not assess the dynamics of sexual mixing among the different racial groups in our study population and thus cannot comment further on this issue from our data. Alternatively, geographic distribution could well influence distribution of serovar Ia, which is supported by the paucity of serovar Ia infections reported in countries outside of the United States. Additionally, biologic differences in persons of different races and in chlamydial organisms of different serovars could influence acquisition and transmission of infection. This is supported by prior findings of lower chlamydial IFU counts in blacks and in C-class serovar (includes Ia) infections, respectively.10 Further investigation is needed to delineate such possible biologic differences.
Our study has limitations worth noting. Because the population we studied consisted of a high-risk group of patients attending an STD clinic, our findings may not be generalizable to other predominantly black populations with different behavioral and other characteristics. Also, the primary objective of this study was to describe the chlamydial serovar distribution in Birmingham, and the sample size used for this purpose may have had limited power in addressing secondary study objectives such as detecting associations of chlamydial serovars with demographic characteristics.
In conclusion, we found C. trachomatis serovar E to be the most prevalent serovar in a predominantly black population in Birmingham, Alabama, and that the serovar distribution differed by race. The association of serovar Ia with the black race, like found in our study and others in the United States, deserves further study to identify the biologic, geographic, or behavioral factors contributing to this association. Chlamydial genotyping would be of interest in future studies to better define strain variation and transmission dynamics that may improve our understanding of the association of chlamydial serotypes with race.
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