Infectious Diseases in Obstetrics and GynecologyVolume 2011 (2011), Article ID 842652, 10 pagesdoi:10.1155/2011/842652

Clinical Study

Association between Semen Exposure and Incident Bacterial Vaginosis

Maria F. Gallo,¹ Lee Warner,¹ Caroline C. King,¹ Jack D. Sobel,² Robert S. Klein,³ Susan Cu-Uvin,⁴ Anne M. Rompalo,⁵ and Denise J. Jamieson¹

¹Division of Reproductive Health, Centers for Disease Control and Prevention, 4770 Buford Highway, Mail Stop K-34, Atlanta, GA 30341-3724, USA²Department of Medicine, Wayne State University School of Medicine, Harper Hospital, 2990 John R. Street, 5 Hudson, Detroit, MI 48201, USA³Mount Sinai School of Medicine, One Gustave L. Levy Place, Box 1057, New York, NY 10029, USA⁴Brown University School of Medicine, The Miriam Hospital, 164 Summit Avenue, Providence, RI 02906, USA⁵Johns Hopkins Bayview Medical Center, Johns Hopkins University School of Medicine, 5200 Eastern Avenue, MFL Building, Center Tower, Suite 4000, Baltimore, MD 21224, USA

Received 9 August 2011; Accepted 27 September 2011

Academic Editor: Harold Wiesenfeld

Copyright © 2011 Maria F. Gallo et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract.

Objective. To identify correlates of incident bacterial vaginosis (BV) diagnosed with Nugent scoring among high-risk women. Study Design. We conducted both cohort and case-crossover analyses, stratified by HIV infection status, based on 871 HIV-infected and 439 HIV-uninfected participants in the HIV Epidemiology Research Study, conducted in 4 US sites in 1993–2000. Results. BV incidence was 21% and 19% among HIV-infected and -uninfected women, respectively. Fewer correlates of BV were found with case-crossover than with cohort design. Reporting frequent coitus (regardless of consistency of condom use) was correlated with BV in cohort analyses but not in case-crossover analyses. The sole correlate of BV in both types of analyses was the detection of spermatozoa on Gram stain, which is a marker of semen exposure. Conclusion. The inconsistent association between condom use and BV in prior studies could be from reporting bias. We found evidence of a relationship between semen exposure and incident BV.

1. Introduction

Bacterial vaginosis (BV) is a common vaginal condition with an estimated prevalence of 29% among U.S. women during 2001–2004 [1]. BV has been linked to a range of adverse reproductive outcomes, including infertility, spontaneous abortion, preterm premature rupture of the membranes, amniotic fluid infection, low birth weight, and preterm delivery [2–8]. BV also might increase women’s risk for pelvic inflammatory disease although evidence on this possible association is inconsistent [9–11]. In addition, evidence suggests that BV increases women’s risk for sexually transmitted infections (STIs), including gonorrhea, chlamydial infection, trichomoniasis, human papillomavirus, herpes simplex virus, and HIV [12–19].

Although the etiology of BV remains unknown, two competing hypotheses currently prevail [20–22]. In the first, BV is viewed as an imbalance of the vaginal microbiota caused by the colonization of endogenous organisms from the intestinal tract [23]. This imbalance could be precipitated by a variety of events, including coitus and vaginal cleansing or douching. The second hypothesis holds that BV is caused by the sexual transmission of a specific pathogen (e.g., Gardnerella vaginalis or unknown bacteria). The similarity between the epidemiology of BV and that of STIs supports the hypothesis that BV is sexually transmitted. For example, BV has been associated with risky sexual behaviors, including having new or a relatively high number of sexual partners, having sex frequently, not using condoms, using drugs during sex, and having sex with uncircumcised partners [1, 16, 22, 24, 25]. However, because these associations often have been found in observational studies, they could be the result of uncontrolled confounding. Use of a case-crossover analysis (in which each woman serves as her own control) would minimize the effects of time-independent confounders [25]. We conducted both cohort and case-crossover analyses to identify time-variant correlates of BV among a cohort of high-risk women in the U.S., who participated in a longitudinal study of the effects of HIV infection on women’s health [26].

2. Materials and Methods

We analyzed data from the HIV Epidemiology Research Study (HERS), which was conducted at 4 U.S. sites (Bronx, NY; Detroit, MI; Baltimore, MD; and Providence, RI, USA) in 1993–2000 [26]. Participants consisted of 871 HIV-infected women and 439 uninfected women who, at the time they enrolled in the study, were 16–55 years of age, did not have an AIDS-defining clinical diagnosis, and either injected drugs or engaged in high-risk sexual behaviors (i.e., had >5 sexual partners in the previous 5 years, traded sex for money or drugs, or had sex with a male who injected drugs or who was suspected of being or known to be infected with HIV). After enrollment, participants completed follow-up visits scheduled at 6-month intervals. During these visits, HERS staff conducted interviews to collect demographic, health, and behavioral information, conducted physical examinations, and collected specimens to be tested for infections, including BV, HIV, human papillomavirus (HPV), and trichomoniasis. Study visits were not used to diagnose or treat symptoms, and less than 1% of participants reported using metronidazole or topical clindamycin [27]. Ethical review boards at the study sites and the Centers for Disease Control and Prevention approved the study, and only women who gave informed consent were enrolled.

Gram-stained slides prepared from swabs of posterior vaginal fornix specimens were air dried, fixed in methanol, and shipped to a central laboratory where a single technician used oil immersion with ×1000 magnification to quantify and score the specimens. Specimens with a Nugent score of 7–10 were considered positive for BV [28]. Gram stains also were evaluated for morphological identification of spermatozoa, which is specific for recent exposure to semen [29]. Spermatozoa usually clear from vaginal secretions by 12–36 hours after exposure to semen although they have been detected microscopically up to 10 days after exposure [30, 31]. Wet mount was used for diagnosing trichomoniasis, and vaginal specimens were cultured for Candida organisms. Aliquots of cervicovaginal lavage fluid were frozen for later testing for HPV by polymerase chain reaction.

We limited our analyses to data collected during participants’ first 10 follow-up visits and excluded the 12 women who HIV seroconverted during the study. Participants’ incident BV status was assessed at follow-up visits only if Nugent scores of samples collected at their preceding visit indicated that they were BV negative. If they tested positive for BV or their Nugent scores were missing, their incident BV status was coded as missing. We used unconditional (using generalized estimating equations to account for intrasubject correlation from multiple visits) and conditional logistic regression to analyze the data as if they were derived from a cohort and case-crossover study, respectively. For both analyses, we constructed individual models to evaluate the correlates of incident BV for HIV-infected and -uninfected women separately. While the analytic population for the cohort analysis included all follow-up visits with nonmissing data on incident BV, the case-crossover analysis was limited to follow-up visits from women who had ≥1 follow-up visit with and ≥1 follow-up visit without incident BV.

For both the cohort and case-crossover analyses, we fitted individual models to assess the bivariable relationship between each potential correlate and incident BV. For the multivariable analyses, we fitted full models with all potential correlates and used manual, backward elimination to exclude factors that were not significantly associated (based on an alpha of 0.05) with incident BV. Potential correlates were selected because of their prior identification in the literature. The cohort analyses included both time-independent and -dependent variables. However, because individual participants in the case-crossover analyses served both as case subjects and matching control subjects, the variables evaluated in these analyses were limited to time-dependent factors, which had the potential to vary between the participant’s visits.

3. Results

Because of the differences in findings by HIV status in both the cohort and case-crossover analyses, we present results separately for HIV-infected and -uninfected participants. The cohort analyses were based on data collected during 3,050 visits by 799 HIV-infected women and 1,564 visits by 375 uninfected women. The case-crossover analyses were based on data collected during 1,543 visits by 332 HIV-infected women and 753 visits by 159 uninfected women. The incidence of BV during the study follow-up period was 21% among HIV-infected women and 19% among uninfected women.

3.1. HIV-Infected Participants

The four time-independent variables assessed (i.e., study site, age at baseline, race, and education at baseline) were significantly associated with incident BV in the bivariable, cohort analyses among HIV-infected women (Table 1). All variables except for study site were also significantly associated with incident BV in the multivariable analysis, the results of which showed risk for incident BV to be higher among women younger than 45 years of age than among those older, higher among black women than among white women, and higher among women with a high-school education or less than among those with post-high-school education. Seven time-dependent variables were correlated with incident BV in the bivariable, cohort analyses. Except for current injection drug use and cigarette use within six months, these also were associated with incident BV in the multivariable, cohort analysis. Visits with trichomoniasis at the preceding visit (adjusted odds ratio [aOR], 1.8; 95% confidence interval [CI], 1.4–2.4), HPV at the preceding visit (aOR, 1.3; 95% CI, 1.0–1.5), and spermatozoa detected (aOR, 1.5; 95% CI, 1.1–2.1) had more incident BV than visits without these diagnoses. Also, visits in which the woman reported inconsistent condom use and frequent coitus (aOR, 1.6; 95% CI, 1.2–2.2) or consistent condom use and frequent coitus (aOR, 1.6; 95% CI, 1.2–2.1) were associated with more incident BV than visits in which women reported no sexual activity. Finally, crack use within six months correlated with incident BV (aOR, 1.9; 95% CI, 1.4–2.6).

Table 1: Results of cohort analyses of associations between selected factors and incident bacterial vaginosis among HIV-infected U.S. women, HERS, 1993–2000*.

Control visits
Case visits
Bivariable model
Multivariable model†

No.
No.
OR
(95% CI)
aOR
(95% CI)

Time-independent factors

Study site

    Site 1
749
138
Referent
Referent

    Site 2
502
170
1.8
(1.3, 2.4)
1.3
(0.9, 1.8)

    Site 3
410
173
2.4
(1.7, 3.2)
1.4
(1.0, 2.0)

    Site 4
761
147
1.1
(0.8, 1.5)
1.1
(0.8, 1.6)

Age at baseline (in years)

    16–24
43
16
2.2
(1.1, 4.3)
1.4
(0.7, 3.0)

    25–34
664
217
2.2
(1.6, 3.0)
2.0
(1.4, 2.8)

    35–44
1235
324
1.7
(1.3, 2.3)
1.6
(1.2, 2.1)

    45+
480
71
Referent
Referent

Race

    Black
1239
430
2.1
(1.6, 2.6)
1.7
(1.3, 2.3)

    Other
1183
198
Referent
Referent

Education at baseline

    <High school
906
289
1.8
(1.4, 2.4)
1.6
(1.2, 2.1)

    High school
763
209
1.5
(1.2, 2.0)
1.5
(1.1, 1.9)

    >High school
750
129
Referent
Referent

Time-dependent factors

Visit (0–10)

0.9
(0.9, 1.0)
1.0
(0.9, 1.0)

CD4+ group (cells/μL)

    0–199
567
147
1.0
(0.8, 1.3)

    200–499
1213
308
1.1
(0.9, 1.5)

    ≥500
598
166
Referent

Trichomoniasis at preceding visit

    Yes
180
117
2.2
(1.7, 2.8)
1.8
(1.4, 2.4)

    No
2240
510
Referent
Referent

Vaginal Candidal culture at preceding visit

    Yes
804
247
1.2
(1.0, 1.4)

    No
1612
377
Referent

Human papillomavirus at preceding visit

    Yes
1468
432
1.3
(1.0, 1.5)
1.3
(1.0, 1.5)

    No
934
188
Referent
Referent

Spermatozoa detected on Gram stain

    Yes
139
70
1.7
(1.3, 2.4)
1.5
(1.1, 2.1)

    No
2283
558
Referent
Referent

Sexual behavior during previous 6 months‡

    Frequent coitus, inconsistent condom use
331
127
2.0
(1.6, 2.7)
1.6
(1.2, 2.2)

    Frequent coitus, consistent condom use
431
146
1.8
(1.4, 2.4)
1.6
(1.2, 2.1)

    Infrequent coitus, inconsistent condom use
240
65
1.4
(1.0, 1.9)
1.2
(0.9, 1.7)

    Infrequent coitus, consistent condom use
488
127
1.4
(1.1, 1.8)
1.3
(1.0, 1.7)

    Not sexually active
920
162
Referent
Referent

Female sex partner during previous 6 months

    Yes
86
25
1.1
(0.8, 1.7)

    No
2320
601
Referent

Douching within previous 48 hours

    Yes
72
24
1.0
(0.6, 1.6)

    No
2340
602
Referent

Current hormonal contraception use

    Yes
101
29
1.2
(0.8, 1.8)

    No
2319
599
Referent

Current injection drug use

    Yes
304
131
1.7
(1.4, 2.2)

    No
2114
497
Referent

Crack use during previous 6 months

    Yes
199
117
2.2
(1.6, 2.9)
1.9
(1.4, 2.6)

    No
2219
511
Referent
Referent

Cigarette use during previous 6 months

    Yes
1639
484
1.5
(1.2, 1.9)

    No
779
144
Referent

HERS = HIV Epidemiology Research Study; OR: odds ratio; aOR: adjusted odds ratio; CI: confidence interval.

*Findings from unconditional logistic regression model, using generalized estimating equations, based on 628 case visits (i.e., visits with incident bacterial vaginosis) and 2422 control visits (i.e., visits without incident bacterial vaginosis) completed by 799 participants.

†Adjusted for all variables in column.

‡Frequent coitus defined as ≥4 times per month and infrequent coitus defined as <4 times per month.

The bivariable, case-crossover analyses yielded four correlates of incident BV among HIV-infected women: spermatozoa detection, sexual behaviors, current injection drug use, and crack use within six months (Table 2). Except for sexual behaviors, these variables remained significantly associated in the multivariable, case-crossover analysis. Visits with spermatozoa detection (aOR, 1.6; 95% CI, 1.1–2.5), reports of current injection drug use (aOR, 1.9; 95% CI, 1.1–3.3) and reports of crack use within six months (aOR, 1.6; 95% CI, 1.0–2.7) were more likely to have incident BV than visits without these factors.

Table 2: Results of case-crossover analyses of associations between selected factors and incident bacterial vaginosis among HIV-infected U.S. women, HERS, 1993–2000*.

Control visits
Case visits
Bivariable model
Multivariable model†

No.
No.
OR
(95% CI)
aOR
(95% CI)

Time-dependent factors

CD4+ group (cells/μL)

    0–199
257
112
0.8
(0.4, 1.3)

    200–499
544
228
0.8
(0.5, 1.2)

    ≥500
249
128
Referent

Trichomoniasis at preceding visit

    Positive
100
74
1.2
(0.8, 1.9)

    Negative
967
400
Referent

Vaginal Candidal culture at preceding visit

    Positive
371
181
1.1
(0.9, 1.5)

    Negative
695
290
Referent

Human papillomavirus at preceding visit

    Positive
668
326
0.8
(0.6, 1.2)

    Negative
392
143
Referent

Spermatozoa detected on Gram stain

    Yes
70
48
1.5
(1.0, 2.3)
1.6
(1.1, 2.5)

    No
998
427
Referent
Referent

Sexual behavior during previous 6 months‡

    Frequent coitus, inconsistent condom use
151
92
1.9
(1.2, 3.1)

    Frequent coitus, consistent condom use
177
113
1.7
(1.1, 2.8)

    Infrequent coitus, inconsistent condom use
118
46
1.2
(0.7, 2.0)

    Infrequent coitus, consistent condom use
225
96
1.2
(0.8, 1.8)

    Not sexually active
388
127
Referent

Female sex partner during previous 6 months

    Yes
51
20
1.5
(0.6, 3.9)

    No
1010
453
Referent

Douching within 48 hours

    Yes
38
19
0.7
(0.4, 1.3)

    No
1025
454
Referent

Current hormonal contraception use

    Yes
30
23
2.4
(0.9, 6.7)

    No
1037
452
Referent

Current injection drug use

    Yes
159
97
2.1
(1.2, 3.5)
1.9
(1.1, 3.3)

    No
907
378
Referent
Referent

Crack use during previous 6 months

    Yes
107
81
1.8
(1.1, 2.9)
1.6
(1.0, 2.7)

    No
959
394
Referent
Referent

Cigarette use during previous 6 months

    Yes
758
364
1.4
(0.7, 2.7)

    No
308
111
Referent

*Findings from conditional logistic regression model based on 475 case visits (i.e., visits with incident bacterial vaginosis) and 1068 control visits (i.e., visits without incident bacterial vaginosis) completed by 332 women with at least one case and one control visit.

†Adjusted for all variables in column.

‡Frequent coitus defined as ≥4 times per month and infrequent coitus defined as <4 times per month.

3.2. HIV-Uninfected Participants

Among HIV-uninfected participants, study site and race were the only time-independent variables significantly associated with BV risk in the bivariable, cohort analyses, and both remained significantly associated with BV risk in the multivariable, cohort analysis (Table 3). Black women had a higher risk than women of other races (aOR, 1.9; 95% CI, 1.3–2.7). Results of the bivariable, cohort analyses results showed six time-dependent variables to be significantly associated with incident BV risk. All except one (crack use within the previous 6 months) also were associated with BV risk in the multivariable analyses. Factors significantly associated with incident BV risk in the multivariable analyses were trichomoniasis at the preceding visit (aOR, 1.7; 95% CI, 1.1–2.6), spermatozoa detection (aOR, 1.9; 95% CI, 1.3–2.9), coitus ≥4 times per month during the previous 6 months and either inconsistent condom use (aOR, 1.9; 95% CI, 1.3–2.8) or consistent condom use (aOR, 1.9; 95% CI, 1.2–3.1) cigarette use during previous 6 months (aOR, 1.5; 95% CI, 1.0–2.1), and current use of hormonal contraception (aOR, 0.4; 95% CI, 0.2–0.8).

Table 3: Results of cohort analyses of associations between selected factors and incident bacterial vaginosis among HIV-uninfected U.S. women, HERS, 1993–2000*.

Control visits
Case visits
Bivariable model
Multivariable model†

No.
No.
OR
(95% CI)
aOR
(95% CI)

Time-independent factors

Study site

    Site 1
437
65
Referent
Referent

    Site 2
185
77
2.7
(1.7, 4.2)
1.9
(1.2, 3.1)

    Site 3
271
94
2.4
(1.5, 3.7)
1.7
(1.0, 2.7)

    Site 4
372
63
1.1
(0.7, 1.7)
1.5
(1.0, 2.3)

Age at baseline (in years)

    16–24
51
7
0.9
(0.3, 2.1)

    25–34
386
106
1.4
(0.9, 2.2)

    35–44
597
139
1.1
(0.7, 1.6)

    45+
231
47
Referent

Race

    Black
568
201
2.5
(1.8, 3.5)
1.9
(1.3, 2.7)

    Other
697
98
Referent
Referent

Education at baseline

    <High school
367
99
1.4
(1.0, 2.1)

    High school
436
101
1.1
(0.8, 1.6)

    >High school
457
99
Referent

Time-dependent factors

Visit (0–15)

0.9
(0.9, 1.0)
0.94
(0.9, 1.0)

Trichomoniasis at preceding visit

    Yes
100
60
2.1
(1.4, 3.2)
1.7
(1.1, 2.6)

    No
1163
239
Referent
Referent

Vaginal Candidal culture at preceding visit

    Yes
393
91
1.0
(0.8, 1.3)

    No
864
208
Referent

Human papillomavirus at preceding visit

    Yes
278
84
1.3
(0.9, 1.7)

    No
975
210
Referent

Spermatozoa detected on Gram stain

    Yes
76
43
2.3
(1.6, 3.3)
1.9
(1.3, 2.9)

    No
1189
256
Referent
Referent

Sexual behavior during previous 6 months‡

    Frequent coitus, inconsistent condom use
411
139
2.3
(1.6, 3.3)
1.9
(1.3, 2.8)

    Frequent coitus, consistent condom use
129
35
1.8
(1.1, 2.9)
1.9
(1.2, 3.1)

    Infrequent coitus, inconsistent condom use
244
49
1.4
(0.9, 2.0)
1.2
(0.8, 1.8)

    Infrequent coitus, consistent condom use
138
23
1.1
(0.6, 1.9)
1.1
(0.6, 2.0)

    Not sexually active
341
52
Referent
Referent

Female sex partner during previous 6 months

    Yes
92
25
1.2
(0.7, 1.8)

    No
1169
272
Referent

Douching within 48 hours

    Yes
44
13
1.1
(0.6, 2.0)

    No
1215
285
Referent

Current hormonal contraception use

    Yes
92
9
0.4
(0.2, 0.9)
0.4
(0.2, 0.8)

    No
1171
289
Referent
Referent

Current injection drug use

    Yes
160
58
1.5
(1.0, 2.2)

    No
1104
241
Referent

Crack use during previous 6 months

    Yes
139
66
2.0
(1.3, 2.9)

    No
1125
233
Referent

Cigarette use during previous 6 months

    Yes
837
230
1.7
(1.2, 2.4)
1.5
(1.0, 2.1)

    No
427
69
Referent
Referent

*Findings from unconditional logistic regression model, using generalized estimating equations, based on 299 case visits (i.e., visits with incident bacterial vaginosis) and 1265 control visits (i.e., visits without incident bacterial vaginosis) completed by 375 participants.

†Adjusted for all variables in column.

‡Frequent coitus defined as ≥4 times per month and infrequent coitus defined as <4 times per month.

Two factors were associated with incident BV in the bivariable, case-crossover analyses among HIV-uninfected participants, and both remained associated in the multivariable analysis (Table 4). Visits with spermatozoa detected were more likely to have incident BV (aOR, 2.1; 95% CI, 1.1–4.0) than visits without its detection. Visits with self-reported frequent coitus and either inconsistent condom use (aOR, 3.0; 95% CI, 1.5–5.9) or consistent condom use (aOR, 3.1; 95% CI, 1.3–7.4) had more incident BV than visits with self-reported lack of sexual activity.

Table 4: Results of case-crossover analyses of associations between selected factors and incident bacterial vaginosis among HIV-uninfected U.S. women, HERS, 1993–2000*.

Control visits
Case visits
Bivariable model
Multivariable model†

No.
No.
OR
(95% CI)
aOR
(95% CI)

Time-dependent factors

Trichomoniasis at preceding visit

    Yes
76
41
1.2
(0.7, 2.1)

    No
446
189
Referent

Vaginal Candidal culture at preceding visit

    Yes
143
70
1.1
(0.7, 1.6)

    No
377
160
Referent

Human papillomavirus at preceding visit

    Yes
126
55
1.0
(0.6, 1.5)

    No
393
171
Referent

Spermatozoa detected on Gram stain

    Yes
36
29
2.4
(1.3, 4.5)
2.1
(1.1, 4.0)

    No
487
201
Referent
Referent

Sexual behavior during previous 6 months‡

    Frequent coitus, inconsistent condom use
159
100
3.3
(1.7, 6.6)
3.0
(1.5, 5.9)

    Frequent coitus, consistent condom use
46
29
3.1
(1.3, 7.3)
3.1
(1.3, 7.4)

    Infrequent coitus, inconsistent condom use
114
36
1.7
(0.8, 3.4)
1.6
(0.8, 3.2)

    Infrequent coitus, consistent condom use
58
19
1.1
(0.5, 2.4)
1.1
(0.5, 2.5)

    Not sexually active
145
45
Referent
Referent

Female sex partner during previous 6 months

    Yes
48
25
1.7
(0.6, 4.6)

    No
472
204
Referent

Douching within 48 hours

    Yes
23
10
0.8
(0.3, 2.2)

    No
497
220
Referent

Current hormonal contraception use

    Yes
22
8
0.4
(0.1, 1.3)

    No
500
221
Referent

Current injection drug use

    Yes
93
42
1.5
(0.7, 3.2)

    No
429
188
Referent

Crack use during previous 6 months

    Yes
93
45
1.4
(0.8, 2.5)

    No
429
185
Referent

Cigarette use during previous 6 months

    Yes
373
173
1.4
(0.6, 3.5)

    No
149
57
Referent

*Findings from conditional logistic regression model based on 230 case visits (i.e., visits with incident bacterial vaginosis) and 523 control visits (i.e., visits without incident bacterial vaginosis) completed by 159 women with at least one case and one control visit.

†Adjusted for all variables in column.

‡Frequent coitus defined as ≥4 times per month and infrequent coitus defined as <4 times per month.

4. Discussion

The sole correlate of incident BV that emerged in both the cohort and case-crossover analyses among HIV-infected and -uninfected women was the detection of spermatozoa on Gram stain, which is a biological marker of recent exposure to semen. The cohort analyses among HIV-infected and -uninfected women also found that incident BV was more common among those reporting frequent coitus (regardless of the consistency of condom use); however, this association was not found in the case-crossover analyses. A protective effect of condoms against BV has been demonstrated in some prior studies (including a case-crossover analysis), but not in all studies [22, 32–34]. The failure to find a relationship between condom use and decreased BV risk in earlier studies could have been the result of misclassification in participant reporting of coitus and condom use. This misclassification could have occurred if studies collected inaccurate reports of condom use, as a result of social desirability or recall bias, or did not collect comprehensive data on condom use, including possible malfunctions or misuse. A protective effect of condoms against BV also could have been obscured in previous studies because of the role of recurring cases of BV. That is, if unprotected coitus can cause incident BV but is not a necessary component for its recurrence, establishing the link between unprotected coitus, and incident BV could be difficult.

We found fewer correlates of incident BV in our case-crossover analyses than in our cohort analyses. Results of the adjusted case-crossover analyses of incident BV among HIV-infected women showed only spermatozoa detection, current injection drug use, and crack use within the previous 6 months to be associated with incident BV, whereas results of the cohort analyses among HIV-infected women also showed trichomoniasis at the previous visit, HPV at the previous visit and coitus ≥4 times per month during the previous 6 months to be associated with incident BV. Similarly, results of the case-crossover analyses of risk among HIV-uninfected women only found spermatozoa detection and coitus ≥4 times per month during the previous 6 months to be associated with incident BV, whereas results of the cohort analyses among HIV-uninfected women also showed trichomoniasis at the previous visit, current hormonal contraception use, and cigarette use within the previous 6 months to be associated with incident BV. The case-crossover analyses might have identified fewer correlates of incident BV as a result of reduced confounding from each woman serving as both a case subject (visits with incident BV) and a matching control subject (visits without incident BV) [25]. Alternatively, reduced power in the case-crossover analyses might have prevented the detection of correlates of BV.

A major study limitation was that BV was only assessed at six-month intervals. Studies with frequent sampling have suggested that women may have rapid fluctuations in vaginal microbiota, including short episodes of BV that resolve spontaneously [35, 36]. Thus, our study might have missed cases of BV. We were also unable to determine the temporal relationship between exposure to semen and the development of BV; as a result, we cannot rule out the possibility that the association between the two factors is the result of BV causing longer persistence of spermatozoa in vaginal fluid rather than semen exposure actually causing BV. Previous case-crossover analyses also suggest that recent menses, use of vaginal lubricants, rectal sex, douching for cleansing after menstruation, and psychosocial stress could be risk factors for incident BV [36–38]. None of these factors, though, were evaluated in the present analysis. Finally, although the detection of spermatozoa is specific for recent exposure to semen, it is not a sensitive marker and cases of exposure might have been missed [29, 39]. Strengths of our study included our use of data from a large, prospective study in which semen exposure was assessed by an objective measure and our use of case-crossover analyses, which allowed us to reduce possible effects of unmeasured time-independent confounding by comparing women to their own control visits.

The detection of BV among women who have reported being sexually abstinent has been an argument against the role of sexual activity as a necessary component in causing BV [40, 41]. However, results of a recent study among young adults with a laboratory-diagnosed case of chlamydial infection, gonorrhea, or trichomoniasis showed that 10% reported having abstained from penile-vaginal intercourse in the previous year and that 6% reported never having had intercourse [42]. Thus, imperfect validity of self reports could explain the occurrence of BV among women reporting abstinence in prior studies. While study findings implicate the role of sexual exposure in the development of incident BV, this does not necessarily mean that BV is caused by the transmission of specific organism(s) during intercourse. Semen exposure could also increase women’s risk for incident BV by increasing vaginal pH levels, changing the growth patterns in bacteria populations, or exposing women to an unidentified component of semen. The present study found biological evidence of an association between semen exposure and incident BV, which provides new support for the sexual transmission of BV; however, the mechanism remains unknown.

Acknowledgments

This study was supported by cooperative agreement Nos. U64/CCU106795, U64/CCU206798, U64/CCU306802, and U64/CCU506831 with the Centers for Disease Control and Prevention. The HIV Epidemiology Research Study Group consists of the following: Robert S. Klein, MD, Ellie Schoenbaum, MD, Julia Arnsten, MD, MPH, Robert D. Burk, MD, Chee Jen Chang, PhD, Penelope Demas, PhD, and Andrea Howard, MD, MSc, from Montefiore Medical Center and the Albert Einstein College of Medicine; Paula Schuman, MD, and Jack Sobel, MD, from the Wayne State University School of Medicine; Anne Rompalo, MD, David Vlahov, PhD, and David Celentano, PhD, from the Johns Hopkins University School of Medicine; Charles Carpenter, MD, and Kenneth Mayer, MD, from the Brown University School of Medicine; Ann Duerr, MD, Lytt I. Gardner, PhD, Scott Holmberg, MD, Denise Jamieson, MD, MPH, Jan Moore, PhD, Ruby Phelps, Dawn Smith, MD, and Dora Warren, PhD, from the Centers for Disease Control and Prevention; Katherine Davenny, PhD, from the National Institute of Drug Abuse. The findings and conclusions in this report are those of the authors and do not necessarily represent the official views of the Centers for Disease Control and Prevention.

References

E. H. Koumans, M. Sternberg, C. Bruce et al., “The prevalence of bacterial vaginosis in the United States, 2001–2004; associations with symptoms, sexual behaviors, and reproductive health,” Sexually Transmitted Diseases, vol. 34, no. 11, pp. 864–869, 2007.

J. Mania-Pramanik, S. C. Kerkar, and V. S. Salvi, “Bacterial vaginosis: a cause of infertility?” International Journal of STD & AIDS, vol. 20, no. 11, pp. 778–781, 2009.

J. Schoeman, P. S. Steyn, H. J. Odendaal, and D. Grové, “Bacterial vaginosis diagnosed at the first antenatal visit better predicts preterm labour than diagnosis later in pregnancy,” Journal of Obstetrics & Gynaecology, vol. 25, no. 8, pp. 751–753, 2005.

H. Leitich, B. Bodner-Adler, M. Brunbauer, A. Kaider, C. Egarter, and P. Husslein, “Bacterial vaginosis as a risk factor for preterm delivery: a meta-analysis,” American Journal of Obstetrics & Gynecology, vol. 189, no. 1, pp. 139–147, 2003.

S. G. Ralph, A. J. Rutherford, and J. D. Wilson, “Influence of bacterial vaginosis on conception and miscarriage in the first trimester: cohort study,” British Medical Journal, vol. 319, no. 7204, pp. 220–223, 1999.

S. L. Hillier, R. P. Nugent, D. A. Eschenbach et al., “Association between bacterial vaginosis and preterm delivery of a low-birth-weight infant,” The New England Journal of Medicine, vol. 333, no. 26, pp. 1737–1742, 1995.

P. E. Hay, R. F. Lamont, D. Taylor-Robinson, D. J. Morgan, C. Ison, and J. Pearson, “Abnormal bacterial colonisation of the genital tract and subsequent preterm delivery and late miscarriage,” British Medical Journal, vol. 308, no. 6924, pp. 295–298, 1994.

M. G. Gravett, H. P. Nelson, T. DeRouen, C. Critchlow, D. A. Eschenbach, and K. K. Holmes, “Independent associations of bacterial vaginosis and Chlamydia trachomatis infection with adverse pregnancy outcome,” Journal of the American Medical Association, vol. 256, no. 14, pp. 1899–1903, 1986.

R. B. Ness, K. E. Kip, S. L. Hillier et al., “A cluster analysis of bacterial vaginosis-associated microflora and pelvic inflammatory disease,” American Journal of Epidemiology, vol. 162, no. 6, pp. 585–590, 2005.

R. B. Ness, S. L. Hillier, K. E. Kip et al., “Bacterial vaginosis and risk of pelvic inflammatory disease,” Obstetrics & Gynecology, vol. 104, no. 4, pp. 761–769, 2004.

J. F. Peipert, A. B. Montagno, A. S. Cooper, and C. J. Sung, “Bacterial vaginosis as a risk factor for upper genital tract infection,” American Journal of Obstetrics & Gynecology, vol. 177, no. 5, pp. 1184–1187, 1997.

R. M. Brotman, M. A. Klebanoff, T. R. Nansel et al., “Bacterial vaginosis assessed by gram stain and diminished colonization resistance to incident gonococcal, chlamydial, and trichomonal genital infection,” Journal of Infectious Diseases, vol. 202, no. 12, pp. 1907–1915, 2010.

J. Atashili, C. Poole, P. M. Ndumbe, A. A. Adimora, and J. S. Smith, “Bacterial vaginosis and HIV acquisition: a meta-analysis of published studies,” AIDS, vol. 22, no. 12, pp. 1493–1501, 2008.

S. Rugpao, S. Sriplienchan, K. Rungruengthanakit et al., “Risk factors for bacterial vaginosis incidence in young adult thai women,” Sexually Transmitted Diseases, vol. 35, no. 7, pp. 643–648, 2008.

R. Kaul, N. J. Nagelkerke, J. Kimani et al., “Prevalent herpes simplex virus type 2 infection is associated with altered vaginal flora and an increased susceptibility to multiple sexually transmitted infections,” Journal of Infectious Diseases, vol. 196, no. 11, pp. 1692–1697, 2007.

P. Thorsen, I. Vogel, K. Molsted et al., “Risk factors for bacterial vaginosis in pregnancy: a population-based study on Danish women,” Acta Obstetricia et Gynecologica Scandinavica, vol. 85, no. 8, pp. 906–911, 2006.

D. H. Watts, M. Fazzari, H. Minkoff, et al., “Effects of bacterial vaginosis and other genital infections on the natural history of human papillomavirus infection among HIV-1-infected and high-risk HIV-1-uninfected women,” Journal of Infectious Diseases, vol. 191, no. 7, pp. 1129–1139, 2005.

T. L. Cherpes, L. A. Meyn, M. A. Krohn, J. G. Lurie, and S. L. Hillier, “Association between acquisition of herpes simplex virus type 2 in women and bacterial vaginosis,” Clinical Infectious Diseases, vol. 37, no. 3, pp. 319–325, 2003.

H. C. Wiesenfeld, S. L. Hillier, M. A. Krohn, D. V. Landers, and R. L. Sweet, “Bacterial vaginosis is a strong predictor of Neisseria gonorrhoeae and Chlamydia trachomatis infection,” Clinical Infectious Diseases, vol. 36, no. 5, pp. 663–668, 2003.

J. M. Marrazzo, D. H. Martin, D. H. Watts et al., “Bacterial vaginosis: identifying research gaps proceedings of a workshop sponsored by DHHS/NIH/NIAID,” Sexually Transmitted Diseases, vol. 37, no. 12, pp. 732–744, 2010.

W. E. Josey and J. R. Schwebke, “The polymicrobial hypothesis of bacterial vaginosis causation: a reassessment,” International Journal of STD & AIDS, vol. 19, no. 3, pp. 152–154, 2008.

K. A. Fethers, C. K. Fairley, J. S. Hocking, L. C. Gurrin, and C. S. Bradshaw, “Sexual risk factors and bacterial vaginosis: a systematic review and meta-analysis,” Clinical Infectious Diseases, vol. 47, no. 11, pp. 1426–1435, 2008.

E. Holst, “Reservoir of four organisms associated with bacterial vaginosis suggests lack of sexual transmission,” Journal of Clinical Microbiology, vol. 28, no. 9, pp. 2035–2039, 1990.

R. S. McClelland, B. A. Richardson, S. M. Graham et al., “A prospective study of risk factors for bacterial vaginosis in HIV-1-seronegative African women,” Sexually Transmitted Diseases, vol. 35, no. 6, pp. 617–623, 2008.

M. Maclure, “The case-crossover design: a method for studying transient effects on the risk of acute events,” American Journal of Epidemiology, vol. 133, no. 2, pp. 144–153, 1991.

D. K. Smith, D. L. Warren, D. Vlahov et al., “Design and baseline participant characteristics of the human immunodeficiency virus epidemiology research (HER) study: a prospective cohort study of human immunodeficiency virus infection in US women,” American Journal of Epidemiology, vol. 146, no. 6, pp. 459–469, 1997.

D. J. Jamieson, A. Duerr, R. S. Klein et al., “Longitudinal analysis of bacterial vaginosis: findings from the HIV epidemiology research study,” Obstetrics & Gynecology, vol. 98, no. 4, pp. 656–663, 2001.

R. P. Nugent, M. A. Krohn, and S. L. Hillier, “Reliability of diagnosing bacterial vaginosis is improved by a standardized method of gram stain interpretation,” Journal of Clinical Microbiology, vol. 29, no. 2, pp. 297–301, 1991.

W. W. Young, A. C. Bracken, M. A. Goddard, and S. Matheson, “Sexual assault: review of a national model protocol for forensic and medical evaluation,” Obstetrics & Gynecology, vol. 80, no. 5, pp. 878–883, 1992.

E. M. Silverman and A. G. Silverman, “Persistence of spermatozoa in the lower genital tracts of women,” Journal of the American Medical Association, vol. 240, no. 17, pp. 1875–1877, 1978.

A. Davies and E. Wilson, “The persistence of seminal constituents in the human vagina,” Forensic Science, vol. 3, no. 1, pp. 45–55, 1974.

K. B. Hutchinson, K. E. Kip, and R. B. Ness, “Condom use and its association with bacterial vaginosis and bacterial vaginosis-associated vaginal microflora,” Epidemiology, vol. 18, no. 6, pp. 702–708, 2007.

M. Yotebieng, A. N. Turner, T. H. Hoke, K. Van Damme, J. R. Rasolofomanana, and F. Behets, “Effect of consistent condom use on 6-month prevalence of bacterial vaginosis varies by baseline BV status,” Tropical Medicine & International Health, vol. 14, no. 4, pp. 480–486, 2009.

M. Riggs, M. Klebanoff, T. Nansel, J. Zhang, J. Schwebke, and W. Andrews, “Longitudinal association between hormonal contraceptives and bacterial vaginosis in women of reproductive age,” Sexually Transmitted Diseases, vol. 34, no. 12, pp. 954–959, 2007.

G. J. F. Priestley, B. M. Jones, J. Dhar, and L. Goodwin, “What is normal vaginal flora?” Genitourinary Medicine, vol. 73, no. 1, pp. 23–28, 1997.

R. M. Brotman, J. Ravel, R. A. Cone, and J. M. Zenilman, “Rapid fluctuation of the vaginal microbiota measured by Gram stain analysis,” Sexually Transmitted Infections, vol. 86, no. 4, pp. 297–302, 2010.

R. M. Brotman, K. G. Ghanem, M. A. Klebanoff, T. E. Taha, D. O. Scharfstein, and J. M. Zenilman, “The effect of vaginal douching cessation on bacterial vaginosis: a pilot study,” American Journal of Obstetrics & Gynecology, vol. 198, no. 6, pp. 628.e1–628.e7, 2008.

T. R. Nansel, M. A. Riggs, K. F. Yu, W. W. Andrews, J. R. Schwebke, and M. A. Klebanoff, “The association of psychosocial stress and bacterial vaginosis in a longitudinal cohort,” American Journal of Obstetrics & Gynecology, vol. 194, no. 2, pp. 381–386, 2006.

J. F. Culhane, P. Nyirjesy, K. McCollum, G. Casabellata, M. Di Santolo, and S. Cauci, “Evaluation of semen detection in vaginal secretions: comparison of four methods,” American Journal of Reproductive Immunology, vol. 60, no. 3, pp. 274–281, 2008.

S. Yen, M. A. Shafer, J. Moncada, C. J. Campbell, S. D. Flinn, and C. B. Boyer, “Bacterial vaginosis in sexually experienced and non-sexually experienced young women entering the military,” Obstetrics & Gynecology, vol. 102, no. 5, part 1, pp. 927–933, 2003.

R. C. Bump and W. J. Buesching, “Bacterial vaginosis in virginal and sexually active adolescent females: evidence against exclusive sexual transmission,” American Journal of Obstetrics & Gynecology, vol. 158, no. 4, pp. 935–939, 1988.

R. J. DiClemente, J. M. Sales, F. Danner, and R. A. Crosby, “Association between sexually transmitted diseases and young adults' self-reported abstinence,” Pediatrics, vol. 127, no. 2, pp. 208–213, 2011.