Evaluation of Clinical Methods for Diagnosing
Bacterial Vaginosis
Robert E. Gutman, MD, Jeffrey F. Peipert, MD, MPH, Sherry Weitzen, PhD, and Jeffrey Blume, PhD
OBJECTIVE: To determine whether the current clinical crite-
ria for diagnosing bacterial vaginosis can be simplified by
using 2 clinical criteria rather than the standard 3 of 4
criteria (Amsels criteria).
METHODS: This was a prospective observational study of
269 women undergoing a vaginal examination in the
Womens Primary Care Center, Division of Research, or
Colposcopy Clinic at Women & Infants Hospital. All 4
clinical criteria for diagnosing bacterial vaginosis were
collected, and Gram stain was used as the gold standard.
Sensitivity and specificity were calculated for each individ-
ual criterion, combinations of criteria, and a colorimetric
pH and amine card. Receiver operating characteristic
curve was generated to estimate the preferred pH and
percentage of clue cells for diagnosing bacterial vaginosis.
RESULTS: The prevalence of bacterial vaginosis in our
study population was 38.7%. Vaginal pH was the most
sensitive of all the criteria, at 89%, and a positive amine
odor was the individual criteria with the highest specificity,
at 93%. Similar specificity was seen with combinations of 2
criteria and Amsels criteria. Receiver operating character-
istic curve analysis yielded a preferred pH and percentage
of clue cells of 5.0 and 20%, respectively. However, a pH of
4.5 or greater improves sensitivity with minimal loss of
specificity.
CONCLUSION: The clinical criteria for diagnosing bacterial
vaginosis can be simplified to 2 clinical criteria without loss
of sensitivity and specificity. (Obstet Gynecol 2005;105:
551 6. 2005 by The American College of Obstetricians
and Gynecologists.)
LEVEL OF EVIDENCE: II-2
Bacterial vaginosis is the most common vaginal infec-
tion, with a prevalence of 9 37%, depending on the
population studied.1,2 Bacterial vaginosis has been asso-
From the Division of Research, Department of Obstetrics and Gynecology, Women
& Infants Hospital, and the Center for Statistical Sciences, Brown University,
Medical School, Providence, Rhode Island.
Preliminary data presented at the Infectious Disease Society of Obstetrics and
Gynecology (IDSOG) Annual Meeting in Quebec City, Canada, August 9, 2001.
The authors thank CooperSurgical for the donation of the FemExam cards used in
this study.
VOL. 105, NO. 3, MARCH 2005
2005 by The American College of Obstetricians and Gynecologists.
Published by Lippincott Williams & Wilkins.
ciated with increased risk of preterm delivery,3 prema-
ture rupture of membranes,4 amniotic fluid infection,5
chorioamnionitis,6 postpartum endometritis,79 pelvic
inflammatory disease,10 12 and postoperative infec-
tions.13,14
Currently, the clinical criteria for diagnosing bacterial
vaginosis requires the presence of 3 of the following 4
criteria: 1) a thin, homogeneous discharge, 2) vaginal pH
greater than 4.5, 3) a positive whiff test or release of
amine odor with the addition of base, and 4) clue cells on
microscopic evaluation of saline wet preparation. These
criteria are based on the original work of Amsel et al.15
The Gram stain is believed by many to be the gold
standard for diagnosing bacterial vaginosis. However,
interpreting the Gram stain requires experience, and it is
often difficult to get timely results for the clinical diagno-
sis of bacterial vaginosis. Thus, it is important to have
simple and reliable clinical criteria that clinicians can use
in practice. Because many clinicians do not routinely
evaluate patients for all 4 of Amsels criteria, it would be
helpful to know if different combinations of criteria can
be used to accurately diagnose bacterial vaginosis.
Although many research studies have looked at the
sensitivity and specificity of Amsels criteria, few have
looked at these criteria individually and in various com-
binations using the Gram stain as the gold standard. We
conducted a MEDLINE search from 1966 to November
2003, using the terms bacterial vaginosis, BV, and
nonspecific vaginitis. We were unable to find any
studies that evaluate all of the different combinations of
clinical criteria for the purpose of simplifying and possi-
bly improving the accuracy of current clinical methods
for diagnosing bacterial vaginosis.
We hypothesize that the current clinical criteria for
diagnosing bacterial vaginosis can be simplified by using
2 clinical criteria, rather than the standard 3 of 4 criteria
(Amsels criteria), without loss of sensitivity or specificity.
Furthermore, we believe that the addition of the FemExam
card (CooperSurgical Inc, Trumbull, CT), a colorimetric
pH and amine card, will have better sensitivity and speci-
ficity when compared with Amsels criteria.
0029-7844/05/$30.00 551
doi:10.1097/01.AOG.0000145752.97999.67
MATERIALS AND METHODS
We performed a prospective observational study of 269
women recruited from various locations at Women &
Infants Hospital over a 20-month period, beginning Au-
gust 1, 2000. Any woman undergoing a speculum exam-
ination at the Womens Primary Care Center, Colpos-
copy Clinic, or Division of Research was eligible to
participate in the study. Because all specimens were
taken in patients who were already undergoing exami-
nations, the study was approved by our Institutional
Review Board as a residual tissue study. Thus, a sepa-
rate, specific informed consent was not obtained, and
information from the study was not used for clinical care
or the management of the patient. Women were ex-
cluded if there was a large amount of vaginal bleeding on
examination. Routine pelvic examinations were per-
formed by select second- through fourth-year obstetric
and gynecology residents, research nurses, or an attend-
ing gynecologist (J.F.P.). All of the examining staff were
trained in the use of the FemExam card by a CooperSur-
gical representative.
Descriptive variables obtained during evaluation in-
cluded age, pregnancy status, parity, ethnicity, presence
or absence of symptoms, and a sexually transmitted dis-
eases history. The following procedures were used to estab-
lish the presence or absence of the 4 Amsel criteria: vaginal
discharge was characterized according to its color and the
presence of a thin homogeneous discharge. Swabs of the
vaginal secretions were taken from the sidewalls. We
placed the secretions from one swab on ColorpHast pH
indicator strips (EM Science, Gibbstown, NJ) with a pH
range of 4.0 7.0 to determine vaginal pH. Vaginal secre-
tions were then combined with 2 drops of normal saline on
a slide and covered with a coverslip for the wet mount.
Examination under high-powered microscopy identified
the percentage of clue cells. We added 10% potassium
hydroxide solution to the vaginal secretions from the sec-
ond swab, and the release of a fishy amine odor signified
a positive whiff test. We then rolled the third swab along
a glass slide and allowed it to air dry for the Gram stain. An
assistant placed vaginal secretions from the final swab on
the FemExam card. This card has 2 colorimetric test circles
for the detection of vaginal pH of 4.7 or greater and volatile
vaginal fluid amines. The individual test circles form a blue
plus () sign when the result is positive and blue minus ()
sign when the result is negative. To minimize bias of the
clinical evaluation, the examiner read the FemExam card
immediately after recording all other information.
Patients found to have bacterial vaginosis by clinical
criteria were treated according to the current standard of
care. We sent the Gram stain to Magee Womens Hos-
pital research laboratory where a standardized 0 10
552 Gutman et al Clinical Diagnosis of Bacterial Vaginosis
point score was assigned. According to Nugents crite-
ria,16 a score of 7 or greater defined the gold standard
diagnosis of bacterial vaginosis.
We calculated the sensitivity, specificity, and 95%
confidence intervals for each of the individual criterion,
combinations of the criteria, Amsels criteria, and the
FemExam card. Patient characteristics were compared
between women with and those without bacterial vagi-
nosis by using the Student t test and Wilcoxon rank-sum
test for continuous data and 2 test for nonparametric
categorical data. We generated a receiver operating char-
acteristic curve to estimate the preferred pH and percent-
age of clue cells for the diagnosis of bacterial vaginosis.
RESULTS
The prevalence of bacterial vaginosis in our study pop-
ulation was 38.7%. Table 1 displays the demographic
and clinical characteristics of the participants. Women
with bacterial vaginosis had slightly higher mean age
(25.4 versus 23.3 years, P .03) and median parity (1.0
versus 0, P .01). Black and Hispanic women had a
higher prevalence of bacterial vaginosis than white
women (P .02). Our population consisted of 37.5%
white women, 30.4% black women, and 27.6% Hispanic
women. Women with bacterial vaginosis were more
likely to be symptomatic, with individual symptoms of
vaginal discharge and foul odor being more common
(P .01). Those with bacterial vaginosis were also more
likely to have a history of Neisseria gonorrhoeae infection
(P .04) and trichomoniasis (P .02), but a history of at
least one sexually transmitted disease was not signifi-
cantly increased in the women with bacterial vaginosis.
Examination and recruitment location did not affect the
prevalence of bacterial vaginosis.
The sensitivity and specificity and their respective
95% confidence intervals of individual criterion and
combinations of criteria are shown in Table 2. Vaginal
pH was the criteria with the highest sensitivity, at 89%.
The sensitivity of the remaining 3 individual criteria
ranged from 67% to 79%, and any combination of 2
criteria ranged from 61% to 69%. Amsels criteria had a
sensitivity of 69%. Positive amine odor was the individ-
ual criteria with the highest specificity, at 93%. Similar
specificity was seen with combinations of 2 criteria and
Amsels criteria (86 95%). Thin, homogeneous dis-
charge had the lowest specificity, at 54%.
The sensitivity, specificity, and 95% confidence inter-
vals of the FemExam card criteria are shown in Table 3.
The FemExam card had a similar sensitivity, but lower
specificity for pH, and a similar specificity but lower
sensitivity for the presence of amines when compared to
the whiff test. When 1 of the 2 criteria was present on
OBSTETRICS & GYNECOLOGY
Table 1. Demographic and Clinical Characteristics of Patients
Bacterial Vaginosis No Bacterial Vaginosis
(n 104) (n 165) P
Age (y) (mean SD) 25.4 7.8 23.3 6.5 .027
Median parity (range) 1.0 (05) 0 (07) .008
Pregnancy status .085
Pregnant 9 (9) 26 (16)
Nonpregnant 95 (91) 137 (84)
Ethnicity .015
White 28 (27) 75 (46)
Black 37 (35) 43 (26)
Hispanic 31 (30) 41 (25)
Asian 0 2 (1)
Other 8 (8) 4 (2)
Symptomatology
Any symptoms 47 (45) 41 (25) .001
Vaginal discharge 39 (38) 25 (15) .001
Foul smelling odor 26 (25) 12 (7) .001
Vaginal itching 5 (5) 12 (7) .418
Vaginal burning 1 (1) 6 (4) .180
History of STD 57 (55) 78 (47) .223
Trichomonas 14 (13) 9 (5) .022
Gonorrhea 19 (18) 16 (10) .042
Chlamydia 45 (43) 54 (33) .081
Herpes 4 (4) 10 (6) .426
HIV 0 1 (1) .426
Other 9 (9) 16 (10) .774
SD, standard deviation; STD, sexually transmitted disease; HIV, human immunodeficiency virus.
Data are presented as n (%) unless otherwise indicated.

the card, the sensitivity was 89% and specificity was 61%. A receiver operating characteristic curve was gener-
When both of the criteria were present, the sensitivity ated for pH and clue cells on wet preparation. We
was 40% and the specificity was 95%. visually estimated the point of inflection on the receiver

Table 2. Sensitivity, Specificity, and 95% Confidence Intervals of the Clinical Criteria for Diagnosing Bacterial Vaginosis
Sensitivity 95% CI Specificity 95% CI AUC 95% CI
Thin homogeneous discharge 0.79 0.690.87 0.54 0.460.62 0.77 0.690.84
pH 4.5 0.89 0.820.95 0.74 0.660.80 0.82 0.760.86
Positive amine odor 0.67 0.570.76 0.93 0.880.97 0.80 0.750.85
Clue cells present ( 20% per hpf) 0.74 0.650.82 0.86 0.800.91 0.80 0.750.85
pH 4.5 and thin homogeneous discharge 0.69 0.590.79 0.86 0.800.91 0.78 0.720.83
pH 4.5 and amine odor 0.64 0.540.73 0.95 0.910.98 0.80 0.740.84
pH 4.5 and clue cells 0.69 0.590.78 0.92 0.860.95 0.80 0.750.85
Clue cells and amine odor 0.63 0.530.73 0.95 0.900.97 0.79 0.740.84
Clue cells and thin homogeneous discharge 0.61 0.500.71 0.91 0.860.95 0.76 0.700.81
Amine odor and thin homogeneous 0.58 0.470.68 0.94 0.890.97 0.76 0.700.81
discharge
Amsels criteria ( 3 of 4 criteria) 0.69 0.590.78 0.93 0.870.96 0.81 0.760.86
CI, confidence interval; AUC, area under the curve; hpf, high-power field.

Table 3. Sensitivity, Specificity, and 95% Confidence Intervals of the FemExam Card for Diagnosing Bacterial Vaginosis
Sensitivity 95% CI Specificity 95% CI AUC 95% CI
pH positive 0.88 0.800.93 0.64 0.560.72 0.76 0.700.81
Amine positive 0.41 0.320.51 0.91 0.850.95 0.66 0.600.72
pH or amine positive 0.89 0.810.94 0.61 0.530.68 0.75 0.690.79
pH and amine positive 0.40 0.310.50 0.95 0.900.97 0.67 0.610.73
CI, confidence interval; AUC, area under the curve.

VOL. 105, NO. 3, MARCH 2005 Gutman et al Clinical Diagnosis of Bacterial Vaginosis 553

Fig. 1. Receiver operating characteristic curve for pH.
Gutman. Clinical Diagnosis of Bacterial Vaginosis. Obstet Gynecol 2005.
operating characteristic curve. A pH of 5.0 or greater
maximized sensitivity (83%) and specificity (82%). How-
ever, a pH of 4.5 or more improves sensitivity to over
89% while still maintaining a relatively high specificity of
74% (Fig. 1). Sensitivity and specificity of clue cells appears
to be maximized with a cutoff of more than 20% clue cells
on saline wet preparation (Fig. 2; sensitivity 74%, specific-
ity, 86%).
DISCUSSION
The prevalence of bacterial vaginosis was higher in our
study (39%) than in most studies,1,2,17 and thus repre-
sents a high-risk population. Although the ethnic diver-
sity of this population is not representative of most U.S.
communities, it provides us with a better sampling of
Fig. 2. Receiver operating characteristic curve for clue
cells.
Gutman. Clinical Diagnosis of Bacterial Vaginosis. Obstet Gynecol 2005.
554 Gutman et al Clinical Diagnosis of Bacterial Vaginosis
black and Hispanic women, who appear to be at higher
risk of having bacterial vaginosis than white women.17
We confirmed the findings seen in another large cross-
sectional study, which showed that patients with bacte-
rial vaginosis are more likely to have vaginal symptoms,
specifically vaginal discharge and foul odor.17,18 A study
by Larsson et al19 revealed similar risk factors for sexu-
ally transmitted diseases and bacterial vaginosis, includ-
ing lower age of first intercourse and higher number of
lifetime sexual partners. Yen et al17 displayed a lower
rate of bacterial vaginosis among the nonsexually ex-
perienced and a higher rate among those with multiple
sexual partners in the past 3 months. However, in our
study and in Yens, a history of a sexually transmitted
disease was not associated with an increased prevalence
of bacterial vaginosis.
We confirmed the findings of Eschenbach et al18 that
vaginal pH had the highest sensitivity, but the lowest
specificity, of all clinical diagnostic criteria. The low
specificity is not surprising considering that elevated pH
may be altered by semen, cervical mucus, and blood. An
elevated pH is more common in postmenopausal wom-
en20 and can also be seen with trichomoniasis infections.
The high sensitivity should serve as an asset in screening
algorithms. Our findings for amine odor also support
previous studies that found it to have the highest speci-
ficity and lowest sensitivity of the individual criteria.18,21
The combination of any 2 criteria decreased sensitivity
and increased specificity. However, the sensitivity and
specificity of each pair of criteria is similar to those
observed for Amsels criteria. This implies that there is
no clear advantage to using Amsels criteria ( 3 of the 4
criteria). This supports our hypothesis that the current
clinical criteria for diagnosing bacterial vaginosis (Am-
sels criteria) can be simplified by using 2 clinical criteria,
without significant loss of sensitivity or specificity.
One study evaluating an objective colorimetric test for
amines found a sensitivity of 87% and specificity of 98%
compared with Gram-stain criteria.22 The authors con-
cluded that these tests are more sensitive than the whiff
test for detecting the presence of amines. We found the
FemExam card to have less sensitivity than the whiff
test for detecting amines and lower specificity for detect-
ing an elevated vaginal pH. The difference in pH may be
due to the different cutoff points: 4.5 for our study and
4.7 for the FemExam card. Overall, we felt the majority
of the differences could be explained by the difficulty in
reading and interpreting the results of the card. Although
it is conceptually easy to distinguish between a plus sign
and a minus sign, the results often had very faint blue
signals that were difficult to interpret. The findings may
also be biased because the card was read after the clinical
criteria were collected and recorded, although we would
OBSTETRICS & GYNECOLOGY
have expected this to falsely raise the sensitivity and
specificity. Also, the slight delay in reading the cards may
have resulted in suboptimal outcomes. Ultimately, the
pH measurements are not statistically different based on
the area under the receiver operating characteristic curve
and 95% CI, but the amine measurements appear to be
worse for the card because of the reasons listed above.
This difference persists when pH and amine are both
positive because of the large discrepancy in the sensitiv-
ity of amine measurements. The findings of this study
fail to support routine use of the FemExam card to
improve sensitivity and specificity.
The receiver operating characteristic curve confirms
previously set cutoff points for pH and percentage of clue
cells. The presence of greater than 20% clue cells on
saline wet preparation appears to optimize sensitivity
and specificity (Fig. 2). Although a pH of 5.0 or greater
appears to optimize sensitivity and specificity, we prefer
a pH of 4.5 or greater to improve sensitivity with an
acceptable decline in specificity (Fig. 1). This is consistent
with the majority of studies that use a pH of 4.5 or 4.7 for
the cutoff of clinically diagnosing bacterial vaginosis.
Before initiating the study, we did a power and sample
size calculation. Based on our desire to have relatively
narrow 95% confidence intervals ( 7%) with an esti-
mated prevalence of 25%, we calculated a sample size of
approximately 500 patients. Our population, however,
had a higher prevalence of bacterial vaginosis and ulti-
mately resulted in a diagnosis of bacterial vaginosis by
Gram-stain criteria in 104 patients. Consequently, de-
spite having 131 fewer patients than our target, we had
only 21 fewer patients with bacterial vaginosis. This
resulted in slightly larger 95% confidence intervals than
anticipated. There is overlap between the confidence
intervals for sensitivity and specificity and the area under
the curve among individual criteria and combinations of
criteria (Table 2). Because of these similarities, a much
larger sample size would be required to detect a statisti-
cally significant difference. The relatively large number
of patients with bacterial vaginosis, the use of Gram stain
as the gold standard diagnosis of bacterial vaginosis, and
careful analytic methods, including receiver operating
characteristic curve analysis, are strengths of this study.
The high prevalence of bacterial vaginosis, however,
may limit the generalizability of the results.
Because of the superior sensitivity of pH for the detec-
tion of bacterial vaginosis, we recommend an algorithm
that involves initial diagnostic testing for bacterial vagi-
nosis with pH, using test paper with an appropriate pH
range (pH 4.0 6.0). Testing for one of the other 3
criteria should then be performed. If the first 2 tests are
positive for bacterial vaginosis, the diagnosis can be
made and appropriate treatment given. If one or both
VOL. 105, NO. 3, MARCH 2005
tests are negative, then additional tests should be per-
formed until a diagnosis is made. This algorithm is only
indicated for the detection of bacterial vaginosis and
cannot be used to rule out other infectious processes that
may occur in conjunction with bacterial vaginosis. Ap-
propriate testing, including microscopy, should be per-
formed as clinically indicated to rule out other infectious
etiologies. In more complicated cases of recurrent or
persistent bacterial vaginosis, a Gram stain should be
considered to corroborate the diagnosis.
In conclusion, we believe that the evidence in this
study supports simplifying the clinical diagnosis of bac-
terial vaginosis. We recommend using any 2 of the 4
clinical criteria to achieve a test performance similar to
that achieved by using the criteria established by Amsel
et al.15
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Reprints are not available. Address correspondence to: Robert
E. Gutman, MD, 4940 Eastern Avenue, Room 125, Baltimore,
MD 21224-2780; e-mail: rgutman1@jhmi.edu.
Received June 3, 2004. Received in revised form September 7, 2004.
Accepted October 7, 2004.
OBSTETRICS & GYNECOLOGY