Incidence & Predictors of Treatment Failure for Bacterial Vaginosis

JM Marrazzo, KK Thomas, K Ringwood, T Fiedler, DN Fredricks

University of Washington & Fred Hutchinson Cancer Research Center, Seattle WA

jmm2@u.washington.edu

Background Most common

Cause of vaginal symptoms prompting medical evaluation for vaginitis

Cause of vaginitis >10% of women experience BV May cause 11% of preterm deliveries in U.S.

Etiology not understood, but associated with douching, new male partner, unprotected sex in heterosexual women, sex between women

Frequently persists after treatment (15% - 20%) Very frequently recurs after successful treatment

(30% - 60%) History of BV, regular partner throughout follow-up,

and female partner implicated Sobel 2006; Bradshaw 2006

Background

Traditionally defined as overgrowth of commensal anaerobic flora (classically G. vaginalis, Prevotella, Mobiluncus, M. hominis) relative to H2O2+ lactobacilli

Molecular (cultivation-independent) approaches have recently expanded spectrum of BV microbiology

Some bacteria highly specific for BV in recent studies Some with known high-level resistance to metronidazole

Atopobium vaginae “New” bacteria not yet incorporated into prospective analyses for

BV persistence or recurrence

Verstraelen 2004, Ferris 2004, Fredricks 2005

BV-associated bacteria (BVAB 1, 2, 3) and their relationship to

bacteria in the Clostridium phylum

Clostridium Cluster XIVa

BVAB1

BVAB2 BVAB3

Methods

Subjects recruited to research clinic 16-29 years Sex with woman (prior year) Recruited through ads (self-referred) and partners

Underwent computer-assisted self-interview (CASI) at enrollment

BV diagnosed by Amsel criteria and treated with vaginal metronidazole BV confirmed by Nugent criteria (later)

Vaginal fluid collected with polyurethane foam swab and saline lavage (0.5 cc)

Methods

All women with BV asked to return at 1 month for repeat procedures (CASI, vaginal fluid collection) BV persistence = Nugent >6 Abnormal vaginal flora = Nugent >3

All women asked to return at 3 months BV recurrence defined by Nugent score >6 among

women whose baseline BV was cured at 1 month

MethodsVaginal Fluid Collected at All Visits: Traditional culture

Used to define H2O2 production by lactobacilli Typical panel of anaerobic flora

Bacterium-specific PCR assays based on cloned 16s rDNA sequences derived from earlier analysis of clones by RFLP, sequencing; all positive results sequence-confirmed

BVAB1 Leptotrichia spp Mobiluncus curtisii

BVAB2 Peptoniphilus spp Mobiluncus mulieris

BVAB3 P. lacrimalis Prevotella G1

Megasphaera phylotype 1 (elsdensii-like)

Megasphaera phylotype 2 (micronucliformis-like)

Lactobacillus crispatus

Lactobacillus iners

Eggerthella-like uncultured bacterium

Gardnerella vaginalis Atopobium spp

ResultsBaseline and Persistent BV

239 women enrolled

BVN = 12 / 72 (17%)

No BVN = 60 / 72 (83%)

Characteristics:Median age 25 yNonwhite: 25%

Sex with male last 3 mos: 28%Douche last 3 mos: 6.4%

At 1 month

BV @ baseline

N = 66 (28%)

No BVN = 173 (72%)

New BV during study

N = 6+

• 28% had BV at baseline

• Overall f/u at 1 month = 90%

• 83% responded to vaginal MTZ

• Persistent BV incidence: 17%

72 women with BV

0102030405060708090

100

BVAB1 BVAB2 BVAB3

0102030405060708090

100

Age >25 Black

YesNo

Baseline Vaginal Microbiology (PCRs)

Demographics

OR (95% CI)

Age 26-33 y 3.9 (0.9, 16.4)

Black race 7.0 (3.9, 12.4)

P=0.15P=0.67

P=0.009

P=0.06

OR (95% CI)

BVAB1 2.3 (0.8, 6.5))

BVAB2 2.2 (0.3, 15.5)

BVAB3 4.4 (1.5, 13.4)

ResultsFactors Associated with BV Persistence

Incidence of BV

Recurrence (%

)

P=0.03

0102030405060708090

100

BVAB1+ Adherentto Tx

Any Sex VaginalSex*

YesNo

OR (95% CI)

BVAB1 2.6 (1.3, 5.2)

Adherence 0.36 (0.2, 0.7)

Any sex 4.1 (0.6, 28)

Vaginal sex 2.1 (1.1, 4.2)

P=0.05

*Intercourse with male partner

ResultsAdditional Factors Associated with Persistence of

Abnormal Flora (N = 22)

Incidence of BV

Recurrence (%

)

P=0.02

P=0.05

P=0.09

ResultsBaseline and Persistent BV

239 women enrolled

BVN = 12 / 72 (17%)

No BVN = 60 / 72 (83%)

At 1 month

BV @ baseline

N = 66 (28%)

No BVN = 173 (72%)

New BV during study

N = 6+

• 15% who initially responded to MTZ had BV recurrence

72 women with BV

Recurrent BVN = 8 / 53 (15%)

Conclusions• Distinct risks predict BV persistence

– Demographics• Black women consistently at high risk for BV

– Reasons unclear; not related to douching in our study

• Older age

– Vaginal microbiology• BVAB3, possibly BVAB1

– Sex • Vaginal fluid exchange, intercourse with men [Sanchez 2004,

Bradshaw 2006]

• Further study needed– Additional accrual of subjects to substantiate preliminary

trends and assess risks for BV recurrence– Cultivation of BVAB to assess pathogenicity and antibiotic

susceptibilities

Limitations• Small number of subjects who were self-

referred or referred by partners– Limits on reproducibility, generalizability – However, excellent retention and diverse sexual

practices

• Development of bacterium-specific PCRs directed by initial clone analysis– May not include all relevant species

Acknowledgements

Study personnel• Nancy Dorn • Dana Varon• Lauren Asaba• Susan Heideke• Corey Fish• Kathy Agnew• Becca Hutcheson

Support• Dave Eschenbach• Sharon Hillier• King Holmes• Larry Corey

Associations Between Bacteria Detected and BV

Bacterium-specific PCR Result

Sensi tivity (95% CI)

Specificity (95% CI)

Adjusted OR (95% CI)

BVAB 1 40.7 (22.4-61.2) 97.8 (88.5-99.9) 19.0 (2.2-910.7) BVAB 2 88.9 (70.8-97.6) 95.7 (85.2-99.5) 106.1 (14.3-4755.1) BVAB 3 40.7 (22.4-61.2) 97.8 (88.5-99.9) 21.9 (2.5-1056.4) Gardnerella 100.0 (89.5-100.0) 41.3 (27.0-56.8) 27.7 (3.9-∞) Atopobium sp. 96.3 (81.0-99.9) 80.4 (66.1-90.6) 95.0 (14.6-∞) Eggerthella-like 92.6 (75.7-99.1) 91.3 (79.2-97.6) 103.8 (13.5-4812.8) uncultured bact. Leptotrichia sp. 85.2 (70.8-97.6) 95.7 (85.2-99.5) 330.6 (23.1-∞) Megasphaera- a 96.3 (81.0-99.9) 91.3 (79.2-97.6) 134.4 (16.6-6509.8)

Bacterium-specific PCR assays for vaginal fluid samples from 40 subjects with BV and 65 without BV. ORs adjusted for age, site of enrollment, vaginal symptoms, report of

sex with men