pubmed.ncbi.nlm.nih.gov

Composition of the vaginal microbiota in women of reproductive age--sensitive and specific molecular diagnosis of bacterial vaginosis is possible? - PubMed

  • ️Tue Jan 01 2013

Composition of the vaginal microbiota in women of reproductive age--sensitive and specific molecular diagnosis of bacterial vaginosis is possible?

Elena Shipitsyna et al. PLoS One. 2013.

Abstract

Background and objective: Bacterial vaginosis (BV) is the most common vaginal disorder, characterized by depletion of the normal lactobacillus-dominant microbiota and overgrowth of commensal anaerobic bacteria. This study aimed to investigate the composition of the vaginal microbiota in women of reproductive age (healthy women and women with BV), with the view of developing molecular criteria for BV diagnosis.

Materials and methods: Vaginal samples from 163 women (79 control, 73 BV and 11 intermediate (Lactobacillary grade II flora) cases) were analyzed using 454 pyrosequencing of the hypervariable regions V3-V4 of the 16S rRNA gene and 16 quantitative bacterial species/genus-specific real-time PCR assays. Sensitivities and specificities of potential BV markers were computed using the Amsel criteria as reference standard for BV. The use of quantitative thresholds for prediction of BV, determined for both relative abundance measured with 454 pyrosequencing and bacterial load measured with qPCR, was evaluated.

Results: Relative to the healthy women, the BV patients had in their vaginal microbiota significantly higher prevalence, loads and relative abundances of the majority of BV associated bacteria. However, only Gardnerella vaginalis, Atopobium vaginae, Eggerthella, Prevotella, BVAB2 and Megasphaera type 1 detected at or above optimal thresholds were highly predictable for BV, with the best diagnostic accuracy shown for A. vaginae. The depletion of Lactobacillus species combined with the presence of either G. vaginalis or A. vaginae at diagnostic levels was a highly accurate BV predictor.

Conclusions: Quantitative determination of the presence of G. vaginalis, A. vaginae, Eggerthella, Prevotella, BVAB2 and Megasphaera type 1 as well as the depletion of Lactobacillus was highly accurate for BV diagnosis. Measurements of abundance of normal and BV microbiota relative to total bacteria in vaginal fluid may provide more accurate BV diagnosis, and be used for test-of-cure, rather than qualitative detection or absolute counts of BV related microorganisms.

PubMed Disclaimer

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Heat map based on taxon visualized with MultiExperiment Viewer.

The figure shows clustering of the samples based on the Pearson correlation using absolute distances and complete linkage clustering with a cut-off level set to 1%. The individuals are shown on the x-axis and the different taxons on the y-axis. The relative abundance of each taxon is correlated to the color of the dot, where black is 1% and red is 100%. The most distinct taxons are shown to the right, colored according to the different phyla to the left.

Figure 2
Figure 2. Relative abundances of bacterial species/genera by 454 pyrosequencing.

The figure presents the average relative abundances of 16 bacterial species/genera by 454 pyrosequencing in the vaginal samples from healthy control women (n = 79), intermediate cases (n = 11) and BV patients (n = 73). The healthy control women had vaginal microbiota dominated by Lactobacillus species, accounting for nearly 90% of all sequences, with L. crispatus and L. iners highly prevailing over other lactobacilli. In the intermediate cases, L. iners and G. vaginalis were dominating over other species, accounting together for nearly 75% of all sequences. The BV patients possessed a diverse array of bacteria, with the most abundant species being G. vaginalis, Prevotella, Megasphaera type 1, A. vaginae, L. amnionii, L. iners, BVAB1, BVAB2, S. sanguinegens and Eggerthella.

Figure 3
Figure 3. Prevalence of vaginal bacterial species/genera by real-time PCR.

The qualitative detection rates of 16 bacteria by species/genus-specific PCR assays in healthy control women (n = 79), intermediate cases (n = 11) and BV patients (n = 73) are presented. L. iners was the most frequently detected bacterium in the healthy control women (86%), followed by G. vaginalis (78%), A. vaginae (63%), Prevotella (62%), and F. magna (58%). In the intermediate subjects, the most common species/genera were G. vaginalis (100%), L. iners (91%), A. vaginae (91%), Prevotella (82%) and L. amnionii (82%). G. vaginalis and A. vaginae were both found in 100% of the subjects with BV. Other highly prevalent bacteria in the BV patients were Prevotella (96%), Megasphaera type 1 (96%), L. iners (93%), Eggerthella (93%), BVAB2 (90%), L. amnionii (88%) and S. sanguinegens (81%).

Figure 4
Figure 4. Genome/gene copy numbers determined by specific quantitative PCR of bacterial species/genera in vaginal samples by species/genus-specific qPCR.

The scatter plots present DNA concentrations of the 16 bacteria by qPCR in the samples from healthy control women (n = 79), intermediate cases (n = 11) and BV patients (n = 73). Bars represent median concentrations. L. iners had the highest median concentration in the samples from both healthy (2.8×105 copies/µl) and intermediate (1.7×106 copies/µl) subjects. In the intermediate subjects, also G. vaginalis was present in high loads (median concentration 1.5×105 copies/µl). In the BV subjects, the highest median concentrations were displayed by G. vaginalis and A. vaginae (7.6×105 and 4.6×105 copies/µl, respectively), followed by L. iners, Megasphaera type 1 and BVAB2 (3.5×105, 2.2×105 and 1.2×105 copies/µl, respectively).

Similar articles

Cited by

References

    1. Morris M, Nicoll A, Simms I, Wilson J, Catchpole M (2001) Bacterial vaginosis: a public health review. Br J Obstet Gynaecol 108: 439–450. - PubMed
    1. Hillier SL, Nugent RP, Eschenbach DA, Krohn MA, Gibbs RS, et al. (1995) Association between bacterial vaginosis and preterm delivery of a low-birth-weight infant. The Vaginal Infections and Prematurity Study Group. N Engl J Med 333: 1737–1742. - PubMed
    1. Silver HM, Sperling RS, St Clair PJ, Gibbs RS (1989) Evidence relating bacterial vaginosis to intraamniotic infection. Am J Obstet Gynecol 161: 808–812. - PubMed
    1. Hillier SL, Martius J, Krohn M, Kiviat N, Holmes KK, et al. (1988) A case-control study of chorioamnionic infection and histologic chorioamnionitis in prematurity. N Engl J Med 319: 972–978. - PubMed
    1. Sweet RL (1995) Role of bacterial vaginosis in pelvic inflammatory disease. Clin Infect Dis 20: S271–S275. - PubMed

Publication types

MeSH terms

Substances

Grants and funding

This work was supported by general grants from the Söderberg’s Foundation, the Örebro County Council Research Committee and the Foundation for Medical Research at Örebro University Hospital, Sweden. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.