Molecular Identification of Bacteria Associated with Bacterial Vaginosis

TLDR

Bacterial vaginosis affects millions of women, is linked to serious health conditions, and its cause remains poorly understood. The study aimed to identify bacteria associated with bacterial vaginosis in vaginal fluid using culture‑independent methods. Researchers used broad‑range 16S rDNA PCR with clone analysis, bacterium‑specific 16S rDNA PCR, and fluorescence in situ hybridization on vaginal fluid samples. Women with bacterial vaginosis exhibited markedly higher bacterial diversity, with 9–17 phylotypes per sample and 35 unique species—including several uncultivated and Clostridiales members—while such diversity was absent in healthy controls.

Abstract

Bacterial vaginosis affects millions of women and is associated with several serious health conditions. The cause of bacterial vaginosis remains poorly understood despite numerous studies based on cultures. Bacteria in microbial communities can be identified without cultivation by characterizing their ribosomal DNA (rDNA) sequences.We identified bacteria in samples of vaginal fluid with a combination of broad-range polymerase-chain-reaction (PCR) amplification of 16S rDNA with clone analysis, bacterium-specific PCR assay of 16S rDNA, and fluorescence in situ hybridization (FISH) performed directly on vaginal fluid from 27 subjects with bacterial vaginosis and 46 without the condition. Twenty-one subjects were studied with the use of broad-range PCR of 16S rDNA, and 73 subjects were studied with the use of bacterium-specific PCR.Women without bacterial vaginosis had 1 to 6 vaginal bacterial species (phylotypes) in each sample (mean, 3.3), as detected by broad-range PCR of 16S rDNA, and lactobacillus species were the predominant bacteria noted (83 to 100 percent of clones). Women with bacterial vaginosis had greater bacterial diversity (P<0.001), with 9 to 17 phylotypes (mean, 12.6) detected per sample and newly recognized species present in 32 to 89 percent of clones per sample library (mean, 58 percent). Thirty-five unique bacterial species were detected in the women with bacterial vaginosis, including several species with no close cultivated relatives. Bacterium-specific PCR assays showed that several bacteria that had not been previously described were highly prevalent in subjects with bacterial vaginosis but rare in healthy controls. FISH confirmed that newly recognized bacteria detected by PCR corresponded to specific bacterial morphotypes visible in vaginal fluid.Women with bacterial vaginosis have complex vaginal infections with many newly recognized species, including three bacteria in the Clostridiales order that were highly specific for bacterial vaginosis.