Early pregnancy vaginal microbiome trends and preterm birth

Abstract

Background: Despite decades of attempts to link infectious agents to preterm birth, an exact causative microbe or community of microbes remains elusive. Nonculture 16S ribosomal RNA gene sequencing suggests important racial differences and pregnancy specific changes in the vaginal microbial communities. A recent study examining the association of the vaginal microbiome and preterm birth documented important findings but was performed in a predominantly white cohort. Given the important racial differences in bacterial communities within the vagina as well as persistent racial disparities in preterm birth, it is important to examine cohorts with varied demographic compositions.

Objective: To characterize vaginal microbial community characteristics in a large, predominantly African-American, longitudinal cohort of pregnant women and test whether particular vaginal microbial community characteristics are associated with the risk for subsequent preterm birth.

Study design: This is a nested case-control study within a prospective cohort study of women with singleton pregnancies, not on supplemental progesterone, and without cervical cerclage in situ. Serial mid-vaginal swabs were obtained by speculum exam at their routine prenatal visits. Sequencing of the V1V3 region of the 16S rRNA gene was performed on the Roche 454 platform. Alpha diversity community characteristics including richness, Shannon diversity, and evenness as well as beta diversity metrics including Bray Curtis Dissimilarity and specific taxon abundance were compared longitudinally in women who delivered preterm to those who delivered at term.

Results: A total of 77 subjects contributed 149 vaginal swabs longitudinally across pregnancy. Participants were predominantly African-American (69%) and had a preterm birth rate of 31%. In subjects with subsequent term delivery, the vaginal microbiome demonstrated stable community richness and Shannon diversity, whereas subjects with subsequent preterm delivery had significantly decreased vaginal richness, diversity, and evenness during pregnancy (P < .01). This change occurred between the first and second trimesters. Within-subject comparisons across pregnancy showed that preterm birth is associated with increased vaginal microbiome instability compared to term birth. No distinct taxa were associated with preterm birth.

Conclusion: In a predominantly African-American population, a significant decrease of vaginal microbial community richness and diversity is associated with preterm birth. The timing of this suppression appears early in pregnancy, between the first and second trimesters, suggesting that early gestation may be an ecologically important time for events that ordain subsequent term and preterm birth outcomes.

Keywords: pregnancy; preterm birth; vaginal microbiome.

Figure 1. Community Richness, Shannon Diversity, and Pielou Evenness (Genus Level) In All Subjects

Legend: Richness, Shannon Diversity, and Pielou evenness (genus level V1V3) of the full cohort. Blue = term birth Yellow = preterm birth Lines represent regression lines from mixed models Shaded areas represent 95% confidence intervals Term birth shows stable richness (p=0.14) and stable Shannon Diversity (p=0.07). Pielou evenness decreases slightly (p=0.04) over pregnancy. Preterm birth shows significant decrease in richness (p<0.001), Shannon Diversity (p<0.001), and Pielou evenness (p<0.001) over pregnancy. Term versus preterm birth in single trimester comparisons showed no significant difference in pairwise comparisons within a single trimester.

Figure 2. Community Richness, Shannon Diversity, and Pielou Evenness (Genus Level) In African-American Subjects

Legend: Richness, Shannon Diversity, and Pielou evenness (genus level V1V3) of the African American cohort. Blue = term birth Yellow = preterm birth Lines represent regression lines from mixed models Shaded areas represent 95% confidence intervals Term birth shows stable richness (p=0.11), stable Shannon Diversity (p=0.09), and stable Pielou evenness (p=0.08) over pregnancy. Preterm birth shows significant decrease in richness (p<0.001), Shannon Diversity (p=0.003), and Pielou evenness (p<0.001) over pregnancy. Term versus preterm birth in single trimester comparisons showed no significant difference in pairwise comparisons within a single trimester.

Figure 3. Bray Curtis Dissimilarity Within Subjects Across Pregnancy

Legend: Bray Curtis dissimilarity for (A) entire cohort and (B) African American cohort for subjects with paired samples in trimester 1 and 2 and paired samples in trimester 2 and 3. Yellow = preterm birth Blue = term birth. In both the full cohort and the African American cohort preterm birth samples show significantly more dissimilarity to each other in the second and third trimesters than term birth samples suggesting that preterm birth outcomes are associated with a less stable vaginal community.

Figure 4. Heat map of all samples in cohort clustered by term and preterm birth and trimester of pregnancy

Legend: Heat map of all samples from cohort showing all organisms comprising at least 1% of the community. Panel A V1V3 amplification, Panel B V3V5 amplification. Left bar: blue=term birth, yellow= preterm birth. Right bar: light gray=trimester 1, medium gray=trimester 2, black=trimester 3. Histogram: darkest red = highest abundance, shades of red =relatively less abundance, pale yellow=low abundance or not present. This heatmap demonstrates Lactobacillus dominant communities and Lactobacillus poor communities are present in all trimesters of pregnancy and in both term and preterm birth outcomes. In the most Lactobacillus dominant communities there is a paucity of other taxa present whereas in Lactobacillus poor communities there is a higher abundance of other taxa such as Prevotella, Sneathia, Atopobium, Mycoplasma and others. Gardnerella abundance (seen best in panel B) is more common in Lactobacillus poor communities.