Characterisation of Mid-Gestation Amniotic Fluid Cytokine and Bacterial DNA Profiles in Relation to Pregnancy Outcome in a Small Australian Cohort

Abstract

A well-established association exists between intrauterine bacteria and preterm birth. This study aimed to explore this further through documenting bacterial and cytokine profiles in Australian mid-gestation amniotic fluid samples from preterm and term births. Samples were collected during amniocenteses. DNA was extracted and the full-length 16S rRNA gene was amplified and sequenced. Levels of the cytokines IL-1β, IL-6, IL-10, TNF-α and MCP-1 were determined using the Milliplex MAGPIX system. Bacterial DNA profiles were low in diversity and richness, with no significant differences observed between term and preterm samples. No differences in the relative abundance of individual OTUs between samples were identified. IL-1β and TNF-α levels were significantly higher in samples containing reads mapping to Sphingomonas sp.; however, this result should be interpreted with caution as similar reads were also identified in extraction controls. IL-6 levels were significantly increased in samples with reads that mapped to Pelomonas sp., whilst TNF-α levels were elevated in fluid samples from pregnancies that subsequently delivered preterm. Bacterial DNA unlikely to have originated from extraction controls was identified in 20/31 (64.5%) mid-gestation amniotic fluid samples. Bacterial DNA profiles, however, were not predictive of preterm birth, and although cytokine levels were elevated in the presence of certain genera, the biological relevance of this remains unknown.

Keywords: PCR (polymerase chain reaction); amniotic fluid; bacteria; cytokines; inflammation; microbiome; preterm birth.

Figure 1

TNF-α concentrations (pg/mL) in mid-trimester amniotic fluid samples collected from women who went on to deliver at term (light blue boxes) or preterm (dark blue boxes).

Figure 2

Shannon diversity and richness (number of OTUs) in mid-trimester amniotic fluid samples collected from women who went on to deliver at term (light blue boxes) or preterm (dark blue boxes).

Figure 3

Relative abundances (%) of taxa which made up >0.5% of amniotic fluid bacterial DNA profiles in term (T) and preterm (PT) samples.

Figure 4

Relative abundances (%) of taxa which made up >0.5% of amniotic fluid bacterial DNA profiles in term (T) and preterm (PT) samples and were not present in extraction controls in high numbers.