Exploratory and confirmatory analysis to investigate the presence of vaginal metabolome expression of microbial invasion of the amniotic cavity in women with preterm labor using high-performance liquid chromatography

Background: While the influence of microbial invasion of the amniotic cavity on the development of spontaneous preterm delivery is unquestionable, the use of an invasive procedure to diagnosis the status of infection limits its clinical translation.

Objective: The aim of the study was to use exploratory and confirmatory analysis to investigate the presence of vaginal metabolome expression of microbial invasion of the amniotic cavity in women diagnosed with preterm labor using high-performance liquid chromatography.

Study design: In a total of 140 women with singleton pregnancies and a diagnosis of preterm labor below 34 weeks of gestation, we analyzed vaginal amino acid concentrations using high-performance liquid chromatography. Vaginal samples were collected shortly after the amniocentesis performed at admission to rule out microbial invasion of the amniotic cavity. Data was normalized for the median of all the amino acid concentrations evaluated. Microbial invasion of the amniotic cavity was defined as a positive aerobic/anaerobic amniotic fluid culture for bacteria or yeast or the presence of Ureaplasma spp. or Mycoplasma hominis in mycoplasma culture or a positive polymerase chain reaction result for 16S rRNA gene sequence. Exploratory analysis was performed in half the sample and confirmatory analysis in the other half. We compared vaginal amino acid concentrations between women with and without microbial invasion of the amniotic cavity in both cohorts. The area under curve (AUC) with the 95% confidence interval was calculated for vaginal amino acids with significant differences.

Results: In the exploratory cohort (2014-2015), 17/76 (22.3%) women had microbial invasion of the amniotic cavity compared to 14/72 (19.4%) in the confirmatory cohort (2016-2017). In the exploratory cohort, we found significantly higher amino acid concentrations of vaginal taurine, lysine and cysteine and significantly lower concentrations of vaginal glutamate, aspartate and the aspartate/asparagine ratio. These significant differences were confirmed in the confirmatory cohort. The AUC of these vaginal amino acids to predict microbial invasion of the amniotic cavity ranged between 0.72 and 0.79, being cysteine the amino acid with the best performance with an AUC of 0.79 (95% CI 0.71-0.88).

Conclusion: We found vaginal metabolome expression of microbial invasion of the amniotic cavity in women with preterm labor and intact membranes. These findings might open the possibility to develop non-invasive diagnostic tools of microbial invasion of the amniotic cavity with the aim of selecting women who would most likely benefit from an amniocentesis for this indication.

Keywords: Preterm labor; high-performance liquid chromatography; metabolomics; microbial invasion of amniotic cavity; minimally invasive diagnoses; vaginal fluid.