Introduction: Changes in the vaginal microecology of commensal and pathogenic organisms may influence the initiation of preterm birth (PTB),¹ and can be detected in the cervicovaginal fluid (CVF) metabolite profile. We characterized and validated the metabolite profile of CVF of a cohort of pregnant women presenting with symptoms of preterm labor by both ¹H Magnetic Resonance spectroscopy (MRS) and spectrophotometric assay, and determined their predictive capacity for PTB.

Methods: CVF was obtained by high-vaginal swabs from 86 pregnant women presenting with symptoms of preterm labor (i.e. regular uterine contractions and < 3 cm dilated cervix), but with intact membranes and no evidence of genital infection, between 24 and 36 weeks (wks) gestation. Vaginal secretions were dissolved in 600 µl sterile phosphate buffered saline and a 400 µl MR sample containing 20 µl D₂O was scanned with a 9.4T Bruker Avance III MR spectrometer (Bruker BioSpin GmbH, Karlsruhe, Germany), with 5mm BBO probe using a Watergate water suppression pulse sequence (NS = 256, D1 = 5s, AQ = 1s, SW = 20.6 ppm, TD = 16446). Acetate and other metabolites were identified in the MR spectrum, integrated for peak area and normalized to the total spectrum integral (excluding water signal). Acetate concentrations (AceConc) were also determined from a randomly selected subset of the same samples (n = 57), by spectrophotometric absorption of NADH using acetic acid assay kit (K-ACETGK 08/14, Megazyme, IE).

Results: Acetate normalized integral (P = 0.002) and AceConc (P = 0.006) were significantly higher in the women who delivered preterm compared to their term counterparts (Fig. 1). The MR derived acetate integrals were strongly correlated with the AceConc estimated from the spectrophotometric technique (r = 0.69; P < 0.0001) (Fig. 2). Both methods were predictive of PTB < 37 wks (acetate integral: AUC = 0.75, LR+ = 4.0, LR- = 0.3; AceConc: AUC = 0.74, LR+ = 2.5, LR- = 0.4) (Fig. 3). Additionally, acetate 1H integrals and AceConc were predictive of delivery within 2 weeks of the index assessment (acetate integral: AUC = 0.77, LR+ = 2.0, LR- = 0.5; AceConc: AUC = 0.68, LR+ = 3.1, LR- = 0.3) with an optimal cut-off of > 0.54 g/l (Fig. 4). Normalized integrals of succinate, formate, lactate, glucose, glutamine/glutamate, alanine and branched chain amino acids did not differ between term- and preterm-delivered cohorts.

Conclusion: Elevated CVF acetate in women with symptoms of preterm labor appears predictive of preterm delivery, as well as delivery within 2 weeks of presentation. In these women, a clinical assay of acetate in CVF may prove of clinical utility for predicting PTB.