Detection of ureaplasmas by the polymerase chain reaction in the amniotic fluid of patients with cervical insufficiency

Abstract

Aims: The purpose of this study was to determine the clinical significance of detecting microbial footprints of ureaplasmas in amniotic fluid (AF) using specific primers for the polymerase chain reaction (PCR) in patients presenting with cervical insufficiency.

Methods: Amniocentesis was performed in 58 patients with acute cervical insufficiency (cervical dilatation, > or =1.5 cm) and intact membranes, and without regular contractions (gestational age, 16-29 weeks). AF was cultured for aerobic and anaerobic bacteria as well as genital mycoplasmas. Ureaplasmas (Ureaplasma urealyticum and Ureaplasma parvum) were detected by PCR using specific primers. Patients were divided into three groups according to the results of AF culture and PCR for ureaplasmas: those with a negative AF culture and a negative PCR (n=44), those with a negative AF culture and a positive PCR (n=10), and those with a positive AF culture regardless of PCR result (n=4).

Results: 1) Ureaplasmas were detected by PCR in 19.0% (11/58) of patients, by culture in 5.2% (3/58), and by culture and/or PCR in 22.4% (13/58); 2) Among the 11 patients with a positive PCR for ureaplasmas, the AF culture was negative in 91% (10/11); 3) Patients with a negative AF culture and a positive PCR for ureaplasmas had a significantly higher median AF matrix metalloproteinase-8 (MMP-8) concentration and white blood cell (WBC) count than those with a negative AF culture and a negative PCR (P<0.001 and P<0.05, respectively); 4) Patients with a positive PCR for ureaplasmas but a negative AF culture had a higher rate of spontaneous preterm birth within two weeks of amniocentesis than those with a negative AF culture and a negative PCR (P<0.05 after adjusting for gestational age at amnio-centesis); 5) Of the patients who delivered within two weeks of amniocentesis, those with a positive PCR for ureaplasmas and a negative AF culture had higher rates of histologic amnionitis and funisitis than those with a negative AF culture and a negative PCR (P<0.05 after adjusting for gestational age at amniocentesis, for each); 6) However, no significant differences in the intensity of the intra-amniotic inflammatory response and perinatal outcome were found between patients with a positive AF culture and those with a negative AF culture and a positive PCR.

Conclusions: 1) Cultivation techniques for ureaplasmas did not detect most cases of intra-amniotic infection caused by these microorganisms (91% of cases with cervical insufficiency and microbial footprints for ureaplasmas in the amniotic cavity had a negative AF culture); 2) Patients with a negative AF culture and a positive PCR assay were at risk for intra-amniotic and fetal inflammation as well as spontaneous preterm birth.

Figure 1

Amniotic fluid (AF) matrix metalloproteinase-8 (MMP-8) concentrations and white blood cell (WBC) counts according to the results of AF culture and polymerase chain reaction (PCR) assay for ureaplasmas. (A) AF MMP-8 concentrations (group 1: median, 51.4 ng/mL [range, 0.5–1801.4 ng/mL]; group 2: median, 178.4 ng/mL [range, 45.7–6142.6 ng/mL]; group 3: median, 547.3 ng/mL [range, 129.1–770.6 ng/mL]; P<0.01 by Kruskal-Wallis analysis of variance test) and (B) AF WBC counts (group 1: median, 3 cells/mm³ [range, 0–>1000 cells/mm³]; group 2: median, 501 cells/mm³ [range, 2–>1000 cells/mm³]; group 3: median, 775 cells/mm³ [range, 62–>1000 cells/mm³]; P <0.001 by Kruskal-Wallis analysis of variance test).