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, ≥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 amniocentesis); 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.

Mid-trimester preterm premature rupture of membranes (PPROM): etiology, diagnosis, classification, international recommendations of treatment options and outcome

“…However, recent publications have demonstrated that bacterial culture had a lower rate of detection (27.1%) compared to PCR (up to 72.9%) [35,40]. The use of amniotic fluid culture alone could miss up to 91% of genital mycoplasma [41]. PCR methodology offers a number of advantages over bacterial culture techniques: species-and serovar-specific identification even in the absence of viable organisms (positivity even after antibiotic treatment), higher sensitivity and quicker results (within 24 h).…”

“…After the occurrence of the PPROM in the second trimester, bacteria rapidly colonize the surfaces of amniotic membrane, chorion, decidua, fetal skin and mucosa, as well as the umbilical cord (Figure 3). The PCRbased assays for bacterial presence in the amniotic fluid have a superior sensitivity compared to standard culture methods and the positive predictive value for predicting the incidence of neonatal complications of 60% vs. 35% for the standard test [41,84,85]. Walker et al [86] found that only 33% of infants remained undelivered after one week following a PPROM history.…”

Occurrence of Septic Shock in a Patient Submitted to Emergency Cerclage following a Negative Amniocentesis: Report of a Case

“…However, the risk of adverse pregnancy outcome after emergency cerclage seems to be increased also among those with no detectable amniotic fluid infection but with a latent intra-amniotic inflammatory process. Several markers of intra-amniotic inflammation have been shown to predict early cerclage failure and impending delivery including IL-6, metalloproteinase-8, glucose concentration, CRP, white blood cell, LDH and neutrophil counts [4,5,8,10,11]. …”

“…The patient agreed to undergo a preliminary amniocentesis in order to exclude an actual amniotic infection and to proceed with the emergency cerclage only if negative results were obtained. Then, we retrieved 10 ml of clear amniotic fluid and obtained the following results within 12 h: normal glucose level (19 mg/dl), interleukin-6 (IL-6; 8.68 ng/ml), CRP (<0.03 mg/dl; normal values <0.6 mg/dl [8]; the CRP level was measured with a latex-enhanced turbidimetric immunoassay). A sample was additionally sent for culture (bacteria and Mycoplasma ) to the Microbiology Department.…”

The Effect of Inflammation on Preterm Birth

“…Various findings suggest that in some pregnancies, the infective organism may be present in placental tissues from mid-gestation. In fact, it has been suggested that intrauterine infections may originate pre-conceptually, possibly arising from organisms residing in the endometrium prior to implantation which subsequently activate the maternal innate immune system as a result of a change in growth or immune surveillance (Oh et al, 2010). It is possible that an altered cytokine expression can cause per se preterm labor, namely an overproduction of cytokines can be harmful to the fetus because of the activation of the cytokine flow (Vrachnis et al, 2010).…”