Clinical implications of detection of Ureaplasma urealyticum in the amniotic cavity with the polymerase chain reaction

OBJECTIVE: The objective of this study was to determine the frequency and clinical significance of the detection of Ureaplasma urealyticum by means of the polymerase chain reaction with specific primers in the amniotic fluid of patients with preterm premature rupture of membranes. STUDY DESIGN: Amniocentesis was performed in 154 patients with preterm premature rupture of membranes. Amniotic fluid was cultured for aerobic and anaerobic bacteria and for mycoplasmas. Ureaplasma urealyticum was detected by means of the polymerase chain reaction with specific primers. Patients were divided into the following 3 groups according to the results of amniotic fluid culture and polymerase chain reaction for U. urealyticum: those with a negative amniotic fluid culture and a negative polymerase chain reaction (n = 99), those with a negative amniotic fluid culture but a positive polymerase chain reaction (n = 18), and those with a positive amniotic fluid culture regardless of the results of the polymerase chain reaction (n = 37). Contingency table and survival techniques were used for analysis. RESULTS: (1) U. urealyticum was detected by polymerase chain reaction in 28% (43/154) of patients and by culture in 16% (25/154). (2) Among the 43 patients with a positive polymerase chain reaction for U urealyticum, amniotic fluid culture was negative in 42% (18/43). (3) Patients with a negative amniotic fluid culture for U urealyticum but a positive polymerase chain reaction had a significantly shorter median interval from amniocentesis to delivery and a higher amniotic fluid interleukin 6 and white blood cell count than did those with a negative amniotic fluid culture and a negative polymerase chain reaction (interval to delivery; median, 53 hours; range, 0.3-335 hours; vs. median, 141 hours; range, 0.1-3552 hours; P<.05; amniotic fluid white blood cell count: median, 513 cells/mm(3); range, 1-2295 cells/mm(3); vs. median, 1 cell/mm(3); range, 0-7956 cells/mm(3); amniotic fluid interleukin 6: median, 16.6 ng/mL; range, 0.3-53.0 ng/mL; vs. median 0.4 ng/mL; range, 0-69.8 ng/mL; P<.0001 for all). (4) Patients with a positive polymerase chain reaction for U. urealyticum but a negative amniotic fluid culture had a higher rate of significant neonatal morbidity than did those with both a negative culture and a negative polymerase chain reaction (P<.05). (5) No significant differences in perinatal outcome were observed between patients with a negative culture but a positive polymerase chain reaction and those with a positive amniotic fluid culture. CONCLUSION: (1) Culture techniques for mycoplasmas missed 40% of cases of microbial invasion of the amniotic cavity with U. urealyticum. (2) Patients with a positive polymerase chain reaction but a negative amniotic fluid culture are at risk for adverse outcomes. (3) The use of molecular microbiologic techniques is likely to increase the detection of infection among patients with obstetric complications.     

Infection and labor. V. Prevalence, microbiology, and clinical significance of intraamniotic infection in women with preterm labor and intact membranes

Amniotic fluid was retrieved by amniocentesis from 264 patients with preterm labor and intact membranes admitted to Yale-New Haven Hospital from Jan. 1, 1985, to July 31, 1988. The prevalence of a positive amniotic fluid culture was 9.1% (24/264). A total of 111 patients (42%) delivered preterm neonates, and 24 (21.6%) of those had positive amniotic fluid cultures. The diagnostic indexes of the Gram stain of amniotic fluid in the prediction of a positive amniotic fluid culture were as follows: sensitivity, 79.1%; specificity, 99.6%; positive predictive value, 95%; and negative predictive value, 98%. Endotoxin was detected with the limulus amebocyte lysate assay in 4.9% (13/264) of patients with preterm labor. All patients with endotoxin in the amniotic fluid delivered preterm neonates. The three most frequently isolated organisms were Ureaplasma urealyticum (n = 6), Fusobacterium species (n = 5), and Mycoplasma hominis (n = 4). Clinical chorioamnionitis was present in only 12.5% of the patients with positive amniotic fluid cultures. Women with positive amniotic fluid cultures had lower gestational ages and more advanced cervical dilatation on admission than women with negative cultures. Preterm infants born to mothers with positive amniotic fluid cultures had a higher incidence of respiratory distress syndrome and infectious complications than preterm neonates born after negative amniotic fluid cultures. These data underscore the frequency and importance of intraamniotic infections in women with preterm labor.     

Infection and labor. VI. Prevalence, microbiology, and clinical significance of intraamniotic infection in twin gestations with preterm labor

The purpose of this study was to establish the prevalence, microbiology, and outcome of microbial invasion of the amniotic cavity in twin gestation presenting with preterm labor and intact membranes. Amniocenteses were performed on both sacs of 46 women with twin gestations, preterm labor, and intact membranes. Indigo carmine was injected to ensure sampling of both amniotic sacs. Amniotic fluid was cultured for aerobic and anaerobic bacteria, Mycoplasma hominis, and Ureaplasma urealyticum. A positive amniotic fluid culture of at least one sac was noted in 10.8% (5/46) of patients admitted in preterm labor and in 11.9% (5/42) of women delivered of preterm neonates. Of the five patients with microbial invasion of the amniotic cavity, three had microorganisms isolated from both sacs. The presenting sac was involved in all cases, supporting an ascending route for microbial invasion of the amniotic cavity in twin gestation. Polymicrobial infection was found in three of the eight amniotic sacs with positive cultures. In two cases different organisms were isolated from each sac. All patients with positive amniotic fluid cultures were delivered of preterm infants within 48 hours of amniocentesis. Patients with positive amniotic fluid cultures presented with preterm labor at an earlier gestational age and with more advanced cervical dilatation than did women with negative amniotic fluid cultures. Clinical evidence of chorioamnionitis subsequently developed in two of five women with positive amniotic fluid cultures. The interval between amniocentesis and delivery was shorter in women with positive amniotic fluid cultures than in women with negative amniotic fluid cultures (median: 3.5 vs 168 hours, p less than 0.0001). Infants born to women with microbial invasion of the amniotic cavity had a lower median birth weight and a higher incidence of respiratory distress syndrome than those born to women with negative amniotic fluid cultures (birth weight: 1085 vs 1975 gm, p = 0.024; respiratory distress syndrome: 37.5% vs 8.3%, p = 0.04).     

Microbial invasion of the amniotic cavity at birth is associated with adverse short-term outcome of preterm infants

AIMS: To determine the frequency and clinical significance of microbial invasion of the amniotic cavity at the time of delivery in preterm infants. METHODS: Prospective cohort study during June 2001 and January 2002. Preterm infants < 33 + 6 weeks of gestation who had amniotic fluid and placental tissue sampled for culture during cesarean section were included. RESULTS: Of a total of 80 neonates, 42 had negative culture results, 22 had growth of Ureaplasma urealyticum, and 16 had growth of other pathogens. Isolation of Ureaplasma urealyticum was associated with a decreased risk of developing hyaline membrane disease after birth but a more than 20 times increased risk of developing chronic lung disease. Patients with growth of other pathogens had a significantly higher mortality than patients with negative culture results. CONCLUSIONS: Isolation of miroorganisms from the amniotic cavity at birth is associated with an adverse outcome of the preterm infant. In the light of extremely small numbers of positive blood cultures in preterm infants after birth, we consider it reasonable to recommend routine culturing of amniotic cavity tissues/fluid obtained during cesarean section in order to increase the identification rate of pathogens potentially involved in the pathogenesis of perinatal infections.     

Biovar diversity of Ureaplasma urealyticum in amniotic fluid: distribution,intrauterine inflammatory response and pregnancy outcomes

OBJECTIVE: The objective of this study was to determine the distribution of two biovars of Ureaplasma urealyticum (parvo and T960) in human amniotic fluid and to examine whether the magnitude of the intrauterine inflammatory response and pregnancy outcomes are different between patients with microbial invasion of the amniotic cavity with "parvo biovar" and those with "T960 biovar". STUDY DESIGN: This cohort included 77 preterm singleton pregnancies (gestational age < 37 weeks) in whom U. urealyticum was detected from amniotic fluid using the polymerase chain reaction (PCR). Amniotic fluid was obtained by transabdominal amniocentesis. Amniotic fluid was cultured for aerobic and anaerobic bacteria as well as mycoplasmas. U. urealyticum was biotyped by PCR methods. Amniotic fluid inflammatory response was determined by amniotic fluid white blood cell count and interleukin-6 concentration. RESULTS: 1) The "parvo biovar" was detected in 82% (63/77) and "T960 biovar" was in 18% (14/77) of cases; 2) U. urealyticum was isolated by conventional culture method from amniotic fluid in 56% (35/63) of cases with positive for "parvo biovar" and in 50% (7/14) of cases with positive for "T960 biovar"; 3) There were no significant differences in the median gestational age at amniocentesis, gestational age at delivery, birth weight, amniotic fluid white blood cell count, amniotic fluid interleukin-6 concentration and the rates of clinical chorioamnionitis, histologic chorioamnionitis, funisitis and neonatal morbidity between patients in the two biovar groups. CONCLUSIONS: 1) The "parvo biovar" is more frequently isolated from amniotic fluid of preterm gestations than the "T960 biovar"; 2) Biovar diversity of U. urealyticum in amniotic fluid was not associated with different pregnancy outcome and magnitude of the intraamniotic inflammatory response.     

Mycoplasma hominis and Ureaplasma urealyticum in midtrimester amniotic fluid: association with amniotic fluid cytokine levels and pregnancy outcome

OBJECTIVE: The association between the detection of Mycoplasma hominis or Ureaplasma urealyticum in midtrimester amniotic fluid and amniotic fluid cytokine concentrations and subsequent pregnancy outcome were examined. STUDY DESIGN: Amniocentesis was performed between 15 and 19 weeks of gestation in 179 asymptomatic women. Aliquots were assayed for M hominis and U urealyticum by polymerase chain reaction coupled to enzyme-linked immunosorbent assay. Intra-amniotic levels of interleukin-1beta, interleukin-1 receptor antagonist, interleukin-4, interleukin-6, and tumor necrosis factor-alpha were determined by enzyme-linked immunosorbent assay. Pregnancy outcomes were obtained after the completion of all testing. RESULTS: U urealyticum was detected in 22 of 172 amniotic fluids (12.8%); M hominis was present in 11 of 179 amniotic fluids (6.1%). There was no relationship between U urealyticum detection and the concentration of any cytokine. Detection of M hominis was associated with elevated intra-amniotic concentrations of interleukin-4 ( P = .01). Preterm premature rupture of membranes that was followed by preterm birth occurred in 5 women (2.8%); 5 women (2.8%) had a spontaneous preterm birth with intact membranes. All 5 of the women with preterm premature rupture of membranes (100%) tested positive for either U urealyticum or M hominis , as opposed to none of the women with spontaneous preterm birth and to 27 of 161 women (16.8%) with a term birth ( P = .0002). CONCLUSION: The detection of M hominis or U urealyticum in midtrimester amniotic fluid by polymerase chain reaction-enzyme-linked immunosorbent assay may be a risk factor for subsequent preterm premature rupture of membranes.     

Interleukin-6 and interleukin-8 in cervical fluid in a population of Swedish women in preterm labor: relationship to microbial invasion of the amniotic fluid, intra-amniotic inflammation, and preterm delivery

BACKGROUND: Intrauterine infection and inflammation in women with preterm labor are related to adverse perinatal outcome. Due to its subclinical nature, a correct diagnosis depends on retrieval of amniotic fluid. Amniocentesis is, however, not performed as a clinical routine because of its invasiveness. Hypothetically, cytokines in the cervical fluid may represent an alternative diagnostic approach. The aim was to examine cervical interleukin (IL)-6 and IL-8 in relation to microbial invasion of the amniotic fluid, intra-amniotic inflammation, and preterm birth in women in preterm labor. METHODS: Women with singleton pregnancies in preterm labor (<34 weeks of gestation) and intact membranes were included. Cervical (n = 91) and amniotic fluids (n = 56) were collected. Polymerase chain reaction for Ureaplasma urealyticum and Mycoplasma hominis and culture for aerobic and anaerobic bacteria were performed. IL-6 and IL-8 were analyzed with enzyme-linked immunosorbent assay. RESULTS: Non-lactobacillus-dominated biota was detected in cervical secretion in 25% (22/89) and the presence of micro-organisms in the amniotic fluid in 16% (9/56) of the patients. The presence of U. urealyticum in the cervical fluid (21/46) was associated with significantly higher levels of IL-6 in the secretion. IL-6 and IL-8 were significantly higher in cervical fluid of women with intra-amniotic infection and inflammation and in women who delivered < or =7 days and/or before 34 weeks of gestation. Cervical IL-6 > or = 1.7 ng/ml was related to intra-amniotic inflammation (relative risk: 2.67; range: 1.50-4.74) and had a sensitivity, specificity, positive predictive value, and negative predictive value of 58, 83, 75, and 69%, respectively, in the identification of intra-amniotic inflammation. Similar data were obtained for IL-8 > or = 6.7 ng/ml. CONCLUSIONS: High levels of cervical IL-6 and IL-8 are moderately predictive of intrauterine infection/inflammation and preterm delivery.     

Clinical significance of intra-amniotic inflammation in patients with preterm labor and intact membranes.

OBJECTIVE: The purpose of this study was to determine the frequency and clinical significance of intraamniotic inflammation in patients with preterm labor and intact membranes. STUDY DESIGN: Amniocentesis was performed in 206 patients with preterm labor and intact membranes. Amniotic fluid was cultured for aerobic and anaerobic bacteria and mycoplasmas. The diagnosis of intraamniotic inflammation was made in patients with a negative amniotic fluid culture on the basis of amniotic fluid concentrations of interleukin-6 (>2.6 ng/mL, derived from receiver operating characteristic curve analysis). Statistical analysis was conducted with contingency tables and survival techniques. RESULTS: Intra-amniotic inflammation (negative amniotic fluid culture but elevated amniotic fluid interleukin-6) was more common than intra-amniotic infection (positive amniotic fluid culture regardless of amniotic fluid interleukin-6 concentration; 21% [44/206 women] vs 10% [21/206 women]; P <.001). The amniocentesisto-delivery interval was significantly shorter in patients with intra-amniotic inflammation than in patients with a negative culture and without an inflammation (median, 20 hours [range, 0.1-2328 hours] vs median, 701 hours [range, 0.1-3252 hours], respectively; P <.0001). Spontaneous preterm delivery of <37 weeks was more frequent in patients with intra-amniotic inflammation than in those with a negative culture and without inflammation (98% vs 35%; P <.001). Patients with intra-amniotic inflammation had a significantly higher rate of adverse outcome than patients with a negative culture and without intra-amniotic inflammation. Adverse outcomes included clinical and histologic chorioamnionitis, funisitis, early preterm birth, and significant neonatal morbidity. There were no significant differences in the rate of adverse outcomes between patients with a negative culture but with intra-amniotic inflammation and patients with intra-amniotic infection (positive culture regardless of amniotic fluid interleukin-6 concentration). CONCLUSION: Intra-amniotic inflammation/infection complicates one third of the patients with preterm labor (32%; 65/206 women), and its presence is a risk factor for adverse outcome. The outcome of patients with microbiologically proven intra-amniotic infection is similar to that of patients with intra-amniotic inflammation and a negative amniotic fluid culture. We propose that the treatment of patients in preterm labor be based on the operational diagnosis of intra-amniotic inflammation rather than the diagnosis of intra-amniotic infection because the latter diagnosis cannot be undertaken rapidly.     

Amnionitis with Ureaplasma urealyticum or other microbes leads to increased morbidity and prolonged hospitalization in very low birth weight infants

OBJECTIVE: To investigate the influence of culture proven intrauterine infection on preterm morbidity and to test the effect of antimicrobial treatment. METHODS: Retrospective cohort study conducted between October 1997 and February 2001 in patients with preterm premature rupture of membranes or preterm labor. Vaginal swabs were sampled and amniocentesis for microbiologic culture of the amniotic fluid was performed. Patients with Ureaplasma urealyticum in the amniotic fluid were treated with josamycin. Infants were followed post partum according to birth weight, gestational age, APGAR score and infant morbidity. RESULTS: In 49 eligible patients, 40% of cultures were positive, 22% for Ureaplasma urealyticum, 12% for other bacteria and 6% for candida. Children of mothers with positive amniotic fluid cultures had significantly lower gestational ages (26+4 weeks for Ureaplasma urealyticum [p=0.04] and 25+5 weeks for other microorganisms [p=0.0017] versus 28+6 weeks for mothers with negative amniotic fluid cultures) and lower birth weights (975 g [n.s.] and 828 g [p=0.0072] versus 1,041 g) but were appropriate for their gestational ages. 33.3% and 66.7% versus 24% of the children were mechanically ventilated [n.s.], duration of mechanical ventilation was 5.3 [p=0.02] and 10.1 days [p=0.04] versus 1.4 days, and prevalence of chronic lung disease was 38% and 33% versus 11% [n.s.]. Prevalence of severe intraventricular hemorrhage (12.5% [n.s.] and 33% [p=0.04] versus 3.4%) and nosocomial infections (50% for both groups of positive cultures versus 10.3% for negative cultures, p=0.02 and 0.03, respectively) was higher and median length of stay was significantly longer (121 [p=0.02] and 107 days [p=0.03] versus 60 days) in these patients. Maternal positive vaginal swab cultures were not associated with any of the above-mentioned factors. In none of the patients treated with macrolids for proven Ureaplasma urealyticum amnionitis could the microbes be eradicated. CONCLUSION: Maternal positive amniotic fluid cultures have been associated with lower gestational age and lower birth weight. Rate of infant morbidity was higher and length of stay was significantly longer in this group. Positive vaginal swabs were not predictive for infant morbidity. Treatment of mothers showing positive amniotic fluid cultures with macrolids was not effective.     

A sonographic short cervix as the only clinical manifestation of intra-amniotic infection

OBJECTIVE: A sonographically short cervix is a powerful predictor of spontaneous preterm delivery. However, the etiology and optimal management of a patient with a short cervix in the mid-trimester of pregnancy remain uncertain. Microbial invasion of the amniotic cavity (MIAC) and intra-amniotic inflammation are frequently present in patients with spontaneous preterm labor or acute cervical insufficiency. This study was conducted to determine the rate of MIAC and intra-amniotic inflammation in patients with a cervical length < 25 mm in the mid-trimester. STUDY DESIGN: A retrospective cohort study was conducted of patients referred to our high risk clinic because of a sonographic short cervix or a history of a previous preterm birth. Amniocenteses were performed for the evaluation of MIAC and for karyotype analysis in patients with a short cervix. Fluid was cultured for aerobic and anaerobic bacteria, as well as genital mycoplasmas. Patients with MIAC were treated with antibiotics selected by their physician. RESULTS: Of 152 patients with a short cervix at 14-24 weeks, 57 had amniotic fluid analysis. The prevalence of MIAC was 9% (5/57). Among these patients, the rate of preterm delivery (< 32 weeks) was 40% (2/5). Microorganisms isolated from amniotic fluid included Ureaplasma urealyticum (n=4) and Fusobacterium nucleatum (n=1). Patients with a positive culture for Ureaplasma urealyticum received intravenous Azithromycin. Three patients with Ureaplasma urealyticum had a sterile amniotic fluid culture after treatment, and subsequently delivered at term. The patient with Fusobacterium nucleatum developed clinical chorioamnionitis and was induced. CONCLUSION: (1) Sub-clinical MIAC was detected in 9% of patients with a sonographically short cervix (< 25 mm); and (2) maternal parenteral treatment with antibiotics can eradicate MIAC caused by Ureaplasma urealyticum. This was associated with delivery at term in the three patients whose successful treatment was documented by microbiologic studies.     

The frequency and clinical significance of intra-amniotic inflammation in patients with a positive cervical fetal fibronectin.

OBJECTIVE: A positive fetal fibronectin result in cervicovaginal fluid is a powerful predictor of preterm delivery and is considered a marker for upper genital tract infection (ie, intrauterine infection). Treatment with antimicrobial agents is being considered in patients with a positive fetal fibronectin test of cervico/vaginal fluid. This study was undertaken to determine the frequency and clinical significance of intra-amniotic infection/inflammation in patients with a positive fetal fibronectin. STUDY DESIGN: A total of 1709 pregnant women (gestational age, 23-31 weeks) were screened for cervical fetal fibronectin. Patients with a positive fibronectin were offered amniocentesis for the diagnosis of intra-amniotic infection and treatment with antibiotics. Amniocentesis was performed in 58 patients with a positive fibronectin test (>50 ng/mL). Amniotic fluid was cultured for aerobic/anaerobic bacteria and mycoplasmas. Polymerase chain reaction assay for Ureaplasma urealyticum was performed. Interleukin-6 concentrations were measured by a specific immunoassay. Nonparametric statistics were used for analysis. RESULTS: None of the patients with a positive fibronectin had a positive amniotic fluid culture. U urealyticum was detected in 1 case (1.8%) with the polymerase chain reaction assay. Amniotic fluid IL-6 was elevated (>2.5 ng/mL) in 5.3% of patients (3/57 patients); all of these patients delivered preterm neonates. There was no relationship between amniotic fluid IL-6 and cervical fibronectin concentration (r = 0.14;P: >.1). Patients who delivered preterm (<34 weeks) had higher median amniotic fluid IL-6 and cervical fetal fibronectin concentrations than those patients who delivered after 34 weeks (IL-6: median, 2.1 ng/mL [range, 0.1-25.3 ng/mL] vs median, 0.3 ng/mL [0.03-2.4 ng/mL]; P <.05; fibronectin: median, 509 ng/mL [260->1000 ng/mL] vs median, 155 ng/mL [50-889 ng/mL]; P <.01). CONCLUSION: Intra-amniotic infection was detected in 1.8% of cases with a positive fibronectin in the cervical fluid; intra-amniotic inflammation was present in 5.3% of cases. All patients with a positive fetal fibronectin and intra-amniotic inflammation delivered preterm neonates.     

Tumor necrosis factor in preterm and term labor.

OBJECTIVE: Our objective was to determine if labor (term and preterm) and microbial invasion of the amniotic cavity were associated with changes in amniotic fluid concentrations of tumor necrosis factor. STUDY DESIGN: Amniotic fluid was retrieved by transabdominal amniocentesis from 269 women in the following groups: midtrimester (n = 38), preterm labor with intact membranes (n = 52), preterm premature rupture of membranes (n = 74), term in active labor (n = 84), and term not in labor (n = 21). Fluid was cultured for aerobic and anaerobic bacteria and for Mycoplasma species. Tumor necrosis factor was measured with a commercially available enzyme-linked immunosorbent assay validated for amniotic fluid (sensitivity 60 pg/ml). RESULTS: Amniotic fluid from pregnant women in the second and third trimesters who were not in labor did not contain tumor necrosis factor. Among women in preterm labor, 92.3% (12/13) of patients with a positive amniotic fluid culture had detectable tumor necrosis factor in the amniotic fluid (median 820 pg/ml, range less than 60 to 2340 pg/ml). In contrast, only 10.2% (4/39) of women with a negative amniotic fluid culture had detectable tumor necrosis factor. Histopathologic chorioamnionitis was found in all patients who had a positive amniotic fluid culture, and tumor necrosis factor was detectable in the amniotic fluid of all but one of these patients. Among women in active labor at term, 25% (21/84) had detectable tumor necrosis factor in the amniotic fluid. Tumor necrosis factor was detected more frequently in the amniotic fluid of patients with a positive amniotic fluid culture than in patients with a negative culture (46.6% [7/15] vs 20.2% [14/69], p = 0.047). Amniotic fluid concentrations of tumor necrosis factor were significantly higher in patients with preterm premature rupture of membranes, labor, and a positive amniotic fluid culture than in the other subgroups of patients with preterm premature rupture of membranes. CONCLUSION: Parturition in the setting of microbial invasion of the amniotic cavity is associated with activation of the cytokine network as demonstrated by the detection of tumor necrosis factor in human amniotic fluid.     

Performance characteristics of putative tests for subclinical chorioamnionitis

OBJECTIVE: To evaluate amniotic fluid glucose, matrix metalloproteinase (MMP)-9, interleukin (IL)-6, and IL-12 for diagnosing subclinical chorioamnionitis in women with preterm labor. METHODS: Forty-four women in preterm labor at 22-35 weeks gestation with suspected subclinical chorioamnionitis underwent amniocentesis. Amniotic fluid analysis included Gram stain, culture, and determination of glucose, MMP-9, IL-6, and IL-12 concentrations. Median values of these analytes were compared using the Mann-Whitney U test. Sensitivity, specificity, and positive and negative predictive values were calculated for tests using a positive amniotic fluid culture or delivery within 24 hours as the key outcome variables. RESULTS: Amniotic fluid concentrations of glucose, MMP-9, and IL-6 correlated closely with positive culture or delivery within 24 hours. IL- 12 concentrations did not correlate with either a positive culture or delivery within 24 hours. CONCLUSIONS: Amniotic fluid glucose, MMP-9, and IL-6 reliably predict microbial invasion of the amniotic cavity or imminent delivery. IL- 12 values did not correlate with amniotic fluid culture results or imminent delivery.     

IL-8 concentrations in maternal serum, amniotic fluid and cord blood in relation to different pathogens within the amniotic cavity

OBJECTIVE: The association between elevated interleukin (IL)-8 concentrations in amniotic fluid and preterm delivery is well described. Little consideration has been given to the impact of different groups of microorganisms within the amniotic cavity on IL-8 concentration. METHODS: We collected amniotic fluid, placental tissue and amniotic membranes during preterm cesarean sections for bacterial culture. In addition, we determined IL-8 concentrations in maternal serum, amniotic fluid and cord blood and correlated them with the various intra-amniotic pathogens isolated by bacterial culture. RESULTS: IL-8 concentrations were determined in amniotic fluid in 107 cases, in cord blood in 185 cases and in maternal blood in 158 cases. Women with intra-amniotic Ureaplasma urealyticum infection had significantly higher amniotic fluid concentrations of IL-8 than those without (P< 0.001). In cord blood, we found significantly elevated IL-8 concentrations due to intra-amniotic infection with U. urealyticum (P=0.045) and other pathogens (P=0.04). In maternal sera, we found no significant elevation of maternal IL-8 in any of the groups. CONCLUSION: Intrauterine infection with U. urealyticum seems to play a profound role in the cascade of inflammation and increases IL-8 concentrations in amniotic fluid and cord blood.     

Interleukin 6 determinations in cervical fluid have diagnostic and prognostic value in preterm premature rupture of membranes

OBJECTIVE: Our aim was to determine whether interleukin-6 concentrations in cervical fluid samples are of value in the identification of microbial invasion of the amniotic cavity, prediction of the duration of the latency period, and assessment of the risk of neonatal complications in preterm premature rupture of membranes. STUDY DESIGN: A cohort study was performed in 86 patients with preterm premature rupture of membranes. Amniotic fluid and cervical fluid were collected. Amniotic fluid was cultured for aerobic and anaerobic bacteria, as well as mycoplasmas. Interleukin 6 was measured by a sensitive and specific immunoassay. The receiver operating characteristic curve, logistic regression, and survival techniques were used for analysis. RESULTS: (1) Patients with a positive amniotic fluid culture had a significantly higher median cervical fluid interleukin 6 concentration than those with negative results (median, 528 pg/mL; range, 174-825 pg/mL; vs median, 169 pg/mL; range, 8-986 pg/mL; P <.0001). (2) A cervical fluid interleukin 6 concentration of >350 pg/mL had a sensitivity of 92% and a specificity of 78% in the identification of a positive amniotic fluid culture. (3) Patients with a cervical fluid interleukin 6 concentration of >350 pg/mL had a significantly shorter median interval to delivery and higher rate of funisitis, preterm delivery within 2 days and 7 days, and the occurrence of significant neonatal morbidity than did those with a cervical fluid interleukin 6 concentration of <350 pg/mL (P <.05 for each). (4) The increased perinatal morbidity remained significant after adjustment for gestational age (P <.05). (5) There was a strong correlation between cervical fluid concentrations and amniotic fluid concentrations of interleukin 6 (P <.001). CONCLUSION: Cervical fluid interleukin 6 determinations are of value in the assessment of the likelihood of microbial invasion of the amniotic cavity, impending preterm delivery, and the occurrence of significant neonatal complications in the setting of preterm premature rupture of membranes.     

Microbial invasion and cytokine response in amniotic fluid in a Swedish population of women in preterm labor

BACKGROUND: Previous studies indicate an association between intra-amniotic microbial invasion and/or inflammation and spontaneous preterm birth, but there is a limited amount of data available from Europe. The aim of this study was to investigate the occurrence of intra-amniotic microorganisms and cytokines (interleukin-6 and interleukin-8) in a Swedish population of women in preterm labor and their correlation with preterm birth. METHODS: Amniotic fluid was retrieved transabdominally from 61 patients in preterm labor before 34 weeks of gestation. Polymerase chain reaction analyses for Ureaplasma urealyticum and Mycoplasma hominis and culture for aerobic and anaerobic bacteria were performed. Interleukin-6 and interleukin-8 were analyzed with enzyme-linked immunosorbent assay. RESULTS: Microorganisms in amniotic fluid were detected in 10 patients (16%). Patients with detected bacteria in the amniotic fluid had significantly higher levels of interleukin-6 and interleukin-8. There was also an association between interleukin-6/-8, the amniocentesis-delivery interval (or= 1.5 ng/mL or interleukin-8 >or= 1.3 ng/mL was associated with an increased risk of delivery within 7 days (interleukin-6: relative risk 7.3; 95% confidence interval: 2.8-19; sensitivity 83%, specificity 87%; interleukin-8: relative risk 14, 95% confidence interval: 3.6-55, sensitivity 91%, specificity 87%). CONCLUSIONS: The occurrence of intra-amniotic microbial invasion and inflammation in this population of Swedish women in preterm labor was similar to data reported from populations with a higher incidence of preterm delivery. Amniotic interleukin-6 and interleukin-8 correlated with the presence of microorganisms and with preterm birth.     

Human beta-defensin-2: a natural antimicrobial peptide present in amniotic fluid participates in the host response to microbial invasion of the amniotic cavity.

OBJECTIVE: Human beta-defensin-2 (HBD-2) is a potent antimicrobial peptide that is part of the innate immune response. The purpose of this study was to determine whether HBD-2 is present in amniotic fluid and if its concentration changes with microbial invasion of the amniotic cavity (MIAC) and labor. STUDY DESIGN: Amniotic fluid was retrieved by amniocentesis from 318 patients in the following groups: (1) mid-trimester (n=75); (2) term not in labor (n=28) and in labor (n=51); (3) preterm labor and intact membranes without MIAC who delivered at term (n=36), who delivered preterm without MIAC (n=52), and preterm labor with MIAC who delivered preterm (n=25); and (4) preterm premature rupture of membranes (preterm PROM) with (n=25) and without MIAC (n=26). MIAC was defined as a positive amniotic fluid culture for microorganisms. Amniotic fluid HBD-2 concentrations were determined using a sensitive and specific ELISA. Non-parametric statistics were used for analysis. RESULTS: (1) HBD-2 was detected in all amniotic fluid samples; (2) the concentration of HBD-2 did not change with gestational age from mid-trimester to term (p=0.8); (3) intra-amniotic infection was associated with a significant increase in amniotic fluid concentrations of HBD-2 in both women with preterm labor and intact membranes, and women with preterm PROM (p<0.05 for each comparison); (4) patients with preterm labor and a negative amniotic fluid culture who delivered preterm had a higher median amniotic fluid HBD-2 concentration than those with preterm labor who delivered at term (p=0.001); and (5) among patients with preterm labor without MIAC, those who had intra-amniotic inflammation (amniotic fluid white blood cell count>100 cells per mL) had a higher median amniotic fluid concentration of HBD-2 than those without this condition (p<0.002). CONCLUSION: (1) Amniotic fluid contains HBD-2, a natural antimicrobial peptide, and this may account for some of the antimicrobial activity of amniotic fluid; (2) amniotic fluid HBD-2 concentrations are increased in women with MIAC, regardless of the membrane status (intact membranes or PROM); and (3) we propose that amniotic fluid HBD-2 is part of the innate immune system within the amniotic cavity.     

Evidence for the participation of interstitial collagenase (matrix metalloproteinase 1) in preterm premature rupture of membranes

OBJECTIVE: Rupture of membranes is thought to result from the effects of physical forces in localized areas of the membranes weakened by the degradation of structural collagens. Matrix metalloproteinases are enzymes that degrade extracellular matrix components and have been implicated in membrane rupture. The objective of this study was to determine whether spontaneous rupture of membranes is associated with a change in the amniotic fluid concentration of interstitial collagenase (matrix metalloproteinase 1 [MMP-1]), a major collagenase. STUDY DESIGN: A cross-sectional study was conducted to determine MMP-1 concentrations in amniotic fluid from 353 women in the following categories: (1) term with intact membranes not in labor and in labor, (2) preterm labor who delivered at term, (3) preterm labor who delivered preterm without microbial invasion of the amniotic cavity, (4) preterm labor who delivered preterm with microbial invasion of the amniotic cavity, (5) preterm premature rupture of membranes with and without microbial invasion of the amniotic cavity, (6) term premature rupture of membranes not in labor and in labor, and (7) mid trimester of pregnancy. Microbial invasion of the amniotic cavity was determined by an amniotic fluid culture positive for microorganisms. MMP-1 concentrations in amniotic fluid were determined by means of sensitive and specific immunoassays. RESULTS: (1) MMP-1 was detectable in 81.3% of amniotic fluid samples (287/353), and its concentrations increased with advancing gestational age (r = 0.4; P <.001). (2) Preterm premature rupture of membranes was associated with a significant increase in the median amniotic fluid concentration of MMP-1 (P =.02). (3) Women with term premature rupture of membranes had a significantly lower amniotic fluid MMP-1 concentration than those with intact membranes at term not in labor (P <.001). (4) Microbial invasion of the amniotic cavity in patients in preterm labor with intact membranes and in patients with preterm premature rupture of membranes was also associated with significant increases in the median amniotic fluid MMP-1 concentrations (P <.05 and P <.01, respectively). (5) Patients with preterm premature rupture of membranes and microbial invasion of the amniotic cavity had a significantly higher median amniotic fluid MMP-1 concentration than those with intact membranes and microbial invasion of the amniotic cavity (P =.01). (6) Neither term nor preterm parturition was associated with changes in amniotic fluid MMP-1 concentrations (P =.6 and P =.3, respectively). CONCLUSION: (1) Collagenase 1 (MMP-1) is a physiologic constituent of amniotic fluid. (2) Preterm premature rupture of membranes (in both the presence and absence of infection) was associated with an increase in the amniotic fluid MMP-1 concentrations. (3) Neither term nor preterm parturition was associated with a significant increase in the amniotic fluid concentration of MMP-1.     

Idiopathic vaginal bleeding during pregnancy as the only clinical manifestation of intrauterine infection.

OBJECTIVE: To determine the frequency and clinical significance of microbial invasion of the amniotic cavity (MIAC) in patients with vaginal bleeding in the absence of placenta previa, preterm labor or preterm premature rupture of membranes (PROM). STUDY DESIGN: This retrospective cohort study included patients who presented with vaginal bleeding between 18 and 35 weeks, and underwent an amniocentesis shortly after admission for the assessment of the microbiologic status of the amniotic cavity and/or fetal lung maturity. Amniotic fluid was cultured for aerobic and anaerobic bacteria, as well as genital mycoplasmas. Patients presenting with preterm labor, preterm PROM, placenta previa, overt placental abruption, and an intrauterine device in situ were excluded, as well as those with local cervical bleeding. MIAC was defined as a positive amniotic fluid culture. Analysis was conducted with non-parametric statistics. RESULTS: One hundred and fourteen patients met the entry criteria. MIAC was detected in 14% of cases (16/114). Patients with vaginal bleeding and a gestational age < 28 weeks at the time of amniocentesis had a significantly higher frequency of MIAC than those with a gestational age 28 weeks [25% (13/52) vs. 4.8% (3/62), respectively; p < 0.01]. Ureaplasma urealyticum was the microorganism most frequently isolated from the amniotic fluid. Except for one case admitted at 33 weeks, all patients with MIAC had an early preterm delivery 32 weeks. Patients with vaginal bleeding and MIAC had a shorter procedure-to-delivery interval than those without MIAC [MIAC, median survival 19 days (95% CI 10-27 days) vs. no MIAC, median survival 50 days (95% CI 37-62 days); p < 0.0001]. Patients with vaginal bleeding and MIAC had a significantly lower gestational age at delivery and lower birth weight than those with vaginal bleeding and negative amniotic fluid cultures (for gestational age, median 25 weeks, range 21-33 weeks vs. median 37 weeks, range 19-42 weeks, respectively; p < 0.01, and for birth weight, median 750 grams, range 520-1820 grams vs. 2800 grams, range 520-4880 grams, respectively; p < 0.01), as well as a higher frequency of subsequent preterm PROM [81.3% (13/16) vs. 9.2% (9/98); p < 0.01]. CONCLUSIONS: MIAC was detected in 14% of patients with 'idiopathic' vaginal bleeding and was associated with subsequent preterm PROM and early preterm delivery. Vaginal bleeding may be the only clinical manifestation of MIAC, and it predisposes to adverse outcome.     

Amniotic fluid glucose concentration: a marker for infection in preterm labor and preterm premature rupture of membranes

Amniotic fluid Gram stain and culture have been utilized as laboratory tests of microbial invasion of the amniotic cavity. The Gram stain of amniotic fluid has a low sensitivity in the detection of clinical infection or microbial invasion of the amniotic cavity, and amniotic fluid culture results are not immediately available for management decisions. Glucose concentration is used to diagnose infection in other sites such as cerebrospinal fluid.Objective: The purpose of this study was to evaluate the usefulness of amniotic fluid glucose concentration in detecting microbial invasion of the amniotic cavity associated with preterm labor and preterm premature rupture of membranes.Methods: Amniocentesis was performed in 60 women with preterm labor and/or preterm premature rupture of membranes. Gram stain and culture for Mycoplasma hominis, Ureaplasma urealyticum, aerobic, and anaerobic bacteria were performed. Subjects were studied prospectively for the development of positive amniotic fluid cultures and the development of clinical chorioamnionitis.Results: The diagnosis of clinical chorioamnionitis was made in 25% (15/60) of women entered into the study. Low amniotic fluid glucose concentration Was considered < 15 mg/dl. The sensitivity, specificity, and positive predictive value of low amniotic, fluid glucose concentration to predict clinical chorioamnionitis were 73.3%, 88.1%, and 68.8% respectively, while positive amniotic fluid culture, hada sensitivity of 43.8%, specificity of 79.5%, and positive predictive value of 43.8%.Conclusions: Amniotic fluid glucose concentration was more sensitive in predicting chorioamnionitis than either Gram stain or culture. Amniotic fluid glucose concentration was better in predicting clinical chorioamnionitis than predicting positive amniotic fluid culture results. Gestational age-dependent normal ranges and pathologic conditions that may alter amniotic fluid glucose concentrations should be considered when interpreting amniotic fluid glucose values to diagnose microbial invasion of the amniotic cavity.