Detection Of Intra-amniotic Infection Research Articles

Nicotinamide phosphoribosyltransferase and intra-amniotic inflammation in preterm prelabor rupture of membranes

Introduction: The amniotic fluid nicotinamide phosphoribosyltransferase (NAMPT) levels have not been compared among women with preterm prelabor rupture of membranes (PPROM) comorbid with intra-amniotic infection, sterile intra-amniotic inflammation (IAI), colonization, or without IAI and microbial invasion of the amniotic cavity (MIAC). Therefore, the main aim was to quantify the amniotic fluid NAMPT in women with PPROM complicated by intra-amniotic infection, sterile IAI, or colonization. The second aim was to characterize the diagnostic indices of NAMPT to reveal IAI. The third aim was to determine whether the cervical fluid and maternal serum NAMPT quantitation might be of value in the identification of intra-amniotic inflammatory complications in PPROM.Methods of study: NAMPT levels in amniotic fluid, cervical fluid, and maternal serum were assessed in three independent cohorts of women with singleton pregnancies complicated by PPROM between 24+0 and 36+6 weeks of gestation consisting of 88, 121, and 88 women, respectively. Amniotic fluid samples were obtained by transabdominal amniocentesis, cervical fluid samples were obtained using a Dacron polyester swab and maternal blood was obtained by venipuncture of the cubital vein. The NAMPT levels were measured by an enzyme-linked immunosorbent assay. Testing for MIAC and IAI was performed on all women, who were then categorized into four subgroups: intra-amniotic infection (MIAC and IAI), sterile IAI (IAI alone), colonization (MIAC alone), and without MIAC and IAI.Results: Women with intra-amniotic infection and women with sterile IAI had higher NAMPT levels than did women with colonization and women without MIAC and IAI (intra-amniotic infection: median 73.6 ng/mL, sterile IAI: median 55.5 ng/mL, colonization: median 12.1 ng/mL, without MIAC and IAI: 10.6 ng/mL; p < .0001). An amniotic fluid NAMPT level of 37 ng/mL was the best value for the detection of intra-amniotic infection in women with PPROM. Cervical fluid (p = .51) and maternal serum (p = .50) NAMPT levels did not reflect intra-amniotic inflammatory complications in women with PPROM.Conclusions: Intra-amniotic infection and sterile IAI are associated with higher NAMPT levels in amniotic fluid but not in cervical fluid or maternal serum in women with PPROM. Amniotic fluid NAMPT might be a marker for invasive identification of IAI in PPROM.

The diagnostic performance of the beta-glucan assay in the detection of intra-amniotic infection with Candida species

Introduction: A bioassay based on the detection of beta-glucan, a constituent of the cell wall of fungi, has been successfully used to diagnose fungal infections in a variety of biological fluids but not yet in the amniotic fluid.Objective: To determine the diagnostic performance of a beta-glucan bioassay in the detection of Candida species in the amniotic fluid of women who either did or did not have an intrauterine contraceptive device (IUD) in place during an episode of spontaneous preterm parturition.Methods: The study population comprised women who had a singleton pregnancy without congenital or chromosomal abnormalities, who experienced preterm labor or preterm prelabor rupture of the fetal membranes, and who underwent a transabdominal amniocentesis for clinical indications. Samples of amniotic fluid were cultured for aerobic and anaerobic bacteria, genital mycoplasmas, and Candida species, and assayed for beta-glucan, using the (1→3)-beta-d-glucan-specific Limulus amebocyte lysate test (beta-glucan assay) in all cases. Amniotic fluid interleukin (IL)-6 assay results were also available for all cases. The beta-glucan assay takes about 1 hour to run: a concentration >80 pg/mL was considered positive for fungi. Sterile intra-amniotic inflammation of the amniotic cavity was defined by the presence of an amniotic fluid IL-6 concentration ≥2.6 ng/mL and a negative amniotic fluid culture.Results: (1) One hundred ninety-seven (197) women met the study criteria, of whom 58 (29.4%) had an IUD in place; (2) 20 (10.2%) women had a culture of proven intra-amniotic Candida species-related infection, 19 of whom had a positive beta-glucan assay [sensitivity, 95% (19/20; 95% confidence interval (CI): 75.1–99.9%)]; and (3) the specificity of the beta-glucan assay was 75.1% [133/177; 95% CI: 68.1–99.9%]. It was affected by the presence of nonfungal intra-amniotic infections and an IUD, but not by the presence of sterile intra-amniotic inflammation, and there was a significant interaction between the presence of an IUD and nonfungal intra-amniotic infections (estimated for the interaction effect = 2.1923, p value =.026). The assay’s specificity was reduced when nonfungal intra-amniotic infections were diagnosed but only in women who did not have an IUD. Among women without an IUD, the assay’s specificity was 91.4% (117/128); it was 93% (106/114) for those without intra-amniotic infection, and 78.6% (11/14) for those with a nonfungal intra-amniotic infection; the difference was not significant (p = .09). Among women with an IUD, the assay’s specificity was 32.7% (16/49); 42.9% (9/21) for those with a nonfungal intra-amniotic infection; and 25% (7/28) for those without intra-amniotic infection; and the difference was significant (p = .03).Conclusions: The beta-glucan assay is a sensitive, rapid, point-of-care test used to diagnose intra-amniotic Candida species-related infection, and it has a high specificity in pregnant women who did not have an IUD in place.

Detection of intra-amniotic infection in a rabbit model by proteomics-based amniotic fluid analysis

This study was undertaken to identify intra-amniotic infection caused by several different organisms in a rabbit model by using proteomics. Twenty infected and 18 uninfected amniotic fluid samples were subjected to proteomic analysis by surface-enhanced laser desorption ionization (SELDI-TOF, Ciphergen Biosystems, Fremont, Calif), 1- and 2-dimensional gel electrophoresis, and tandem mass spectrometry (MS/MS). Detailed SELDI-TOF spectra revealed a constitutive 4.0 kd peak in all animals. Infected samples also displayed a signature double peak at 3.6 kd. A SELDI-TOF signature profile for intra-amniotic infection predicted positive amniotic fluid and/or fetal cultures with a sensitivity of 90% and specificity of 83%. Similar proteomic profiles were obtained regardless of the infecting organism. The 3.6 kd peak appeared to contain rabbit calgranulin C and rabbit calcyclin, members of the S100 family of calcium binding proteins. Amniotic fluid proteomic analysis was able to detect intra-amniotic infection in this experimental rabbit model. S100 proteins may be involved in the host inflammatory response to intra-amniotic infection.

Bacterial 16S rDNA polymerase chain reaction in the detection of intra-amniotic infection.

Bacterial polymerase chain reaction (PCR) was used to detect early subclinical intraamhiotic infection. We used universal primers which amplify a DNA fragment of 16S ribosomal DNA (rDNA) from all known bacteria and sequenced the positive samples to identify the bacterial species. Transabdominally obtained amniotic fluid samples from 20 pregnant women with prelabour rupture of the fetal membranes (PROM), showing no signs of clinical infection, and 16 control samples were analysed with universal bacterial PCR. In addition, routine bacterial culture and amniotic fluid glucose were studied. Out of 20 PROM patients, five were positive in the PCR. PCR detected Ureaplasma urealyticum in two cases, Haemophilus influenzae in one case, Streptococcus oralis in one case and Fusobacterium sp. in one case. Only two of these were positive in a routine bacterial culture. Both were multibacterial infections, which caused discrepancies between the PCR and culture results. Two patients developed infectious complications: both were identified with the PCR assay. Amniotic fluid glucose was lower in PCR positive patients compared with PCR negative patients. Bacterial 16S rDNA PCR, in properly controlled conditions, promises to be a fast and reliable test for early intra-amniotic infection especially concerning Ureaplasma urealyticum.

Evaluation of rapid diagnostic tests in the detection of microbial invasion of the amniotic cavity

Abstract OBJECTIVE: We sought to determine and compare the value of several rapid diagnostic tests in the detection of intraamniotic infection. STUDY DESIGN: Gram stain, intraamniotic glucose level determination, leukocyte esterase assay, and the Limulus amebocyte lysate assay were performed on 144 amniotic fluid specimens retrieved by transabdominal amniocentesis in 136 patients with preterm premature rupture of the membranes or preterm labor. Diagnostic indices for a positive amniotic fluid culture and the development of clinical infection were calculated for each rapid test. Receiver-operator characteristic curves were generated to help select the optimal glucose level and combination of tests to detect intraamniotic infection. RESULTS: The greatest sensitivity for predicting either a positive culture or subsequent clinical infection in preterm labor patients and in predicting clinical infection in patients with preterm premature rupture of the membranes was demonstrated by a low glucose level. The Gram stain provided the greatest positive predictive value in patients with preterm labor. Combining the Gram stain and measurement of intraamniotic glucose levels did not improve sensitivity above glucose alone or positive predictive value above Gram stain alone. CONCLUSION: Leukocyte esterase determination and Limulus amebocyte lysate assay are insensitive indicators of intraamniotic infection. Selection of Gram stain or glucose level measurement alone or in combination as an appropriate screen for intraamniotic infection will depend on the clinicians' false-positive rate threshold.

Evaluation of rapid diagnostic tests in the detection of microbial invasion of the amniotic cavity

OBJECTIVE: We sought to determine and compare the value of several rapid diagnostic tests in the detection of intraamniotic infection. STUDY DESIGN: Gram stain, intraamniotic glucose level determination, leukocyte esterase assay, and the Limulus amebocyte lysate assay were performed on 144 amniotic fluid specimens retrieved by transabdominal amniocentesis in 136 patients with preterm premature rupture of the membranes or preterm labor. Diagnostic indices for a positive amniotic fluid culture and the development of clinical infection were calculated for each rapid test. Receiver-operator characteristic curves were generated to help select the optimal glucose level and combination of tests to detect intraamniotic infection. RESULTS: The greatest sensitivity for predicting either a positive culture or subsequent clinical infection in preterm labor patients and in predicting clinical infection in patients with preterm premature rupture of the membranes was demonstrated by a low glucose level. The Gram stain provided the greatest positive predictive value in patients with preterm labor. Combining the Gram stain and measurement of intraamniotic glucose levels did not improve sensitivity above glucose alone or positive predictive value above Gram stain alone. CONCLUSION: Leukocyte esterase determination and Limulus amebocyte lysate assay are insensitive indicators of intraamniotic infection. Selection of Gram stain or glucose level measurement alone or in combination as an appropriate screen for intraamniotic infection will depend on the clinicians' false-positive rate threshold.

Detection of intra-amniotic infection by gas-liquid chromatography.

The use of gas-liquid chromatography to detect short-chain organic acids in the amniotic fluid of patients with amnionitis has been previously described. Most of the studies describe patients in the early third trimester with such infections. The purpose of the current study was to confirm the correlation of infection with increased production of organic acids and to assess the effect of gestational age on the presence of these short-chain fatty acids in the amniotic fluid. Six patients with confirmed chorioamnionitis were used as positive control subjects. Seventy-two patients at various gestational ages from 18 to 42 weeks with negative Gram's stain and culture results from the amniotic fluid were used as negative control subjects. The data revealed an increased production of organic acids, particularly pyruvic, oxalic and succinic, in patients with amnionitis regardless of gestational age. Interestingly, patients with noninfected amniotic fluid also revealed an increase in the concentrations of volatile organic acids between 27 and 32 weeks' gestation. It appears from this study that previous results correlating chorioamnionitis with an increased production of organic acids in the amniotic fluid may have been confounded by gestational age.

192 Usefulness of amniotic fluid glucose measurement in detection of intraamniotic infection

192 Usefulness of amniotic fluid glucose measurement in detection of intraamniotic infection

Diagnosis of Intra-Amniotic Infection: The Acridine Orange Stain

The rapid diagnosis of intra-amniotic infection in patients with premature rupture of the membranes (PROM) and preterm labor is of utmost importance. The Gram's stain examination of amniotic fluid can only detect half of these infections. The acridine orange (AO) stain has been claimed to be superior to the Gram's stain in the detection of bacteria in biologic fluids. Therefore, we undertook the present study to examine the value of AO in the detection of bacteria in amniotic fluid. One hundred and thirty-eight patients with a culture-documented prevalence of intra-amniotic infection of 23.2% were studied. The diagnostic indices of the AO were: sensitivity, 43.8%, specificity, 97.2%, positive predictive value, 82.8%, and negative predictive value, 85.1%; the diagnostic indices for the Gram's stain were: sensitivity, 46.8%, specificity, 98.1%, positive predictive value, 88.2%, and negative predictive value, 85.9%. There was no difference in the diagnostic values of the tests. The agreement between the two techniques was substantial, as indicated by a kappa index of 0.72 (SE = 0.17, p less than 0.001). The AO offered no significant advantage over the Gram's stain examination of amniotic fluid in the detection of intra-amniotic infection. However, the AO stain was able to identify mycoplasma infections that escaped detection by the Gram's stain. Therefore, the AO is presently the only microscopic technique capable of detecting mycoplasma in amniotic fluid.

The value of the leukocyte esterase test in diagnosing intra-amniotic infection.

This communication examines the diagnostic value of the leukocyte esterase activity (LEA) test in the detection of intra-amniotic infection. Amniotic fluid from 171 consecutive patients with premature rupture of membranes (n = 149) and preterm labor (n = 22) was obtained through amniocentesis. Gram stain, bacterial and mycoplasma cultures, and LEA tests were performed. The LEA had a sensitivity of 19%, a specificity of 86.7%, a positive predictive value (PPV) of 42.3% and a negative predictive value (NPV) of 67.6% in the prediction of a positive amniotic fluid culture (prevalence of positive cultures = 33.9%). The Gram stain had a sensitivity of 36.2%, specificity of 94.7%, PPV of 77.8%, and NPV of 74.3%. When both tests were combined, a significant increase in sensitivity to 50% was observed. This was associated with a drop in specificity to 81.4%. There was a correlation between the number of white blood cells in the amniotic fluid and the result of the LEA test. A positive LEA assay was associated with an increased likelihood of postpartum endometritis, but not with clinical chorioamnionitis or neonatal infectious morbidity.

Do blood and meconium affect the detection of endotoxin in amniotic fluid with the limulus amebocyte gel clot assay?

The limulus amebocyte lysate (LAL), assay is the most sensitive technique for the detection of endotoxin in biological fluids. Because endotoxin is a component of gram-negative bacteria, the assay has been employed in the detection of gram-negative bacterial contamination of biological fluids. The LAL assay is rapid, inexpensive, easy to perform, and requires little laboratory expertise. When used in conjunction with the gram stain examination of amniotic fluid, it improves the detection of intra-amniotic infection before the availability of culture results. However, the usefulness of the LAL assay in the detection of endotoxin in other body fluids is limited by the presence of an inhibitor to the gelation of the assay. The studies reported in this communication were undertaken to establish if amniotic fluid contains such an inhibitor. Sterile amniotic fluid (AF) samples obtained from 93 patients by transabdominal amniocentesis before labor were used to determine the ED 50 dose of endotoxin necessary for a positive LAL result. The ED 50 dose of endotoxin required for gelation was significantly higher when AF--rather than pyrogen-free saline--was used as the diluent, implying that inhibitors are in fact present (ED 50 = 58.3 pgm/ml). The presence of blood or meconium in the AF did not enhance inhibition significantly: ED 50 doses were 58.3 pgm/ml and 56.2 pgm/ml, respectively. This is not significantly different from the ED 50 of clear amniotic fluid.