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J. Perinat. Med. 38 (2010) 275–279 • Copyright by Walter de Gruyter • Berlin • New York. DOI 10.1515/JPM.2010.001

The frequency and clinical significance of intra-amniotic and/or in women with placenta previa and vaginal : an unexpected observation

Ichchha Madan1,2, Roberto Romero1–3,*, Juan Results: 1) The prevalence of intra-amniotic infection was Pedro Kusanovic1,2, Pooja Mittal1,2, Tinnakorn 5.7% (2/35), and that of IAI was 17.9% (5/28); 2) the ges- Chaiworapongsa1,2, Zhong Dong1, Shali Mazaki- tational age at delivery was lower in patients with IAI than Tovi1,2, Edi Vaisbuch1,2, Zeynep Alpay Savasan1,2, in those without IAI w29.4 weeks, interquartile range (IQR): Lami Yeo1,2, Chong Jai Kim1,4 and Sonia S. 23.1–34.7 vs. 35.4 weeks, IQR: 33.9–36.9; Ps0.028x; and Hassan1,2 3) patients with placenta previa and IAI had a higher rate of s 1 delivery within 48 h (80% (4/5) vs. 19% (4/21); P 0.008) Perinatology Research Branch, NICHD/NIH/DHHS, than those without IAI. Bethesda, Maryland and Detroit, MI, USA 2 Conclusions: Patients with placenta previa presenting with Department of Obstetrics and Gynecology, Wayne State vaginal bleeding have intra-amniotic infection in 5.7% of the University School of Medicine, Detroit, MI, USA 3 cases, and IAI in 17.9%. IAI in patients with placenta previa Center for Molecular Medicine and Genetics, Wayne and vaginal bleeding is a risk factor for preterm delivery State University, Detroit, MI, USA within 48 h. 4 Department of Pathology, Wayne State University School of Medicine, Detroit, MI, USA Keywords: Chorioamnionitis; cytokines; deciduitis; idiopathic vaginal bleeding; intra-amniotic inflammation; IL-6; prematurity; preterm delivery; preterm labor. Abstract

Objective: Idiopathic vaginal bleeding, a common compli- cation of , increases the risk of small-for-gestation- Introduction al age (SGA) neonate, preeclampsia and preterm delivery and can be the only clinical manifestation of intra-amniotic Placenta previa complicates 0.3–0.5% of w23, infection and/or inflammation (IAI). Placenta previa is 25x, and in ;70% of cases, women will experience at least thought to be protective against ascending intrauterine infec- one episode of vaginal bleeding during the second and third tion, yet an excess of histologic chorioamnionitis has been trimesters of pregnancy w25, 30x. Contractions associated reported in this condition. The aim of this study was to deter- with cervical change can cause partial detachment of the pla- mine the frequency and clinical significance of IAI in women centa and it has been proposed that this is the mechanism with placenta previa and vaginal bleeding in the absence of whereby vaginal bleeding occurs in these cases w30x. How- preterm labor. ever, patients with placenta previa can also present with an Study design: A retrospective cohort study including 35 episode of vaginal bleeding in the absence of contractions. women with placenta previa and vaginal bleeding Idiopathic vaginal bleeding is a common complication of -37 weeks of gestation who underwent was pregnancy and has been associated with adverse pregnancy undertaken. Patients with multiple gestations were excluded. outcome w18x, such as preeclampsia w29x, small-for-gesta- Intra-amniotic infection was defined as a positive culture for tional age (SGA) neonate w1, 21, 44x and preterm delivery microorganisms, and intra-amniotic inflammation as an ele- w1, 7, 17, 21, 26, 27, 43, 44x. Recently, idiopathic vaginal vated interleukin (IL)-6 concentration. IL-6 bleeding during pregnancy has been identified as the only concentrations were determined by ELISA in 28 amniotic clinical manifestation of intrauterine infection w17x. Tradi- fluid samples available. Non-parametric statistics were used tionally, it has been believed that placenta previa may be for analysis. protective against ascending intrauterine infection. Yet, pathologic studies have reported an excess of histologic *Corresponding author: chorioamnionitis in this condition w31x. A recent study dem- Roberto Romero, MD Perinatology Research Branch, NICHD, NIH, DHHS onstrated that a subset of patients with placenta previa and Wayne State University/Hutzel Women’s Hospital an episode of spontaneous preterm labor with intact mem- 3990 John R, Box 4 branes have evidence of intra-amniotic infection and/or Detroit, MI 48201 inflammation (IAI) w31x. Thus, the aim of this study was to USA Tel.: q1 (313) 993-2700 determine the frequency and clinical significance of IAI in Fax: q1 (313) 993-2694 women with placenta previa and vaginal bleeding in the E-mail: [email protected] absence of preterm labor.

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276 Madan et al., The frequency and clinical significance of IAI in women with placenta previa and vaginal bleeding

Materials and methods amniotic cavity and/or assessment of fetal lung maturity. Amniotic fluid samples were transported to the laboratory in a sterile capped Study design and population syringe and cultured for aerobic/anaerobic and genital . White cell count, glucose concentration and A retrospective cohort study was conducted by searching our clin- Gram stain were also performed shortly after collection as previ- ical database and bank of biological samples and included patients ously described w37, 38, 41x. The results of these tests were used admitted with the diagnosis of placenta previa and vaginal bleeding for clinical management. Amniotic fluid not required for clinical who met the following criteria: 1) singleton gestation; 2) diagnosis assessment was centrifuged for 10 min at 48C and the supernatant of placenta previa confirmed by ultrasound; 3) gestational age was aliquoted and stored at –708C. Amniotic fluid IL-6 concentra- -37 weeks; and 4) amniocentesis performed. Patients with preterm tions were determined for research purposes in 28 patients whose labor, preterm prelabor (PROM) and those amniotic fluid samples were available for analysis. with suspected were excluded from the study. Between January 1998 and July 2007, 168 patients with placenta Statistical analysis previa and vaginal bleeding were admitted to the Sotero del Rio Hospital in Puente Alto, Santiago, Chile. Patients presenting with The normality of the data was tested using the Shapiro-Wilk and vaginal bleeding to this institution are offered amniocentesis as part Kolmogorov-Smirnov tests. Since the data were not normally dis- of standard obstetrical practice because previous studies have sug- tributed, non-parametric tests were used for analyses. Comparisons gested an association between vaginal bleeding, intra-amniotic between proportions were performed with x2-test, and Mann-Whit- infection and chorioamnionitis w8, 17x. After counseling, 66 patients ney U-test was used for analyses of continuous variables. A P-value underwent amniocentesis, and after exclusion criteria were applied, of -0.05 was considered statistically significant. The statistical 35 patients were left for analysis. All women provided written package used was SPSS v.15.0 (SPSS Inc, Chicago, IL, USA). informed consent prior to the collection of amniotic fluid. The col- lection and utilization of amniotic fluid for research purposes was approved by the Institutional Review Boards of the Sotero del Rio Hospital and the Eunice Kennedy Shriver National Institute of Child Results Health and Human Development, NIH, DHHS. Demographic and clinical characteristics Definitions of the study population

Placenta previa was defined as a placenta that overlies or is proxi- Table 1 displays the demographic and clinical characteristics mate to the internal cervical os based on trans-vaginal ultrasound of patients included in the study. There was no significant w30x. Patients with low lying placenta at the time of amniocentesis difference in the median maternal age and pregestational were not included in the study. Preterm labor was defined by the body mass index, as well as in the proportion of nulliparity presence of regular uterine contractions occurring at a frequency of and history of previous between patients with at least two every 10 min before 37 weeks of gestation. Vaginal and without IAI. Among patients with placenta previa and bleeding was diagnosed by sterile speculum examination confirming vaginal bleeding, those with IAI had a lower gestational age the presence of blood coming through the external os of the cervix. at amniocentesis than those without IAI (Table 1). Intra-amniotic infection was defined as a positive amniotic fluid culture for microorganisms. Intra-amniotic inflammation was diag- nosed by an amniotic fluid interleukin (IL)-6 concentration The prevalence and clinical significance of IAI in G2.6 ng/mL w48x. patients with placenta previa and vaginal bleeding

Sample collection Among patients with placenta previa and vaginal bleeding, the prevalence of intra-amniotic infection was 5.7% (2/35), Amniotic fluid samples were obtained by trans-abdominal amnio- and that of IAI was 17.9% (5/28). The microorganisms centesis performed for evaluation of the microbial status of the recovered from the amniotic cavity were

Table 1 Demographic and clinical characteristics of patients with and without IAI.

With IAI (ns5) Without IAI (ns23) P-value Maternal age (years) 32 (27.5–38.0) 32 (28.0–35.8) 0.9 Pregestational BMI (kg/m2) 23.2 (20.5–28.3) 24.8 (22.3–29.0) 0.5 Smoking (%) 0 (0/5) 4.3 (1/23) 0.6 Nulliparous (%) 0 (0/5) 21.7 (5/23) 0.3 Previous preterm delivery (%) 0 (0/5) 21.7 (5/23) 0.3 Gestational age at amniocentesis (weeks) 23.6 (23.0–30.7) 31.1 (28.1–33.9) 0.03 Gestational age at delivery (weeks) 29.4 (23.1–34.7) 35.4 (33.9–36.9) 0.03 Birth weight (g) 1570 (545–2620) 2410 (1970–2913) 0.06 Amniotic fluid WBC count (cells/mm3) 5 (1–430) 2 (0–5) 0.2 Amniotic fluid IL-6 (ng/mL) 58.5 (2.1–145.5) 0.9 (0.5–1.3) 0.03 Results are expressed as median (interquartile range) or percentage (proportion). IAIsintra-amniotic infection and/or inflammation, BMIsbody mass index, WBCswhite blood cell, IL-6sinterleukin-6. Article in press - uncorrected proof

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and viridans. The gestational age at delivery from the same group of investigators w31x in patients with was significantly lower in patients with IAI than in those spontaneous preterm labor and intact membranes in cases of without IAI w29.4 weeks, interquartile range (IQR): placenta previa reported a rate of 4.9%, 16.7% and 19% for 23.1–34.7 vs. 35.4 weeks, IQR: 33.9–36.9; Ps0.028x. intra-amniotic infection, IAI and histologic chorioamnionitis, Patients with placenta previa and vaginal bleeding respectively. Among patients with histologic chorioamnio- -28 weeks of gestation had a higher rate of IAI than those nitis, inflammation of the choriodecidua (exposed to the cer- admitted )28 weeks, although this difference was not sta- vical canal) was present in all cases (8/8), but inflammation tistically significant w30% (3/10) vs. 11.1% (2/18); Ps0.2x. of the chorionic plate existed in 63% (5/8) of them w31x.It Clinical information at the time of delivery was available is possible that placenta previa may act as a mechanical bar- in 33 cases (two patients delivered elsewhere). Overall, the rier preventing, at least partially, ascending intra-amniotic rate of delivery -34 weeks and -37 weeks was 42.4% infection w31x. (14/33) and 81.2% (27/33), respectively. Patients with pla- Idiopathic vaginal bleeding is associated with intra-amni- centa previa and vaginal bleeding, in the presence of IAI, otic infection in 14% of patients w17x. The finding that 5.7% had a significantly higher rate of delivery within 48 h w80% of patients with placenta previa and vaginal bleeding have (4/5) vs. 19% (4/21); Ps0.008), 7 days w80% (4/5) vs. intra-amniotic infection suggest that vaginal bleeding may 28.6% (6/21); Ps0.03x and at -32 weeks of gestation w75% be also a manifestation of microbial invasion in a subset of (3/4) vs. 8.3% (1/12); Ps0.008x than those without IAI. No patients with placenta previa. The microorganisms recovered differences were observed in the rate of delivery at -34 from the amniotic cavity in the two patients in this study w80% (4/5) vs. 43.8% (7/16); Ps0.2x. were Escherichia coli and Streptococcus viridans. These bac- Four patients with IAI delivered within 48 h and at teria have been previously reported in cervical and vaginal F32 weeks. Three cases developed spontaneous preterm flora of pregnant women w22, 45, 46x suggesting that, even labor after admission and one case required an emergency in the presence of placenta previa, ascending infection is still cesarean section due to persistent bleeding. Interestingly, one a plausible mechanism for microorganisms to gain access to case had an elevated amniotic fluid IL-6 concentration the amniotic cavity. Thus, the traditional view that vaginal (3.9 ng/mL) at 23.6 weeks but delivered at term. In addition, bleeding in patients with placenta previa is likely to be due one of the patients with intra-amniotic infection (amniotic to the presence of the placenta previa itself and not of IAI, fluid culture positive for Escherichia coli) had normal amni- needs to be reconsidered. Exclusion of IAI should be con- otic fluid IL-6 concentration (0.26 ng/mL) and delivered sidered as part of the clinical evaluation of women with pla- within 24 h. centa previa presenting with vaginal bleeding, because those patients are at increased risk for adverse pregnancy out- comes. Further studies are required to determine the preva- Discussion lence and significance of intra-amniotic infection using molecular microbiologic techniques in these patients. Principal findings of the study The clinical significance of vaginal bleeding in Among patients with placenta previa and idiopathic vaginal patients with placenta previa bleeding, the prevalence of intra-amniotic infection was 5.7% and that of IAI 17.9%. Similar to pregnant women with Bleeding of unknown etiology in the second half of preg- normal placentation and IAI, patients with placenta previa nancy has been associated with adverse pregnancy outcome and IAI are at increased risk for delivery within 48 h, 7 days including preterm delivery w1, 7, 17, 21, 26, 27, 43, 44x, and at -32 weeks of gestation. preterm PROM w17, 20, 21x, preeclampsia w29x, SGA w1, 21, 44x, w1, 26x, fetal anomalies w1, 7, 26, 43, 44x,as Intra-amniotic infection in the context of placenta well as admission to neonatal intensive care unit, reduced w x previa birth weight and low Apgar scores 21, 27 . This study further demonstrates that, among patients with It has been proposed that microorganisms may gain access placenta previa and idiopathic vaginal bleeding, the gesta- to the amniotic cavity through any of the following path- tional age at delivery was significantly lower in those with ways: 1) ascending infection from the vagina and the cervix; IAI compared to those without IAI, as well as a significantly 2) hematogenous dissemination through the placenta; 3) ret- higher rate of delivery within 48 h, 7 days and at rograde seeding from the peritoneal cavity through the fal- -32 weeks. The amniocentesis-to-delivery interval was not lopian tubes; and 4) accidental introduction at the time of significantly different between the two groups (data not invasive procedures (e.g., amniocentesis, chorionic villous shown). This may be explained due to the following: 1) One sampling, shunting). The most common pathway of intra- patient with an elevated amniotic fluid IL-6 concentration at uterine infection is the ascending route w39x. In a cohort of 23.6 weeks of gestation delivered at term (37.3 weeks); and 206 patients with spontaneous preterm labor and intact mem- 2) since patients with placenta previa are delivered at branes, in the absence of placenta previa, the frequency of ;36–37 weeks of gestation w30x, and the median gestational intra-amniotic infection was 10%, IAI was found in 32% of age at amniocentesis in patients without IAI was 31.1 weeks, cases and 53% of the placentas available for analysis had the amniocentesis-to-delivery interval for this group was histologic chorioamnionitis w48x. Interestingly, a recent study similar to the IAI group. Article in press - uncorrected proof

278 Madan et al., The frequency and clinical significance of IAI in women with placenta previa and vaginal bleeding

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