Population-based estimate of sibling risk for preterm birth, preterm premature rupture of membranes, placental abruption and pre-eclampsia

<p>Abstract</p> <p>Background</p> <p>Adverse pregnancy outcomes, such as preterm birth, preeclampsia and placental abruption, are common, with acute and long-term complications for both the mother and infant. Etiologies underlying such adverse outcomes are not well understood. As maternal and fetal genetic factors may influence these outcomes, we estimated the magnitude of familial aggregation as one index of possible heritable contributions.</p> <p>Using the Missouri Department of Health's maternally-linked birth certificate database, we performed a retrospective population-based cohort study of births (1989–1997), designating an individual born from an affected pregnancy as the proband for each outcome studied. We estimated the increased risk to siblings compared to the population risk, using the sibling risk ratio, λ_s, and sibling-sibling odds ratio (sib-sib OR), for the adverse pregnancy outcomes of preterm birth, preterm premature rupture of membranes (PPROM), placental abruption, and pre-eclampsia.</p> <p>Results</p> <p>Risk to siblings of an affected individual was elevated above the population prevalence of a given disorder, as indicated by λ_S(λ_S(95% CI): 4.3 (4.0–4.6), 8.2 (6.5–9.9), 4.0 (2.6–5.3), and 4.5 (4.4–4.8), for preterm birth, PPROM, placental abruption, and pre-eclampsia, respectively). Risk to siblings of an affected individual was similarly elevated above that of siblings of unaffected individuals, as indicated by the sib-sib OR (sib-sib OR adjusted for known risk factors (95% CI): 4.2 (3.9–4.5), 9.6 (7.6–12.2), 3.8 (2.6–5.5), 8.1 (7.5–8.8) for preterm birth, PPROM, placental abruption, and pre-eclampsia, respectively).</p> <p>Conclusion</p> <p>These results suggest that the adverse pregnancy outcomes of preterm birth, PPROM, placental abruption, and pre-eclampsia aggregate in families, which may be explained in part by genetics.</p

Rapid detection of group B streptococcus and Escherichia coli in amniotic fluid using real-time fluorescent PCR.

OBJECTIVE: To establish reliability and validity of real-time fluorescent PCR for early detection of bacterial invasion of the amniotic cavity. METHODS: Amniotic fluid samples from 40 patients undergoing mid-trimester genetic amniocentesis were incubated for 6 h at 37 degrees C and were cultured on media specific for group B streptococcus (GBS) and E. coli. Concurrently, samples were analyzed with real-time fluorescent PCR (Roche LightCycler) using DNA primers and probes designed to detect the CAMP factor encoding cfb gene and uidA gene of GBS and E. coli, respectively. For positive control and to simulate amniotic fluid colonization, 104 cfu/ml of GBS and E. coli were inoculated on sterile amniotic fluid and incubated for 6 h. Bacterial genomic DNA for the two organisms was extracted and purified via the two-step precipitation method using a commercial kit. The real-time PCR assays were also tested against 25 non-GBS and non-E. coli bacterial species. The lower limit of detection for each pathogen was established using serial dilution of bacterial genomic DNA. RESULTS: All patient samples were negative for evidence of GBS and E. coli with both culture and real-time PCR methods. Amniotic fluid samples inoculated with GBS and E. coli were positive with real-time PCR whereas the 25 bacterial species other than GBS or E. coli tested negative with the assay. Average total sample processing time including the pre-enrichment step was 7 h 40 min. The average cost for DNA extraction and PCR testing was 8.50 dollars per test. CONCLUSION: Real-time fluorescent PCR is a valid and reliable method for detection of specific pathogens in amniotic fluid. This technique is sensitive for low inoculation levels. Real-time fluorescent PCR has potential to impact clinical management as a rapid, reliable detection method for GBS and E. coli in chorioamnionitis

Is fetal gender associated with adverse perinatal outcome in intrauterine growth restriction (IUGR)?

OBJECTIVE: The purpose of this study was to determine if there is a difference in perinatal outcome by gender among growth-restricted fetuses. STUDY DESIGN: This was a retrospective cohort study of intrauterine growth restriction (IUGR) singleton pregnancies over a 5-year period. Clinical outcomes compared by gender included preterm delivery, perinatal mortality (PNM), respiratory distress syndrome (RDS), grade 3 or 4 intraventricular hemorrhage (IVH), necrotizing enterocolitis (NEC), and periventricular leukomalacia (PVL). Statistical analysis included bivariate and multivariable techniques. RESULTS: Seven hundred and twenty-seven singleton pregnancies with IUGR were identified. Three hundred and forty-six (47.6%) were males. Birth weight was similar between the groups. After adjusting for maternal demographics, medical history, gestational age, mode of delivery, and antenatal corticosteroids, adverse perinatal outcomes were similar between the groups. Severity of outcomes was also similar between males and females (P = .66). CONCLUSION: Male fetuses with IUGR have similar outcomes when compared with female IUGR fetuses. Gender does not play a role in perinatal outcome in the setting of fetal growth restriction

A population-based study of race-specific risk for placental abruption

<p>Abstract</p> <p>Background</p> <p>Efforts to elucidate risk factors for placental abruption are imperative due to the severity of complications it produces for both mother and fetus, and its contribution to preterm birth. Ethnicity-based differences in risk of placental abruption and preterm birth have been reported. We tested the hypotheses that race, after adjusting for other factors, is associated with the risk of placental abruption at specific gestational ages, and that there is a greater contribution of placental abruption to the increased risk of preterm birth in Black mothers, compared to White mothers.</p> <p>Methods</p> <p>We conducted a population-based cohort study using the Missouri Department of Health's maternally-linked database of all births in Missouri (1989–1997) to assess racial effects on placental abruption and the contribution of placental abruption to preterm birth, at different gestational age categories (n = 664,303).</p> <p>Results</p> <p>Among 108,806 births to Black mothers and 555,497 births to White mothers, 1.02% (95% CI 0.96–1.08) of Black births were complicated by placental abruption, compared to 0.71% (95% CI 0.69–0.73) of White births (aOR 1.32, 95% CI 1.22–1.43). The magnitude of risk of placental abruption for Black mothers, compared to White mothers, increased with younger gestational age categories. The risk of placental abruption resulting in term and extreme preterm births (< 28 weeks) was higher for Black mothers (aOR 1.15, 95% CI 1.02–1.29 and aOR 1.98, 95% CI 1.58–2.48, respectively). Compared to White women delivering in the same gestational age category, there were a significantly higher proportion of placental abruption in Black mothers who delivered at term, and a significantly lower proportion of placental abruption in Black mothers who delivered in all preterm categories (p < 0.05).</p> <p>Conclusion</p> <p>Black women have an increased risk of placental abruption compared to White women, even when controlling for known coexisting risk factors. This risk increase is greatest at the earliest preterm gestational ages when outcomes are the poorest. The relative contribution of placental abruption to term births was greater in Black women, whereas the relative contribution of placental abruption to preterm birth was greater in White women.</p

Rapid detection of group B streptococcus and Escherichia coli in amniotic fluid using realtime fluorescent PCR. Infect Dis Obstet Gynecol

Objective: To establish reliability and validity of real-time fluorescent PCR for early detection of bacterial invasion of the amniotic cavity. Methods: Amniotic fluid samples from 40 patients undergoing mid-trimester genetic amniocentesis were incubated for 6 h at 378C and were cultured on media specific for group B streptococcus (GBS) and E. coli. Concurrently, samples were analyzed with real-time fluorescent PCR (Roche LightCycler) using DNA primers and probes designed to detect the CAMP factor encoding cfb gene and uidA gene of GBS and E. coli, respectively. For positive control and to simulate amniotic fluid colonization, 104 cfu/ml of GBS and E. coli were inoculated on sterile amniotic fluid and incubated for 6 h. Bacterial genomic DNA for the two organisms was extracted and purified via the two-step precipitation method using a commercial kit. The real-time PCR assays were also tested against 25 non-GBS and non-E. coli bacterial species. The lower limit of detection for each pathogen was established using serial dilution of bacterial genomic DNA. Results: All patient samples were negative for evidence of GBS and E. coli with both culture and real-time PCR methods. Amniotic fluid samples inoculated with GBS and E. coli were positive with real-time PCR whereas the 25 bacterial species other than GBS or E. coli tested negative with the assay. Average total sample processing time including the pre-enrichment step was 7 h 40 min. The average cost for DNA extraction and PCR testing was $8.50 per test. Conclusion: Real-time fluorescent PCR is a valid and reliable method for detection of specific pathogens in amniotic fluid. This technique is sensitive for low inoculation levels. Real-time fluorescent PCR has potential to impact clinical management as a rapid, reliable detection method for GBS and E. coli in chorioamnionitis. Key words: REAL-TIME FLUORESCENT PCR; E. COLI; GBS INTRODUCTION Preterm birth occurs in approximately 11% of all pregnancies. Preterm labor may be initiated by infection in 33% of cases with intact membranes and in 40% with preterm premature rupture of membranes (pPROM). Early preterm birth (5 32 weeks gestation) has the strongest association with infection 1 . Neonatal sepsis occurs more commonly with early preterm birth, accounting for a large portion of perinatal morbidity and mortality. Pathogens that have been isolated from amniotic fluid in these cases include Ureaplasm urealyticum, Mycoplasm hominis, fusobacterium species, prevo- tella species, gardnarella vaginalis, streptococcus agalactiae, anaerobic streptococci and Escherichia coli 2,3 . In the pathogenesis of intra-amniotic infection, or chorioamnionitis, it appears that vaginal organisms overcome available cervical barrier and immunity factors and penetrate the chorioamnion to enter the amniotic fluid. Ultimately, some fetuses become infected 4 . Identifying the gravid patient destined to deliver prematurely because of ascending genital tract infection is difficult. Many patients remain asymptomatic until they present in preterm labor, and most of them do not show signs of chorioamnionitis. Even patients with pPROM often lack clinical evidence of infection upon presentation. Current clinical management of preterm labor and pPROM does not universally include routine amniocentesis because there are no rapid, reliable diagnostic tests for intraamniotic infection and because infection rates vary among clinical populations. Amniotic fluid culture takes 48 to 72 h, and may lack the sensitivity needed to reliably identify infection at a stage when intervention could alter outcome. Amniotic fluid glucose, Gram stain and leukocyte esterase assays provide rapid results, but have poor predictive value for infection-mediated preterm labor, and even less for identifying the neonate destined to develop sepsis. Ideally, accurate identification of bacterial invasion of the amniotic cavity might allow expedient and targeted management of infection-mediated preterm labor. PCR is used to identify genetic material of pathogenic microorganisms by amplification, with highly specific and sensitive results, and has been shown to detect bacteria in amniotic fluid at a higher rate than standard microbial cultures The objective of our study was to develop a valid, reliable test for detection of GBS and E. coli in amniotic fluid. MATERIALS AND METHODS This study was initiated after approval by the Institutional Review Board at David Grant USAF Medical Center. For genetic analysis, 40 amniotic fluid samples were obtained from women undergoing mid-trimester amniocentesis. To test amniotic fluid samples for GBS and E.coli infection by standard culture and conventional and real-time PCR, 2 ml of each sample were incubated for 6 h at 37 C. Samples were frozen, stored at -70 C and batched for later analysis. An aliquot of each sample was cultured according to CDC recommendations for detection of GBS and E.coli. A second aliquot was processed for traditional and realtime fluorescent PCR using a Roche LightCycler. Bacterial DNA was extracted using the Gentra Systems Puregene DNA isolation kit, modified by the addition of a second precipitation step. Pathogen genomic targets The real-time PCR probe for GBS targeted the cfb gene encoding the Christie-Atkins-MunchPetersen (CAMP) factor 7 . Hybridization probes designed to anneal to an internal sequence of the 153 base pair (bp) amplicon were used. The E. coli target for the assay consists of a 186 bp sequence of the uidA gene encoding the b-glucoronidase protein. GBS or E. coli was detected in each sample in triplicate using the Hybridization (GBS) or Taqman (E.coli) probe systems in the LightCycler. Specificity of assays To assess the specificity of the Hybridization and Taqman probe assays, a battery of 25 non-GBS and non-E coli bacterial strains were grown on blood agar plates, and 40 ml of a 104 cfu/ml bacterial suspension of each strain were added to 500 ml amniotic fluid sample and incubated for 6 h at 37 C. The sample was then tested for the presence of GBS and E coli target DNA on the LightCycler, in duplicate. Real-time fluorescent PCR Straka et al. . INFECTIOUS DISEASES IN OBSTETRICS AND GYNECOLOGY Spiked samples Streptococcus agalactiae (ATCC 12386) and E. coli (ATCC 35218) were grown in pure culture on blood agar plates and diluted to 104 cfu/ml. Then 40 ml (25 cfu total) of each of the bacterial suspensions were added to 500 ??mu?l aliquots of amniotic fluid, incubated at 37 C for 6 h and stored at -80 C until analyzed. PCR-grade distilled, de-ionized water was used as negative control for both traditional PCR and real-time PCR. The dilute bacterial suspensions served as the positive control for traditional and real-time PCR. Limit of detection To determine the lower limit of detection of the assays, pure genomic DNA extracted from GBS or E. coli cultured on sheep blood agar was obtained using the Gentra Puregene Isolation kit with the modification noted above. RNAse and proteinase K treatment steps after DNA isolation were added to remove contaminating proteins and RNA. The concentration of DNA was calculated by measuring absorbance at 260 and 280 nm. DNA concentration was adjusted to 1 mM and serial dilution of DNA in Tris buffer was performed. GBS or E. coli was detected from each concentration in triplicate using the Hybridization (GBS) or Taqman (E. coli) probe systems in the LightCycler. RESULTS As was anticipated on the basis of the clinical histories, none of the 40 native amniotic fluid samples had evidence of E.coli or GBS by culture, conventional PCR or real-time PCR. All amniotic fluid samples inoculated with GBS or E. coli were uniformly positive by real-time PCR DISCUSSION We have shown that real-time PCR is a sensitive (100%) and rapid method for detecting pathogens in amniotic fluid that are commonly involved in the pathogenesis of preterm labor and adverse neonatal outcome. We consistently detected artificially created infected amniotic fluid samples and consistently detected greater than 50 cfu/ml of pathogen. We also proved that the real-time PCR system is highly specific (100%), by showing that the probes tested did not detect alternative bacterial species or produce false-positive results in control samples devoid of bacteria. Real-time PCR proved to be valid and reliable when We have also shown with sensitive and specific biomolecular techniques that the amniotic cavity under uncomplicated clinical conditions is sterile with respect to GBS and E. coli in the midtrimester. If confirmed in future studies, the clinical implications of our data may lead to changes in diagnostic methods and clinical management of chorioamnionitis, preterm labor and pPROM. Future research should focus on validating this real-time PCR system in other clinical populations, including cases of suspected preterm labor, pPROM or chorioamnionitis. We are currently planning such a study, to include patients with these diagnoses. We also plan to correlate PCR results with various adverse neonatal outcomes. With further study, it may become possible to more accurately and more rapidly identify earlier in gestation those pregnancies destined to deliver premature neonates that may develop sepsis. Early discovery of intra-amniotic infection may allow specific therapy in utero and maintainance of such pregnancies closer to term, as in the case of listeriosis. Conversely, early identification of intra-amniotic bacterial invasion may indicate expeditious delivery rather than ineffective maternal tocolytic or antibiotic therapy as the best strategy for reducing neonatal infectious morbidity and mortality. . INFECTIOUS DISEASES IN OBSTETRICS AND GYNECOLOGY In summary, the results of our study suggest that real-time PCR technology is a promising method that may improve diagnostic accuracy and rapidity in identifying intra-amniotic infection, and may open new windows for applying therapy to prevent preterm delivery and neonatal morbidity and mortality from prematurity. However, before this technique can be applied clinically, further research is necessary. The sensitivity and specificity of real-time PCR needs to be evaluated and validated in the most important clinical target population, namely women at 24 to 32 weeks of gestation who are at increased risk for preterm delivery or pPROM or are suspected to have preterm labor. In addition, probes for other common and uncommon pathogens need to be tested

Should all pregnant diabetic women undergo a fetal echocardiography? A cost-effectiveness analysis comparing four screening strategies.

OBJECTIVE: To determine if a policy of universal fetal echocardiography for all pregnant diabetic women is cost-effective as a screening tool for congenital heart defects. STUDY DESIGN: Using a decision-analysis model, we compared the cost-effectiveness of four screening strategies: (1) none--no ultrasound is performed; (2) selective fetal echocardiography after abnormal detailed anatomic survey; (3) fetal echocardiography for only high hemoglobin A1C, and (4) universal fetal echocardiography for all diabetics. The sensitivity and specificity for each strategy were derived by literature search. The analysis was from a societal perspective using a willingness-to-pay threshold (50,000 dollars) and a theoretic cohort of 40,000 pregnant diabetics. Costs included costs of tests and the costs of complications and of raising a child with a cardiac defect. Outcomes were reported as cost per quality-adjusted life years (QALY) gained for each congenital heart defect prevented by each strategy and the number of congenital heart defects detected. One-way, multiway and probabilistic sensitivity analyses were performed. RESULTS: Compared with the other strategies, selective fetal echocardiography after abnormal detailed anatomic survey costs less per QALY gained for cardiac defect screening. Although universal fetal echocardiography was associated with a higher detection rate for cardiac defects, it was more costly. The sensitivity analyses revealed a robust model over a wide range of values. CONCLUSION: Under the baseline assumptions, selective fetal echocardiography after an abnormal detailed anatomic survey is more cost-effective compared with universal fetal echocardiography as a screening strategy for cardiac defects in pregnant diabetics