Véronique Houfflin-Debarge

High Sensitivity of Fetal DNA in Plasma Compared to Serum and Nucleated Cells Using Unnested PCR in Maternal Blood

DNA analysis of blood is conventionally performed on cells – plasma and serum are discarded. Free DNA has been demonstrated in serum in cancer and autoimmune disorders and in pregnancy. We investigated possible noninvasive prenatal diagnosis using fetal DNA from maternal plasma and serum in pregnancy. Fetal gender was determined by PCR on DNA from maternal venous blood, serum and plasma of 65 women by boiling with or without phenol/chloroform extraction. When sensitivities were compared for plasma, additional phenol/chloroform extraction proved more sensitive than boiling alone (89 vs. 50%), the observed difference was 50% (CI 19 to 81%). Extracted plasma amplified better than extracted serum (89 vs. 46%), the observed difference being 44% (CI 22 to 66%). In contrast, fetal gender could not reliably be determined from DNA extracted from maternal nucleated blood cells. The size of plasma and serum DNA at 15–17 weeks of gestation was >1,500 bp. This work confirms the presence of fetal DNA in maternal plasma and serum which may be applicable to noninvasive prenatal diagnosis of paternally derived DNA sequences. We conclude that optimal sensitivity requires two methods of DNA extraction and that the use of plasma is preferred to that of serum.

Ultrasound diagnosis, management and prognosis in a consecutive series of 27 cases of fetal hydrops following maternal parvovirus B19 infection.

Introduction: Fetal hydrops caused by anemia from parvovirus B19 infection (FH-B19) is rare. Doppler measurement of the middle cerebral artery peak systolic velocity (PSV-MCA) improves its prenatal diagnosis, but its frequency and prognosis are still poorly known. Despite improved survival due to in utero transfusions, the possibility of late neurological sequelae makes prognosis uncertain. Objectives: To assess the frequency, management and prognosis of a consecutive series of FH-B19 observed over a 15-year period. Methods: Retrospective study of 27 cases of FH-B19, that is, 3/100,000 births, 24 of them discovered during routine second-trimester ultrasound. All but 1 case (96.2%) had at least four of the six ultrasound signs that Saltzman et al. [Obstet Gynecol 1989;74:106–111] suggested as indicators of anemia. Of the fetuses tested, 80% had a PSV-MCA >1.5 MoM, also indicative of anemia. Of the 19 fetuses treated by exchange transfusions, 11 were liveborn compared with 2 of the 6 not so treated (57.8 vs. 33.3%, NS). The survival rate was higher during the second half of the study period (23.1 vs. 71.4%, p < 0.02) for less severe anemia (p < 0.03) and for repeated transfusions (p = 0.03). In our series, 1 case of prenatal cerebral atrophy was identified on screening. All 13 liveborn children appeared healthy at the age of 1 year. Conclusion: In cases of fetal hydrops, Saltzman et al.’s ultrasound criteria and PSV-MCA measurement made it possible to determine the likelihood that anemia is the cause of the hydrops and to measure its intensity. Use of these techniques allowed us to choose the most appropriate treatment (transfusion or not, depending on the degree of anemia), and survival improved notably in our series.

Prise en charge médicale per et post-natale de la hernie congénitale du diaphragme

Resume La physiopathologie de cette malformation est complexe, et malgre les progres de la reanimation, la mortalite neonatale reste elevee proche de 30-40 % essentiellement du fait de l’hypoplasie pulmonaire et de l’hypertension arterielle pulmonaire (HTAP). Une meilleure connaissance physiopathologique est necessaire pour ameliorer la prise en charge de ces enfants. Par ailleurs, la HDC s’accompagne d’une morbidite sequellaire elevee qui touche environ la moitie des enfants vivants. Les principales sequelles observees sont respiratoires (HTAP chronique, dysplasie broncho-pulmonaire, susceptibilite aux infections virales), digestives (reflux gastro – œsophagien, trouble de l’oralite), nutritionnelles (denutrition d’origine multifactorielle : RGO, trouble de l’oralite, insuffisance respiratoire), et orthopedique (scoliose). La prise en charge actuelle de ces complications est tardive : elle necessite une intervention plus precoce, qui doit debuter des la periode neonatale. Elle necessite imperativement un suivi et une prise en charge multidisciplinaire specialisee du fait de l’intrication de ces differentes complications. Ainsi, les objectifs de la prise en charge en periode neonatale sont de reduire la mortalite immediate, essentiellement liee a un echec de l’adaptation a la vie extra-uterine, mais aussi de mettre en place, des la naissance, des mesures de prevention de la morbidite a long terme.Congenital diaphragmatic hernia (CDH) is a 1 out of 3500 live-born malformation with persistent 30-40% mortality rate, related to severe pulmonary hypoplasia and hypertension. Better knowledge on the mechanisms inducing failure of adaptation at birth is a prerequisite for improving CDH prognosis. CDH is also associated with longterm morbidity, including prolonged respiratory failure, failure to growth, oral aversion, and scoliosis. Early prevention starting as soon as the first hours of life are required to reduced long term morbidity. The aims of the management are not only to reduce early mortality, related to persistent pulmonary hypertension, but also to prevent late morbidity.

Feasibility and safety of intact cord resuscitation in newborn infants with congenital diaphragmatic hernia (CDH)

BACKGROUNDnStarting resuscitation before clamping the umbilical cord at birth may progressively increase pulmonary blood flow while umbilical venous blood flow is still contributing to maintenance of oxygenation and left ventricle preload.nnnOBJECTIVEnTo evaluate the feasibility, safety, and effects of intact cord resuscitation (ICR) on cardiorespiratory adaptation at birth in newborn infants with CDH.nnnSTUDY DESIGNnProspective, observational, single-center pilot study.nnnMETHODSnPhysiologic variables and outcomes were collected prospectively in 40 consecutive newborn infants with an antenatal diagnosis of isolated CDH.nnnRESULTSnInfants were managed with immediate cord clamping (ICC group) from 1/2012 to 5/2014 or the cord was clamped after initiation of resuscitation maneuvers (ICR group) from 6/2014 to 4/2016 (20 in each group). Ante- and postnatal markers of CDH severity were similar between groups. Resuscitation before cord clamping was possible for all infants in the ICR group. No increase in maternal or neonatal adverse events was observed during the period of ICR. The pH was higher and the plasma lactate concentration was significantly lower at one hour after birth in the ICR than in the ICC group (pH=7.17±0.1 vs 7.08±0.2; lactate=3.6±2.3 vs 6.6±4.3mmol/l, p<0.05). Mean blood pressure was significantly higher in the ICR than in the ICC group at H1 (52±7.7 vs 42±7.5mmHg), H6 (47±3.9 vs 40±5.6mmHg) and H12 (44±2.9 vs 39±3.3mmHg) (p<0.05).nnnCONCLUSIONnCommencing resuscitation and initiating ventilation while the infant is still attached to the placenta is feasible in infants with CDH. The procedure may support the cardiorespiratory transition at birth in infants with CDH.

Fetal Heart-Rate Variability: Validation of a New Continuous, Noninvasive Computerized Analysis

Objective: We developed a computerized heart-rate variability index related to the fetal parasympathetic activity: the Fetal Stress Index (FSI). The objective was to determine whether the FSI is related to the visual analysis of the fetal heart rate (FHR). Methods: Thirty tracings recorded at a labor ward were classified according to the NICHD categories: (I) normal FHR tracing, (II) intermediate risk of acidosis, and (III) high risk. FSI was calculated as minimum, maximum, and mean, and was evaluated before the onset of the FHR pattern, during the 10 min following, and between 10 and 20 min after that. Results: The FSI for categories II and III was similar to that of category I before the onset of the FHR pattern. FSI min was lower just after the onset of the abnormal FHR in category III, compared with that of category I (33 vs. 43, p < 0.001). Between 10 and 20 min after the onset of the abnormal FHR, we observed a significant reduction in FSI min in categories II and III (44 vs. 39 vs. 29.7, p < 0.0001). Conclusion: Although further studies are necessary for the sake of clinical validation, FSI could constitute an interesting method for the evaluation of fetal well-being.

Effects of Eicosapentaenoic Acid (EPA) and Docosahexaenoic Acid (DHA) on Fetal Pulmonary Circulation: An Experimental Study in Fetal Lambs

Background: Persistent pulmonary hypertension of the newborn (PPHN) causes significant morbidity and mortality in neonates. n-3 Poly-unsaturated fatty acids have vasodilatory properties in the perinatal lung. We studied the circulatory effects of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in fetal sheep and in fetal pulmonary arterial rings. Methods: At 128 days of gestation, catheters were placed surgically in fetal systemic and pulmonary circulation, and a Doppler probe around the left pulmonary artery (LPA). Pulmonary arterial pressure and LPA flow were measured while infusing EPA or DHA for 120 min to the fetus, to compute pulmonary vascular resistance (PVR). The dose effects of EPA or DHA were studied in vascular rings pre-constricted with serotonin. Rings treated with EPA were separated into three groups: E+ (intact endothelium), E− (endothelium stripped) and LNA E+ (pretreatment of E+ rings with l-nitro-arginine). Results: EPA, but not DHA, induced a significant and prolonged 25% drop in PVR (n = 8, p < 0.001). Incubation of vascular rings with EPA (100 µM) caused a maximum relaxation of 60% in the E+ (n = 6), whereas vessel tone did not change in the E− (n = 6, p < 0.001). The vascular effects of EPA were significantly decreased in LNA E+ (n = 6). Incubation with DHA resulted in only a mild relaxation at the highest concentration of DHA (300 µM) compared to E+. Conclusions: EPA induces a sustained pulmonary vasodilatation in fetal lambs. This effect is endothelium- and dose-dependent and involves nitric oxide (NO) production. We speculate that EPA supplementation may improve pulmonary circulation in clinical conditions with PPHN.

Prenatal therapy with pyrimethamine + sulfadiazine vs spiramycin to reduce placental transmission of toxoplasmosis: a multicenter, randomized trial

BACKGROUND: The efficacy of prophylaxis to prevent prenatal toxoplasmosis transmission is controversial, without any previous randomized clinical trial. In France, spiramycin is usually prescribed for maternal seroconversions. A more potent pyrimethamine + sulfadiazine regimen is used to treat congenital toxoplasmosis and is offered in some countries as prophylaxis. OBJECTIVE: We sought to compare the efficacy and tolerance of pyrimethamine + sulfadiazine vs spiramycin to reduce placental transmission. STUDY DESIGN: This was a randomized, open‐label trial in 36 French centers, comparing pyrimethamine (50 mg qd) + sulfadiazine (1 g tid) with folinic acid vs spiramycin (1 g tid) following toxoplasmosis seroconversion. RESULTS: In all, 143 women were randomized from November 2010 through January 2014. An amniocentesis was later performed in 131 cases, with a positive Toxoplasma gondii polymerase chain reaction in 7/67 (10.4%) in the pyrimethamine + sulfadiazine group vs 13/64 (20.3%) in the spiramycin group. Cerebral ultrasound anomalies appeared in 0/73 fetuses in the pyrimethamine + sulfadiazine group, vs 6/70 in the spiramycin group (P = .01). Two of these pregnancies were terminated. Transmission rates, excluding 18 children with undefined status, were 12/65 in the pyrimethamine + sulfadiazine group (18.5%), vs 18/60 in the spiramycin group (30%, P = .147), equivalent to an odds ratio of 0.53 (95% confidence interval, 0.23–1.22) and which after adjustment tended to be stronger (P = .03 for interaction) when treatment started within 3 weeks of seroconversion (95% confidence interval, 0.00–1.63). Two women had severe rashes, both with pyrimethamine + sulfadiazine. CONCLUSION: There was a trend toward lower transmission with pyrimethamine + sulfadiazine, but it did not reach statistical significance, possibly for lack of statistical power because enrollment was discontinued. There were also no fetal cerebral toxoplasmosis lesions in the pyrimethamine + sulfadiazine group. These promising results encourage further research on chemoprophylaxis to prevent congenital toxoplasmosis.

Parasympathetic tone variations according to umbilical cord pH at birth: a computerized fetal heart rate variability analysis

Non-reassuring fetal heart rate tracings reflect an imbalance between the parasympathetic and sympathetic nervous systems. In this situation, fetal asphyxia can be suspected and may be confirmed by metabolic measurements at birth like low pH or high base deficit values. The objective of this study was to determine whether fetal asphyxia during labor is related to parasympathetic nervous system activity. This is a retrospective study of a database collected in 5 centers. Two hundred and ninety-nine fetal heart rate tracings collected during labor were analyzed. Autonomic nervous system, especially the parasympathetic nervous system, was analyzed using an original index: the FSI (Fetal Stress Index). The FSI is a parasympathetic activity evaluation based on fetal heart rate variability analysis. Infants were grouped based on normal or low pH value at birth. FSI was measured during the last 30xa0min of labor before birth and compared between groups. The minimum value of the FSI during the last 30xa0min before delivery was significantly lower in the group with the lower umbilical cord arterial pH value. In this pilot study during labor, FSI was lower in the group of infants with low arterial pH at birth.

Short-Term Variation of the Fetal Heart Rate for Predicting Neonatal Acidosis in Preeclampsia.

Introduction: The aim of this study was to measure the performance of short-term variation (STV) in predicting the onset of neonatal acidosis in fetuses at risk due to maternal preeclampsia. Material and Methods: This retrospective study examined data from a series of 159 women with singleton pregnancies, hospitalized for preeclampsia in a level 3 reference maternity hospital in northern France, with an STV measurement in the 24 h preceding cesarean delivery and a measurement of the newborns arterial cord pH at birth. The main outcome was determined by a correlation between STV and neonatal pH. Results: The last computerized fetal heart rate analysis took place a mean of 7.9 ± 6.3 h before birth, and neonatal acidosis was diagnosed in 38 newborns (23.9%). Although STV and umbilical artery pH at birth were significantly correlated (ρ = 0.16, p < 0.05), the performance of STV in predicting neonatal acidosis was poor, with an area under the ROC curve of 0.63. The sensitivity reached only 50.0% and the specificity 71.9% at the best STV threshold for predicting acidosis. Conclusion: The performance of STV for screening for neonatal acidosis is poor in women with preeclampsia. The divergent results between studies are probably due to the variable intervals between STV measurement and birth.

Suspected hemorrhagic ovarian cyst causing fetal anemia measured by the peak systolic velocity in the middle cerebral artery

We present two case reports of suspicion of hemorrhagic ovarian cyst resulting in fetal anemia. The main risk in acute fetal anemia is essentially cerebral. Only two cases of fetal anemia secondary to intra-cystic bleeding have been described in the literature. In one of the published cases, in utero transfusion was necessary because of severe anemia [1]. In our first patient, ultrasonography at 32 weeks demonstrated in a female fetus a strictly anechoic pelvic rounded image with fine inner walls, 39 mm 37 mm. The presumed diagnosis was an uncomplicated fetal ovarian cyst (FOC). This cyst was isolated, with no ascites or hydramnios. At 34 weeks, intra-cystic hemorrhage was suspected (Fig. 1A) as size of the cyst increased to