Timothy Overton

High failure rate of umbilical vessel occlusion by ultrasound-guided injection of absolute alcohol or enbucrilate gel

The success rate for injected umbilical vascular occlusion in the published literature exceeds 85 per cent. In this study we assessed the efficacy of two forms of injected sclerosants in achieving umbilical vessel occlusion. 12 cases of attempted ultrasound‐guided occlusion over a 2½ year period were reviewed. These were monochorionic (MC) twins (n=6), dichorionic twins (n=3) and singletons (n=3) undergoing fetocide for severe anomalies, or impending fetal demise. Absolute alcohol (n=6), enbucrilate gel (n=5) or both (n=1) were used in an attempt to achieve vascular occlusion. Complete vessel occlusion was achieved in only a third of cases (4/12), three with absolute alcohol and one with enbucrilate gel. In MC twins occlusion was successful in two of six cases. In contrast to previously published data, this large series, containing more cases than the total previously reported, shows considerably poorer success rates for injected umbilical vascular occlusion. Injection of currently available sclerosants can no longer be recommended for umbilical vascular occlusion in human fetuses. Copyright

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.

Detection of fetal messenger ribonucleic acid in maternal blood to determine fetal RhD status as a strategy for noninvasive prenatal diagnosis

OBJECTIVES Our purpose was to test the hypothesis that reverse transcriptase-polymerase chain reaction for fetal messenger ribonucleic acid in maternal blood is more sensitive than polymerase chain reaction from genomic deoxyribonucleic acid in prenatal determination of fetal RhD blood type. STUDY DESIGN Fetal nucleated erythrocytes in peripheral blood from 35 RhD-negative women were enriched by triple-density gradient centrifugation, anti-CD71 magnetic sorted, and deoxyribonucleic acid and ribonucleic acid extracted. Sensitivities of reverse transcriptase-polymerase chain reaction and polymerase chain reaction were compared to predict definitive fetal RhD blood type determined in fetal tissues. RESULTS Reverse transcriptase-polymerase chain reaction was significantly more accurate (P = .03) than genomic-polymerase chain reaction in predicting fetal RhD blood type, both overall (28 of 35 vs 22 of 35) and when the fetus was RhD-positive (12 of 19 vs 6 of 19). CONCLUSIONS Reverse transcriptase-polymerase chain reaction is more sensitive than genomic-polymerase chain reaction in detection of fetal RhD sequences in maternal blood.

Accuracy of Prenatal Determination of Rhd Type Status by Polymerase Chain-Reaction with Amniotic Cells

OBJECTIVE Our purpose was to determine the accuracy of RhD typing by use of amniocytes obtained at amniocentesis in RhD-negative women. STUDY DESIGN One hundred thirty-five RhD-negative women undergoing amniocentesis for management of suspected alloimmunization (n = 95) or routine second-trimester cytogenetic indications (n = 40) were studied. Amniocytes were then used as template to amplify specific portions of the Rh D and Rh CcEe genes by polymerase chain reaction. The fetal RhD type was confirmed by serologic techniques either after fetal blood sampling or cord blood samples at birth. RESULTS Thirty-six fetuses were serologically typed as RhD negative and all 36 were typed RhD negative by polymerase chain reaction. Ninety-eight fetuses were serologically typed as RhD positive; of these, 96 were correctly typed as RhD positive and two were incorrectly typed as RhD negative, with an overall error rate of 1.4%. Both of the errors occurred in a single batch of six samples tested at the same time. In one of these cases the fetus had mild Rh alloimmune disease and required exchange transfusion at birth. In the second case the fetus had severe hydrops fetalis and died in utero at 28 weeks. Deoxyribonucleic acid isolated from fetal blood was tested with the same polymerase chain reaction technique after delivery, and in both cases the fetus was correctly typed as RhD positive. Deoxyribonucleic acid amplification repeatedly failed in one case. CONCLUSION Prenatal fetal RhD typing by polymerase chain reaction with amniotic fluid cells is accurate and reliable. In sensitized pregnancies it allows early management of Rh disease and avoids invasive procedures in RhD-negative fetuses. In nonsensitized pregnancies it avoids the use of anti-RhD immunoglobulin after invasive procedures or during pregnancy. To eliminate the possibility of genetic and laboratory sources of errors, we suggest using different sets of primers at two different loci (e.g., exon 4 to 5 and exon 10).

Management and outcome of pregnancies with parvovirus B19 infection over seven years in a tertiary fetal medicine unit

Objectives: To determine rates of fetal anaemia and pregnancy outcome in susceptible pregnant women infected with human parvovirus B19 infection in a tertiary fetal medicine department over a 7-year period. Additional features enabling identification of fetuses that progress to severe anaemia were also investigated. Methods: Forty-seven susceptible, pregnant women with confirmed parvovirus infection referred to a regional fetal medicine unit, over a 7-year period (1999–2006), were identified. Where possible maternal serum AFP measurements were obtained from second-trimester serum screening and the presence or absence of echogenic bowel noted. Results: Of the 47 cases, one was excluded. Of the remaining 46 cases, 34 (74%) showed no signs of fetal anaemia and delivered at term. The remaining 12 (26%) showed signs of fetal anaemia. Eight of the 12 developed hydrops and underwent fetal blood sampling and transfusion (median pretransfusion Hb 3.6 g/dl). Seven of the 8 transfused fetuses were thrombocytopenic with a platelet count <150 × 109/l, with 2 fetuses having platelet counts <50 × 109/l. The median gestation age at transfusion was 22 weeks (range 18–27 weeks). The median number of weeks between seroconversion and transfusion was 6 (range 3–12). The signs of anaemia resolved after one transfusion in 5 of the 8 transfused fetuses and they subsequently delivered at term. There were 2 fetal deaths during or shortly after transfusion and one neonatal death following delivery at 28 weeks gestation due to severe pre-eclampsia, 5 days after successful transfusion. Conclusions: Following parvovirus seroconversion, the incidence of significant fetal anaemia requiring transfusion was 17%. Seroconversion after 21 weeks did not result in severe fetal anaemia. Significant anaemia requiring intervention did not occur 12 weeks after maternal seroconversion. We did not demonstrate a correlation with either maternal serum AFP or the presence of fetal echogenic bowel and the development of severe fetal anaemia. Because of the association between fetal anaemia and severe thrombocytopenia, it may be prudent to have compatible platelets available at the time of fetal blood sampling.

Prenatal Diagnosis by minimally invasive first-trimester transcervical sampling is unreliable

OBJECTIVE We investigated whether reliable prenatal diagnosis is possible from fetal cells harvested transcervically in first-trimester pregnancies. STUDY DESIGN Fetal cells were obtained transcervically from 87 women undergoing pregnancy termination. Fetal gender was determined in 51 pregnancies with three different polymerase chain reaction techniques and in 36 pregnancies with fluorescent in situ hybridization. In known male pregnancies the number of male fetal cells present was also determined. RESULTS Polymerase chain reaction detected male deoxyribonucleic acid in up to 79% of cases in male pregnancies and up to 45% of cases in female pregnancies. Fetal gender was correctly predicted in up to 72% of cases with fluorescent in situ hybridization. However, fetal cells were identified in < 40% of informative male pregnancies and were present in low numbers-0.7% to 3.4% in swabs and 4.4% to 24.8% in flushes. CONCLUSION The use of fetal cells obtained by minimally invasive first-trimester transcervical sampling is unreliable for prenatal diagnosis.

Neonatal outcomes of pregnancies affected by haemolytic disease of the foetus and newborn and managed with intrauterine transfusion: a service evaluation

BACKGROUND This study, conducted in the tertiary Foetal Medicine Unit at St Michaels Hospital, Bristol, was designed to obtain information regarding neonatal outcomes of pregnancies affected by haemolytic disease of the foetus and newborn and managed by intrauterine transfusion, and to determine whether a change in intrauterine transfusion protocol in 2004 had improved safety. The new protocol included attendance of two Foetal Medicine Unit consultants, foetal sedation and use of the intrahepatic vein as an alternative route to placental cord insertion if deemed safer. MATERIALS AND METHODS Data for pregnancies affected by haemolytic disease of the foetus and newborn as a result of haemolytic red cell alloimmunisation and managed with intrauterine transfusion at St Michaels Hospital between 1999 and 2009 were retrospectively collected using local databases, and medical note review. RESULTS Overall, 256 relevant intrauterine transfusions were performed. The median number of intrauterine transfusions per pregnancy was two. Ninety-three per cent of the live deliveries had 5-minute APGAR scores ≥9 and 98% were admitted to a Neonatal Intensive Care Unit/Special Care Baby Unit, requiring phototherapy (96%), top-up transfusions (44%: 23.2% immediate, 13.4% late, 7.3% both), and exchange transfusion (37%). An association was found between increased intrauterine transfusion number and reduced phototherapy duration and hospital admission: each additional intrauterine transfusion reduced the duration of phototherapy by 16% (95% CI: 0.72-0.98), and Neonatal Intensive Care Unit/Special Care Baby Unit admission by 44% (95% CI: 0.48-0.66). Following the change in intrauterine transfusion protocol, there was a significant reduction in the number of emergency Caesarean sections occurring directly after an intrauterine transfusion (n =5 vs 0; P =0.02). The foetal loss rate within 48 hours of an intrauterine transfusion was 1.9% per pregnancy, or 0.8% per intrauterine transfusion: no losses occurred under the new protocol (n =3 vs 0; P = NS). DISCUSSION Although the majority of neonates required admission to a Neonatal Intensive Care Unit/Special Care Baby Unit and phototherapy, the medium-term outcomes were positive. Importantly, the safety of the intrauterine transfusion procedure has improved significantly since the change in protocol.

Maternal contamination at fetal muscle biopsy

Duchenne muscular dystrophy (DMD) can be diagnosed by fetal muscle biopsy and immunohistochemical staining showing the absence of dystrophin. We report a case of fetal muscle biopsy in which the needle gun was successfully fired within the fetal gluteal muscle but the sample was contaminated by maternal tissue. This was attributed to the design of the biopsy needle, allowing transient opening of the biopsy core as the needle penetrated the maternal rectus sheath, muscle, and myometrium. Histology showed tissue suggestive of maternal origin, confirmed by DNA analysis. Repeat sampling revealed fetal muscle with normal dystrophin expression. We recommend that care be taken during needle insertion to avoid maternal contamination, and tests be used to confirm the fetal source of the sample.

Management of Fetal Hydrops due to Maternal Antibodies against a Low Incidence Paternal Red Cell Antigen: A Novel Insight from Clinical Practice

A rare case of a low incidence red cell antigen causing severe fetal allo-immune red cell disease is presented. Discussion of how this can be diagnosed and successfully managed antenatally using middle cerebral artery Doppler ultrasound and maternal antibody titre levels for fetal surveillance and timing of intervention with intrauterine transfusion.