Continued Validation of Ultrasound Guidance Targeting Tasks: Relationship with Procedure Performance.

RATIONALE AND OBJECTIVES: To determine if deliberative practice with novel ultrasound guidance targeting tasks improves simulated procedural skill. MATERIALS AND METHODS: In a nonrandomized interventional trial first year medical students practiced the previous described dowel and straw targeting tasks 1 hour a week for 4 weeks (training group) or had no training (controls). Afterward, they each performed a simulated amniocentesis (AMN) and chorionic villus sampling (CVS) procedure. Procedures were scored using a global rating scale (GRS) and compared between groups with Mann-Whitney U tests. Two-way random effects intraclass correlation coefficients for the inter- and intra-rater variability were calculated for each item in both GRS's. RESULTS: The training group (n = 22) had higher scores on several aspects and overall performance of AMN compared to controls (n = 15). There were no differences between groups for CVS. The inter-rater and intra-rater reliability of the GRS's for both AMN and CVS ranged from 0.16 to 0.89 with most values demonstrating good to excellent agreement. CONCLUSION: This study demonstrates validity evidence in the content and internal structure domains for the AMN and CVS simulators and their accompanying GRS's. Repetitive practice of the targeting tasks improved student performance in simulated AMN, but modifications are needed for it to be relevant to other procedures such as CVS.

Accurate fetal variant calling in the presence of maternal cell contamination.

High-throughput sequencing of fetal DNA is a promising and increasingly common method for the discovery of all (or all coding) genetic variants in the fetus, either as part of prenatal screening or diagnosis, or for genetic diagnosis of spontaneous abortions. In many cases, the fetal DNA (from chorionic villi, amniotic fluid, or abortive tissue) can be contaminated with maternal cells, resulting in the mixture of fetal and maternal DNA. This maternal cell contamination (MCC) undermines the assumption, made by traditional variant callers, that each allele in a heterozygous site is covered, on average, by 50% of the reads, and therefore can lead to erroneous genotype calls. We present a panel of methods for reducing the genotyping error in the presence of MCC. All methods start with the output of GATK HaplotypeCaller on the sequencing data for the (contaminated) fetal sample and both of its parents, and additionally rely on information about the MCC fraction (which itself is readily estimated from the high-throughput sequencing data). The first of these methods uses a Bayesian probabilistic model to correct the fetal genotype calls produced by MCC-unaware HaplotypeCaller. The other two methods "learn" the genotype-correction model from examples. We use simulated contaminated fetal data to train and test the models. Using the test sets, we show that all three methods lead to substantially improved accuracy when compared with the original MCC-unaware HaplotypeCaller calls. We then apply the best-performing method to three chorionic villus samples from spontaneously terminated pregnancies.

Amniocentesis and chorionic villus sampling for prenatal diagnosis.

BACKGROUND: During pregnancy, fetal cells suitable for genetic testing can be obtained from amniotic fluid by amniocentesis (AC), placental tissue by chorionic villus sampling (CVS), or fetal blood. A major disadvantage of second trimester amniocentesis is that the results are available relatively late in pregnancy (after 16 weeks' gestation). Earlier alternatives are chorionic villus sampling (CVS) and early amniocentesis, which can be performed in the first trimester of pregnancy. OBJECTIVES: The objective of this review was to compare the safety and accuracy of all types of AC (i.e. early and late) and CVS (e.g. transabdominal, transcervical) for prenatal diagnosis. SEARCH METHODS: We searched the Cochrane Pregnancy and Childbirth Group's Trials Register (3 March 2017), ClinicalTrials.gov, the WHO International Clinical Trials Registry Platform (ICTRP; 3 March 2017), and reference lists of retrieved studies. SELECTION CRITERIA: All randomised trials comparing AC and CVS by either transabdominal or transcervical route. DATA COLLECTION AND ANALYSIS: Two review authors independently assessed trials for inclusion and risk of bias, extracted data and checked them for accuracy. The quality of the evidence was assessed using the GRADE approach. MAIN RESULTS: We included a total of 16 randomised studies, with a total of 33,555 women, 14 of which were deemed to be at low risk of bias. The number of women included in the trials ranged from 223 to 4606.Studies were categorized into six comparisons: 1. second trimester AC versus control; 2. early versus second trimester AC; 3. CVS versus second trimester AC; 4. CVS methods; 5. Early AC versus CVS; and 6. AC with or without ultrasound.One study compared second trimester AC with no AC (control) in a low risk population (women = 4606). Background pregnancy loss was around 2%. Second trimester AC compared to no testing increased total pregnancy loss by another 1%. The confidence intervals (CI) around this excess risk were relatively large (3.2% versus 2.3 %, average risk ratio (RR) 1.41, 95% CI 0.99 to 2.00; moderate-quality evidence). In the same study, spontaneous miscarriages were also higher (2.1% versus 1.3%; average RR 1.60, 95% CI 1.02 to 2.52; high-quality evidence). The number of congenital anomalies was similar in both groups (2.0% versus 2.2%, average RR 0.93, 95% CI 0.62 to 1.39; moderate-quality evidence).One study (women = 4334) found that early amniocentesis was not a safe early alternative compared to second trimester amniocentesis because of increased total pregnancy losses (7.6% versus 5.9%; average RR 1.29, 95% CI 1.03 to 1.61; high-quality evidence), spontaneous miscarriages (3.6% versus 2.5%, average RR 1.41, 95% CI 1.00 to 1.98; moderate-quality evidence), and a higher incidence of congential anomalies, including talipes (4.7% versus 2.7%; average RR 1.73, 95% CI 1.26 to 2.38; high-quality evidence).When pregnancy loss after CVS was compared with second trimester AC, there was a clinically significant heterogeneity in the size and direction of the effect depending on the technique used (transabdominal or transcervical), therefore, the results were not pooled. Only one study compared transabdominal CVS with second trimester AC (women = 2234). They found no clear difference between the two procedures in the total pregnancy loss (6.3% versus 7%; average RR 0.90, 95% CI 0.66 to 1.23, low-quality evidence), spontaneous miscarriages (3.0% versus 3.9%; average RR 0.77, 95% CI 0.49 to 1.21; low-quality evidence), and perinatal deaths (0.7% versus 0.6%; average RR 1.18, 95% CI 0.40 to 3.51; low-quality evidence). Transcervical CVS may carry a higher risk of pregnancy loss (14.5% versus 11.5%; average RR 1.40, 95% CI 1.09 to 1.81), but the results were quite heterogeneous.Five studies compared transabdominal and transcervical CVS (women = 7978). There were no clear differences between the two methods in pregnancy losses (average RR 1.16, 95% CI 0.81 to 1.65; very low-quality evidence), spontaneous miscarriages (average RR 1.68, 95% CI 0.79 to 3.58; very low-quality evidence), or anomalies (average RR 0.68, 95% CI 0.41 to 1.12; low-quality evidence). We downgraded the quality of the evidence to low due to heterogeneity between studies. Transcervical CVS may be more technically demanding than transabdominal CVS, with more failures to obtain sample (2.0% versus 1.1%; average RR 1.79, 95% CI 1.13 to 2.82, moderate-quality evidence).Overall, we found low-quality evidence for outcomes when early amniocentesis was compared to transabdominal CVS. Spontaneous miscarriage was the only outcome supported by moderate-quality evidence, resulting in more miscarriages after early AC compared with transabdominal CVS (2.3% versus 1.3%; average RR 1.73, 95% CI 1.15 to 2.60). There were no clear differences in pregnancy losses (average RR 1.15, 95% CI 0.86 to 1.54; low-quality evidence), or anomalies (average RR 1.14, 95% CI 0.57 to 2.30; very low-quality evidence).We found one study that examined AC with or without ultrasound, which evaluated a type of ultrasound-assisted procedure that is now considered obsolete. AUTHORS' CONCLUSIONS: Second trimester amniocentesis increased the risk of pregnancy loss, but it was not possible to quantify this increase precisely from only one study, carried out more than 30 years ago.Early amniocentesis was not as safe as second trimester amniocentesis, illustrated by increased pregnancy loss and congenital anomalies (talipes). Transcervical chorionic villus sampling compared with second trimester amniocentesis may be associated with a higher risk of pregnancy loss, but results were quite heterogeneous.Diagnostic accuracy of different methods could not be assessed adequately because of incomplete karyotype data in most studies.

Early pregnancy failure as a training tool for chorionic villus sampling.

OBJECTIVES: The goal of this study is to evaluate the success of a training program in chorionic villus sampling (CVS) of early pregnancy failure (EPF) for maternal-fetal medicine (MFM) fellows. METHODS: We conducted a retrospective review of a new training program in CVS for MFM fellows. Women with EPF up to 13 weeks estimated gestational age were offered inclusion in our program and counseled on alternatives. Transcervical CVS was performed for both fellow education and cytogenetic diagnosis. The primary outcome was a successful diagnosis, and the secondary outcome was cell growth by fellow experience. RESULTS: Thirty-nine patients diagnosed with EPF from December 2011 to March 2013 underwent CVS. Villi obtained via CVS yielded a diagnosis in 62% of cases. CVS samples with successful karyotype had more villi but were otherwise similar. CVS by experience showed a trend towards increased success and villi volume after the first five procedures. Abnormal results were obtained in 72% of cases, 15% of which led to changes in care. CONCLUSION: CVS of EPF is an option for the training of MFM fellows as it was acceptable to most patients, and the majority of CVS specimens yielded a karyotype diagnosis.

[Maternal cell contamination of prenatal samples and the potential effects on prenatal diagnosis results].

OBJECTIVE: To assess the frequency and significance of maternal cell contamination (MCC) in the invasive prenatal diagnosis, and to analysis the MCC effect on prenatal diagnosis results. METHODS: Totally 519 amniotic fluid samples from second trimester pregnancy, 57 chorionic villus samples from first trimester pregnancy, and 576 blood samples from corresponded pregnant women were collected and genotyped by Promega PowerPlex 16 system. MCC was determined according to the genotyping results. Karyotypic and molecular diagnosis results were contrasted between MCC and non-MCC specimen of the same fetal. RESULTS: MCC presented in 3.1% (16/519) uncultured amniotic fluid, 1.3% (7/519) cultured amniotic fluid and 5% (3/57) villi specimens. In the study of fetal karyotype, MCC had no significant effect on normal female fetus; but for male fetus and abnormal female fetus, there were risk of erroneous results of mosaics. As to molecular diagnosis, MCC resulted in more complex effects for the different diagnostic methods. And 10%MCC had led to misdiagnosis. CONCLUSIONS: For the prenatal cytogenetic tests, MCC should be excluded when there were mosaicism karyotype results or suspicious MCC of chorionic villi samples. The effects of MCC had more seriously impact on prenatal molecular testing, which suggesting the recommend regular identity test for MCC should be carried out.

Analysis of fetal and maternal results from fetal genetic invasive procedures: an exploratory study at a University Hospital.

OBJECTIVE: To characterize the indications of pregnant women who sought the Fetal Medicine Services of the Hospital das Clínicas, at the Medical School of the Universidade de São Paulo for performing invasive diagnostic procedures, and to evaluate the results of fetal karyotypes and their pregnancies. METHODS: A retrospective and observational study on pregnant women who underwent chorionic villus sampling (CVS), amniocentesis, and cordocentesis in the period from February, 2005 to December, 2009. Other diagnostic or therapeutic procedures were not included. The result of pregnancy was obtained by consulting patient electronic records, medical records, and/or telephone call. RESULTS: 713 procedures were performed (113 CVS, 340 amniocenteses, and 260 cordocenteses). The main indication for performing invasive procedures was the presence of structural changes in fetuses, followed by increased values of nuchal translucency, and advanced maternal age. Fetal karyotype was altered in 186 cases (26.1%). The 18 trisomy was the commonest aneuploidy followed by the 21 trisomy, X monosomy, and 13 trisomy. There were 4.9% cases of miscarriage, 25.7% cases of stillborn infants, and 13% cases of neonatal deaths. Eight pregnant women opted for legally induced abortion. 99% of pregnant women whose fetuses did not present abnormalities and presented normal fetal karyotype had infants who were born alive.

Análise dos resultados maternos e fetais dos procedimentos invasivos genéticos fetais: um estudo exploratório em Hospital Universitário / Analysis of fetal and maternal results from fetal genetic invasive procedures: an exploratory study at a University Hospital

OBJETIVO: Caracterizar as indicações das gestantes que procuraram o serviço de Medicina Fetal do Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo para realização de procedimentos invasivos diagnósticos e avaliar os resultados dos cariótipos fetais e de suas gestações. MÉTODOS: Estudo observacional retrospectivo das gestantes que realizaram biópsia de vilo corial (BVC), amniocentese e cordocentese no período de fevereiro de 2005 a dezembro de 2009. Não foram incluídos outros procedimentos diagnósticos ou procedimentos terapêuticos. O resultado da gestação foi obtido através de consulta de prontuário eletrônico e/ou físico e/ou contato telefônico. RESULTADOS: Foram realizados 713 procedimentos (113 BVC, 340 amniocenteses e 260 cordocenteses). A principal indicação para a realização dos procedimentos invasivos foi a presença de alterações estruturais nos fetos, seguido por valores aumentados da translucência nucal e pela idade materna avançada. O cariótipo fetal esteve alterado em 186 casos (26,1%). A trissomia do cromossomo 18 foi a aneuploidia mais comum, seguida pela trissomia do 21, a monossomia do X e a trissomia do cromossomo 13. Ocorreram 4,9% de abortamento, 25,7% de natimortos e 13% de neomortos. Oito gestantes optaram pela interrupção judicial, e 99% das gestantes cujos fetos não apresentavam malformação e que apresentavam cariótipo fetal normal tiveram nativivos.

OBJECTIVE: To characterize the indications of pregnant women who sought the Fetal Medicine Services of the Hospital das Clínicas, at the Medical School of the Universidade de São Paulo for performing invasive diagnostic procedures, and to evaluate the results of fetal karyotypes and their pregnancies. METHODS: A retrospective and observational study on pregnant women who underwent chorionic villus sampling (CVS), amniocentesis, and cordocentesis in the period from February, 2005 to December, 2009. Other diagnostic or therapeutic procedures were not included. The result of pregnancy was obtained by consulting patient electronic records, medical records, and/or telephone call. RESULTS: 713 procedures were performed (113 CVS, 340 amniocenteses, and 260 cordocenteses). The main indication for performing invasive procedures was the presence of structural changes in fetuses, followed by increased values of nuchal translucency, and advanced maternal age. Fetal karyotype was altered in 186 cases (26.1%). The 18 trisomy was the commonest aneuploidy followed by the 21 trisomy, X monosomy, and 13 trisomy. There were 4.9% cases of miscarriage, 25.7% cases of stillborn infants, and 13% cases of neonatal deaths. Eight pregnant women opted for legally induced abortion. 99% of pregnant women whose fetuses did not present abnormalities and presented normal fetal karyotype had infants who were born alive.

10 años de experiencia en diagnóstico prenatal invasivo en el Instituto Dexeus / Invasive prenatal diagnostic practice: a review of a ten years experience at Dexeus Institut

Objetivos. Describir nuestra experiencia en la realización y resultados de las técnicas invasivas (TI) de diagnóstico prenatal (DP). Métodos. Se trata de un estudio descriptivo retrospectivo de las pacientes sometidas a una TI de DP desde enero de 1999 a diciembre de 2008, incluyendo todos los casos consecutivos de biopsia corial (BC) y amniocentesis (AC) genéticas. Resultados. Hemos analizado un total de 9.340 TI (8.928 AC y 412 BC). La edad gestacional ha sido de 15,9 (rango 10-38) y 11,6 (rango 9-16) semanas, respectivamente. Las principales indicaciones de referencia son la edad materna avanzada y la ansiedad. El porcentaje de cultivos no informativos ha sido de 0,49% para BC y 0,1% para AC. Se han diagnosticado un total de 380 anomalías cromosómicas (4,1%), 269 clínicamente relevantes. El valor predictivo positivo (VPP) para cromosomopatía relevante es del 2,9% (2,14% en la serie de AC y 19,6% en la serie de BC). El mayor VPP se obtiene ante las indicaciones de referencia ecográficas. Se han recogido un total de 88 complicaciones atribuibles a la TI (0,94%), con una tasa de pérdida fetal pos-procedimiento del 0,6%. La tasa de pérdida gestacional es del 0,9% (0,8% en la serie de AC y 4,3% en BC). Cuando analizamos los aspectos relacionados con la tasa de complicaciones y pérdidas fetales pos-procedimiento, únicamente el n.° de fetos, aspecto del LA, la indicación de la TI y la experiencia del operador son factores significativos. Conclusiones. El seguimiento y monitorización de las TI de DP constituye un primer paso en el proceso de control de calidad, recientemente introducido en la práctica médica en el área del diagnóstico prenatal(AU)

Objective. To describe invasive prenatal diagnostic practice in a single-center over a 10-years period. Methods. Included in this study were all consecutive pregnancies with genetic amniocentesis (AC) or chorionic villous sampling (CVS) procedure done during the period January 1999-December 2008. Results. A total of 8928 AC and 412 CVS were performed. Main indications for referral were increased maternal age and anxiety. The mean gestational age in which the procedure was done was 15,9 weeks (range 10-38) in AC and 11,6 weeks (range 9-16) in CVS. In this series, 380 cases of chromosome abnormalities (CA) were detectable by conventional cytogenetic analysis (4,1%), 269 of them considered clinically significant. The positive predictive value (PPV) for significant CA was 2,9%, 2,1% and 19,6% in the overall, AC and CVS group, respectively. The highest PPV were obtained for sonographic referral indications. A total of 88 complications during the 4 weeks’ period after the procedure were register (0,94%). The post procedural loss rate was 0,6% (0,4% after AC and 4,1% after CVS), 0,3% in singles and 1,6% in twin pregnancies AC group. The overall loss rate was 0,9% (0,8% in AC group, 4,3% in CVS group). There was a significant correlation between post procedure complications and number of fetuses, technical aspects (dark amniotic fluid), indications for referral and operator's experience. Conclusions. Monitoring of prenatal diagnostic invasive tests is the first step in the process of quality control, recently introduced into medical practice in the area of prenatal diagnosis(AU)

Reacción en cadena de la polimerasa cuantitativa y cultivo largo: ¿el mejor método para el estudio citogenético de muestras de vellosidad corial? / Quantitative Polymerase Chain Reaction and long term culture: the best method for cytogenetic study of chorionic villus sampling?

El diagnóstico prenatal citogenético durante el primer trimestre de gestación se realiza a partir de biopsias de vellosidad corial. Para la obtención de metafases se utilizan dos métodos: el cultivo corto o semidirecto (STC) y cultivo largo (LTC). La principal ventaja del STC es que no presenta contaminación materna y la del LTC es que no hay descritos en la literatura falsos negativos. Se considera que la combinación de las dos técnicas (STC y LTC) es la estrategia diagnóstica más eficaz para este tipo de estudios. La técnica de PCR cuantitativa fluorescente (QF-PCR) permite evaluar las aneuploidías más frecuentemente implicadas en el diagnóstico prenatal en 24-48 horas en muestras de vellosidad corial. El objetivo de este trabajo es evaluar la combinación de QF-PCR y LTC como sustituto de las clásicas STC y LTC para el diagnóstico prenatal en muestras de vellosidad corial. Para ello presentamos nuestra experiencia en 900 muestras de vellosidad corial(AU)

First trimester cytogenetic prenatal diagnosis is performed on chorionic villus biopsies. Two methods are used to obtain metaphases: the short-term or semi-direct culture (STC) and long term culture (LTC). The main advantage of STC is that there is no risk of maternal contamination, and of LTC that no false-negative findings are described in the literature. It is considered that the combination of the two techniques (STC and LTC) is the most effective diagnostic strategy for this type of study. The technique of quantitative fluorescent PCR (QF-PCR) allows the evaluation of aneuploidy most frequently involved in prenatal diagnosis in 24-48 hours in chorionic villus samples. The aim of this study is to evaluate the combination of QF-PCR and LTC as a substitute for classical STC and LTC for prenatal diagnosis in chorionic villus samples. We present our experience in 900 chorionic villus samples(AU)

[Feasibility of chorionic villous sampling outside a prenatal diagnosis center]. / Faisabilité de la biopsie de trophoblaste en maternité de niveau 2 hors centre de référence de diagnostic prénatal.

OBJECTIVES: According to new recommendations, a high combined risk for Down syndrome in the first trimester of pregnancy must indicate the need for a prenatal diagnosis. This is possible thanks to chorionic villous sampling. The objective of our study was to show that chorionic villous sampling is achievable in everyday practice, even outside research centers for pre-natal diagnosis. PATIENTS AND METHODS: It was a descriptive, retrospective study. All the patients who underwent a chorionic villous sampling in our level II maternity center from November 2005 to September 2009 were included. Success and complications rates linked with the procedure were calculated. RESULTS: One hundred and fourteen pregnancies were included. A definitive diagnosis was given in 98.25% of cases. A secondary amniocentesis was necessary in 1.75% of cases. A medical termination of the pregnancy was done in 18.42% of cases. Without accounting for underlying pathology, fetal loss rate was up to 5.75%. Only one case of unexpected fetal loss was noted (1.15% of the ongoing pregnancies). CONCLUSION: Our study shows that the presence of trained professional allows for onsite performance chorionic villous sampling.

The role of molecular microsatellite identity testing to detect sampling errors in prenatal diagnosis.

OBJECTIVE: The objective was to determine the risk of sampling error in amniocentesis and chorionic villus sampling (CVS) in singleton and multiple pregnancies. Data from this and other published studies were used to discuss current practice guidelines for molecular identity testing. METHOD: Clinical and laboratory records of all patients undergoing molecular-based identity testing in our clinical laboratory from July 2002 until March 2008 were reviewed. DNA microsatellite testing was performed to determine zygosity in multiple pregnancies and maternal cell contamination (MCC) in both singleton and multiple pregnancies. RESULTS: MCC was detected in 6/148 (4%) CVS and 1/87 (1%) amniotic fluids from singleton pregnancies. In two of the CVS, only maternal cells were found. In 2/24 (8%) twin pregnancies, the same fetus was tested twice. In a total of 285 pregnancies (235 singleton, 24 twin, 26 with >or= 3 fetuses), without molecular identity testing, four women would have received erroneous results. CONCLUSION: Current guidelines recommend molecular identity testing for MCC in conjunction with molecular diagnostic testing, but not for cytogenetic testing. No published guidelines were found for zygosity testing in multiple pregnancies. We suggest that identity testing be considered for all prenatal testing of multiple pregnancies, especially if CVS is performed.