Search

Introduction: Haemophilias are X-linked inherited bleeding disorders, due to de novo F8/F9 gene variants in 30-50% of cases. The identification of causative variant in index cases (IC) is crucial for genetic counselling in related women. Over the last 20 years the Emilia-Romagna Regional Haemophilia Registry documented high proportions of sporadic severe haemophilia., Aim: To clarify if carriers' reproductive choices influence the sporadic/familial ratio of severe haemophilia., Methods: Genetic counselling and genotyping in 221 relatives of severe IC were retrospectively reviewed, retrieving reproductive choices and pregnancy history of childbearing-age carriers from familial and sporadic pedigrees and according to the IC degree of relationship (mothers, daughters, II/other)., Results: Carriers' identification rates were lower in sporadic women and in other-degree relatives. Among childbearing age women (n = 140), carriers were 37/48 (77%) and 57/92 (62%) of familial and sporadic relatives, respectively. Forty-five/57 sporadic carriers experienced 67 pregnancies, while 21/37 familial carriers had 39 pregnancies (four voluntary terminations), with a significantly higher number of affected sons in the former (40/67 vs. 12/35, P = .025). Prenatal diagnosis was chosen by 40% and 47% of sporadic and familial aware carriers, respectively. Sporadic mothers often avoided further pregnancies (17/38, 45%) after a firstborn affected child, while familial mothers tended to face pregnancies without prenatal approaches (6/10, 60%)., Conclusion: In this cohort sporadic offspring account for more than 70% of severe haemophilia cases. This increasing proportion is likely to reflect the influence in reproductive choices of awareness of carriers' status, particularly in sporadic mothers, and of prenatal diagnosis options., (© 2022 John Wiley & Sons Ltd.)

BACKGROUND In pregnant women with advanced maternal age (AMA) and fetuses with ultrasonographic (USG) soft markers it is always challenging to decide whether to implement chromosomal microarray analysis (CMA) or not. It is unclear whether CMA should be used in the fetuses with isolated USG soft markers, and there is still a lack of extensive sample research. MATERIAL AND METHODS We enrolled 1521 cases in our research and divided them into 3 groups as follows: pregnant women with isolated AMA (group 1, n=633), pregnant women whose fetuses had isolated USG soft markers (group 2, n=750), and pregnant women with AMA whose fetuses had isolated USG soft markers (group 3, n=138). All pregnant women underwent prenatal ultrasound and amniocentesis, and fetal cells in the amniotic fluid were used for genetic analysis of CMA. All participants signed a written informed consent prior to CMA. RESULTS Abnormal findings were detected by CMA in 330 (21.70%) fetuses, including 37 (2.43%) clinically significant copy number variations (CNVs), 52 (3.42%) benign or likely benign CNVs, and 240 (15.78%) variants of unknown significance. The frequency of clinically significant CNVs in group 1 and group 2 were significantly lower than that in group 3 (2.37% and 2.0% vs 5.07%, P<0.01). More than a half (59.46%, 22/37) of the pregnant women decided to continue their pregnancy despite having a fetus diagnosed with clinically significant CNV. CONCLUSIONS CMA can increase the diagnostic yield of fetal chromosomal abnormality for pregnant women with isolated AMA or/and their fetuses had isolated USG soft markers.

Prenatal genetic testing and analysis in the past was usually only offered when a particular fetal phenotype was noted or suspected, meaning that filtering and interpretation of genetic variants identified could be anchored in attempts to explain an existing health concern. Advanced genomic testing is now increasingly used in "low-risk" pregnancies, producing information on genotype adrift of the phenotypic data that is necessary to give it meaning, thus increasing the difficulty in predicting whether and how particular genetic variants might affect future development and health. A challenge to healthcare scientists, clinicians, and parents therefore is deciding what qualities prenatal genotypic variation should have in order to be constructed as a 'result.' At the same time, such tests are often re requested in order to make binary decisions about whether to continue a pregnancy or not. As a range of professional organizations develop guidelines on the use of advanced genomic testing during pregnancy, we highlight the particular difficulties of discovering ambiguous findings such as variants with uncertain clinical significance, susceptibility loci for neurodevelopmental problems and susceptibility to adult-onset diseases. We aim to foster international discussions about how decisions around disclosure are made and how uncertainty is communicated.

Introduction: Prenatal anxiety is a prevalent condition that is harmful for women and a strong predictor of postpartum depression. This trial assesses an intervention initiated in early pregnancy to mid pregnancy among women with clinical or subclinical symptoms of anxiety in Pakistan. METHODS AND ANALYSIS: Happy Mother, Healthy Baby (HMHB) is a phase three, two-arm, single-blind, individual randomised clinical trial conducted in the outpatient department of Holy Family Hospital, a large public tertiary care facility affiliated with Rawalpindi Medical University (RMU). Pregnant women (enrolled at ≤22 weeks of gestation) receive six individual HMHB sessions based on cognitive-behavioral therapy (CBT) and relaxation techniques that are administered by non-specialist providers and tailored to address anxiety symptoms. Two to six booster sessions are given between the fifth consecutive weekly core session and the sixth core session that occurs in the third trimester. Apart from baseline data, data are collected in the third trimester, at birth and at 6-weeks postpartum. Primary outcomes include diagnoses of postpartum common mental disorders. Secondary outcomes include symptoms of anxiety and of depression, and birth outcomes including small-for-gestational age, low birth weight and preterm birth. An economic analysis will determine the cost effectiveness of the intervention., Ethics: Ethics approval was obtained from the Johns Hopkins Bloomberg School of Health Institutional Review Board (Baltimore, USA), the Human Development Research Foundation Ethics Committee (Islamabad, Pakistan), the RMU Institutional Research Forum (Rawalpindi, Pakistan) and the National Institute of Mental Health-appointed Global Mental Health Data Safety and Monitoring Board., Dissemination: Results from this trial will build evidence for the efficacy of a CBT-based intervention for pregnant women delivered by non-specialised providers. Identification of an evidence-based intervention for anxiety starting in early pregnancy to mid pregnancy may be transferable for use and scale-up in other low-income and middle-income countries., Trial Registration Number: NCT03880032., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Objective: To assess inter-manufacturer automated immunoassays for soluble FMS-like tyrosine kinase 1 (sFlt-1) to placental growth factor (PlGF)., Methods: sFlt-1 and PlGF levels were measured using the AutoDelfia PlGF1-2-3 (PerkinElmer Inc. Turku, Finland), BRAHMS Kryptor sFlt-1, PlGF plus and PlGF-2 (BRAHMS ThermoFisher, Germany) and Cobas e411 Elecsys® sFlt-1 and PLGF (Roche Diagnostics GmbH, Mannheim, Germany) in 965 asymptomatic pregnancies between 20 and 39 weeks of gestation and in in-vitro samples with predefined levels of glycosylated PlGF isomers (1, 2 and 3), sFlt-1 in human male serum. Percentage PlGF isoform recovery and cross-reactivities were determined. Paired Bland-Altman and Passing-Bablok analyses were performed to determine bias, precision and accuracy. Inter-manufacturer sFlt-1:PlGF ratio were compared., Results: PlGF-1 isomer recovery ranged from 36 to 39% for Elecsys® to 52-60% for PlGF plus and PlGF-1-2-3 assays. PlGF-2 and PlGF-3 isoform cross-reactivity was assay dependent, ranging from 10 to 21% and 16-36% respectively. BRAHMS PlGF-2 assay had high cross-reactivity to PlGF-1 (37-41%) and PlGF-3 isomers (48-65%). Elecsys® recovery of sFlt-1 was 13% vs 6% for BRAHMS. Passing-Bablok indicated significant proportional and systematic differences between all paired PlGF assay comparisons. PlGF Bland-Altman percentage biases ranged from 12 to 37% for PlGF and 18% for sFlt-1. A linear relationship existed between log transformed sFlt-1:PlGF ratios. The clinical equivalent of the BRAHMS sFlt-1:PlGF plus to the Elecsys® sFlt-1:PlGF ratios of 38 and 110 are 55 and 188 respectively., Conclusion: Inter-manufacture immunoassay differences are significantly different. sFlt-1:PlGF rule in/rule out criteria are manufacturer specific, not interchangeable and require separate clinical validation., (Copyright © 2019 International Society for the Study of Hypertension in Pregnancy. Published by Elsevier B.V. All rights reserved.)

Maternal prenatal symptoms of depression and anxiety have been suggested to impose differential effects on later offspring development, depending on their characteristics, such as timing, intensity and persistence. Paternal symptoms have been less investigated. While knowledge on these trajectory characteristics is essential for improved comprehension of prenatal stress, prospective studies including both expecting parents have been scarce. We aim at identifying and comparing the trajectories of prenatal depressive and anxiety symptoms in both parents in a pregnancy cohort design. The sample included 3202 mothers and 2076 fathers who were recruited to the FinnBrain Birth Cohort study (www.finnbrain.fi). Depressive symptoms were assessed using the Edinburgh Postnatal Depression Scale (EPDS) and general anxiety by the anxiety scale of the Symptom Checklist -90 (SCL-90) repeatedly at 14, 24, and 34 gestational weeks. Five differential depressive and four anxiety symptom trajectories were identified across pregnancy both in mothers and in fathers. The trajectories of consistently low depressive or anxiety symptoms were associated with higher educational level in both parents, and with nulliparity and non-smoking during pregnancy in mothers. Parents with consistently high or increasing levels of symptoms had more often prenatal SSRI medication. The congruences between elevated depressive and anxiety symptoms at any point in pregnancy, as well as parental trajectories within families were low. However, in this population-based sample, the self-reported symptom levels of both parents were generally very low. Variance in timing and persistence of parent-reported prenatal depressive and anxiety symptoms is potentially important, while symptom trajectories are very similar in mothers and fathers. These differential symptom trajectories and the significance of their correlates should be acknowledged when studying prenatal stress exposures and the related outcomes in children., Competing Interests: The authors have declared that no competing interests exist.

Objective: To determine whether abnormal levels of first-trimester maternal serum free β-hCG and PAPP-A are associated with significant copy number variants (CNVs) on chromosomal microarray analysis (CMA)., Methods: Retrospective cohort of singleton prenatal CMA studies (n = 2880). Cases with an abnormal karyotype, benign familial or de novo variants, and absence of heterozygosity were excluded. The prevalence of abnormal serum analytes was compared between patients with significant CNVs (n = 56) and those with normal CMA (n = 884). Odds ratios (ORs) and 95% confidence intervals (CI) were calculated using Fisher's exact test. Mantel-Haenszel method was utilized to adjust ORs for prenatal diagnostic procedure type and indications for testing. Statistical significance was determined as P value < 0.05., Results: Abnormally low serum free β-hCG (≤0.45 MoM) was associated with an increased risk of significant CNVs (OR 3.53, 95% CI, 1.25-8.66, P < 0.01). This association remained significant after adjusting for abnormal nuchal translucency and advanced maternal age (AMA) (adjusted OR 3.04, 95% CI, 1.05-7.48, P < 0.05) or procedure type and AMA (adjusted OR 3.21, 95% CI 1.13-8.16, P < 0.05). The associations of abnormally high serum free β-hCG, low PAPP-A, and high PAPP-A with significant CNVs were not statistically significant., Conclusion: Low first-trimester serum β-hCG is associated with an increased risk of significant CNVs on CMA., (© 2018 John Wiley & Sons, Ltd.)

Background: In New Zealand, haemoglobin A1c measurements are routinely offered at booking, preferably before 20 weeks gestation, to detect pre-existing hyperglycaemia. A haemoglobin A1c <5.9% (41 mmol/mol) is considered normal based on the reference range for the non-pregnant population., Aims: To determine pregnancy-specific haemoglobin A1c centiles by gestation and ethnicity., Materials and Methods: This is a population-based observational study of pregnancies uncomplicated by diabetes (pre-existing or gestational) with ≥1 haemoglobin A1c measurement. Haemoglobin A1c centiles were calculated from data extracted from electronic laboratory and clinical records for pregnancies during 2008-2010., Results: Included were 6800 pregnancies, European 80% (5462), Māori 6% (415), Pacific Islander 3% (196) and 11% (727) 'Others' (mostly Asian). Haemoglobin A1c levels fell with increasing gestation, reaching a nadir at 24 weeks, a trend verified by longitudinal data from 112 women. The 97.5th centile for haemoglobin A1c in European women was 5.76% (39.5 mmol/mol) at 8 +0  weeks, 5.70% (38.8 mmol/mol) at 16 +0  weeks, and 5.65% (38.3 mmol/mol) at 24 +0  weeks. Non-European women had both higher plasma glucose levels (although within the range considered normal) and higher mean haemoglobin A1c levels compared with Europeans; mean (SD) difference in haemoglobin A1c in Māori +0.13% (0.05) (+1.4 mmol/mol (0.5)), Pacific +0.20% (0.03) (+2.2 mmol/mol (0.3)), 'Others' +0.10% (0.03) (+1.1 mmol/mol (0.3))., Conclusions: The New Zealand haemoglobin A1c cut-point ≥5.9% (41 mmol/mol) for identifying hyperglycaemia in early pregnancy is greater than the 97.5th centile in European and 'Other' women. Utilising population haemoglobin A1c centiles adjusted by gestation may thus better guide management decisions., (© 2017 The Royal Australian and New Zealand College of Obstetricians and Gynaecologists.)

Objectives: To assess whether serum concentrations of saturated (SFAs), polyunsaturated (PUFAs), and monounsaturated (MUFAs) fatty acids are associated with changes in blood pressure (BP) throughout pregnancy., Study Design: Prospective cohort., Main Outcome Measures: Longitudinal measurements of systolic (SBP) and diastolic (DBP) BP., Methods: Two hundred twenty-three healthy pregnant women were recruited in a public health center in Rio de Janeiro, Brazil between 2009 and 2011. Fasting blood samples and BP measurements were obtained at the 1st (5th-13th weeks), 2nd (20 th -26th) and 3rd trimester (30th-36th). Crude and adjusted (maternal age, education, energy intake, gestational body weight change, leptin concentrations, early pre-pregnancy BMI, leisure time physical activity prior to pregnancy and linear and quadratic gestational weeks) longitudinal linear mixed-effects models were employed., Results: SBP and DBP decreased from the 1st to the 2nd trimester and slightly increased from the 2nd to the 3rd trimester (P < 0.001). In the adjusted model (ß and 95% CI), total SFAs [0.005 (0.001-0.008); P = 0.008], total MUFAs [0.005 (0.001-0.009); P = 0.019] and total n-6 PUFAs [0.005 (0.001-0.009); P = 0.025] were positively associated with SBP throughout pregnancy., Conclusions: Maternal serum concentrations of total SFAs, MUFAs and n-6 PUFAs were positively associated with BP levels in normotensive pregnant women., (Copyright © 2018 International Society for the Study of Hypertension in Pregnancy. Published by Elsevier B.V. All rights reserved.)

Introduction: Balanced information before prenatal diagnosis (PND) aims to help expectant parents to make an informed choice. However, it is important that the information does not increase the expectant parents' psychological distress. The aim was to examine psychological distress among expectant parents, before and after receiving information about PND, to evaluate the possible differences between two different procedures of information giving, and to evaluate the association between satisfaction with the information and psychological distress., Material and Methods: A longitudinal design, based on questionnaire data from 380 expectant parents from four counties in Sweden. The measurement points; T1, before the information about PND was given and T2, 2 weeks after the prenatal screening or 15 weeks of gestation. The Hospital Anxiety and Depression Scale (HADS) and the Swedish version of the Cambridge Worrying Scale (CWS) measured psychological distress. The Satisfaction with Genetic Counseling Scale (SCS) measured satisfaction with information about PND., Results: The rate of psychological distress was stable among the pregnant women, but decreased among their partners, after the information was received. General anxiety and the social-medical dimension of pregnancy-related worry decreased among the participants who received information, using the more distinct two-stage process (group A), but was unchanged in group B (less distinct two-stage process). Health-related worry decreased in both groups, whereas relational worry and level of depressive symptoms were unchanged in both groups., Conclusion: Information about PND does not increase the psychological distress among expectant parents. A more distinct two-stage process of information giving might even decrease their anxiety., (© 2017 Nordic Federation of Societies of Obstetrics and Gynecology.)

Objectives To examine associations between depression and preterm birth and small-for gestational age (SGA) among women of predominantly Puerto Rican descent, a population who experiences disparities in adverse birth outcomes and one of the highest infant mortality rates in the United States. Methods Proyecto Buena Salud (PBS) was a prospective cohort study conducted from 2006 to 2011 at a large tertiary care center in Western Massachusetts. Caribbean Islander (i.e., Puerto Rican and Dominican Republic) women were interviewed in early, mid and late pregnancy. Among 1262 participants, associations between depression, assessed using the Edinburgh Postnatal Depression Scale, and risk of preterm birth and small-for-gestational age (SGA) were evaluated. Results Women with at least probable minor depression [odds ratio (OR) = 1.77 (95% confidence interval (CI) = 1.02, 3.07)] or probable major depression [OR = 1.82 (95% CI = 1.01, 3.25)] in mid-pregnancy had an increased risk of SGA compared to non-depressed women in adjusted analyses. Borderline significant associations were observed between increasing levels of depressive symptom scores in early and mid-pregnancy [OR = 1.05 (95% CI = 1.00, 1.11) and OR = 1.04 (95% CI = 1.00, 1.09), respectively] and each additional trimester of exposure to probable major depression across mid- to late pregnancy [OR = 1.66 (95% CI = 1.00, 2.74)] and SGA. Late pregnancy depression was not associated with SGA; depression during pregnancy was not associated with preterm birth. Conclusions for Practice In this population of predominantly Puerto Rican women, mid-pregnancy depression increased risk for SGA. Findings can inform culturally appropriate, targeted interventions to identify and treat pregnant women with depression.

Human chorionic gonadotropin (hCG) is a pregnancy hormone secreted by the placental synctiotrophoblast cell layer that has been linked to fetal growth and various placental, uterine and fetal functions. In order to investigate the effects of hCG on clinical endpoints, knowledge on reference range (RR) methodology and determinants of gestational hCG levels is crucial. Moreover, a better understanding of gestational hCG physiology can improve current screening programs and future clinical management. Serum total hCG levels were determined in 8195 women participating in the Generation R Study. Gestational age specific RRs using 'ultrasound derived gestational age' (US RRs) were calculated and compared with 'last menstrual period derived gestational age' (LMP RRs) and a model-based RR. We also investigated which pregnancy characteristics were associated with hCG levels. Compared to the US RRs, the LMP RRs were lower, most notably for the median and lower limit levels. No considerable differences were found between RRs calculated in the general population or in uncomplicated pregnancies only. Maternal smoking, BMI, parity, ethnicity, fetal gender, placental weight and hyperemesis gravidarum symptoms were associated with total hCG. We provide gestational RRs for total hCG and show that total hCG values and RR cut-offs during pregnancy vary depending on pregnancy dating methodology. This is likely due to the influence of hCG on embryonic growth, suggesting that ultrasound based pregnancy dating might be less reliable in women with high/low hCG levels. Furthermore, we identify different pregnancy characteristics that influence total hCG levels considerably and should therefore be accounted for in clinical studies.

Thyroid hormones are critical for normal brain development. This study examined autism spectrum disorders (ASD) and thyroid stimulating hormone (TSH) levels measured in mid-pregnancy maternal serum and infant blood after birth. Three groups of children born in Orange County, CA in 2000-2001 were identified: ASD (n = 78), developmental delay (n = 45), and general population controls (GP) (n = 149). Samples were retrieved from prenatal and newborn screening specimen archives. Adjusted logistic regression models showed inverse associations between ASD and log transformed TSH levels in maternal serum samples (ASD vs. GP: OR [95 % CI] 0.33 [0.12-0.91], Early Onset ASD vs. GP: 0.31 [0.10-0.98]). Results for thyroid levels in newborn blood samples were similar though not significant (ASD vs. GP: 0.61 [0.18-2.04]).

The discovery of cell-free fetal DNA in maternal plasma opened up new possibilities for noninvasive prenatal testing (NIPT). Conceptual advances in single-molecule counting have resulted in robust methods for the NIPT of fetal chromosomal aneuploidies and subchromosomal aberrations. Such methods are employed worldwide and are among the most rapidly adopted genomic tests. Furthermore, approaches for fetal whole-genome sequencing from maternal plasma, as well as for targeted detection of many single-gene disorders, have been reported. Recently, fetal methylome and transcriptome sequencing from maternal plasma have also been achieved, potentially allowing fetal physiological and pathological processes to be monitored noninvasively using maternal blood. These advances herald exciting future applications in prenatal medicine., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Objective: To determine the diagnostic accuracy, validity, current limitations of, and possible solutions to, fetal RhD genotyping from maternal blood based on existing studies written in English., Methods: A literature search was conducted that described fetal RhD determination from maternal blood. The number of samples tested, fetal RhD genotype, the source of cell-free fetal DNA, gestational age and fetal Rh type were examined in each study to calculate the accuracy, sensitivity and specificity of fetal RhD genotyping., Results: Forty-one publications, which included 11,129 samples with non-invasive Rh genotyping of cell-free fetal DNA from maternal blood, were selected. After the exclusion of 352 inconclusive samples, the overall diagnostic accuracy was 98.5% (10,611/10,777), and sensitivity and specificity were 99% and 98%, respectively. First trimester diagnosis showed an accuracy of 99%, higher than second and third trimester diagnosis. Thirty studies reported a 100% diagnostic accuracy of fetal RhD genotyping., Conclusion: Non-invasive fetal RhD genotyping from maternal blood has high accuracy, sensitivity and specificity. METHODS reducing false results have been explored and applied in research. These achievements indicate that this technique will be widely used in routine clinical care.

Objective: To explore the major risk factors and early prediction methods in the pathogenesis of early onset preeclampsia through combining prenatal screening markers and epidemiological characteristics., Methods: Prenatal screening was performed in second trimester using enzyme-linked immunosorbent assay in 1,011 gravidas and epidemiological correlation factors were got by telephone with prospective cohort study. Predictive model of early onset preeclampsia was established and evaluated by single and multiple factor logistic analysis in 30 cases of preeclampsia and 867 cases of normal gravidas., Results: As compared with the control group, the maternal serum level of human chorionic gonadotropin (hCG) in second trimester of patients with early onset preeclampsia elevated significantly (P < 0.001). Pregestational BMI ≥ 24 kg/m(2) (OR = 3.649, 95 % CI 1.600-8.321, P = 0.002), history of hypertension, diabetes and nephritis (OR = 55.724, 95 % CI 8.223-377.614, P < 0.001), family history of hypertension (OR = 6.777, 95 % CI 2.917-15.742, P < 0.001), and risk coefficient for trisomy 21 (OR = 3.688, 95 % CI 1.013-13.429, P = 0.048) were major risk factors of early onset preeclampsia. The sensitivity and specificity of predictive model were 70.0 and 75.1 %, when cutoff point was 0.249. The diagnostic accuracy of the logistic model was better than hCG., Conclusions: In order to early prevent the onset and development of EOPE, it is necessary to strengthen pregestational and prenatal care for women in these aspects including pregestational BMI ≥ 24 kg/m(2), history of hypertension, diabetes, nephritis, family history of hypertension, and high risk for trisomy 21 syndrom.

Background: Non-invasive fetal RHD genotyping is an important tool to assess the risk of fetuse's hemolytic disease of anti-D allo-immunized pregnant woman by non-invasive method. A method of genotyping has been developed in the laboratory of Lyon-GHE according to Minon's team (J Gynecol Obstet Biol Reprod 2005): exon 4, 5, and 10 are amplified by real time PCR. At first, genotyping results of 200 pregnant women have been compared with RH1 phenotype at birth. The most important parameters of validation have been tested: the sensibility and the specificity; the negative predictive value; the correlation study permitted to define criteria of biological interpretation. The validation of this method permitted to determine critical points and the limits of the method due to the minor amount of fetal DNA in the maternal plasma and existence of many variant forms of the RHD gene., Conclusion: We worked too in the perspective to the accreditation for our genetic laboratory., (Copyright © 2014. Published by Elsevier SAS.)

Down syndrome is the most common cause of mental retardation and has an incidence of between 1:600 and 1:800 pregnancies. It is the condition for which prenatal diagnosis is requested the most and most developed countries have adopted a screening program based around maternal plasma/serum testing and ultrasound. Advances have been made recently to eliminate invasive testing for genetic diagnosis of this condition based on the analysis of free fetal DNA in maternal plasma. But, routine noninvasive prenatal diagnosis for trisomy 21 still appears to be years away. Screening based on assessment of various biomarkers present in maternal plasma represents a front-line test to assess the risk of the mother carrying an aneuploid fetus. Recent comparative proteomics techniques have resulted in studies that have assessed maternal plasma from mothers carrying normal and trisomy 21 fetuses and various gestational ages. Over 100 biomarker candidates have been described, but little consensus has emerged. This may be due to a number of compounded factors, but interesting to note that other neurological disorders have overlapping biomarkers. This article describes these developments and how these biomarkers could contribute to future screening in an emerging era where next-generation sequencing of free fetal DNA will be established in prenatal diagnostics, which appears imminent., (Copyright 2013 Prous Science, S.A.U. or its licensors. All rights reserved.)

Objective: To evaluate the extent of fetal structural abnormalities, other than neural tube and abdominal wall defects (AWDs), identified by California's Prenatal Screening Program., Methods: The Quad Marker Prenatal Screening records of 516,172 women were examined for screening interpretation and the diagnosis of structural abnormalities detected via follow-up. Women who were screen-positive for trisomy 21, trisomy 18, neural tube defects (NTDs) or Smith-Lemli-Opitz syndrome (SLOS) received follow-up services at state-approved Prenatal Diagnosis Centers (PDCs). Detailed reports of services and diagnostic information were linked in a database to the original screening results., Results: A total of 26 323 women received follow-up ultrasound services at the PDCs in the study time period. Of these women, 1085 (4.1%) were identified as having fetuses with significant structural abnormalities, other than NTDs (n = 207) or AWDs (n = 254). In addition to the structural abnormalities, 225 cases of fetal demise, 4 molar pregnancies, 15 cases of twin-to-twin transfusion, and 92 cases with placental abnormalities were identified., Conclusion: While Prenatal Screening Programs do not explicitly screen for structural abnormalities other than NTDs and AWDs, clearly many other structural abnormalities may be associated with a screen-positive status. Thus, the detection of these additional structural defects can be considered an ancillary program benefit., (© 2010 John Wiley & Sons, Ltd.)

The authors report a case of minimal prenatal trauma producing a large subdural hematoma in the fetus, which was diagnosed in utero by MR imaging. The occurrence of such a complication is extremely rare in the absence of significant maternal trauma. Prenatally diagnosed intracranial hemorrhages, particularly those that are subdural in origin, have a poor prognosis in most cases. After birth, brain compression required a complex neurosurgical intervention because simple hematoma evacuation was not possible. The clinical and neurological outcome at 6 months was excellent, as confirmed by the neuroimaging findings.

Objective: To re-evaluate in a larger cohort of patients if the maternal serum biochemical markers used in first trimester aneuploidy screening have the same marker distributions in pregnancies with a previous history of aneuploidy compared with those that have no previous history., Methods: Information related to previous pregnancy history is routinely recorded as part of first trimester screening in three centres King George, Kings College and Fetal Medicine Centre, London. From the database, records were extracted for women who had a previous pregnancy diagnosed with trisomies 13, 18 or 21. For each woman with a previous aneuploidy, five unaffected pregnancies in women of the same maternal age and with no previous aneuploidy pregnancy were selected as controls. A comparison was made between the marker distributions for pregnancy associated plasma protein-A (PAPP-A) and free beta-human chronic gonadotrophin (beta-hCG) amongst the cases and controls using nonparametric statistical tests., Results: A series of 8240 controls were compared against group of 1032 cases with a previous trisomy 21, 293 with a previous trisomy 18 and 158 with a previous trisomy 13. Cases with multiple previous trisomies were excluded. There were no significant differences in the level of free beta-hCG; however, in cases of trisomy 21 and trisomy 13 the levels of PAPP-A were increased by 5 and 16%, respectively., Conclusion: Risk calculation algorithms may need to take account of the increased PAPP-A levels in women with a previous trisomy 21 or trisomy 13., (Copyright (c) 2009 John Wiley & Sons, Ltd.)

Purpose of Review: The purpose of the review is to argue for and against introduction of HPA-1 typing of all pregnant women to reduce morbidity and mortality caused by foetal/neonatal alloimmune thrombocytopenia (FNAIT)., Recent Finding: Several groups have done HPA-1 typing in cohorts of pregnant women. Results from a Norwegian study (>100,000 pregnancies) indicate that screening combined with simple intervention decreases morbidity and mortality due to FNAIT and is cost effective in Norway. Results from this study and several other studies show that there is correlation between the level of anti-HPA-1a antibodies in the mother and the severity of thrombocytopenia in the newborn. An important finding is that about 75% of women with antibodies are immunized in connection with delivery. Only 25% of the women are immunized during pregnancy., Summary: Screening for FNAIT does not fully meet the criteria presented by the WHO. Nevertheless, the results of the Norwegian study strongly indicate that morbidity and mortality related to FNAIT can be reduced. If the recent attempts to make a vaccine aimed at prevention of immunization and/or tolerizing peptides or neutralizing antibodies for already immunized women are shown to be successful, screening must be implemented.

Purpose: Genetic counseling of carriers with individual chromosome translocation requires information on how balanced reciprocal chromosome translocations (RCT) will segregate, what possible form of unbalanced embryo/fetus/child can occur, and the survival rates that have been observed in the particular families. We collected new empirical data and evaluated pedigrees of RCT carriers involving 9p in order to improve risk figures., Material and Methods: Empirical data on 241 pregnancies of 70 carriers were collected from 32 pedigrees of carriers of RCT at risk for a single 9p segment imbalance (RCT9p) from the literature and unpublished data. The probability rates of particular types of pathology have been calculated according to the method of Stengel-Rutkowski and Stene. Cytogenetic interpretation was based on GTG, RBG and FISH techniques., Results: The probability rate for unbalanced offspring at birth for the whole group of pedigrees was calculated as 17.8+/-3% (33/185) (high risk). Considering the size of the imbalanced segment of 9p, the probability rates for RCT carriers with a breakpoint position at 9p22 at 9p13 and at 9p11.2 were estimated separately, and were found as 21.2+/-4.4% (18/85), 25+/-8.8% (6/24) and 11.8+/-3.7% (9/76), respectively. For unbalanced fetuses at 2nd prenatal diagnosis, we found the risk value as 57.9+/-11.3 % (11/19). The risk value for unkaryotyped stillbirths/early deaths of newborns and miscarriages were 5.4+/-1.7% (10/185) (medium risk) and 13+/-2.8% (rate 24/185) (high risk) respectively., Conclusions: Our results showed that the recurrence probability rates are different for particular categories of unfavorable pregnancy outcomes. How much they are dependent on the size of 9p chromosome segments taking part in the imbalance needs further studies based on a larger number of observations.

Objectives: Detection of free fetal DNA (ffDNA) in maternal blood during pregnancy has given rise to the possibility of developing new noninvasive approaches for early prenatal diagnosis. On a large-scale study, two protocols of real-time polymerase chain reaction (PCR) were compared in order to establish which Y-specific locus, either multicopy DYS14 or single copy SRY sequence, was the most suitable for developing a test with high diagnostic efficiency for early fetal gender assessment. The second aim was to assess whether the combination of the two detection systems could increase the performance of the prenatal test., Methods: We analyzed 145 plasma samples from healthy pregnant women between 11 and 12 weeks of singleton gestation. For each sample, fetal gender was determined by using both protocols (DYS14 and SRY) during the same real-time PCR run., Results: The data obtained by the DYS14 and SRY assays showed an efficiency in fetal gender prediction of 97.9 and 80%, respectively. It is not advisable to combine the two protocols because this association does not help in further improvements in fetal gender prediction., Conclusions: DYS14 assay is the best approach for early fetal gender assessment because it is more sensitive, accurate, and efficient than the SRY assay.

Objective: Australian women conceiving with ART are at fourfold risk of admission to early parenting treatment programs compared with those conceiving spontaneously. This study aimed to identify prevalence and determinants of antenatal mood disturbance and other risks for early parenting difficulties after assisted conception., Design: A prospective longitudinal investigation from conception to 18 months postpartum using telephone interviews and self-report questionnaires., Setting: Melbourne IVF and Royal Women's Hospital Reproductive Services, Victoria, Australia., Patient(s): A consecutive cohort of English-speaking women with ultrasound-confirmed ART-conceived pregnancies., Main Outcome Measure(s): Standardized psychometric measures of mood, quality of marital relationship, mother-to-fetus emotional attachment, and personality., Intervention(s): None., Result(s): Of the 288 women with confirmed pregnancies, 239 were contactable, and 183 (77%) were recruited, 95% of whom completed both early and late pregnancy assessments. Participants were socioeconomically advantaged, had very good pregnancy health, exceptional marital relationships, normal personality styles, and intense affectionate attachment to the fetus. Very few (<5%) had clinically significant mood disturbance in late pregnancy., Conclusion(s): There were low rates of antenatal mood disturbance and other risk factors for postpartum depression. Pregnancy and motherhood might be idealized after ART conception, and preparation for the realities of infant care might then be insufficient.

During pregnancy periodical hormonal, metabolic, hematological, vascular and immunological changes can be observed, that can influence the function of the eye. Some of them can be observed in an eye due to pregnancy itself and would resolved spontaneously postpartum. Pregnancy can also induce or aggravate the preexisting conditions such as diabetic retinopathy or uveal melanoma. The autoimmune intraocular inflammation is modified by pregnancy manifesting by regression of the ocular symptoms. Pregnancy can have also beneficial effects on glaucoma because intraocular pressure has been reported to decrease during pregnancy. The presence of any ocular symptoms in a pregnant woman requires urgent ophthalmic examination and further management.

Background: The discovery of the presence of fetal genetic material in maternal blood has opened up a new approach to prenatal diagnosis. One approach that has been extensively investigated over the past few decades is the isolation of fetal cells from maternal blood (Herzenberg et al. Proc Natl Acad Sci USA. 1979;76:1453-5; Bianchi et al. Proc Natl Acad Sci USA. 1990;87:3279-83; Cha et al. Prenat Diagn. 2005;25:586-91). As the fetal cells are scarce and the enrichment is of low efficiency, the technique could not be implemented in clinics. In 1997, Lo et al. (Lancet 1997;350(9076):485-7) discovered that cell-free fetal DNA is present in the plasma and serum of pregnant women. This discovery suggests that maternal plasma/serum DNA may be a useful source of material for non-invasive prenatal diagnosis. The objective of our study was to investigate the feasibility of using fetal DNA in maternal plasma for prenatal diagnosis., Methods: Plasma DNA in 277 blood samples of 40 pregnant women at the gestational period from 5 to 40 weeks and 24 h after delivery were extracted by column separation. FQ-PCR was used to amplify the SRY sequence in 237 plasma samples of 30 pregnant women. Fluorescent PCR was used to amplify 9 short tandem repeat loci simultaneously in 40 plasma samples of 10 pregnant women, and genomic DNA samples from their husbands were amplified by the same method., Results: The fetal SRY sequence could be detected from the 7th week of gestation, with a concentration that increased with progressing gestational age, attaining its highest peak before delivery. Twenty-four hours after delivery, fetal SRY sequence could not be detected in the maternal plasma. The concordance rate of the SRY sequence amplification results of plasma-free DNA, with real fetal gender was 100%. Analysis of maternal plasma samples collected during pregnancy revealed the presence of paternally inherited fetal-specific alleles. Among the 30 collected plasma samples, fetal-specific alleles were detected in 23 plasma DNA samples. The rate of positive results was 76% (23/30), and the frequency of positive results was 6/10 in early pregnancy, 8/10 in middle pregnancy, and 9/10 in late pregnancy. Short tandem repeats could not be detected from the maternal plasma 24 h after delivery., Conclusion: Fluorescent PCR can be used for amplification of fetal SRY sequence and STRs in maternal plasma to obtain fetal genetic information, which may have implications for non-invasive prenatal diagnosis of certain hereditary diseases.

Unlabelled: Hepatitis C is the most common chronic bloodborne infection in the United States. The diagnosis of vertical transmission is reliably established by a positive serum hepatitis C virus (HCV) RNA on 2 occasions 3 to 4 months apart after the infant is at least 2 months old and/or by the detection of anti-HCV antibodies after the infant is 18 months old. Vertical transmission in HCV RNA-negative pregnant women is approximately 1% to 3% versus approximately 4% to 6% in HCV RNA-positive women. From the standpoint of vertical transmission, no critical HCV RNA titer has been established. Coinfection with HIV has been shown to increase the risk of vertical transmission of HCV, but highly active antiretroviral therapy may decrease the risk significantly. In HIV-negative women, route of delivery does not influence vertical transmission. In HCV/HIV-coinfected women, decisions regarding mode of delivery should be based on HIV status. There is no association between vertical transmission of HCV and gestational age at delivery or the presence of chorioamnionitis. The use of a scalp electrode has been associated with vertical transmission and this practice is discouraged. Data are conflicting regarding duration of ruptured membranes and the risk of vertical transmission of hepatitis C. When the duration of membrane rupture exceeds 6 hours, the risk may be increased. There is no evidence demonstrating an increased risk of HCV transmission in HIV-negative women who breast feed. In HCV/HIV-coinfected women, breast feeding is discouraged in women who have consistent access to safe infant formula., Target Audience: Obstetricians & Gynecologists, Family Physicians., Learning Objectives: After completion of this article, the reader should be able to recall that vertical transmission of hepatitis C (HCV) does occur, state that coinfection with HIV increases the transmission rate, and summarize that there is no association between gestational age or presence of chorioamnionitis and no evidence that a cesarean delivery prevents transmission.

Objective: To review the literature to produce a catalog of solutes measured in paired extraembryonic celomic fluid and maternal serum samples., Study Design: We conducted a MEDLINE and PUBMED literature search for January 1990 through March 2004 using the keywords coelomic fluid, celomic fluid, coelocentesis, celocentesis and maternal serum to select articles that met the objective criteria. The following information was extracted from these publications: gestational age at sampling, number of samples, mean concentration of the solute measured in maternal serum and extra-embryonic celomic fluid, authors and year of publication. The catalog is presented in 3 tables based on the gradients found between compartments., Results: We found 73 publications and selected 42 in which paired celomic fluid and maternal serum samples had been obtained. Ninety solutes were grouped into 3 tables using a 30% cutoff difference in favor of either of the 2 compartments., Conclusion: A catalog of solutes measured in human extraembryonic celomic fluid between the 5th and 14th weeks of pregnancy was generated. This catalog can provide guidance for future manipulations of extracelomic fluid for therapeutic purposes.

Objective: To develop a nested polymerase chain reaction (PCR) technique for fetal SRY gene identification using cell-free fetal DNA in maternal plasma., Methods: Peripheral blood samples were obtained from 30 pregnant women and cell-free DNA was extracted by the phenol/chloroform method from plasma. The nested PCR was carried out to amplify the fragment of SRY gene by two sets of PCR primer pairs. Direct sequencing analysis was then performed on the PCR product., Results: Among the 17 women bearing male fetuses, SRY sequences were detected in 15 plasma samples after nested PCR amplification, while none of the 13 women bearing female fetuses had the positive results. The accuracy and sensitivity were 93.3% (28/30) and 88.2% (15/17), respectively., Conclusion: The phenol/chloroform extraction for fetal DNA in maternal plasma was effective and simple. And the nested PCR amplification of SRY sequence is a convenient and low-cost approach for the non-invasive early prenatal diagnosis of sex-linked inheritant diseases.

Background: Large quantities of cell-free foetal DNA have been detected in amniotic fluid, and it has been proposed that this material may contribute to the pool of cell-free foetal DNA in maternal plasma., Methods: Twelve maternal blood samples were obtained from pregnant women about to undergo an amniocentesis. Cell-free DNA was extracted from the maternal plasma samples and the matched amniotic fluid samples. The amount of cell-free foetal DNA was quantified by real-time PCR assays for the SRY and RHD genes., Results: Amniotic fluid was found to contain vast quantities of cell-free DNA (median concentration = 3,978 copies/ml amniotic fluid). The concentration of cell-free foetal DNA in maternal plasma was much lower (median concentration = 96.6 copies/ml maternal plasma). No significant correlation could, however, be determined between these two pools of cell-free foetal DNA., Conclusions: Our data confirm that amniotic fluid contains prodigious quantities of cell-free foetal DNA, but as no relationship exists between this material and that in the maternal circulation, it is unlikely that the amnion contributes to the presence of cell-free foetal DNA in maternal plasma.

Objective: To gain an understanding of the experience of pregnancy while carrying a child with a known, nonlethal congenital abnormality., Design: Phenomenological study., Setting: The participants were interviewed in their own homes, a quiet clinic room, or another location comfortable for them., Participants: Fifteen pregnant women carrying a child with a nonlethal, congenital abnormality participated., Results: The pregnancy experience was of a paradoxical nature. Knowledge of the fetal diagnosis resulted in both positive and negative consequences. Three major themes were common to the experience for women: Time is good, but it is also the enemy; you grieve, but you do not grieve; my baby's not perfect, but (s)he's still mine., Conclusion: Because of the paradoxical feelings experienced, women need as much information as they request regarding the abnormality, its treatment, and prognosis. An appointment during pregnancy with pediatric specialists to discuss the plan for care of the infant after delivery is beneficial. Health care providers should support women's prenatal attachment during this unique pregnancy.

Objective: We examined the relationship between maternal plasma transforming growth factor-beta1 (TGF-beta1) concentrations and risk of preeclampsia among women delivering at Harare Maternity Hospital in Zimbabwe. We evaluated the relationship in the context of maternal systemic inflammation using plasma tumor necrosis factor-a soluble receptor p55 (sTNFp55) as a marker., Methods: 132 women with preeclampsia and 180 controls were included in this case-control study analysis. Maternal post-diagnosis plasma TGF-beta1 and sTNFp55 concentrations were determined using immunoassays. Logistic regression procedures were used to estimate odds ratios (OR) and 95% confidence intervals (CI) adjusted for confounders., Results: A linear increase in preeclampsia risk was observed with increasing quartiles of TGF-beta1 concentrations (p<0.01). Women whose TGF-beta1 concentrations were >or=25.1 ng/ml (quartile 4) had a 2.5-fold (95% CI 1.2-5.6) increased risk of preeclampsia as compared with those women whose concentrations were <11.2 ng/ml (quartile 1). Relative to women with no evidence of systemic inflammation and no elevated TGF-beta1 concentrations, those women who were jointly positive for elevated TGF-beta1 and sTNFp55 concentrations experienced a 5.3-fold (95% CI 2.3-12.0) increased risk of preeclampsia., Conclusion: Overall, we noted that elevated TGF-beta1 is associated with an increased risk of preeclampsia. We also noted that the preeclampsia risk is exaggerated in the presence of maternal systemic inflammation.

Objective: Maternal serum pregnancy-associated plasma protein-A (PAPP-A) and free beta-human chorionic gonadotrophin (beta-hCG) are useful markers in the screening of Down syndrome in the first trimester. We investigated the effect of intracytoplasmic sperm injection (ICSI), freezing and thawing of embryos on the levels of these two analytes in assisted reproduction pregnancies., Methods: We recruited 149 women who conceived after assisted reproduction with fresh embryos (92 from conventional IVF and 57 from ICSI), 85 women who conceived with frozen-thawed embryos (54 from conventional IVF and 31 from ICSI) and 401 women with spontaneous conceptions as controls. The concentrations of PAPP-A and free beta-hCG were measured between 10 and 14 weeks and were converted to multiples of medians (MoM) for comparisons., Results: Median PAPP-A MoMs were significantly reduced in ICSI pregnancies in the fresh and frozen-thawed embryo subgroups (0.70 and 0.66 MoM respectively) and in the IVF fresh embryo subgroups (0.83 MoM), as compared to controls (1.00 MoM). Free beta-hCG MoM was significantly reduced in the IVF fresh embryos subgroup (0.87 MoM), but not in the other three subgroups., Conclusion: Further studies for exploring the underlying pathophysiology and adjustment in the marker levels for screening of Down syndrome are warranted in pregnancies conceived after assisted reproduction., (Copyright (c) 2005 John Wiley & Sons, Ltd.)

Inhibin A is secreted in significant quantities by the corpus luteum and fetoplacental unit, suggesting a role in fertility and pregnancy. Negative feedback regulation of follicle-stimulating hormone during pregnancy is one expected function of inhibin A, but the complete repertoire of actions of this hormone in pregnancy, including paracrine and autocrine actions, is yet to be fully understood. Inhibin A levels have been carefully described throughout normal pregnancy and studied in association with maternal and fetal complication such as intrauterine growth restriction, preterm labor or delivery, and preeclampsia. The first clinical application of inhibin A measurement in pregnancy has been its use as a second-trimester maternal serum marker for Down syndrome. Our laboratory was among the first, in 1998, to implement Quad marker screening, inhibin A measurement in conjunction with alpha-fetoprotein, unconjugated estriol, and human chorionic gonadotropin, to assess patients' risk of having a Down syndrome baby. The test performance of the Quad test has been validated by several large studies, detecting about 80% of Down syndrome pregnancies at a 5% false-positive rate. The present review describes Down syndrome and the use of inhibin A in second-trimester prenatal screening. We also address the method used for inhibin A measurement, the biology of inhibin A in Down syndrome pregnancy, and the effects of covariates and other fetal abnormalities on inhibin A levels.

Objective: Folic acid fortification of breads and grains was implemented in the United States in 1998 in an attempt to reduce the incidence of neural tube defects. Outcome data from birth registries have shown a 20% drop-less than originally predicted. In this study, we ascertain if the impact of folic acid fortification is better seen at the time of midtrimester prenatal diagnosis by looking at incidence of high maternal serum alpha-fetoprotein (MSAFP) values., Methods: Data regarding MSAFP levels in 61,119 patients undergoing maternal serum screening at a large commercial laboratory were categorized by multiples of the median (MoM). The data were compared between 2 groups: before mandatory supplementation in the United States in 1997 and after mandatory supplementation in 2000. High MSAFP values were further categorized as high (2.75-4.00 MoM) or very high (more than 4.00 MoM). Data were analyzed by chi(2) analysis., Results: Comparative data showed a 32% decrease of patients with MoM greater than 2.75 + (2.5% -1.7%). Further categorizations revealed similar decreases., Conclusion: The introduction of folic acid fortification has produced a profound decrease in the number of high MSAFP values, reflective of a decreased incidence of neural tube defects. Our results help to validate the decision to fortify food with folic acid, which represents a highly successful public health policy for primary prevention of birth defects., Level of Evidence: II-2

Aim: The present study investigates both the antenatal prediction of the occurrence of depression during the first year postpartum and the course of depression in populations at different degrees of risk., Methods: In a population-based prospective study, 1618 women were screened during mid-pregnancy for risk factors with regard to depression. High-risk and low-risk women were identified, and depression (Research Diagnostic Criteria, RDC) was assessed at 32 weeks gestation and at 3, 6, and 12 months postpartum., Results: In the high-risk group (n=97), 25% of the women were depressed during the first year postpartum compared to 6% of the low-risk women (n=87). At 3 months postpartum, significantly more high-risk (17%) than low-risk women (1%) were depressed. While prevalence rates decreased after 3 months postpartum in the high-risk group, no significant fluctuations of prevalence rates were found in the low-risk group. Two risk factors were independently predictive of depression during the postpartum period: a personal history of depression, and high depressive symptomatology during mid-pregnancy., Conclusions: Women at high risk and low risk for depression during the early postpartum period can be detected during pregnancy. High-risk women were only at particular risk during the first 3 months postpartum.

Objectives: Maternal serum alpha-fetoprotein (AFP) levels have been known to be increased in red blood cell alloimmunisation. Since AFP is now thought be a pro-erythropoietic factor, we wished to evaluate fetal serum levels of AFP in cases of alloimmunised pregnancies., Methods: We studied AFP levels in 32 fetal serum samples from women with red blood cell alloimmunisation at the time of the first fetal blood sampling. We expressed the levels of AFP and haemoglobin as absolute numbers and as delta values (number of SDs by which the observed value differed from the normal mean for the same gestation). Main outcome measures were fetal serum AFP levels and fetal haemoglobin concentration., Results: Overall, fetal AFP level was higher than normal in the cases (delta values 2.4 +/- 5.5 SD). However, mildly affected non-hydropic cases had higher levels than severely affected fetuses with hydrops., Conclusions: Fetuses affected by red blood cell alloimmunisation have increased levels of serum AFP but these levels fall back to low levels in severe anaemia especially with hydrops, which could represent the failing of a compensatory erythropoietic mechanism. Our results suggest that fetal haematopoiesis is activated early in red blood cell alloimmunisation but was subsequently impaired., (Copyright 2003 John Wiley & Sons, Ltd.)

Although fetal cells have been known to escape to the maternal circulation for a number of years, research attempts to use them for prenatal diagnosis have not had any consistent success. This review attempts to trace the history of such attempts and to document their progress and reasons for success or failure. The opinions of recent conferences including that of the US National Institute of Child Health and Human Development, a sponsor of major US research in the field, are reported and discussed. It is concluded that although basic work has demonstrated the biologic availability of both fetal cells and their free DNA representatives in the maternal circulation at gestational ages relevant to prenatal diagnosis, much work remains to develop practical technology for their consistent recovery and assay., (Copyright 2003 John Wiley & Sons, Ltd.)