Prenatal diagnosis of metabolic diseases on chorionic villi obtained before the ninth week of pregnancy

Nine pregnancies at risk for various metabolic disorders were monitored by prenatal diagnosis on chorionic villi obtained between the sixth and ninth weeks of pregnancy. A diagnosis of an affected fetus was made in five cases (Sandhoff, Tay—Sachs (2), Pompe's, GMI), while metachromatic leukodystrophy, GM1 (2), and Pompe's were excluded in four cases. It is concluded that chorionic villi are a reliable tissue for prenatal diagnosis of metabolic disorders also when obtained before the ninth week.     

Prenatal diagnosis of inherited metabolic disorders by quantitation of characteristic metabolites in amniotic fluid: Facts and future

An attempt is made to summarize as completely as possible what is known about the prenatal diagnosis of amino– and organic acidurias by direct measurement of characteristic metabolites in amniotic fluid, and to indicate which disorders can potentially be diagnosed prenatally by direct quantitation of metabolites. Furthermore, the disorders are mentioned in which the prenatal diagnosis was proven to be unsuccessful by this approach. The prenatal diagnoses of a case of propionic acidemia and a case of tyrosinemia type I in the 11th and 12th week of gestational age, respectively, are reported and the prospects of performing amniocentesis in the first trimester for prenatal diagnosis are discussed.     

Chromosome banding in direct preparations of chorionic villi

Chorionic villus sampling (CVS) is now currently offered for first trimester prenatal diagnosis of genetic disorders. Chromosome analysis of CVS in direct and culture preparations is possible using modifications of standard banding techniques. We summarize our experience in applying QFQ, GTG, RBG, CBG, DA/DAPI, NOR, and SC differentiation protocols to direct preparations. Characteristic chromosome regions are properly labelled by these techniques, and analysis of 300 band stage karyotypes is consistently achievable on GTG banded direct preparations. However, banding of CVS direct chromosomes has proved to be difficult, and the analysis needs to be backed up by culture preparations.     

Prenatal screening for haemoglobin disorders

The technology has been available to detect carriers of haemoglobin disorders since the late 1960s. Prenatal diagnosis has been available since 1978. First trimester diagnosis by chorionic villus sampling and DNA analysis was introduced in 1982, and subsequent simplifications in DNA technology have made screening, counselling and prenatal diagnosis cost-effective at the community level, in countries at all levels of development. Audit of prenatal diagnosis for haemoglobin disorders in countries which have the resources and infrastructure necessary for genetic population screening (such as the UK and other European countries), has shown that the number of prenatal diagnoses actually performed fall far short of expectation. The demonstration that this reflects failures in delivering information, screening and counselling to the populations at risk, rather than rejection of prenatal diagnosis, shows the importance of placing more emphasis on the organisational and social requirements for genetic population screening. In some countries current attitudes towards abortion exclude provision of prenatal diagnosis within the health service, but in many such cases it has been set up in the private sector. It is also being introduced through combined private and charitable efforts in an increasing number of developing countries, including some with extremely limited health resources: such centres are likely to act as nuclei for emergence of genetics services in these communities. A particularly notable recent achievement is the introduction of prenatal diagnosis in Nigeria, where 1–2% of all children born suffer from sickling disorders.     

Delivering an accredited non-invasive prenatal diagnosis service for monogenic disorders and recommendations for best practice

The identification of cell-free fetal DNA circulating in maternal blood combined with technological developments, in particular next-generation sequencing, is enabling the development of safer prenatal diagnosis. While this technology has been widely applied as a highly sensitive screening test for aneuploidy, there has been relatively little clinical application for the diagnosis of monogenic disorders. In the UK, we have established non-invasive prenatal diagnosis (NIPD) as a clinical service for a range of inherited disorders. The results from NIPD do not require confirmation by invasive testing and are welcomed by patients and health professionals alike. Here, we describe the technical approaches used, current practice and outline recommendations for best practice when delivering an NIPD service from an accredited laboratory. © 2017 John Wiley & Sons, Ltd.     

Preimplantation genetic diagnosis at 20 years

First reported in 1990, PGD has evolved into a complementary form of prenatal diagnosis offering novel indications. DNA for PGD can be recovered with equal safety and facility from polar bodies I and II, blastomere (8 cell embryo) and trophectoderm (5–6 day blastocyst). Diagnostic accuracy is very high (>99%) for both chromosomal abnormalities and single gene disorders. Traditional application of FISH with chromosome specific probes for detecting aneuploidy and translocations may be replaced or complemented by array comparative genome hybridization (array CGH); biopsied embryos can now be cryopreserved (vitrification) while analysis proceeds in orderly fashion. PGD has been accomplished for over 200 different single gene disorders. Novel indications for PGD not readily applicable by traditional prenatal genetic diagnosis include avoiding clinical pregnancy termination, performing preconceptional diagnosis (polar body I), obtaining prenatal diagnosis without disclosure of prenatal genotype (nondisclosure), diagnosing adult-onset disorders particularly cancer, and identifying HLA compatible embryos suitable for recovering umbilical cord blood stem cells. Copyright © 2010 John Wiley & Sons, Ltd.     

Attitude of Saudi families affected with hemoglobinopathies towards prenatal screening and abortion and the influence of religious ruling (Fatwa)

Hemoglobinopathies are common inherited disorders in Saudi Arabia. Prenatal diagnosis for such diseases is specific and sensitive but not yet implemented in Saudi Arabia. Saudis are Muslims with a very high rate of consanguinity and inherited genetic disorders. To examine the attitude of Saudi families affected with hemoglobinopathies towards prenatal diagnosis and abortion, and to evaluate the effect of education on religious ruling on such attitudes, 32 families were interviewed using a pre-structured questionnaire. The majority accepted prenatal diagnosis (81.3%). The attitude towards abortion was greatly affected by religious values. Education about religious ruling significantly affected parents' attitude towards accepting abortion and prenatal diagnosis. No other factors were found to influence the outcome. Although the majority of families received some kind of formal genetic counseling [23/32 (71.9%)], none of them was informed about the possibility of prenatal or preimplantation diagnosis prior to the interview. Therefore for prevention of genetic disorders, the emphasis in countries with a vast majority of Muslims such as Saudi Arabia has probably to be placed on public awareness about genetic risks, the risk of consanguinity, availability of services, and so on, while at the same time taking into consideration the religious beliefs and education of the target population Copyright © 2001 John Wiley & Sons, Ltd.     

First trimester diagnosis of lesch-nyhan syndrome: Applications to other disorders of purine metabolism

The prenatal detection of hypoxanthine phosphoribosyltransferase (HPRT, EC 2.4.2.8) deficiency, the Lesch-Nyhan syndrome, during the first trimester of an affected pregnancy through the use of chorionic villus sampling is reported. Quantitation of reaction products formed by villus cell extracts from exogenous hypoxanthine-8-[¹⁴C] or adenine-8-[¹⁴C] is used in diagnosis. We report the diagnosis of Lesch-Nyhan syndrome using a chorionic villus specimen and confirmation of that diagnosis. In addition, adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP), enzymes deficient in inherited immune disorders, are detected in chorionic villus samples. These heritable disorders also appear amenable to early prenatal diagnosis.     

不同肥料施用对设施菠菜地NH3挥发和N2O排放的影响

以湖南设施菠菜地为研究对象,采用通气法和静态箱-气相色谱法研究了设施菠菜生长季施用工厂化堆肥(SC)、农民传统堆肥(SS)和单施化肥(FC)对土壤NH_3和N_2O排放的影响.结果表明在同等施氮水平下,与FC相比,SC处理能减少NH_3和N_2O排放量,分别减少52.9%和95.12%(P0.01);SS处理则增加NH_3挥发量24.8%,减少N_2O排放量48.8%.不同处理NH_3损失率分别为SS(10.97%)FC(4.19%)SC(2.74%);N_2O排放系数分别为FC(4.50%)SC(2.21%)SS(0.60%);与FC处理相比,SC处理条件下菠菜产量和作物吸氮量分别降低19.61%和13.20%,差异不显著;SS处理则显著降低27.9%和40.0%(P0.05).在该地区冬季设施菠菜田中,影响NH_3和N_2O排放的主要因素为土壤水分,环境温度不是此研究中影响气体排放的主要因素.因此,畜禽养殖固体废弃物经过不同处理后作为有机肥施入菠菜田,SC处理的菠菜可保证产量的同时,减少NH_3挥发和N_2O排放损失.     

Molecular prenatal diagnosis: the impact of modern technologies

Originally prenatal diagnosis was confined to the diagnosis of metabolic disorders and depended on assaying enzyme levels in amniotic fluid. With the development of recombinant DNA technology, molecular diagnosis became possible for some genetic conditions late in the 1970s. Here we briefly review the history of molecular prenatal diagnostic testing, using Duchenne muscular dystrophy as an example, and describe how over the last 30 years we have moved from offering testing to a few affected individuals using techniques, such as Southern blotting to identify deletions, to more rapid and accurate PCR-based testing which identifies the precise change in dystrophin for a greater number of families. We discuss the potential for safer, earlier prenatal genetic diagnosis using cell free fetal DNA in maternal blood before concluding by speculating on how more recent techniques, such as next generation sequencing, might further impact on the potential for molecular prenatal testing. Progress is not without its challenges, and as cytogenetics and molecular genetics begin to unite into one, we foresee the main challenge will not be in identifying the genetic change, but rather in interpreting its significance, particularly in the prenatal setting where we frequently have no phenotype on which to base interpretation. Copyright © 2010 John Wiley & Sons, Ltd.     

Circulating ‘trophoblast’ cells in pregnancy have maternal genetic markers

A syncytiotrophoblast-associated antigen identified by the monoclonal antibody (McAb) H315 is detectable on the surface of a low proportion of peripheral blood cells in pregnant women, raising the possibility of a new approach to prenatal diagnosis of genetic disorders. We aimed at verifying the trophoblastic origin of H315⁺ cells and their use for prenatal diagnosis of β-thalassaemia. H315 ⁺ cells were separated from the peripheral blood of pregnant women: the DNA obtained from these cells in two selected cases was shown to have genetic markers indistinguishable from those of the mother and definitely different from the fetus. Our results suggest that H315 antigen is expressed by maternal cells and that prenatal diagnosis on peripheral blood of the mother using H315 McAb is not feasible.     

Prenatal diagnosis by chorionic villus sampling: Lessons of the first 600 cases

Chorionic villus sampling (CVS) has emerged as a first trimester alternative to amniocentesis for the prenatal detection of genetic disorders. We report our experience in 600 consecutive CVS procedures to better delineate the safety, efficacy and reliability of this new method of prenatal diagnosis. Adequate samples were obtained at the initial visit in 97 per cent of the cases, and successful cultures were established in 98.7 per cent of these patients. Chromosome abnormalities were detected in 5.9 per cent of those pregnancies tested because of advanced maternal age (≥ 35 years). A discrepancy between the villus karyotype and that of the fetus was found in 2.0 per cent of cases, and most commonly consisted of mosaicism in the villus sample for a chromosomal abnormality that was not found in fetal samples. The risk of spontaneous abortion following the procedure was 6.3 per cent. We conclude that chorionic villus sampling is an acceptably safe and reliable procedure, but further investigation is needed before it can become an established technique in prenatal diagnosis.     

First-trimester biochemical and molecular diagnoses using chorionic villi: High accuracy in the U.S. collaborative study

The accuracy of biochemical and molecular prenatal diagnoses using chorionic villi as the fetal source was assessed by seven centres participating in the NICHD collaborative study on the safety and accuracy of chorionic villus sampling (CVS) and amniocentesis. Of 601 pregnancies studied, biochemical methods were used to determine the diagnosis in 283 fetuses at risk for 35 different metabolic disorders. Fifteen different lysosomal storage diseases accounted for 81 per cent of the biochemical prenatal diagnoses performed, with 57 per cent of these pregnancies at risk for Tay-Sachs disease. No errors were made in the biochemical diagnoses that predicted affected or unaffected fetuses. However, the diagnoses of certain disorders (e.g., mucopolysacchariodosis type IH, metachromatic leukodystrophy, and Krabbe disease) occasionally required confirmatory studies in cultured amniocytes because the enzyme results were inconclusive in direct and/or cultured villi or due to the presence of a pseudodeficiency allele. Of these, only the diagnosis of a fetus at risk for Krabbe disease remained inconclusive after special studies to discriminate between mutant and pseudodeficiency alleles. Recombinant DNA techniques were used to predict the diagnosis of 318 fetuses at risk for 16 different disorders in which the defective disease gene could be detected either directly or by linkage analysis to a nearby polymorphic marker. Of these, 32 per cent were for haemoglobinopathies, 25 per cent for cystic fibrosis, 24 per cent for Duchenne or Becker muscular dystrophy, and 7 per cent for haemophilias. Pregnancies at risk for known disorders with specific molecular lesions (e.g., sickle cell disease) were accurately diagnosed in direct and/or cultured villi. Diagnoses requiring analyses with closely linked polymorphic markers were occasionally uninformative or inconclusive. Maternal contamination was not reported in any biochemical or molecular-based diagnosis. These studies document the high accuracy and rapidity of both biochemical and mutation-specific prenatal diagnoses with direct and cultured chorionic villi.     

Ultrastructure of first trimester chorionic villi with regard to the prenatal diagnosis of genodermatoses

Hopes are held out for chorion villus sampling, a technique which is gaining more and more importance for the first trimester prenatal diagnosis of chromosomal aberrations and metabolic abnormalities. A variety of inherited skin diseases can be diagnosed postnatally and prenatally (in the second trimester) by ultrastructural diagnostic markers. For evaluation of prenatal diagnosis in the first trimester, we investigated chorionic villi derived from the trophoblast layer of the early pregnancy by light microscopy and conventional electron microscopy. The ultrastructure of the cellular layers covering the villi, i.e., the inner cytotrophoblast and the outer syncytiotrophoblast, as well as that of the connective tissue of the inner extraembryonic mesoderm, are thoroughly described in relation to the ultra-structural changes in certain genodermatoses including epidermolyses and keratinization disorders. We found that chorionic villi have only a few of the characteristics differentiated in skin, and none of the structures which are relevant to the diagnosis of genodermatoses. In our view, the ultrastructural approach is not suitable for first trimester prenatal diagnosis of genodermatoses in chorionic villi.     

Prenatal diagnosis of inborn errors in peroxisomal β-oxidation

In recent years, an increasing number of inherited diseases in man have been recognized in which there is an impairment in the peroxisomal β-oxidation of very-long-chain fatty acids. In general, these disorders are associated with severe neurological and physical abnormalities and death within the first years of life. In this paper we describe our experience with regard to the prenatal diagnosis of a number of different inborn errors of peroxisomal β-oxidation. Eleven pregnancies at risk were monitored by measuring very-long-chain fatty acid levels as well as very-long-chain fatty acid β-oxidation in cultured chorionic villous fibroblasts and/or amniotic fluid cells. Five affected fetuses were identified. It is concluded that prenatal diagnosis in this group of diseases can be done reliably using cultured chorionic villous fibroblasts or amniotic fluid cells.     

Prenatal diagnosis of haemoglobin disorders by cordocentesis at 12 weeks' gestation

Prenatal diagnosis of haemoglobin disorders is accepted to be a useful procedure to avoid births of infants with homozygous diseases. Advances in sampling and molecular techniques, such as polymerase chain reaction (PCR) and chorionic villus sampling (CVS), have made earlier and safer first-trimester prenatal diagnosis possible. However, these procedures need previous studies of at-risk couples, which can be very time-consuming when a number of different β-thalassaemia mutations occur in the region. We describe the possibility of making a first-trimester prenatal diagnosis by cordocentesis and fetal blood analysis at the 12th week of gestation. We found no statistically significant difference (p>0.05) between β/γ values in fetuses at the 12th and 18th weeks of gestation. In seven affected fetuses aborted at the 12th week of gestation, the diagnosis was confirmed in all cases by PCR analysis. These findings suggest that early cordocentesis could be an alternative procedure to CVS and PCR analysis.     

Future use and development of prenatal diagnosis. Consumers' attitudes

The rapid progress of prenatal diagnosis and genetic tests makes it important to investigate attitudes towards this development. A total of 40 women and 20 men with personal experience of prenatal diagnosis for chromosome aberrations were interviewed about their moral opinion of the development. The majority (88 per cent) considered it certain or probable that all new methods developed will also be used in the future. The majority (62 per cent) were hesitant about testing for common disorders, e.g., diabetes mellitus and rheumatoid arthritis, but regarded it justified in some situations. One-third of the individuals (31 per cent) wanted some kind of restrictions for the use of the tests, but only 13 per cent recommended legislation for this purpose. The majority (84 per cent) believed that ethical principles are influenced by scientific progress. In most aspects, the women and their partners had similar attitudes. However, 82 per cent of the women but only 20 per cent of the men considered that the couple itself should decide about prenatal diagnosis. The results demonstrated a considerable respect regarding the difficult choices associated with the development of prenatal diagnosis, also among those who have decided in favour of the test. The study indicated a greater need for autonomy among women than among men.     

Prenatal diagnosis of carbamoyl-phosphate synthetase deficiency by fetal liver biopsy

In a pregnancy at risk for carbamoyl-phosphate synthetase (CPS) deficiency, prenatal diagnosis was attempted by fetal liver biopsy, performed at 18 weeks of gestation. CPS activity was absent and the diagnosis was confirmed after termination of the pregnancy. The technique employed for fetal liver biopsy is described together with an evaluation of its possible role in prenatal diagnosis.     

Prenatal diagnosis of Huntington's disease (HD): Experiences with six cases and PCR

In the course of a 2-year predictive testing programme for Huntington's disease (HD), six couples from a total of 52 applicants requested prenatal testing. In each case, the pregnancy was in the first or second trimester when the couples were referred for DNA diagnosis. In five cases, exclusion testing was offered; in one case, a person at risk with an increased risk of being a gene carrier requested prenatal diagnosis. In all cases, informative markers for prenatal testing could be determined. Whenever possible, the newer technique of polymerase chain reaction (PCR) for D4S125 was applied to perform rapid prenatal diagnosis. Two couples withdrew before chorionic villus sampling was undertaken; prenatal diagnosis was completed in the remaining four cases. After exclusion testing, two pregnancies were determined to have an increased risk and two fetuses to have a low risk of being HD gene carriers.     

Light microscopic visualization of peroxisomes and plasmalogens in first trimester chorionic villi

Peroxisomes and their metabolites the plasmalogens were visualized in chorionic villi by quick and simple procedures. Villi were stained and mounted in toto, and can also be embedded. These methods may contribute to early prenatal diagnosis of peroxisomal disorders (Zellweger syndrome, neonatal adrenoleukodystrophy, infantile Refsum's disease a.o.).