Exome sequencing identifies mutations in KIF14 as a novel cause of an autosomal recessive lethal fetal ciliopathy phenotype.

Gene discovery using massively parallel sequencing has focused on phenotypes diagnosed postnatally such as well-characterized syndromes or intellectual disability, but is rarely reported for fetal disorders. We used family-based whole-exome sequencing in order to identify causal variants for a recurrent pattern of an undescribed lethal fetal congenital anomaly syndrome. The clinical signs included intrauterine growth restriction (IUGR), severe microcephaly, renal cystic dysplasia/agenesis and complex brain and genitourinary malformations. The phenotype was compatible with a ciliopathy, but not diagnostic of any known condition. We hypothesized biallelic disruption of a gene leading to a defect related to the primary cilium. We identified novel autosomal recessive truncating mutations in KIF14 that segregated with the phenotype. Mice with autosomal recessive mutations in the same gene have recently been shown to have a strikingly similar phenotype. Genotype-phenotype correlations indicate that the function of KIF14 in cell division and cytokinesis can be linked to a role in primary cilia, supported by previous cellular and model organism studies of proteins that interact with KIF14. We describe the first human phenotype, a novel lethal ciliary disorder, associated with biallelic inactivating mutations in KIF14. KIF14 may also be considered a candidate gene for allelic viable ciliary and/or microcephaly phenotypes.

Deletion in Xp22.11: PTCHD1 is a candidate gene for X-linked intellectual disability with or without autism.

Submicroscopic chromosomal anomalies play an important role in the aetiology of intellectual disability (ID) and have been shown to account for up to 10% of non-syndromic forms. We present a family with two affected boys compatible with X-linked inheritance of a phenotype of severe neurodevelopmental disorder co-segregating with a deletion in Xp22.11 exclusively containing the PTCHD1 gene. Although the exact function of this gene is unknown to date, the structural overlap of its encoded patched domain-containing protein 1, the transmembrane protein involved in the sonic hedgehog pathway, and its expression in human cortex and cerebellum as well as in mice and drosophila brain suggests a causative role of its nullisomy in the developmental phenotype of our family. Our findings support the recent notions that PTCHD1 may play a role in X-linked intellectual disability (XLID) and autism disorders.

Genotype-phenotype dilemma in a case of sudden cardiac death with the E1053K mutation and a deletion in the SCN5A gene.

Mutations in the cardiac sodium channel gene SCN5A may result in various arrhythmia syndromes such as long QT syndrome type 3 (LQTS), Brugada syndrome (BrS), sick sinus syndrome (SSS), cardiac conduction diseases (CCD) and possibly dilated cardiomyopathy (DCM). In most of these inherited cardiac arrhythmia syndromes the phenotypical expression may range from asymptomatic phenotypes to sudden cardiac death (SCD). A 16-year-old female died during sleep. Autopsy did not reveal any explanation for her death and a genetic analysis was performed. A variant in the SCN5A gene (E1053K) that was previously described as disease causing was detected. Family members are carriers of the same E1053K variant, some even in a homozygous state, but surprisingly did not exhibit any pathological cardiac phenotype. Due to the lack of genotype-phenotype correlation further genetic studies were performed. A novel deletion in the promoter region of SCN5A was identified in the sudden death victim but was absent in other family members. These findings demonstrate the difficulties in interpreting the results of a family-based genetic screening and underline the phenotypic variability of SCN5A mutations.

[New options in prenatal and preimplantation diagnosis of genetic disorders]. / Neue Möglichkeiten der pränatalen genetischen Diagnostik inklusive Präimplantationsdiagnostik.

During recent years the progress with the most important practical impact in prenatal diagnosis has been the implementation of first trimester risk screening for common aneuploidies leading to a much improved identification of pregnancies at risk. Molecular methods for a rapid, cost-effective, but selective aneuploidy diagnosis such as interphase FISH or QF-PCR have been around for years, do have their specific indications, but will unlikely replace conventional cytogenetic tools in routine diagnosis. They apparently do also play a role as marketing instruments in the competition among cytogenetic laboratories. The most thrilling issue for all cytogeneticists in the years to come will be the introduction of array-based methods in the prenatal routine diagnosis of chromosomal abnormalities. Polar body diagnosis has been the only option available for preimplantation genetic diagnosis in german speaking countries. The overwhelming majority of all professionals involved and many families concerned share the hope that the legal situation will improve in these countries to allow the examination of early embryos in high risk situations.

[Recent advances in prenatal diagnostics]. / Neue Möglichkeiten in der pränatalen Diagnostik.

During the last years, technical improvements have increased the possibilities in prenatal ultrasound. During the eighties and nineties, fetal malformations were increasingly detected and specified. Since a few years, the measurement of the fetal nuchal translucency between 11 and 14 weeks of gestation has been implemented to calculate the individual risk, in combination with most recent biochemical markers. Today, the sonographic measurement of the nuchal translucency is regarded as a valuable screening tool for chromosomal anomalies in prenatal medicine. Beside standardized examinations, a profound information and counseling of the pregnant women should be emphasized. With the improvement of the specific maternal risk calculation, using the sonographic measurement of the nuchal translucency, the biochemical markers and the maternal age, unnecessary invasive examinations may be prevented and their overall number can significantly be reduced. The same trend is seen in the whole field of prenatal medicine, illustrated by the detection of the fetal rhesus D status from the maternal blood and the use of Doppler ultrasound in the management of fetal anemia.

Autosomal recessive deafness with skeletal dysplasia and facial appearance of Marshall syndrome.

We report on two sibs born to consanguineous parents with clinical and radiological features closely resembling those previously described by Insley and Astley [1974]. This observation provides further evidence for a distinct autosomal recessive condition with the facial appearance of Marshall syndrome, deafness, and skeletal dysplasia.

Distal limb deficiency following chorionic villus sampling?

We have reviewed the follow-up of almost 3000 completed pregnancies in the Münster CVS Program and identified 4 children with distal limb deficiencies. Two cases involved only minor anomalies of distal digital phalanges. One child had a Hanhart anomaly (hypoglossia hypodactylia). We also reviewed 24 cases of limb defects following CVS reported previously. With the exception of a single series, neither the overall incidence of affected children nor cases reported from larger series provide unambiguous evidence of an increased risk of distal limb deficiency caused by CVS.

Estimating frequency of Kallmann syndrome among hypogonadic and among anosmic patients.

The frequency of Kallmann syndrome (hypogonadotropic hypogonadism and anosmia, HHA) was estimated in patients presenting with hypogonadism and patients with anosmia. Of 791 hypogonadal males 19 had HHA. The frequency of HHA was about 1:25 (n = 8/189) in outpatients questioned about their sense of smell, about 1:50 (n = 11/579) in patients whose blood samples were sent to us for chromosome analysis, and about 1:30 (n = 19/605) in males with hypogonadism and 46,XY chromosomes. The relation of patients with HHA to those with Klinefelter syndrome was 1:10 (n = 19/186). From 24 patients presenting with anosmia we found 1 hitherto undiagnosed case of HHA. The mean age at diagnosis was 24.8 and 24.9 years in our cases and cases from literature, respectively. These data provide evidence that Kallmann syndrome is not infrequent and that most patients remain undiagnosed until the third decade of life. Earlier diagnosis is emphasized by questioning each hypogonadal patient about his sense of smell because therapeutic success seems to be age dependent.

Validity of cytogenetic analyses from trophoblast tissue throughout gestation.

Cytogenetic findings in the Münster Chorionic Villi Sampling (CVS) program are presented after 1,184 first trimester transcervical samplings and 131 second and third trimester placentacenteses. In the first trimester series the abnormality rate is low (2.4%) in patients with only an age-dependent aneuploidy risk. In this group terminations were performed in only 1.6% because of aneuploidy. True mosaicism was found more frequently after CVS, and the risk of maternal cell contamination seems higher as compared to amniocentesis. There are no obvious differences in the overall rate of diagnostic errors after both procedures, when metaphases after direct preparation and chorionic cell cultures are analysed and doubtful findings such as mosaicism are adequately followed up by amniocentesis. The cytogenetic techniques also offer a very rapid approach to karyotyping in the second and third trimester. We found a high rate of aneuploidy (15%) when placentacentesis was performed after sonographic diagnosis of fetal abnormalities. We conclude that cytogenetic analysis from trophoblast tissue is an accurate diagnostic tool applicable from first to third trimester of pregnancy.

Low rates of pregnancy termination for prenatally diagnosed Klinefelter syndrome and other sex chromosome polysomies.

Over the past 9 years we counseled 55 couples whose unborn child was found to carry a sex chromosome polysomy. We performed a survey of postcounseling parental decisions about continuation or termination of these pregnancies. Of the 55 embryos or fetuses, 23 had the karyotype 47,XXY, 10 had 47,XYY, and 12 had 47,XXX. In addition, there were 10 instances of true mosaicism, i.e. 47,XXY/46,XY (n = 5), 47,XYY/46,XY (n = 2), or 47,XXX/46,XX (n = 3). Mean gestational age (+/-standard deviation) at diagnosis was 18.3+/-3.0 weeks. After comprehensive genetic counseling 48 (87.3%) of these pregnancies were carried to term. In seven cases (12.7%) the parents elected a pregnancy termination. Two of 31 pregnancies (6.5%) primarily ascertained at our center were aborted, whereas amongst the 24 referred cases, 5 couples (20.8%) opted for a termination. The mean gestational age of the terminated pregnancies was 19.7 weeks. The overall termination rate of 12.7% appears low in comparison with literature data. Most reports from other institutions present termination rates between 32 and 66%. The reason for the low rate of induced abortions in our study cohort is not clear. Cultural differences in parental perception of sex chromosomal polysomies may be of importance, and peculiarities of genetic counseling at our institution could also play a role. Although counseling was nondirective, we did put emphasis on providing prospective parents with information from unbiased follow-up studies of children with Klinefelter syndrome and other sex chromosome polysomies.

Parental origin of the extra haploid chromosome set in triploidies diagnosed prenatally.

The parental origin of the additional chromosome complement in a total of 17 cases of triploidy was determined mainly using highly polymorphic microsatellites. Maternal origin of the triploidy was demonstrated in most cases. To the best of our knowledge, this is the first systematic evaluation of the parental origin of chromosome sets in fetuses who survived until a cytogenetic diagnosis was established. In contrast to previous investigations this study documented a predominance of maternal origin of the extra haploid set mainly due to longer survival time for digynic triploidies. The concept of 2 distinct fetal phenotypes in triploidy is clearly supported by this study.

Transabdominal placental biopsy in the second and third trimesters of pregnancy: what is the risk of maternal contamination in DNA diagnosis?

We investigated the risk of maternal contamination in antenatal DNA diagnosis after second- and third-trimester transabdominal placental biopsy. For this purpose, we compared the restriction fragment length polymorphism (RFLP) patterns of 11 chorionic villus DNA samples after late chorionic villus sampling (CVS) with those of the corresponding maternal DNA. Ten of 11 aspirated tissue samples were not separated from maternal contamination before DNA extraction. All 11 mother-embryo pairs were informative for analysis of maternal contamination, ie, the mother showed one RFLP allele not present in the embryo. In none of the 11 cases did the fetal DNA show maternal contamination after molecular hybridization, although ten samples were contaminated with maternal tissue macroscopically and microscopically. Despite some maternal tissue admixture, the risk of contamination seems to be lower in the second- and third-trimester CVS than in first-trimester CVS, based on previous reports and our own experiences. This is most likely due to the anatomically closer contact of villi and decidua in the first trimester of pregnancy.

Beta-thalassemia problems in the Turkish population in the F.R.G.

A sample of 165 normal Turkish probands (79 males, 86 females) living in the F.R.G. was screened for the presence of heterozygous beta-thalassemia as well as their knowledge and perception of thalassemias in general. Hematologic studies revealed two persons to be heterozygous for beta-thalassemia who had not known this before. This incidence of 1.2% in our sample of Turkish probands living in the F.R.G. is in accordance with the average range of 0.2-6% given by the World Health Organization for Turkey. The probands' places of origin in Turkey correlate well with the pattern of migration within the country, with most people coming from central Anatolia which is an area with a relatively low incidence of beta-thalassemia. Interviews conducted by a Turkish doctor revealed that the knowledge about thalassemias and the function of blood in general was extremely limited in our study population. On the other hand, 50% of the respondents would refrain from marriage or choose a different partner if both partners knew that they were heterozygous for thalassemia. 19% would still marry in this situation but refrain from having children. When asked about their most likely decision in the case of an early diagnosis of homozygous thalassemia, 78% said they would have a termination and only 15% would definitely go on with the pregnancy. About 65% felt that consanguineous marriage is a predisposing factor for diseases in the offspring. From our study it can be concluded that a large-scale screening program in the high-risk population which exists in most Mediterranean countries would be difficult regarding the more than one million people from Turkey living in the F.R.G. and would have to take into account the cultural and educational conditions of the Turkish minority. In general, if no Turkish-speaking specialist is available to explain the diagnostic possibilities, probably pre-pregnancy or early pregnancy testing would be the most appropriate way of reaching the couples at risk in the German medical system.

A novel syndrome involving primary skeletal growth and retardation in siblings.

An identical pattern of malformations was found in two brothers both having microcephaly and severe developmental delay. Additionally, they had hypotelorism, epicanthic folds, and convergent strabismus. There was shortening of either the radius or the tibia and shortening of the first metacarpals. Persistently dorsally flexed fingers and toes were noted, all of which are unusually long. Both boys had a high-pitched voice and were unable to communicate verbally at the age of 4.5 years. They both developed short stature. One brother has anal atresia; the other had a pulmonary artery atresia, VSD, ASD, and an over-riding aorta. This apparently new syndrome is possibly an autosomal, or a X-linked recessive trait.

[Is sterility a genetic burden?]. / Ist Sterilität eine Erblast?

Genetic causes of infertility are probably not rare. Today only a fraction of genes directly or indirectly involved in reproduction including sex determination and differentiation are known. Nevertheless, the list of well-defined genetic disorders impairing fertility is impressing already today and growing rapidly. Gonosomal aneuploidy and structural rearrangements represent a significant portion of the genetic causes of infertility in both sexes. Other chromosomal conditions include autosomal balanced structural changes (e.g. translocations), probably due to pairing disturbances of the affected chromosomes during meiosis. Some fundamental mechanisms in sex determination and differentiation have been characterized in recent years. Mutations in some of the genes involved in this process may lead to familial infertility. Genetic defects in gametogenesis of both sexes are currently being investigated using mouse models. Male specific causes of infertility include microdeletion within the AZF region of the euchromatic part of the long arm of the Y chromosome and obstructive azoospermia due congenital aplasia of the vas deferens in the presence of mutations in the CFTR gene.

[Genetic testing in pregnancy]. / Genetische Untersuchungen während der Schwangerschaft.

First trimester risk screening is probably the major methodological advance in identifying pregnancies at increased risk for genetic disease during recent years with an impact on all pregnancies. The high detection rate with moderate false positive rates will reduce the over-all number of invasive procedures as compared to the traditional approach based on maternal age exclusively, in particular considering the demographic shift towards higher mean maternal age. Non-invasive prenatal diagnosis from fetal cells or DNA in the maternal circulation remains an experimental approach, despite a growing number of reports on successful diagnoses of single gene disorders. In the lab molecular cytogenetic approaches have considerably broadened the diagnostic spectrum of conventional karyotyping and facilitated a rapid diagnosis of selected frequent aneuploidies. Molecular genetic testing, in particular on chorionic villi, allows an early and reliable diagnosis of a growing number of severe monogenic conditions. A restrictive legislation has hampered the development of preimplantation genetic diagnosis in German speaking countries, only a few groups work on polar body diagnosis, a legal but restricted alternative.

[Prenatal diagnosis]. / Pränatale Diagnostik.

A growing number of methods for the in-utero diagnosis of fetal disease became available during recent decades, mainly due to rapid advances in ultrasound technology and laboratory methods. Exclusion of fetal aneuploidy in pregnancies of women beyond 34 years is still by far the most common reason for an invasive procedure event at a time when the number of diagnosable inborn errors of metabolism and monogenetic disorders is increasing rapidly. Great efforts have been made to improve the poor predictive value of maternal age as an indicator for an increased risk for fetal aneuploidy. Maternal serum screening using various parameters [e.g. AFP, hCG, uE3] has been found effective in identifying the majority of pregnancies with Down syndrome in the second trimester. Current research goals are the optimal choice of markers and the introduction of maternal serum screening in the first trimester of pregnancy. Ultrasound is another suitable tool to identify pregnancies at high risk for aneuploidy at least in the hands of experienced operators. The diagnostic use of fetal cells in the maternal circulation is presently investigated in a large collaborative trial. Second trimester amniocentesis still is the most widely applied invasive technique in pregnancies with maternal age related risks for fetal aneuploidy. Safety and diagnostic accuracy are well established and amniotic fluid can easily be shipped. Without major changes in conventional sampling and laboratory techniques the procedure can be performed at 13 weeks of gestation and later. The safety and efficacy of first-trimester amniocentesis has still to be established in larger series. First-trimester chorionic villus sampling is a well-established alternative to amniocentesis with comparable procedure-related risks. It is the method of first choice in pregnancies at high risk for aneuploidy, inborn errors of metabolism and monogenic disorders, since uncultured villi can be used for a rapid diagnosis. Placental biopsy should also be considered in the second and third trimester of pregnancy for these indications whenever time is a critical issue. Fetal blood obtained by ultrasound-guided cordocentesis has successfully been used for the diagnosis of fetal infection and is at the same time another good source of cells for rapid karyotyping. Other invasive procedures such as fetal skin or liver biopsies for electron microscopy or specific metabolic tests are restricted to the relatively rare instances of diseases where DNA diagnosis is currently not available or uninformative. Providing information on the different alternatives in an individual situation is one of the critical issues in pregnancy care today.

Dihydropyrimidine dehydrogenase deficiency caused by a novel genomic deletion c.505_513del of DPYD.

Dihydropyrimidine dehydrogenase (DPD) deficiency is an autosomal recessive disorder of the pyrimidine degradation pathway. In a patient presenting with convulsions, psychomotor retardation and Reye like syndrome, strongly elevated levels of uracil and thymine were detected in urine. No DPD activity could be detected in peripheral blood mononuclear cells. Analysis of the gene encoding DPD (DPYD) showed that the patient was homozygous for a novel c.505_513del (p.169_171del) mutation in exon 6 of DPYD.