Autosomal recessive hereditary spastic paraplegia type SPG35 due to a novel variant in the FA2H gene in a Czech patient.

Biallelic pathogenic variants in FA2H gene have been repeatedly described as a cause of hereditary spastic paraplegia (HSP) type35 (SPG35). Targeted massive parallel sequencing (MPS) of the HSP genes panel revealed a novel homozygous variant c.130C > T (p.P44S) in the FA2H gene in the 30-year-old patient presenting with spastic paraplegia. The patient originated form the Czech minority in Romania. The patient manifests typical clinical signs for SPG35 (youth onset gait impairment, progressive spastic paraparesis on lower limbs, dysarthria, white matter changes in MRI).

Genome-wide uniparental diploidy of all paternal chromosomes in an 11-year-old girl with deafness and without malignancy.

Approximately 20 cases of genome-wide uniparental disomy or diploidy (GWUPD) as mosaicism have previously been reported. We present the case of an 11-year-old deaf girl with a paternal uniparental diploidy or isodisomy with a genome-wide loss of heterozygosity (LOH). The patient was originally tested for non-syndromic deafness, and the novel variant p.V234I in the ESRRB gene was found in a homozygous state. Our female proband is the seventh patient diagnosed with GWUPD at a later age and is probably the least affected of the seven, as she has not yet presented any malignancy. Most, if not all, reported patients with GWUPD whose clinical details have been published have developed malignancy, and some of those patient developed malignancy several times. Therefore, our patient has a high risk of malignancy and is carefully monitored by a specific outpatient pediatric oncology program. This observation seems to be novel and unique in a GWUPD patient. Our study is also unique as it not only provides very detailed documentation of the genomic situations of various tissues but also reports differences in the mosaic ratios between the blood and saliva, as well as a normal biparental allelic situation in the skin and biliary duct. Additionally, we were able to demonstrate that the mosaic ratio in the blood remained stable even after 3 years and has not changed over a longer period.

Diagnóstico genético preimplantacional de la enfermedad de Charcot-Marie-Tooth ligada al cromosoma X mediante análisis de ligamiento indirect / Preimplantation genetic diagnosis of X-linked Charcot-Marie-Tooth disease by indirect linkage analysis

Objetivo: Presentar un enfoque metodológico del diagnóstico genético preimplantacional (DGP) como opción para embarazos no afectados en parejas en edad reproductiva con riesgo genético de neuropatía periférica dominante por enfermedad de Charcot-Marie-Tooth tipo 1 ligada al cromosoma X. Pacientes y métodos: Llevamos a cabo el DGP de enfermedad de Charcot-Marie-Tooth tipo 1 ligada al cromosoma X utilizando un análisis de ligamiento indirecto/haplotificación, durante el cual logramos excluir los embriones portadores de un haplotipo de alto riesgo ligado a la mutación causal p.Leu9Phe en el gen GJB1. Resultados: Dentro del ciclo de DGP, examinamos 4 blastómeros biopsiados de los embriones en fase de división, y recomendamos la transferencia de 3 embriones. Dos embriones fueron implantados en el útero; sin embargo, el resultado fue un embarazo único con un descendiente varón. Transcurridos 3 años, la pareja regresó con un embarazo espontáneo. La biopsia coriónica de este embarazo reveló el sexo femenino y una inversión pericéntrica del cromosoma 5 en el 70% de las células fetales cultivadas. Conclusión: Utilizando el análisis de ligamiento indirecto, el DGP puede ayudar a identificar durante el cribado los defectos genéticos ligados al cromosoma X, eludiendo por tanto los problemas potenciales con el aborto (AU)

Objective: To present methodical approach of preimplantation genetic diagnosis (PGD) as an option for an unaffected pregnancy in reproductive-age couples who have a genetic risk of the X-linked dominant peripheral neuropathy Charcot-Marie-Tooth type 1 disease. Patients and methods: We performed PGD of X-linked Charcot-Marie-Tooth type 1 disease using haplotyping/indirect linkage analysis, when during analysis we reach to exclude embryos that carry a high-risk haplotype linked to the causal mutation p.Leu9Phe in the GJB1 gene. Results: Within the PGD cycle, we examined 4 blastomeres biopsied from cleavage-stage embryos and recommended 3 embryos for transfer. Two embryos were implanted into the uterus; however, it resulted in a singleton pregnancy with a male descendant. Three years later, the couple returned again with spontaneous gravidity. A chorionic biopsy examination of this gravidity ascertained the female sex and a pericentric inversion of chromosome 5 in 70% of the cultivated foetal cells. Conclusion: Using indirect linkage analysis, PGD may help to identify genetic X-linked defects within embryos during screening, thereby circumventing the potential problems with abortion (AU)

Preimplantation genetic diagnosis of X-linked Charcot-Marie-Tooth disease by indirect linkage analysis. / Diagnóstico genético preimplantacional de la enfermedad de Charcot-Marie-Tooth ligada al cromosoma X mediante análisis de ligamiento indirecto.

OBJECTIVE: To present methodical approach of preimplantation genetic diagnosis (PGD) as an option for an unaffected pregnancy in reproductive-age couples who have a genetic risk of the X-linked dominant peripheral neuropathy Charcot-Marie-Tooth type 1 disease. PATIENTS AND METHODS: We performed PGD of X-linked Charcot-Marie-Tooth type 1 disease using haplotyping/indirect linkage analysis, when during analysis we reach to exclude embryos that carry a high-risk haplotype linked to the causal mutation p.Leu9Phe in the GJB1 gene. RESULTS: Within the PGD cycle, we examined 4 blastomeres biopsied from cleavage-stage embryos and recommended 3 embryos for transfer. Two embryos were implanted into the uterus; however, it resulted in a singleton pregnancy with a male descendant. Three years later, the couple returned again with spontaneous gravidity. A chorionic biopsy examination of this gravidity ascertained the female sex and a pericentric inversion of chromosome 5 in 70% of the cultivated foetal cells. CONCLUSION: Using indirect linkage analysis, PGD may help to identify genetic X-linked defects within embryos during screening, thereby circumventing the potential problems with abortion.

OmniPlex--a new QF-PCR assay for prenatal diagnosis of common aneuploidies based on evaluation of the heterozygosity of short tandem repeat loci in the Czech population.

OBJECTIVES: The aim of our study was to assess the utility of commonly used multiplex assays of short tandem repeat markers used for quantitative fluorescent polymerase chain reaction (QF-PCR) for prenatal rapid aneuploidy detection (RAD) in routine prenatal diagnosis in the Czech population. METHODS: Two previously published RAD multiplexes (2M test) were tested on 2906 local prenatal samples and used to calculate the rates of heterozygosity within this population. Most of the markers used in both multiplexes were highly informative. However, some had little utility, either due to a low heterozygosity rate (D21S499, D18S978 and P39) or because they were difficult to evaluate (DXS1283E). RESULTS: After evaluation of the 2M test results, a new multiplex assay (OmniPlex) was designed, developed and tested on 960 samples. This new assay was evaluated for heterozygosity rates and for the probability of having two or more informative markers on each chromosome. CONCLUSIONS: OmniPlex assay significantly improved the QF-PCR methodology for rapid prenatal aneuploidy detection in the Czech population. Based on detected heterozygosity of markers used for QF-PCR in this population, OmniPlex is a robust assay for the detection of chromosomes 13, 18, 21, X and Y in a single reaction.

QF-PCR-based prenatal detection of common aneuploidies in the Czech population: five years of experience.

We present the results from the largest clinical application of QF-PCR for antenatal rapid aneuploidy detection (RAD) in routine prenatal diagnosis in the Czech Republic. QF-PCR was performed in addition to karyotyping (dual testing) in two settings: the first was a single multiplex reaction testing only trisomy 21 and amelogenin X/Y alleles in the second trimester screened positive cases (T21 test), and the second setting consisted of two multiplexes (2M test) for common aneuploidies (13, 18, 21, X and Y) in cases with other RAD indications such as ultrasound findings, late booking or maternal anxiety. Dual testing was performed in 6349/12,778 (49.7%) of prenatal samples using either T21 or 2M test between 2002 and 2007. The clinical acceptability of our dual testing policy, methodological efficiency of RAD and residual risks of other chromosomal aberrations (CHAs) were evaluated. QF-PCR detected 92% (175/190) of significant CHAs. The 2M test identified 93.5% and the T21 test identified 87.5% of the significant CHAs with complete specificity. The residual risk of significant CHA was 1/231 in the 2M test and 1/565 in the T21 test. If RAD for all common aneuploidies is used as the sole prenatal diagnosis method, the odds of missing a CHA of any type are 1:90 and the odds of missing significant CHA with no ultrasound findings are 1:1513. If prenatal karyotyping were used as an additional procedure to RAD in cases only with ultrasound findings, 186/190 (97.8%) of the significant CHAs would be detected when 15.7% cases were karyotyped, according to our data. We consider RAD directed towards trisomy 21 alone (our T21 test) as an economically and clinically acceptable part of second trimester screening for Down syndrome. Both RAD tests allow fast alleviation of maternal anxiety with low residual risk when the test results are negative, and allow fast decision making if the results are positive. However, replacement of dual testing with only the RAD procedure in specific indications accepted in some countries (Great Britain) remains in the Czech Republic a theme for debate.