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Prenatal genetic diagnosis of monogenic diseases is a process involving the use of a variety of molecular techniques for the molecular characterization of a potential monogenic disease in the fetus during pregnancy. Prenatal genetic diagnosis can be performed through invasive and non-invasive methods. A distinction must be made between “NIPD” (non-invasive prenatal diagnosis), which is considered to be diagnostic, from “NIPT” (non-invasive prenatal test), which is a screening test that requires subsequent confirmation by invasive methods. The different techniques currently available aim at detecting either, previously characterized pathogenic mutations in the family, the risk haplotype associated with the familial mutation, or potential pathogenic mutation(s) in a gene associated with a diagnostic suspicion. An overview is provided of relevant aspects of prenatal genetic diagnosis of monogenic diseases. The objective of this paper is to describe the main molecular techniques currently available and used in clinical practice. A description is provided of the indications, limitations and analytical recommendations regarding these techniques, and the standards governing genetic counseling. Continuous rapid advances in the clinical applications of genomics have provided increased access to comprehensive molecular characterization. Laboratories are struggling to keep in pace with technology developments.

El diagnóstico genético prenatal de enfermedades monogénicas es un proceso que engloba el conjunto de técnicas moleculares dirigidas a caracterizar molecularmente una posible enfermedad monogénica en el feto durante el embarazo. Actualmente, el diagnóstico genético prenatal puede realizarse de manera invasiva o no invasiva. Debemos diferenciar “NIPD” (del inglés: non invasive prenatal diagnosis) que se considera diagnóstico de “NIPT” (del inglés: non invasive prenatal test) que se considera prueba de cribado y tendrá que confirmarse por técnicas invasivas. Las diferentes metodologías disponibles y empleadas pueden ir dirigidas a la detección de variante/s genética/s causal/es previamente caracterizada/s en la familia, la detección del haplotipo de riesgo asociado a la mutación familiar y/o la búsqueda de posibles variante/s patogénicas en un gen asociado a una sospecha diagnóstica. Se revisarán los aspectos relevantes del diagnóstico genético prenatal de las enfermedades monogénicas. El objetivo es la exposición de las principales técnicas moleculares disponibles y empleadas en la actualidad, detallando sus indicaciones, limitaciones y recomendaciones analíticas, así como la normativa que regula el asesoramiento genético. La evolución permanente y exponencial en la aplicación asistencial de las técnicas genómicas, facilita progresivamente el acceso a una caracterización molecular exhaustiva, obligando a una constante actualización homogénea de los laboratorios.

Pathogenic variants in the AP4B1 gene lead to a rare form of hereditary spastic paraplegia (HSP) known as SPG47. We report on a patient with a clinical suspicion of complicated HSP of the lower limbs with intellectual disability, as well as a novel homozygous noncanonical splice site variant in the AP4B1 gene, in which the effect on splicing was validated by RNA analysis. We sequenced 152 genes associated with HSP using Next‐Generation Sequencing (NGS). We isolated total RNA from peripheral blood and generated cDNA using reverse transcription‐polymerase chain reaction (RT‐PCR). A region of AP4B1 mRNA was amplified by PCR and the fragments obtained were purified from the agarose gel and sequenced. We found a homozygous variant of uncertain significance in the AP4B1 gene NM_006594.4: c.1511‐6C>G in the proband. Two different AP4B1 mRNA fragments were obtained in the patient and his carrier parents. The shorter fragment was the predominant fragment in the patient and revealed a deletion with skipping of the AP4B1 exon 10. The patient's longer fragment corresponded to an insertion of the last five nucleotides of AP4B1 intron 9. We confirmed that this variant affects the normal splicing of RNA, sustaining the molecular diagnosis of SPG47 in the patient. [ABSTRACT FROM AUTHOR]

Background Osteogenesis imperfecta ( OI) is a heterogeneous bone disorder characterized by recurrent fractures. Although most cases of OI have heterozygous mutations in COL1A1 or COL1A2 and show autosomal dominant inheritance, during the last years there has been an explosion in the number of genes responsible for both recessive and dominant forms of this condition. Herein, we have analyzed a cohort of patients with OI, all offspring of unaffected parents, to determine the spectrum of variants accounting for these cases. Twenty patients had nonrelated parents and were sporadic, and 21 were born to consanguineous relationships. Methods Mutation analysis was performed using a next-generation sequencing gene panel, homozygosity mapping, and whole exome sequencing ( WES). Results Patients offspring of nonconsanguineous parents were mostly identified with COL1A1 or COL1A2 heterozygous changes, although there were also a few cases with IFITM5 and WNT1 heterozygous mutations. Only one sporadic patient was a compound heterozygote for two recessive mutations. Patients offspring of consanguineous parents showed homozygous changes in a variety of genes including CRTAP, FKBP10, LEPRE1, PLOD2, PPIB, SERPINF1, TMEM38B, and WNT1. In addition, two patients born to consanguineous parents were found to have de novo COL1A1 heterozygous mutations demonstrating that causative variants in the collagen I structural genes cannot be overlooked in affected children from consanguineous couples. Further to this, WES analysis in probands lacking mutations in OI genes revealed deleterious variants in SCN9A, NTRK1, and SLC2A2, which are associated with congenital indifference to pain ( CIP) and Fanconi-Bickel syndrome ( FBS). Conclusion This work provides useful information for clinical and genetic diagnosis of OI patients with no positive family history of this disease. Our data also indicate that CIP and FBS are conditions to be considered in the differential diagnosis of OI and suggest a positive role of SCN9A and NTRK1 in bone development. [ABSTRACT FROM AUTHOR]