Primary periodic paralysis (PPP) is an inherited disorders of ion channel dysfunction characterized by recurrent episodes of flaccid muscle weakness, which can classified as hypokalemic (HypoPP), ...normokalemic (NormoPP), or hyperkalemic (HyperPP) according to the potassium level during the paralytic attacks. However, PPP is charactered by remarkable clinical and genetic heterogeneity, and the diagnosis of suspected patients is based on the characteristic clinical presentation then confirmed by genetic testing. At present, there are only limited cohort studies on PPP in the Chinese population.
We included 37 patients with a clinical diagnosis of PPP. Eleven (29.7%) patients were tested using a specific gene panel and 26 (70.3%) by the whole-exome sequencing (WES). Twenty-two cases had a genetic variant identified, representing a diagnostic rate of 59.5% (22/37). All the identified mutations were either in the SCN4A or the CACNA1S gene. The overall detection rate was comparable between the panel (54.5%: 6/11) and WES (61.5%: 16/26). The remaining patients unresolved through panel sequencing were further analyzed by WES, without the detection of any mutation. The novel atypical splicing variant c.2020-5G > A affects the normal splicing of the SCN4A mRNA, which was confirmed by minigene splicing assay. Among 21 patients with HypoPP, 15 patients were classified as HypoPP-2 with SCN4A variants, and 6 HypoPP-1 patients had CACNA1S variants.
Our results suggest that SCN4A alleles are the main cause in our cohort, with the remainder caused by CACNA1S alleles, which are the predominant cause in Europe and the United States. Additionally, this study identified 3 novel SCN4A and 2 novel CACNA1S variants, broadening the mutation spectrum of genes associated with PPP.
The next generation sequencing (NGS) based non-invasive prenatal test (NIPT) has outplayed the traditional serum biochemical tests (SBT) in screen of fetal aneuploidies with a high sensitivity and ...specificity. However, it has not been widely used as a primary screen tool due to its high cost and the cheaper SBT is still the choice for primary screen even with well-known shortages in sensitivity and specificity. Here, we report a multiplex droplet digital PCR NIPT (dPCR-NIPT) assay that can detect trisomies 21, 18 and 13 (T21, T18 and T13) in a single tube reaction with a better sensitivity and specificity than the SBT and a much cheaper price than the NGS-NIPT.
In this study, the dPCR-NIPT assay's non-clinical characteristics were evaluated to verify the cell free fetal DNA (cffDNA) fraction enrichment efficiencies, the target cell free DNA (cfDNA) concentration enrichment, the analytical sensitivity, and the sample quality control on the minimum concentration of cfDNA required for the assay. We validated the clinical performance for this assay by blindly testing 283 clinical maternal plasma samples, including 36 trisomic positive samples, from high risk pregnancies to access its sensitivity and specificity. The cost effectiveness of using the dPCR-NIPT assay as the primary screen tool was also analyzed and compared to that of the existing contingent strategy (CS) using the SBT as the primary screen tool and the strategy of NGS-NIPT as the first-tier screen tool in a simulating situation.
For the non-clinical characteristics, the sample processing reagents could enrich the cffDNA fraction by around 2 folds, and the analytical sensitivity showed that the assay was able to detect trisomies at a cffDNA fraction as low as 5% and the extracted cfDNA concentration as low as 0.2 ng/μL. By testing the 283 clinical samples, the dPCR-NIPT assay demonstrated a detection sensitivity of 100% and a specificity of 95.12%. Compared to the existing CS and the NGS-NIPT as the first-tier screen strategy, dPCR-NIPT assay used as a primary screen tool followed by the NGS-NIPT rescreen is the most economical approach to screen pregnant women for fetal aneuploidies without sacrificing the positive detection rate.
This is the first report on a dPCR-NIPT assay, consisting of all the necessary reagents from sample processing to multiplex dPCR amplification, can detect T21, T18 and T13 in a single tube reaction. The study results reveal that this assay has a sensitivity and specificity superior to the SBT and a cost much lower than the NGS-NIPT. Thus, from both the test performance and the economic benefit points of views, using the dPCR-NIPT assay to replace the SBT as a primary screen tool followed by the NGS-NIPT rescreen would be a better approach than the existing CS for detection of fetal aneuploidies in maternal plasma.
Leber congenital amaurosis (LCA) and juvenile retinitis pigmentosa (RP) are inherited retinal diseases that cause early onset severe visual impairment. An accurate molecular diagnosis can refine the ...clinical diagnosis and allow gene specific treatments.
We developed a capture panel that enriches the exonic DNA of 163 known retinal disease genes. Using this panel, we performed targeted next generation sequencing (NGS) for a large cohort of 179 unrelated and prescreened patients with the clinical diagnosis of LCA or juvenile RP. Systematic NGS data analysis, Sanger sequencing validation, and segregation analysis were utilised to identify the pathogenic mutations. Patients were revisited to examine the potential phenotypic ambiguity at the time of initial diagnosis.
Pathogenic mutations for 72 patients (40%) were identified, including 45 novel mutations. Of these 72 patients, 58 carried mutations in known LCA or juvenile RP genes and exhibited corresponding phenotypes, while 14 carried mutations in retinal disease genes that were not consistent with their initial clinical diagnosis. We revisited patients in the latter case and found that homozygous mutations in PRPH2 can cause LCA/juvenile RP. Guided by the molecular diagnosis, we reclassified the clinical diagnosis in two patients.
We have identified a novel gene and a large number of novel mutations that are associated with LCA/juvenile RP. Our results highlight the importance of molecular diagnosis as an integral part of clinical diagnosis.
Leber congenital amaurosis (LCA) is a blinding retinal disease that presents within the first year after birth. Using exome sequencing, we identified mutations in the nicotinamide adenine ...dinucleotide (NAD) synthase gene NMNAT1 encoding nicotinamide mononucleotide adenylyltransferase 1 in eight families with LCA, including the family in which LCA was originally linked to the LCA9 locus. Notably, all individuals with NMNAT1 mutations also have macular colobomas, which are severe degenerative entities of the central retina (fovea) devoid of tissue and photoreceptors. Functional assays of the proteins encoded by the mutant alleles identified in our study showed that the mutations reduce the enzymatic activity of NMNAT1 in NAD biosynthesis and affect protein folding. Of note, recent characterization of the slow Wallerian degeneration (Wld(s)) mouse model, in which prolonged axonal survival after injury is observed, identified NMNAT1 as a neuroprotective protein when ectopically expressed. Our findings identify a new disease mechanism underlying LCA and provide the first link between endogenous NMNAT1 dysfunction and a human nervous system disorder.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Background
Oligohydramnios or polyhydramnios, is associated with chromosomal aberrations, particularly aneuploidy. However, its correlation with copy number variation (CNV) remains unclear.
Methods
...We retrospectively analyzed 428 cases with an abnormal level of amniotic fluid, comprising of 139 cases of single ultrasound findings (SU group) and 289 cases of multiple ultrasound findings (MU group), by CNV sequencing (CNV‐Seq) and followed their pregnancy outcomes.
Results
The overall detection rate of clinically significant findings was 8%, with 5% in the SU group and 11% in MU group. Besides, 18 microdeletion/microduplication syndromes were detected, with the highest rate of renal cysts and diabetes syndrome (22%, 4/18). Also, the rate of termination of pregnancy in MU group was much higher than that in the SU group (29% vs. 10%, ***p < 0.001), and in the MU‐oligohydramnios subgroup, it was the highest (34%), regardless of cases with chromosomal anomaly and lost to follow‐up.
Conclusion
Our results showed that the abnormal level of amniotic fluid, especially combined with other ultrasound abnormalities, is closely related to chromosomal abnormalities and genetic CNVs. CNV‐Seq may be useful in investigating pregnancies with an abnormal amniotic fluid level.
The overall detection rate of clinically significant findings was 8%, with 5.0% in the isolated group and 11.0% in non‐isolated group.
Non‐isolated group especially oligohydramnios subgroup is inclined to obtained more pathogenicity genetic anomalies and to terminate their pregnancy outcome.
To identify the cause of retinitis pigmentosa (RP) in UTAD003, a large, six-generation Louisiana family with autosomal dominant retinitis pigmentosa (adRP).
A series of strategies, including ...candidate gene screening, linkage exclusion, genome-wide linkage mapping, and whole-exome next-generation sequencing, was used to identify a mutation in a novel disease gene on chromosome 10q22.1. Probands from an additional 404 retinal degeneration families were subsequently screened for mutations in this gene.
Exome sequencing in UTAD003 led to identification of a single, novel coding variant (c.2539G>A, p.Glu847Lys) in hexokinase 1 (HK1) present in all affected individuals and absent from normal controls. One affected family member carries two copies of the mutation and has an unusually severe form of disease, consistent with homozygosity for this mutation. Screening of additional adRP probands identified four other families (American, Canadian, and Sicilian) with the same mutation and a similar range of phenotypes. The families share a rare 450-kilobase haplotype containing the mutation, suggesting a founder mutation among otherwise unrelated families.
We identified an HK1 mutation in five adRP families. Hexokinase 1 catalyzes phosphorylation of glucose to glucose-6-phosphate. HK1 is expressed in retina, with two abundant isoforms expressed at similar levels. The Glu847Lys mutation is located at a highly conserved position in the protein, outside the catalytic domains. We hypothesize that the effect of this mutation is limited to the retina, as no systemic abnormalities in glycolysis were detected. Prevalence of the HK1 mutation in our cohort of RP families is 1%.
•Diagnostic yield of WES and mitochondrial genome assessment was studied.•Proband-only WES provided molecular diagnosis for a large cohort of patients.•WES simultaneously analyzed SNVs, exons, ...mitochondrial genomes, and CNVs.•It improved diagnostic yield and could facilitate the identification of novel genes.
Whole exome sequencing (WES) is becoming more widely used as a diagnostic tool in the field of medicine. In this article, we reported the diagnostic yield of WES and mitochondrial genome assessment in 2226 consecutive cases in a single clinical laboratory.
We retrospectively analyzed consecutive WES reports from 2226 patients with various genetic disorders. WES-process was focused exclusively on the probands and aimed at a higher diagnostic capacity. We determined the diagnostic rate of WES overall and by phenotypic category, mode of inheritance, mitochondrial genome variant, and copy number variants (CNVs).
Among the 2226 patients who had diagnostic WES proband-only, the overall diagnostic yield of WES was 34.59% (770/2226). The highest diagnostic yield was observed in autosomal dominant disorders, at 45.58% (351/770), followed by autosomal recessive at 31.95%(246/770), X-linked disorder at 9.61%(74/770), and mitochondrial diseases at a notably lower 0.65%(5/770). The 12.21% (94/770) diagnoses were based on a total of 94 copy number variants reported from WES data. CNVs in children accounted for 67.02% of the total CNVs. While majority of the molecular diagnoses were related to nuclear genes, the inclusion of mitochondrial genome sequencing in the WES test contributed to five diagnoses. all mitochondrial diseases were identified in adults.
The proband-only WES provided a definitive molecular diagnosis for 34.59% of a large cohort of patients while analysis of WES simultaneously analyzed the SNVs, exons, mitochondrial genome, and CNVs, thereby improving the diagnostic yield significantly compared to the single-detection WES method; and facilitating the identification of novel candidate genes.
Abstract Objective We aimed to investigate the yield of prenatal exome sequencing (pES) in morphologically normal fetuses. Method This retrospective study analyzed 254 families with morphologically ...normal fetuses who underwent prenatal trio exome sequencing based on parental request between September 2020 and October 2023. Results Overall, abnormal findings were detected in 8 families (3.1%, 8/254) by pES. Among these, 6 families (2.3%, 6/254) were found to have fetuses affected with monogenic disorders (2 autosomal recessive conditions and 4 autosomal dominant conditions), while 2 families (0.8%, 2/254) were incidentally found to be couples at risk of having a future pregnancy with a recessive condition. Among the six fetuses detected with monogenic disorders, two fetuses carried a de novo variant in OPA1 and NF1 , which are known to cause Optic atrophy 1 and Neurofibromatosis, respectively. One fetus was detected with a maternally inherited variant in PKD2 related to polycystic kidney disease 2 (not known to the mother until then). One fetus was detected with a maternally inherited variant in SDHB associated with Pheochromocytoma. Two fetuses carried compound heterozygous variants in NAGLU and GJB2 associated with Mucopolysaccharidosis type IIIB and Deafness, respectively. In the 2 families where parents were found to be carriers but the fetuses were unaffected, heterozygous variants in the GJB2 and SERPINB7 genes were detected in the parents, respectively, which are associated with deafness and palmoplantar keratoderma. Conclusion Our research indicated that pES can provide significant critical information for families with morphologically normal fetuses. Prenatal screening with exome sequencing requires careful management and detailed pre‐test and post‐test genetic counseling.
Key points What is already known about this topic? Previous studies have demonstrated the increased diagnostic yield and significant clinical impacts of prenatal exome sequencing (pES) for structurally abnormal fetuses. However, few studies have published on the yield of pES for morphologically normal fetuses. What does this study add? The yield of pES in morphologically normal fetuses was found to be 3.1%. Prenatal ES can provide significant critical information for morphologically normal fetuses but detailed pre‐test and post‐test genetic counseling is necessary.
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