Targeted long-read sequencing enriches disease-relevant genomic regions of interest to provide complete Mendelian disease diagnostics.

Despite advances in sequencing technologies, a molecular diagnosis remains elusive in many patients with Mendelian disease. Current short-read clinical sequencing approaches cannot provide chromosomal phase information or epigenetic information without further sample processing, which is not routinely done and can result in an incomplete molecular diagnosis in patients. The ability to provide phased genetic and epigenetic information from a single sequencing run would improve the diagnostic rate of Mendelian conditions.

Substitution of a single non-coding nucleotide upstream of TMEM216 causes non-syndromic retinitis pigmentosa and is associated with reduced TMEM216 expression.

Genome analysis of individuals affected by retinitis pigmentosa (RP) identified two rare nucleotide substitutions at the same genomic location on chromosome 11 (g.61392563 [GRCh38]), 69 base pairs upstream of the start codon of the ciliopathy gene TMEM216 (c.-69G>A, c.

Clinician-Driven Reanalysis of Exome Sequencing Data From Patients With Inherited Retinal Diseases.

Importance: Despite advances in next-generation sequencing (NGS), a significant proportion of patients with inherited retinal disease (IRD) remain undiagnosed after initial genetic testing. Exome sequencing (ES) reanalysis in the clinical setting has been suggested as one method for improving diagnosis of IRD.

Objective: To investigate the association of clinician-led reanalysis of ES data, which incorporates updated clinical information and comprehensive bioinformatic analysis, with the diagnostic yield in a cohort of patients with IRDs in Korea.

Coding and non-coding variants in the ciliopathy gene cause early-onset non-syndromic retinal degeneration.

Inherited retinal degenerations are blinding genetic disorders characterized by high genetic and phenotypic heterogeneity. The implementation of next-generation sequencing in routine diagnostics, together with advanced clinical phenotyping including multimodal retinal imaging, have contributed to the increase of reports describing novel genotype-phenotype associations and phenotypic expansions. In this study, we describe sixteen families with early-onset non-syndromic retinal degenerations in which affected probands carried rare bi-allelic variants in , a ciliary gene previously associated with syndromic recessive Jeune syndrome.

Biallelic variation in the choline and ethanolamine transporter underlies a pleiotropic disease spectrum from adult neurodegeneration to severe developmental disorders.

encodes Feline leukemia virus subgroup C receptor 1 (FLVCR1), a solute carrier (SLC) transporter within the Major Facilitator Superfamily. FLVCR1 is a widely expressed transmembrane protein with plasma membrane and mitochondrial isoforms implicated in heme, choline, and ethanolamine transport. While knockout mice die with skeletal malformations and defective erythropoiesis reminiscent of Diamond-Blackfan anemia, rare biallelic pathogenic variants are linked to childhood or adult-onset neurodegeneration of the retina, spinal cord, and peripheral nervous system.

Non-syndromic Retinal Degeneration Caused by Pathogenic Variants in Joubert Syndrome Genes.

Inherited retinal degenerations (IRDs) are a group of genetic disorders characterized by progressive dysfunction and loss of photoreceptors. IRDs are classified as non-syndromic or syndromic, depending on whether retinal degeneration manifests alone or in combination with other associated symptoms. Joubert syndrome (JBTS) is a genetically and clinically heterogeneous disorder affecting the central nervous system and other organs and tissues, including the neuroretina.

Identification of a novel large multigene deletion and a frameshift indel in as the underlying cause of early-onset recessive rod-cone degeneration.

A family, with two affected identical twins with early-onset recessive inherited retinal degeneration, was analyzed to determine the underlying genetic cause of pathology. Exome sequencing revealed a rare and previously reported causative variant (c.1923_1969delinsTCTGGG; p.

Novel variants causing later-onset non-syndromic retinal dystrophy with macular chorioretinal atrophy.

Background: Variants in were recently described to cause a retinal dystrophy with only eight families described to date and a predominant phenotype of macular atrophy and peripheral reticular degeneration. Here, we further evaluate the genotypic and phenotypic characteristics of biallelic -associated retinal dystrophy in a North American clinic population.

Methods: A retrospective analysis of genetic and clinical features was performed in individuals with biallelic variants in .

Broadening INPP5E phenotypic spectrum: detection of rare variants in syndromic and non-syndromic IRD.

Pathogenic variants in INPP5E cause Joubert syndrome (JBTS), a ciliopathy with retinal involvement. However, despite sporadic cases in large cohort sequencing studies, a clear association with non-syndromic inherited retinal degenerations (IRDs) has not been made. We validate this association by reporting 16 non-syndromic IRD patients from ten families with bi-allelic mutations in INPP5E.

In or Out? New Insights on Exon Recognition through Splice-Site Interdependency.

Noncanonical splice-site mutations are an important cause of inherited diseases. Based on in vitro and stem-cell-based studies, some splice-site variants show a stronger splice defect than expected based on their predicted effects, suggesting that other sequence motifs influence the outcome. We investigated whether splice defects due to human-inherited-disease-associated variants in noncanonical splice-site sequences in , , and could be rescued by strengthening the splice site on the other side of the exon.

Lactoferrin and oral pathologies: a therapeutic treatment.

The oral cavity is a non-uniform, extraordinary environment characterized by mucosal, epithelial, abiotic surfaces and secretions as saliva. Aerobic and anaerobic commensal and pathogenic microorganisms colonize the tongue, teeth, jowl, gingiva, and periodontium. Commensals exert an important role in host defenses, while pathogenic microorganisms can nullify this protective function causing oral and systemic diseases.

A combined RNA-seq and whole genome sequencing approach for identification of non-coding pathogenic variants in single families.

Inherited retinal degenerations (IRDs) are at the focus of current genetic therapeutic advancements. For a genetic treatment such as gene therapy to be successful, an accurate genetic diagnostic is required. Genetic diagnostics relies on the assessment of the probability that a given DNA variant is pathogenic.

Late-Onset Stargardt Disease Due to Mild, Deep-Intronic ABCA4 Alleles.

Purpose: To investigate the role of two deep-intronic ABCA4 variants, that showed a mild splice defect in vitro and can occur on the same allele as the low penetrant c.5603A>T, in Stargardt disease (STGD1).

Methods: Ophthalmic data were assessed of 18 STGD1 patients who harbored c.

ABCA4-associated disease as a model for missing heritability in autosomal recessive disorders: novel noncoding splice, cis-regulatory, structural, and recurrent hypomorphic variants.

Purpose: ABCA4-associated disease, a recessive retinal dystrophy, is hallmarked by a large proportion of patients with only one pathogenic ABCA4 variant, suggestive for missing heritability.

Methods: By locus-specific analysis of ABCA4, combined with extensive functional studies, we aimed to unravel the missing alleles in a cohort of 67 patients (p), with one (p = 64) or no (p = 3) identified coding pathogenic variants of ABCA4.

Results: We identified eight pathogenic (deep-)intronic ABCA4 splice variants, of which five are novel and six structural variants, four of which are novel, including two duplications.

Deep-intronic ABCA4 variants explain missing heritability in Stargardt disease and allow correction of splice defects by antisense oligonucleotides.

Purpose: Using exome sequencing, the underlying variants in many persons with autosomal recessive diseases remain undetected. We explored autosomal recessive Stargardt disease (STGD1) as a model to identify the missing heritability.

Methods: Sequencing of ABCA4 was performed in 8 STGD1 cases with one variant and p.

The Common ABCA4 Variant p.Asn1868Ile Shows Nonpenetrance and Variable Expression of Stargardt Disease When Present in trans With Severe Variants.

Purpose: To assess the occurrence and the disease expression of the common p.Asn1868Ile variant in patients with Stargardt disease (STGD1) harboring known, monoallelic causal ABCA4 variants.

Methods: The coding and noncoding regions of ABCA4 were sequenced in 67 and 63 STGD1 probands respectively, harboring monoallelic ABCA4 variants.

Identification and Rescue of Splice Defects Caused by Two Neighboring Deep-Intronic ABCA4 Mutations Underlying Stargardt Disease.

Sequence analysis of the coding regions and splice site sequences in inherited retinal diseases is not able to uncover ∼40% of the causal variants. Whole-genome sequencing can identify most of the non-coding variants, but their interpretation is still very challenging, in particular when the relevant gene is expressed in a tissue-specific manner. Deep-intronic variants in ABCA4 have been associated with autosomal-recessive Stargardt disease (STGD1), but the exact pathogenic mechanism is unknown.

midigenes reveal the full splice spectrum of all reported noncanonical splice site variants in Stargardt disease.

Stargardt disease is caused by variants in the gene, a significant part of which are noncanonical splice site (NCSS) variants. In case a gene of interest is not expressed in available somatic cells, small genomic fragments carrying potential disease-associated variants are tested for splice abnormalities using in vitro splice assays. We recently discovered that when using small minigenes lacking the proper genomic context, in vitro results do not correlate with splice defects observed in patient cells.

MIB2 variants altering NOTCH signalling result in left ventricle hypertrabeculation/non-compaction and are associated with Ménétrier-like gastropathy.

We performed whole exome sequencing in individuals from a family with autosomal dominant gastropathy resembling Ménétrier disease, a premalignant gastric disorder with epithelial hyperplasia and enhanced EGFR signalling. Ménétrier disease is believed to be an acquired disorder, but its aetiology is unknown. In affected members, we found a missense p.

Photoreceptor Progenitor mRNA Analysis Reveals Exon Skipping Resulting from the ABCA4 c.5461-10T→C Mutation in Stargardt Disease.

Purpose: To elucidate the functional effect of the ABCA4 variant c.5461-10T→C, one of the most frequent variants associated with Stargardt disease (STGD1).

Design: Case series.

Heterozygous deep-intronic variants and deletions in ABCA4 in persons with retinal dystrophies and one exonic ABCA4 variant.

Variants in ABCA4 are responsible for autosomal-recessive Stargardt disease and cone-rod dystrophy. Sequence analysis of ABCA4 exons previously revealed one causative variant in each of 45 probands. To identify the "missing" variants in these cases, we performed multiplex ligation-dependent probe amplification-based deletion scanning of ABCA4.

Mutations in MFSD8, encoding a lysosomal membrane protein, are associated with nonsyndromic autosomal recessive macular dystrophy.

Purpose: This study aimed to identify the genetic defects in 2 families with autosomal recessive macular dystrophy with central cone involvement.

Design: Case series.

Participants: Two families and a cohort of 244 individuals with various inherited maculopathies and cone disorders.

Autosomal dominant Ménétrier-like disease.

Background: Familial occurrence of Ménétrier disease is rare and has been reported only in few instances.

Methods: Affected patients from a large pedigree were evaluated at the clinical, endoscopic, and pathological levels.

Results: Affected members presented with gastropathy of variable severity but without protein loss.