Suzanne de Bruijn

51 The impact of modern technologies on molecular diagnostics significance. 56,101 However, an important caveat is that a reliable database should be frequently updated, and uploaded sequencing data should adhere to quality control criteria. An example of non-pathogenic variants mistakenly reported as pathogenic has been highlighted in a study performed by Hanany et al. 102 The authors extracted up-to- date allele frequencies from gnomAD of variants in genes associated with dominantly inherited RD and concluded that the pathogenicity of variants in 19% of these genes should be debated. Inherited HL, on the other hand, is a more common condition, than RD and therefore the expected maximum allele frequency for a pathogenic variant should be adjusted accordingly. 103 Once a potentially disease-causing variant is identified, a rich source of available scientific and medical literature can be assessed. A first important step entails thorough comparisons between the observed phenotype in the investigated proband and phenotypic observations described in literature. Most well-described phenotype- genotype correlations can be found in data repositories: Online Mendelian Inheritance in Man (OMIM) 104 , ClinGen 92 and ClinVar. 93 Strong phenotype-genotype correlations are complicated by a phenomenon called allelism: different phenotypes can result from different alleles of the same gene. 105 For example, autosomal recessive Stargardt disease (STGD1), which is due to two variants or alleles in ABCA4 , shows a wide clinical spectrum of maculopathies. 106 The most severe form is early-onset (onset <10 years) STGD1 or panretinal cone-rod dystrophy which is due to two deleterious ABCA4 alleles. Classical or intermediate STGD1 (onset between 10 and 40 years) is due to a combination of a deleterious variant and a mild variant. Finally, late-onset STGD1 (onset >40 years) is caused by a deleterious variant and a mild variant (p.Asn1868Ile) showing reduced penetrance. 106-108 Truncating variants in the CDH23 gene are assumed to cause Usher syndrome type 1D, which consists of HL and retinitis pigmentosa, whereas missense variants cause non-syndromic HL. 109 However, several exceptions to this rule have been reported. 110,111 For pathogenic variants in the USH2A gene that can cause both non-syndromic retinitis pigmentosa and Usher syndrome type IIa, the correlation with missense and truncating variants with the associated phenotypic expression is not always clear, although truncating USH2A variants are more frequently reported in patients diagnosed with a syndromic phenotype. 112,113 Additionally, variants affecting genes that are implicated in ciliopathies (e.g. BBS1, CEP290 , IQCB1 ) can cause a wide range of variable symptoms that are part of a (syndromic) phenotype. Symptoms described for ciliopathies often include retinal degeneration and less frequently HL (reviewed in (9)). To date, >80 forms of syndromic RD have been described which are linked to approximately 200 RD-associated genes 114 ; for syndromic HL, these numbers are suggested to be even higher. 115