Objectives: Diagnosis of Wilson disease (WD) is difficult and, as early detection may prevent all symptoms, it is essential to know the exact prevalence to evaluate the cost-efficacy of a screening program. As the number of WD patients was high in our population, we wished to estimate prevalence by determining the carrier frequency for clinically relevant ATP7B mutations. Methods: To estimate prevalence, screening for the most prevalent mutation was performed in 1661 individuals with ancestry in Gran Canaria, and the frequency of other mutations was estimated from patient records. Alternatively, ATP7B mutations were detected from exomes and genomes from 851 individuals with Canarian ancestry, 236 from Gran Canaria, and a public Spanish exome database. Results: Estimated carrier frequencies in Gran Canaria ranged from 1 in 20 to 28, depending on the method used, resulting in prevalences of 1 case per 1547 to 3140 inhabitants. Alternatively, the estimated affected frequencies were 1 in 5985 to 7980 and 1 in 6278 to 16,510 in the archipelago or mainland Spain respectively. Conclusions: The number of carriers predicts much higher prevalences than reported, suggesting that WD is underdiagnosed; specific mutations may remain unnoticed due to low penetrance or no signs of disease at all; regional prevalence rather than national prevalence should be considered in cost-efficacy models to approach preventive screening in the asymptomatic population and genetic screening strategies will have to deal with the genetic heterogeneity of ATP7B in the general population and in patients.

A hurdle of exome sequencing is its limited capacity to represent the entire exome. To ascertain the diagnostic power of this approach we determined the extent of coverage per individual sample. Using alignment data (BAM files) from 15 exome samples, sequences of any length that were below a determined sequencing depth coverage (DP) were detected and annotated with the Ensembl exon database using MIST, a novel software tool. Samples sequenced at 50X mean coverage had, on average, up to 50% of the Ensembl annotated exons with at least one nucleotide (L=1) with a DP<20, improving to 35% at 100X mean coverage. In addition, almost 15% of annotated exons were never sequenced (L=50, DP<1) at 50x mean coverage, reaching down to 5% at 100x. The diagnostic utility of this approach was tested for hypertrophic cardiomyopathy, a genetically heterogeneous disease, where exome sequencing covered as much as 80% of all candidate genes exons at DP≥20. This report stresses the value of identifying, precisely, which sequences are below a specific depth in an individual´s exome, and provides a useful tool to assess the potential and pitfalls of exome sequencing in a diagnostic or gene discovery setting.