Format

Send to

Choose Destination
Genet Med. 2019 Jan;21(1):53-61. doi: 10.1038/s41436-018-0016-6. Epub 2018 Aug 13.

Development of an evidence-based algorithm that optimizes sensitivity and specificity in ES-based diagnostics of a clinically heterogeneous patient population.

Author information

1
Centogene AG, Rostock, Germany. peter.bauer@centogene.com.
2
Centogene AG, Rostock, Germany.
3
Albrecht-Kossel-Institute for Neuroregeneration, Medical University Rostock, Rostock, Germany.

Abstract

PURPOSE:

Next-generation sequencing (NGS) is rapidly replacing Sanger sequencing in genetic diagnostics. Sensitivity and specificity of NGS approaches are not well-defined, but can be estimated from applying NGS and Sanger sequencing in parallel. Utilizing this strategy, we aimed at optimizing exome sequencing (ES)-based diagnostics of a clinically diverse patient population.

METHODS:

Consecutive DNA samples from unrelated patients with suspected genetic disease were exome-sequenced; comparatively nonstringent criteria were applied in variant calling. One thousand forty-eight variants in genes compatible with the clinical diagnosis were followed up by Sanger sequencing. Based on a set of variant-specific features, predictors for true positives and true negatives were developed.

RESULTS:

Sanger sequencing confirmed 81.9% of ES-derived variants. Calls from the lower end of stringency accounted for the majority of the false positives, but also contained ~5% of the true positives. A predictor incorporating three variant-specific features classified 91.7% of variants with 100% specificity and 99.75% sensitivity. Confirmation status of the remaining variants (8.3%) was not predictable.

CONCLUSIONS:

Criteria for variant calling in ES-based diagnostics impact on specificity and sensitivity. Confirmatory sequencing for a proportion of variants, therefore, remains a necessity. Our study exemplifies how these variants can be defined on an empirical basis.

KEYWORDS:

Genetic testing; Laboratory standards; Sensitivity; Specificity; exome sequencing

PMID:
30100613
DOI:
10.1038/s41436-018-0016-6

Supplemental Content

Full text links

Icon for Nature Publishing Group
Loading ...
Support Center