539Mendelian diseases are considered to be rare, yet genetic disorders are estimated
to occur at a rate of 40–82 per 1,000 live births. Epidemiologic studies show that if
all congenital anomalies are considered as part of the genetic load, then ~8 % of per-
sons are identifi ed as having a genetic disorder before reaching adulthood. Most of
these cases remain undiagnosed. Genomic sequencing with the use of massively par-
allel NGS is an effective alternative to locus-specifi c and gene-panel tests in a
research setting for establishing a genetic basis of disease. Initial applications of NGS
to clinical diagnosis posed many challenges. Technical, bioinformatic, interpretive,
and validation pipelines have been developed for whole-exome sequencing (WES) in
a certifi ed clinical laboratory to identify sequence variants underlying disease pheno-
types in patients (Yang et al. 2013 ). Application of WES to the diagnoses of 250
unselected consecutive patients resulted in a molecular diagnostic yield of 25 %,
which is higher than the positive rates of other genetic tests, such as karyotype analy-
sis (5–15 %), chromosomal microarray analysis (15–20 %), and Sanger sequencing
for single genes (3–15 %). Among the 500 additional clinical exomes completed dur-
ing the review process, the authors obtained a similar diagnostic yield (26 %). Results
of this study support the use of WES as a diagnostic test for patients with nonspecifi c
or unusual disease presentations of possible genetic cause and for patients with clini-
cal diagnoses of heterogeneous genetic conditions. Cost-effectiveness, accuracy,
yield, and integration of genome-based diagnosis in medical care must be addressed
in future studies and will require prospective study designs. Although this seems logi-
cal, a prospective study design involving a million variants per person, long-term
follow-up periods for proving effectiveness, and suffi cient power to test whether
knowledge of any given variant has an effect on clinical outcome would be a chal-
lenge with traditional randomized, case control designs. Appropriate study designs
that can distinguish pathogenic from benign variants and test the effect of genetic
knowledge on clinical outcomes require serious consideration (Jacob 2013 ).
DNA Sequencing for Prenatal Disorders
In high-risk pregnant women, noninvasive prenatal testing with the use of massively
parallel sequencing of maternal plasma cell-free DNA (cfDNA) accurately detects
fetal autosomal aneuploidy. In a blinded multicenter study in the US, blood samples
from women with singleton pregnancies who were undergoing standard aneuploidy
screening (serum biochemical assays with or without nuchal translucency measure-
ment) were collected and massively parallel sequencing was performed to deter-
mine the chromosome dosage for each sample (Bianchi et al. 2014 ). The results
showed that prenatal testing with use of cfDNA had signifi cantly lower false posi-
tive rates and higher positive predictive values for detection of trisomies 21 and 18
than standard screening with ultrasound and biochemical blood tests. Positive results
from standard screening methods, such as ultrasound and testing the mother’s blood
for proteins associated with fetal deformities, accurately point to Down syndrome in
only about 4 % of cases. And if one of those screens indicates a problem, invasive
methods, such as amniocentesis, must be performed to substantiate the results.
Sequencing in Genetic Disorders