👤 Thomas Rehm

As DNA sequencing costs decline, genetic testing options have expanded. Whole exome sequencing and whole genome sequencing (WGS) are entering clinical use, posing questions about their incremental value compared with disease-specific multigene panels that have been the cornerstone of genetic testing. Forty-one patients with hypertrophic cardiomyopathy who had undergone targeted hypertrophic cardiomyopathy genetic testing (either multigene panel or familial variant test) were recruited into the MedSeq Project, a clinical trial of WGS. Results from panel genetic testing and WGS were compared. In 20 of 41 participants, panel genetic testing identified variants classified as pathogenic, likely pathogenic, or uncertain significance. WGS identified 19 of these 20 variants, but the variant detection algorithm missed a pathogenic 18 bp duplication in myosin binding protein C ( WGS detected nearly all variants identified on panel testing, provided 1 new diagnostic finding, and allowed interrogation of posited disease genes. Several variants of uncertain clinical use and numerous secondary genetic findings were also identified. Whereas panel testing and WGS provided similar diagnostic yield, WGS offers the advantage of reanalysis over time to incorporate advances in knowledge, but requires expertise in genomic interpretation to appropriately incorporate WGS into clinical care. URL: https://clinicaltrials.gov. Unique identifier: NCT01736566. Show less

Familial involvement is common in dilated cardiomyopathy (DCM) and >40 genes have been implicated in causing disease. However, the role of genetic testing in clinical practice is not well defined. We examined the experience of clinical genetic testing in a diverse DCM population to characterize the prevalence and predictors of gene mutations. We studied 264 unrelated adult and pediatric DCM index patients referred to 1 reference lab for clinical genetic testing. Up to 10 genes were analyzed (MYH7, TNNT2, TNNI3, TPM1, MYBPC3, ACTC, LMNA, PLN, TAZ, and LDB3), and 70% of patients were tested for all genes. The mean age was 26.6 ± 21.3 years, and 52% had a family history of DCM. Rigorous criteria were used to classify DNA variants as clinically relevant (mutations), variants of unknown clinical significance (VUS), or presumed benign. Mutations were found in 17.4% of patients, commonly involving MYH7, LMNA, or TNNT2 (78%). An additional 10.6% of patients had VUS. Genetic testing was rarely positive in older patients without a family history of DCM. Conversely in pediatric patients, family history did not increase the sensitivity of genetic testing. Using rigorous criteria for classifying DNA variants, mutations were identified in 17% of a diverse group of DCM index patients referred for clinical genetic testing. The low sensitivity of genetic testing in DCM reflects limitations in both current methodology and knowledge of DCM-associated genes. However, if mutations are identified, genetic testing can help guide family management. Show less

The myosin-binding protein C isoform 3 (MYBPC3) variant Arg502Trp has been identified in multiple hypertrophic cardiomyopathy (HCM) cases, but compelling evidence to support or refute the pathogenicity of this variant is lacking. To determine the prevalence, origin and clinical significance of the MYBPC3 Arg502Trp variant. The prevalence of MYBPC3 Arg502Trp was ascertained in 1414 sequential HCM patients of primarily European descent. MYBPC3 Arg502Trp was identified in 34 of these 1414 unrelated HCM patients. Segregation of MYBPC3 Arg502Trp with clinical status was assessed in family members. Disease haplotypes were examined in 17 families using two loci flanking MYBPC3. Family studies identified an additional 43 variant carriers, many with manifest disease, yielding a calculated odds ratio of 11 000:1 for segregation of MYBPC3 Arg502Trp with HCM. Analyses in 17 families showed at least 4 independent haplotypes flanked MYBPC3 Arg502Trp. Eight individuals (4 probands and 4 family members) also had another sarcomere protein gene mutation. Major adverse clinical events occurred in approximately 30% of MYBPC3 Arg502Trp carriers by age 50; these were significantly more likely (P<0.0001) when another sarcomere mutation was present. MYBPC3 Arg502Trp is the most common and recurrent pathogenic mutation in a diverse primarily European descent HCM cohort, occurring in 2.4% of patients. MYBPC3 Arg502Trp conveys a 340-fold increased risk for HCM by 45 years of age, when more than 50% of carriers have overt disease. HCM prognosis worsens when MYBPC3 Arg502Trp occurs in the setting of another sarcomere protein gene mutation. Show less

The childhood onset of idiopathic cardiac hypertrophy that occurs without a family history of cardiomyopathy can portend a poor prognosis. Despite morphologic similarities to genetic cardiomyopathies of adulthood, the contribution of genetics to childhood-onset hypertrophy is unknown. We assessed the family and medical histories of 84 children (63 boys and 21 girls) with idiopathic cardiac hypertrophy diagnosed before 15 years of age (mean [+/-SD] age, 6.99+/-6.12 years). We sequenced eight genes: MYH7, MYBPC3, TNNT2, TNNI3, TPM1, MYL3, MYL2, and ACTC. These genes encode sarcomere proteins that, when mutated, cause adult-onset cardiomyopathies. We also sequenced PRKAG2 and LAMP2, which encode metabolic proteins; mutations in these genes can cause early-onset ventricular hypertrophy. We identified mutations in 25 of 51 affected children without family histories of cardiomyopathy and in 21 of 33 affected children with familial cardiomyopathy. Among 11 of the 25 children with presumed sporadic disease, 4 carried new mutations and 7 inherited the mutations. Mutations occurred predominantly (in >75% of the children) in MYH7 and MYBPC3; significantly more MYBPC3 missense mutations were detected than occur in adult-onset cardiomyopathy (P<0.005). Neither hypertrophic severity nor contractile function correlated with familial or genetic status. Cardiac transplantation and sudden death were more prevalent among mutation-positive than among mutation-negative children; implantable cardioverter-defibrillators were more frequent (P=0.007) in children with family histories that were positive for the mutation. Genetic causes account for about half of presumed sporadic cases and nearly two thirds of familial cases of childhood-onset hypertrophy. Childhood-onset hypertrophy should prompt genetic analyses and family evaluations. Show less

Amplified fragment length polymorphism (AFLP) typing was applied to 116 Streptococcus suis isolates with different clinical backgrounds (invasive/pneumonia/carrier/human) and with known profiles of virulence-associated genes (cps1, -2, -7 and -9, as well as mrp, epf and sly). A dendrogram was generated that allowed identification of two clusters (A and C) with different subclusters (A1, A2, C1 and C2) and two heterogeneous groups of strains (B and D). For comparison, three strains from each AFLP subcluster and group were subjected to multilocus sequence typing (MLST) analysis. The closest relationship and lowest diversity were found for patterns clustering within AFLP subcluster A1, which corresponded with sequence type (ST) complex 1. Strains within subcluster A1 were mainly invasive cps1 and mrp+ epf+ (or epf*) sly+ cps2+ strains of porcine or human origin. A new finding of this study was the clustering of invasive mrp* cps9 isolates within subcluster A2. MLST analysis suggested that A2 correlates with a single ST complex (ST87). In contrast to A1 and A2, subclusters C1 and C2 contained mainly pneumonia isolates of genotype cps7 or cps2 and epf- sly-. In conclusion, this study demonstrates that AFLP allows identification of clusters of S. suis strains with clinical relevance. Show less

Definition of virulent Streptococcus suis strains is controversial. One successful approach for identification of virulent European strains is differentiation of capsular serotypes (or the corresponding cps types) and subsequent detection of virulence-associated factors, namely the extracellular factor (EF, epf), the muramidase-released protein (MRP, mrp) and the hemolysin suilysin (SLY, sly). In this work we present a novel multiplex PCR (MP-PCR) and an mrp variant PCR for identification and characterization of virulent S. suis strains. These new methods were used to identify association of disease with particular profiles of virulence-associated genes. The MP-PCR allowed identification of S. suis through detection of the housekeeping gene gdh, differentiation of four cps types (1, 2, 7 and 9), and detection of epf, mrp, sly and arcA (arginine deiminase from S. suis). Furthermore, this study describes the first PCR assay for differentiation of at least six mrp variants. Expression of the corresponding size variants of MRP was shown for four of the six mrp variants, but was undetectable for the two larger mrp variants in the particular strains investigated. The results of this study suggest that cps7 strains are associated with pneumonia and that variation of mrp is very pronounced among these strains. Gene profiles of invasive, pneumonia and carrier S. suis isolates by combination of PCR assays allowed differentiation of 24 different genotypes among cps1, 2, 7 and 9 strains. Forty-five percent of the invasive S. suis diseases investigated in this study were caused by only two of these genotypes, namely cps2/mrp+/epf+/sly+ and cps9/mrp(*)/epf-/sly+. Thus, this study demonstrates for the first time a uniform profile of the particular virulence-associated genes for the vast majority of the investigated invasive cps9 strains. Show less