Comprehensive population screening in the Ashkenazi Jewish population for recurrent disease‐causing variants

The Ashkenazi Jewish (AJ) population has an increased risk for a variety of recessive diseases due to historical founder effects and genetic drift. For some, the disease‐causing founder mutations have been identified and well‐characterized, but for others, further study is necessary. The purpose of this study is to assess the carrier frequencies of 85 pathogenic variants causative of 29 recessive conditions in the AJ population. Up to 3000 AJ individuals were genotyped by Luminex MagPlex^®‐TAG^? bead array or Agena Bioscience^? MassARRAY assays. We identified seven conditions with carrier frequencies higher than 1 in 100, nine between 1 in 100 and 1 in 200, and four between 1 in 200 and 1 in 500. Variants in nine conditions had a detected carrier rate of less than 1 in 500 or were not identified in approximately 2000 AJ individuals. We assessed the combined AJ carrier frequency for 18 relatively prevalent diseases to be 1 in 6, and the risk of AJ individuals to be a carrier couple for one of these 18 diseases as 1 in 441. We note additional recessive genetic conditions should be considered for AJ carrier screening panels.     

Premarital and prenatal screening for cystic fibrosis: experience in the Ashkenazi Jewish population.

PURPOSE: Since the early 1990s, Dor Yeshorim (DY) and the Mount Sinai School of Medicine (MSSM) have conducted premarital and prenatal carrier screening for cystic fibrosis (CF) in the Ashkenazi Jewish (AJ) population as part of their genetic testing programs, respectively. Together, over 170,000 screenees have been tested. In this study, we report the CF mutation frequencies in over 110,000 screenees who reportedly were of 100% AJ descent from the DY program and MSSM. In addition, the CF mutation frequencies in a group of > 7,000 screenees for AJ diseases who were of < 100% AJ descent are reported. METHODS: Testing for CF mutations was performed by either PCR and restriction digestion or ASO hybridization analyses at MSSM or sent to various academic and commercial laboratories by DY. RESULTS: The overall (and individual) carrier frequency for the five common AJ mutations, W1282X (0.020), DeltaF508 (0.012), G542X (0.0024), 3849+10kb C>T (0.0020), and N1303K (0.0016), among screenees who were 100% AJ was 1 in 26; when D1152H and the rare 1717-1G>A were included, the overall carrier frequency increased to approximately 1 in 23. In four families with D1152H, five compound heterozygotes for D1152H and W1282X (n = 2), DeltaF508 (1) or 3849+10kb C>T (1) were identified. In contrast, the carrier frequency for screenees reporting < 100% AJ descent was approximately 1 in 30 for the seven mutations. CONCLUSIONS: The carrier frequency for five common CF mutations in a large 100% AJ sample increased from 1 in 26 to 1 in 23 when D1152H was included in the panel. Addition of D1152H to mutation panels when screening the AJ population should be considered because compound heterozygosity is associated with a variable disease phenotype. Further studies to delineate the phenotype of CF patients with this mutation are needed.     

Molecular screening for diseases frequent in Ashkenazi Jews: lessons learned from more than 100,000 tests performed in a commercial laboratory.

PURPOSE: To determine the frequency of carriers of Ashkenazi Jewish (AJ) genetic diseases in the US population and compare these numbers with previously published frequencies reported in smaller more isolated cohorts. METHODS: A database containing more than 100,000 genotyping assays was queried. Assays for 10 separate AJ genetic diseases where comparisons were made with published data. RESULTS: As expected, we observed lower carrier frequencies in a general, US population than those reported in literature. In 2427 patients tested for a panel of 8 AJ diseases, 20 (1:121) were carriers of two diseases and 331 (1:7) were carriers of a single disease. Fifty-three of 7184 (1:306) individuals tested for Gaucher disease had 2 Gaucher Disease mutations indicating a potentially affected phenotype. CONCLUSIONS: As the number of AJ diseases increases, progressively more individuals will be identified as carriers of at least one disease.     

Canavan disease: carrier-frequency determination in the Ashkenazi Jewish population and development of a novel molecular diagnostic assay

Canavan disease (CD) is an autosomal recessive progressive neurodegenerative disorder prevalent in the Ashkenazi Jewish (AJ) population. The carrier rate for the most common mutations that cause CD in the AJ population is often quoted as 1:37-1:40. This is not supported by our finding of only two diagnosed cases of CD in the last 20 years in the Toronto AJ population of 160,000 and an estimated birth rate of 1,500-2,000 per year. Therefore, we embarked on a prevalence cross-sectional screening study to determine the carrier rate of CD in this population. In order to perform low-cost, high-throughput population testing for CD using molecular techniques, we first developed a novel molecular assay using multiplex fluorescent allele specific polymerase chain reaction (PCR) to test for the three most common mutations causing CD in the AJ population (A854C, C693A, C914A) and a neutral polymorphism at the site of the C693A mutation. During testing it was noted that individuals who were carriers of the A854C mutation also had a T polymorphism at the site of the C693A mutation (Y231X). We confirmed that in all A854C carriers the 854C mutation was in disequilibrium with the 693T polymorphism, indicating a founder chromosome for the A854C mutation in the AJ population. Twenty-five carriers were found from 1,423 samples yielding a carrier rate of 1:57, differing from the widely quoted frequency of 1:40 and supporting our observed frequency of disease.     

Clinical experience with carrier screening in a general population: support for a comprehensive pan-ethnic approach

PurposeTo present results from a large cohort of individuals receiving expanded carrier screening (CS) in the United States.MethodsSingle-gene disorder carrier status for 381,014 individuals was determined using next-generation sequencing (NGS) based CS for up to 274 genes. Detection rates were compared with literature-reported values derived from disease prevalence and carrier frequencies. Combined theoretical affected pregnancy rates for the 274 screened disorders were calculated.ResultsFor Ashkenazi Jewish (AJ) diseases, 81.6% (4434/5435) of carriers identified did not report AJ ancestry. For cystic fibrosis, 44.0% (6260/14,229)of carriers identified had a variant not on the standard genotyping panel. Individuals at risk of being a silent spinal muscular atrophy carrier, not detectable by standard screening, comprised 1/39 (8763/344,407) individuals. For fragile X syndrome, compared with standard premutation screening, AGG interruption analysis modified risk in 83.2% (1128/1356) premutation carriers. Assuming random pairing across the study population, approximately 1/175 pregnancies would be affected by a disorder in the 274-gene screening panel.ConclusionCompared with standard screening, NGS-based CS provides additional information that may impact reproductive choices. Pan-ethnic CS leads to substantially increased identification of at-risk couples. These data support offering NGS-based CS to all reproductive-aged women.     

Prepregnancy body mass index and risk of multiple congenital anomalies

Familial dysautonomia (FD) is an autosomal recessive congenital neuropathy that occurs almost exclusively in the Ashkenazi Jewish (AJ) population. Mutations in the IκB kinase complex‐associated protein (IKBKAP) gene cause FD. Two IKBKAP mutations, IVS20^{+6T → C} and R696P, have been identified in FD patients of AJ descent. The splice site mutation IVS20^{+6T → C} is responsible for > 99.5% of known AJ patients with FD, and haplotype analyses were consistent with a common founder. In contrast, the R696P mutation has been identified in only a few AJ patients. To facilitate carrier detection, a single PCR and allele‐specific oligonucleotide (ASO) hybridization assay was developed to facilitate the detection of both the IVS20^{+6T → C} and R696P mutations. Screening of 2,518 anonymous AJ individuals from the New York metropolitan area revealed a carrier frequency for IVS20^{+6T → C} of 1 in 32 (3.2%; 95% CI, 2.5–3.9%), similar to the previously estimated carrier frequency (3.3%) based on disease incidence. No carrier was identified for the R696P lesion, indicating that the mutation was rare in this population (< 1 in 2,500). This sensitive and specific assay should facilitate carrier screening for FD mutations in the AJ community, as well as postnatal diagnostic testing. © 2002 Wiley‐Liss, Inc.     

Mutation frequencies for glycogen storage disease Ia in the Ashkenazi Jewish population

Glycogen storage disease type Ia (GSDIa) is a severe autosomal recessive disorder caused by deficiency of the enzyme D-glucose-6-phosphatase (G6Pase). While numerous mutations have been found in cosmopolitan European populations, Ashkenazi Jewish (AJ) patients appear to primarily carry the R83C mutation, but possibly also the Q347X mutation found generally in Caucasians. To determine the frequency for both these mutations in the AJ population, we tested 20,719 AJ subjects for the R83C mutation and 4,290 subjects for the Q347X mutation. We also evaluated the mutation status of 30 AJ GSDIa affected subjects. From the carrier screening, we found 290 subjects with R83C, for a carrier frequency for this mutation of 1.4%. This carrier frequency translates into a predicted disease prevalence of 1 in 20,000, five times higher than for the general Caucasian population, confirming a founder effect and elevated frequency of GSDIa in the AJ population. We observed no carriers of the Q347X mutation. Among the 30 GSDIa affected AJ subjects, all were homozygous for R83C. These results indicate that R83C is the only prevalent mutation for GSDIa in the Ashkenazi population.     

The GALT rush: high carrier frequency of an unusual deletion mutation of the GALT gene in the Ashkenazi population

Classic galactosemia is an autosomal recessive disorder of galactose metabolism manifesting in the first weeks of life following exposure to a milk-based diet. Despite the benefit of avoidance of lactose, many patients suffer from long-term complications including neurological deficits and ovarian failure. To date, over 230 mutations have been described in the GALT gene resulting in galactosemia. Recently, an unusual mutation was characterized causing a 5.5 kb deletion, with a relatively high carrier rate in subjects of Ashkenazi Jewish (AJ) descent. The aim of this study was to estimate the carrier frequency of this mutation in the AJ population in Israel. For this purpose we developed a high-throughput methodology to genotype both normal and deleted alleles using a chip-based matrix-assisted laser desorption-time-of-flight (MALDI-TOF) mass spectrometer and Multiplex PCR. DNA samples of 760 anonymous AJ subjects were submitted for analysis, subsequently detecting six individuals heterozygous for the GALT deletion mutation, giving a carrier frequency of 1 in 127 (0.79%). Based on these results, we suggest that the method described here provides a basis for genetic screening and prenatal counseling and can potentially reduce the morbidity and mortality associated with delayed diagnosis of galactosemia in this patient population.     

A deleterious mutation in the PEX2 gene causes Zellweger syndrome in individuals of Ashkenazi Jewish descent

Zellweger syndrome is known to be caused by numerous mutations that occur in at least 12 of the PEX genes. While phenotypes vary, many are severely debilitating, and death can result in affected newborns. Since the disease follows an autosomal recessive pattern of inheritance, carrier screening can be done for at‐risk couples, but the number of potential mutations sites to screen can be daunting. Ethnicity‐specific studies can help narrow this range by highlighting mutations that are present at higher percentages in certain populations. In this article, the carrier frequencies for two mutations causative of the severe Zellweger syndrome spectrum phenotype that occur in the PEX2 gene, c.355C>T and c.550del, were studied in individuals of Ashkenazi Jewish descent in order to advise on inclusion in existing carrier screening mutation panels for this population. The screening was performed for 2093 individuals through the use of TaqMan genotyping assays, real‐time PCR, and allelic discrimination. Results indicated a carrier frequency of 0.813% (±0.385%) for the c.355C>T mutation and a carrier frequency of 0.00% (±0.00%) for the c.550del mutation. On the basis of these frequencies, we believe that the c.355C>T mutation should be considered for inclusion in carrier screening panels for the Ashkenazi population.     

Familial dysautonomia: detection of the IKBKAP IVS20(+6T --> C) and R696P mutations and frequencies among Ashkenazi Jews

Familial dysautonomia (FD) is an autosomal recessive congenital neuropathy that occurs almost exclusively in the Ashkenazi Jewish (AJ) population. Mutations in the IkappaB kinase complex-associated protein (IKBKAP) gene cause FD. Two IKBKAP mutations, IVS20(+6T --> C) and R696P, have been identified in FD patients of AJ descent. The splice site mutation IVS20(+6T --> C) is responsible for > 99.5% of known AJ patients with FD, and haplotype analyses were consistent with a common founder. In contrast, the R696P mutation has been identified in only a few AJ patients. To facilitate carrier detection, a single PCR and allele-specific oligonucleotide (ASO) hybridization assay was developed to facilitate the detection of both the IVS20(+6T --> C) and R696P mutations. Screening of 2,518 anonymous AJ individuals from the New York metropolitan area revealed a carrier frequency for IVS20(+6T --> C) of 1 in 32 (3.2%; 95% CI, 2.5-3.9%), similar to the previously estimated carrier frequency (3.3%) based on disease incidence. No carrier was identified for the R696P lesion, indicating that the mutation was rare in this population (< 1 in 2,500). This sensitive and specific assay should facilitate carrier screening for FD mutations in the AJ community, as well as postnatal diagnostic testing.     

Using Reference Databases of Genetic Variation to Evaluate the Potential Pathogenicity of Candidate Disease Variants

The potential pathogenicity of genetic variants identified in disease‐based resequencing studies is often overlooked where variants have previously been reported in dbSNP, the 1000 genomes project, or the National Heart, Lung and Blood Institute Exome Sequencing Project (ESP). In this work, we estimate that collectively, these databases capture ～52% of mutations (dbSNP 50.4%; 1000 genomes 4.8%; and ESP 10.2%) reported as disease causing within phenotype‐based locus‐specific databases (LSDBs). To investigate whether these mutations may simply represent benign population variants, we evaluated whether the carrier frequencies associated with mutations implicated in amyotrophic lateral sclerosis were higher than what could be accounted for by high‐penetrance disease models. In doing so, we have questioned the veracity of 51 mutations, but also demonstrated that each of the three databases included credible disease variants. Our results demonstrate the benefits of using databases such as dbSNP, the 1000 genomes project, and the ESP to evaluate the pathogenicity of putative disease variants, and suggest that many disease mutations reported across LSDBs may not actually be pathogenic. However, they also demonstrate that even in the context of rare Mendelian disorders, the potential pathogenicity of variants reported by these databases should not be overlooked without proper evaluation.     

Carrier frequency of two BBS2 mutations in the Ashkenazi population

Bardet–Biedl syndrome (BBS) is known to be caused by numerous mutations that occur in at least 15 of the BBS genes. As the disease follows an autosomal recessive pattern of inheritance, carrier screening can be performed for at‐risk couples, but the number of potential mutation sites to screen can be daunting. Ethnic studies can help to narrow this range by highlighting mutations that are present at higher percentages in certain populations. In this article, the carrier frequency for two mutations that occur in the BBS2 gene, c.311A>C and c.1895G>C were studied in individuals of Ashkenazi Jewish descent in order to advise on including them in existing mutation panels for this population. Carrier screenings were performed on individuals from the Ashkenazi Jewish population using a combination of TaqMan genotyping assays followed by real‐time polymerase chain reaction (PCR) and allelic discrimination, and allele‐specific PCR confirmed by restriction analysis. The combined results indicated carrier frequencies of 0.473% (±0.0071%) for the c.311A>C mutation and 0.261% (±0.0064%) for the c.1895G>C mutation. On the basis of these frequencies, we believe that the two mutations should be considered for inclusion in screening panels for the Ashkenazi population.     

Pan-ethnic carrier screening and prenatal diagnosis for spinal muscular atrophy: clinical laboratory analysis of >72,400 specimens

Spinal muscular atrophy (SMA) is a leading inherited cause of infant death with a reported incidence of ~1 in 10,000 live births and is second to cystic fibrosis as a common, life-shortening autosomal recessive disorder. The American College of Medical Genetics has recommended population carrier screening for SMA, regardless of race or ethnicity, to facilitate informed reproductive options, although other organizations have cited the need for additional large-scale studies before widespread implementation. We report our data from carrier testing (n = 72,453) and prenatal diagnosis (n = 121) for this condition. Our analysis of large-scale population carrier screening data (n = 68,471) demonstrates the technical feasibility of high throughput testing and provides mutation carrier and allele frequencies at a level of accuracy afforded by large data sets. In our United States pan-ethnic population, the calculated a priori carrier frequency of SMA is 1/54 with a detection rate of 91.2%, and the pan-ethnic disease incidence is calculated to be 1/11,000. Carrier frequency and detection rates provided for six major ethnic groups in the United States range from 1/47 and 94.8% in the Caucasian population to 1/72 and 70.5% in the African American population, respectively. This collective experience can be utilized to facilitate accurate pre- and post-test counseling in the settings of carrier screening and prenatal diagnosis for SMA.     

Estimation of carrier frequencies of six autosomal-recessive Mendelian disorders in the Korean population

Although many studies have been performed to identify mutations in Korean patients with various autosomal-recessive Mendelian disorders (AR-MDs), little is known about the carrier frequencies of AR-MDs in the Korean population. Twenty common mutations from six AR-MDs, including Wilson disease (WD), non-syndromic hearing loss (NSHL), glycogen storage disease type Ia (GSD Ia), phenylketonuria (PKU), congenital hypothyroidism (CH), and congenital lipoid adrenal hyperplasia (CLAH) were selected to screen for based on previous studies. A total of 3057 Koreans were genotyped by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry followed by confirmation using the Sanger sequencing. We found 201 and 8 carriers with either one or two mutations in different genes, respectively, yielding a total carrier frequency of 1 in 15 (6.7%). Of the six AR-MDs, NSHL has the highest carrier frequency followed by WD, CH, CLAH, GSD Ia, and PKU. As carrier screening tests are becoming prevalent and the number of mutations known and tested is rising, a priori data on the carrier frequencies in different ethnic groups is mandatory to plan a population screening program and to estimate its efficiency. In light of this, the present results can be used as a basis to establish a screening policy for common AR-MRs in the Korean population.     

An empirical estimate of carrier frequencies for 400+ causal Mendelian variants: results from an ethnically diverse clinical sample of 23,453 individuals

PurposeRecent developments in genomics have led to expanded carrier screening panels capable of assessing hundreds of causal mutations for genetic disease. This new technology enables simultaneous measurement of carrier frequencies for many diseases. As the resultant rank-ordering of carrier frequencies impacts the design and prioritization of screening programs, the accuracy of this ranking is a public health concern.MethodsA total of 23,453 individuals from many obstetric, genetics, and infertility clinics were referred for routine recessive disease carrier screening. Multiplex carrier screening was performed and results were aggregated for this study.ResultsTwenty-four percent of individuals were identified as carriers for at least one of 108 disorders, and 5.2% were carriers for multiple disorders. We report tabulations of carrier frequency by self-identified ethnicity and disease.ConclusionTo our knowledge, this study of a large, ethnically diverse clinical sample provides the most accurate measurements to date of carrier frequencies for hundreds of recessive alleles. The study also yields information on the clinical considerations associated with routine use of expanded panels and provides support for a pan-ethnic screening paradigm that minimizes the use of “racial” categories by the physician, as recommended by recent guidelines.Genet Med 2013:15(3):178–186     

A common spinal muscular atrophy deletion mutation is present on a single founder haplotype in the US Hutterites

Spinal muscular atrophy (SMA) is an autosomal recessive (AR) neuromuscular disease that is one of the most common lethal genetic disorders in children, with carrier frequencies as high as ～1 in 35 in US Whites. As part of our genetic studies in the Hutterites from South Dakota, we identified a large 22 Mb run of homozygosity, spanning the SMA locus in an affected child, of which 10 Mb was also homozygous in three affected Hutterites from Montana, supporting a single founder origin for the mutation. We developed a haplotype-based method for identifying carriers of the SMN1 deletion that leveraged existing genome-wide SNP genotype data for ～1400 Hutterites. In combination with two direct PCR-based assays, we identified 176 carriers of the SMN1 deletion, one asymptomatic homozygous adult and three carriers of a de novo deletion. This corresponds to a carrier frequency of one in eight (12.5%) in the South Dakota Hutterites, representing the highest carrier frequency reported to date for SMA and for an AR disease in the Hutterite population. Lastly, we show that 26 SNPs can be used to predict SMA carrier status in the Hutterites, with 99.86% specificity and 99.71% sensitivity.     

Strategies for the prevention of hereditary diseases in a highly consanguineous population

Autosomal recessive hereditary diseases are relatively common in the Saudi population. The consanguinity rate is in excess of 50% and is a practice that remains strongly embedded within Saudi culture. The impact of this practice is recognized and is being addressed. Early detection and treatment of diseases can reduce mortality and minimize morbidity. This is the basis of successful neonatal screening for inborn errors of metabolism where treatment or modification of lifestyle can modulate disease. Ultimately, understanding the genetics of these diseases will provide opportunities for prevention. Options such as prenatal screening can be used to reduce the incidence of live births with inherited diseases. However, prenatal diagnosis and associated intervention is unacceptable to wide sections of all societies. Carrier detection and genetic counselling programmes have been very successful in reducing the incidence of inherited disorders in many populations. These programmes are most successful when they are sensitive to the cultural backgrounds of populations in which they are applied. In Saudi society, premarital screening to identify carrier status and the provision of appropriate counselling has tremendous potential to prevent inherited disease.     

Cystic fibrosis carrier screening in a North American population

PurposeThe aim of this study was to compare the mutation frequency distribution for a 32-mutation panel and a 69-mutation panel used for cystic fibrosis carrier screening. Further aims of the study were to examine the race-specific detection rates provided by both panels and to assess the performance of extended panels in large-scale, population-based cystic fibrosis carrier screening. Although genetic screening for the most common CFTR mutations allows detection of nearly 90% of cystic fibrosis carriers, the large number of other mutations, and their distribution within different ethnic groups, limits the utility of general population screening.MethodsPatients referred for cystic fibrosis screening from January 2005 through December 2010 were tested using either a 32-mutation panel (n = 1,601,308 individuals) or a 69-mutation panel (n = 109,830).ResultsThe carrier frequencies observed for the 69-mutation panel study population (1/36) and Caucasian (1/27) and African-American individuals (1/79) agree well with published cystic fibrosis carrier frequencies; however, a higher carrier frequency was observed for Hispanic-American individuals (1/48) using the 69-mutation panel as compared with the 32-mutation panel (1/69). The 69-mutation panel detected ~20% more mutations than the 32-mutation panel for both African-American and Hispanic-American individuals.ConclusionExpanded panels using race-specific variants can improve cystic fibrosis carrier detection rates within specific populations. However, it is important that the pathogenicity and the relative frequency of these variants are confirmed.Genet Med16 7, 539–546.     

FMR1 premutation frequency in a large,ethnically diverse population referred for carrier testing

Fragile X syndrome (FXS) is the most common inherited cause of intellectual disability and is caused by an expansion of cytosine‐guanine‐guanine (CGG) repeats in the FMR1 gene. Female premutation allele carriers (55–200 CGG repeats) are at risk to have an affected child. Currently, specific population‐based carrier screening for FXS is not recommended. Previous studies exploring female premutation carrier frequency have been limited by size or ethnicity. This retrospective study provides a pan‐ethnic estimate of the Fragile X premutation carrier frequency in a large, ethnically diverse population of women referred for routine carrier screening during a specified time period at Progenity, Inc. Patient ethnicity was self‐reported and categorized as: African American, Ashkenazi Jewish, Asian, Caucasian, Hispanic, Native American, Other/Mixed/Unknown, or Sephardic Jewish. FXS test results were stratified by ethnicity and repeat allele category. Total premutation carrier frequency was calculated and compared against each ethnic group. A total of 134,933 samples were included. The pan‐ethnic premutation carrier frequency was 1 in 201. Only the Asian group differed significantly from this frequency. Using the carrier frequency of 1 in 201, a conservative pan‐ethnic risk estimate for a male fetus to have FXS can be calculated as 1 in 2,412. This risk is similar to the highest ethnic‐based fetal risks for cystic fibrosis and spinal muscular atrophy, for which population‐wide screening is currently recommended. This study adds to the literature and supports further evaluation into specific population‐wide screening recommendations for FXS.     

Homocystinuria in the Arab population of Israel: identification of two novel mutations using DGGE analysis

This study describes, for the first time, a thorough genetic investigation in Israeli Arab homocystinuric patients. By using a DGGE methodology and sequencing we were able to identify the disease causing mutation in all. Of the mutations that were detected, two are novel: a 785C>G transversion in exon 7 (T262R) and a 5-bp deletion in the 5' of IVS17 including the T in the +2 position that is crucial for correct splicing (g18327-18331del5). In spite of the highly consanguineous nature of this population several different mutations were found. This may suggest that the mutations arose only recently in the population. The results of our study would enable early prenatal diagnosis, genetic counseling and screening for the mutations in population at risk. Hum Mutat 16:372, 2000.