Objectives
To evaluate the performance of cell‐free DNA (cfDNA) screening for common fetal aneuploidies, choice of prenatal procedure, and chromosome conditions identified during pregnancy after ...low‐risk cfDNA screening.
Method
A single‐center prenatal cfDNA screening test was employed to detect trisomies 21, 18, and 13 (T21, T18, T13) and sex chromosome aneuploidies (SCAs). Test performance, choice of prenatal procedure, and cytogenetic results in pregnancies with low‐risk cfDNA screening were reviewed.
Results
CfDNA screening of 38,289 consecutive samples identified 720 (1.9%) pregnancies at increased risk for aneuploidy. Positive predictive values (PPVs) for high‐risk singleton pregnancies were 98.5% (T21), 92.5% (T18) and 55.2% (T13). PPVs for SCAs ranged from 30.6% to 95.2%. Most women elected chorionic villus sampling for prenatal diagnosis of T21, T18 and T13; amniocentesis and/or postnatal testing were commonly chosen for SCAs. Cytogenetic tests from 616 screen‐negative pregnancies identified 64 cases (12.7%) with chromosome conditions not detected by cfDNA screening, including triploidy (n = 30) and pathogenic and likely pathogenic copy number variants (n = 34). A further 15 (0.04%) false‐negative common aneuploidy results were identified.
Conclusions
CfDNA screening was highly accurate for detecting fetal aneuploidy in this general‐risk obstetric population. Fetal ultrasound and prenatal diagnostic testing were important in identifying chromosome conditions in pregnancies screened as low‐risk.
Key points
What's already known about this topic?
Prenatal cell‐free DNA (cfDNA) screening for common autosomal trisomies has high sensitivity and specificity. Fewer data are available for sex chromosome aneuploidies (SCAs), which show greater variability in screening performance.
Chorionic villus sampling and amniocentesis are available for prenatal diagnosis of high‐risk results.
Other chromosome conditions not screened for, including copy number variants (CNVs), may not be recognized until later in pregnancy or after birth.
What does this study add?
cfDNA screening had high positive predictive values (PPVs) for common trisomies and most SCAs. Trisomy screening performance was high at fetal fractions ≥2.5%.
Most women elected chorionic villus sampling for prenatal diagnosis of autosomal trisomies. Amniocentesis or postnatal testing were more common for SCAs.
Diagnostic testing in pregnancies with low‐risk cfDNA screening results identified 64/616 (12.7%) cases with chromosome conditions not detected by cfDNA screening, and 0.04% false‐negative cases within the entire cohort.
Abstract
The view of maternal effects (nongenetic maternal environmental influence on offspring phenotype) has changed from one of distracting complications in evolutionary genetics to an important ...evolutionary mechanism for improving offspring fitness. Recent studies have shown that maternal effects act as an adaptive mechanism to prepare offspring for stressful environments. Although research into the magnitude of maternal effects is abundant, the molecular mechanisms of maternal influences on offspring phenotypic variation are not fully understood. Despite recent work identifying DNA methylation as a potential mechanism of nongenetic inheritance, currently proposed links between DNA methylation and parental effects are indirect and primarily involve genomic imprinting. We combined a factorial breeding design and gene-targeted sequencing methods to assess inheritance of methylation during early life stages at 14 genes involved in growth, development, metabolism, stress response, and immune function of Chinook salmon (Oncorhynchus tshawytscha). We found little evidence for additive or nonadditive genetic effects acting on methylation levels during early development; however, we detected significant maternal effects. Consistent with conventional maternal effect data, maternal effects on methylation declined through development and were replaced with nonadditive effects when offspring began exogenous feeding. We mapped methylation at individual CpG sites across the selected candidate genes to test for variation in site-specific methylation profiles and found significant maternal effects at selected CpG sites that also declined with development stage. While intergenerational inheritance of methylated DNA is controversial, we show that CpG-specific methylation may function as an underlying molecular mechanism for maternal effects, with important implications for offspring fitness.
Rapid genomic testing is a valuable new diagnostic tool for acutely unwell infants, however exome sequencing does not deliver clinical-grade mitochondrial genome sequencing and may fail to diagnose ...mitochondrial disorders caused by mitochondrial DNA (mtDNA) variants. Rapid mitochondrial genome sequencing and analysis are not routinely available in rapid genomic diagnosis programmes. We present two critically ill neonates with transfusion-dependent anaemia and persistent lactic acidosis who underwent rapid mitochondrial genome sequencing in tandem with exome sequencing as part of an exome sequencing-based rapid genomic diagnosis programme. No diagnostic variants were identified on examination of the nuclear exome data for either infant. Mitochondrial genome sequencing identified a large mtDNA deletion in both infants, diagnosing Pearson syndrome within 74 and 55 h, respectively. Early diagnosis in the third week of life allowed the avoidance of a range of other investigations and appropriate treatment planning. Rapid mitochondrial genome analysis provides additional diagnostic and clinical utility and should be considered as an adjunct to exome sequencing in rapid genomic diagnosis programmes.
Whilst next generation sequencing can report point mutations in fixed tissue tumour samples reliably, the accurate determination of copy number is more challenging. The conventional Multiplex ...Ligation-dependent Probe Amplification (MLPA) assay is an effective tool for measurement of gene dosage, but is restricted to around 50 targets due to size resolution of the MLPA probes. By switching from a size-resolved format, to a sequence-resolved format we developed a scalable, high-throughput, quantitative assay. MLPA-seq is capable of detecting deletions, duplications, and amplifications in as little as 5ng of genomic DNA, including from formalin-fixed paraffin-embedded (FFPE) tumour samples. We show that this method can detect BRCA1, BRCA2, ERBB2 and CCNE1 copy number changes in DNA extracted from snap-frozen and FFPE tumour tissue, with 100% sensitivity and >99.5% specificity.
ABSTRACT
Conventional means of identifying variants in high‐throughput sequencing align each read against a reference sequence, and then call variants at each position. Here, we demonstrate an ...orthogonal means of identifying sequence variation by grouping the reads as amplicons prior to any alignment. We used AmpliVar to make key‐value hashes of sequence reads and group reads as individual amplicons using a table of flanking sequences. Low‐abundance reads were removed according to a selectable threshold, and reads above this threshold were aligned as groups, rather than as individual reads, permitting the use of sensitive alignment tools. We show that this approach is more sensitive, more specific, and more computationally efficient than comparable methods for the analysis of amplicon‐based high‐throughput sequencing data. The method can be extended to enable alignment‐free confirmation of variants seen in hybridization capture target‐enrichment data.
In this study, AmpliVar was applied to amplicon sequencing data derived from acute myeloid leukemia, breast and ovarian cancer samples. Using a grouped read and quality filtering approach, AmpliVar identified variants with the highest sensitivity and effective false positive reduction. It displayed superior processing speed and was able to be used as an orthogonal means of validation for hybridization capture data.
Brassica ASTRA is a public database for genomic information on Brassica species. The database incorporates expressed sequences with Swiss-Prot and GenBank comparative sequence annotation as well as ...secondary Gene Ontology (GO) annotation derived from the comparison with Arabidopsis TAIR GO annotations. Simple sequence repeat molecular markers are identified within resident sequences and mapped onto the closely related Arabidopsis genome sequence. Bacterial artificial chromosome (BAC) end sequences derived from the Multinational Brassica Genome Project are also mapped onto the Arabidopsis genome sequence enabling users to identify candidate Brassica BACs corresponding to syntenic regions of Arabidopsis. This information is maintained in a MySQL database with a web interface providing the primary means of interrogation. The database is accessible at http://hornbill.cspp.latrobe.edu.au.
Salmon farming is one of Canada's fastest growing industries and contributes to Canada's economy as well as creating jobs in rural areas; however, the industry is challenged by the need to balance ...production economics against environmental impacts. While Atlantic salmon (Salmo salar) are the most commonly farmed species on the west coast of Canada, Chinook salmon (Oncorhynchus tshawytscha) are a valuable alternative, as they fill a niche market and generate reduced environmental concerns because they are a native species. However, Chinook salmon have not been systematically domesticated, and their performance remains highly variable. Here we report on the results of a research program designed to develop a performance-enhanced hybrid Chinook salmon stock. Growth and survival were estimated for seven domestic-wild hybrid Chinook salmon crosses at various freshwater stages and during 15 months of saltwater rearing at a British Columbia Chinook salmon farm and compared with domestic-domestic crosses (control). The project included 8640 individually (PIT) tagged offspring from the domestic stock and seven domestic-wild hybrid stocks originating from the Lower Fraser Valley, Lower Mainland Vancouver, and Vancouver Island, British Columbia, Canada. Within each stock, milt from 10 sires was used to fertilize eggs pooled from 15 highly inbred domestic females to produce 80 half-sib families. Our breeding design allows the partitioning of stock and sire effects, and minimises maternal genetic and maternal environment effects. Replicates of all families were reared under common environmental conditions in both fresh- and salt water and monitored for body size and survival. There was significant variation in survival, body size, and saltwater biomass among the Chinook salmon hybrid stocks. The performance of some of the hybrid crosses exceeded that of the fully domesticated stock, although the pattern of performance varied with rearing stage. Overall, two hybrid stocks consistently outperformed the domestic stock in terms of survival, growth, and biomass estimates. We systematically assess production performance across a wide range of wild-domestic hybrid crosses in a Pacific salmon species, and our results highlight opportunities to improve the production performance of Chinook salmon culture.
•Chinook salmon (Oncorhynchus tshawytscha) fill a niche aquaculture market, but performance remains variable.•Domestic fish were outcrossed with seven regional stocks and growth and survival surveyed during fresh- and saltwater stages.•Breeding design partitioned stock and sire effects, and minimized maternal-genetic and maternal-environment effects.•Significant performance variation across hybrids and rearing stage highlight prospects for improved Chinook salmon culture.
Whilst next generation sequencing can report point mutations in fixed tissue tumour samples reliably, the accurate determination of copy number is more challenging. The conventional Multiplex ...Ligation-dependent Probe Amplification (MLPA) assay is an effective tool for measurement of gene dosage, but is restricted to around 50 targets due to size resolution of the MLPA probes. By switching from a size-resolved format, to a sequence-resolved format we developed a scalable, high-throughput, quantitative assay. MLPA-seq is capable of detecting deletions, duplications, and amplifications in as little as 5ng of genomic DNA, including from formalin-fixed paraffin-embedded (FFPE) tumour samples. We show that this method can detect BRCA1, BRCA2, ERBB2 and CCNE1 copy number changes in DNA extracted from snap-frozen and FFPE tumour tissue, with 100% sensitivity and >99.5% specificity.
The availability of sequence data derived from shotgun sequencing programs enables mining for simple sequence repeats (SSRs), providing useful genetic markers for crop improvement. This study ...presents the development and characterization of 40 SSR markers from Brassica oleracea shotgun sequence and their cross-amplification across Brassica species. The markers show reliable amplification, genome specificity and considerable polymorphism, demonstrating the utility of SSRs for genetic analysis of commercial Brassica germplasm.
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