Fragmentomic cfDNA Patterns in Noninvasive Prenatal Testing and Beyond

The release of fetoplacental cell-free DNA (cfDNA) into the maternal bloodstream opened up new avenues towards noninvasive prenatal testing (NIPT) for aneuploidies, hereditary DNA mutations and other pregnancy-related developmental disorders. Increasingly, cfDNA catches interest for its noninvasive screening value in other areas as well, including oncology. Although there are indications that cfDNA fragmentation is a non-random process, the etiology and different structural aspects of cfDNA are still not well known. The emerging field of cfDNA fragmentomics investigates the existence of tissue and disease specific cfDNA signatures and the chemistry and biology underlying the fragmentation process. This review sheds light on recent developments in cfDNA fragmentomics and illustrates their significance in NIPT improvement and beyond.We discuss aspects of fragment size distributions, epigenetic correlations and putatively enriched cfDNA fragment-end compositions. Combinatorial fragmentomic efforts have provided more insights into the roles of different enzymes that contribute to the fragmentation process in the tissue of origin and in the bloodstream. Altogether, these studies revealed multiple fragmentomic-related biomarkers that can be used to make noninvasive screening and other types of clinical use of cfDNA more robust, by raising its distinctive capacities. This includes multiple complementary approaches to determine the fetal fraction, a key determinant in NIPT. Furthermore, these developments translate to a better understanding of the encountered cfDNA patterns and will catalyze the expansion of screening possibilities in NIPT and beyon

Monitoring and predictive modelling of estuarine benthic macrofauna and their relevance to resource management problems

Practical considerations in estuarine management, as well as prediction of the consequences of global change on coastal protection, urgently require a better understanding and better modeling of estuarine ecosystems as influenced by ecological, physical, chemical and morphological processes. Recent Dutch examples of such questions are: the impact of enhanced dredging in the Schelde estuary, the impact of sea level rise on the Wadden Sea and Delta area, concerns about the loss of salt marsh habitats, etc. Benthic communities are good indicators of biotic integrity and reflect the present state of the estuarine ecosystem. The analysis of benthic infauna is a key element of many marine and estuarine monitoring programs. In the Dutch Delta area (SW-Netherlands) there is a relatively long tradition on estuarine macrozoobenthos monitoring, such as implemented e.g. in the BIOMON program. This program was designed to detect long-term trends in the average density, biomass and species composition of large parts of different systems (e.g. Schelde estuary, Oosterschelde, Grevelingen), in order to obtain insight in the natural development of estuarine and coastal areas and the anthropogenic influences on these systems. Running now for over a decade, these programs, together with other field campaigns, provide a unique data set on benthic macrofauna (e.g. for the Schelde estuary over 5000 samples are available at the moment). Until recently these data were hardly processed and used for further analysis. However, such data sets offer the opportunity to analyze and predict patterns in occurrence of benthic macrofauna in a much more profound way. Recently, within a cooperation between decision makers (Rijkswaterstaat, Directie Zeeland), advisers (RIKZ) and scientists (NIOO-CEMO), the possibilities and limitations of using these data sets for the predictions of benthic macrofauna at scales relevant to resource management problems are evaluated. In our approach we use different statistical methodologies to quantify, model and predict patterns at different spatial and temporal scales, going from patterns on a single tidal flat to inter-estuary comparisons and from monthly patterns to decennial trends. Several examples are shown that illustrate the use of these data, going from simple classification techniques to more sophisticated predictive modeling: Changes and shifts in benthos communities are shown for a land reclamation area of Rotterdam harbour in the Haringvliet-delta using classification techniques. Ordination analysis on the saline lake Grevelingen, a former estuary, showed long-term changes in macrobenthic community structure as a consequence of changes in salinity, light penetration, etc. This case study will be dealt with in more detail in a separate contribution. In the Schelde estuary, a detailed study was performed to unravel the use of environmental data in predicting benthic macrofaunal species distributions at different spatial scales (from a single tidal flat to the whole estuary). Statistical techniques such as geostatistics, hierarchical analysis and logistic regression were applied. At these different scales a distinct relation between the environment (e.g. salinity, sediment characteristics) on the one hand and macrofaunal species distributions on the other hand was observed. As a consequence, predictions of macrofaunal distributions can be made quite successful from environmental data within the Schelde estuary. An inter-estuary comparison between the Schelde estuary and Oosterschelde revealed that predictive models should also incorporate system-wide properties of estuarine systems, such as primary production and suspended matter concentrations, in order to perform in a more generic way. The results clearly show their use in making more sensible long-term decisions about matters having direct environmental effects. The results also provide information on how the design of monitoring programs could be improved or optimized, depending on the questions asked. As such, a more synergetic and flexible approach is urgently necessary, in which decision makers, advisers and scientists communicate in a more efficient way

Conserved structured domains in plant non-coding RNA enod40, their evolution and recruitment of sequences from transposable elements

Plant long noncoding RNA enod40 is involved in the regulation of symbiotic associations with bacteria, in particular, in nitrogen-fixing root nodules of legumes, and with fungi in phosphate-acquiring arbuscular mycorrhizae formed by various plants. The presence of enod40 genes in plants that do not form such symbioses indicates its other roles in cell physiology. The molecular mechanisms of enod40 RNA function are poorly understood. Enod40 RNAs form several structured domains, conserved to different extents. Due to relatively low sequence similarity, identification of enod40 sequences in plant genomes is not straightforward, and many enod40 genes remain unannotated even in complete genomes. Here, we used comparative structure analysis and sequence similarity searches in order to locate enod40 genes and determine enod40 RNA structures in nitrogen-fixing clade plants and in grasses. The structures combine conserved features with considerable diversity of structural elements, including insertions of structured domain modules originating from transposable elements. Remarkably, these insertions contain sequences similar to tandem repeats and several stem-loops are homologous to microRNA precursors.</p

Feasibility of preconception screening for thalassaemia in Indonesia: exploring the opinion of Javanese mothers

Background. Thalassaemia has become a major public health issue in Indonesia. It has been estimated that up to 10% of the population carries a gene associated with beta-thalassaemia. Currently, there is no formal recommendation for thalassaemia screening. This study aimed to explore awareness of thalassaemia, and to explore attitudes regarding carrier testing among Javanese mothers. Methods. A quantitative questionnaire, designed using constructs of the Theory of Planned Behaviour, was applied cross-sectionally. Results. Out of 191 mothers who were invited, 180 agreed to participate (RR 94%), of whom 74 had a child affected with thalassaemia. Both attitudes towards receiving information about thalassaemia, and attitudes towards carrier testing were very positive. Awareness of thalassaemia was poor. Mothers, both those with and without an affected child, had barely heard of thalassaemia, nor of carrier testing. However, all mothers, including those with an affected child expressed high levels of interest in carrier testing. Respondents did not perceive that they had any control over carrier testing, and feared stigmatization and being discriminated against if their carrier status was identified. Attitudes towards carrier testing explained 23% of future reproductive intentions, in addition to perceived stigmatization, education level and ‘mother’s age’ (R2 0.44; p 0.001). Conclusion. Responding mothers expressed high levels of interest in receiving information on both thalassaemia and carrier testing. The less educated and the more deprived they were, the keener they were to receive this information. Overall, awareness of thalassaemia was low. Even mothers with affected children seemed unaware of the inheritance pattern and the recurrent risk of having an affected child in a subsequent pregnancy, showing the need for genetic counselling in Indonesia. It is therefore recommended not only to raise awareness about thalassaemia, but to improve the education of healthcare professionals as well. Keywords: thalassaemia; carrier screening; Javanese mothers; genetic awareness; Theory of Planned Behaviour (TpB); awareness; Indonesi

Additional value of advanced ultrasonography in pregnancies with two inconclusive cell-free DNA draws

Objective: We aimed to evaluate the additional value of advanced fetal anatomical assessment by ultrasound in pregnancies with twice inconclusive noninvasive testing (NIPT) due to low fetal fraction (FF). Methods: We performed a multicenter-retrospective study between 2017 and 2020 including 311 pregnancies with twice inconclusive NIPT due to low FF ≤ 1%. Women were offered invasive testing and advanced fetal anatomical assessment at ≤18 weeks' gestation. Ultrasound findings, genetic testing, and pregnancy/postnatal outcomes were evaluated. Results: Ninety-two/311 (29.6%) women underwent invasive testing. Structural anomalies were diagnosed in 13/311 (4.2%) pregnancies (nine at the first scan and four at follow-up). In 6/13 (46.2%) cases, genetic aberrations were confirmed (one case of Trisomy 13 (detectable by NIPT), two of Triploidy, one of 16q12-deletion, HCN4-mutation and UPD(16) (nondetectable by NIPT). Genetic aberrations were found in 4/298 (1.3%) structurallynormal pregnancies (one 47XYY, two microscopic aberrations, one monogenic disorder found postpartum). Structural anomalies in genetically normal fetuses (2.0%) were not more prevalent compared to the general pregnant population (OR 1.0 [0.4–2.2]). Conclusion: In pregnancies with twice inconclusive NIPT due to low FF, fetal structural anomalies are not more prevalent than in the general obstetric population. The detailed anatomical assessment has the added value to detect phenotypical features suggestive of chromosomal/genetic aberrations and identify pregnancies where advanced genetic testing may be indicated

A novel CCM2 variant in a family with non-progressive cognitive complaints and cerebral microbleeds

Lobar cerebral microbleeds are most often sporadic and associated with Alzheimer's disease. The aim of our study was to identify the underlying genetic defect in a family with cognitive complaints and multiple lobar microbleeds and a positive family history for early onset Alzheimer's disease. We performed exome sequencing followed by Sanger sequencing for validation purposes on genomic DNA of three siblings with cognitive complaints, reduced amyloid-beta-42 in CSF and multiple cerebral lobar microbleeds. We checked for the occurrence of the variant in a cohort of 363 patients with early onset dementia and/or microbleeds. A novel frameshift variant (c.236_237delAC) generating a premature stop codon in the CCM2 gene shared by all three siblings was identified. Pathogenicity of the variant was supported by the presence of cerebral cavernous malformations in two of the siblings and by the absence of the variant exome variant databases. Two siblings were homozygous for APOE-ϵ4; one heterozygous. The cognitive complaints, reduced amyloid-beta-42 in CSF and microbleeds suggest preclinical Alzheimer's disease, but the stability of the cognitive complaints does not. We hypothesize that the phenotype in this family may be due to a combination of the CCM2 variant and the APOE status. © 2016 The Authors. American Journal of Medical Genetics Part B: Neuropsychiatric Genetics Published by Wiley Periodicals, Inc