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Crooked tail (Cd) mice bear a gain-of-function mutation in Lrp6, a co-receptor for canonical WNT signaling, and are a model of neural tube defects (NTDs), preventable with dietary folic acid (FA) supplementation. Whether the FA response reflects a direct influence of FA on LRP6 function was tested with prenatal supplementation in LRP6-deficient embryos. The enriched FA (10 ppm) diet reduced the occurrence of birth defects among all litters compared with the control (2 ppm FA) diet, but did so by increasing early lethality of Lrp6(-/-) embryos while actually increasing NTDs among nulls alive at embryonic days 10-13 (E10-13). Proliferation in cranial neural folds was reduced in homozygous Lrp6(-/-) mutants versus wild-type embryos at E10, and FA supplementation increased proliferation in wild-type but not mutant neuroepithelia. Canonical WNT activity was reduced in LRP6-deficient midbrain-hindbrain at E9.5, demonstrated in vivo by a TCF/LEF-reporter transgene. FA levels in media modulated the canonical WNT response in NIH3T3 cells, suggesting that although FA was required for optimal WNT signaling, even modest FA elevations attenuated LRP5/6-dependent canonical WNT responses. Gene expression analysis in embryos and adults showed striking interactions between targeted Lrp6 deficiency and FA supplementation, especially for mitochondrial function, folate and methionine metabolism, WNT signaling and cytoskeletal regulation that together implicate relevant signaling and metabolic pathways supporting cell proliferation, morphology and differentiation. We propose that FA supplementation rescues Lrp6(Cd/Cd) fetuses by normalizing hyperactive WNT activity, whereas in LRP6-deficient embryos, added FA further attenuates reduced WNT activity, thereby compromising development.

The review lists the genetic diseases reported in Lebanese individuals, surveys genetic programs and services, and highlights the absence of basic genetic health services at the individual and community level. The incidence of individual diseases is not determined, yet the variety of genetic diseases reported is tremendous, most of which follow autosomal recessive inheritance reflecting the social norms in the population, including high rates of consanguinity, which favor the increase in incidence of these diseases. Genetic services including all activities for the diagnosis, care, and prevention of genetic diseases at community level are extremely inadequate. Services are limited to some clinical and laboratory diagnostic services with no genetic counseling. These services are localized within the capital thus preventing their accessibility to high-risk communities. Screening programs, which are at the core of public health prevention services, are minimal and not nationally mandated. The absence of adequate genetic services is attributed to many factors undermining the importance of genetic diseases and their burden on society, the most important of which is genetic illiteracy at all levels of the population, including high-risk families, the general public, and most importantly health care providers and public health officials. Thus, a country like Lebanon, where genetic diseases are expected to be highly prevalent, is in utmost need for community genetics services. Strategies need to be developed to familiarize public health officials and medical professionals with medical genetics leading to a public health infrastructure that delivers community genetics services for the prevention and care of genetic disorders at community level.

BackgroundNeural tube defects (NTDs) are the second most common birth defect in humans. Dietary folic acid (FA) supplementation effectively and safely reduces the incidence of these often debilitating congenital anomalies. FA plays an established role in folate and homocysteine metabolism, but the means by which it suppresses occurrence of NTDs is not understood. In addition, many cases remain resistant to the beneficial effects of folic acid supplementation. To better understand the molecular, biochemical and developmental mechanisms by which FA exerts its effect on NTDs, characterized mouse models are needed that have a defined genetic basis and known response to dietary supplementation.ResultsWe examined the effect of FA supplementation, at 5-fold the level in the control diet, on the NTD and vertebral phenotypes in Apob tm1Unc and Vangl2 Lp mice, hereafter referred to as Apob and Lp respectively. The FA supplemented diet did not reduce the incidence or severity of NTDs in Apob or Lp mutant homozygotes or the loop-tail phenotype in Lp mutant heterozygotes, suggesting that mice with these mutant alleles are resistant to FA supplementation. Folic acid supplementation also did not affect the rate of resorptions or the size of litters, but instead skewed the embryonic genotype distribution in favor of wild-type alleles.ConclusionSimilar genotypic biases have been reported for several NTD models, but were interpreted as diet-induced increases in the incidence and severity of NTDs that led to increased embryonic lethality. Absence of differences in resorption rates and litter sizes argue against induced embryonic lethality. We suggest an alternative interpretation, namely that FA supplementation led to strongly skewed allelic inheritance, perhaps from disturbances in polyamine metabolism that biases fertilization in favor of wild-type gametes.

Pyruvate dehydrogenase complex (PDC) and pyruvate carboxylase (PC) are mitochondrial enzymes that provide the initial steps of the two main alternatives for pyruvate metabolism: oxidative decarboxylation vs. anaplerotic carboxylation, gluconeogenesis, and glycerogenesis. Assays of the enzymatic activity of these two enzymes in cells and tissues are described in this chapter, based on evolution or fixation of (14)CO(2). These assays are both suitable for use in crude homogenates of cultured skin fibroblasts, lymphocytes, and frozen muscle (PDC) or liver (PC). Activities of these two enzymes are related to spectrophotometric assays of two other mitochondrial enzymes, dihydrolipoamide dehydrogenase (E3) and citrate synthase (CS), providing initial indices of sample integrity and mitochondrial content. These parameters have proven useful for initial detection of inherited human disorders due to deficiencies of these enzymes, and in combination with available genetic analyses can lead to confirmation of specific diagnoses.

Purpose Clinical genome sequencing (cGS) followed by orthogonal confirmatory testing is standard practice. While orthogonal testing significantly improves specificity, it also results in increased turnaround time and cost of testing. The purpose of this study is to evaluate machine learning models trained to identify false positive variants in cGS data to reduce the need for orthogonal testing. Methods We sequenced five reference human genome samples characterized by the Genome in a Bottle Consortium (GIAB) and compared the results with an established set of variants for each genome referred to as a truth set. We then trained machine learning models to identify variants that were labeled as false positives. Results After training, the models identified 99.5% of the false positive heterozygous single-nucleotide variants (SNVs) and heterozygous insertions/deletions variants (indels) while reducing confirmatory testing of nonactionable, nonprimary SNVs by 85% and indels by 75%. Employing the algorithm in clinical practice reduced overall orthogonal testing using dideoxynucleotide (Sanger) sequencing by 71%. Conclusion Our results indicate that a low false positive call rate can be maintained while significantly reducing the need for confirmatory testing. The framework that generated our models and results is publicly available at https://github.com/HudsonAlpha/STEVE.

The coronavirus disease 2019 (COVID-19) emerged in early 2020 and since, has brought about tremendous cost to economies and healthcare systems universally. Reports of pediatric patients with inherited conditions and COVID-19 infections are emerging. Specific risks for morbidity and mortality that this pandemic carries for different categories of genetic disorders are still mostly unknown. Thus, there are no specific recommendations for the diagnosis, management, and treatment of patients with genetic disorders during the COVID-19 or other pandemics. Emerging publications, from Upper-Middle Income countries (UMIC), discuss the recent experiences of genetic centers in the continuity of care for patients with genetic disorders in the context of this pandemic. Many measures to facilitate the plan to continuous genetic care in a well-developed health system, may not be applicable in Low and Middle Income countries (LMIC). With poorly structured health systems and with the lack of established genetic services, the COVID-19 pandemic will easily exacerbate the access to care for patients with genetic disease in these countries. This article focuses on the unique challenges of providing genetic healthcare services during emergency situations in LMIC countries and provides practical preparations for this and other pandemic situations.

Purpose: Neurodevelopmental disorders (NDDs) often result from rare genetic variation, but genomic testing yield for NDDs remains around 50%, suggesting some clinically relevant rare variants may be missed by standard analyses. Here we analyze poison exons (PEs) which, while often absent from standard gene annotations, are alternative exons whose inclusion results in a premature termination codon. Variants that alter PE inclusion can lead to loss-of-function and may be highly penetrant contributors to disease. Methods: We curated published RNA-seq data from developing mouse cortex to define 1,937 PE regions conserved between humans and mice and potentially relevant to NDDs. We then analyzed variants found by genome sequencing in multiple NDD cohorts. Results: Across 2,999 probands, we found six clinically relevant variants in PE regions that were previously overlooked. Five of these variants are in genes that are part of the sodium voltage-gated channel alpha subunit family (SCN1A, SCN2A, and SCN8A), associated with epilepsies. One variant is in SNRPB, associated with Cerebrocostomandibular Syndrome. These variants have moderate to high computational impact assessments, are absent from population variant databases, and were observed in probands with features consistent with those reported for the associated gene. Conclusion: With only a minimal increase in variant analysis burden (most probands had zero or one candidate PE variants in a known NDD gene, with an average of 0.77 per proband), annotation of PEs can improve diagnostic yield for NDDs and likely other congenital conditions.