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Neurosurgeons at NINS further classified cases based on examination, observations during surgery, and review of neuroimaging studies. Cases were eligible to enroll in the study if they were diagnosed with myelomeningocele or meningocele following examination by a physician, were less than a year old, received medical care from NINS, and if parents were able to identify the primary water source used during early pregnancy. Trio designs, which are suited to studying health outcomes that develop early in life (Lupo et al., 2017), can be used to detect de novo mutations and permit assessment using the transmission disequilibrium test to determine if alleles are over-transmitted from parents to case children (Wolujewicz et al., 2021). Biological pathways examined in candidate gene studies of neural tube defects involve the non-canonical Wnt/PCP and SHH signaling pathways (Wolujewicz & Ross, 2019), similar to the animal genetic models of neural tube defects.

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Data from all samples available after filtering for duplicates and failed samples were then merged into a single multi-sample PED file for GWAS analysis. Of the remaining 749 saliva samples with GSA data, 36 duplicates were removed from further analysis. Of the 760 samples sent for analysis, four did not have sufficient quantities of DNA and were excluded from the genetic analysis. Seven hundred sixty saliva DNA samples were shipped to the Broad Institute (Cambridge, MA, USA) for DNA quantification and genetic analysis (Figure 1). Trained study staff collected saliva samples from children and their parents using the Oragene® OGR-575 kit (DNA Genotek Inc., Ottawa, ON, Canada), following the manufacturer’s instructions.

• Genetic association studies, including case-control and child-parent trio designs, have been implemented to explore the relationship between genetic variants and neural tube defect risk in humans.
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• Serum folate concentrations were higher among mothers who reported folic acid use during pregnancy compared to those who did not report folic acid use (10.20 ng/ml and 7.96 ng/ml, respectively).

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A subset of individuals from the case-control study had saliva DNA of sufficient quality and were included in the genetic analyses (Figure 1). Control children were excluded from the main study if the child was diagnosed with cancer or a condition considered to be related to arsenic exposure (i.e. arsenic-related keratosis). Various candidate gene studies of neural tube defects in humans have examined one or a few genes (Finnell et al., 2021), particularly those that are orthologs of risk genes in mice or genes involved in folate pathways (Greene & Copp, 2014). Determinants of neural tube defects include a combination of environmental, nutritional, and genetic risk factors (Greene & Copp, 2014; Harris & Juriloff, 2007). At minimum, more surveillance for spina bifida is needed in high arsenic areas such as Bangladesh.

• We did not find a main effect of mother’s arsenic exposure on spina bifida risk.
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• We hope to validate the findings from this study with future follow-up genetic studies in humans and animals through existing collaborations, and to explore how folate status and arsenic exposure potentially modify the relationship between genetics and spina bifida risk.

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Table 1

Trained staff visited these sites and collected water samples in polyethylene containers. At enrollment, mothers were asked to identify the primary drinking water source they used when they learned they were pregnant. Folate status was additionally assessed in serum samples using Chemiluminescent Microparticle Immunoassay at NINS&H (ARCHITECT plus ci4100, Abbott Company, Abbott Park, IL, USA). We measured infant’s weight and head circumference using standardized protocols, and mother’s medication and vitamin intake using a structured questionnaire that included the seven major types of folic acid-containing tablets in Bangladesh.

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We included families with children with lipomeningocele as controls because our study was primarily designed to investigate associations between arsenic and spina bifida (myelomeningocele and meningocele), and lipomeningocele would be present in the population that would give rise to cases. In our analysis including genetic variants spanning the genome, we did not observe significant differences in measured serum folate among mothers of cases and controls and did not identify any significant associations between genetic variants https://wakelet.com/@spinracasinoen in genes relevant in folate pathways and case status in the child, maternal, or trio models. The genetic association analysis includes data from child-parent trios, allowing us to explore family-based, genome-wide association analyses in a unique study population with myelomeningocele and meningocele, subtypes of spina bifida, from Bangladesh, a country with relatively high prevalence of neural tube defects with a population that is underrepresented in genome-wide association studies (Gurdasani et al., 2019). In the current study, we build upon this previous analysis, performing a genome-wide scan of variants to assess genetic contributors to neural tube defect risk in a larger case-control population. In this study, we investigated the association of genetic variants spanning the genome with risk of spina bifida (i.e., myelomeningocele and meningocele) in a subset of families enrolled from December 2016 through December 2022 in a case-control study in Bangladesh, a population often underrepresented in genetic studies. A recent study from an area of Turkey with high arsenic exposures reported higher levels of arsenic in plasma samples of 100 mothers of NTD cases as compared to mothers from 70 controls .

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The study involved a hospital visit that included a physical examination of the child and their parents, the administration of questionnaires, and the collection of biological samples. BFour samples failed Boston Children’s Hospital Research Computing QC due to one sex mismatch or issues with familial relatedness analysis (i.e. two individuals were determined to have wrong family relationship statuses, and one additional child sample was removed due to subsequent unavailability of parent information). Flowchart of saliva DNA sample selection and case/control family member inclusion in the study.

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