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Here, we aimed to validate a noninvasive method using capture sequencing for prenatal diagnosis of congenital adrenal hyperplasia due to 21-Hydroxylase deficiency (21-OHD). Noninvasive prenatal diagnosis (NIPD) of 21-OHD was based on 14 plasma samples collected from 12 families, including four plasma sample collected during the first trimester. Targeted capture sequencing was performed using genomic DNA from the parents and child trios to determine the pathogenic and wild-type alleles associated with the haplotypes. Maternal plasma DNA was also sequenced to determine the fetal inheritance of the allele using hidden Markov model-based haplotype linkage analysis. The effect of fetal DNA fraction and sequencing depth on the accuracy of NIPD was investigated. The lower limit of fetal DNA fraction was 2% and the threshold mean sequence depth was 38, suggesting potential advantage if used in early gestation. The CYP21A2 genotype of the fetus was accurately determined in all the 14 plasma samples as early as day 1 and 8 weeks of gestation. Results suggest the accuracy and feasibility of NIPD of 21-OHD using a small target capture region with a low threshold for fetal DNA fraction and sequence depth. Our method is cost-effective and suggests diagnostic applications in clinical practice.

During the drum screening of Fritillaria ussuriensis Maxim (FUM), collisions between FUM bulbs are the main cause of mechanical damage. The effects of the mass relationship and the position, direction, and height of the collision on the absorbed energy and mass–loss ratio of FUM were analyzed through collision tests. The results of the constructed regression models showed that the collision direction has no significant effect on the mass–loss ratio, whereas the mass relationship and the position, direction, and height of the collision showed significance. Under different mass relationships, the collision positions and direction, and the collision height are positively correlated with the absorbed energy and mass–loss ratio, and the absorbed energy is also positively correlated with the mass–loss ratio. In addition, the influence of the mass relationship and collision height, position, and direction on the absorbed energy are highly significant. The research findings serve as a valuable reference for designing and developing low‐loss drum screening processes. Practical applications: Fritillaria ussuriensis Maxim (FUM) is a traditional Chinese medicine. The planting area is wide and the economic benefit is high. The annual economic benefit for farmers is about 1.8 billion yuan. However, the existing production method has a low degree of mechanization, especially the harvest stage, which mainly relies on manual harvesting, is limited by the number of labor force and the harvest time is in the rainy season (FUM is easily spoiled in water), which is easy to cause economic losses. To determine better working parameters, the damage factors of FUM during screening were analyzed by developing and testing the drum‐type FUM harvester. It is helpful to improve the production efficiency of FUM, and reduce the labor demand and the damage of screening to FUM. [ABSTRACT FROM AUTHOR]

The impact damage is the most destructive effect on FUM during the drum screening. To explore the impact damage of FUM during drum screening, a free-drop test device was built to simulate the impact damage of FUM. Adopting an L16(45) orthogonal experiment scheme, we investigated the effect of mass, dropping height, collision material, and number of collisions on the bruise depth and specific surface area of bruise, impact peak force, and Ratio of amplitude attenuation. Then, we analyzed the influence of these four factors using a single factor test for each factor. Orthogonal experiments identified the main and secondary factors affecting each outcome; the number of collisions had no significant influence on the impact peak force, while the collision material was the only factor with a significant influence on the Ratio of amplitude attenuation. Single-factor test results indicate that the bruising depth and maximum impact force increase with FUM mass, and the specific surface area of the bruise first increases and then reduces. The maximum impact force and the bruising depth increase with dropping height, and the specific surface area of the bruise periodically changes. The bruising depth and damage surface area of FUM increased with an increased hardness of the collision material, and the maximum impact force and ratio of amplitude attenuation decreased with an increased hardness of the collision material. The specific surface area of the bruise periodically changes with the drop time, and the bruising depth first increases and then decreases. [ABSTRACT FROM AUTHOR]

Objective: This study aimed to validate the feasibility of haplotype-based noninvasive prenatal diagnosis (NIPD) of cobalamin C (cblC) deficiency.Method: This method includes three steps: First, targeted sequencing was performed on 21 families affected by cblC deficiency (including the couples and probands). Second, parental haplotypes linked with the pathogenic variant were determined using the genotypes of trios. Then, the fetal haplotypes were inferred through a parental haplotype assisted hidden Markov model (HMM). The NIPD results were confirmed by using the invasive procedures.Results: Twenty-one fetal genotypes were successfully inferred by NIPD including three compound heterozygotes with cblC deficiency, nine heterozygote carriers of cblC deficiency, and nine normal fetuses. The NIPD results were confirmed using the invasive procedures with 100% concordant rate.Conclusion: This result has shown that haplotype-based NIPD of cblC deficiency has high concordant rate and indicated potential clinical utility as a pregnancy diagnosis method for high-risk carrier couples. [ABSTRACT FROM AUTHOR]

Background: Since the discovery of cell-free foetal DNA in the plasma of pregnant women, many non-invasive prenatal testing assays have been developed. In the area of skeletal dysplasia diagnosis, some PCR-based non-invasive prenatal testing assays have been developed to facilitate the ultrasound diagnosis of skeletal dysplasias that are caused by de novo mutations. However, skeletal dysplasias are a group of heterogeneous genetic diseases, the PCR-based method is hard to detect multiple gene or loci simultaneously, and the diagnosis rate is highly dependent on the accuracy of the ultrasound diagnosis. In this study, we investigated the feasibility of using targeted capture sequencing to detect foetal de novo pathogenic mutations responsible for skeletal dysplasia. Methodology/Principal Findings: Three families whose foetuses were affected by skeletal dysplasia and two control families whose foetuses were affected by other single gene diseases were included in this study. Sixteen genes related to some common lethal skeletal dysplasias were selected for analysis, and probes were designed to capture the coding regions of these genes. Targeted capture sequencing was performed on the maternal plasma DNA, the maternal genomic DNA, and the paternal genomic DNA. The de novo pathogenic variants in the plasma DNA data were identified using a bioinformatical process developed for low frequency mutation detection and a strict variant interpretation strategy. The causal variants could be specifically identified in the plasma, and the results were identical to those obtained by sequencing amniotic fluid samples. Furthermore, a mean of 97% foetal specific alleles, which are alleles that are not shared by maternal genomic DNA and amniotic fluid DNA, were identified successfully in plasma samples. Conclusions/Significance: Our study shows that capture sequencing of maternal plasma DNA can be used to non-invasive detection of de novo pathogenic variants. This method has the potential to be used to facilitate the prenatal diagnosis of skeletal dysplasia. [ABSTRACT FROM AUTHOR]

Here we developed a haplotype-based noninvasive prenatal diagnosis method for hyperphenylalaninemia (HPA) and demonstrated its accuracy and feasibility during early pregnancy. Capture sequencing was performed on genomic DNA from parents and probands using customized hybridization probes targeting highly heterozygous single-nucleotide polymorphisms located within the 1 M region flanking phenylalanine hydroxylase (PAH) and 6-pyruvoyltetrahydropterin (PTS) and its coding region to determine the parental haplotypes and linkage to pathogenic mutations. Maternal plasma DNA obtained at 12-20 weeks of gestation was also subjected to targeted sequencing to deduce the fetal haplotypes based on the parental haplotypes. The fetal genotypes were further validated by invasive prenatal diagnosis. Haplotype-based noninvasive prenatal testing was successfully performed in 13 families. Five fetuses were identified to harbor bi-allelic pathogenic variants of PAH, four fetuses were carriers of one heterozygous PAH variant, three fetuses were normal, and the fetus of the 6-pyruvoyl tetrahydrobiopterin synthase family was identified as normal. The fetal genotypes at two gestational weeks from the same PAH family were identical. All results were consistent with the prenatal diagnosis based on amniotic fluid. Haplotype-based noninvasive prenatal testing for HPA through targeted sequencing is accurate and feasible during early gestation.