Unrelated Cases Research Articles

Impaired muscle morphology in a Drosophila model of myosin storage myopathy was supressed by overexpression of an E3 ubiquitin ligase.

ABSTRACTMyosin is vital for body movement and heart contractility. Mutations in MYH7, encoding slow/β-cardiac myosin heavy chain, are an important cause of hypertrophic and dilated cardiomyopathy, as well as skeletal muscle disease. A dominant missense mutation (R1845W) in MYH7 has been reported in several unrelated cases of myosin storage myopathy. We have developed a Drosophila model for a myosin storage myopathy in order to investigate the dose-dependent mechanisms underlying the pathological roles of the R1845W mutation. This study shows that a higher expression level of the mutated allele is concomitant with severe impairment of muscle function and progressively disrupted muscle morphology. The impaired muscle morphology associated with the mutant allele was suppressed by expression of Thin (herein referred to as Abba), an E3 ubiquitin ligase. This Drosophila model recapitulates pathological features seen in myopathy patients with the R1845W mutation and severe ultrastructural abnormalities, including extensive loss of thick filaments with selective A-band loss, and preservation of I-band and Z-disks were observed in indirect flight muscles of flies with exclusive expression of mutant myosin. Furthermore, the impaired muscle morphology associated with the mutant allele was suppressed by expression of Abba. These findings suggest that modification of the ubiquitin proteasome system may be beneficial in myosin storage myopathy by reducing the impact of MYH7 mutation in patients.

DYNC2H1 hypomorphic or retina-predominant variants cause nonsyndromic retinal degeneration

PurposeDetermining the role of DYNC2H1 variants in nonsyndromic inherited retinal disease (IRD). MethodsGenome and exome sequencing were performed for five unrelated cases of IRD with no identified variant. In vitro assays were developed to validate the variants identified (fibroblast assay, induced pluripotent stem cell [iPSC] derived retinal organoids, and a dynein motility assay). ResultsFour novel DYNC2H1 variants (V1, g.103327020_103327021dup; V2, g.103055779A>T; V3, g.103112272C>G; V4, g.103070104A>C) and one previously reported variant (V5, g.103339363T>G) were identified. In proband 1 (V1/V2), V1 was predicted to introduce a premature termination codon (PTC), whereas V2 disrupted the exon 41 splice donor site causing incomplete skipping of exon 41. V1 and V2 impaired dynein-2 motility in vitro and perturbed IFT88 distribution within cilia. V3, homozygous in probands 2–4, is predicted to cause a PTC in a retina-predominant transcript. Analysis of retinal organoids showed that this new transcript expression increased with organoid differentiation. V4, a novel missense variant, was in trans with V5, previously associated with Jeune asphyxiating thoracic dystrophy (JATD). ConclusionThe DYNC2H1 variants discussed herein were either hypomorphic or affecting a retina-predominant transcript and caused nonsyndromic IRD. Dynein variants, specifically DYNC2H1 variants are reported as a cause of non syndromic IRD.

Convex hulls in hamming space enable efficient search for similarity and clustering of genomic sequences

BackgroundIn molecular epidemiology, comparison of intra-host viral variants among infected persons is frequently used for tracing transmissions in human population and detecting viral infection outbreaks. Application of Ultra-Deep Sequencing (UDS) immensely increases the sensitivity of transmission detection but brings considerable computational challenges when comparing all pairs of sequences. We developed a new population comparison method based on convex hulls in hamming space. We applied this method to a large set of UDS samples obtained from unrelated cases infected with hepatitis C virus (HCV) and compared its performance with three previously published methods.ResultsThe convex hull in hamming space is a data structure that provides information on: (1) average hamming distance within the set, (2) average hamming distance between two sets; (3) closeness centrality of each sequence; and (4) lower and upper bound of all the pairwise distances among the members of two sets. This filtering strategy rapidly and correctly removes 96.2% of all pairwise HCV sample comparisons, outperforming all previous methods. The convex hull distance (CHD) algorithm showed variable performance depending on sequence heterogeneity of the studied populations in real and simulated datasets, suggesting the possibility of using clustering methods to improve the performance. To address this issue, we developed a new clustering algorithm, k-hulls, that reduces heterogeneity of the convex hull. This efficient algorithm is an extension of the k-means algorithm and can be used with any type of categorical data. It is 6.8-times more accurate than k-mode, a previously developed clustering algorithm for categorical data.ConclusionsCHD is a fast and efficient filtering strategy for massively reducing the computational burden of pairwise comparison among large samples of sequences, and thus, aiding the calculation of transmission links among infected individuals using threshold-based methods. In addition, the convex hull efficiently obtains important summary metrics for intra-host viral populations.

Comprehensive Description of Monoallelic GP1BA Variants Associated with Thrombocytopenia

GP1BA encodes the glycoprotein (GP) 1bα subunit of the GP1b/IX/V complex, the platelet receptor for von Willebrand factor (VWF) and a mediator of outside-in signalling that contributes to platelet activation. Biallelic variants in GPIBA that prevent surface expression of GPIB/IX/V cause the recessive disorder Bernard Soulier Syndrome (BSS; OMIM #23100) in which there is often severe bleeding caused by thrombocytopenia and platelet dysfunction. Thrombocytopenia has also been associated with monoallelic missense variants in the GP1bα R loop (residues 227-242) which enhance VWF binding (platelet-type von Willebrand disease (pVWD); OMIM #177820). Other monoallelic GPIBA missense variants such as the Bolzano variant (p.Ala172Val; OMIM #153670) have been associated with dominant thrombocytopenia without altered VWF levels. In order to better describe the relationship between monoallelic GPIBA variants and thrombocytopenia we evaluated data from the NIHR BioResource for Rare Diseases, (NBR-RD) and the Inherited Platelet Disorders programmes at the Universities of Birmingham (UK), Murcia (Spain) and Toronto (Canada) in which patients with unexplained platelet disorders underwent diagnostic next generation sequencing. Using the BeviMed method for genetic association we compared the genotypes at rare nonsynoymous variants of 105 unrelated thrombocytopenia cases (platelet count (PLT) <130x109/L or the human phenotype ontology term 'HP 001873 thrombocytopenia') in the NBR-BR with those of 10,152 unrelated participants without thrombocytopenia. There was a strong statistical association (posterior probability 0.97) between dominantly inherited GPIBA variants and thrombocytopenia. Considering the five variants within the NBR-BR that drove this association alongside genotype data from the other case collections, we identified a total of 21 different monoallelic GPIBA coding variants that were rare (gnomAD allele frequency <1/1000) and had a CADD pathogenicity score >15. These were present in 29 index cases (18 females, median age 37 years) with unexplained thrombocytopenia and included the variant predicting the p.Asn150Ser substitution which was identified in seven independent index cases. The variants comprised 18 missense, two inframe insertions (one with an additional missense change) and one insertion resulting in a frame shift. Seven had been previously associated with thrombocytopenia. Sixteen of the variants predicted amino acid substitutions or single residue insertions within the GP1bα leucine rich repeat region (LRR; residues 48-200) or in adjacent regions of the LRR N- and C-terminal caps. Cases with variants in the LRR region typically displayed mild mucocutaneous bleeding, abnormal bleeding after minor trauma or surgery and heavy menstrual bleeding. The platelet count was typically 60-100 x109/L and mean platelet volume greater than 11 fL. There were two missense variants in the GP1bα R loop that have been previously associated with pVWD and two further previously unreported missense variants in the cytoplasmic domain. The final variant was a monoallelic single nucleotide insertion predicted to cause a frame shift and absent expression of GP1bα and has been previously reported as a biallelic variant associated with BSS. These data indicate that in a multicentre cross-sectional cohort of cases with otherwise unexplained thrombocytopenia, monoallelic missense GPIBA variants affecting the LRR or adjacent regions were more prevalent than R loop variants associated with pVWD or variants causing absent GP1bα expression associated with biallelic BSS. The LRR region contains the GP1bα-VWF binding site which is predicted to be disrupted by the missense variants observed in this study. The LRR region is also the location of the GP1bα Bolzano variant and six other monoallelic variants previously associated with thrombocytopenia in previous single case reports. These observations extend the repertoire of GPIBA variants associated with thrombocytopenia and although requiring experimental confirmation of pathogenicity, suggest a common molecular pathogenesis for dominant GPIBA-associated thrombocytopenia in which reduced circulating platelet numbers arises from defective GP1bα-VWF interactions. Figure Disclosures Turro: Tachyon Ventures: Other: Partner & Scientific Director.

Incident Reports of Naming Errors among Two Sets of Infant Twins.

Introduction:Newborns are at high risk for identification errors due to their inability to speak and indistinguishable features. To reduce this risk, The Joint Commission requires hospitals to use a distinct identification method for newborns. Most hospitals create medical records for newborns at birth using temporary naming conventions, resulting in patients with similar identifiers. Typically, multiple-birth infants are distinguished from their siblings by a single character (1, 2, or A, B), placing them at higher risk for identification errors, which can delay care and compromise patient safety.Methods:We present 2 unrelated cases involving naming errors in sets of infant twins receiving care in a healthcare system using Joint Commission compliant distinct temporary naming convention.Results:In the 2 cases, system failures contributed to naming errors in 2 sets of infant twins, which resulted in delayed care. In the first case, twins were inadvertently assigned the same temporary name. In the second case, an infant’s blood specimen label did not include a single character, which distinguishes a multiple-birth infant from their sibling. Further safeguards are needed to reduce this risk. These cases illustrated the potential for misidentification related to newborn naming conventions during the registration process, especially between siblings of multiple-birth infants.Conclusions:Further research is needed to determine strategies to prevent newborn identification errors. Potential strategies to reduce this risk and protect newborns include improving the design of newborn identifiers, systems-level interventions such as verification alerts, and improved registration processes.

Consensus Multilocus Sequence Typing Scheme for Pneumocystis jirovecii.

Pneumocystis jirovecii is an opportunistic human pathogenic fungus causing severe pneumonia mainly in immunocompromised hosts. Multilocus sequence typing (MLST) remains the gold standard for genotyping of this unculturable fungus. However, the lack of a consensus scheme impedes a global comparison, large scale population studies and the development of a global MLST database. To overcome this problem this study compared all genetic regions (19 loci) currently used in 31 different published Pneumocystis MLST schemes. The most diverse/commonly used eight loci, β-TUB, CYB, DHPS, ITS1, ITS1/2, mt26S and SOD, were further assess for their ability to be successfully amplified and sequenced, and for their discriminatory power. The most successful loci were tested to identify genetically related and unrelated cases. A new consensus MLST scheme consisting of four genetically independent loci: β-TUB, CYB, mt26S and SOD, is herein proposed for standardised P. jirovecii typing, successfully amplifying low and high fungal burden specimens, showing adequate discriminatory power, and correctly identifying suspected related and unrelated isolates. The new consensus MLST scheme, if accepted, will for the first time provide a powerful tool to investigate outbreak settings and undertake global epidemiological studies shedding light on the spread of this important human fungal pathogen.

Bi-Allelic Novel Variants in CLIC5 Identified in a Cameroonian Multiplex Family with Non-Syndromic Hearing Impairment

DNA samples from five members of a multiplex non-consanguineous Cameroonian family, segregating prelingual and progressive autosomal recessive non-syndromic sensorineural hearing impairment, underwent whole exome sequencing. We identified novel bi-allelic compound heterozygous pathogenic variants in CLIC5. The variants identified, i.e., the missense [NM_016929.5:c.224T>C; p.(L75P)] and the splicing (NM_016929.5:c.63+1G>A), were validated using Sanger sequencing in all seven available family members and co-segregated with hearing impairment (HI) in the three hearing impaired family members. The three affected individuals were compound heterozygous for both variants, and all unaffected individuals were heterozygous for one of the two variants. Both variants were absent from the genome aggregation database (gnomAD), the Single Nucleotide Polymorphism Database (dbSNP), and the UK10K and Greater Middle East (GME) databases, as well as from 122 apparently healthy controls from Cameroon. We also did not identify these pathogenic variants in 118 unrelated sporadic cases of non-syndromic hearing impairment (NSHI) from Cameroon. In silico analysis showed that the missense variant CLIC5-p.(L75P) substitutes a highly conserved amino acid residue (leucine), and is expected to alter the stability, the structure, and the function of the CLIC5 protein, while the splicing variant CLIC5-(c.63+1G>A) is predicted to disrupt a consensus donor splice site and alter the splicing of the pre-mRNA. This study is the second report, worldwide, to describe CLIC5 involvement in human hearing impairment, and thus confirms CLIC5 as a novel non-syndromic hearing impairment gene that should be included in targeted diagnostic gene panels.

Novel GLI3 pathogenic variants in complex pre- and postaxial polysyndactyly and Greig cephalopolysyndactyly syndrome.

Polydactyly is a limb malformation and can occur as nonsyndromic polydactyly, syndromic polydactyly, or along with other limb defects. A few genes have been identified that cause various forms of syndromic and nonsyndromic polydactyly, of which GLI3 has been extensively explored. In the present study, GLI3 gene was screened by direct resequencing in 15 polydactyly cases with or without other anomalies. GLI3 screening revealed two novel pathogenic variants, NM_000168.6:c.3414delC [p.(H1138Qfs*68)] and NM_000168.6:c.1862C>T [p.(P621L)], found in two unrelated cases of familial complex pre- and postaxial polysyndactyly and sporadic Greig cephalopolysyndactyly syndrome (GCPS), respectively. The first pathogenic GLI3 variant, NM_000168.6:c.3414delC, causes premature protein truncation at the C-terminal domain of GLI3. Alternatively, the second pathogenic variant, NM_000168.6:c.1862C>T, lies in the DNA binding domain of GLI3 protein and may affect its hydrophobic interaction with DNA. Both pathogenic GLI3 variants had reduced transcriptional activity in HEK293 cells that likely had led to haploinsufficiency and, consequently, the clinical phenotypes. Overall, the present study reports a novel familial case of complex pre- and postaxial polysyndactyly and the underlying novel pathogenic GLI3 variant expanding the clinical criteria for GLI3 mutational spectrum to complex pre- and postaxial polysyndactyly. Furthermore, this study also reports a novel GLI3 pathogenic variant linked to GCPS, highlighting the known genotype-phenotype correlation.

Landscape of Public T Cell Receptors Associated with Recovery from COVID-19

T cells play an essential role in COVID-19, but characteristics of beneficial clones remain unclear. By analysis of clonal trajectories, we identified recovery-associated expanding T cell clones many of which carrying public T cell receptors (TCRs). Mining the immune repertoires of unrelated COVID-19 cases for these sequences revealed T cells with exact TCR complementarity-determining region 3 identity that were more abundant in recovering than in fatal cases. These TCRs were also found in subjects not previously exposed to the virus, with lower representation in repertoires from risk groups like aged individuals or cancer patients. Together, our data indicate that a significant part of the recovery-associated T cell response in COVID-19 may be mediated by public TCRs that are present in repertoires of unexposed individuals. The lower representation of these clones in repertoires of risk groups or failure to expand such clones may contribute to more unfavorable clinical COVID-19 courses.Funding: This project was partially funded by the CRC 841 of the German Research Foundation (toMB) as well as by the Martin-Luther-University Halle (Saale).Conflict of Interest: The authors declare no competing interests.Ethical Approval: Blood collection of COVID-19 patients was performed under institutional review board approvals number 2020-039 and 11/17. Blood collection of pre-pandemic samples was performed under approval of ethics committee of the medical association Hamburg (project number PV4767) and the ethics committee of the medical faculty of MartinLuther-University Halle-Wittenberg (project number 2014-75). Written informed consent was received from all participants. The study has been performed in accordance with the declaration of Helsinki of 1975.

CONGENITAL MYOPATHIES 2: P.93 Novel TTN variants identified by whole exome sequencing in patients with prenatal onset titinopathy

The TTN gene encodes the largest human protein, titin, which is of functional and structural importance at the sarcomere. Pathogenic variants in TTN have been reported for a variety of skeletal muscle disorders and cardiomyopathies and exhibit both autosomal dominant and recessive inheritance. Next generation sequencing (NGS) and whole exome sequencing (WES) have allowed for routine sequencing of a large number of genes, including the full TTN gene. These advancements have helped to delineate the phenotypic spectrum in patients with mutations in several neuromuscular genes, including TTN. In 2014, the first patient with arthrogryposis multiplex congenita (AMC) and pathogenic variants in TTN was reported, but additional published cases are limited. Utilizing WES, we have identified four unrelated cases in which the proband presented prenatally or neonatally with features suggestive of a severe neuromuscular condition such as polyhydramnios, hydrops fetalis, talipes equinovarus, arthrogryposis, hypotonia, and/or respiratory insufficiency. In one of these cases, the maternal family history was also suggestive for dilated cardiomyopathy in two siblings. Case one - homozygous c.25063+1G>A (I-band). Case two - c.100296dup (p.Glu33433*, M-band) and c.36402del (p.Val12135Cysfs*23, I-band). Case three - c.38442dupA (p.Pro12815Thrfs*37, I-band) and c.88594+1G>T (A-band). Case four has a maternally inherited c.61921C>T (p.Arg20641*, A-band); however, a second paternal variant was not identified. Although truncating variants in the Z-disc, I-band and M-band in TTN may not confer cardiomyopathy risk, some of these variants may be pathogenic for severe, recessive neuromuscular conditions. TTN variants, in particular protein truncating variants, should be considered in cases of lethal congenital contracture syndrome, AMC, congenital myopathies, and hydrops fetalis. These cases expand our knowledge and understanding of the more severe phenotypic spectrum for titinopathies.

Impact of vitaminD receptor gene polymorphisms on vitiligo susceptibility and clinical features inaSoutheastern European Caucasian population.

An association of vitamin D receptor (VDR) polymorphisms and vitiligo has been suggested. However, previous studies have reported contradictory results while including limited data among Caucasians. The aim of this single-center study was to evaluate the effect of three common VDR gene polymorphisms (FokI, TaqI and BsmI) on suscep-tibility and clinical aspects of vitiligo in a Southeastern European Caucasian population. A total of 110 unrelated vitiligo cases and 509 general population controls were enrolled from October 2018 to November 2019. Genomic DNA was extracted from whole blood after de-identification and anonymization of the samples and genotyped for the selected VDR polymorphisms by the qPCR (melting curve analysis). Subgroup analysis by clinical features among subsets of patients indicated that, compared to subjects with the FokI TT genotype or T allele, carriers of the FokI CC genotype or C allele exhibited significantly decreased risk of developing vitiligo before the age of 30 [TT vs. CC: odds ratio (OR)=0.286, 95% confidence interval (CI): 0.083-0.984, P=0.041; T vs. C: OR=0.545, 95% CI: 0.313-0.948, P=0.031]. Intra-patient analysis also revealed that, compared to T allele, the presence of TaqI C allele was adversely associated with the incidence of concurrent leukotrichia (T vs. C: OR=1.874, 95% CI: 1.018-3.451, P=0.042). Comparisons between the case and control groups showed no evidence to support an association between susceptibility to vitiligo and the VDR BsmI, TaqI, and FokI polymorphisms in this cohort. Thus, the studied VDR polymorphisms might indirectly impact the clinical course and treatment decision-making despite their lack of association with vitiligo per se. Further research with larger sample sizes, especially across Caucasian individuals, should be performed to confirm these findings.

Young-onset colorectal cancer is associated with a personal history of type 2 diabetes.

Colorectal cancer (CRC) is rising in incidence in young adults, and this observation is currently unexplained. We investigated whether having a personal history of type 2 diabetes mellitus (T2D) was a potential risk factor for young-onset colorectal cancer (YOCRC). The South Australian Young Onset (SAYO) CRC study is a series of young adults with CRC below age 55. Ninety unrelated YOCRC cases were recruited to the study. Personal history and detailed family history of T2D were obtained at face-to-face interview and confirmed from medical records. Whole exome sequencing was conducted on germline DNA from each CRC case. Controls for personal history studies of T2D were 240 patients with proven clear colonoscopies and no known CRC predispositions. The median age of YOCRC cases was 44 years (18-54) and of controls was 45 years (18-54), and 53% of both cases and controls were females (P=0.99). Left-sided (distal) CRC was seen in 67/89 (75%) of cases. A personal history of T2D was confirmed in 17/90 (19%) YOCRC patients compared with controls (12/240, 5%; P<0.001; odds ratio = 4.4; 95% confidence interval, 2.0-9.7). YOCRC patients frequently reported at least one first-degree relative with T2D (32/85, 38%). Ten of 87 (12%) of YOCRC cases had CRC-related pathogenic germline variants, however, no pathogenic variants in familial diabetes-associated genes were seen. Though the mechanism remains unclear, our observations suggest that there is enrichment for personal history of T2D in YOCRC patients. A diagnosis of T2D could therefore potentially identify a subset of young adults at increased risk for CRC and in whom early screening might be appropriate.

CLL-264: CLL Patients with Identical IGHV Genes

Background Studies over the last 20 years demonstrate a significant IGHV repertoire narrowing in CLL. For this disease, the existence of more than 200 subsets of tumor IGHV antigen-binding sites, so called stereotypical antigen receptors (SAR), has been shown among unrelated cases. Most common subsets (19 SARs) are responsible for more than 12% of all cases of CLL. Methods We assessed nucleotide sequences and CDR3 composition in 1038 CLL patients diagnosed and followed up from 2008 to 2020. Results 132 patients had one of major 19 SARs. Among all patients with SAR, three pairs of patients were identified whose CDR3 amino acid sequences matched 100%. In the first pair (with CLL#1), the IGHV complete nucleotide sequences were differed significantly. One patient had 100% homology with IGHV1-3*01 gene, the other had 99.7% similarity with IGHV1-2*02 gene. There were 34 differences in their nucleotide sequences; 4 were in the CDR3 region. In a pair of patients with CLL#2 SAR, both expressed IGHV3-21*01 gene with homology 96.6% and 97.2%. As a result, the complete nucleotide sequences of these patients were differed in 14 positions; 1 was in the CDR3 region. The most interesting case was the third pair, with the expression of CLL#28A. Their nucleotide sequences were composed from IGHV1-2*06 (100% homology), IGHD1-26*01 and IGHJ6*02 and were almost identical, differing by 1 nucleotide in the CDR3 region only. Conclusions These findings support additional arguments in favour of the significant role of antigen stimulation in the pathogenesis of CLL. Studies over the last 20 years demonstrate a significant IGHV repertoire narrowing in CLL. For this disease, the existence of more than 200 subsets of tumor IGHV antigen-binding sites, so called stereotypical antigen receptors (SAR), has been shown among unrelated cases. Most common subsets (19 SARs) are responsible for more than 12% of all cases of CLL. We assessed nucleotide sequences and CDR3 composition in 1038 CLL patients diagnosed and followed up from 2008 to 2020. 132 patients had one of major 19 SARs. Among all patients with SAR, three pairs of patients were identified whose CDR3 amino acid sequences matched 100%. In the first pair (with CLL#1), the IGHV complete nucleotide sequences were differed significantly. One patient had 100% homology with IGHV1-3*01 gene, the other had 99.7% similarity with IGHV1-2*02 gene. There were 34 differences in their nucleotide sequences; 4 were in the CDR3 region. In a pair of patients with CLL#2 SAR, both expressed IGHV3-21*01 gene with homology 96.6% and 97.2%. As a result, the complete nucleotide sequences of these patients were differed in 14 positions; 1 was in the CDR3 region. The most interesting case was the third pair, with the expression of CLL#28A. Their nucleotide sequences were composed from IGHV1-2*06 (100% homology), IGHD1-26*01 and IGHJ6*02 and were almost identical, differing by 1 nucleotide in the CDR3 region only. These findings support additional arguments in favour of the significant role of antigen stimulation in the pathogenesis of CLL.

Analysis of rare variants of autosomal-dominant genes in a Chinese population with sporadic Parkinson's disease.

BackgroundTo date, several studies have suggested that genes involved in monogenic forms of Parkinson's disease (PD) contribute to unrelated sporadic cases, but there is limited evidence in the Chinese population.MethodsWe performed a systematic analysis of 12 autosomal‐dominant PD (AD‐PD) genes (SNCA, LRRK2, GIGYF2, VPS35, EIF4G1, DNAJC13, CHCHD2, HTRA2, NR4A2, RIC3, TMEM230, and UCHL1) using panel sequencing and database filtration in a case‐control study of a cohort of 391 Chinese sporadic PD patients and unrelated controls. We evaluated the association between candidate variants and sporadic PD using gene‐based analysis.ResultsOverall, 18 rare variants were discovered in 18.8% (36/191) of the index patients. In addition to previously reported pathogenic mutations (LRRK2 p.Arg1441His and p.Ala419Val), another four unknown variants were found in LRRK2, which also contribute to PD risk (p = 0.002; odds ratio (OR) = 7.83, 95% confidence intervals (CI) = 1.76–34.93). The cumulative frequency of undetermined rare variants was significantly higher in PD patients (14.1%) than in controls (3.5%) (p = 0.0002; OR=4.54, 95% CI = 1.93‐10.69).ConclusionOur results confirm the strong impact of LRRK2 on the risk of sporadic PD, and also provide considerable evidence of the existence of additional undetermined rare variants in AD‐PD genes that contribute to the genetic etiology of sporadic PD in a Chinese cohort.

Expanding the phenotype of CRYAA nucleotide variants to a complex presentation of anterior segment dysgenesis

BackgroundMutations in CRYAA, which encodes the α-crystallin protein, are associated with a spectrum of congenital cataract–microcornea syndromes.ResultsIn this study, we performed clinical examination and subsequent genetic analysis in two unrelated sporadic cases of different geographical origins presenting with a complex phenotype of ocular malformation. Both cases manifested bilateral microphthalmia and severe anterior segment dysgenesis, primarily characterized by congenital aphakia, microcornea, and iris hypoplasia/aniridia. NGS-based analysis revealed two novel single nucleotide variants occurring de novo and affecting the translation termination codon of the CRYAA gene, c.520T > C and c.521A > C. Both variants are predicted to elongate the C-terminal protein domain by one-third of the original length.ConclusionsOur report not only expands the mutational spectrum of CRYAA but also identifies the genetic cause of the unusual ocular phenotype described in this report.

Identification of novel candidate genes in heterotaxy syndrome patients with congenital heart diseases by whole exome sequencing

Heterotaxy syndrome (HS) involves dysfunction of multiple systems resulting from abnormal left-right (LR) body patterning. Most HS patients present with complex congenital heart diseases (CHD), the disability and mortality of HS patients are extremely high. HS has great heterogeneity in phenotypes and genotypes, which have rendered gene discovery challenging. The aim of this study was to identify novel genes that underlie pathogenesis of HS patients with CHD. Whole exome sequencing was performed in 25 unrelated HS cases and 100 healthy controls; 19 nonsynonymous variants in 6 novel candidate genes (FLNA, ITGA1, PCNT, KIF7, GLI1, KMT2D) were identified. The functions of candidate genes were further analyzed in zebrafish model by CRISPR/Cas9 technique. Genome-editing was successfully introduced into the gene loci of flna, kmt2d and kif7, but the phenotypes were heterogenous. Disruption of each gene disturbed normal cardiac looping while kif7 knockout had a more prominent effect on liver budding and pitx2 expression. Our results revealed three potential HS pathogenic genes with probably different molecular mechanisms.

An exome-wide exploration of cases of primary ovarian insufficiency uncovers novel sequence variants and candidate genes.

Primary ovarian insufficiency (POI) implies the cessation of menstruation for several months in women before the age of 40 years and is a major cause of infertility. The study of the contribution of genetic factors to POI has been fueled by the use of whole exome sequencing (WES). Here, to uncover novel causative pathogenic variants and risk alleles, WES has been performed in 12 patients with familial POI (eight unrelated index cases and two pairs of sisters) and six women with early menopause and family history of POI (four index cases and one pair of sisters). Likely causative variants in NR5A1 and MCM9 genes were identified as well as a variant in INHA that requires further investigation. Moreover, we have identified more than one candidate variant in 3 out of 15 familial cases. Taken together, our results highlight the genetic heterogeneity of POI and early menopause and support the hypothesis of an oligogenic inheritance of such conditions, in addition to monogenic inheritance.

Multiplex melanoma families are enriched for polygenic risk.

Cancers, including cutaneous melanoma, can cluster in families. In addition to environmental etiological factors such as ultraviolet radiation, cutaneous melanoma has a strong genetic component. Genetic risks for cutaneous melanoma range from rare, high-penetrance mutations to common, low-penetrance variants. Known high-penetrance mutations account for only about half of all densely affected cutaneous melanoma families, and the causes of familial clustering in the remainder are unknown. We hypothesize that some clustering is due to the cumulative effect of a large number of variants of individually small effect. Common, low-penetrance genetic risk variants can be combined into polygenic risk scores. We used a polygenic risk score for cutaneous melanoma to compare families without known high-penetrance mutations with unrelated melanoma cases and melanoma-free controls. Family members had significantly higher mean polygenic load for cutaneous melanoma than unrelated cases or melanoma-free healthy controls (Bonferroni-corrected t-test P = 1.5 × 10−5 and 6.3 × 10−45, respectively). Whole genome sequencing of germline DNA from 51 members of 21 families with low polygenic risk for melanoma identified a CDKN2A p.G101W mutation in a single family but no other candidate high-penetrance melanoma susceptibility genes. This work provides further evidence that melanoma, like many other common complex disorders, can arise from the joint action of multiple predisposing factors, including rare high-penetrance mutations, as well as via a combination of large numbers of alleles of small effect.

Genetic polymorphism of HPA1-17 alloantigen system in the Achang and Jingpo populations population in Yunnan.

Human platelet alloantigen (HPA) is part of the platelet membrane structure, named HPA1- 17 system. This study aimed to investigate platelet alloimmunity in the Achang and Jingpo population, ethnic minority specific, to establish a typed platelet donor data bank in Yunnan's ethnic minority areas. In this study, samples from 139 unrelated healthy cases from the Achang population, 148 cases of the Jingpo population, and 150 healthy cases from Yunnan's Han population were collected as a control. PCR-sequence specific primers (PCR-SSP) methods were respectively adopted to genotype HPA-1-17. The frequency of genes and genotypes was calculated separately, and the frequency distribution of alleles in the Achang population was compared with that of the Han population. Monomorphic HPA-7-14 and HPA-16 and 17 were found in the samples from the Han population, while HPA-b was not found in any of these samples. In HPA-1, 2, 4, 5, and 6, aa homozygosity was predominant. Monomorphic HPA-7, HPA-9-14 and HPA-16 were found in the samples from the Achang population, while HPA-b was not found. In HPA-1, 2, 4, 5, 6, 8, and 17, aa homozygosity was predominant. The genotype results of the HPA-4, 7, HPA9-12, HPA14, 16-17 antigen systems in the Jingpo population were all aa, while the HPA-b was not detected. In HPA-1, 2, 5, 6, 8 and 13, aa homozygosity was predominant. HPA-3, 15 showed the greatest heterozygosity among the three populations. The frequency of HPA-1a in the Achang population in Yunnan was significantly different from that in the Han population (P<0.05). There was no difference in the HPA system of the Jingpo population, but the HPA-2a system was different between the Achang and Jingpo populations (P<0.05). In Yunnan, the distribution of the allele polymorphism of HPA-1-17 in the Achang and Jingpo population is similarly distributed to that in the Han population and exhibits their own characteristic. Therefore, a database of platelet donor typing of the ethnic group should be established.

A Novel Link between Early Life Allergen Exposure and Neuroimmune Development in Children

As COVID-19 unprecedented situation significantly increased the time families spend indoors, the awareness of unhealthy living conditions negatively impacting immune system and early neurodevelopment of children is of crucial importance. We retrospectively reviewed unrelated cases of the children with confirmed multiple indoor allergen sensitization due to prolonged exposure to unhealthy indoor environment with infestation and water damage, who, in addition to multiple health problems related to allergy and asthma, also developed neuroimmune complications and growth delay. Documented early in life atypical neurologic and behavioral changes were common in all cases. Clinical analysis did not establish other causative reason aside from prenatal and early life exposure to unhealthy living conditions. Alternaria Alternara and Penicillium/Aspergillus molds were found in all homes and sensitization was confirmed in all cases. Significant similarities in the symptoms recorded in all three families led us to a hypothesis that, likely, a significant level of the immune response to external immunogenic pathological stimulus such as mold spore protein, mycotoxin protein, dust mite protein, decay-related volatile particles (VOC) skewed a balance of the neuroimmune interactions, and further affected neuronal network establishment. As all children exhibited significant spectrum of the systemic inflammatory conditions early in life, coupled with inability to follow normal neurodevelopment, we hypothesize that an overwhelming activation of the aggressive immune mechanisms by the epigenetic factors led to glia activation, cytokine storm and break of tolerance. We hypothesize that developing immune system exhibited aggressive responses due to environmental danger signals, subsequently TH-1 or TH-2 switch enables multiple clinical syndromes development with atypical presentation due to the described novel mechanism. An increased due to the COVID-19 lock-down may increase an amount of exposure of vulnerable people to indoor biological particles and volatile organic compounds present in unhealthy buildings. It is of crucial importance to identify and remediate indoor exposure factors that can decrease immune protection, especially against infectious pathogens such as novel coronavirus.