Heterogeneous genetic patterns in bilateral perisylvian polymicrogyria: insights from a Finnish family cohort.

Bilateral perisylvian polymicrogyria is the most common form of regional polymicrogyria within malformations of cortical development, constituting 20% of all malformations of cortical development. Bilateral perisylvian polymicrogyria is characterized by an excessive folding of the cerebral cortex and abnormal cortical layering. Notable clinical features include upper motoneuron dysfunction, dysarthria and asymmetric quadriparesis. Cognitive impairment and epilepsy are frequently observed. To identify genetic variants underlying bilateral perisylvian polymicrogyria in Finland, we examined 21 families using standard exome sequencing, complemented by optical genome mapping and/or deep exome sequencing. Pathogenic or likely pathogenic variants were identified in 5/21 (24%) of families, of which all were confirmed as de novo. These variants were identified in five genes, i.e. DDX23, NUS1, SCN3A, TUBA1A and TUBB2B, with NUS1 and DDX23 being associated with bilateral perisylvian polymicrogyria for the first time. In conclusion, our results confirm the previously reported genetic heterogeneity of bilateral perisylvian polymicrogyria and underscore the necessity of more advanced methods to elucidate the genetic background of bilateral perisylvian polymicrogyria.

A novel variant in CYFIP2 in a girl with severe disabilities and bilateral perisylvian polymicrogyria.

Bilateral perisylvian polymicrogyria (BPP) is a structural malformation of the cerebral cortex that can be caused by several genetic abnormalities. The most common clinical manifestations of BPP include intellectual disability and epilepsy. Cytoplasmic FMRP-interacting protein 2 (CYFIP2) is a protein that interacts with the fragile X mental retardation protein (FMRP). CYFIP2 variants can cause various brain structural abnormalities with the most common clinical manifestations of intellectual disability, epileptic encephalopathy and dysmorphic features. We present a girl with multiple disabilities and BPP caused by a heterozygous, novel, likely pathogenic variant (c.1651G>C: p.(Val551Leu) in the CYFIP2 gene. Our case report broadens the spectrum of genetic diversity associated with BPP by incorporating CYFIP2.

A loss of function variant in AGPAT3 underlies intellectual disability and retinitis pigmentosa (IDRP) syndrome.

Intellectual disability (ID) and retinal dystrophy (RD) are the frequently found features of multiple syndromes involving additional systemic manifestations. Here, we studied a family with four members presenting severe ID and retinitis pigmentosa (RP). Using genome wide genotyping and exome sequencing, we identified a nonsense variant c.747 C > A (p.Tyr249Ter) in exon 7 of AGPAT3 which co-segregates with the disease phenotype. Western blot analysis of overexpressed WT and mutant AGPAT3 in HEK293T cells showed the absence of AGPAT3, suggesting instability of the truncated protein. Knockdown of Agpat3 in the embryonic mouse brain caused marked deficits in neuronal migration, strongly suggesting that reduced expression of AGPAT3 affects neuronal function. Altogether, our data indicates that AGPAT3 activity is essential for neuronal functioning and loss of its activity probably causes intellectual disability and retinitis pigmentosa (IDRP) syndrome.

Variants in EFCAB7 underlie nonsyndromic postaxial polydactyly.

Polydactyly is the most common limb malformation that occurs in 1.6-10.6 per one thousand live births, with incidence varying with ancestry. The underlying gene has been identified for many of the ~100 syndromes that include polydactyly. While for the more common form, nonsydromic polydactyly, eleven candidate genes have been reported. We investigated the underlying genetic cause of autosomal recessive nonsyndromic postaxial polydactyly in four consanguineous Pakistani families. Some family members with postaxial polydactyly also present with syndactyly, camptodactyly, or clinodactyly. Analysis of the exome sequence data revealed two novel homozygous frameshift deletions in EFCAB7: [c.830delG;p.(Gly277Valfs*5)]; in three families and [c.1350_1351delGA;p.(Asn451Phefs*2)] in one family. Sanger sequencing confirmed that these variants segregated with postaxial polydactyly, i.e., family members with postaxial polydactyly were found to be homozygous while unaffected members were heterozygous or wild type. EFCAB7 displays expressions in the skeletal muscle and on the cellular level in cilia. IQCE-EFCAB7 and EVC-EVC2 are part of the heterotetramer EvC complex, which is a positive regulator of the Hedgehog (Hh) pathway, that plays a key role in limb formation. Depletion of either EFCAB7 or IQCE inhibits induction of Gli1, a direct Hh target gene. Variants in IQCE and GLI1 have been shown to cause nonsyndromic postaxial polydactyly, while variants in EVC and EVC2 underlie Ellis van Creveld and Weyers syndromes, which include postaxial polydactyly as a phenotype. This is the first report of the involvement of EFCAB7 in human disease etiology.

Nonsense variant in a consanguineous family expands the phenotype of KPTN gene-related syndrome to include hearing impairment.

A short report with two affected siblings from consanguineous family born with intellectual disability, motor disability, language deficit, and hearing impairment and found to carry biallelic nonsense variant in KPTN gene known to be associated with KPTN gene related syndrome.

A Novel Variant in VPS13B Underlying Cohen Syndrome.

Pathogenic variants in vacuolar protein sorting 13 homolog B (VPS13B) cause Cohen syndrome (CS), a clinically diverse neurodevelopmental disorder. We used whole exome and Sanger sequencing to identify disease-causing variants in a Pakistani family with intellectual disability, microcephaly, facial dysmorphism, neutropenia, truncal obesity, speech delay, motor delay, and insomnia. We identified a novel homozygous nonsense variant c.8841G > A: p.(W2947∗) in VPS13B (NM_017890.5) which segregated with the disease. Sleep disturbances are commonly seen in neurodevelopmental disorders and can exacerbate medical issues if left untreated. We demonstrate that individuals with Cohen syndrome may also be affected by sleep disturbances. In conclusion, we expand the genetic and phenotypic features of Cohen syndrome in the Pakistani population.

Soil Properties Correlate with Microbial Community Structure in Qatari Arid Soils.

This is the first detailed characterization of the microbiota and chemistry of different arid habitats from the State of Qatar. Analysis of bacterial 16S rRNA gene sequences showed that in aggregate, the dominant microbial phyla were Actinobacteria (32.3%), Proteobacteria (24.8%), Firmicutes (20.7%), Bacteroidetes (6.3%), and Chloroflexi (3.6%), though individual soils varied widely in the relative abundances of these and other phyla. Alpha diversity measured using feature richness (operational taxonomic units [OTUs]), Shannon's entropy, and Faith's phylogenetic diversity (PD) varied significantly between habitats (P = 0.016, P = 0.016, and P = 0.015, respectively). Sand, clay, and silt were significantly correlated with microbial diversity. Highly significant negative correlations were also seen at the class level between both classes Actinobacteria and Thermoleophilia (phylum Actinobacteria) and total sodium (R = -0.82 and P = 0.001 and R = -0.86, P = 0.000, respectively) and slowly available sodium (R = -0.81 and P = 0.001 and R = -0.8 and P = 0.002, respectively). Additionally, class Actinobacteria also showed significant negative correlation with sodium/calcium ratio (R = -0.81 and P = 0.001). More work is needed to understand if there is a causal relationship between these soil chemical parameters and the relative abundances of these bacteria. IMPORTANCE Soil microbes perform a multitude of essential biological functions, including organic matter decomposition, nutrient cycling, and soil structure preservation. Qatar is one of the most hostile and fragile arid environments on earth and is expected to face a disproportionate impact of climate change in the coming years. Thus, it is critical to establish a baseline understanding of microbial community composition and to assess how soil edaphic factors correlate with microbial community composition in this region. Although some previous studies have quantified culturable microbes in specific Qatari habitats, this approach has serious limitations, as in environmental samples, approximately only 0.5% of cells are culturable. Hence, this method vastly underestimates natural diversity within these habitats. Our study is the first to systematically characterize the chemistry and total microbiota associated with different habitats present in the State of Qatar.

Rare-variant association analysis reveals known and new age-related hearing loss genes.

Age-related (AR) hearing loss (HL) is a prevalent sensory deficit in the elderly population. Several studies showed that common variants increase ARHL susceptibility. Here, we demonstrate that rare-variants play a crucial role in ARHL etiology. We analyzed exome and imputed data from white-European UK Biobank volunteers, performing both single-variant and rare-variant aggregate association analyses using self-reported ARHL phenotypes. We identified and replicated associations between ARHL and rare-variants in KLHDC7B, PDCD6, MYO6, SYNJ2, and TECTA. PUS7L and EYA4 also revealed rare-variant associations with ARHL. EYA4, MYO6, and TECTA are all known to underline Mendelian nonsyndromic HL. PDCD6, a new HL gene, plays an important role in apoptosis and has widespread inner ear expression, particularly in the inner hair cells. An unreplicated common variant association was previously observed for KHLDC7B, here we demonstrate that rare-variants in this gene also play a role in ARHL etiology. Additionally, the first replicated association between SYNJ2 and ARHL was detected. Analysis of common variants revealed several previously reported, i.e., ARHGEF28, and new, i.e., PIK3R3, ARHL associations, as well as ones we replicate here for the first time, i.e., BAIAP2L2, CRIP3, KLHDC7B, MAST2, and SLC22A7. It was also observed that the odds ratios for rare-variant ARHL associations, were higher than those for common variants. In conclusion, we demonstrate the vital role rare-variants, including those in Mendelian nonsyndromic HL genes, play in the etiology of ARHL.

Syntaxin 4 is essential for hearing in human and zebrafish.

Congenital hearing impairment (HI) is a genetically highly heterogeneous disorder in which prompt recognition and intervention are crucial to optimize outcomes. In this study, we used exome sequencing to investigate a large consanguineous Pakistani family with eight affected individuals showing bilateral severe-to-profound HI. This identified a homozygous splice region variant in STX4 (c.232 + 6T>C), which causes exon skipping and a frameshift, that segregated with HI (two-point logarithm of odds (LOD) score = 5.9). STX4, a member of the syntaxin family, is a component of the SNARE machinery involved in several vesicle transport and recycling pathways. In silico analysis showed that murine orthologue Stx4a is highly and widespread expressed in the developing and adult inner ear. Immunofluorescent imaging revealed localization of STX4A in the cell body, cell membrane and stereocilia of inner and outer hair cells. Furthermore, a morpholino-based knockdown of stx4 in zebrafish showed an abnormal startle response, morphological and developmental defects, and a disrupted mechanotransduction function in neuromast hair cells measured via FM1-43 uptake. Our findings indicate that STX4 dysfunction leads to HI in humans and zebrafish and supports the evolutionary conserved role of STX4 in inner ear development and hair cell functioning.

A novel autosomal dominant GREB1L variant associated with non-syndromic hearing impairment in Ghana.

BACKGROUND: Childhood hearing impairment (HI) is genetically heterogeneous with many implicated genes, however, only a few of these genes are reported in African populations. METHODS: This study used exome and Sanger sequencing to resolve the possible genetic cause of non-syndromic HI in a Ghanaian family. RESULTS: We identified a novel variant c.3041G > A: p.(Gly1014Glu) in GREB1L (DFNA80) in the index case. The GREB1L: p.(Gly1014Glu) variant had a CADD score of 26.5 and was absent from human genomic databases such as TopMed and gnomAD. In silico homology protein modeling approaches displayed major structural differences between the wildtype and mutant proteins. Additionally, the variant was predicted to probably affect the secondary protein structure that may impact its function. Publicly available expression data shows a higher expression of Greb1L in the inner ear of mice during development and a reduced expression in adulthood, underscoring its importance in the development of the inner ear structures. CONCLUSION: This report on an African individual supports the association of GREB1L variant with non-syndromic HI and extended the evidence of the implication of GREB1L variants in HI in diverse populations.

A splice site variant in TCTN3 underlies an atypical form of orofaciodigital syndrome IV.

Orofaciodigital syndrome (OFD) is clinically heterogeneous and is characterized by abnormalities in the oral cavity, facial features, digits, and central nervous system. At least 18 subtypes of the condition have been described in the literature. OFD is caused by variants in several genes with overlapping phenotypes. We studied a consanguineous Pakistani family with two affected siblings with an atypical form of OFD type 4 (OFD4). In addition to the typical features of OFD4 that include limb defects and growth retardation, the siblings displayed rare features of scaphocephaly and seizures. Exome sequencing analysis revealed a novel homozygous splice site variant c.257-1G>A in TCTN3 that segregated with disease. This homozygous splice site variant in TCTN3 is most likely the underlying cause of the atypical form of OFD4 observed in this family. Our results contribute to the phenotypic spectrum of TCTN3 associated ciliopathies and will facilitate better clinical diagnosis.

SCN1A Variants as the Underlying Cause of Genetic Epilepsy with Febrile Seizures Plus in Two Multi-Generational Colombian Families.

Genetic epilepsy with febrile seizures plus (GEFS+) is an autosomal dominant disorder with febrile or afebrile seizures that exhibits phenotypic variability. Only a few variants in SCN1A have been previously characterized for GEFS+, in Latin American populations where studies on the genetic and phenotypic spectrum of GEFS+ are scarce. We evaluated members in two multi-generational Colombian Paisa families whose affected members present with classic GEFS+. Exome and Sanger sequencing were used to detect the causal variants in these families. In each of these families, we identified variants in SCN1A causing GEFS+ with incomplete penetrance. In Family 047, we identified a heterozygous variant (c.3530C > G; p.(Pro1177Arg)) that segregates with GEFS+ in 15 affected individuals. In Family 167, we identified a previously unreported variant (c.725A > G; p.(Gln242Arg)) that segregates with the disease in a family with four affected members. Both variants are located in a cytoplasmic loop region in SCN1A and based on our findings the variants are classified as pathogenic and likely pathogenic, respectively. Our results expand the genotypic and phenotypic spectrum associated with SCN1A variants and will aid in improving molecular diagnostics and counseling in Latin American and other populations.

Exome sequencing of families from Ghana reveals known and candidate hearing impairment genes.

We investigated hearing impairment (HI) in 51 families from Ghana with at least two affected members that were negative for GJB2 pathogenic variants. DNA samples from 184 family members underwent whole-exome sequencing (WES). Variants were found in 14 known non-syndromic HI (NSHI) genes [26/51 (51.0%) families], five genes that can underlie either syndromic HI or NSHI [13/51 (25.5%)], and one syndromic HI gene [1/51 (2.0%)]. Variants in CDH23 and MYO15A contributed the most to HI [31.4% (16/51 families)]. For DSPP, an autosomal recessive mode of inheritance was detected. Post-lingual expression was observed for a family segregating a MARVELD2 variant. To our knowledge, seven novel candidate HI genes were identified (13.7%), with six associated with NSHI (INPP4B, CCDC141, MYO19, DNAH11, POTEI, and SOX9); and one (PAX8) with Waardenburg syndrome. MYO19 and DNAH11 were replicated in unrelated Ghanaian probands. Six of the novel genes were expressed in mouse inner ear. It is known that Pax8-/- mice do not respond to sound, and depletion of Sox9 resulted in defective vestibular structures and abnormal utricle development. Most variants (48/60; 80.0%) have not previously been associated with HI. Identifying seven candidate genes in this study emphasizes the potential of novel HI genes discovery in Africa.

Phenotype Expansion for Atypical Gaucher Disease Due to Homozygous Missense PSAP Variant in a Large Consanguineous Pakistani Family.

Atypical Gaucher disease is caused by variants in the PSAP gene. Saposin C is one of four homologous proteins derived from sequential cleavage of the saposin precursor protein, prosaposin. It is an essential activator for glucocerebrosidase, which is deficient in Gaucher disease. Although atypical Gaucher disease due to deficiency of saposin C is rare, it exhibits vast phenotypic heterogeneity. Here, we report on a Pakistani family that exhibits features of Gaucher disease, i.e., prelingual profound sensorineural hearing impairment, vestibular dysfunction, hepatosplenomegaly, kyphosis, and thrombocytopenia. The family was investigated using exome and Sanger sequencing. A homozygous missense variant c.1076A>C: p.(Glu359Ala) in exon 10 of the PSAP gene was observed in all affected family members. In conclusion, we identified a new likely pathogenic missense variant in PSAP in a large consanguineous Pakistani family with atypical Gaucher disease. Gaucher disease due to a deficiency of saposin C has not been previously reported within the Pakistani population. Genetic screening of patients with the aforementioned phenotypes could ensure adequate follow-up and the prevention of further complications. Our finding expands the genetic and phenotypic spectrum of atypical Gaucher disease due to a saposin C deficiency.

Autosomal recessive nonsyndromic hearing impairment in two Finnish families due to the population enriched CABP2 c.637+1G>T variant.

BACKGROUND: The genetic architecture of hearing impairment in Finland is largely unknown. Here, we investigated two Finnish families with autosomal recessive nonsyndromic symmetrical moderate-to-severe hearing impairment. METHODS: Exome and custom capture next-generation sequencing were used to detect the underlying cause of hearing impairment. RESULTS: In both Finnish families, we identified a homozygous pathogenic splice site variant c.637+1G>T in CAPB2 that is known to cause autosomal recessive nonsyndromic hearing impairment. Four CABP2 variants have been reported to underlie autosomal recessive nonsyndromic hearing impairment in eight families from Iran, Turkey, Pakistan, Italy, and Denmark. Of these variants, the pathogenic splice site variant c.637+1G>T is the most prevalent. The c.637+1G>T variant is enriched in the Finnish population, which has undergone multiple bottlenecks that can lead to the higher frequency of certain variants including those involved in disease. CONCLUSION: We report two Finnish families with hearing impairment due to the CABP2 splice site variant c.637+1G>T.

ADAMTS1, MPDZ, MVD, and SEZ6: candidate genes for autosomal recessive nonsyndromic hearing impairment.

Hearing impairment (HI) is a common disorder of sensorineural function with a highly heterogeneous genetic background. Although substantial progress has been made in the understanding of the genetic etiology of hereditary HI, many genes implicated in HI remain undiscovered. Via exome and Sanger sequencing of DNA samples obtained from consanguineous Pakistani families that segregate profound prelingual sensorineural HI, we identified rare homozygous missense variants in four genes (ADAMTS1, MPDZ, MVD, and SEZ6) that are likely the underlying cause of HI. Linkage analysis provided statistical evidence that these variants are associated with autosomal recessive nonsyndromic HI. In silico analysis of the mutant proteins encoded by these genes predicted structural, conformational or interaction changes. RNAseq data analysis revealed expression of these genes in the sensory epithelium of the mouse inner ear during embryonic, postnatal, and adult stages. Immunohistochemistry of the mouse cochlear tissue, further confirmed the expression of ADAMTS1, SEZ6, and MPDZ in the neurosensory hair cells of the organ of Corti, while MVD expression was more prominent in the spiral ganglion cells. Overall, supported by in silico mutant protein analysis, animal models, linkage analysis, and spatiotemporal expression profiling in the mouse inner ear, we propose four new candidate genes for HI and expand our understanding of the etiology of HI.

Identification of autosomal recessive nonsyndromic hearing impairment genes through the study of consanguineous and non-consanguineous families: past, present, and future.

Hearing impairment (HI) is one of the most common sensory disabilities with exceptionally high genetic heterogeneity. Of genetic HI cases, 30% are syndromic and 70% are nonsyndromic. For nonsyndromic (NS) HI, 77% of the cases are due to autosomal recessive (AR) inheritance. ARNSHI is usually congenital/prelingual, severe-to-profound, affects all frequencies and is not progressive. Thus far, 73 ARNSHI genes have been identified. Populations with high rates of consanguinity have been crucial in the identification of ARNSHI genes, and 92% (67/73) of these genes were identified in consanguineous families. Recent changes in genomic technologies and analyses have allowed a shift towards ARNSHI gene discovery in outbred populations. The latter is crucial towards understanding the genetic architecture of ARNSHI in diverse and understudied populations. We present an overview of the 73 ARNSHI genes, the methods used to identify them, including next-generation sequencing which revolutionized the field, and new technologies that show great promise in advancing ARNSHI discoveries.

Delineating the genotypic and phenotypic spectrum of HECW2-related neurodevelopmental disorders.

BACKGROUND: Variants in HECW2 have recently been reported to cause a neurodevelopmental disorder with hypotonia, seizures and impaired language; however, only six variants have been reported and the clinical characteristics have only broadly been defined. METHODS: Molecular and clinical data were collected from clinical and research cohorts. Massive parallel sequencing was performed and identified individuals with a HECW2-related neurodevelopmental disorder. RESULTS: We identified 13 novel missense variants in HECW2 in 22 unpublished cases, of which 18 were confirmed to have a de novo variant. In addition, we reviewed the genotypes and phenotypes of previously reported and new cases with HECW2 variants (n=35 cases). All variants identified are missense, and the majority of likely pathogenic and pathogenic variants are located in or near the C-terminal HECT domain (88.2%). We identified several clustered variants and four recurrent variants (p.(Arg1191Gln);p.(Asn1199Lys);p.(Phe1327Ser);p.(Arg1330Trp)). Two variants, (p.(Arg1191Gln);p.(Arg1330Trp)), accounted for 22.9% and 20% of cases, respectively. Clinical characterisation suggests complete penetrance for hypotonia with or without spasticity (100%), developmental delay/intellectual disability (100%) and developmental language disorder (100%). Other common features are behavioural problems (88.9%), vision problems (83.9%), motor coordination/movement (75%) and gastrointestinal issues (70%). Seizures were present in 61.3% of individuals. Genotype-phenotype analysis shows that HECT domain variants are more frequently associated with cortical visual impairment and gastrointestinal issues. Seizures were only observed in individuals with variants in or near the HECT domain. CONCLUSION: We provide a comprehensive review and expansion of the genotypic and phenotypic spectrum of HECW2 disorders, aiding future molecular and clinical diagnosis and management.

Genomic analysis of childhood hearing loss in the Yoruba population of Nigeria.

Although variant alleles of hundreds of genes are associated with sensorineural deafness in children, the genes and alleles involved remain largely unknown in the Sub-Saharan regions of Africa. We ascertained 56 small families mainly of Yoruba ethno-lingual ancestry in or near Ibadan, Nigeria, that had at least one individual with nonsyndromic, severe-to-profound, prelingual-onset, bilateral hearing loss not attributed to nongenetic factors. We performed a combination of exome and Sanger sequencing analyses to evaluate both nuclear and mitochondrial genomes. No biallelic pathogenic variants were identified in GJB2, a common cause of deafness in many populations. Potential causative variants were identified in genes associated with nonsyndromic hearing loss (CIB2, COL11A1, ILDR1, MYO15A, TMPRSS3, and WFS1), nonsyndromic hearing loss or Usher syndrome (CDH23, MYO7A, PCDH15, and USH2A), and other syndromic forms of hearing loss (CHD7, OPA1, and SPTLC1). Several rare mitochondrial variants, including m.1555A>G, were detected in the gene MT-RNR1 but not in control Yoruba samples. Overall, 20 (33%) of 60 independent cases of hearing loss in this cohort of families were associated with likely causal variants in genes reported to underlie deafness in other populations. None of these likely causal variants were present in more than one family, most were detected as compound heterozygotes, and 77% had not been previously associated with hearing loss. These results indicate an unusually high level of genetic heterogeneity of hearing loss in Ibadan, Nigeria and point to challenges for molecular genetic screening, counseling, and early intervention in this population.