OBJECTIVE: This study aims to elucidate the genetic causes of congenital hypogonadotropic hypogonadism (CHH), a rare genetic disorder resulting in GnRH deficiency, in six families from Pakistan.
METHODS: Eighteen DNA samples from six families underwent genome sequencing followed by standard evaluation for pathogenic single nucleotide variants (SNVs) and small indels. All families were subsequently analyzed for pathogenic copy number variants (CNVs) using CoverageMaster.
RESULTS: Novel pathogenic homozygous SNVs in known CHH genes were identified in four families: two families with variants in GNRHR, and two others harboring KISS1R variants. Subsequent investigation of CNVs in the remaining two families identified novel unique large deletions in ANOS1.
CONCLUSIONS: A combined, systematic analysis of single nucleotide and CNVs helps to improve the diagnostic yield for variants in patients with CHH.

BACKGROUND: Oculocutaneous albinism (OCA) is a genetically heterogeneous condition that is associated with reduced or absent melanin pigment in the skin, hair, and eyes, resulting in reduced vision, high sensitivity to light, and rapid and uncontrolled eye movements. To date, seventeen genes have been associated with OCA including syndromic and non-syndromic forms of the condition.
METHODS: Whole exome sequencing (WES) was performed to identify pathogenic variants in nine Pakistani families with OCA, with validation and segregation of candidate variants performed using Sanger sequencing. Furthermore, the pathogenicity of the identified variants was assessed using various in-silico tools and 3D protein structural analysis software.
RESULTS: WES identified biallelic variants in three genes explaining the OCA in these families, including four variants in TYR, three in OCA2, and two in HPS1, including two novel variants c.667C > T: p.(Gln223*) in TYR, and c.2009 T > C: p.(Leu670Pro) in HPS1.
CONCLUSIONS: Overall, this study adds further knowledge of the genetic basis of OCA in Pakistani communities and facilitates improved management and counselling services for families suffering from severe genetic diseases in Pakistan.

BACKGROUND: Breast cancer has emerged as the foremost cause of female mortality worldwide, with triple negative breast cancer accounting for approximately 10-15% of all breast cancer cases. The triple negative breast cancer family has obvious familial heritability, but no potential pathogenic variation was found in BRCA1/2.
METHODS: The patient was a 56-year-old woman of Han ethnicity. The clinical characteristics of this patient with breast cancer were summarized, peripheral blood of one normal female and two patients with breast cancer in this family was collected, DNA was extracted, and the potential pathogenic variation was analyzed by whole exome sequencing. The normal female and two patients with breast cancer in this family shared a maternal grandmother. The proband\'s right breast mass was punctured, and the biopsy showed invasive carcinoma of the right breast, grade II-III, with necrosis. No mutation was found in BRCA1/2 gene test; immunohistochemical of surgical specimens showed triple negative breast cancer. Three mutation types and 17 gene mutation sites were detected through bioinformatics prediction analysis on the basis of co-segregation of genotype and phenotype within the family and whole exome sequencing results. Combined with the Cancer Genome Atlas database comprehensive analysis, the MT1E c.G107A (p.C36Y) mutation may be a potential pathogenic site.
CONCLUSIONS: Through whole exome sequencing, we identified a total of 17 potential pathogenic mutation loci, none of which have been reported thus far. Therefore, our work expanded the gene mutation spectrum of familial hereditary triple negative breast cancer, which can provide more basis for family genetic counseling.

The advent of genome-wide ancient DNA analysis has revolutionized our understanding of prehistoric societies. However, studying biological relatedness in these groups requires tailored approaches due to the challenges of analyzing ancient DNA. READv2, an optimized Python3 implementation of the most widely used tool for this purpose, addresses these challenges while surpassing its predecessor in speed and accuracy. For sufficient amounts of data, it can classify up to third-degree relatedness and differentiate between the two types of first-degree relatedness, full siblings and parent-offspring. READv2 enables user-friendly, efficient, and nuanced analysis of biological relatedness, facilitating a deeper understanding of past social structures.

BACKGROUND: Hereditary spherocytosis (HS, MIM#612641) is one of the most common hereditary hemolytic disorders. This study aimed to confirm a novel variant\'s pathogenicity and reveal a patient\'s genetic etiology.
METHODS: The clinical data of a patient with HS who underwent genetic sequencing at the Children\'s Hospital of Chongqing Medical University were reviewed retrospectively. In silico prediction and in vitro minigene splicing reporter system were then conducted on the detected variant to analyze its intramolecular impact. A summary of the literature related to HS due to SPTB gene variants was also presented.
RESULTS: A novel variant (c.301-2 A > G) in the SPTB gene (NM_001024858.4) was identified in the proband. Using Sanger sequencing, we conclusively confirmed that the inheritance of the variant could not be traced to the biological parents. The in vitro minigene assay revealed three different transcripts derived from the c.301-2 A > G variant: r.301_474del, r.301_306delCCAAAG, and r.301-1_301-57ins. Through a literature review, patients with HS who had been genotypically validated were summarized and the SPTB gene variant profile was mapped.
CONCLUSIONS: We identified a splicing variant of the SPTB gene, thus confirming its aberrant translation. The novel variant was the probable genetic etiology of the proband with HS. Our findings expanded the variant spectrum of the SPTB gene, thus improving the understanding of the associated hereditary hemolytic disorders from a clinical and molecular perspective and contributing to the foundation of genetic counseling and diagnosis.

A 28-year-old woman was found to have coagulation factor Ⅷ activity (FⅧ∶C) <1% and von Willebrand factor antigen (VWF∶Ag) <1% during routine prenatal examinations. No pathogenic variation was found in the exon region of the VWF gene using next-generation sequencing. The clinical presentation of this patient does not match the clinical characteristics of type Ⅲ hemophilia [von Willebrand disease (VWD) ]; therefore, third-generation sequencing technology was used to perform whole-genome sequencing on the patient and her family members. Multiple members of the patient\'s paternal family carried a heterozygous variant of VPS33B, c.869G>C. The family members carrying this variant all had varying degrees of reduced VWF levels (39% -56% ). Moreover, the proband was detected with the heterozygous variant c.1474dupA in GP1BA. The ACMG and Clinvar databases determined that this variation was associated with platelet-type pseudo VWD. The decrease in VWF levels caused by heterozygous variations in VPS33B in families is the first international report, and no previous studies have reported cases of severe decrease in plasma VWF levels caused by double heterozygous variations in VPS33B and GP1BA.
一例28岁女性，孕期常规体检发现凝血因子Ⅷ活性（FⅧ∶C）<1%、血管性血友病因子抗原（VWF∶Ag）<1%。二代测序未发现其VWF基因外显子区域存在致病变异。由于该患者临床表现与Ⅲ型血管性血友病（VWD）临床特征不符，因此采用三代测序技术对该患者及其家系成员进行全基因组测序，发现该患者父系家族中有多位成员中携带VPS33B基因杂合变异c.869G>C，携带该变异的家系成员均有不同程度的VWF水平降低（39%～56%）。同时，先证者还检出GP1BA基因杂合变异c.1474dupA，ACMG及Clinvar数据库判断该变异与\"血小板型假性VWD\"相关。VPS33B基因杂合变异导致的VWF水平降低家系为国际首次报道，VPS33B基因与GP1BA基因双重杂合变异引起血浆VWF水平严重降低病例之前也未见报道。.

OBJECTIVE:  Congenital sucrase-isomaltase deficiency is an autosomal recessive inherited disaccharidase deficiency characterized by chronic osmotic diarrhea. In this study, the genotype-phenotype relationships of close relatives of an index case with congenital sucrase-isomaltase deficiency were investigated.
METHODS:  A 23-month-old female patient with a sucrase-isomaltase gene c.317G>A (p.C106Y) homozygous mutation was diagnosed as an index case and her pedigree analysis was performed subsequently. The family members with and without sucrase- isomaltase gene mutations were compared in terms of clinical symptoms.
RESULTS:  The study included 109 cases [mean age ± SD: 22.6 ± 17.2 years (0.1-75 years), 61 males (56%)] of 130 family members of the index case. Sucrase-isomaltase gene c.317G>A (p.C106Y) heterozygous mutation was detected in 27 cases (24.7%); 14 (51.9%) were male and had a mean age of 23.2 ± 18.3 years. The most common complaints of 12 (44.4%) symptomatic patients with mutations were abdominal pain (37%), gas irritability (33.3%), bloating (22.2%), and foul-smelling stools (18.5%). Compared with the cases without mutation, a statistically significant difference was observed in the incidence of gas irritability, foul-smelling stool, ≥2 gastrointestinal symptoms, postprandial complaints, and food allergy (P = .005, P = .047, P = .049, P = .017, P = .021, respectively). Sacrosidase enzyme replacement was applied to 7 patients whose symptoms did not improve with dietary elimination. Clinical response was obtained after enzyme treatment.
CONCLUSIONS:  Despite its autosomal recessive inheritance, congenital sucrase-isomaltase deficiency can also be symptomatic in heterozygous individuals. Further studies are required to clarify the genotype-phenotype relationship and management of the disease.

Sequence variants in Eyes Shut Homolog (EYS) gene are one of the most frequent causes of autosomal recessive retinitis pigmentosa (RP). Herein, we describe an Italian RP family characterized by EYS-related pseudodominant inheritance. The female proband, her brother, and both her sons showed typical RP, with diminished or non-recordable full-field electroretinogram, narrowing of visual field, and variable losses of central vision. To investigate this apparently autosomal dominant pedigree, next generation sequencing (NGS) of a custom panel of RP-related genes was performed, further enhanced by bioinformatic detection of copy-number variations (CNVs). Unexpectedly, all patients had a compound heterozygosity involving two known pathogenic EYS variants i.e., the exon 33 frameshift mutation c.6714delT and the exon 29 deletion c.(5927þ1_5928-1)_(6078þ1_6079-1)del, with the exception of the youngest son who was homozygous for the above-detailed frameshift mutation. No pathologic eye conditions were instead observed in the proband\'s husband, who was a heterozygous healthy carrier of the same c.6714delT variant in exon 33 of EYS gene. These findings provide evidence that pseudodominant pattern of inheritance can hide an autosomal recessive RP partially or totally due to CNVs, recommending CNVs study in those pedigrees which remain genetically unsolved after the completion of NGS or whole exome sequencing analysis.

Homologous recombination plays pivotal roles in physical attachments and genetic diversity. In the past, it was studied among individuals from different populations. However, only few gametes from individual could generate offspring, which limits its exploration in nature selection. In the last few years, preimplantation blastocysts based on trio SNP-chip data were available in individuals for preimplantation genetic testing (PGT). In this protocol, we demonstrate how to detect meiotic recombination events and construct the genetic map based on trio SNP-chip data, obtained from biopsied blastocysts and their related individuals in PGT cycles, which may allow better understanding of recombination events in nature selection.

Branchio-oto-renal (BOR) and branchio-otic (BO) syndromes are characterized by anomalies affecting the ears, often accompanied by hearing loss, as well as abnormalities in the branchial arches and renal system. These syndromes exhibit a broad spectrum of phenotypes and a complex genomic landscape, with significant contributions from the EYA1 gene and the SIX gene family, including SIX1 and SIX5. Due to their diverse phenotypic presentations, which can overlap with other genetic syndromes, molecular genetic confirmation is essential. As sequencing technologies advance, whole-genome sequencing (WGS) is increasingly used in rare disease diagnostics. We explored the genomic landscape of 23 unrelated Korean families with typical or atypical BOR/BO syndrome using a stepwise approach: targeted panel sequencing and exome sequencing (Step 1), multiplex ligation-dependent probe amplification (MLPA) with copy number variation screening (Step 2), and WGS (Step 3). Integrating WGS into our diagnostic pipeline detected structure variations, including cryptic inversion and complex genomic rearrangement, eventually enhancing the diagnostic yield to 91%. Our findings expand the genomic architecture of BOR/BO syndrome and highlight the need for WGS to address the genetic diagnosis of clinically heterogeneous rare diseases.