Hermansky–Pudlak Syndrome Type 6 Accompanied with Bowel Vascular Malformation: Clinical Case

Background. Hermansky–Pudlak syndrome type 6 is rare hereditary disease caused by pathogenic variants in base sequence, deletions, and insertions in the HPS6 gene encoding the transmembrane protein of the same name. This disease occurs with hemorrhagic syndrome, oculocutaneous albinism, and inflammatory bowel diseases (in some cases). The clinical picture of the disease, including the gastrointestinal tract pathology, has not been studied completely due to the syndrome rarity.Clinical case description. We would like to present the description of clinical case of the patient with Hermansky–Pudlak syndrome type 6 accompanied with bowel vascular malformation. The patient diagnosed with “oculocutaneous albinism” at the age of 4.5 has shown recurrent intestinal bleedings, blood hemoglobin concentration decrease to 45 g/l; platelet count, mean platelet volume and platelet distribution width remained within the reference values. Slight decrease of Quick’s value to 68% (normal range 70–120%) was revealed. The study of platelet morphology has revealed a decrease in the number of dense granules: < 3 in 25% platelets, < 6 — in 64%. Ultrasound investigation has revealed signs of vascular malformation in ascending colon: significant changes of diameter (widening) and shape of intestinal wall vessels. Molecular genetic analysis (NGS) has revealed the nucleotide variant c.1133T>G (p.Leu378Arg) in homozygous state in the HPS6 gene. The same variant in homozygous state was revealed in the younger proband sister who also had vascular changes in the ascending colon wall.Conclusion. Differential diagnosis of Germanic–Pudlak syndrome type 6 should be performed with other types of this syndrome as well as with syndrome and non-syndrome forms of oculocutaneous albinism. Molecular genetic confirmation of the diagnosis is suggested via massive parallel sequencing (NGS) methods (exome sequencing included) due to the rarity of Hermansky–Pudlak syndrome.

Karyotype and COI gene sequence of Chironomus heteropilicornis Wülker, 1996 (Diptera, Chironomidae) from the Gydan Peninsula, Russia

The karyotype features and gene COI sequence of Chironomus heteropilicornis Wülker, 1996 from the Gydan Peninsula are presented for the first time. Nine banding sequences were determined, eight of them hpiA2, hpiB1, hpiC1, hpiC2, hpiD1, hpiE1, hpiF3 and hpiG1 were previously known from European, Georgian (South Caucasus) and Siberian populations. One new banding sequence for Ch. heteropilicornis, hpiB2, was found. The hpiA2 banding sequence was found in all individuals, and this is its second finding after the Georgian population (Karmokov 2019). The hpiF3 banding sequence was found only in the homozygous state. Additional B-chromosomes are absent. The genetic distances (K2P) between Ch. heteropilicornis COI gene sequence from Gydan Peninsula and Norway are 1.1­–1.3%, and Georgia – 1.8%, much lower than the commonly accepted threshold of 3% for species of genus Chironomus Meigen, 1803. The phylogenetic tree for COI gene sequences estimated by Bayesian inference showed geographically determined clusters of Norway and Gydan and a separate lineage of the Georgian population of Ch. heteropilicornis. The analysis of karyotype and COI gene sequences shows that the population of Ch. heteropilicornis from the Gydan Peninsula has an intermediate position within the Ch. pilicornis group between Georgian, Yakutian and Norwegian populations. The position of Ch. pilicornis Fabricius, 1787 from Canada and Greenland on the phylogenetic tree is discussed.

Cystic Fibrosis: Systems Biology Analysis from Homozygous p.Phe508del Variant Patients’ Samples Reveals Perturbations in Tissue-Specific Pathways

Cystic fibrosis (CF) is an autosomal recessive disorder, caused by diverse genetic variants for the CF transmembrane conductance regulator (CFTR) protein. Among these, p.Phe508del is the most prevalent variant. The effects of this variant on the physiology of each tissue remains unknown. This study is aimed at predicting cell signaling pathways present in different tissues of fibrocystic patients, homozygous for p.Phe508del. The study involved analysis of two microarray datasets, E-GEOD-15568 and E-MTAB-360 corresponding to the rectal and bronchial epithelium, respectively, obtained from the ArrayExpress repository. Particularly, differentially expressed genes (DEGs) were predicted, protein-protein interaction (PPI) networks were designed, and centrality and functional interaction networks were analyzed. The study reported that p.Phe508del-mutated CFTR-allele in homozygous state influenced the whole gene expression in each tissue differently. Interestingly, gene ontology (GO) term enrichment analysis revealed that only “neutrophil activation” was shared between both tissues; however, nonshared DEGs were grouped into the same GO term. For further verification, functional interaction networks were generated, wherein no shared nodes were reported between these tissues. These results suggested that the p.Phe508del-mutated CFTR-allele in homozygous state promoted tissue-specific pathways in fibrocystic patients. The generated data might further assist in prediction diagnosis to define biomarkers or devising therapeutic strategies.

Bi-Allelic c.1746G>T; p.Leu582= Variants in TUBGCP4 in a Boy with Autism: Clinical Data and Literature Review

Bi-allelic mutations in the <i>TUBGCP4</i> gene have been recently associated with autosomal recessive microcephaly with chorioretinopathy. However, little is known about the genotype-phenotype characteristics of this disorder. Here, we describe a 5-year-old male patient with autism and a normal occipitofrontal circumference. No retinal abnormalities were observed. Brain MRI revealed the presence of enlarged sheaths of both tortuous optic nerves; both eyes had shorter axial lengths. Whole-exome sequencing in trio revealed synonymous <i>TUBGCP4</i> variants in homozygous state: c.1746G&#x3e;T; p.Leu582=. This synonymous variant has been previously described and probably leads to skipping of exon 16 of <i>TUBGCP4</i>. These results broaden the clinical spectrum of this new syndrome and suggest that <i>TUBGCP4</i> bi-allelic mutations may underlie complex neurodevelopmental disorders.

RELATIONSHIP BETWEEN RFC GENE VARIANTS (RS1051266) AND CLINICAL FEATURES OF SEVERE INTRAVENTRICULAR HEMORRHAGE IN PRETERM INFANTS

The physiological function of the RFC gene is to ensure the processes of intracellular folic acid transport that are extremely important not only for the processes of reproduction, but also for the embryonic develop-ment of a growing foetus. The aim of this study is to investigate the association between the RFC (G80A, rs1051266) gene variants and the features of the neonatal course in premature infants with severe intraven-tricular haemorrhage. Materials and methods. The study included 24 preterm infants with severe intraven-tricular haemorrhages that received standard clinical, laboratory and instrumental examination. Determina-tion of RFC gene variants was performed using the polymerase chain reaction- restriction fragment length polymorphism method. Results. The following frequencies of genotypes were determined according to the G80A variant of the RFC gene: GG – 8 (33.3%), GA – 9 (37.5%) and AA – 7 (29.2%) in the study group. Children with the A allele of the RFC gene had an increased risk of developing hypertension and a higher mean concentration of ionized calcium. Preterm infants with the A allele were more likely to require oxygen therapy with maximum oxygen concentration and in the presence of this allele in a homozygous state (AA genotype) required nCPAP and non-invasive mechanical ventilation. Conclusion. The obtained results sup-port a hypothesis about the influence on variants of RFC genes on severe neonatal period in preterm infants with intraventricular haemorrhages. However, further multifaceted research in this area is required to give more ground in supporting hypothesis.

Optimization of Robust Diagnostic Strategy for Patients with Fanconi Anaemia (FA)- an Indian Perspective

Objective: Fanconi anaemia is a genotypically heterogeneous disease. A fast and reliable genetic diagnosis method helps is important for the clinical care of these patients. The objective of this study was to establish a strategy for expeditious molecular diagnosis for the Indian FA patients. Methods: Exome sequencing was performed for 119 FA patients on Illumina HiSeq X system and the data was analysed by Sentieon (v201808.01) to identify germline variants. Single nucleotide variants (SNVs) with an allele frequency of &lt;3% in various population databases including the Indian population database were analysed and annotated using an in-house pipeline. Copy number variants (CNVs) were detected using ExomeDepth (v1.1.10). Long amplicon next-generation sequencing was performed for FANCA and FANCG genes. Gene dosage analysis was performed for the deletions identified by WES in the FANCT/UBE2T gene. Sanger sequencing was performed for the selected variants identified by WES. Results: SNVs associated with FA were identified in heterozygous states in 93 (78%) patients. Of the remaining 20 patients, 8 had homozygous deletions, and twelve were compound heterozygous with deletions and SNVs. We detected deletions in 20 patients: 7 with heterozygous and 12 with homozygous deletions in FANCA, and one with a homozygous deletion in FANCT (UBE2T). MLPA confirmed the deletions in the FANCA gene. The homozygous deletion in the FANCT gene was confirmed by the lack of amplification by PCR using the primers binding to the deleted region. By combining the SNVs and deletions, disease-causing genotypes were identified in 113 of 119 (95%) patients. A large number of our patients (81.5 %) were homozygous (Figure) due to the high rate of consanguinity in the population. FANCA was found to be the most frequently mutated gene in (57.5%) while mutations in FANCG gene accounted for 14.2% of our patients. FANCC mutations were found at a very low frequency (1.8%) in our patients. Although mutations in FANCL are rare in all the populations, 24 (21.2%) patients with FANCL mutations were identified in our study. A previously reported synonymous splicing mutation FANCL c.1092G&gt;A;p.K364= was found in homozygous state in 22 (19.5%) patients. Two other FANCL mutations identified includes a missense mutation c.827C&gt;T; p.Pro281Leu in the homozygous state in one patient and a nonsense mutation c.997C&gt;T; p.Gln333Ter in the compound heterozygous state with the common FANCL c.1092G&gt;A;p.K364= mutation in another patient. Mutations in rarely mutated FA genes included 1 patient each in FANCT/UBE2T and FANCI and two each in FANCJ/BRIP1 and FANCF gene. Based on the comprehensive genotype analysis, we could design an algorithm for FA in the Indian population. ~20% of the FA patients who have FANCL c.1092G&gt;A;p.K364= mutation can be diagnosed by Sanger sequencing. MLPA can detect FANCA deletions (16.5% of the overall mutations). The results from these two tests can be obtained in 48 hours. For those who are negative for the mutations by these two methods, LA-NGS can detect SNVs in the FANCA and FANCG genes, which constitute &gt;55% of the FA mutations in the population. We have tested this algorithm for expedited molecular diagnosis in 27 FA patients and found that the disease genotypes could be established in 94% of the patients in less than two weeks. Conclusion/Clinical applicability: Exome sequencing identified several novel mutations in the FA pathway genes in the FA patients. A cost-effective and time-saving algorithm was explicitly established for molecular diagnosis of FA in the Indian population. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Whole exome sequencing in 17 consanguineous Iranian pedigrees expands the mutational spectrum of inherited retinal dystrophies

Inherited retinal dystrophies (IRDs) constitute one of the most heterogeneous groups of Mendelian human disorders. Using autozygome-guided next-generation sequencing methods in 17 consanguineous pedigrees of Iranian descent with isolated or syndromic IRD, we identified 17 distinct genomic variants in 11 previously-reported disease genes. Consistent with a recessive inheritance pattern, as suggested by pedigrees, variants discovered in our study were exclusively bi-allelic and mostly in a homozygous state (in 15 families out of 17, or 88%). Out of the 17 variants identified, 5 (29%) were never reported before. Interestingly, two mutations (GUCY2D:c.564dup, p.Ala189ArgfsTer130 and TULP1:c.1199G > A, p.Arg400Gln) were also identified in four separate pedigrees (two pedigrees each). In addition to expanding the mutational spectrum of IRDs, our findings confirm that the traditional practice of endogamy in the Iranian population is a prime cause for the appearance of IRDs.

Clinical, Biochemical, and Genetic Heterogeneity in Glutaric Aciduria Type II Patients

The variants of electron transfer flavoprotein (ETFA, ETFB) and ETF dehydrogenase (ETFDH) are the leading cause of glutaric aciduria type II (GA-II). In this study, we identified 13 patients harboring six variants of two genes associated with GA-II. Out of the six variants, four were missense, and two were frameshift mutations. A missense variant (ETFDH:p.Gln269His) was observed in a homozygous state in nine patients. Among nine patients, three had experienced metabolic crises with recurrent vomiting, abdominal pain, and nausea. In one patient with persistent metabolic acidosis, hypoglycemia, and a high anion gap, the ETFDH:p.Gly472Arg, and ETFB:p.Pro94Thrfs*8 variants were identified in a homozygous, and heterozygous state, respectively. A missense variant ETFDH: p.Ser442Leu was detected in a homozygous state in one patient with metabolic acidosis, hypoglycemia, hyperammonemia and liver dysfunction. The ETFDH:p.Arg41Leu, and ETFB:p.Ile346Phefs*19 variants were observed in a homozygous state in one patient each. Both these variants have not been reported so far. In silico approaches were used to evaluate the pathogenicity and structural changes linked with these six variants. Overall, the results indicate the importance of a newborn screening program and genetic investigations for patients with GA-II. Moreover, careful interpretation and correlation of variants of uncertain significance with clinical and biochemical findings are needed to confirm the pathogenicity of such variants.

Familial Hypomagnesemia with Hypercalciuria, Nephrocalcinosis, and Bilateral Chorioretinal Atrophy in a Patient with Homozygous p.G75S Variant in CLDN19

Introduction Familial hypomagnesemia with hypercalciuria and nephrocalcinosis (FHHNC) is a rare disorder caused by perturbation in renal reabsorption of magnesium and calcium. Biallelic pathogenic variants either in gene CLDN16 or CLDN19 are responsible for molecular defects. Most patients with CLDN19 variants have been associated with ocular involvements (FHHNCOI). Patient and Methods We had a pediatric patient with hypercalciuric hypomagnesemia and bilateral chorioretinal atrophy. Metabolic profiling and radiology examinations were performed, in addition to whole exome sequencing (WES) used for detection of the causative variant. Results Analysis of WES revealed a homozygous c.223G > A (p.G75S) variant in CLDN19. MutationTaster and Combined Annotation-Dependent Depletion support its deleterious effect and SHERLOC's criteria put it in pathogenic category. This variant is previously reported in compound heterozygous state with other known pathogenic variant. As far as we know, it is the first report of this variant in homozygous state. Conclusion The variant found in our patient is pathogenic and compatible with FHHNCOI characteristics. WES is an advantageous tool in molecular diagnosis and finding genetic pathology of this disease. In line with other reports, ocular abnormalities are variable in patients with CLDN19 mutations, and chronic kidney disease and retinal damages must be considered in this group.

Nos défauts génétiques cachés

A systematic study analysing the exomes of several thousand individuals indicates that each of them carries at least one strongly deleterious mutation that is innocuous in a heterozygote but results in a severe phenotype in the homozygous state. Most of these mutations are very rare, while a few are present in 1 or 2% of the population. The frequency of at-risk couples is approximately 1.5%, but increases dramatically to 25% if the partners of the couple are first cousins. This work has important implications for carrier screening and population genetics in general.