Newborn Screening for Severe T and B Cell Lymphopenia Using TREC/KREC Detection: A Large-Scale Pilot Study of 202,908 Newborns.

Newborn screening (NBS) for severe inborn errors of immunity (IEI), affecting T lymphocytes, and implementing measurements of T cell receptor excision circles (TREC) has been shown to be effective in early diagnosis and improved prognosis of patients with these genetic disorders. Few studies conducted on smaller groups of newborns report results of NBS that also include measurement of kappa-deleting recombination excision circles (KREC) for IEI affecting B lymphocytes. A pilot NBS study utilizing TREC/KREC detection was conducted on 202,908 infants born in 8 regions of Russia over a 14-month period. One hundred thirty-four newborns (0.66‰) were NBS positive after the first test and subsequent retest, 41% of whom were born preterm. After lymphocyte subsets were assessed via flow cytometry, samples of 18 infants (0.09‰) were sent for whole exome sequencing. Confirmed genetic defects were consistent with autosomal recessive agammaglobulinemia in 1/18, severe combined immunodeficiency - in 7/18, 22q11.2DS syndrome - in 4/18, combined immunodeficiency - in 1/18 and trisomy 21 syndrome - in 1/18. Two patients in whom no genetic defect was found met criteria of (severe) combined immunodeficiency with syndromic features. Three patients appeared to have transient lymphopenia. Our findings demonstrate the value of implementing combined TREC/KREC NBS screening and inform the development of policies and guidelines for its integration into routine newborn screening programs.

Sudden death due to rupture of an aortic syphilitic aneurysm.

A 26-year-old man who suddenly collapsed and died was found at autopsy to have a ruptured aortic aneurysm which had the classic "tree bark" appearance of tertiary syphilis. Tracking of blood into the pericardial sac had resulted in sudden death from cardiac tamponade. Serological results were consistent with syphilis and HIV was excluded. Sudden death in a young HIV-negative man from the effects of syphilis is exceedingly rare nowadays.

Waxy phenotype evolution in the allotetraploid cereal broomcorn millet: mutations at the GBSSI locus in their functional and phylogenetic context.

Waxy mutants, in which endosperm starch contains ~100% amylopectin rather than the wild-type composition of ~70% amylopectin and ~30% amylose, occur in many domesticated cereals. The cultivation of waxy varieties is concentrated in east Asia, where there is a culinary preference for glutinous-textured foods that may have developed from ancient food processing traditions. The waxy phenotype results from mutations in the GBSSI gene, which catalyzes amylose synthesis. Broomcorn or proso millet (Panicum miliaceum L.) is one of the world's oldest cultivated cereals, which spread across Eurasia early in prehistory. Recent phylogeographic analysis has shown strong genetic structuring that likely reflects ancient expansion patterns. Broomcorn millet is highly unusual in being an allotetraploid cereal with fully waxy varieties. Previous work characterized two homeologous GBSSI loci, with multiple alleles at each, but could not determine whether both loci contributed to GBSSI function. We first tested the relative contribution of the two GBSSI loci to amylose synthesis and second tested the association between GBSSI alleles and phylogeographic structure inferred from simple sequence repeats (SSRs). We evaluated the phenotype of all known GBSSI genotypes in broomcorn millet by assaying starch composition and protein function. The results showed that the GBSSI-S locus is the major locus controlling endosperm amylose content, and the GBSSI-L locus has strongly reduced synthesis capacity. We genotyped 178 individuals from landraces from across Eurasia for the 2 GBSSI and 16 SSR loci and analyzed phylogeographic structuring and the geographic and phylogenetic distribution of GBSSI alleles. We found that GBSSI alleles have distinct spatial distributions and strong associations with particular genetic clusters defined by SSRs. The combination of alleles that results in a partially waxy phenotype does not exist in landrace populations. Our data suggest that broomcorn millet is a system in the process of becoming diploidized for the GBSSI locus responsible for grain amylose. Mutant alleles show some exchange between genetic groups, which was favored by selection for the waxy phenotype in particular regions. Partially waxy phenotypes were probably selected against-this unexpected finding shows that better understanding is needed of the human biology of this phenomenon that distinguishes cereal use in eastern and western cultures.

Reticulate evolution in Panicum (Poaceae): the origin of tetraploid broomcorn millet, P. miliaceum.

Panicum miliaceum (broomcorn millet) is a tetraploid cereal, which was among the first domesticated crops, but is now a minor crop despite its high water use efficiency. The ancestors of this species have not been determined; we aimed to identify likely candidates within the genus, where phylogenies are poorly resolved. Nuclear and chloroplast DNA sequences from P. miliaceum and a range of diploid and tetraploid relatives were used to develop phylogenies of the diploid and tetraploid species. Chromosomal in situ hybridization with genomic DNA as a probe was used to characterize the genomes in the tetraploid P. miliaceum and a tetraploid accession of P. repens. In situ hybridization showed that half the chromosomes of P. miliaceum hybridized more strongly with labelled genomic DNA from P. capillare, and half with labelled DNA from P. repens. Genomic DNA probes differentiated two sets of 18 chromosomes in the tetraploid P. repens. Our phylogenetic data support the allotetraploid origin of P. miliaceum, with the maternal ancestor being P. capillare (or a close relative) and the other genome being shared with P. repens. Our P. repens accession was also an allotetraploid with two dissimilar but closely related genomes, the maternal genome being similar to P. sumatrense. Further collection of Panicum species, particularly from the Old World, is required. It is important to identify why the water-efficient P. miliaceum is now of minimal importance in agriculture, and it may be valuable to exploit the diversity in this species and its ancestors.

Intraoperative electroencephalogram patterns as predictors of postoperative delirium in older patients: a systematic review and meta-analysis.

Background: Postoperative delirium (POD) significantly affects patient outcomes after surgery, leading to increased morbidity, extended hospital stays, and potential long-term cognitive decline. This study assessed the predictive value of intraoperative electroencephalography (EEG) patterns for POD in adults. Methods: This systematic review and meta-analysis followed the PRISMA and Cochrane Handbook guidelines. A thorough literature search was conducted using PubMed, Medline, and CENTRAL databases focusing on intraoperative native EEG signal analysis in adult patients. The primary outcome was the relationship between the burst suppression EEG pattern and POD development. Results: From the initial 435 articles identified, 19 studies with a total of 7,229 patients were included in the systematic review, with 10 included in the meta-analysis (3,705 patients). In patients exhibiting burst suppression, the POD incidence was 22.1% vs. 13.4% in those without this EEG pattern (p=0.015). Furthermore, an extended burst suppression duration associated with a higher likelihood of POD occurrence (p = 0.016). Interestingly, the burst suppression ratio showed no significant association with POD. Conclusions: This study revealed a 41% increase in the relative risk of developing POD in cases where a burst suppression pattern was present. These results underscore the clinical relevance of intraoperative EEG monitoring in predicting POD in older patients, suggesting its potential role in preventive strategies. Systematic Review Registration: This study was registered on International Platform for Registered Protocols for Systematic Reviews and Meta-Analyses: INPLASY202420001, https://doi.org/10.37766/inplasy2024.2.0001.

Fibroblasts and polymer composition are essential for bioengineering of airway epithelium on nonwoven scaffolds.

To make tissue engineering a truly effective tool, it is necessary to understand how the patterns of specific tissue development are modulated by and depend on the artificial environment. Even the most advanced approaches still do not fully meet the requirements of practical engineering of tracheobronchial epithelium. This study aimed to test the ability of the synthetic and natural nonwoven scaffolds to support the formation of morphological sound airway epithelium including the basement membrane (BM). We also sought to identify the potential role of fibroblasts in this process. Our results showed that nonwoven scaffolds are generally suitable for producing well-differentiated tracheobronchial epithelium (with cilia and goblet cells), while the structure and functionality of the equivalents appeared to be highly dependent on the composition of the scaffolds. Unlike natural scaffolds, synthetic ones supported the formation of the epithelium only when epithelial cells were cocultured with fibroblasts. Fibroblasts also appeared to be obligatory for basal lamina formation, regardless of the type of the nonwoven material used. However, even in the presence of fibroblasts, the synthetic scaffolds were unable to support the formation of the epithelium and of the BM (in particular, basal lamina) as effectively as the natural scaffolds did.

Decreased TREC and KREC levels in newborns with trisomy 21.

Newborn screening (NBS) for severe combined immunodeficiency (SCID) has been widely implemented to enable early detection and intervention. Trisomy 21, commonly known as Down syndrome (DS), poses unique challenges in NBS due to its frequent association with T and/or B cell lymphopenia. The pilot NBS screening program recently conducted in Russia was aimed to identify both severe T and B cell deficiencies by measuring TREC and KREC. This study aims to evaluate the incidence of DS in newborns who participated in the pilot program, assess their TREC and KREC values, and determine the proportion of DS newborns potentially identifiable through T/B lymphopenia NBS. We conducted a retrospective analysis of the data obtained during the pilot NBS program, involving 202,908 newborns from eight regions of Russia. The study identified 157 patients with trisomy 21 among the screened cohort, resulting in a DS birth prevalence of 1:1,284. Median TREC and KREC values did not significantly differ between full-term and pre-term subgroups of DS patients. TREC values in DS newborns were decreased and comparable to those of the extremely preterm newborns. DS newborns also demonstrated significant differences in KREC values as compared to the general cohort regardless of gestational age. Our data suggests abnormalities of T- and B-cell lineages development and requires further investigation. This article highlights the need for increased awareness of the intrinsic immunological defects associated with DS. The findings underscore the importance of continued follow-up and comprehensive support by healthcare teams for individuals with DS.

Short-term and medium-term clinical outcomes of multisystem inflammatory syndrome in children: a prospective observational cohort study.

BACKGROUND: Even though the incidence of Multisystem Inflammatory Syndrome in children (MIS-C) is decreasing cases are still reported across the world. Studying the consequences of MIS-C enhances our understanding of the disease's prognosis. The objective of this study was to assess short- and medium-term clinical outcomes of MIS-C. METHODS: Prospective observational cohort study at Municipal Children's Hospital Morozovskaya, Moscow, Russia. All children meeting the Royal College of Paediatrics and Child Health (RCPCH), Centers for Disease Control and Prevention (CDC), or the World Health Organization (WHO) MIS-C case definition admitted to the hospital between 17 May and 26 October 2020 were included in the study. All survivors were invited to attend a clinic at 2 and 6 weeks after hospital discharge. RESULTS: 37 children median age 6 years (interquartile range [IQR] 3.3-9.4), 59.5% (22/37) boys were included in the study. 48.6% (18/37) of patients required ICU care. One child died. All children had increased levels of systemic inflammatory markers during the acute event. Echocardiographic investigations identified abnormal findings in 35.1% (13/37) of children. 5.6% (2/36) of children were presenting with any symptoms six weeks after discharge. By six weeks the inflammatory markers were within the reference norms in all children. The echocardiographic evaluation showed persistent coronary dilatation in one child. CONCLUSIONS: Despite the severity of their acute MIS-C, the majority of children in our cohort fully recovered with none having elevated laboratory markers of inflammation at 6 weeks, few (< 10%) reporting persistent symptoms at 6 weeks, and only one with persistent echocardiographic abnormalities.

Epidemiology of Spinal Muscular Atrophy Based on the Results of a Large-Scale Pilot Project on 202,908 Newborns.

BACKGROUND: This study presents the findings of a newborn screening (NBS) pilot project for 5q-spinal muscular atrophy (5q-SMA) in multiple regions across Russia for during the year 2022. The aim was to assess the feasibility and reproducibility of NBS for SMA5q in diverse populations and estimate the real prevalence of 5q-SMA in Russia as well as the distribution of patients with different number of SMN2 copies. METHODS: The pilot project of NBS here was based on data, involving the analysis of 202,908 newborns. SMA screening assay was performed using a commercially available real-time polymerase chain reaction kit, the Eonis SCID-SMA. RESULTS: In one year, 202,908 newborns were screened, identifying 26 infants with homozygous deletion of SMN1 exon 7, yielding an estimated 5q-SMA incidence of 1:7804 newborns. It was found that 38.46% had two SMN2 copies, 42.31% had three copies, 15.38% had four copies, and 3.85% had five copies of SMN2. Immediate treatment was proposed for patients with two or three SMN2 copies. Infants with four or more SMN2 copies warranted further investigation on management and treatment. Short-term monitoring after gene therapy showed motor function improvements. Delays in treatment initiation were observed, including the testing for adeno-associated virus 9 antibodies and nonmedical factors. CONCLUSIONS: The study emphasizes the need for a standardized algorithm for early diagnosis and management through NBS to benefit affected families. Overall, the NBS program for 5q-SMA in Russia demonstrated the potential to improve outcomes and transform SMA from a devastating disease to a chronic condition with evolving medical requirements.

Population genomic structure of Eurasian and African foxtail millet landrace accessions inferred from genotyping-by-sequencing.

Foxtail millet [Setaria italica (L.) P. Beauv.] is the second most important millet species globally and is adapted to cultivation in diverse environments. Like its wild progenitor, green foxtail [S. viridis (L.) P. Beauv.], it is a model species for C4 photosynthetic pathways and stress tolerance genes in related bioenergy crops. We addressed questions regarding the evolution and spread of foxtail millet through a population genomic study of landraces from across its cultivated range in Europe, Asia, and Africa. We sought to determine population genomic structure and the relationship of domesticated lineages relative to green foxtail. Further, we aimed to identify genes involved in environmental stress tolerance that have undergone differential selection between geographical and genetic groups. Foxtail millet landrace accessions (n = 328) and green foxtail accessions (n = 12) were sequenced by genotyping-by-sequencing (GBS). After filtering, 5,677 single nucleotide polymorphisms (SNPs) were retained for the combined foxtail millet-green foxtail dataset and 5,020 for the foxtail millet dataset. We extended geographic coverage of green foxtail by including previously published GBS sequence tags, yielding a 4,515-SNP dataset for phylogenetic reconstruction. All foxtail millet samples were monophyletic relative to green foxtail, suggesting a single origin of foxtail millet, although no group of foxtail millet was clearly the most ancestral. Four genetic clusters were found within foxtail millet, each with a distinctive geographical distribution. These results, together with archaeobotanical evidence, suggest plausible routes of spread of foxtail millet. Selection scans identified nine candidate genes potentially involved in environmental adaptations, particularly to novel climates encountered, as domesticated foxtail millet spread to new altitudes and latitudes.

Resequencing of global Tartary buckwheat accessions reveals multiple domestication events and key loci associated with agronomic traits.

BACKGROUND: Tartary buckwheat (Fagopyrum tataricum) is a nutritionally balanced and flavonoid-rich crop plant that has been in cultivation for 4000 years and is now grown globally. Despite its nutraceutical and agricultural value, the characterization of its genetics and its domestication history is limited. RESULTS: Here, we report a comprehensive database of Tartary buckwheat genomic variation based on whole-genome resequencing of 510 germplasms. Our analysis suggests that two independent domestication events occurred in southwestern and northern China, resulting in diverse characteristics of modern Tartary buckwheat varieties. Genome-wide association studies for important agricultural traits identify several candidate genes, including FtUFGT3 and FtAP2YT1 that significantly correlate with flavonoid accumulation and grain weight, respectively. CONCLUSIONS: We describe the domestication history of Tartary buckwheat and provide a detailed resource of genomic variation to allow for genomic-assisted breeding in the improvement of elite cultivars.

Identification of novel variants in the LDLR gene in Russian patients with familial hypercholesterolemia using targeted sequencing.

Familial hypercholesterolemia (FH) is caused by mutations in various genes, including the LDLR, APOB and PSCK9 genes; however, the spectrum of these mutations in Russian individuals has not been fully investigated. In the present study, mutation screening was performed on the LDLR gene and other FH-associated genes in patients with definite or possible FH, using next-generation sequencing. In total, 59 unrelated patients were recruited and sorted into two separate groups depending on their age: Adult (n=31; median age, 49; age range, 23-70) and children/adolescent (n=28; median age, 11; age range, 2-21). FH-associated variants were identified in 18 adults and 25 children, demonstrating mutation detection rates of 58 and 89% for the adult and children/adolescent groups, respectively. In the adult group, 13 patients had FH-associated mutations in the LDLR gene, including two novel variants [NM_000527.4: c.433_434dupG p.(Val145Glyfs*35) and c.1186G>C p.(Gly396Arg)], 3 patients had APOB mutations and two had ABCG5/G8 mutations. In the children/adolescent group, 21 patients had FH-causing mutations in the LDLR gene, including five novel variants [NM_000527.4: c.325T>G p.(Cys109Gly), c.401G>C p.(Cys134Ser), c.616A>C p.(Ser206Arg), c.1684_1691delTGGCCCAA p.(Pro563Hisfs*14) and c.940+1_c.940+4delGTGA], and 2 patients had APOB mutations, as well as ABCG8 and LIPA mutations, being found in different patients. The present study reported seven novel LDLR variants considered to be pathogenic or likely pathogenic. Among them, four missense variants were located in the coding regions, which corresponded to functional protein domains, and two frameshifts were identified that produced truncated proteins. These variants were observed only once in different patients, whereas a splicing variant in intron 6 (c.940+1_c.940+4delGTGA) was detected in four unrelated individuals. Previously reported variants in the LDLR, APOB, ABCG5/8 and LIPA genes were observed in 33 patients. The LDLR p.(Gly592Glu) variant was detected in 6 patients, representing 10% of the FH cases reported in the present study, thus it may be a major variant present in the Russian population. In conclusion, the present study identified seven novel variants of the LDLR gene and broadens the spectrum of mutations in FH-related genes in the Russian Federation.

Plasma Treatment of Poly(ethylene terephthalate) Films and Chitosan Deposition: DC- vs. AC-Discharge.

Plasma treatment is one of the most promising tools to control surface properties of materials tailored for biomedical application. Among a variety of processing conditions, such as the nature of the working gas and time of treatment, discharge type is rarely studied, because it is mainly fixed by equipment used. This study aimed to investigate the effect of discharge type (direct vs. alternated current) using air as the working gas on plasma treatment of poly(ethylene terephthalate) films, in terms of their surface chemical structure, morphology and properties using X-ray photoelectron spectroscopy, scanning electron microscopy, atomic force microscopy and contact angle measurements. The effect of the observed changes in terms of subsequent chitosan immobilization on plasma-treated films was also evaluated. The ability of native, plasma-treated and chitosan-coated films to support adhesion and growth of mesenchymal stem cells was studied to determine the practicability of this approach for the biomedical application of poly(ethylene terephthalate) films.

The spectrum of pathogenic variants of the ATP7B gene in Wilson disease in the Russian Federation.

BACKGROUND: Wilson's disease (WD) is a rare inherited disorder caused by mutations in the ATP7B gene resulting in copper accumulation in different organs. However, data on ATP7B mutation spectrum in Russia and worldwide are insufficient and contradictory. The objective of the present study was estimation of the frequency of ATP7B gene mutations in the Russian population of WD patients. MATERIALS AND METHODS: 75 WDpatients were examined by next-generation sequencing (NGS). A targeted panel NimbleGen SeqCap EZ Choice: 151012_HG38_CysFib_EZ_HX3 (ROCHE)was designed for analysis of ATP7B gene and possible modifier genes. Retrospective assessment of a diagnostic WD score (Leipzig, 2001) was also performed. RESULTS: 31 mutations in ATP7B gene were detected. Two most frequent mutations were c.3207C > A (51,85% of alleles) and c.3190 G > A (8,64% of alleles). Single rare mutations were detected in 29% of cases. In 96% cases mutations of both copies of the ATP7B were revealed. We also observed 3 novel potentially pathogenic variants which were not previously described (c.1870-8A > G, c.3655A > T (p.Ile1219Phe), c.3036dupC (p.Lys1013fs). For 25% of patients at the time of the manifestation the diagnosis WD could not be established using the earlier proposed diagnostic score. There was a remarkable delay in diagnosis for the majority of patients. Only 33% of patients WD was diagnosed in three months after the first symptoms, 29%patients - in 3-12 months, 30% - in 1-10 years, in 8% - more than 10 years. Generally, clinical appearance of WD may be rather variable at manifestation and genetic profiling at this step is the only way to confirm the presence of WD.

Analysis of the Spectrum of ACE2 Variation Suggests a Possible Influence of Rare and Common Variants on Susceptibility to COVID-19 and Severity of Outcome.

OBJECTIVES: In March 2020, the World Health Organization declared that an infectious respiratory disease caused by a new severe acute respiratory syndrome coronavirus 2 [SARS-CoV-2, causing coronavirus disease 2019 (COVID-19)] became a pandemic. In our study, we have analyzed a large publicly available dataset, the Genome Aggregation Database (gnomAD), as well as a cohort of 37 Russian patients with COVID-19 to assess the influence of different classes of genetic variants in the angiotensin-converting enzyme-2 (ACE2) gene on the susceptibility to COVID-19 and the severity of disease outcome. RESULTS: We demonstrate that the European populations slightly differ in alternative allele frequencies at the 2,754 variant sites in ACE2 identified in the gnomAD database. We find that the Southern European population has a lower frequency of missense variants and slightly higher frequency of regulatory variants. However, we found no statistical support for the significance of these differences. We also show that the Russian population is similar to other European populations when comparing the frequencies of the ACE2 variants. Evaluation of the effect of various classes of ACE2 variants on COVID-19 outcome in a cohort of Russian patients showed that common missense and regulatory variants do not explain the differences in disease severity. At the same time, we find several rare ACE2 variants (including rs146598386, rs73195521, rs755766792, and others) that are likely to affect the outcome of COVID-19. Our results demonstrate that the spectrum of genetic variants in ACE2 may partially explain the differences in severity of the COVID-19 outcome.

Experimental animal model for assessment of tracheal epithelium regeneration.

OBJECTIVES/HYPOTHESIS: To develop an experimental model in rabbits for assessment of tracheal epithelium regeneration through application of either natural or artificial polymer scaffolds. STUDY DESIGN: First, we identified the size of full-thickness mucosal defect, which does not allow self-healing (a "critical defect"), thus representing an adequate experimental model for regenerative therapy of tracheal epithelium damage. Then, two methods of polymer scaffold fixation at the site of the epithelium defect were compared: suturing and fixation with a stent. This was done through: 1) formation of a full-thickness anterolateral mucosal defect by tracheal mucosa excision; and 2) fixation of the scaffold at the site of the tracheal epithelium defect using sutures (through a tracheal wall "window") or a vascular stent (through a small tracheal incision). RESULTS: The dimension of a critical anterolateral mucosal defect of the trachea for rabbits was found to be 1.5 cm in length and more than 50% of the tracheal circumference. Fixation of the scaffold with a stent proved to be more efficient due to a uniform distribution of the pressure over the entire surface of the scaffold, whereas the suturing of the scaffold provided unsatisfactory results. In addition, fixation of the scaffold by suturing required formation of a large "window" in the tracheal wall. Thus, using the stent appeared to be technically less complicated and much less traumatic as compared to suturing. CONCLUSION: We present an experimental in vivo animal model of tracheal epithelium injury and recovery. It can be effectively used with certain further modifications as a basis for routine testing of bioengineered constructs. LEVEL OF EVIDENCE: NA Laryngoscope, 129:E213-E219, 2019.

Non-woven bilayered biodegradable chitosan-gelatin-polylactide scaffold for bioengineering of tracheal epithelium.

OBJECTIVES: The conversion of tissue engineering into a routine clinical tool cannot be achieved without a deep understanding of the interaction between cells and scaffolds during the process of tissue formation in an artificial environment. Here, we have investigated the cultivation conditions and structural features of the biodegradable non-woven material in order to obtain a well-differentiated human airway epithelium. MATERIALS AND METHODS: The bilayered scaffold was fabricated by electrospinning technology. The efficiency of the scaffold has been evaluated using MTT cell proliferation assay, histology, immunofluorescence and electron microscopy. RESULTS: With the use of a copolymer of chitosan-gelatin-poly-l-lactide, a bilayered non-woven scaffold was generated and characterized. The optimal structural parameters of both layers for cell proliferation and differentiation were determined. The basal airway epithelial cells differentiated into ciliary and goblet cells and formed pseudostratified epithelial layer on the surface of the scaffold. In addition, keratinocytes formed a skin equivalent when seeded on the same scaffold. A comparative analysis of growth and differentiation for both types of epithelium was performed. CONCLUSIONS: The structural parameters of nanofibres should be selected experimentally depending on polymer composition. The major challenges on the way to obtain the well-differentiated equivalent of respiratory epithelium on non-woven scaffold include the following: the balance between scaffold permeability and thickness, proper combination of synthetic and natural components, and culture conditions sufficient for co-culturing of airway epithelial cells and fibroblasts. For generation of skin equivalent, the lack of diffusion is not so critical as for pseudostratified airway epithelium.

Reproductive History of a Woman With 8p and 18p Genetic Imbalance and Minor Phenotypic Abnormalities.

We report on the phenotype and the reproductive history of an adult female patient with an unbalanced karyotype: 8p23 and 18p11.3 terminal deletions and 8p22 duplication. The indication for karyotyping of the 28-year-old patient was a structural rearrangement in her miscarriage specimen: 45,ХХ,der(8;18)t(8;18)(p23;p11.3). Unexpectedly, the patient had the same karyotype with only one normal chromosome 8, one normal chromosome 18, and a derivative chromosome, which was a product of chromosomes 8 and 18 fusion with loss of their short arm terminal regions. Fluorescence in situ hybridization revealed that derivative chromosome was a pseudodicentric with an active centromere of chromosome 8. Array comparative genomic hybridization confirmed 8p and 18p terminal deletions and additionally revealed 8p22 duplication with a total of 43 OMIM annotated genes being affected by the rearrangement. The patient had minor facial and cranial dysmorphia and no pronounced physical or mental abnormalities. She was socially normal, had higher education and had been married since the age of 26 years. Considering genetic counseling, the patient had decided to conceive the next pregnancy through in vitro fertilization (IVF) with preimplantation genetic testing for structural chromosomal aberrations (PGT-SR). She underwent four IVF/PGT-SR cycles with a total of 25 oocytes obtained and a total of 10 embryos analyzed. Only one embryo was balanced regarding chromosomes 8 and 18, while the others were unbalanced and demonstrated different combinations of the normal chromosomes 8 and 18 and the derivative chromosome. The balanced embryo was transferred, but the pregnancy was not registered. After four unsuccessful IVF/PGT-SR cycles, the patient conceived naturally. Non-invasive prenatal testing showed additional chromosome 18. The prenatal cytogenetic analysis of chorionic villi revealed an abnormal karyotype: 46,ХХ,der(8;18)t(8;18)(p23;p11.3)mat,+18. The pregnancy was terminated for medical reasons. The patient has a strong intention to conceive a karyotypically normal fetus. However, genetic counseling regarding this issue is highly challenging. Taking into account a very low chance of balanced gametes, emotional stress caused by numerous unsuccessful attempts to conceive a balanced embryo and increasing age of the patient, an IVF cycle with a donor oocyte should probably be considered.

Whole­exome sequencing in Russian children with non­type 1 diabetes mellitus reveals a wide spectrum of genetic variants in MODY­related and unrelated genes.

The present study reports on the frequency and the spectrum of genetic variants causative of monogenic diabetes in Russian children with non­type 1 diabetes mellitus. The present study included 60 unrelated Russian children with non­type 1 diabetes mellitus diagnosed before the age of 18 years. Genetic variants were screened using whole­exome sequencing (WES) in a panel of 35 genes causative of maturity onset diabetes of the young (MODY) and transient or permanent neonatal diabetes. Verification of the WES results was performed using PCR­direct sequencing. A total of 38 genetic variants were identified in 33 out of 60 patients (55%). The majority of patients (27/33, 81.8%) had variants in MODY­related genes: GCK (n=19), HNF1A (n=2), PAX4 (n=1), ABCC8 (n=1), KCNJ11 (n=1), GCK+HNF1A (n=1), GCK+BLK (n=1) and GCK+BLK+WFS1 (n=1). A total of 6 patients (6/33, 18.2%) had variants in MODY­unrelated genes: GATA6 (n=1), WFS1 (n=3), EIF2AK3 (n=1) and SLC19A2 (n=1). A total of 15 out of 38 variants were novel, including GCK, HNF1A, BLK, WFS1, EIF2AK3 and SLC19A2. To summarize, the present study demonstrates a high frequency and a wide spectrum of genetic variants causative of monogenic diabetes in Russian children with non­type 1 diabetes mellitus. The spectrum includes previously known and novel variants in MODY­related and unrelated genes, with multiple variants in a number of patients. The prevalence of GCK variants indicates that diagnostics of monogenic diabetes in Russian children may begin with testing for MODY2. However, the remaining variants are present at low frequencies in 9 different genes, altogether amounting to ~50% of the cases and highlighting the efficiency of using WES in non­GCK­MODY cases.