Genus-Level Analysis of Gut Microbiota in Children with Autism Spectrum Disorder: A Mini Review.

Autism is a global health problem, probably due to a combination of genetic and environmental factors. There is emerging data that the gut microbiome of autistic children differs from the one of typically developing children and it is important to know which bacterial genera may be related to autism. We searched different databases using specific keywords and inclusion criteria and identified the top ten bacterial genera from the selected articles that were significantly different between the studied patients and control subjects studied. A total of 34 studies that met the inclusion criteria were identified. The genera Bacteroides, Bifidobacterium, Clostridium, Coprococcus, Faecalibacterium, Lachnospira, Prevotella, Ruminococcus, Streptococcus, and Blautia exhibited the most substantial data indicating that their fluctuations in the gastrointestinal tract could be linked to the etiology of autism. It is probable that autism symptoms are influenced by both increased levels of harmful bacteria and decreased levels of beneficial bacteria. Interestingly, these genera demonstrated varying patterns of increased or decreased levels across different articles. To validate and eliminate the sources of this fluctuation, further research is needed. Consequently, future investigations on the causes of autism should prioritize the examination of the bacterial genera discussed in this publication.

Initial Effect of Recombinant Human Growth Hormone Treatment in a Patient with Löwe Syndrome.

OBJECTIVES: Löwe syndrome (the oculocerebrorenal syndrome of Löwe, OCRL, OMIM #309000, ORPHA: 534) is a very rare multisystem X-linked disorder characterized by ocular, kidney and nervous system anomalies. CASE PRESENTATION: We present the first Bulgarian genetically confirmed patient with OCRL. The patient had facial dysmorphism, cryptorchidism, congenital cataracts, nystagmus, delayed physical and mental development, and poor nutritional status. He had severe rickets, metabolic acidosis, hypokalaemia, hypophosphataemia, and low IGF-1 levels at the age of three, in addition to his developmental delay. The molecular-genetic analysis reported a pathogenic variant c.1124A>G, p.H375R in the OCRL gene. This variant was inherited from the mother, who was a carrier. Following the diagnosis of OCRL, treatment with potassium citrate, phosphate, and calcitriol was initiated, along with an increase in caloric intake. Following general physical and biochemical improvement, therapy with rhGH started 4 years ago, and current results are presented. CONCLUSIONS: The patient with Löwe syndrome who was presented with a 6-year follow-up demonstrates the complexity of rare disease cases and the value of multidisciplinary care together with growth hormone treatment for better results in these patients.

Congenital myasthenic syndrome due to a genetic mutation.

ABSTRACT: Congenital myasthenic syndrome (CMS) is a group of rare genetic disorders that mimics the symptoms of myasthenia gravis, but it is due to a genetic defect. We present a case of a male CMS patient, and the course of the disease through the years. The patient initially presented with generalized muscle weakness and difficulty swallowing. During the follow-up, he developed difficulty in chewing, bilateral external ophthalmoparesis with an almost full block of eye movements and bulbar syndrome. The case illustrates both the clinical heterogeneity and the progressive worsening of the symptoms of the disease over the years. The optimal treatment for CMS is based on the molecular defect and its localization in the neuromuscular junction. In our case, treatment with pyridostigmine resulted in good long-term control of symptoms. As a result of the patient's good compliance with treatment, he was not admitted to hospital because of respiratory distress. The lack of a unified protocol for the treatment of CMS highlights the need for a more personalized approach when dealing with patients with rare diseases.

Oxford Nanopore Technology and its Application in Liquid Biopsies.

Advanced medical technologies are transforming the future of healthcare, in particular, the screening and detection of molecular-genetic changes in patients suspected of having a neoplasm. They are based on the assumption that neoplasms release small amounts  of  various  neoplasm-specific molecules, such as tumor DNA, called circulating DNA (cirDNA), into the extracellular space and subsequently into the blood. The detection of tumor-specific molecules and specific molecular changes in body fluids in a noninvasive or minimally invasive approach is known as "liquid biopsy." The aim of this review is to summarize the current knowledge of the application of ONT for analyzing circulating DNA in the field of liquid biopsies among cancer patients. Databases were searched using the keywords "nanopore" and "liquid biopsy" and by applying strict inclusion criteria. This technique can be used for the detection of neoplastic disease, including metastases, guiding precision therapy, and monitoring its effects. There are many challenges, however, for the successful implementation of this technology into the clinical practice. The first one is the low amount of tumor-specific molecules in the body fluids. Secondly, a tumor molecular signature should be discriminated from benign conditions like clonal hematopoiesis of unknown significance. Oxford Nanopore Technology (ONT) is a third-generation sequencing technology that seems particularly promising to complete these tasks. It offers rapid sequencing thanks to its ability to detect changes in the density of the electric current passing through nanopores. Even though ONT still needs validation technology, it is a promising approach for early diagnosis, therapy guidance, and monitoring of different neoplasms based on analyzing the cirDNA.

Factor V Leiden, Factor II, Protein C, Protein S, and Antithrombin and Ischemic Strokes in Young Adults: A Meta-Analysis.

Ischemic strokes are one of the leading causes of death worldwide. The aim of this meta-analysis is to elaborate on the role of inherited predisposition to thrombophilia in the etiology of ischemic strokes in young adults. The keywords factor V Leiden (FVL), factor II, prothrombin (PT), protein C (PC), protein S (PS), antithrombin (AT), ischemic stroke, and young were used to search different databases. We selected studies with participants who were between 18 and 65 years. A total of 104 studies were eligible for inclusion in the meta-analysis. All the studied genetic markers were risk factors for ischemic stroke according to our results (FVL OR = 1.74; PT OR = 1.95; PC OR = 10.20; PS OR = 1.74; AT OR = 3.47; p < 0.05). There was moderate heterogeneity for most of the results, and subgroup analyses were conducted by dividing the studies according to the geographic location, gender ratio, and selection criteria of the performed study. There were no significant differences between the groups, but different geographic location was a probable source of heterogeneity. All of the studied markers-FVL, prothrombin, PC, PS, and AT-were significantly associated with increased risk of ischemic stroke in young adults and, if tested, could improve the quality of care.

Potential role of dynein-related genes in the etiology of male infertility: A systematic review and a meta-analysis.

BACKGROUND: The dynein-related genes may have a role in the etiology of male infertility, particularly in cases of impaired sperm motility. OBJECTIVES: The goal of this review is to compile a list of the most important dynein-related candidate genes that may contribute to male factor infertility. MATERIALS AND METHODS: Databases were searched using the keywords "dynein," "male," "infertility," and by applying strict inclusion criteria. A meta-analysis was also performed by using the eligible case-control studies. The odd ratios (ORs), the Z-test score, and the level of significance were determined using a fixed model with a p value of 0.05. Funnel plots were used to check for publication bias. RESULTS: There were 35 studies that met the inclusion criteria. There were a total of 15 genes responsible for the production of dynein structural proteins, the production of dynein assembling factors, and potentially associated with male infertility. A total of five case-control studies were eligible for inclusion in the meta-analysis. Variants in the dynein-related genes were linked to an increased the risk of male infertility (OR = 21.52, 95% confidence interval 8.34-55.50, Z test = 6.35, p < 0.05). The percentage of heterogeneity, I2 , was 47.00%. The lack of variants in the dynein genes was an advantage, and this was statistically significant. DISCUSSION: The results from the present review illustrate that pathogenic variants in genes both for dynein synthesis and for dynein assembly factors could be associated with isolated cases of male infertility without any other symptoms. CONCLUSIONS: The genes addressed in this study, which are involved in both the production and assembly of dynein, could be used as molecular targets for future research into the etiology of sperm motility problems.

Molecular screening for fragile X syndrome in children with unexplained intellectual disability and/or autistic behaviour.

INTRODUCTION: Fragile X syndrome (FXS, OMIM #300624) is the most common inherited form of intellectual disability and the leading monogenic cause of autism.

Variants of uncertain significance in the era of next-generation sequencing.

ABSTRACT: Next-generation sequencing (NGS) is now widely used in diagnosing rare diseases. However, it has some limitations, such as variants of uncertain significance (VUS). This can present difficulties even for nurse practitioners involved in clinical genetics. We present three cases from our clinical practice: two targeted panel testing and one exome sequencing. Whole blood samples were collected and sent for NGS analysis. In case 1, a VUS was found in the LITAF gene, which is associated with autosomal dominant Charcot-Marie-Tooth disease type 1C. In case 2, a VUS was reported in the MEFV gene, which is associated with autosomal recessive and autosomal dominant familial Mediterranean fever. In these cases, the reported VUS corresponded to the clinical diagnosis. In case 3, two variants in the heterozygous state were found in the ATP7B gene, which is associated with Wilson disease, and the disorder was later clinically recognized. According to the published guidelines, VUSs should not be discussed as a cause for an observed genetic condition. Nevertheless, if the reported variant is in a gene associated with the clinically diagnosed disorder, and there is a strong genotype-phenotype correlation, it could be suggestive of the etiological role of this variant.

Single Nucleotide Polymorphisms in microRNA Genes and Colorectal Cancer Risk and Prognosis.

There is growing interest in single nucleotide polymorphisms (SNPs) in the genes of microRNAs (miRNAs), which could be associated with susceptibility to colorectal cancer (CRC) and therefore for prognosis of the disease and/or treatment response. Moreover, these miRNAs-SNPs could serve as new, low-invasive biomarkers for early detection of CRC. In the present article, we performed a thorough review of different SNPs, which were investigated for a correlation with the CRC risk, prognosis, and treatment response. We also analyzed the results from different meta-analyses and the possible reasons for reported contradictory findings, especially when different research groups investigated the same SNP in a gene for a particular miRNA. This illustrates the need for more case-control studies involving participants with different ethnic backgrounds. According to our review, three miRNAs-SNPs-miR-146a rs2910164, miR-27a rs895819 and miR-608 rs4919510-appear as promising prognostic, diagnostic and predictive biomarkers for CRC, respectively.

CFTR gene variants as a reason for impaired spermatogenesis: a pilot study and a Meta-analysis of published data.

There is increasing data that IVS8-5T variand and TG repeats could lead to impaired spermatogenesis. To investigate this we performed Sanger sequencing on 50 Bulgarian men with a sperm count below 5 × 106/mL and 20 normal fertile men. Frequencies of the results were compared among the two groups. A meta-analysis was perfomed by using the data for 6,423 patients and 5,834 control subjects, tested for the IVS8-5T polymorphism. One case subject (2.0%) was homozygote for the 5 T/5T variant whereas two (4.0%) were heterozygotes for the 5 T/7T variant. No 5 T alleles were found in the control group. The genotypes of the two groups showed a statistically significant difference (p = 0.04, α < 0.05). Also, the odds ratio was 3.73, but this was unsignificant (p = 0.38). All control subjects had 11 TG repeats and for the test group: 47 (94.0%) men with 11 TG repeats and three (6.00%) with 10 TG repeats. Fisher's test showed no significant difference (p = 0.55). The meta-analysis showed that IVS8-5T variant was a risk factor for impaired spermatogenesis (OR = 2.84, p < 0.05) and this was more prominent for non-European (OR = 4.50, p < 0.05) compared to European (OR = 1.28, p < 0.05) men. The IVS8 - 5 T variant could be associated with disorders of sperm production.

Oncogenic Functions and Clinical Significance of Circular RNAs in Colorectal Cancer.

Colorectal cancer (CRC) is ranked as the second most commonly diagnosed disease in females and the third in males worldwide. Therefore, the finding of new more reliable biomarkers for early diagnosis, for prediction of metastasis, and resistance to conventional therapies is an important challenge in overcoming the disease. The current review presents circular RNAs (circRNAs) with their unique features as potential prognostic and diagnostic biomarkers in CRC. The review highlights the mechanism of action and the role of circRNAs with oncogenic functions in the CRC as well as the association between their expression and clinicopathological characteristics of CRC patients. The comprehension of the role of oncogenic circRNAs in CRC pathogenesis is growing rapidly and the next step is using them as suitable new drug targets in the personalized treatment of CRC patients.

16p11.2 Duplication Syndrome - a Case Report.

16p11.2 duplication syndrome is a rare disorder, often associated with intellectual disability, attention deficit, hyperactivity disorder, and a predisposition to epilepsy and schizophrenia. There are no specific dysmorphic features for this genetic condition, but micro-cephaly, micrognathia and hypertelorism could be present. We report a case of 16p11.2 duplication syndrome which has the typical clinical presentation - slight facial dysmorphism, impaired intellectual development, and autistic behavior. Whole-exome sequencing was performed, but no pathogenic or likely pathogenic mutations were identified. Array comparative genomic hybridization analysis established the diagnosis of 16p11.2 duplication syndrome, which illustrates the importance of this method when diagnosing children with unexplained intellectual disability.

Association between polymorphic markers human leucocyte antigen-G and tumour necrosis factor alpha and susceptibility to recurrent miscarriages among Bulgarian women.

OBJECTIVE: To analyze the role of 14 base pair (bp) insertion (ins)/deletion (del) and tumour necrosis factor alpha (TNF-α) G/A polymorphisms as risk factors for spontaneous miscarriage in patients with two or more unsuccessful pregnancies and a group of control women with at least two normal live births. MATERIALS AND METHODS: To investigate the role of these mutations, 50 patients with two or more idiopathic recurrent miscarriages and 50 normal fertile women were tested for the presence of human leucocyte antigen-G (HLA-G) 14 bp ins/del and TNF-α -308 G/A variants. The frequencies of the studied polymorphisms were compared between the two groups. RESULTS: Individuals with a history of miscarriages had a significantly higher prevalence of 14 bp insertion alleles compared with control patients (p=0.04). There was also a two times higher relative risk for miscarriage among carriers of this variant. No statistical difference in allele frequencies of the TNF-α -308 G/A polymorphism was established between controls and study patients (p=0.78). CONCLUSION: The 14 bp ins HLA-G variant could be associated with a higher risk for unsuccessful pregnancy according to the results from the study. There is no association between the studied TNF-α -308 GA polymorphism and rate of spontaneous abortions.

Comparison between thrombophilic gene polymorphisms among high risk patients.

INTRODUCTION: The purpose of this study was to compare the role of the thrombophilic variants among two groups of high risk patients with vascular disorders and recurrent pregnancy loss. METHODS: 200 patients, including 76 with thrombotic accidents and 124 with two or more idiopathic recurrent miscarriage during the first trimester, were tested for the presence of Factor V (F V) Leiden G1691A, Factor II (F II) G20210A, plasminogen activator inhibitor (PAI) 4G/5G, and methylenetetrahydrofolate reductase (MTHFR) C677T polymorphisms using Real time polymerase chain reaction (RT - PCR) in the Laboratory of Medical Genetics, Varna, Bulgaria between June 2016 and May 2019. Frequencies of thrombophilic gene polymorphisms were compared among the two populations and to the expected genotype frequencies. RESULTS: Individuals with a history of vascular disorders had a significantly higher frequency of F V Leiden variant compared to women with recurrent miscariage. There was no statistical difference between the analyzed patients for the other three thrombophilic polymorphisms. The allelic frequencies and the expected genotype frequencies of the F V, F II and MTHFR polymorphisms were calculated according to Hardy-Weinberg equilibrium. The percentages of the homozygotes for F V and F II were higher than expected in the two groups of patients. For the MTHFR there was no difference. CONCLUSION: F V Leiden remains the strongest risk factor for vascular disorders and recurrent pregnancy loss. Screening for this variant should be recommended to patients with thrombotic accidents and women with repeated miscarriage. The role of F II, PAI and MTHFR remains controversial.