Host genetic variants associated with COVID-19 reconsidered in a Slovak cohort.

We present the results of an association study involving hospitalized coronavirus disease 2019 (COVID-19) patients with a clinical background during the 3rd pandemic wave of COVID-19 in Slovakia. Seventeen single nucleotide variants (SNVs) in the eleven most relevant genes, according to the COVID-19 Host Genetics Initiative, were investigated. Our study confirms the validity of the influence of LZTFL1 and 2'-5'-oligoadenylate synthetase (OAS)1/OAS3 genetic variants on the severity of COVID-19. For two LZTFL1 SNVs in complete linkage disequilibrium, rs17713054 and rs73064425, the odds ratios of baseline allelic associations and logistic regressions (LR) adjusted for age and sex ranged in the four tested designs from 2.04 to 2.41 and from 2.05 to 3.98, respectively. The OAS1/OAS3 haplotype 'gttg' carrying a functional allele G of splice-acceptor variant rs10774671 manifested its protective function in the Delta pandemic wave. Significant baseline allelic associations of two DPP9 variants in all tested designs and two IFNAR2 variants in the Omicron pandemic wave were not confirmed by adjusted LR. Nevertheless, adjusted LR showed significant associations of NOTCH4 rs3131294 and TYK2 rs2304256 variants with severity of COVID-19. Hospitalized patients' reported comorbidities were not correlated with genetic variants, except for obesity, smoking (IFNAR2), and hypertension (NOTCH4). The results of our study suggest that host genetic variations have an impact on the severity and duration of acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Considering the differences in allelic associations between pandemic waves, they support the hypothesis that every new SARS-CoV-2 variant may modify the host immune response by reconfiguring involved pathways.

Identification of a novel RPGR mutation associated with retinitis pigmentosa and primary ciliary dyskinesia in a Slovak family: a case report.

Background: The mutations in the RPGR (retinitis pigmentosa GTPase regulator) gene are the most common cause of X-linked retinitis pigmentosa (XLRP), a rare genetic disorder affecting the photoreceptor cells in the retina. Several reported cases identified this gene as a genetic link between retinitis pigmentosa (RP) and primary ciliary dyskinesia (PCD), characterised by impaired ciliary function predominantly in the respiratory tract. Since different mutations in the same gene can result in various clinical manifestations, it is important to describe a correlation between the gene variant and the observed phenotype. Methods: Two young brothers from a non-consanguineous Slovak family with diagnosed retinal dystrophy and recurrent respiratory infections were examined. Suspected PCD was diagnosed based on a PICADAR questionnaire, nasal nitric oxide analysis, transmission electron microscopy, high-speed video microscopy analysis, and genetic testing. Results: We identified a novel frameshift RPGR mutation NM_001034853: c.309_310insA, p.Glu104Argfs*12, resulting in a complex X-linked phenotype combining PCD and RP. In our patients, this mutation was associated with normal ultrastructure of respiratory cilia, reduced ciliary epithelium, more aciliary respiratory epithelium, shorter cilia, and uncoordinated beating with a frequency at a lower limit of normal beating, explaining the clinical manifestation of PCD in our patients. Conclusion: The identified novel pathogenic mutation in the RPGR gene expands the spectrum of genetic variants associated with the X-linked PCD phenotype overlapping with RP, highlighting the diversity of mutations contributing to the disorder. The described genotype-phenotype correlation can be useful in clinical practice to recognise a broader spectrum of PCD phenotypes as well as for future research focused on the genetic basis of PCD, gene interactions, the pathways implicated in PCD pathogenesis, and the role of RPGR protein for the proper functioning of cilia in various tissues throughout the body.

Diagnostic value of clot formation parameters determined by rotational thromboelastometry in 63 patients with congenital dysfibrinogenemia.

Rotational thromboelastometry (ROTEM) is a global hemostasis assay. The diagnosis added value of ROTEM in congenital dysfibrinogenemia remains to be established. The aim of this study was to analyze clot formation by ROTEM in a cohort of dysfibrinogenemic patients and to establish correlations with genotype, clinical features, and coagulation parameters. The study included genetically confirmed congenital dysfibrinogenemia cases (n = 63) and healthy controls ( n  = 50). EXTEM, INTEM, FIBTEM tests were used to measure ROTEM parameters, that is, clotting time (CT), clot formation time (CFT), maximal clot firmness (MCF) and amplitude 10 min after CT (A10). The ISTH bleeding assessment tool was used to determine bleeding episodes. CT (INTEM) was statistically significantly shorter in congenital dysfibrinogenemia patients compared to controls while CFT (EXTEM) was prolonged. Patients's MCF in EXTEM, INTEM, and FIBTEM were similar to controls while A10 (FIBTEM) was statistically significantly lower. Fibrinogen activity was positively correlated with fibrinogen antigen, A10 and MCF in all three assays. Bleeding phenotypes were observed in 23 (36.5%) patients. Only CFT in EXTEM and CT in INTEM were statistically different in patients with bleeding phenotype versus controls. Carriers of the FGA mutation p.Arg35His had a CT (EXTEM) slightly prolonged and a reduced A10 (FIBTEM) compared to controls. Some ROTEM parameters were able to distinguish congenital dysfibrinogenemia patients from controls, and patients with a bleeding phenotype. Prolonged CFT in EXTEM were associated with congenital dysfibrinogenemia and bleeding phenotype. Bleeding episodes in most patients were generally mild and prevalence of thrombosis was very low.

Anxiety and depression among women with newly diagnosed vulvar cancer - A nationwide longitudinal study.

INTRODUCTION: Our objective was to investigate the trajectories of anxiety, depression, emotional and social functioning in women with newly diagnosed vulvar cancer from the time of diagnosis to 12 months after treatment. A further aim was to identify risk factors for high levels of anxiety. MATERIAL AND METHODS: PROVE (PROspective Vulvar Cancer Evaluation) is a nationwide longitudinal cohort study investigating quality of life in women with newly diagnosed vulvar cancer by the following validated patient-reported outcome measures at diagnosis, and 3 and 12 months after treatment: The Hospital Anxiety and Depression Scale, the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire C30, and the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire Vulvar module VU34. Mean scores, changes over time and associations were analyzed by generalized estimated equations and log-linear regression models, adjusted for possible confounders. RESULTS: Between 2019 and 2021, 105 (69%) women completed the questionnaires at all three time points. At diagnosis, 42% of the women reported elevated anxiety levels, decreasing significantly to 30% during the first 12 months. Insomnia, persisting vulvar symptoms and high information needs were significantly associated with a high level of anxiety (relative risk [RR] 2.1, 95% CI 1.2-3.7 for insomnia; RR 2.8, 95% CI 1.7-4.6 for vulvar symptoms, RR 2.7, 95% CI 1.5-4.9 for information needs). We found a trend towards a higher level of anxiety in younger women (<65 years: RR 1.5, 95% CI 1.0-2.5). Participants reported a low and stable prevalence of depression (14%) and high social functioning throughout the study period. CONCLUSIONS: Women with newly diagnosed vulvar cancer report a high level of anxiety at diagnosis. Despite a significant improvement, anxiety remains widely prevalent during the first year of follow-up. Targeting insomnia, vulvar symptoms and unmet needs may decrease anxiety during surveillance.

Polysomnographic profile in children diagnosed with celiac disease before starting on a gluten free diet.

STUDY AIMS: The study assessed the presence of sleep abnormalities in children who had recently been diagnosed with celiac disease (CD) and not started a gluten free diet (GFD). The children's polysomnographic profiles were also characterized and further compared with healthy children of the same age. METHODS: This prospective cross-sectional study involved 46 pediatric subjects (aged 1-19 years) who had recently been diagnosed with CD and not started a GFD. The control group consisted of 32 healthy children (aged 2-17 years). All children underwent anthropometric measurement, laboratory testing and standard overnight observation with in-laboratory video-PSG. The study and control group were divided into subgroups according to the subjects' median ages (8.1 years): celiac children aged less than 8.1 years (n = 23) and more than 8.1 years (n = 23), healthy children less aged than 8.1 years (n = 16) and more than 8.1 years (n = 16). RESULTS: No significant differences in the basic demographic and anthropometric parameters between the celiac and control group were observed. Significantly prolonged sleep latency (SOL) was evident in the celiac subjects (21.89 ± 20.77 min. vs. 10.99 ± 7.94 min, p = 0.02), with a probability of prolonged SOL of 4.23-fold greater (OR = 4.23; 95 % CI 1.1-16.22) than the healthy controls, especially in the subgroup of older celiac patients. No significant differences in the sleep period time (SPT), total sleep time (TST), wake during sleep (WASO), sleep efficiency (SE) and sleep stage distribution and cyclization were found. The respiratory rates during sleep indicated a significantly greater incidence of the central apnea-hypopnea index (CAHI) (0.54 ± 0.78 vs. 0.18 ± 0.24, p = 0.03) with a 3.16-fold greater probability of pathological CAHI (OR = 3.16; 95 % CI 1.02-9.77) than the control group. An increased incidence of CSA in the subgroup of younger celiac patients compared to younger healthy controls was especially evident. CONCLUSIONS: The findings of our study suggest a difference in sleep architecture and an increased incidence of CSA in children with untreated CD, but additional research is required to verify the results.

Differential gene expression of immunity and inflammation genes in colorectal cancer using targeted RNA sequencing.

Introduction: Colorectal cancer (CRC) is a heterogeneous disease caused by molecular changes, as driver mutations, gene methylations, etc., and influenced by tumor microenvironment (TME) pervaded with immune cells with both pro- and anti-tumor effects. The studying of interactions between the immune system (IS) and the TME is important for developing effective immunotherapeutic strategies for CRC. In our study, we focused on the analysis of expression profiles of inflammatory and immune-relevant genes to identify aberrant signaling pathways included in carcinogenesis, metastatic potential of tumors, and association of Kirsten rat sarcoma virus (KRAS) gene mutation. Methods: A total of 91 patients were enrolled in the study. Using NGS, differential gene expression analysis of 11 tumor samples and 11 matching non-tumor controls was carried out by applying a targeted RNA panel for inflammation and immunity genes containing 475 target genes. The obtained data were evaluated by the CLC Genomics Workbench and R library. The significantly differentially expressed genes (DEGs) were analyzed in Reactome GSA software, and some selected DEGs were used for real-time PCR validation. Results: After prioritization, the most significant differences in gene expression were shown by the genes TNFRSF4, IRF7, IL6R, NR3CI, EIF2AK2, MIF, CCL5, TNFSF10, CCL20, CXCL11, RIPK2, and BLNK. Validation analyses on 91 samples showed a correlation between RNA-seq data and qPCR for TNFSF10, RIPK2, and BLNK gene expression. The top differently regulated signaling pathways between the studied groups (cancer vs. control, metastatic vs. primary CRC and KRAS positive and negative CRC) belong to immune system, signal transduction, disease, gene expression, DNA repair, and programmed cell death. Conclusion: Analyzed data suggest the changes at more levels of CRC carcinogenesis, including surface receptors of epithelial or immune cells, its signal transduction pathways, programmed cell death modifications, alterations in DNA repair machinery, and cell cycle control leading to uncontrolled proliferation. This study indicates only basic molecular pathways that enabled the formation of metastatic cancer stem cells and may contribute to clarifying the function of the IS in the TME of CRC. A precise identification of signaling pathways responsible for CRC may help in the selection of personalized pharmacological treatment.

Multi-faceted CRISPR/Cas technological innovation aspects in the framework of 3P medicine.

Since 2009, the European Association for Predictive, Preventive and Personalised Medicine (EPMA, Brussels) promotes the paradigm change from reactive approach to predictive, preventive, and personalized medicine (PPPM/3PM) to protect individuals in sub-optimal health conditions from the health-to-disease transition, to increase life-quality of the affected patient cohorts improving, therefore, ethical standards and cost-efficacy of healthcare to great benefits of the society at large. The gene-editing technology utilizing CRISPR/Cas gene-editing approach has demonstrated its enormous value as a powerful tool in a broad spectrum of bio/medical research areas. Further, CRISPR/Cas gene-editing system is considered applicable to primary and secondary healthcare, in order to prevent disease spread and to treat clinically manifested disorders, involving diagnostics of SARS-Cov-2 infection and experimental treatment of COVID-19. Although the principle of the proposed gene editing is simple and elegant, there are a lot of technological challenges and ethical considerations to be solved prior to its broadly scaled clinical implementation. This article highlights technological innovation beyond the state of the art, exemplifies current achievements, discusses unsolved technological and ethical problems, and provides clinically relevant outlook in the framework of 3PM.

Sensitive SARS-CoV-2 detection, air travel Covid-19 testing, variant determination and fast direct PCR detection, using ddPCR and RT-qPCR methods.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) monitoring in air traffic is important in the prevention of the virus spreading from abroad. The gold standard for SARS-CoV-2 detection is RT-qPCR; however, for early and low viral load detection, a much more sensitive method, such as droplet digital PCR (ddPCR), is required. Our first step was to developed both, ddPCR and RT-qPCR methods, for sensitive SARS-CoV-2 detection. Analysis of ten swab/saliva samples of five Covid-19 patients in different stages of disease showed positivity in 6/10 samples with RT-qPCR and 9/10 with ddPCR. We also used our RT-qPCR method for SARS-CoV-2 detection without the need of RNA extraction, obtaining results in 90-120 minutes. We analyzed 116 self-collected saliva samples from passengers and airport staff arriving from abroad. All samples were negative by RT-qPCR, while 1 was positive, using ddPCR. Lastly, we developed ddPCR assays for SARS-CoV-2 variants identification (alpha, beta, gamma, delta/kappa) that are more economically advantageous when compared to NGS. Our findings demonstrated that saliva samples can be stored at ambient temperature, as we did not observe any significant difference between a fresh sample and the same sample after 24 hours (p = 0.23), hence, saliva collection is the optimal route for sampling airplane passengers. Our results also showed that droplet digital PCR is a more suitable method for detecting virus from saliva, compared to RT-qPCR. Keywords: COVID-19; RT-PCR; ddPCR; SARS-CoV-2; nasopharyngeal swab; saliva.

Comparison of the methods for isolation and detection of SARS-CoV-2 RNA in municipal wastewater.

Introduction: Coronavirus SARS-CoV-2 is a causative agent responsible for the current global pandemic situation known as COVID-19. Clinical manifestations of COVID-19 include a wide range of symptoms from mild (i.e., cough, fever, dyspnea) to severe pneumonia-like respiratory symptoms. SARS-CoV-2 has been demonstrated to be detectable in the stool of COVID-19 patients. Waste-based epidemiology (WBE) has been shown as a promising approach for early detection and monitoring of SARS-CoV-2 in the local population performed via collection, isolation, and detection of viral pathogens from environmental sources. Methods: In order to select the optimal protocol for monitoring the COVID-19 epidemiological situation in region Turiec, Slovakia, we (1) compared methods for SARS-CoV-2 separation and isolation, including virus precipitation by polyethylene glycol (PEG), virus purification via ultrafiltration (Vivaspin®) and subsequent isolation by NucleoSpin RNA Virus kit (Macherey-Nagel), and direct isolation from wastewater (Zymo Environ Water RNA Kit); (2) evaluated the impact of water freezing on SARS- CoV-2 separation, isolation, and detection; (3) evaluated the role of wastewater filtration on virus stability; and (4) determined appropriate methods including reverse transcription-droplet digital PCR (RT-ddPCR) and real-time quantitative polymerase chain reaction (RT-qPCR) (targeting the same genes, i.e., RdRp and gene E) for quantitative detection of SARS-CoV-2 in wastewater samples. Results: (1) Usage of Zymo Environ Water RNA Kit provided superior quality of isolated RNA in comparison with both ultracentrifugation and PEG precipitation. (2) Freezing of wastewater samples significantly reduces the RNA yield. (3) Filtering is counterproductive when Zymo Environ Water RNA Kit is used. (4) According to the specificity and sensitivity, the RT-ddPCR outperforms RT-qPCR. Discussion: The results of our study suggest that WBE is a valuable early warning alert and represents a non-invasive approach to monitor viral pathogens, thus protects public health on a regional and national level. In addition, we have shown that the sensitivity of testing the samples with a nearer detection limit can be improved by selecting the appropriate combination of enrichment, isolation, and detection methods.

Analysis of HIF-1α expression and genetic polymorphisms in human clear cell renal cell carcinoma.

Introduction: Clear cell renal cell carcinoma (ccRCC) is mostly diagnosed incidentally and has relatively high recurrence rates. Alterations in VHL/HIF and mTOR pathways are commonly present in ccRCC. The present study attempted to identify potential diagnostic markers at the biochemical and molecular level. Methods: In total, 54 subjects (36 patients with ccRCC and 18 cancer-free controls) were enrolled. ELISA was used to measure the levels of HIF-1α in the tumor and healthy kidney tissue. The association between five selected SNPs (rs779805, rs11549465, rs2057482, rs2295080 and rs701848) located in genes of pathologically relevant pathways (VHL/HIF and mTOR) and the risk of ccRCC in the Slovak cohort was studied using real-time PCR. Results: Significant differences in HIF-1α tissue levels were observed between the tumor and healthy kidney tissue (p < 0.001). In the majority (69%) of cases, the levels of HIF-1α were higher in the kidney than in the tumor. Furthermore, the concentration of HIF-1α in the tumor showed a significant positive correlation with CCL3 and IL-1ß (p (R2) 0.007 (0.47); p (R2) 0.011 (0.38). No relationship between intratumoral levels of HIF-1α and clinical tumor characteristics was observed. Rs11549465, rs2057482 in the HIF1A gene did not correlate with the expression of HIF-1α either in the tumor or in the normal kidney. None of the selected SNPs has influenced the susceptibility to ccRCC. Conclusion: More research is neccesary to elucidate the role of HIF-1α in the pathogenesis of ccRCC and the association between selected SNPs and susceptibility to this cancer.

DNA Polymorphisms in Pregnant Women with Sticky Platelet Syndrome.

Sticky platelet syndrome (SPS) is a thrombophilia caused by the increased aggregability of platelets in response to the addition of low concentrations of epinephrine (EPI) and/or adenosine diphosphate (ADP). Some of the single nucleotide polymorphisms (SNP), alleles and haplotypes of platelet glycoprotein receptors were proved to have a role in the etiology of thrombotic episodes When comparing SPS and the control group, in VEGFA rs3025039, the p value for both CC vs. TT and CT vs. TT analyses was <0.001. Interestingly, no minor TT genotype was present in the SPS group, suggesting the thrombotic pathogenesis of recurrent spontaneous abortions (RSA) in these patients. Moreover, we found a significant difference in the presence of AT containing a risky A allele and TT genotype of ALPP rs13026692 (p = 0.034) in SPS patients when compared with the controls. Additionally, we detected a decreased frequency of the GG (CC) genotype of FOXP3 rs3761548 in patients with SPS and RSA when compared with the control group (p value for the CC (GG) vs. AA (TT) 0.021). This might indicate an evolutionary protective mechanism of the A (T) allele in the SPS group against thrombotic complications in pregnancy. These results can be used for antithrombotic management in such pregnant patients.

Is the Physiological Composition of the Vaginal Microbiome Altered in High-Risk HPV Infection of the Uterine Cervix?

BACKGROUND: Cervical cancer is the fourth most common malignancy and fourth leading cause of cancer death in women worldwide. More than 99.7% of cases are caused by human papillomavirus (HPV), while HPV types 16 and 18 cause over 70% of all cervical cancer cases. In this preliminary study, we aimed to investigate the presence of HPV infection and diversity of bacteria associated with bacterial vaginosis. METHODS: Cervical swabs (n = 21) taken from women aged 21-47 years, in seventeen cases, with different degrees of cervical abnormality, and from four healthy women, were tested for the presence of HPV DNA, as well as the bacterial strains associated with bacterial vaginosis, using the real-time PCR method. RESULTS: HPV16 was the dominant genotype in 53% (9/17) of patients with confirmed precancerous lesions (ASCUS, LSIL, and HSIL). In specimens with confirmed cytological abnormalities and hrHPV infection, we detected a wide diversity of microbes, while the most common species were Gardnerella vaginalis, Atopobium vaginae, Prevotella bivia, Ureaplasma parvum, Ureaplasma urealyticum, Leptotrichia amnionii, Bacteroides ureolyticus, and Sneathia sanguinegens. The presence of pathogens did not differ, depending on the degree of precancerous lesions or HPV type. CONCLUSION: In our work, HPV16 dominated in patients with cervical precancerous lesions. We also suggest an increased bacterial diversity of the vaginal microbiome in patients with cervical lesions, for which the HPV virus is largely responsible.

Interaction of cervical microbiome with epigenome of epithelial cells: Significance of inflammation to primary healthcare.

One pillar of the predictive, preventive, and personalized medicine framework strategies is the female health. The evaluation of women's lifestyle and dietary habits in context with genetic and modifiable risk factors may reflect the prevention of cervical cancer before the occurrence of clinical symptoms and prediction of cervical lesion behavior. The main aim of this review is to analyze publications in the field of precision medicine that allow the use of research knowledge of cervical microbiome, epigenetic modifications, and inflammation in potential application in clinical practice. Personalized approach in evaluating patient's risk of future development of cervical abnormality should consider the biomarkers of the local microenvironment characterized by the microbial composition, epigenetic pattern of cervical epithelium, and presence of chronic inflammation. Novel sequencing techniques enable a more detailed characterization of actual state in cervical epithelium. Better understanding of all changes in multiomics level enables a better assessment of disease prognosis and selects the eligible targeted therapy in personalized medicine. Restoring of healthy vaginal microflora and reversing the outbreak of cervical abnormality can be also achieved by dietary habits as well as uptake of prebiotics, probiotics, synbiotics, microbial transplantation, and others.

Heterogeneity of Genotype-Phenotype in Congenital Hypofibrinogenemia-A Review of Case Reports Associated with Bleeding and Thrombosis.

Congenital fibrinogen disorders are diseases associated with a bleeding tendency; however, there are also reports of thrombotic events. Fibrinogen plays a role in the pathogenesis of thrombosis due to altered plasma concentrations or modifications to fibrinogen's structural properties, which affect clot permeability, resistance to lysis, and its stiffness. Several distinct types of genetic change and pathogenetic mechanism have been described in patients with bleeding and a thrombotic phenotype, including mutations affecting synthesis or processing in three fibrinogen genes. In this paper, we focused on familial hypofibrinogenemia, a rare inherited quantitative fibrinogen disorder characterized by decreased fibrinogen levels with a high phenotypic heterogeneity. To begin, we briefly review the basic information regarding fibrinogen's structure, its function, and the clinical consequences of low fibrinogen levels. Thereafter, we introduce 15 case reports with various gene mutations derived from the fibrinogen mutation database GFHT (French Study Group on Hemostasis and Thrombosis), which are associated with congenital hypofibrinogenemia with both bleeding and thrombosis. Predicting clinical presentations based on genotype data is difficult. Genotype-phenotype correlations would be of help to better understand the pathologic properties of this rare disease and to provide a valuable tool for the identification of patients who are not only at risk of bleeding, but also at risk of a thrombotic event.

Circulating miRNA expression over the course of colorectal cancer treatment.

Colorectal cancer (CRC) is the third-most common cancer type in males and the second-most common cancer type in females, and has the second-highest overall mortality rate worldwide. Approximately 50% of patients in stage I-III develop metastases, mostly localized to the liver. All physiological conditions occurring in the organism are also reflected in the levels of circulating microRNAs (miRNAs/miRs) in patients. miRNAs are a class of small, non-coding, single-stranded RNAs consisting of 18-25 nucleotides, which have important roles in various cellular processes. The aim of the present study was to evaluate a panel of seven circulating miRNAs (miR-106a-5p, miR-210-5p, miR-155-5p, miR-21-5p, miR-103a-3p, miR-191-5p and miR-16-5p) as biomarkers for monitoring patients undergoing adjuvant treatment of CRC. Total RNA was extracted from the plasma of patients with CRC prior to surgery, in the early post-operative period (n=60) and 3 months after surgery (n=14). The levels of the selected circulating miRNAs were measured with the miRCURY LNA miRNA PCR system and fold changes were calculated using the standard ∆∆Cq method. DIANA-miRPath analysis was used to evaluate the role of significantly deregulated miRNAs. The results indicated significant upregulation of miR-155-5p, miR-21-5p and miR-191-5p, and downregulation of miR-16-5p directly after the surgery. In paired follow-up samples, the most significant upregulation was detected for miR-106a-5p and miR-16-5p, and the most significant downregulation was for miR-21-5p. Pathway analysis outlined the role of the differentially expressed miRNAs in cancer development, but the same pathways are also involved in wound healing and regeneration of intestinal epithelium. It may be suggested that these processes should also be considered in studies investigating sensitive and easily detectable circulating biomarkers for recurrence in patients.

Congenital Afibrinogenemia and Hypofibrinogenemia: Laboratory and Genetic Testing in Rare Bleeding Disorders with Life-Threatening Clinical Manifestations and Challenging Management.

Congenital fibrinogen disorders are rare pathologies of the hemostasis, comprising quantitative (afibrinogenemia, hypofibrinogenemia) and qualitative (dysfibrinogenemia and hypodysfibrinogenemia) disorders. The clinical phenotype is highly heterogeneous, being associated with bleeding, thrombosis, or absence of symptoms. Afibrinogenemia and hypofibrinogenemia are the consequence of mutations in the homozygous, heterozygous, or compound heterozygous state in one of three genes encoding the fibrinogen chains, which can affect the synthesis, assembly, intracellular processing, stability, or secretion of fibrinogen. In addition to standard coagulation tests depending on the formation of fibrin, diagnostics also includes global coagulation assays, which are effective in monitoring the management of replacement therapy. Genetic testing is a key point for confirming the clinical diagnosis. The identification of the precise genetic mutations of congenital fibrinogen disorders is of value to permit early testing of other at risk persons and better understand the correlation between clinical phenotype and genotype. Management of patients with afibrinogenemia is particularly challenging since there are no data from evidence-based medicine studies. Fibrinogen concentrate is used to treat bleeding, whereas for the treatment of thrombotic complications, administered low-molecular-weight heparin is most often. This review deals with updated information about afibrinogenemia and hypofibrinogenemia, contributing to the early diagnosis and effective treatment of these disorders.

Sensenbrenner Syndrome Presenting with Severe Anorexia, Failure to Thrive, Chronic Kidney Disease and Angel-Shaped Middle Phalanges in Two Siblings.

Sensenbrenner syndrome is a very rare autosomal recessive disorder caused by variants in genes involved in the functional development of primary cilia. Typical clinical manifestations include craniofacial and skeletal abnormalities, hence the alternative name cranioectodermal dysplasia. Chronic kidney disease due to progressive tubulointerstitial nephritis (nephronophthisis) has been described in these patients. The authors present 2siblings with severe anorexia, failure to thrive, chronic kidney disease, and angel-shaped middle phalanges. Two previously described variants p.(Leu641*) and p.(Asp841Val) were identified in the WDR35 gene which is most commonly affected in this condition. Analysis of all coding exons of the GDF5 gene was normal. This is the first report of Sensenbrenner syndrome presenting with severe anorexia and failure to thrive at early age. Angel-shaped middle phalanges in the absence of the GDF5 variant may represent an overlapping phenotypic manifestation of ciliopathy.

Comparison of SARS-CoV-2 Detection by Rapid Antigen and by Three Commercial RT-qPCR Tests: A Study from Martin University Hospital in Slovakia.

The global pandemic of coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is having a tremendous impact on the global economy, health care systems and the lives of almost all people in the world. The Central European country of Slovakia reached one of the highest daily mortality rates per 100,000 inhabitants in the first 3 months of 2021, despite implementing strong prophylactic measures, lockdowns and repeated nationwide antigen testing. The present study reports a comparison of the performance of the Standard Q COVID-19 antigen test (SD Biosensor) with three commercial RT-qPCR kits (vDetect COVID-19-MultiplexDX, gb SARS-CoV-2 Multiplex-GENERI BIOTECH Ltd. and Genvinset COVID-19 [E]-BDR Diagnostics) in the detection of infected individuals among employees of the Martin University Hospital in Slovakia. Health care providers, such as doctors and nurses, are classified as "critical infrastructure", and there is no doubt about the huge impact that incorrect results could have on patients. Out of 1231 samples, 14 were evaluated as positive for SARS-CoV-2 antigen presence, and all of them were confirmed by RT-qPCR kit 1 and kit 2. As another 26 samples had a signal in the E gene, these 40 samples were re-isolated and subsequently re-analysed using the three kits, which detected the virus in 22, 23 and 12 cases, respectively. The results point to a divergence not only between antigen and RT-qPCR tests, but also within the "gold standard" RT-qPCR testing. Performance analysis of the diagnostic antigen test showed the positive predictive value (PPV) to be 100% and negative predictive value (NPV) to be 98.10%, indicating that 1.90% of individuals with a negative result were, in fact, positive. If these data are extrapolated to the national level, where the mean daily number of antigen tests was 250,000 in April 2021, it points to over 4700 people per day being misinterpreted and posing a risk of virus shedding. While mean Ct values of the samples that were both antigen and RT-qPCR positive were about 20 (kit 1: 20.47 and 20.16 for Sarbeco E and RdRP, kit 2: 19.37 and 19.99 for Sarbeco E and RdRP and kit 3: 17.47 for ORF1b/RdRP), mean Ct values of the samples that were antigen-negative but RT-qPCR-positive were about 30 (kit 1: 30.67 and 30.00 for Sarbeco E and RdRP, kit 2: 29.86 and 31.01 for Sarbeco E and RdRP and kit 3: 27.47 for ORF1b/RdRP). It confirms the advantage of antigen test in detecting the most infectious individuals with a higher viral load. However, the reporting of Ct values is still a matter of ongoing debates and should not be conducted without normalisation to standardised controls of known concentration.

An association between fibrinogen gene polymorphisms and diabetic peripheral neuropathy in young patients with type 1 diabetes.

In complex etiopathogenesis of diabetic peripheral neuropathy (DPN), hemostatic dysfunction and subclinical inflammation play a possible role. Fibrinogen is involved in both the hemostatic and inflammatory pathways, so we hypothesize that fibrinogen gene polymorphisms might be associated with DPN. A total of 127 young patients with type 1 diabetes (T1D) (average age, 18.5 ± 4.65 years; average diabetes duration, 14.5 ± 2.26 years) and 90 healthy controls were enrolled into the study. Basic biochemical and coagulation parameters were measured and gene polymorphisms of fibrinogen alpha (rs6050) and beta (rs1800790) were established. DPN was diagnosed in 38 diabetic patients by neurological examination. AA genotype and A allele of rs1800790 polymorphism of fibrinogen beta were associated with increased risk of DPN (odds ratio [OR] 4.537, 95% confidence interval [95CI] 1.14-19.94, p = 0.019 and OR 1.958, 95CI 1.038-3.675, p = 0.029, respectively). No association was found between DPN and rs6050 gene polymorphisms. Plasma fibrinogen concentration significantly correlated with HbA1c (Spearman's correlation coefficient [r] = 0.54) and HDL cholesterol (r = - 0.67). A allele and AA genotype of rs1800790 seem to be associated with DPN in young patients with T1D. Further studies are appropriate to elucidate the role of fibrinogen gene polymorphisms in the complex etiology of DPN.