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Background. According to WHO guidelines, one of the criteria for diagnosis of polycythemia vera is the presence of somatic mutations in exon 12 of the JAK2 gene, but to date there is no universally accepted simple method to analyze these mutations. We have previously proposed two methods for screening such mutations based on heteroduplex and HRM (High Resolution Melt) assays, which are relatively cheap and fast compared to sequencing.Aim. To analyze the sensitivity of these screening methods.Materials and methods. The study used cloned DNA samples from 6 patients with various mutations in exon 12 of the JAK2 gene that we had previously identified, as well as a clone of the corresponding wild-type DNA segment. Dilution of the cloned mutant samples with wild-type clones was performed to obtain samples with different levels of allele burden: 100, 50, 25, 12.5, 6.25, 3.13, 1.56 and 0.78 %. Heteroduplex analysis followed by PAGE (polyacrylamide gel) and HRM analysis was then performed with the diluted samples.Results. The sensitivity threshold of the heteroduplex analysis was found to be between 3.13–6.25 % allele burdens depending on the specific mutation, the sensitivity threshold of the HRM assay was 6.25–12.5 % similarly.Conclusion. Our proposed methods of heteroduplex analysis followed by PAGE and HRM-analysis for the detection of polycythemia vera-specific mutations in exon 12 of the JAK2 gene allow increasing the efficiency of using different types of sequencing and can be used as simpler and less expensive methods of preliminary screening of these mutations.

Relevance. The identification of determinants of the human genome, such as single nucleotide polymorphisms (SNPs), in association with various disease patterns, including infectious diseases, is one of the most actively developing areas of scientific research in the world.. Hepatitis C (HC), which remains a serious global health problem, belongs to the number of infections that attract the attention of specialists.Aims. Determination of genetic markers of hepatitis C virus (HCV) natural elimination and assessment of their role as a monitoring parameter of the epidemiological surveillance system.Materials and methods. The study included 660 people divided into 2 groups: persons with chronic HC (CHC) and blood donors (indicator group of the healthy population). In the studied groups, the following SNPs were typed: rs1143634, rs1143627 (IL-1B); rs4251961, rs419598 (IL1RN); rs1800795 (IL6); rs1800896 (IL-10); rs4986790 (TLR4); rs4374383 (MERTK). The associative relationship between SNPs and CHC alleles was identified using logistic regression analysis within four models (codominant, dominant, recessive, and overdominant). Additionally, the significance of polymorphisms at the intragenic and intergenic levels was assessed using modern bioinformatic resources in the field of functional genomics.Results. In this study, genotypes associated with the natural elimination of HCV were identified. Paired combinations of IL 1RA/IL-1B genotypes associated with the probability of the formation of CHC have been established. It is shown that synonymous SNPs can be associated with any characteristics of the pathological process, which can be explained by disequilibrium in coupling with functionally significant alleles of other genetic loci.Conclusion. The detection of the association of SNPs with clinical manifestations of the pathological process is not final and requires further study taking into account ONP coupling groups.

Background. The development of myelofibrosis (MF) is driven by complex molecular genetic events that include driver somatic mutations responsible for the constitutive activation of the JAK/STAT signaling pathway (JAK2, CALR, and MPL), additional mutations affecting epigenetic regulators (TET2, ASXL1, IDH1/2, etc.) and RNA splicing (SRSF2, U2AF1, SF3B1, etc.), as well as genetic aberrations that contribute to genomic instability and disease progression.Aim. To analyze driver (JAK2, CALR, MPL) and prognostic (ASXL1) somatic mutations in patients with MF and evaluate their impact on survival.Materials and methods. The study included 29 patients diagnosed with MF, selected by hematologists from the City Clinical Hospital No. 7 and Regional Clinical Hospital (Krasnoyarsk).Results. 26 (89.6 %) out of 29 examined patients had some driver mutations in JAK2, CALR, MPL genes. The p.V617F mutation in the JAK2 gene was found in 20 (68.9 %) patients. Mutations in the CALR gene were detected in 4 (13.8 %) patients, mutations in the MPL gene were found in 3 patients (10.3 %). In 1 of 26 patients, 2 driver mutations were present simultaneously. 3 (10.3 %) patients were triple negative. Mutations in the ASXL1 gene were detected in 12 (41.4 %) out of 29 examined patients. Conducted targeted NGS (next generation sequencing) for 13 out of 29 patients revealed additional genetic variants that contribute to the understanding of the development mechanism and disease course. When evaluating the overall survival in the groups of patients diagnosed with MF examined by us, depending on the combination of driver (JAK2, CALR, MPL) and prognostic (ASXL1) mutations, no statistically significant differences were found (p = 0.12). This appears to be due to the small sample size. At the same time, assessment of patient survival depending on ASXL1 status showed that in the presence of mutations in the ASXL1 gene, the median survival was 45 months (range 7–120 months), while in the absence of mutations it was 48 months (range 21–359 months) (p = 0.03).Conclusion. The results obtained allow us to assume that the presence of mutations in the ASXL1 gene is an unfavorable factor in the course of the disease.

Introduction. Cervical cancer (CC) is one of the most common cancers in women. The CC etiological agent is the high-risk oncogenic human papillomavirus. In the meantime, not all women infected with this virus can develop cancer, thus suggesting that there is genetic predisposition to CC.The aim of the study was to analyze information about single nucleotide polymorphisms associated with the CC risk.Materials and methods. The performed search was focused on genome-wide association studies (GWAS) and meta-analyses conducted over the last 10 years and addressing the genetic risk of CC in the Caucasian population.Results. The most significant associations with CC were found in the following single nucleotide polymorphisms. Based on the GWAS data, they involve risk alleles rs138446575-T (OR = 2.39) TTC34; rs73728618-T (OR = 1.48) HLA-DQA1; rs3130196-C (OR = 1.4) HLA-DPB1; rs2516448-T (OR = 1.39 and 1.44) MICA and protective alleles rs9271898-A (OR = 0.64) and 9272143-C (OR = 0.65) between HLA-DRB1 and HLA-DQA1, rs55986091-A HLADQB1 (OR = 0.66). Based on the meta-analysis data, they involve genotype rs4646903-СС (OR = 4.65) CYP1A1 and protective alleles rs1801133-T (OR = 0.77) MTHFR, rs2333227-AA (OR = 0.57) MPO.Conclusion. The obtained data are critically important for development of laboratory techniques and reagent kits allowing for a personalized approach to identification of risk groups, which could benefit from compulsory vaccination and screening for pre-cancers of the cervix.

Background. The ongoing pandemic of a new coronavirus infection (COVID-19) determines the relevance of the analysis of epidemiological patterns of SARS-CoV-2 spread among the population of the Russian Federation.Aim — study of the manifestations of the epidemic process of COVID-19 in the Russian Federation in 2020–2022.Materials and methods. A retrospective epidemiological analysis of the incidence of COVID-19 in the Russian Federation was carried out from 03/30/2020 to 04/24/2022. The data from the Rospotrebnadzor report No. 970 “Information on cases of infectious diseases in persons with suspected new coronavirus infection”, information portal Stopcoronavirus.rf, etc. were used. The presence of SARS-CoV-2 RNA was confirmed by real-time RT-PCR.Results and discussion. The analysis of the manifestations of the epidemic process of COVID-19 in the Russian Federation in 2020–2022 showed the presence of two stages which differed depending on the influence of the biological factor and the ongoing anti-epidemic measures. There was a pronounced trend in the development of the epidemic process, starting from megacities (Moscow, Moscow region and St. Petersburg), which are major transport hubs and centers of migration activity of the population, to the regions of the Russian Federation. The SARS-CoV-2 pathogenicity has been shown to decrease with each subsequent cycle of the rise in the incidence of COVID-19 against the background of the increased contagiousness of the virus.Conclusion. As a result of the study, risk areas (megacities) and risk groups were identified.

Introduction. Hepatitis B retains the status of socially significant infection and remains a major health problem worldwide, including the Russian Federation. The improvement of the effectiveness of the current complex of preventive measures, especially vaccination, is an important task for public health. Although vaccination against hepatitis B is highly successful, 5% to 10% of individuals do not experience a response to vaccine with an adequate level of antibodies to hepatitis B surface antigen (anti-HBs). One of the key factors determining the absence or insufficiency of post-vaccination immunity against hepatitis B may be the single-nucleotide polymorphisms (SNPs) that change gene sequences, including those that determine the mechanism of immunogenesis. Such genetic changes may affect the signaling pathways and result in significant decrease in antibody response to hepatitis B vaccine. Assessment of epidemiological significance of such SNPs is an important task, considering its possible associations with failure to respond adequately to vaccination.The aim of the study was to determine the effect of SNPs of IL1B (rs1143634, rs1143627), IL1RN (rs4251961, rs419598), IL6 (rs1800795), IL10 (rs1800896), TULP1 (rs9380516), TLR4 (rs4986790), MERTK (rs4374383) genes on the formation of post-vaccination immunity against hepatitis B.Materials and methods. Healthcare workers (n = 271) of the Treatment and Rehabilitation Center of the Ministry of Health of the Russian Federation with known vaccination history, data on age, work experience and department of the medical institution were included in this research. The presence and levels of anti-HBs and anti-HBcore IgG antibodies were determined by the ELISA method using the DS-ELISA-ANTI-HBs and DS-ELISA-ANTI-HBc kits, according to the manufacturer’s instructions. Genotyping was performed by real time polymerase chain reaction. Statistical analysis of data was carried out using the "Statistica 6.0" software.Results. Statistically significant differences in the frequencies of CC (rs9380516) genotypes (p = 0.034; OR 0.497; 95% CI 0.261–0.949) and CT (p = 0.044; OR 1.967; 95% CI 1.015–3.812) of the TULP1 gene in the group of individuals with anti-HBs concentrations of 10–100 IU/l were found in association with the intensity of the post-vaccination response against hepatitis B. Also, for this group, differences were found in the structure of the TT/CT genotype pair of IL-10/TULP1 genes (rs1800896/rs9380516) (p = 0.003; OR = 5.39; 95% CI 1.7–17.4) and for the combination of AA/TT SNP MERTK/IL1RN genotypes (rs4374383/rs4251961) (p = 0.003; OR = 7.96; 95% CI 1.7–37.6).Conclusion. Our study revealed that above variants of genotypes could play a role in predicting an increased risk of low (or absence) post-vaccination immune response against hepatitis B. It seems appropriate to use the relationship between the gene polymorphisms and a low concentration of post-vaccination anti-HBs antibodies in assessing scenarios for the development of the epidemic process of hepatitis B, since the identified associations allow to quantify the risks of poor herd immunity against this infection.

JAK2 mutations can be associated with any phenotypic form of chronic myeloproliferative neoplasia, while MPL and CALR mutations occur, as a rule, in cases of essential thrombocythemia and primary myelofibrosis and they are not observed in polycythemia vera. In this article we describe a clinical case of CALR mutation (c.1154_1155insGTGTC; p.E386fs*46) presence in a JAK2-negative polycythemia vera patient at age 36. In January 2018 changes in his hemogramm were recorded for the first time. In June 2018, based on a diagnostic study of bone marrow trepanobiopsy, a diagnosis of polycythemia vera was made. Molecular genetic study of the patient’s DNA didn’t reveal mutations in the JAK2 (12 and 14 exons) and the MPL genes. CALR mutation was revealed during the screening by heteroduplex analysis with the electrophoresis in polyacrylamide gel. Then the mutation was identified by Sanger’s DNA sequencing as с.1154_1155insGTGTC; p.E386fs*46. The allelic burden level as determined by pyrosequencing was 20 % (June 2018). In conclusion we can suppose that the revealed CALR mutation с.1154_1155insGTGTC; p.E386fs*46 plays its role in our patient’s polycythemia phenotype.

The objective of this study was to characterize and compare antigenic and genetic characteristics of Streptococcus pneumoniae strains isolated from patients with invasive and non-invasive pneumococcal infections (PIs) by using the data of high-throughput sequencing.Materials and methods. A total of 158 S. pneumoniae strains were studied. All of them were isolated during different stages of the PEHASus multicenter study performed in 2015-2020. The data analysis was based on the information about whole-genome sequences of 46 strains isolated during the above study. Real-time PCR methods and high-throughput sequencing (the Illumina platform) were used for identification of serotypes. The SeroBA, PneumoCaT software and PubMLST.org website resources were used in the data processing.Results and discussion. The serotypes of all the studied strains were identified. A number of discrepancies among serotypes in serogroup 6 and one discordant result were revealed by the analysis of whole-genome sequences using 2 programs. The PCR methods were effectively used to characterize serotypes in 87% and 69% of the pathogens of invasive and non-invasive PIs, respectively. The serotypes contained in PCV13 accounted for 59% and 37%, while PPV23 serotypes accounted for 78% and 53% of the strains isolated from patients with invasive and non-invasive PIs, respectively. The data analysis was unable to identify either the dominant sequence type (a total of 81 sequence types have been identified) or clonal complexes, except for serotype 3 strains, thus demonstrating consistency with the data from previous studies suggesting the absence of a well-represented clonal structure of S. pneumoniae associated with pneumococcal meningitis in Russia.Conclusion. The obtained data made it possible to identify the distribution of the circulating serotypes and genetic characteristics of the strains isolated from PI patients, thus being instrumental for assessment of the effectiveness of the existing polyvalent vaccines and providing information for improvement of the PCR-based methods of serotyping.

Introduction. Detection of somatic mutations in the BRAF gene can be used in clinical oncology to clarify the diagnosis, select therapy and assess the prognosis of the disease. Pyrosequencing technology makes it possible to identify both already known and new mutations, as well as to determine the mutant allele ratio in the sample.The aim of the study was to develop the pyrosequencing-based method for detecting mutations in 592–601 codons of the BRAF gene.Material and Methods. The nucleotide sequences were obtained using «PyroMark Q24» instrument. The sensitivity and specificity of the method were estimated using dilutions of plasmid DNA samples containing the intact BRAF gene fragment mixed with sequence containing one of the mutations V600E, V600R, V600K, V600M, and K601E. The clinical testing was performed on 200 samples from thyroid nodules.Results. The developed method makes it possible to determine samples containing 2 % of the mutant allele for mutations V600K and V600R, 3 % for V600E and V600M, and 10 % for K601E. The pyrogram signal values for samples without mutations ranged from 0 to 19.5 % for different mutations. An analysis algorithm was developed to confirm the presence and differentiation of mutations in the 600 codon at a low proportion of the mutant allele based on the signals ratio on the pyrogram. The 47 clinical samples with mutations were found, 45 with V600E and 1 with V600_K601>E, for one sample, the type of mutation in the 600 codon could not be determined. The proportion of the mutant allele was 3.5–45 %. The concentration of extracted DNA less than 10 copies per mkl was obtained in 47 samples, of which 8 samples were found to have the mutations.Conclusion. The pyrosequencing-based method was developed for the detection of somatic mutations in 592–601 codons of the BRAF gene. The technique provided sufficient sensitivity to detect frequent mutations in the 600 codon and allowed the detection of rare mutations. Extraction of DNA from clinical samples obtained by fine-needle aspiration biopsy in most cases provided a sufficient concentration of DNA, which made it possible to use the technique in combination with cytological analysis without additional sampling. This approach can be applied to determine somatic mutations in DNA fragments of same length for other oncogenes.

Relevance. Unencapsulated strains of meningococcus (NmNG) very rarely cause invasive meningococcal disease. A new ciprofloxacin-resistant strain NmNG ST-175 cc175, which has recently caused several cases of invasive meningococcal infection in Europe, has been discovered in the Russian Federation.Aim. To compare the new Russian strains of NmNG ST-175 with the already characterized NmNG ST-175 and to analyze the genetic markers associated with antibiotic resistance.Materials and methods. The nucleotide sequences of NmNG ST-175 strains of more than 2 million base pairs were exported from the PubMLST database. Comparison was carried out for 1605 core genome loci using the N. meningitidis cgMLST v1.0 ". Genetic relationships of 127 NmNG ST-175 strains, including 8 Russian carrier strains, were visualized using the SplitsTree software (version 4.16.2).Results. Of the eight Russian strains, six found themselves in a cluster with German and Swedish isolates that caused invasive meningococcal infection. Two more Russian strains were closest to the isolates of the cluster, including carrier isolates from England. Seven strains showed resistance to ciprofloxacin and possessed the gyrA-187 and gyrA-152 alleles, which, based on the phylogenetic analysis of the alleles, belonged to the genetic branches of Neisseria cinerea and Nm.Conclusion. Protein vaccines alone could potentially provide protection against ST-175 NmNG. It seems promising to study the antigenic characteristics of Russian Nm strains, including NmNG ST-175, to assess the potential vaccination coverage with existing protein vaccines, the possibility of their registration on the territory of the Russian Federation, as well as the development of domestic vaccines.

Introduction. Borrelia miyamotoi is a pathogen causing erythema-free ixodid tick-borne borreliosis (ITBB), a disease widespread in Russia. The genome of B. miyamotoi contains genes of multiple variable major proteins (Vmps). Vmps fall into two families — Vsps and Vlps (with subfamilies δ, γ, α and β). At a particular time, a single B. miyamotoi expresses only one variant of Vmp gene.The purpose of the work is to develop a technique for identification of the Vmp present at the expression site.Materials and methods. The technique is designed in the format of a real-time multiplex PCR. It was tested by using B. miyamotoi DNA samples extracted from blood collected from 172 ITBB patients and 109 ticks. The samples were collected in 14 regions of Russia.Results. The new technique made it possible to identify the expressed Vmp in 82% of the examined samples, thus having demonstrated its efficiency. Negative results were much less often observed with samples from patients than with samples from ticks. At the same time, the percentage of samples with one type of Vmp is identical for clinical samples and ticks, while the percentage of samples containing concurrently two types of Vmps is significantly higher among samples from patients with the most frequent occurrence of the Vlp-δ and Vsp combination.Discussion. The frequent occurrence of the combination of two Vmp types in the blood samples can indicate the concurrent presence of several subpopulations of B. miyamotoi in ITBB patients. A new antigenic Vmp variant is synthesized after protective antibodies have been produced for the major protein of the strain transmitted by a tick. This phenomenon known as immune evasion allows the pathogen to persist within a host.Conclusion. The developed technique of real-time multiplex PCR allows to simultaneous detect of several antigenic variants of the variable basic surface proteins of B. miyamotoi. The study of the antigenic spectrum of B. miyamotoi strains in comparison with the characteristics of conserved regions of the genome by the method of multilocus sequencing will clarify the stages of evolution and distribution of B. miyamotoi sensu lato.

Background. In accordance with the World health organization clinical guidelines, the analysis of somatic mutations in the CALR gene, as well as mutations in the JAK2 and MPL genes, are included in the list of criteria for the Ph-myeloproliferative neoplasms diagnosis.More than 50 different mutation variants have been found in the CALR gene, among which the most frequent are a 52 bp deletion (c.1092_1143del), also called type 1, and a 5 bp insertion (c.1154_1155insTTGTC), also called type 2 (88 %).The remaining 12 % are other type less frequent indels or combinations thereof.It is most convenient to use sequencing methods to identify all possible variants of CALR mutations. It is also important to develop inexpensive screening test that can detect any mutations in the analyzed DNA fragment of CALR gene. This method can be heteroduplex analysis followed by electrophoresis on polyacrylamide gel (PAGE).The objective: to develop and demonstrate the feasibility of using heteroduplex analysis with separation of the PCR product by electrophoresis on non-denaturing PAGE for the CALR exon 9 mutations detection as the screening test. Materials and methods. DNA samples of 13 CALR-positive patients with different phenotypic variants of Ph-myeloproliferative neoplasms were screened by heteroduplex analysis. For the most common variants of CALR mutations (c.1092_1143del and c.1154_1155insTTGTC), a threshold determination of the mutant allele presence was analyzed.Nucleotide sequence of exon 9 fragment was determined using Sanger sequencing. Also, all 13 samples were analyzed using the pyrosequencing method to assess the allelic burden level.Results. Heteroduplex analysis revealed mutations in exon 9 of the CALR gene in all 13 patients. The threshold determinations of the method in the case of the c.1154_1155insTTGTC and c.1092_1143del analysis are 6.25 % and 3.13 % of the mutant allele presence in the patient sample, respectively.Conclusion. The proposed variant of the heteroduplex analysis with separation of the PCR product by electrophoresis on non-denaturing PAGE can be recommended for use as the preliminary screening test which is carried out before the confirming sequencing methods for the different indels (or combinations thereof) CALR mutations determine.The presence of heteroduplexes indicates the presence of a mutation, even if the mutant product is not visualized (in case of small mutations).

Antipsychotic drugs are a critical modality in managing of schizophrenia. Although medications can be highly effective, response varies and some patients derive considerably less benefit than others. Long-term use of antipsychotic drugs is associated with the development of adverse reactions. Te safety advantages of the atypical drugs havebeen questioned because of their propensity to induce weight gain and alter glucose and lipid metabolism. Antipsychotic-induced weight gain is a common cause of self-discontinuation of treatment and a significant deterioration in the quality of life in patients with schizophrenia. Te severity of adverse reactions when taking antipsychotics in different patients varies, which be associated with genetic factors. Antipsychotic induced weight gain is a major health concern and unfortunately, there is no predictive tool to identify who are high risk individuals. Te LEP, LEPR and NRY genes represents a compellings candidates for genetic studies of antipsychotic-induced weight gain. Candidate gene selection should rely on current knowledge on the molecular pathways to weight gain, antipsychotic pharmacokinetics and pharmacodynamics, as well as possible disease-related genetic links to the side effects under study. Pharmacogenetics will provide rational treatment based on matching antipsychotics to a patient’s DNA profile, thus, potentially providing effective treatment with minimal side effects to outliers and mean responders to a given antipsychotic medication.

Relevance. Population migration can play a crucial role in the spread of invasive strains of meningococcus, initiating outbreaks of meningococcal infection, and changing the incidence at the local level.Aim. To assess the prevalence of meningococcal carriage among migrants arriving in Moscow and to characterize the antigenic and genetic properties of carrier strains of meningococcus.Materials and methods. The study was conducted in March 2020 at the bases of the Multifunctional Migration Center of Moscow and the Federal Budget Institution of Science «Central Research Institute of Epidemiology». Samples of nasopharyngeal mucus were collected from 352 people. Nasopharyngeal strains of meningococcus were identified and identified using microbiological, serological, and molecular biological methods.Results. The overall level of the carriage was 5.7%. Of the twenty selected strains, 10 have a serogroup defined: Y – 5 strains, W - 3, A, and B – 1 each. The obtained genetic and antigenic characteristics do not allow talking about the import into the RF of representatives of known hypervirulent clonal complexes. In this study, strains were identified that are part of the clonal complex ST-175 complex, which has not been previously described in the Russian Federation.Conclusion. It seems promising to continue the dynamic monitoring of carriage of meningococcus in various groups, including among people entering the country to obtain a migration patent, as well as identifying risk factors for acquiring carriage. The data obtained will supplement current information on the incidence of the generalized form of meningococcal infection and will be crucial for determining the epidemiology at the country level, the population groups responsible for the transmission of the disease, and the need for targeted vaccination.

Background. There are problems related to both quantitative assessment of an allele burden level of a mutant gene and interpretation of results in DNA samples with the burden level of the mutant allele less than 15–20 %, when using Sanger sequencing for analyzing somatic mutations. Applied Biosystems (USA) has developed new software Minor Variant Finder, which allows determining mutations with the allele burden level from 5 %.The objective: to determine the allele burden level and identification of minor variants of somatic mutations in the ASXL1, JAK2 genes and BCR-ABL oncogene using Minor Variant Finder software in patients with myeloproliferative neoplasms.Materials and methods. The level of mutant allele burden for 15 patients with myeloproliferative neoplasms was determined by the identified mutations using the Minor Variant Finder software, after analysis of point somatic mutations in the ASXL1, JAK2 genes and BCR-ABL oncogene by Sanger sequencing.Results. The allele burden level in all 5 ASXL1-positive samples and BCR-ABL-positive sample was determined as higher than 20 % using the Minor Variant Finder software. The allele burden level in 2 cases was higher than 20 % and in 7 cases lower than 20 %, when we analyzed 9 JAK2-positive samples.Conclusion. Minor Variant Finder software can be used to estimate the allele burden level and to identify minor variants of somatic mutations in the ASXL, JAK2 and BCR-ABL genes.

Relevance. Gilbert’s syndrome (GS) is a disease with an autosomal recessive type of inheritance caused by either impaired expression of the UGT1A1 gene, which encodes the isoform of the uridine-5-diphosphate glucuronosyltransferase (UDP-GTA1), or structural modifications of UDP-GTA1. GS is characterized by unconjugated hyperbilirubinemia; drug metabolism disorders and the development of drug-drug interactions. For diagnosis of GS, molecular biological methods are used to determine single nucleotide polymorphisms (SNP). Data on the prevalence of SNP related to GS in Russia are scarce. Study objective: Detection of genetic variant (TA)5/6/7/8 (rs8175347) in the UGT1A1 gene (Gilbert’s syndrome) by pyrosequencing in outpatient practice.Material and methods: 200 outpatients were examined. Of whom: men — 107 (53.5 %), women — 93 (46.5 %) aged 15 to 86 years; patients from 30 years and older formed the majority — 175 (87.5 %). Detection of the genetic variant (TA)5/6/7/8 (rs8175347) in the UGT1A1 gene (GS) was carried out by pyrosequencing using the PyroMark AmpliSens® Pyroscreen UGT1A1 genetic analysis system (manufactured by the Federal Budgetary Scientific Institution Central Research Institute of Epidemiology of Rospotrebnadzor, Russia). For comparison, sequencing according to F. Sanger was used.Results: Normal (TA)6/(TA)6 genotype was found in 71 (35.5 %) patients, (TA)6/(TA)7 genotype was found in 81 (40.5 %) (heterozygous status) and (TA)7/(TA)7 genotype — in 48 (24 %) (homozygous status). Rare (TA)5/(TA)6, (TA)5/(TA)7, (TA)6/(TA)8 and (TA)7/(TA)8 genotypes were not found. The results of the determination of (TA)6/(TA)7 genotypes in the homo- and heterogeneous status by pyrosequencing and Sanger sequencing were the same in all cases. In 30 out of 48 patients, GS was newly diagnosed, and in half of the cases these patients were persons of the older group. None of them showed an increase in bilirubin level.Conclusion: The incidence of GS in outpatients was 24 %. Pyrosequencing allows us to identify various variants of the (TA)5/6/7/8 polymorphism in the homo- and heterozygous status. AmpliSens® Pyroscreen UGT1A1 kit can be used in clinical practice to diagnose GS and to assess side effects of prescribed drugs.

Aim. Estimation of the prevalence of Ixodes tick-borne borreliosis (ITBB) without erythema migrans in the Krasnoyarsk Territory in comparison with the prevalence of ITBB with erythema, that is Lyme disease (LD). Detection of cases of «new» infection caused by Borrelia miyamotoi (ITBB-BM) among patients with suspected tick-borne infection, primarily among patients with ITBB without erythema (ITBB-W/O-E). Materials and methods. We analyzed the database of Rospotrebnadzor containing individualized information about cases of ITBB officially registered in the Krasnoyarsk Territory in 2011 — 2016. Blood samples from 18 patients with suspected of tick-borne infection hospitalized in the Krasnoyarsk Emergency Hospital in 2017 were collected and studied. ITBB-BM was diagnosed if B. miyamotoi DNA was found in patient’s blood and/or the antibodies to specific antigens of B. miyamotoi were detected. Results. With an average annual incidence rate of ITBB of about 10 cases per 100,000 population, the proportion of ITBB-W/O-E among all ITBB in the Krasnoyarsk Territory is about 45%. Epidemiological characteristics of ITBB-W/O-E and LD are similar, but these diseases are very different clinically. All cases of ITBBW/O-E and some cases of tick-borne infections of unknown etiology, subjected to our enhanced laboratory diagnostics, were caused by B. miyamotoi. Conclusion. ITBB-W/O-E is a widespread disease in the Krasnoyarsk Territory, caused mainly by B. miyamotoi. The results of the study significantly expand the known geographic area of the ITBB-BM.

Hemorrhagic fever with renal syndrome (HFRS) is a zoonotic infection disease caused by Orthohantavirus which belongs to Hantaviridae family. This article is a brief review of recent data about genetic factors which play a role in individual predisposition toward HFRS. There are reports discovered associations of polymorphic sites with HFRS severity and risk complications. Polymorphic sites in genes which code proteins of immune (МНС, TNF, IL1) and endothelial (VE-cadherin) systems, blood coagulation (SERPINE1, ITGA2B, NOS) and detoxification (CYP1A1, GSTP1) systems and their links with disease are described in this article. HLA haplotypes B*08-DRB1*03 and B*46-DRB1*09, B*51-DRB1*09 are associated with severe forms of HFRS-PUUV and HFRS-HTNV respectively. TNF A-allele and AA-genotype in -308G>A SNP (rs1800629), CDH5 ТТ-genotype in 1550T>C SNP, SERPINE1 G-allele in -844A>G SNP (rs2227631), alleles HPA3 b, NOS2A*11 and NOS2A*11/NOS2A*12-genotype, CYP1A1 1А2С-genotype in SNP (rs1048943) and GSTP AG-genotype in SNP (rs1695) demonstrated associations with severe HFRS. Differences in the expression levels of GATA3, T-BET, CD3, IFNβ, NFkB, STAT1 and MxA genes in cell cultures stimulated by hantavirus. Expression of GATA3 was significantly higher in cell cultures of patients with severe HFRS than with a mild form. In contrast, MxA gene expression was up-regulated in cell cultures of patients with mild HFRS-PUUV. Considering individual genetic factors of HFRS patients would allow defining the best tactic of therapy and prophylaxis in this way. And as a result of applying this treatment in the clinical practice decrease of unfavorable disease outcome would occur.

Schizophrenia is a socially significant mental disorder characterized by early onset and high time and financial expenditure on treatment. The basic drugs in these patients are antipsychotics that are highly effective against the positive and negative symptoms of schizophrenia, but at the same time have a wide range of adverse reactions (ARs). The clinical effect and tolerability of antipsychotics are variable and depend on the characteristics of genetically determined mechanisms (transportation, biotransformation, and elimination).The paper describes a clinical case of a female patient with schizophrenia who has been noted to be unresponsive to antipsychotic therapy for some years after the onset of the disease. After pharmacogenetic testing, she was found to be homozygous for the nonfunctional allelic variant CYP2D6*4 (1934 G>A, rs3892097), which was the reason for the complete shutdown of isoenzyme 2D6 activity and the development of ARs in the use of initial doses of antipsychotic drugs, as well as for an increase in the severity of ARs with aggravation of psycho-producing symptoms with an even slow titration of the daily dose.

Introduction. The invasive meningococcal disease (meningitis and/or septicemia) is actual problem of public health in Russia. Neisseria meningitidis isolates are classified into serogroups, PorA/FetA VRs, sequence types and clonal complexes. The growth of the invasive forms of meningococcal infection caused by isolates with «W: P1.5,2: F1-1: ST-11 (cc11)» profile requires attention for extended genotyping because the discriminating ability of classical MLST and antigens typing does not allow to answer the question about genetic and antigenic features of the pathogens and their epidemic potential. Materials and Methods. Four N. meningitidis serogroup W isolates associated with invasive meningococcal disease in Moscow (Russia) were characterized by next-generation sequencing. Three isolates were sequenced directly from cerebrospinal fluid samples and one -as a bacterial culture. All isolates were characterized earlier and the data were published in the PubMLST data base (id38565, id38573, id50225 and id50241). Genomic DNA was sequenced on Illumina MiSeq instrument. Results and Discussion. Obtained sequences allowed us to characterize four meningococci isolates for more than 1400 loci from the core genome MLST scheme. We have analyzed the core genome MLST scheme information about surface-antigen coding sequences. Housekeeping genes sequences were used to determine eMLST profile, ribosomal protein genes and some antibiotic resistance associated genes. We have characterized some ribosomal protein genes and antibiotic resistance associated genes. Based on eMLST profiles we noticed that there are at list two clones of N. meningitidis serogroup W inside complex ST-11/ET-37 clonal complex circulating in Moscow during 2016. An eMLST profile of isolates id50225 and id50241 differs in 3 loci out of 20. Application of the approach based on next-generation sequencing in routine epidemiological surveillance dramatically increases the amount of data and genotyping discriminating ability.

Intensive development of DNA analysis technologies and largescale genome-wide association studies have led to accumulation of a large array of data on the relationship between genetic factors and various phenotypic manifestations, including monogenic and polygenic hereditary diseases. This greatly has extended the capabilities of clinical diagnostics and predictive medicine in the field of socially significant diseases. For example, a role of agenetic component of the risk for such a multifactorial and polyetiologic disease as stroke is now actively explored. Large-scale studies have revealed both general and specific genetic markers associated only with a certain type and subtype of stroke. This review analyzes the current state of the problem of using genetic markers for diagnosis of stroke predisposition, complex issues associated with multiplicity of risk factors for stroke, and possible development in this area.

As it was reported by World Health Organization, the annual number of deaths caused by pneumococci reaches 1,6 million people including 700 thousand of children of 0–5 years old. Pneumococcal meningitis is one of the most severe manifestations of pneumococcal infection. S. pneumoniae keeps one of the leading positions in aetiological structure of bacterial meningitis. Data on serotypes distribution of pneumococci in children of the early age in the Russian Federation are limited. In this study serotypes of pneumococci from pediatric patients with pneumococcal meningitis (children aged 0–7 years, n =35) isolated in 1981–2010 in Moscow were identified. Screening for sensitivity of strains to penicillin was performed, as well as molecular typing by MLST approach. The most frequent serotypes causing pneumococcal meningitis in children aged 0–7 years were 19F, 7F, 14 and 1. Most of MDR-strains were collected in 2007–2009. Among MDR-strains and those demonstrating increased resistance to antibiotics, several new allelic patterns were described.