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Acute COVID-19 is a viral infection caused by a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that results in dramatically decreased peripheral blood CD3+T cell count apparently due to alterations of thymic T cell maturation, that can persist long term afterwards. Therefore, we analyzed the levels of peripheral blood TRECs (T-cell receptor excision circles), and investigated the main alterations in peripheral blood T cell subsets in COVID-19 convalescents. We performed molecular quantification of TRECs with “TREC/KREC-AMP PS” kit and flow cytometric analysis of peripheral blood lymphocytes from three groups of patients. The first group contained 109 samples from COVID-19 convalescents (6–12 month post-acute COVID-19) with normal levels of TRECs (TRECn); the second was formed from COVID-19 convalescents (6–12 month post-acute COVID-19) with decreased levels of TRECs (TREClow, n = 29), and healthy control group (HC, n = 18). We noticed no significant differences between all three groups in CD3+T cell relative and absolute numbers. However, CD4+T cell frequencies were decreased in TREClow and TRECn groups compared to HC (40.8% (31.6; 50.1) and 46.4% (40.0; 53.0) vs 53.5% (47.36; 56.9), p 0.001 and р = 0.004, respectively). Furthermore, Th cell levels were decreased in TREClow patients vs HC and TRECn groups (701 cell/1 µL (478; 807) vs 1005 cell/1 µL (700; 1419), р = 0.020, and 876 cell/ 1 µL (661; 1046), р = 0.008, respectively). Finally, both groups of COVID-19 convalescents had increased frequencies of circulating CD8+T cells — 29.4% (20.7; 39.7) in TREClow group, 26.5% (21.1; 32.7) in TRECn group vs 21.3% (17.1; 26.0) in healthy controls (p = 0.024 and р = 0.026, respectively). In TRECn group, CD8+T cell count was elevated vs control range (508 cell/1 µL (372; 622) vs 356 cell/1 µL (247; 531), р = 0.044). Thus, COVID-19 convalescents (6–12 month post-acute COVID-19) showed an imbalance in CD4+and CD8+T cell level even at 6–12 months post-acute SARS-CoV-2 infection, and the observed changes in peripheral blood T cells could be closely related to the alterations in thymic T cell maturation and differentiation. Such a long-term decline in TREC levels in the circulation may have a profound impact on immune system functions and requires immunocorrection therapy.

The main cause of edema in hereditary angioedema (HAE) is due to elevated bradykinin levels, caused either by C1-INH deficiency/change in functional activity and caused by mutations in the SERPING1 gene or by mutations in the F12, PLG, ANGPT1, KNG1, MYOF and HS3ST6 genes with a normal level and functionality of the C1-esterase inhibitor. The aim of the work was in silico prognostic analysis of the rare synonymous variant NC_000003.12:g.186725098T>C in the KNG1 gene and its impact on the development of HAE symptoms. The material was a whole blood sample obtained from a woman with clinical manifestations of hereditary angioedema without a decrease in the levels and function of the C1 inhibitor. The research methods included whole exome sequencing, bioinformatic analysis of the KNG1 gene mutation using a number of databases and web resources. Results. When processing full-exome sequencing data, we detected a synonymous variant in the KNG1 gene (exon 4, isoform 1): NC_000003.12:g.186725098T>C. The patient is a heterozygous carrier of the variant, with a frequency of 0.000004 (1:264690). Presumably, the identified variant can lead to the development of sporadic edema through several pathways that are associated with the formation of bradykinin or its analogues. Therefore, (1) the mutant high-molecular-weight kininogen is more easily activated by kallikrein and becomes a source of bradykinin formation through the kallikrein-kinin system; (2) the mechanism of bradykinin formation undergoes significant changes and results in the formation of functionally active but aberrant bradykinin, which alters its inactivation by enzymes with a consequent increase in its half-life, (3) the changes in positions 380-389 bring about modifications in Lys-bradykinin reproduction such that in subsequent steps it is “easily” cleaved to bradykinin by arginine aminopeptidase. The results of our study therefore indicate a possible role of the identified variant in the KNG1 gene in the development of HAE.

Leptospirosis is a zoonotic disease found virtually worldwide. Microscopic Agglutination Test with live leptospira (MAT) is the reference method for the serological diagnosis of leptospirosis. MAT is based on assessing serum potential to agglutinate live reference serovar Leptospira maintained at a reference laboratory. At some laboratories having own collections of isolated and reference Leptospira strains applicable for serological diagnosis, those microorganisms are maintained for many years by repeated subculturing, that increases markedly a chance of strain cross-contamination. The lack of adequate quality control for reference strains may affect data of epidemiological studies. Control of Leptospira spp. reference strains purity and stability of their antigenic composition is very important for diagnosis of leptospirosis. The study objective was to compare the 16S rRNA gene nucleotide sequences of some Leptospira strains from the collection of the St. Petersburg Pasteur Institute to with relevant sequences uploaded to GenBank. In this study, 38 Leptospira strains were investigated. Nucleotide sequences of 36 strains were deposited in the international GenBank database, inconsistencies were revealed in two strains. The study found that the control Leptospira strains from the collection of the St. Petersburg Pasteur Institute had minimal dissimilarities from international control strains. The analysis of the resultant 16S rRNA sequences has shown the presence of point mutations, transitions, deletions and insertions, regardless of the strain species. The open leptospira pan-genome demonstrates high genomic variability in species due to the capability of leptospira for lateral gene transfer in order to adapt to changing environmental conditions. The massive acquisition and loss of genes give rise to an increased species diversity. The 16S rRNA gene is suitable for screening diagnostics; however, high level of the fragment similarity and close phylogenetic relationship between different species put bounds to its use in genotyping. The presence of point nucleotide mutations is most likely associated with the evolutionary mechanisms of leptospira, their ability to horizontal gene transfer and crossing-over, including ribosomal genes, but this assumption necessitates additional research. For specimen genotyping it is necessary to select alternative genes with high specificity and sufficient level of nucleotide divergence. The study shows a need for genetic analysis of collection strains in order to control the purity of cultures.

The use of dried blood spots (DBS) obtained from the heels of infants has many advantages over the collection of whole blood samples. DNA extracted from DBS can be used to detect genetic diseases by PCR, which has contributed to the development of population-based newborn screening worldwide. Since January 2023, the list of identified diseases includes a group of primary immunodeficiencies (PIDs), associated with the absence or decrease in the levels of T and/or B lymphocytes, determined as part of screening by the levels of TREC and KREC molecules in peripheral blood, respectively. Quantitative analysis requires special attention to biological material. The aim is to evaluate the impact of the preanalytical step on the quantitative analysis of TREC/KREC levels in peripheral blood.The material included 5219 DBS obtained from infants on days 3-4 of life, as well as DBS prepared from the whole blood of 100 apparently healthy individuals aged 18 to 29 years. A comparative analysis of the TREC/KREC molecules number in correctly and incorrectly collected DBS from newborns and adults, as well as depending on the volume of applied blood, was carried out by RT-PCR using test systems to assess the levels of TREC and KREC in peripheral blood. DBS quality was assessed visually.In the first months of the project, a significant number of incorrectly taken samples were identified – over a third of all DNA maps received for each corresponding month. As a result of additional training of medical staff, the amount of incorrectly collected material decreased to a level not exceeding 1% of all monthly samples collected. When using DNA extracted from DBS with application errors, the majority of samples (64% for newborns, 78% for adults) failed to obtain a result. In the remaining cases, the results obtained were significantly lower than the normal levels of TREC/KREC determined in the same samples with correct DBS collection (all p < 0.0001, 95% CI). The volume of blood used when correctly applied to Guthrie cards did not affect the results obtained, TREC and KREC levels were comparable; when comparing the medians calculated for each group of samples, no significant differences were identified (p > 0.05).When quantitatively analyzing TREC/KREC levels in peripheral blood, correctly taken material is fundamental importance to obtain reliable indicators, primarily to exclude false-positive results. To minimize errors in the preanalytical stage, additional training of medical personnel is necessary to control and/or correct errors.

Neonatal screening is a mandatory newborn screening procedure that detects the presence of genetic diseases. Dry blood stains are used for mass screening of children. This technology is the most affordable and convenient for the transportation and storage of biological material. DNA extraction is one of the important steps in molecular diagnostics, the accuracy of which is particularly important in genetic analysis. Different DNA extraction kits offer different protocols and reagents, which may vary in efficiency and quality of extraction.The aim of our work was to perform a comparative analysis of reagent kits for DNA extraction from dried blood spots.The materials were dried blood drop samples on Guthrie cards obtained from healthy preterm infants on day 3-4 of life as part of a newborn screening program.The study methods included spectrophotometric analysis to determine the concentration and purity of DNA, the simplicity of protocols, the duration of isolation, and the possibility of automating the process were also evaluated. The efficiency of DNA isolation using different reagent kits was additionally monitored by real-time PCR results using a test system to assess the level of TREC and KREC in peripheral blood, since quantitative analysis requires more attention to the material under study.In assessing the purity of the nucleic acid extracted using four kits analyzed, successful deproteinization of DNA samples and relative purity could be observed. The average DNA purity for the “Extra-DNA-Bio” set was 2.2±0.23, for “EXTRA-prep PS” – 1.89±0.23, for “MagnoPrime UNI” with manual and automatic isolation – 2.31±0.21 and 2.85±0.09, respectively. The average DNA concentration for the Extra-DNA-Bio kit was 15.28 μg/mL, for the EXTRA-Prep PS kit it was 16.26 μg/mL, and for the MagnoPrime UNI for manual and automated isolation it was 62.5 μg/mL and 102.28 μg/mL, respectively. According to the applied Kraskell-Wallis criterion and Dunn’s test, significant differences in both TREC and KREC parameters are present between the group of DNA samples extracted using the “MagnoPrime UNI” reagent kit for manual extraction and the groups of samples extracted by other methods (“MagnoPrime UNI” for automatic extraction) or “Extra-DNA-Bio” and “EXTRA-Prep PS” kits.In the course of the present study, four comparable reagent sets demonstrated a high level of convergence of the obtained data, satisfying all the necessary parameters for further molecular genetic analysis, can be used for neonatal screening and in other areas of research requiring DNA extraction from a dried blood spots.

The aim of the work was to estimate the prevalence of HIV, HBV and HCV markers among pregnant women and their male partners in the Republic of Guinea.Materials and methods. The material of the study was blood plasma samples from 140 pregnant women living in Kindia prefecture and N’Zerekore prefecture, as well as 60 male partners who reported sexual contact with HIV-infected persons. The samples were examined for the presence of serological (HBsAg, HBeAg, antibodies anti-HBs IgG, anti-HBcore IgG, anti-HBe IgG, anti-HCV IgG, Ag/Ab-HIV) and molecular (HBV DNA, HCV RNA, HIV RNA) markers.Results and discussion. The age of the examined pregnant women ranged from 13 to 55 years and was on average (26.29±9.67) years. The age of men varied from 15 to 60 years, on average – (29.05±11.99) years. When assessing the prevalence of serological markers, antibodies to HCV were detected in 2.14 % cases in women and in 3.33 % cases in men. Antibodies to HIV were found in 6.43 % and 6.67 % women and men, respectively. Serological markers associated with HBV were detected in 80.71 % (HBsAg – 13.57 %) of women and 81.67 % (15 %) of men. In the pregnant women, HCV RNA was not detected, HIV RNA was revealed in 1 case, HBV DNA was identified in 26 cases (18.57 %), including 5 % HBsAg-negative hepatitis B cases. In the men group, HCV RNA and HIV RNA were detected in 3.33 % and 6.67 % cases, respectively. HBV DNA was determined in 16.67 % of men, including latent hepatitis B in one person. A significantly higher incidence of HIV in men compared to women is shown (χ2=3.907 at p

Hereditary angioedema (HAE) is a genetically determined disease characterized by recurrent attacks of edema affecting the subcutaneous and/or submucosal layers of tissue, face, lips, neck, extremities of the body, oral cavity, intestine and/or larynx. In the latter case, the disease becomes life-threatening. The majority of HAE cases are associated with decreased levels of C1 (C1-esterase inhibitor), there are also descriptions of HAE with dysfunctional C1 inhibitor and HAE with normal C1 inhibitor. In the first and second variants, mutations in the C1NH gene are the cause of the disease. HAE with normal quantitative and functional levels of C1-inhibitor has the same clinical manifestations but with mutations in other genes, including F12, PLG, ANGPT1, KNG1, MYOF, and HS3ST6. Currently, mutations in the HS3ST6 gene remain poorly understood; only one missense mutation (p.Thr144Ser, rs746467957) associated with the development of HAE has been described.The aim of our work was to study new mutations in the HS3ST6 gene and analyze in silico their prognostic nature and clinical significance for the development of hereditary angioedema.The material was whole blood samples obtained from 13 patients with symptoms of hereditary angioedema without reduced levels and function of C1-INH.Whole exome sequencing of patients, bioinformatic analysis of HS3ST6 gene mutations using a number of databases and Web resources to predict the effect of mutations on the protein and assess the conservatism of the positions of the mutations detected was involved in study methods.Mutations in the HS3ST6 gene were identified in four patients, including two cases with two mutations simultaneously. Application of bioinformatic analysis allowed us to obtain new data on four missense mutations in the studied gene. Potential pathogenetic significance was determined for three of them. The mutation NC_000016.9:g.1962132G>A (p.A163V) is most likely to be involved in pathogenesis of HAE by indirect disruption of heparan sulfate O-sulfation directly within the protein. The NC_000016.9:g.1962024G>A mutation (p.P199L) appears to lead to the development of the disease through disruption of docking with SDC2 heparan sulfate. In the NC_000016.9:g.1962046C>T (p.A192T) mutation, destabilization of the 192 amino acid position next to PAPS, may contribute to disruption of heparan sulfate O-sulfation through disruption of protein functional activity and, therefore, catalysis transfer of sulfo group to heparan sulfate syndecan-2. Thus, in all three cases, the formation of HAE appears to be possible due to disruption of the O-sulfation steps of heparan sulfate syndecan-2.Considering that in silico methods offer new opportunities to assess the pathogenetic significance of mutations, the application of bioinformatic analysis can contribute to a detailed investigation of the causes of hereditary angioedema. The present work convincingly demonstrates that rare mutations in the HS3ST6 gene may be involved in the pathogenesis of HAE and provoke edema due to increased bradykinin release.

Increasing attention is being paid to methods for detecting primary and secondary T and/or B cell immunodeficiencies. Their implementation into laboratory diagnostics would contribute to the early diagnostics of immunodeficiencies. Currently, the number of identified adult patients with immunodeficiencies of various origins is steadily increasing. Age, gender and ethnicity of patients may be significant factors of immunity. Hence, determination of the population reference intervals for TREC and KREC DNA excision rings in peripheral blood of adult persons is an urgent laboratory task for in-depth examination of both congenital and acquired immunodeficiency conditions. Our purpose was to determine the reference intervals for the quantitative assay of TREC and KREC fragments in peripheral blood among the adult population of St. Petersburg. We studied whole blood samples obtained from 717 apparently healthy volunteers aged 18 to 108 years within the program of population immunity assessment among residents of St. Petersburg. The exclusion criterion included immunodeficiency of any origin, viral hepatitis A, B, C, HIV infection. Quantitation of the target TREC and KREC DNA fragments was carried out using a set of reagents for the quantitative determination of excisional rings TREC and KREC by Real-time PCR (TREC/KREC-AMP PS). The reference intervals were determined by the direct method according to the recommendations of the International Federation of Clinical Chemistry and the Russian State Standard (GOST) R 53022.3-2008. The volunteers were divided into six age groups: 18-29, 30-39, 40-49, 50-59, 60-69 years old, and the persons over 70. The amounts of TREC and KREC in each blood sample were determined for all age groups. Upon correlation analysis, we have revealed a negative relationship between the concentration of TREC molecules in blood samples, and the age of study participants (Spearman correlation coefficient r = -0.80 (p-value < 0.0001)). Significant differences in TREC levels between different age groups were revealed. No correlations were detected between KREC contents in blood samples and age as well as any differences between age groups. Reference intervals of the TREC level were determined for each mentioned age group. A unified reference range was established for the KREC levels. The established reference intervals for TREC and KREC molecules in adults are significantly lower than in newborns. The obtained results enable determination of reference intervals for TREC and KREC levels among adults, thus contributing to effective personalized laboratory diagnosis of immunodeficiency states of various origins.

Introduction. The immune status is a multifaceted parameter quantitatively and qualitatively analyzing functional activity immune system state in immune organs as well as some non-specific mechanisms of antimicrobial protection. Peripheral blood level of T-receptor excision rings (TREC) and B-cell excision rings (KREC), respectively, can serve as surrogate markers of T- and B-cell maturation. Currently, the diagnostic kits available on the market have two significant disadvantages: i) the kits are aimed at diagnosing immunodeficiency conditions only in newborns and children, while keeping adult patients uncovered; ii) essentially, use solely single reference normalization gene for data normalization resulting in increased variability and decreased sensitivity of the assay data. The aim: to develop a highly sensitive method for laboratory assessment of the state of immunity in immunodeficient patients by using real-time PCR for assessing TREC and KREC level in children and adults. Materials and methods. There were used whole blood and dry blood spot samples obtained from newborns and adults, apparently healthy individuals as well as patients with verified PID and HIV-infection. A total of 2577 samples were examined. Commercial kits were used as comparison methods. Results. Multiplex PCR was carried out, analyzing the number of target molecules TREC and KREC, as well as fragments of the HPRT and RPP30 normalization genes analyzed with the developed series of plasmid calibrators. The established analytical range of TREC/KREC DNA measurements comprised 103 to 109 copies/mL. The accuracy of measurements on a tablet-type instrument (CFX) was 95.84%, on a rotary-type instrument (Rotor-Gene 3000) 95.11%, which corresponds to the standard indicator. The equivalence between the data obtained after assessing whole blood samples and dry blood drops was shown. The data analysis allowed to find out 100%-diagnostic specificity and sensitivity of the method proposed. Conclusion. The method developed by us allows to diagnose decline in T- and/or B-cell immunity in children and adults and can be used to detect TREC and KREC molecules both in peripheral whole blood samples and dry blood spots using Guthrie cards. Moreover, the uniform values of reference norms can be used regardless of the type of analyzed clinical material. The study data evidence about potential for effective use of multiplex PCR diagnostics both for complex primary testing/screening of newborns and assessing state of immunity to identify adult patients with PID and as a part of the diagnostic monitoring of patients with secondary immunodeficiencies, e.g., HIV infection.

Primary immunodeficiencies (PID) are a heterogeneous group of hereditary diseases that lead to impaired immune defense. Often, the diagnosis cannot be made without identifying mutations that lead to the development of the disease. For many PIDs, there is no clear understanding of the etiology, pathogenesis, and genes involved. There is an obvious need to identify candidate genes potentially capable of leading to the development of PIDs.Hereditary angioedema (HAE) is a rare genetically determined disease, accompanied by recurrent edema of soft tissues and submucosal membranes, posing a threat to the life of patients. Diagnosis is based on the clinical presentation, family history, laboratory values of C1-esterase inhibitor, complement component 4, complement component 1q, antibodies to C1 and genetic testing for a number of mutations in the genes SERPING1, F12, PLG, ANGPT1, KNG1, MYOF, HS3ST6. However, pathogenesis may involve other genes in which the negative effect of mutations has not yet been studied. HAE is a non-monogenic disease that may involve an extensive network of genes. It seems important to determine the groups of the most probable candidate genes presumably involved in the development of pathology.Aim – to identify, using bioinformatics analysis, candidate genes for the development/pathogenesis of HAE and to reveal their biological context.The analysis was based on a group of genes, mutations in which are significantly associated with HAE: SERPING1, F12, PLG, ANGPT1, KNG1, MYOF, HS3ST6. To analised genetic and protein–protein networks and identify the biological context of the selected candidate genes, a number of web resources were used: HumanNetv3, GeneMania, FUMA GWAS in the GENE2FUNC mode.One hundred potential candidate genes in which mutations can be associated with HAE have been identified. The biological context of the identified genes was determined. The data of the biological context, genetic and protein-protein interactions made it possible to exclude a number of genes from the list of the most likely participants in pathogenesis and divide the remaining ones into groups with a greater or lesser potential for involvement. The group of the most likely HAO candidate genes includes PLAT, HRG, SERPINA1, SERPINF2, MASP2, GRB14, C1QBP, DOK2, KLKB1, F11, TEK, KLK10, KRT1, APOH, CPB2, F2.The results obtained can provide significant assistance in the study of the HAE molecular mechanism, as well as in the diagnosis and prognosis of the disease course. The identified candidate genes have the potential to serve as diagnostic biomarkers for patients with unexplained angioedema.The use of bioinformatic methods makes it possible to determine the list of candidate genes that are presumably involved in the disease pathogenesis or aggravate its course, and to obtain up-to-date information about the biological context of the identified genes. Understanding the genetic underpinnings and pathophysiology of PID may help define new diagnostic and therapeutic targets.

The aim of the study was to determine the tropism of the human immunodeficiency virus in patients with virological failure of antiretroviral therapy (ART) from the Arkhangelsk Region based on the analysis of the env gene V3 loop nucleotide sequence.Materials and methods. We used blood plasma samples obtained from 76 HIV-infected persons from the Arkhangelsk Region with virological failure of antiretroviral therapy. The nucleotide sequences of the HIV env gene C2-V3-C3 region were studied by PCR followed by sequencing. The genotype of the studied strains was determined based on the analysis of their phylogenetic relations with reference sequences from the international GenBank database, as well as using specialized programs. To predict viral tropism, the Garrido rule and the online bioinformatic tool Geno2Pheno[coreceptor] were used. The Geno2Pheno[coreceptor] algorithm, determines the false positive rate (FPR) based on the analysis of the env gene V3 loop nucleotide sequence. Results and discussion. Significantly lower representation of R5X4/X4-tropic HIV variants in long-term infected persons with subsubtype A6 virus compared to subtype B virus has been shown. For all FPR cut-off algorithms, a significant correlation between subtype and HIV tropism was observed (p=0.0014 and p=0.013 for FPR 10 % and FPR 20 %, respectively). While among subtype B strains, at least 57 % were identified as R5X4/X4-tropic variants (for an FPR of 10 %), including two strains classified as X4-tropic; among HIV subsubtype A6 even at an FPR of 20 %, the frequency of R5X4/X4-tropic samples only slightly exceeded 22 %. It can be assumed that the dynamics of changes in HIV tropism depends on the virus subtype. Significant differences in the distribution of amino acid residues of the V3 region sequences in the examined group between R5-tropic and R5X4/X4-tropic strains of subsubtype A6 for positions 18 (χ2=7.616, p=0.0058), 21 (χ2=7.281, p=0.007), 24 (χ2=5.587, p=0.0181), and 34 (χ2=5.144, p=0.0233) have been demonstrated. Among the R5X4/X4-tropic strains of the A6 subsubtype, amino acid substitutions were registered at positions 6, 19, 21, 26, 29, 30, which were not found in the R5-tropic A6 strains. The high occurrence frequency of a number of mutations previously described as presumably associated with resistance to maraviroc and similar drugs may indicate a natural polymorphism characteristic of the A6 subsubtype, which does not correlate with resistance to CCR5 co-receptor antagonists.

Introduction. The problem of transfusion safety in relation to parenteral viral hepatitis still remains relevant. Viral hepatitis B (HB) remains the most common viral infection transmitted through transfusion procedures. One of the natural phases of chronic hepatitis B (CHB) is occult hepatitis B infection (OBI), characterized by an undetectable HBsAg (regardless of the other serological markers content) in the presence of hepatitis B virus (HBV) DNA in the liver tissue and an extremely low, up to undetectable, level of viral load in the blood. In the Republic of Guinea, as in most countries on the continent, the prevention of HBV transmission through transfusion is still based on HBsAg serological testing of donors only. In this connection, OBI remains as a potential threat to blood transfusion safety. Detection of HBV DNA is a reliable preventive measure against transmission of the virus from donors with HBsAg-negative HBV infection, especially in highly endemic regions. In this regard, the study was conducted to substantiate recommendations for improving blood safety against the background of significant HBV prevalence in the Republic of Guinea. The aim of the work was the evaluation of serological and molecular markers of HBV infection in blood donors in the Republic of Guinea. Material and methods. We examined 250 blood samples obtained from donors living in Conakry, Republic of Guinea. Samples were tested for the presence of serological (surface antigen, HBsAg; antibodies (ABs) to surface (anti-HBs IgG) and core (anti-HBc IgG) antigens) and molecular (DNA) markers of HBV infection. Results and discussion. The overall detection rate of hepatitis B markers was 83.2%; HBsAg was detected in 16.4% of all individuals. The high incidence of HBsAg in men (19.55%) compared to women (8.45%) was shown, the relative risk of HBV infection with the formation of HBsAg-positive chronic hepatitis B in males was also significantly higher. The prevalence of the HBV DNA in the study group was 30.4%, the OBI cases accounted for 15.6%. The prevalence of this form of the disease was shown in donors aged 30–49 years (24.78%), in the group of people younger than 30 years, the incidence was lower (8.73%), and at the age of over 50 years, OBI was not detected. Based on the phylogenetic analysis of 76 virus isolates, it was shown that genotype E prevails in the examined group (85.53%). Cases of pathogen DNA detection occurred in HBsAg-negative blood donors in the presence of anti-HBs IgG (n = 4), as well as in the simultaneous presence of ABs anti-HBs IgG and anti-HBc IgG (n = 7). The viral load exceeded 200 IU/ml in OBI samples. Escape mutations were detected by sequencing in each OBI sample, contributing to the virus escaping from diagnostic based on screening for HBsAg. Conclusion. Assessment of the prevalence viral hepatitis B markers in blood donors, determination of genotypes and clinically significant mutations of virus variants are necessary to ensure safe medical manipulations, control and prevention of the spread of this infectious agent.

Introduction. The spread of the human immunodeficiency virus type 1 (HIV-1) has become a global concern and has approached the pandemic status. St. Petersburg, a major transportation, tourist, cultural, industrial center, and a border city, is characterized by high migration of the population. The growing number of migrants can contribute to importation and spread of new genetic variants of the virus and trigger recombination processes in the virus population in St. Petersburg and the Leningrad Region. The aim is to characterize the present-day HIV-1 subtype-specific profile and drug-resistance mutations among patients with virological failure on antiretroviral therapy (ART) in the Leningrad Region. Materials and methods. The study performed in 20162018 was based on clinical material from HIV-infected individuals living in the Leningrad Region and having confirmed virological failure on ART. The genetic diversity and distribution of drug-resistance mutations of the HIV-1 isolates were assessed through analysis of nucleotide sequences of the virus pol gene fragment that included regions encoding protease and the reverse transcriptase region. Results. In the group (n = 138), most of the patients had sub-subtype A6 (97.4%) common in Russia, though a few patients had subtype B and a recombinant containing circulating recombinant form CRF_03AB and sub-subtype A1. The tests showed that 95.79% of patients had at least one significant drug-resistance mutation; in most cases (73%) the virus was resistant to 2 classes of antiretroviral drugs and in some cases (8%) to 3 classes. A total of 105 different drug-resistance mutations were found at 35 positions of the virus genome. Conclusions. The high prevalence of HIV-1 drug-resistance mutations among ART patients with virological failure calls attention to surveillance of drug resistance of the virus both among ART-experienced patients and ARTnave individuals.

The aim of this study was to assess the prevalence of serological and molecular biological markers of viral hepatitis B and C among apparently healthy residents of the Southern Vietnam. The study material was represented by 397 blood serum samples collected from apparently healthy residents of the Southern Vietnam. The ELISA examination for presence of HBV and HCV markers involved HBsAg, anti-HBs IgG, anti-HBcore IgG, and anti-HCV qualitative determination. For HBV DNA and HCV RNA detection, nucleic acids were extracted from serum blood, and a test for virus detection was carried out by real-time PCR with hybridization fluorescence detection. Amplification and subsequent sequencing of HBV and HCV were performed using nested PCR with paired overlapping primers jointly flanking the target regions. Analysis of the overall prevalence of serological markers showed that among the apparently healthy individuals anti-HBsAg and anti-HCV antibodies were detected in 12.3% (95% CI: 9.27–15.99%) and 3.27% (95% CI: 1.76–5.53%) of individuals, respectively. The prevalence of HBsAg in men (19.1%) significantly exceeded that of found in women (5.9%), χ2 = 14.688 with p = 0.0001, df = 1, calculated odds ratio OR = 3.751 (95% CI: 1.892–7.439). Among apparently healthy patients, taking into account HBsAg-positive and negative samples, HBV DNA was detected in 26.95% (95% CI: 22.65–31.6%). HBV phylogenetic analysis showed that subtype B4 prevalence comprised 64.49%. Subtypes C1 (14.95%), B2 (9.35%), C2 (6.54%), C3 (0.93%), and C5 (3.74%) were also identified. HCV RNA was detected in 7 samples, which accounted for 1.76% (95% CI: 0.71–3.6%). Phylogenetic analysis showed that all HCV isolates belong to genotype 6, subtype 6a (100%).

The aim of this study was to characterize mutations in the hepatitis B virus (HBV) genome associated with HBsAg-negative form of the disease in patients receiving hemodialysis replacement therapy. Materials and methods. We used blood plasma samples obtained from hemodialysis centers in St. Petersburg, Russia – 173 patients and 108 patients from Belgrade, Republic of Serbia. The samples were examined for the presence of serological (HBsAg, antibodies anti-HBs IgG, anti-HBcore IgG) and molecular-genetic (HBV DNA) markers of HBV followed by whole-genome sequencing and determination of clinically significant virus mutations. Results and discussion. Antibodies to hepatitis B were detected in 7.5 % and 11.1 % of patients from St. Petersburg and Belgrade, respectively. HBsAg was identified only in 1.1 % of cases in the group from Russia and in 0.9 % of cases in the group from Serbia. HBV DNA was determined in 2.8 % of the studied samples from both, patients from Saint-Petersburg and Belgrade. Phylogenetic analysis of 9 viral isolates showed that genotype D virus (88.9 %) prevailed as compared to genotype A (11.1 %) in the examined group. Among the samples obtained from patients from St. Petersburg, four belonged to the D2 sub-genotype, one to the D3 genotype. Four samples obtained from Belgrade patients belonged to different sub-genotypes – D1, D2, D3, A2, respectively. When analyzing the nucleotide sequences of the HBV genomes, mutations in the MHR region were detected in all cases, but only in HBsAg-negative isolates, mutations were revealed in the region of 124–147 amino acids, including mutations P120T, R122K, A128V, Q129R, M133I, G145R affecting the recognition of HBsAg by anti-HBs antibodies and associated with the resistance of the virus to the vaccine. The results of this study indicate that transmission of blood-borne viral hepatitis agent in the hemodialysis departments of the Russian Federation and the Republic of Serbia still exists. The prevalence of the latent chronic hepatitis B, coupled with the presence of vaccine escape mutations in all identified cases, indicates the need to pay close attention to the occurrence of the virus mutant variants in hemodialysis centers.

Aim: To study the hepatitis A (HAV) and hepatitis Е (HEV) prevalence in the Southern region of Vietnam based on the frequency analysis of the antibodies to hepatitis A and E viruses detection in the local population and groups at increased risk of infection.Materials and methods. Serological markers of enteral viral hepatitis were determined in blood serum samples from adults aged 18 to 65 years of three groups: conditionally healthy individuals (n = 397), HIV-infected (n = 316), and patients with chronic viral hepatitis (n = 268). The ELISA method was used for the qualitative detection of anti-HAV IgG, anti-HAV IgM, anti-HEV IgG, anti-HEV IgM.Results. When analyzing the prevalence of anti-HAV IgG in samples obtained from conditionally healthy, HIV-infected, and patients with chronic viral hepatitis, no differences were found between the groups. The incidence of anti-HAV IgG in the general group (n = 981) was 80.1%, in the absence of anti-HAV IgM. There were no gender-age differences in the frequency of anti-HAV IgG in the examined groups. Antibodies anti-HEV IgG in the groups of conditionally healthy, patients with chronic viral hepatitis, and HIV-infected were present in the samples in 36,2%, 33,2%, and 39,8% of cases, respectively. The prevalence of anti-HEV IgM in these groups was 3,27%, 4,1%, and 3,79%, respectively. In the general group (n = 981), anti-HEV IgG was detected in 36,6% of cases, anti-HEV IgM in 3,66%, which corresponds to the prevalence of antibodies to HEV in endemic regions.Conclusion. A high incidence of enteral viral hepatitis markers was shown in residents of South Vietnam, including the groups of conditionally healthy, patients with chronic viral hepatitis, and HIV-infected. There is an obvious need for further studies of the spread extent of hepatitis A and hepatitis E in the Socialist Republic of Vietnam using currently available highly sensitive diagnostic methods, including sequencing of the virus›s nucleotide sequences.

Hepatitis B, despite of being a controlled infection today, is one of the most common form of hepatitis in the world. According to the experts' evaluation there are about 3 million patients with chronic hepatitis B in our country. The global strategy of the World Health Organization includes the elimination of viral hepatitis by 2030.The program used in the North-West Federal District to eliminate acute hepatitis B has reduced the incidence rate due to the widespread vaccination coverage of children and the annual increase in adult immunization coverage.However, the relevance of HBV-infection in children still remains high which is associated with a high infection by hepatitis B virus in women of childbearing age and the possibility of the transmission of the infection from mother to her child. In case of perinatal infection the formation of chronic hepatitis B in children reaches up to 90%. The natural course of chronic hepatitis B is characterized by a change in pathogenetically determined phases, and HBsAg-negative infection, which is a latent (occult) form of hepatitis, was added to them in 2009. Occult hepatitis B is an epidemiological danger, for the child as well, if the mother suffers from this form of chronic hepatitis B. Monitoring of pregnant women is often limited to identifying only HBsAg, which is not enough to detect occult hepatitis B. Lately diagnosed occult HBV-infection can become a source of the infection for the baby, especially in cases when due to some reasons there are disorders in hepatitis B vaccination schedule after birth. The article presents an interesting clinical case of family hepatitis B.

According to the WHO, there is an increase in the number of cases of dengue fever worldwide. In many countries, where dengue fever is an endemic disease, blood-borne infections associated with hepatitis B and C viruses and HIV are widespread. The Socialist Republic of Vietnam is an endemic region for these pathogens. The unique epidemiological situation in the country provides an excellent opportunity to study the interaction between Arboviruses, agents of parenteral viral hepatitis B and C, and HIV infection in the body of sick people.The aim of this review was to analyze the literature data on the detection of cases of simultaneous infection with Dengue viruses of 1–4 types and agents of blood-borne infections in sick people in the Socialist Republic of Vietnam. Despite the fact that the simultaneous circulation of these pathogens in the patient's body can affect the pathophysiological mechanisms of the disease development, there were very few works devoted to co-infection with Dengue viruses and HIV, hepatitis C or B viruses, including in the regions adjacent to Vietnam. Therefore, research in this direction is promising for both fundamental science and practical medicine.

Rationale. Countries of Africa, especially countries in sub-Saharan Africa, represent a region characterized by high incidence of chronic hepatitis B virus (HBV) and hepatitis C virus (HCV) infections. Methods for detection of HBV and HCV in low and middle-income countries differ from those that are used in countries having access to high-cost technologies. The Republic of Guinea is a region with high prevalence of hepatotropic viruses, however, the information on HBV and HCV prevalence in the area is extremely limited, thus emphasizing the significance of this study.The purpose of the study is to evaluate the need for improving laboratory diagnostics of parenteral HBV and HCV infections in the Republic of Guinea.Materials and methods. A total of 2,616 samples of blood serum were tested, the samples were collected from apparently healthy residents of the Republic of Guinea during the routine medical checkup. The testing included qualitative detection of HBsAg, anti-HBs IgG, anti-HBcore IgG, anti-HCV IgG antibodies as well as HBV DNA and HCV RNA.Results. The detection frequency of serological markers of HBV and HCV infections was 80.77% and 18%, respectively. However, HBsAg was detected only in 16.01% of individuals. Tests for detection of HBV DNA were performed among seropositive patients and patients seronegative by other HBV markers, HBV DNA was detected in 22.36% of cases, including 6.07% of HBsAg-negative cases. HCV RNA was detected in 2.2% of cases. Both HCV RNA and HBV DNA were detected in 27 people, including 19 HBsAg-negative cases, thus accounting for 1.03% of the examined group.Conclusions. The markers that are currently used for laboratory detection of HBV and HCV in the Republic of Guinea are not efficient enough to diagnose reliably all cases. Undoubtedly, there is an urgent need to improve laboratory diagnostics for timely detection of parenteral viral hepatitis. Routine laboratory operations need assays for additional serological and molecular markers of HCV and HBV infections.

The aim of our work was to compare the HBV, HCV and HIV vertical transmission risk in the Republic of Guinea.Materials and methods. The material for the study was 305 blood plasma samples from pregnant women living in Conakry, Republic of Guinea. The samples were examined for the presence of serological (HBsAg, antibodies antiHBs IgG, anti-HBcore IgG, anti-HCV IgG, Ag/Ab-HIV) and molecular (HBV DNA, HCV RNA, HIV RNA) markers.Results and discussion. When assessing the overall prevalence of serological markers among patients, the incidence of HBV markers was 76.06 %. Antibodies to HCV were detected only in 1 case, which amounted to 0.32 %. HIV markers were detected in 3 cases, which amounted to 0.98 %. The prevalence of HBsAg in the group under examination significantly differed between the groups of pregnant women aged 13–19 years (17.33 %) and 20–24 years (12.12 %), p

Background. The development of direct acting antivirals (DAAs) has spurred a revolution in treatment of patients with chronic hepatitis C. However, there are cases showing no response to treatment. In 5% of cases, the viral breakthrough is most likely caused by DAA resistance mutations in the hepatitis C virus genome.The purpose of the study is to detect drug resistance mutations of hepatitis C virus in patients with DAA treatment failure.Materials and methods. The study was performed on plasma samples from 3 patients diagnosed with chronic hepatitis C virus infection and demonstrating DAA virological treatment failure. All isolates had genotype 1b. Drug resistance mutations were detected by using direct sequencing of NS3, NS5A, and NS5B genome regions. The detection technique was developed at the Pasteur Research Institute of Epidemiology and Microbiology.Results. Drug resistance mutations were detected in all cases. By using the Geno2pheno [hcv] 0.92 tool, nucleotide substitutions were detected in different viral genome regions and presumably caused resistance or decreased sensitivity to antivirals both present and absent in the sofosbuvir + daclatasvir combination therapy. Antiviral treatment failure in patients with chronic hepatitis C is caused by drug resistance mutations.Conclusions. The developed technique is efficient for detection of drug resistance mutations in NS3, NS5A, and NS5B regions in cases of virological failure of DAA treatment.

Objective of our work was to assess prevalence of the primary HCV drug resistance mutations in the NS5b gene in patients with newly diagnosed HIV infection.Materials and methods. The study material was 196 blood plasma samples from patients living in the North-Western Federal District with newly diagnosed HIV. Samples were examined for the anti-HCV antibodies and HCV RNA presence. If HCV RNA was detected, amplification was performed using three primers pairs that co-flanked the NS5b gene. After sequencing the indicated gene nucleotide sequence, the virus subtype was determined and drug resistance mutations were detected.Results and discussion. Antibodies to HCV were detected in 18.87 % of HIV-infected individuals. HCV RNA was detected in 18.36 % of the patients, including 89.18 % anti-HCV-positive and 1.88 % anti-HCV-negative. It was shown that co-infection is more common in men (77.8 %) compared to women (22.2 %) – χ2 = 3.996 at p = 0.0456, df = 2. The difference in the HIV viral load between the groups with HIV monoinfection and with HIV + HCV coinfection was demonstrated (χ2 = 6.284 at p = 0.0432, df = 2). A significant difference between the groups by the CD4 + lyphocytes number was shown. In the phylogenetic analysis, the HCV subtypes are distributed as follows: HCV 1b – 47.2 %, HCV 3a – 30.6 %, HCV 1a – 13.9 %, HCV 2a – 5.5 % and only one sample was defined as HCV 2k – 2.8 %, respectively. Nine samples (25 %) presented NS5b mutations in the positions related to the development of drug resistance of HCV, including two samples among HCV genotypes 1a and 3a (i.e., 5.6 % of the total HIV + HCV group), as well as five samples among HCV 1b (13.9 % of the total group). Mutations among HCV 1a were C316Y and N444D substitutions. Among HCV 1b, C316N, C451S, S556N/G substitutions were identified. Among patients with HCV 3a, 2 samples (5.6 %) with a D310N mutation associated with an unfavorable disease prognosis were found. The introduction of direct sequencing of HCV nucleotide sequences into the routine laboratory diagnostics will allow us to estimate the primary drug resistance mutations prevalence in risk groups to predict the HCV life-threatening complications development – fibrosis, cirrhosis, hepatocellular carcinoma, as well as the outcome of antiviral therapy prognosis. The data obtained can be rationally used to assess the dynamics of the HCV primary pharmacoresistance prevalence among HIV-infected individuals.

Currently, along with the increasing need of medical organizations for blood preparations, algorithms for laboratory testing of blood donors are not available for all infections with hemo-contact mechanism of transmission. A representative example is infection caused by parvovirus В19. Purpose of the study. The article presents the results of the original study, the purpose of which was to study the prevalence of antibodies to parvovirus B19 and the activity of the circulation of this virus in socially important categories of the population. Material and methods. The materials of the study were blood samples from blood donors of Saint Petersburg, as well as parvovirus В19 sequences isolated from DNA-positive plasma samples. Results and discussion. According to the results of the laboratory examination, a high proportion of carriers of virus-specific IgG antibodies was found in studied group of donors, which confirms the previous infection of parvovirus B19 in them and illustrates the high prevalence of infection in this socially significant group. Based on the results of the blood preparations testing, the presence of parvovirus DNA В19 in a significant number of samples was determined by polymerase chain reaction method. This indicates an current parvovirus infection in the examined donors and points to a high epidemiological risk of the blood products obtained from them. Sequencing and phylogenetic analysis of a fragment of the VP1 gene demonstrated that the studied isolates belonged to А1 genotype and its subtype 1А2, which correlates with the genotypes of parvovirus В19 circulating in the European Union and Asia. In addition, two previously unknown В19 parvovirus isolates were isolated, the nucleotide sequences of which were deposited into the international GenBank database. Conclusion. Based on the results of the study, it is justified to include testing of blood samples for markers of В19 parvovirus infection in existing algorithms of laboratory examination of donors, which will ensure prevention of hemo-contact infection of blood recipients with parvovirus В19.

Objective. The purpose of our work was molecular genetic characterization of the hepatitis D virus isolates, circulating in the region with high prevalence of HBV + HDV super-infection. Materials and methods. The study material was 64 blood serum samples obtained from Kyrgyz Republic residents - patients with chronic viral hepatitis B+D. The hepatitis D virus complete genomes were sequenced, followed by phylogenetic analysis. Results and discussion. Based on the phylogenetic analysis of 64 HDV samples, it was shown that HDV genotype 1 (96.9 %) predominates in the examined group compared with HDV genotype 2 (3.1 %). Sequences were submitted to GenBank under access No MN984407 through MN984470. When assessing the genetic variability over the examined HDV genotype 1 samples, the maximum genetic distance was 12,49 %, and the minimum – 7,41 %. Within individual clusters, the genetic distance averaged from 2.6 % to 8.5 %. Among the sequences in GenBank, the closest resemblance to the HDV-2 Kyr41 and Kyr43 samples (nucleotide identity was 92.31 % and 89.57 %, respectively) was shown for the virus described earlier in Yakutia (AJ309880). To study the genetic relationships between the analyzed HDV genotype 1 strains in comparison with the HDV reference sequences, the predicted amino acid sequence was studied (111–214). Although hepatitis B preventive measures, including vaccination, have reduced the hepatitis D incidence, there is no effective way to prevent HDV infection in HBV carriers in endemic areas. The HDV sequence molecular-genetic characterization in this study, as well as the viral genomic sequence phylogenetic analysis, will help identify pathogen transmission pathways to control and / or prevent the spread of infection.

Aim. To estimate the distribution of genotypes and subgenotypes of the hepatitis B virus among military personnel with chronic viral hepatitis B. Materials and methods. The work used samples of blood plasma and biopsy material obtained from 90 active or retired military personnel with chronic viral hepatitis B with various degrees of fibrosis undergoing treatment in St. Petersburg. Primary detection of HBV was carried out by isolating nucleic acids (NK) from the blood plasma using the «AmplePrime Ribo-prep» commercial kit (FBIS CRIE, Moscow). Specific primers were used for the amplification and sequencing reaction. Overlapping primer pairs were used, jointly flanking 1475 base pairs (bp) fragment, including the recommended for HBV genotyping the 1169 bp Pre-S1/Pre-S2/S. Results. Among 90 samples from patients with chronic viral hepatitis B from different regions of the Russian Federation, HBV subgenotypes are represented in the following ratios: D2 = 45.6% (n=41), D1 = 32.2% (n=29), D3 = 13.3% (n=12), A2 = 6.7% (n=6), D4 and A1 by 1.1%, respectively. The distribution of HBV subgenotypes from the North Caucasian federal district (D1 – 63.6%, D2, D3, D4, A2 – by 9.1%) was significantly different from the distribution among patients from the Central and North-Western federal districts (D1-20, 9%, D2 – 58%, D3 – 16.3%, A2 – 4.8%) (χ2=11,9 при p=0,0076, df=3). Uncharacteristic for the Russian Federation subgenotypes D4 and A1, representing single imported cases. The tendency to shift the distribution of genovariants due to imports of the corresponding HBV subgenotypes from other countries, including the Central Asian countries, is discussed. Conclusion. A systematic study of the HBV isolates phylogeny provides new information about the HBV subgenotypes distribution among certain population groups, including military personnel.

Aim. Molecular genetic characterization of hepatitis B virus and human immunodeficiency virus in patients with HIV / HBV co-infection living in the Republic of Guinea. Materials and methods. 2168 blood serum samples obtained from the Republic of Guinea residents – blood donors and conditionally healthy people, without suspicion of Ebola virus disease, UK RUSAL employees and their families, as part of their routine medical examination. The presence of serological and molecular biological markers of HIV and HBV was examined. When HIV/HBV co-infection was detected, the nucleotide sequences of the complete HBV genomes and the HIV pol gene fragment were sequenced. Results and discussion. HIVserological markers were detected in 239 people (11.02 %). HIV RNA was detected in 31 people, which accounted for 12.9 % of patients in the seropositive group (1.43 % of the total group). HBV serological markers among HIV RNAs-positive individuals were detected in 29.03 % of patients, including 16.12 % HBsAg and 12.9 % anti-HBcore IgG. HBV DNA was detected in all HBsAg-positive and in two anti-HBcore IgG-positive patients, as well as in 12 people negative for all HBV serological markers analyzed in the work. Thus, HBV DNA was found in 61.29 % of HIV RNA-positive individuals. Based on the pol gene fragment nucleotide sequences analysis of 19 HIV samples, it was shown that the HIV circulating recombinant form CRF02_AG prevails in the examined group (52.63 %) compared with HIV A1 (42.1 %), one sample was an independent recombinant of genotypes A1 and G. HBV phylogenetic analysis of the studied samples showed that genotype E prevails – 47.36 %, compared with HBV D1 – 21.05 %, D2 – 15.78 %, D3 – 10.52 % and A2 – 5.26 %. HIV and HBV samples have been detected that carry drug resistance mutations despite the antiretroviral therapy absence. HIV and HBV drug resistance mutations identification in ART-naive patients emphasizes the need for HIV surveillance programs as well as routine testing for HBV and HIV and HBV drug resistance before starting antiretroviral therapy in the clinical management of patients in the country.

Background and aims: to estimate concentration of sMadCAM-1 in peripheral blood at patients with chronic hepatitis C with excess body weight.Materials and methods: The research included 88 patients (67 men, 21 women 41.4±3.2 years of age) with chronic hepatitis C (CHC) and excess body weight (the index of body mass is ³25 kg/m2, and abdominal circumference more than 94 cm in men, and 80 cm in women) with various morfofunktsionalny changes in a liver and a small bowel. From them men there were 67 people, women – 21, middle age was 41.4±3.2 years.To all the patients complex clinical, biochemical, virologic, morphological trial was carried out. The functional condition of intestines was estimated by identification of a small intestinal bacterial overgrowth (SIBO) when carrying out the hydrogen respiratory test (HRT) with lactulose and existence of endoscopic signs of inflammation of a mucous membrane of intestines at a fibroezofagogastroduodenoskopiya. The quantitative assessment of a mucosal addressin cell adhesion molecule -1 was carried out by the definition concentration of its soluble form (sMadCAM-1) in a blood plasma by enzyme immunoassay method.Results: the sMadCAM-1 level of peripheral blood at the patients with excess body weight increased in process of progressing of a stage of chronic hepatitis C (F0 – 349.10 (324.27-373.92) ng/ml; F1/2 – 439.69 (406.43-472.94) ng/ml; F3/4 – 1057.82 (593.38-1522.26) ng/ml; p˂0.05), existence of a syndrome of excess bacterial growth and endoscopic signsof a duodenitis. Besides, patients had its concentration more with the biochemical signs characterizing cytolytic (at ALT˃N: 502.54 (432.04-573.03) ng/ml against 381.04(345.49-416.58) at the ALT normal values), cholestatic (at GGTP˃N: 550.59 (453.31-647.88) ng/ml against 400.86(365.13-436.59) atnormal GGTP, p values 0.05; at ALP N: 572.2 (353.7-790.8) ng/ml against 468.7 (408.5-528.9) ng/ml at normal ALP, p values 0.05) and metabolic syndromes (at glucose of blood, TG, VLDL N: 562.93 (369.59-756.27) ng/ml, 681.15 (387.81-974.49) ng/ml, 809.65(124.04-1495.28) against (438.34(391.36-485.31) ng/ml), (421.69(379.41-463.97) ng/ml), 434.47(389.45-479.48), p values 0.05 at normal values of these indicators respectively). Conclusion: Progressing of fibrosis and functional disturbances in intestines are interconnected with increase in concentration of MadCAM-1 in blood that allows to consider pathological changes in intestines of various genesis as the accessory factor promoting progressing of СHC at patients with excess body weight. Besides, definition of concentration of sMadCAM-1 in peripheral blood can be used as one of markers of noninvasive diagnostics of a stage of fibrosis at the patients with СHC and excess body weight.

Aim. Genotyping and molecular genetic characteristics of parvovirus B19, circulating in the NorthWest Federal District. Material and Methods. The material of the study is based on serum samples from 821 patients with maculopapular rash negative for antibodies IgM-measles and IgM-rubella were received at the St. Petersburg Regional Center for Measles and Rubella Surveillance in 2009-2017. In the present study we used genotyping by direct sequencing of the NS1/VP1 region of Parvovirus B19 genome. Results. DNA of the virus was detected in 59 (42.4%) of seropositive patients. To assess the heterogeneity of isolates, 14 samples were selected from patients from geographically remote regions of the NWFO, of different ages, regardless of gender, with a high viral load (106-107 copies/ml). Based on the phylogenetic analysis of the isolates showed that only the genotype 1A was detected in all isolates. The nucleotide sequences can be divided into two subgroups: 13 isolates (92.8%) belong to the subgroup 1A2, one isolate to the subgroup 1A1. Conclusion. The introduction of B19 screening from patients with fever / rash can provide meaningful information on the prevalence of parvovirus infection in the Russian Federation. Identifying new mutations of the virus and further analysis of their possible relationships with the course of the mutation disease can help in the development of medicines, as well as in the development of an effective vaccine against the parvovirus B19.

Rationale: Enteric hepatitis including hepatitis A and hepatitis E are globally prevalent diseases with large social and economic burden. In some patient categories these may be lethal. Hepatitis A (HAV) and hepatitis E (HEV) viruses are heterogeneous and have high geographic variability.Aim: To assess current epidemiological, molecular and genetic characteristics of enteric viral hepatitis in the Russian Federation.Materials and methods: We analyzed the data of the State statistical reports on the incidence of infectious diseases in the Russian Federation and of the analytical tables. Also, genotype identification of the HAVs isolated from 31 patients living in St. Petersburg was performed with specific primers flanking VP1/2A region.Results: In the last 20 years (from 1997 to 2017), the epidemic process of hepatitis A in the Russian Federation has been changing substantially, including first of all its intensity and age distribution of the patients. Unlike hepatitis A, the official registration of hepatitis E in the Russian Federation was initiated only in 2013. During the analyzed time period, the incidence of hepatitis E varied from 0.06 to 0.08⁰⁄₀₀₀₀, with non-significant trend towards its increase in 2017. It has been found that from 2013 to 2015, two sub-genotypes of the virus (1a and 1b) circulated in St. Petersburg, with prevailing genotype 1a. The local cases of hepatitis E were caused by HEV genotype 3, whereas the imported cases, by HEV genotype 1.Conclusion: In the Russian Federation, there has been a long-standing and stable trend toward decreased incidence of hepatitis A. However this has led to weakening of collective immunity, mainly among the adult population. In combination with poor levels of municipal infrastructures and services in some territories, this may lead to an increase of the hepatitis A incidence. Despite that Russia is a non-endemic region for hepatitis E, local cases of the disease are increasingly identified.

The prevalence of HBV infection is estimated by the frequency of occurrence of HBsAg and varies depending on the geographic region. Chronic infection is characterized by a stable presence of HBsAg for 6 months, with the exception of the occult form of the disease, characterized by the absence of HBsAg, an extremely low level of HBV DNA in the blood serum. The problem of identifying occult HBV (ocHBV) is especially relevant because of the development of transplantology and transfusiology. However, serological screening of donor blood used in the Russian Federation and Central Asian countries does not reveal HBV seronegative donors. Since HBV infection is possible with the introduction of small doses of the virus, the importance of using complex molecular methods for detecting donor ocHBV is obvious, despite the low viral load, since donor blood is used predominantly in patients with severe course of various diseases characterized by increased susceptibility to HBV because of immunosuppression. The aim of our work was to study the characteristics of the genetic structure of the ocHBV in donors in Astana, Kazakhstan. A total of 500 blood plasma samples from HBsAg-negative donors were obtained in 2012 from residents of Kazakhstan, Astana. Using the method, we proposed to detect HBV DNA with a low viral load, HBV was detected in 9.4% of donors. Serological markers were found in 12.7% of patients with HBV DNA, 8.5% had HBcor IgG antibodies, 4.2% had HBcor IgG and HBe IgG antibodies at the same time. Thus, in 41 (87.3%) of the blood donor, ocHBV was seronegative. Based on the phylogenetic analysis of the 47 isolates showed that the HBV of genotype D (95.75%) prevails in the examined group in comparison with HBV of genotype A (4.25%). HBV subgenotypes are represented in the following ratios: D1 — 46.8%, D2 — 17.05%, D3 — 31.9%, A2 — 4.25%. In a comparative analysis, the distribution of HBV subgenotypes in the group with ocHBV and in the case of the manifest form in donors in the Republic of Kazakhstan significantly differed — χ2 = 14.027 at p = 0.0072, df = 4. The incidence of HBV D3 with ocHBV (31.9%) exceeded that of patients with a manifest form (7.4%). The relative risk of occult form of disease in patients with the subgenotype D3 is significantly higher (RR = 1.572, CI: 1.179–2.096, p = 0.0208). When assessing the picture of HBV diversity on the material of the group including HBsAg-negative and HBsAg-positive blood donors, it is evident that the genetic relationship of the manifestations of the manifest HBV and the ocHBV of genotype D is obvious. Among the isolates are both similar in nucleotide sequences with those previously described in various regions of Europe and Central Asia, and circulating in the territory of the Republic of Kazakhstan, which indicates an independent homologous evolution of HBV in the region. The high incidence of ocHBV among HBsAg-negative blood donors is indicative not only of the widespread prevalence of the occult form of the disease course in the population and the inadequacy for the detection of chronic HBV of conventional HBsAg and HBV DNA in the peripheral blood using commercial kits, but also the need to study the characteristics of the immune response with this form of the disease course.

Aim. Epidemiologic examination of the case of group HIV-1 infection based on data obtained by general diagnostics methods for confirmation of the investigation hypothesis of deliberate HIV infection during heterosexual intercourse. Materials and methods. Sera samples from 8 HIV infected patients (5 female and 3 male) from Veliky Novgorod sent for epidemiologic investigation were used. Determination was carried out based on 1285 nt sequence analysis of polymerase gene segment (pol). Results. The study allowed to identify HIV in clinical samples sent to the expertise and establish phylogenetic connections between virus isolates obtained from both target and control group patients. Analysis of the results allowed to isolate samples grouped in a separate cluster that indicates tight cordial connections between the vims isolates from clinical material of these patients. Patients of the target group were infected with HIV-1 isolate ofthe circulating recombinant form CRF03_AB from the same origin that is confirmed by high homology ofthe nucleotide sequences. Conclusion. Epidemiologic investigation of a group case of HIV-1 infection has determined that the infection of the women of the target group occurred from the same source. Phylogenetic analysis results indicate the presence of an epidemiologic connection within the examined group that confirms the HIV infection transmission and conclusions ofthe investigation. Use of molecular-phylogenetic analysis of data obtained by laboratory diagnostics methods of HIV resistance to antiretroviral preparations allows with anamnestic and investigation information (in the context of available evidence) to investigate cases of HIV infection.

Aim. Study cases virologic failure ART HIV-1 in children.Materials and methods. The blood plasma samples of 6 patients with HIV infection were used, identified under 1 year of life, from the North-West Federal District of the Russian Federation, received in 2014–2016. Presented in a group of patients were aimed at identifying drug resistance in the virus and correction of ART due to virological failure. In the present study we used genotyping by direct sequencing of the site of the polymerase gene (pol) length of 1285 nt., The gene encoding the protease (PR) length of 465 nt. and a portion of the reverse transcriptase (RT) gene length of 820 nt.Results. In all cases, I was diagnosed with HIV-1 subtype A1, the so-called IDU-A, which is the most common form of HIV-1 in the Russian Federation. Among the obtained isolates of HIV have been identified typical drug resistance mutations to NRTI and NNRTI. Detected multiple natural polymorphic variants from the land of the nucleotide sequences. Thus, all samples show protease mutation M36I, R41K, H69K, and L89M, five samples E35D, four samples I13V. Only one among the surveyed our children with combined variant protease gene mutations L63T + V77I + I93L and reverse transcriptase gene V35I + K166R, despite ongoing therapy and absence of significant drug resistance mutations, according to Stanford University-based data showed high viral load – 2.03×107 copies/ml. The importance of the influence of natural polymorphisms in the drug resistance of the virus is discussed.Conclusion. The high frequency of natural polymorphic variants of the protease and reverse transcriptase genes of the virus in clinical samples submitted. The number of such mutations, apparently does not depend on the number of treatment regimens, but found some correlation with adherence to treatment.

The prevalence of one of the hepatotropic virus, hepatitis B virus (HBV) remains a serious global health problem. Since hepatitis B is transmitted through contact with blood or other fluids of an infected person, blood safety is one of the major public health issues in regions with high virus prevalence. Observed in recent years the trend to a shift in the prevalence of various genotypes of HBV in different geographical areas due to immigration from regions of the world with a high incidence of hepatotropic viruses, makes doctors and epidemiologists to pay close attention to the epidemiological situation in neighboring countries. The aim of our work was to study the characteristics of the genetic structure of the HBV in primary donors in Astana, Kazakhstan. A total of 30 blood plasma samples from newly diagnosed hepatitis B (HBsAg+) of Astana. HBV DNA was detected in 27 samples out of 30. Based on the phylogenetic analysis of the isolates showed that among patients examined HBV identified mainly D genotype, which is the most common genotype of HBV in Central Asia. Thus HBV subtype predominant D1 (85,2%) compared to the HBV subtype D2 (3,7%) and subtype D3 (7,4%), in a single sample was detected HBV genotypes A subtype A1. High similarities identified isolates previously described in Iran, Sudan, Mongolia, Tunisia suggest numerous independent, perhaps mutual, the importation of the virus in the country, including in the major migration waves. First detected at the territory of Kazakhstan HBV subtype A1, uncharacteristic for the region, as well as subtypes D2 and D3, which have a high similarity with the nucleotide sequences of HBV in Russia, show cases of importation of the virus from other countries. Identification of the propagation and the role of «imported» genotypes of HBV virus in circulation may be essential for regions where the prevalence of hepatotropic viruses is high, and the genome structure and the way of their distribution sufficiently studied.

Aim. Analysis of phylogenetic relationships of HIV isolates obtained from a child 8 years old and HIV-positive parents to search for a possible source of infection.Materials and methods. The blood plasma samples of 3 patients (father, mother and child) HIV from Veliky Novgorod were used. Presented in a group of patients were directed to conduct epidemiological investigation cases of HIV-1 infection a child from a dysfunctional family. In the present study we used genotyping by direct sequencing of the site of the polymerase gene (pol) length of 1285 nt., The gene encoding the protease (PR) length of 465 nt. and a portion of the reverse transcriptase (RT) gene length of 820 nt.Results. The study allowed the identification of the HIV virus in clinical samples. HIV-1 subtype A1 (IDU-A) was detected in all cases. Phylogenetic analysis of isolates, where the control samples using HIV-1 isolates obtained previously from Veliky Novgorod, possible to identify grouped in a separate cluster sample from the mother, father and child, which makes it possible to conclude about intrafamily HIV infected child by one of his parents. The nucleotide identity of the samples obtained from the mother and the child, showed a higher percentage of similarity – 98.4%, compared with the identity of the samples between the father and the mother and between father and child (96.2% and 94.2%, respectively). Differences natural polymorphisms in protease and reverse transcriptase genes of the parents and the child are discussed. Conclusion. The analysis of phylogenetic relationships of HIV isolates showed that the source of infection of the child 8 years old, born and living in socially disadvantaged families with HIV-positive parents, is his mother. Way parenteral transmission of infection by random simultaneous trauma in the mother and child.Molecular phylogenetic analysis using routine methods for determining the resistance of HIV to antiretroviral drugs allows to investigate cases of HIV infection, identifying the source of infection.

Aim. Evaluate prevalence of genetic variants of hepatitis В viruses in population of various regions of Uzbekistan with hepatitis of various genesis and different severity levels of liver fibrosis and cirrhosis. Materials and methods. Blood plasma and liver biopsy from 39 patients with different severity levels of liver fibrosis and cirrhosis served as study material. Genotyping based on direct sequencing of Pre-Sl/Pre-S2/S HBV DNA region was applied. Results. Hepatitis В virus was detected in 32 samples ofthe 39 provided, frequency of occurrence - 82%, respectively. Phylogenetic analysis has shown, that only genotype D was detected among the examined patients, hepatitis В virus subtype D1 predominated (84.38%) compared with D2 (3.12%) and D3 (12.5%) subtypes. Conclusion. Large-scale sequencing of HBV in Central Asia will allow to evaluate routes of transmission and time of evolutionary separation of virus isolates. Understanding the epidemiology of the infectious process is important for development of programs for prophylaxis and therapy of the infection.

Aim. Evaluate significance of covalently closed circular DNA of hepatitis В virus as a marker for detection of occult viral hepatitis В in Uzbekistan population with hepatitis of various genesis. Materials and methods. Blood plasma and liver biopsy from 39 patients with different severity levels of liver fibrosis and cirrhosis served as study material. HBV covalently closed circular DNA detection was carried out according to Pollicino T. et al. (2004). Results. Covalently closed circular DNA of hepatitis В virus was detected in 82% of samples, including in 54.5% of patients with chronic viral hepatitis C (CVHC) and in 100% of patients with hepatitis of unknown etiology. Quantitative evaluation of content of covalently closed circular DNA of hepatitis В virus in liver tissue in patients with CVHB has shown an average of 2.5 copies of HBV genome as ccc DNA per cell, in patients with CVHB + D an average of 0.7 copies/cell, in patients with co-infection by HCV and HBV - 0.5 copies/cell, in patients with CVHC an average of 0.12 copies/cell, and in patients with cryptogenic hepatitis - 0.2 copies/cell. Conclusion. Detection of HBV DNA is a complex problem for effective laboratory diagnostics of hepatitis. Detection of H BV ccc DNA as a marker of occult hepatitis В in patients with CVHC and patients with hepatitis of unclear etiology is an important factor for diagnostics, selection of adequate therapy, prognosis of disease outcome and prevention of development of severe liver diseases.

One of the most serious health problems in the world are hepatotropic viruses that cause chronic liver disease. Hepatitis B virus is distributed globally; around 5% of the carriers are also infected with hepatitis delta virus. Co-infection or superinfection of hepatitis viruses B and D significantly associated with a much more severe liver disease, compared with infection only hepatitis B virus. However, examination of hepatitis virus B carriers for the presence of hepatitis D virus in most regions of the world is not mandatory. It should be noted that the complete genotype mapping of viruses hepatitis B and D isolated on the territory of the CIS and the countries of the former Soviet Union, there is not yet, despite the constantly ongoing works devoted genotyping hepatotropic virus in the territory of the Russian Federation and neighboring countries. Due to the fact that one of the prospective ways of spreading viruses is the “labor migration” the inhabitants of Central Asia in other countries, including the Russian Federation, there is a need to pay attention to the situation of viral hepatitis in the region. The aim of our study was to estimate the prevalence of genetic variants and characteristics of molecular epidemiology of chronic viral hepatitis co-infection B + D in Kyrgyzstan. The study involved 30 plasma samples from patients with chronic viral hepatitis B and D from different regions of Kyrgyzstan. Based on the phylogenetic analysis of the isolates showed that among patients examined HBV identified only D genotype. Based on the phylogenetic analysis of the isolates indicated that among the examined patients with chronic viral hepatitis B revealed only genotype D. It is shown prevalence of HBV subtype D1 (73.34%) compared to the HBV subtype D2 (3.33%) and D3 (23.33%). Revealed HDV genotype I with highly variable region of the gene encoding the delta antigen. The high similarity of some isolates with strains specific to neighboring countries endemic for hepatotropic viruses, as well as a dense clustering of other isolates may be an indication of numerous independent drifts of strains into the territory of the country. Also it can talk about the speed of evolution of the virus in a geographically isolated region as Kyrgyzstan. Identification of the propagation characteristics and endemics role in circulation of genotype of hepatitis B and D is great importance.

Objective: To estimate the prevalence of genetic variants and features of the molecular epidemiology of HBV in Yakutia residents suffering from HBV.Materials and methods: The study involved 35 patients with chronic hepatitis B from urban and rural areas of Yakutia while most of the group were representatives of the autochthonous population. In the present study we used genotyping by direct sequencing of the Pre-S1 / Pre-S2 / S region of HBV DNA.Results: Based on the phylogenetic analysis of the isolates showed that among patients examined HBV identified only D genotype, which is the most common genotype of HBV in the Russian Federation. It is shown prevalence of HBV subtype D2 (85,8%) compared to the HBV subtype D3 (14,2%).Conclusion: We identified clearly clustered group of HBV isolates and close ties within the group, which suggests the existence of at least four permanent sources of infection, acting for a few years and decades. The systematic application of complex molecular, virological, epidemiological methods and molecular phylogenetics could contribute to the current understanding of the epidemiology of HBV and improve the quality of the traditional methods of supervision in Russia.

Objective: To evaluate relationships between virological and morphological data in patients with chronic hepatitis B in phase immune control and reactivation.Materials and methods: The study involved 46 patients with chronic hepatitis B, indicators defined were content of surface antigen and hepatitis B virus DNA in the peripheral blood, the level of covalently closed circular HBV DNA in liver tissue and fibrosis stage and histological activity index (METAVIR).Results: The study found a direct correlation between the level of covalently closed circular DNA in liver puncture biopsies (number of copies per cell) and quantitative content of HBsAg in serum (r = 0,51, p = 0,03) in patients with chronic hepatitis B in phase immune control. Also in the immune control phase a direct correlation between the HBV DNA and the level of HBsAg in serum (r = 0.79, p = 0.0001) is shown. The level of covalently closed circular HBV DNA in liver puncture biopsy specimens did not differ in patients with chronic hepatitis B in the phase of immune control in the phase of reactivation (1,02 ± 0,01 copies / cell and 1,03 ± 0,03 copies / cell, p = 0.72). The index of histological activity and fibrosis were not correlated with any of the investigated virological indicator.Conclusion: Results of the study emphasized the complexity and ambiguity of the relationships between both virological and morphological indicators in patients with chronic hepatitis B, determined the direction for further study (in particular the assessment of the epigenetic regulation of the synthesis of circular covalently closed DNA), as well as set the stage for improving the principles of dynamic observing the patients with chronic hepatitis B in a phase of immune control.

Aim. Comparative typing of Leptospira spp. strain collection based on analysis of 16S RNA fragment. Materials and methods. 2 pairs of primers were used for PCR, that jointly flank 1423 b.p. sized fragment. Sequences of Leptospira spp. strain 16S rRNA, presented in the international database, were used for phylogenetic analysis. Results. A high similarity, including interspecies, of the 16S fragment in Leptospira spp. strains was shown independently of the source, serovar and serogroup. Heterogeneity ofthe primary matrix, spontaneous mutations of hotspots and erroneous nucleotide couplings, characteristic for 16S sequence of pathogenic Leptospira spp. strains, are discussed. Molecular-genetic characteristic of certain reference Leptospira spp. strains by 16S sequence is obtained. Conclusion. Results of the studies give evidence on expedience of introduction into clinical practice of identification of Leptospira spp. by 16S sequence directly from the clinical material, that would allow to significantly reduce identification time, dismiss complex type-specific sera and other labor-intensive methods.

Efficiency of the telbivudine administration in the third trimestr of pregnancy for the prevention of perinatal transmission of hepatitis B virus