Репозиторий Университета

© Bionika Media Ltd. Objective. To investigate the clinical efficacy of Elgyna in the medical treatment of human papillomavirus (HPV)- associated cervical intraepithelial neoplasias (CIN). Subject and methods. The results of antibacterial therapy prior to destructive treatment were comparatively analyzed in 53 patients with severe dysbiosis in HPV-associated CIN. A study group included 27 patients who took the combined drug Elgyna as one vaginal tablet twice daily (6 administrations); a comparison group of 27 patients received the combined drug Tergynan as one vaginal tablet twice daily (6 administrations). A control group comprised 20 women with the visually unchanged cervix. All the women underwent standard examinations and vaginal microbiocenosis estimation by RT-PCR assay using the Femoflor test. Results. The use of the combined drug Elgyna during medical treatment before destruction was found to normalize molecular biological parameters at the local level and contributes to a significant reduction in the time of epithelialization and a decline in the number of recurrences. Conclusion. HPV-associated cervical neoplasias are accompanied by the development of pronounced dysbiotic processes in the vagina with the predominant involvement of obligate anaerobes. Taking into account the properties of the combined drug Elgyna that provides a normalizing effect on the parameters of biocenosis in accelerating healing and in preventing a recurrence of the process, it is appropriate to prescribe the drug before destructive treatments.

© 2017 Izdatel'stvo Meditsina. All rights reserved. Chronic hepatitis B is caused by a persistent form of hepatitis B virus, covalentiy closed circular DNA (cccDNA). Stability of cccDNA is associated with intracellular localization of cccDNA and formation of minichromosome, regu-lated by epigenetic mechanisms. One of the key mechanisms in epigenetics is methylation of DNA on CpG islands. Expression levels of DNA-methyltransferases (DNMTs) in chronic hepatitis B patients were shown to be upregu-lated. Nevertheless, the role of DNMTs in the life cycle of HBV and their effects on the cell remain elusive. In this review, we discuss latest achievements on the role of DNMTs in chronic hepatitis B and HBV in vitro models.

© 2017 Izdatel'stvo Meditsina. All rights reserved. The enzyme-linked immunosorbent assay (ELISA) and the neutralization test (NT) are often used to determine the level of seropositive population and to evaluate the immunogenicity of vaccines. ELISA provides information on the total pool of antiviral antibodies, while NT allows the antiviral protection level of a person to be estimated. It is assumed that the 1: 100 titer in ELISA and the 1: 10 titer in NT are protective. Obviously, the ratio of the total pool and virus neutralizing antibodies can vary as a result of natural immunization or vaccination. In this study, two methods were used to study the blood serum samples taken in a group of inhabitants of the Sverdlovsk region aged from 1 to 60 years. The samples were collected before immunization and 30 days after two immunizations with inactivated vaccines against tick-borne encephalitis of different manufacturers. Immunizations were performed either according to a standard scheme (30-day interval between immunizations), or according to an emergency scheme (14-day interval). It was shown that the data on the presence of antiviral antibodies in protective titers obtained by ELISA and NT were consistent in more than 85% of cases. The discrepancies between the data are due, in the first place, to the difference in the sensitivities of the two methods. The proportion of seropositive people according to NT data is always greater than that according to the results of ELISA. Nevertheless, among 174 children, about 5% of recipients after a double immunization were seropositive according to ELISA, but did not have neutralizing antibodies in protective titers.

© 2018, Bionika Media Ltd.. All rights reserved. Objective. To improve cervical cancer screening programs via clinical introduction of a diagnostic vaginal swab self-sampling test for the detection of high-risk human papillomavirus (hrHPV). Subjects and methods. A total of 455 residents of the Lipetsk Region were examined; vaginal swabs tested for HPV were taken both independently by a woman with a Qvintip device and by a gynecologist with a universal urogenital probe. The women were asked whether it was convenient for them to self-collect a vaginal swab. Results. In the surveyed sample of 455 women, there were 35 (7.7%) cases of hrHPV identified on the swabs collected by the Qvintip and 38 (8.4%) women with hrHPV detected on those taken by the physician, out of them 26 (68.5%) women had atypical squamous cells of undetermined significance. 70.3% of women preferred the Qvintip self-sampling method. Conclusion. The Qvintip self-sampling method for hrHPV testing shows high diagnostic efficiency, simplicity, and ease-to-use. The method may be recommended for the detection of hrHPV to increase cervical cancer screening coverage.

© 2018, Bionika Media Ltd.. All rights reserved. The authors have established a relationship between intraepithelial cervical lesions and vaginal biocenotic disorders in pregnant women with human papillomavirus (HPV) infection and determined their management tactics. The HPV–positive pregnant women have a high incidence of vaginal dysbiosis (38.7%) that is, in most of them, characterized by co–infection (65%), mainly by anaerobic and aerobic infections (44.1%). High-rate HPV in the pregnant women is associated with bacterial vaginosis, mixed dysbiosis, and their recurrences and with the frequency of atypical squamous cells of undetermined significance (ASCUS) and low-grade squamous intraepithelial lesions (LSIL) in the presence of an inflammatory response. Restoring the normal microflora contributes to the disappearance of the inflammatory response and to the reduction of ASCUS. Therapy for dysbiosis does not change the rate of LSIL at 3-month follow-up.

© 2018 Ruslania. All rights reserved. Liver cirrhosis and hepatocellular carcinoma are the most common outcomes of chronic hepatitis B. Hepatitis B virus (HBV) induces transformation and cell death in chronic hepatitis B (CHB). DNA double strand breaks (DSBs) represent the most dangerous type of genome damage. It was shown previously that generation of phosphorylated histone H2AX foci is a reliable marker of DSBs. The aim of this study was to analyse generation of yH2AX foci in HBV and hepatitis D virus (HDV) infection in vitro and in liver biopsies of patients with CHB and CHB with delta-agent (CHD). Human hepatoma cell line HepG2-1.1merHBV with activated HBV life cycle was used to perform real-time PCR for analysis of pregenomic RNA, HBV DNA, HBV cccDNA and for immunocytochemical analysis of yH2AX. Liver biopsies from CHB and CHD patients were analyzed to confirm the results. HBV induces multiple discrete yH2AX foci in HepG2-1.1merHBV cells in vitro and in biopsies of CHB and CHB+D patients. The ratio of hepatocytes w/o yH2AX foci is significantly lower (49,9+7-12,3% vs. 85,5+/-0,9%, p<0,05), while the proportion of cells with 1-10 yH2AX foci is higher (49,3+7-12,6% vs. 14,5+/-0,9%, p<0,05) compared to healthy control. There is a significant increase In the mean number of yH2AX foci in biopsies from CHB+D patients (3,5+7-1,1 and 5,5+/-1,5 vs. 0,5+/-0,16 in control hepatocytes, p<0.05). The ratio of hepatocytes w/o yH2AX foci is significantly lower in CHB and CHB+D patients, while percentage of cells with 1-10 yH2AX foci is higher. Rare hepatocytes with multiple (11-30 yH2AX foci per cell) foci appear in CHB and CHB+D patients. In conclusion, yH2AX foci are generated in hepatocytes of CHB and CHB+D patients and can be utilized to assess genome damage, associated with HBV and HDV viral infection.

© 2018 Saint Petersburg Pasteur Institute.All Rights Reserved. Enteroviruses are small RNA viruses, which are ubiquitous and commonly cause outbreaks with various clinical manifestations. In 2006, the Program on enterovirus surveillance was approved in the Russian Federation. Over the last years, molecular-biological and bioinformatics methods for enterovirus epidemiology studies have been developed both in Russia and worldwide. Currently, identification of enteroviruses is carried out by analyzing nucleotide sequence of the full-length VP1 genome region (ca. 900 nt). Routinely, it is sufficient to obtain a partial VP1 genome region sequence (ca. 300 bp) for enteroviruse verification in most cases; however, a more stringent type criterion of 80% sequence identity should be used compared to the 75% sequence identity cut-off for the complete VP1 genome region. Further sequence analysis may be performed by using Bayesian phylogenetic methods, which allow using molecular clock to trace outbreak emergence. Enteroviruses accumulate about 0.5–1% nucleotide substitutions per year. Therefore, a short genome fragment may be used to analyze virus phylodynamics at the level of international transfers and circulating virus variants. On a shorter timescale, a full-length VP1 genome region or a complete genome sequence are preferred for investigating molecular epidemiology, because a short sequence allows to reliably distinguish not more than 1–2 transmission events per year. Thus, determining enterovirus sequences for full-length VP1 genome region or full-genome sequence is preferred for examining viral outbreaks. It is increasingly apparent that analyzing available enterovirus nucleotide sequences reveals limitations related to uneven surveillance efficacy in various countries and short length of genome fragment measured in routine control. As a result, a proper global-scale analysis of enterovirus molecular epidemiology remains problematic. Over the last 20 years, the number of available enterovirus nucleotide sequences increased by hundred times, but understanding emergence of enterovirus infection outbreaks remains limited. Further development of enterovirus surveillance would require new methods for sewage monitoring, affordable high-throughput sequencing and harmonization of global surveillance systems.

© 2018, Silicea-Poligraf. All rights reserved. Aim. To evaluate the prevalence of myocarditis in adult patients with non-compaction myocardium (NCM) of the left ventricle (LV), and its influence on the disease course, results of treatment and outcomes. Material and methods. To the study, 103 adult patients included, with NCM, 61 males, mean age 45,6±14,9 y. o. (from 18 to 78). Mean end diastolic LV size was 6,0±0,8 сm, EF LV 38,8±14,5%. Diagnosis of NCM had been done by echocardiography, multispiral computed tomography (n=81) and magnetic resonance tomography (n=39). DNA-diagnostics was performed by NGS method with further Senger sequencing. Pathogenic mutations were found in 9% of patients in the genes MYH7, MyBPC3, LAMP2, DES, DSP, TTN. The investigation also included anticardiac antibodies, genome of cardiotropic viruses by PCR, coronary arteriography (n=26), scintigraphy (n=25). Morphological assessment of the myocardium was done in 19 patients (12 endomyocardial biopsies), 1 intraoperation biopsy, 3 explanted hearts, incl. 2 after biopsy, and 5 autopsies. Mean follow-up 12 [2; 32] months. Results. Myocarditis was found in 53,4%, incl. virus-positive in 32,7% of those, with morphology done for 19 patients (active myocarditis in 10, borderline in 6; with minimal signs of activeness in 3). Viral genome in myocardium found in 8 patients (42,1%). The prevalence of myocarditis 44,4% in an arrhythmic variant of NCM, 12,5% in chronic ischemic, 57,5% in dilation cardiomyopathy, 50,0% in NCM patients with other cardiomyopathies. Special cases were acute/subacute myocarditis in NCM (10,7% of all), acute necrosis (infarction) in 4,9%. Comorbidity with myocarditis in NCM led to significantly more severe LV dysfunction (CHF FC 2 [1; 3] v 1,75 [0; 2], p<0,01, EF 33,8±13,5 v 44,7±13,6%, p<0,001), more prevalent non-sustained ventricular tachycardia (67,3% v 29,3%, p<0,01), proper shocks (38,9% v 0, p<0,05), deaths (16,4 and 4,2%, OR 5,75, 95% CI 1,21-27,43, p<0,05) and transplantation (7,3% v 2,1%, p>0,05). Only in myocarditis patients, as a result of basis (antiviral, immune suppression) and cardiotropic therapy there was significant increase of EF (in acute myocarditis from 25,4±7,9 to 38,6±9,5%, p<0,01), decrease of LV size and pulmonary systolic pressure. Conclusion. Myocarditis is typical phenomenon developing in patients with the primary, i. e. genetically verified NCM. The nature of myocarditis in NCM varies (primary infectious-autoimmune, secondary as a response on genetic/ischemic damage of cardiomyocytes), however regardless of this, it leads to significant worsening of structural and functional parameters, increase of the life-threatening arrhythmias rate, and outcomes (death + transplantation, proper shocks), demanding for active basic therapy.

© 2017 Elsevier Inc. Rabies virus is endemic to Russia, among other countries. It is therefore critical to develop a high-quality and high-precision diagnostic procedure for the control and prevention of infection. The main objective of the research presented here was to develop a reliable RT-qPCR assay for rabies diagnostics. For this purpose, a RABV strains from various biological and geographical origins were used. In addition, rabies-positive and rabies-negative samples, as well as nucleic acids from other viruses and DNA extracted from the brain tissues of mice, dogs, cats, bats and humans, were studied using the developed assay. The analytical sensitivity of the assay, as assessed using armored recombinant positive control dilutions, was 103 copies/ml, and the sensitivity measured using characterized strains was between 0.1 LD50/ml and 1.0 LD50/ml. A broad range of RNA from RABV strains circulating in different regions of Russia, as well as RNA from RABV-positive primary brain samples from 81 animals and two humans, was detected using the developed assay. No false-positive or false-negative results were obtained. Given that high analytical and diagnostic sensitivities and a high specificity were verified for this assay, it has high potential as a screening test that may be suitable for the epizootiological monitoring of animals and for the fast postmortem diagnosis of rabies.

Репликативный полипротеин 1а вируса желтухи свеклы (ВЖС) содержит консервативные домены лидерной папаин-подобной протеиназы (РСР), метилтрансферазы (MTR) и РНК хеликазы (HEL). Центральный район (central region, CR) между MTR и HEL ранее считали «вариабельным». Нами проведен компьютерный анализ CR, который позволил выявить новый консервативный домен между позициями 1287-1390 (здесь и далее приводится нумерация аминокислотных остатков белка 1а вируса желтухи свеклы, BYV), сохраняющийся у всех представителей рода Closterovirus. Этот домен содержит 4 предсказанных альфа-спиральных участка (альфа А – D) и три строго консервативные позиции – глютамат-1291, пролин-1380 и аргинин-1384. Кроме того, биоинформатический анализ позволил предсказать амфипатическую спираль в позициях 1368-1380 (входящую в состав участка альфа D). Гидрофобный домен CR-2 (позиции 1305-1494 белка 1а), вызывающий при экспрессии в растениях реструктуризацию эндоплазматического ретикулюма и образование подвижных глобул диаметром ~1 мкм, включает участки альфа В, С и D. Установлено, что экспрессия в растениях слитных белков CR-2:GFP и GFP:CR-2 вызывает сходный «глобулообразующий» фенотип, т.е. N-концевое или C-концевое положение маркера GFP в слитном белке не влияет на переформатирование мембран эндоплазматического ретикулюма. Проведен делеционный анализ CR-2 BYV. Показано, что делеционные варианты 1355-1494 и 1325-1484 сохраняют фенотип дикого типа (образование глобул и реструктуризация ЭР вокруг ядра клетки). Варианты 1375-1484, 1368-1484 и 1368-1432 индуцировали образование глобул, но утрачивали способность к реструктуризации ЭР. Внесение замен гидрофобных аминокислотных остатков на остатки серина и глицина в «минимальном» делеционном мутанте 1368-1432 блокировало образование глобул. Предложена рабочая гипотеза о влиянии консервативной амфипатической спирали 1368-1385 в белке 1а BYV на ремоделирование мембран ЭР растительной клетки и создание репликативных платформ при клостеровирусной инфекции.

Wild ducks are known to be able to carry avian influenza viruses over long distances and infect domestic ducks, which in their turn infect domestic chickens. Therefore, prevention of virus transmission between ducks and chickens is important to control the spread of avian influenza. Here we used a low pathogenic wild aquatic bird virus A/duck/Moscow/4182/2010 (H5N3) for prevention of highly pathogenic avian influenza virus (HPAIV) transmission between ducks and chickens. We first confirmed that the ducks orally infected with H5N1 HPAIV A/chicken/Kurgan/3/2005 excreted the virus in feces. All chickens that were in contact with the infected ducks became sick, excreted the virus, and died. However, the ducks orally inoculated with 10⁴ 50% tissue culture infective doses of A/duck/Moscow/4182/2010 and challenged 14 to 90 days later with H5N1 HPAIV did not excrete the challenge virus. All contact chickens survived and did not excrete the virus. Our results suggest that low pathogenic virus of wild aquatic birds can be used for prevention of transmission of H5N1 viruses between ducks and chickens.

Chronic hepatitis B virus (HBV) infection has long remained a critical global health issue. Covalently closed circular DNA (cccDNA) is a persistent form of the HBV genome that maintains HBV chronicity. Decades of extensive research resulted in the two therapeutic options currently available: nucleot(s)ide analogs and interferon (IFN) therapy. A plethora of reliable markers to monitor HBV patients has been established, including the recently discovered encapsidated pregenomic RNA in serum, which can be used to determine treatment end-points and to predict the susceptibility of patients to IFN. Additionally, HBV RNA splice variants and cccDNA and its epigenetic modifications are associated with the clinical course and risks of hepatocellular carcinoma (HCC) and liver fibrosis. However, new antivirals, including CRISPR/Cas9, APOBEC-mediated degradation of cccDNA, and T-cell therapies aim at completely eliminating HBV, and it is clear that the diagnostic arsenal for defining the long-awaited sterilizing cure is missing. In this review, we discuss the currently available tools for detecting and measuring HBV RNAs and cccDNA, as well as the state-of-the-art in clinical implications of these markers, and debate needs and goals within the context of the sterilizing cure that is soon to come. View Full-Text

Wild ducks are known to be able to carry avian influenza viruses over long distances and infect domestic ducks, which in their turn infect domestic chickens. Therefore, prevention of virus transmission between ducks and chickens is important to control the spread of avian influenza. Here we used a low pathogenic wild aquatic bird virus A/duck/Moscow/4182/2010 (H5N3) for prevention of highly pathogenic avian influenza virus (HPAIV) transmission between ducks and chickens. We first confirmed that the ducks orally infected with H5N1 HPAIV A/chicken/Kurgan/3/2005 excreted the virus in feces. All chickens that were in contact with the infected ducks became sick, excreted the virus, and died. However, the ducks orally inoculated with 10⁴ 50% tissue culture infective doses of A/duck/Moscow/4182/2010 and challenged 14 to 90 days later with H5N1 HPAIV did not excrete the challenge virus. All contact chickens survived and did not excrete the virus. Our results suggest that low pathogenic virus of wild aquatic birds can be used for prevention of transmission of H5N1 viruses between ducks and chickens.

Chronic hepatitis B virus (HBV) infection has long remained a critical global health issue. Covalently closed circular DNA (cccDNA) is a persistent form of the HBV genome that maintains HBV chronicity. Decades of extensive research resulted in the two therapeutic options currently available: nucleot(s)ide analogs and interferon (IFN) therapy. A plethora of reliable markers to monitor HBV patients has been established, including the recently discovered encapsidated pregenomic RNA in serum, which can be used to determine treatment end-points and to predict the susceptibility of patients to IFN. Additionally, HBV RNA splice variants and cccDNA and its epigenetic modifications are associated with the clinical course and risks of hepatocellular carcinoma (HCC) and liver fibrosis. However, new antivirals, including CRISPR/Cas9, APOBEC-mediated degradation of cccDNA, and T-cell therapies aim at completely eliminating HBV, and it is clear that the diagnostic arsenal for defining the long-awaited sterilizing cure is missing. In this review, we discuss the currently available tools for detecting and measuring HBV RNAs and cccDNA, as well as the state-of-the-art in clinical implications of these markers, and debate needs and goals within the context of the sterilizing cure that is soon to come. View Full-Text

Репликативный полипротеин 1а вируса желтухи свеклы (ВЖС) содержит консервативные домены лидерной папаин-подобной протеиназы (РСР), метилтрансферазы (MTR) и РНК хеликазы (HEL). Центральный район (central region, CR) между MTR и HEL ранее считали «вариабельным». Нами проведен компьютерный анализ CR, который позволил выявить новый консервативный домен между позициями 1287-1390 (здесь и далее приводится нумерация аминокислотных остатков белка 1а вируса желтухи свеклы, BYV), сохраняющийся у всех представителей рода Closterovirus. Этот домен содержит 4 предсказанных альфа-спиральных участка (альфа А – D) и три строго консервативные позиции – глютамат-1291, пролин-1380 и аргинин-1384. Кроме того, биоинформатический анализ позволил предсказать амфипатическую спираль в позициях 1368-1380 (входящую в состав участка альфа D). Гидрофобный домен CR-2 (позиции 1305-1494 белка 1а), вызывающий при экспрессии в растениях реструктуризацию эндоплазматического ретикулюма и образование подвижных глобул диаметром ~1 мкм, включает участки альфа В, С и D. Установлено, что экспрессия в растениях слитных белков CR-2:GFP и GFP:CR-2 вызывает сходный «глобулообразующий» фенотип, т.е. N-концевое или C-концевое положение маркера GFP в слитном белке не влияет на переформатирование мембран эндоплазматического ретикулюма. Проведен делеционный анализ CR-2 BYV. Показано, что делеционные варианты 1355-1494 и 1325-1484 сохраняют фенотип дикого типа (образование глобул и реструктуризация ЭР вокруг ядра клетки). Варианты 1375-1484, 1368-1484 и 1368-1432 индуцировали образование глобул, но утрачивали способность к реструктуризации ЭР. Внесение замен гидрофобных аминокислотных остатков на остатки серина и глицина в «минимальном» делеционном мутанте 1368-1432 блокировало образование глобул. Предложена рабочая гипотеза о влиянии консервативной амфипатической спирали 1368-1385 в белке 1а BYV на ремоделирование мембран ЭР растительной клетки и создание репликативных платформ при клостеровирусной инфекции.