Vol 11, No 6 (2021)

A wide variety of zoonotic viruses that can cross the interspecies barrier promote the emergence of new, potentially pandemic viruses in the human population that is often accompanied by the disappearance of existing circulating strains. Among the various reasons underlying this phenomenon is the strengthening of herd immunity by expanding the immune layer of population and improving means and methods of medical care. However, natura abhorret vacuum, and new pathogens come to replace disappearing ones. Over the past ten years, humanity has faced two pandemics: swine flu A(H1N1)pdm09 in 2009 and COVID-19 in 2019, providing scientists with a unique opportunity to learn more about a relationship between respiratory viruses and their pathogenesis. Together with viruses of pandemic significance, a large number of seasonal respiratory viruses circulate, which contribute to the structure of human morbidity, and coinfections aggravate the condition of the illness. In the conditions of the spread of new viruses with unexplored characteristics, in the absence of means of prevention and therapy, it is especially important to prevent the aggravation of morbidity due to mixed infections. Here we review the mutual involvement of pandemic influenza A(H1N1)pdm09 and SARS-CoV-2 coronavirus and seasonal respiratory viruses in the epidemic process, discuss some issues related to their spread, potential causes affecting the spread and severity of the morbidity. The given facts testify to the existence of seasonality and temporal patterns of the beginning and end of respiratory viruses circulation. Interestingly, the beginning of circulation of the pandemic influenza A(H1N1)pdm09 virus led to a shift in the timing and intensity of circulation of some respiratory viruses, which is probably caused by existence of “replication conflicts” between them, and did not affect others. Coinfection with SARS-CoV-2-19 and other respiratory viruses, especially respiratory syncytial virus and rhinoviruses, was quite often observed. At the current stage, no aggravating effect of influenza on the course of COVID-19 in mixed infection has been established. Whether this is due to the mild course of influenza infection in the 2020 epidemic season, or the competitive impact of SARS-CoV-2 on influenza viruses is not yet clear. Experts are still at the stage of accumulating facts and working on creating means of effective prevention and treatment of the new coronavirus infection.

H. pylori is one of the most common commensal microorganisms in the human body, colonizing up to 60% of the inhabitants of all continents. Some strains of H. pylori have acquired virulent properties and their presence can significantly complicate the course of atrophic gastritis of type B, gastric ulcer and duodenal ulcer, as well as malignant diseases of the stomach. In such situations, eradication therapy seems to be pathogenetically justified. International recommendations for standard first-line triple eradication therapy, including a proton pump inhibitor (PPI), amoxicillin and clarithromycin in a course of 7–10 days, were proposed in 1996. Until the beginning of the XXI century, it was actively and with high efficiency (up to 90%) used everywhere, but later reports began to appear about a catastrophic decrease in results (up to 60%). Then it turned out that the effectiveness of the three-component (triple) therapy directly correlates with the resistance to clarithromycin, which has increased significantly in recent decades, and so necessitated the creation of new H. pylori elimination schemes. The results of various schemes for H. pylori eradication were analyzed, including variants of modified triple therapy associated with the inclusion of new drugs or an increase in the duration of eradication. In particular, it was proposed to replace amoxicillin with metronidazole. However, further studies have shown that the combination of clarithromycin with amoxicillin seems to be preferable, which is due to the high level of H. pylori resistance to metronidazole in many countries. Attempts to use probiotics in parallel, in particular cultures of various Lactobacillus species, were analyzed, which increases the level of eradication during standard triple therapy from 61.5 to 81.6%, and also significantly reduces the severity of side effects. It has been shown that a promising way to increase the effectiveness of 7-day first-line therapy schemes with clarithromycin is the use of modern effective PPIs (for example, esomeprazole or rabeprazole). The scheme of modified sequential therapy with the replacement of clarithromycin with tetracycline or levofloxacin, which has shown high efficiency, is considered. A variant of standard triple therapy modified into quadrotherapy with the addition of metronidazole or tinidazole was analyzed. It has been shown that the sequential therapy scheme is ineffective for eradication of multidrug-resistant strains. Ideally, the treatment of bacterial infections should be based on endoscopic sampling of gastric mucosa biopsies, followed by microbiological determination of the sensitivity of the isolated isolates to antibacterial drugs in vitro.

Among a whole variety of EBV- and HHV6-linked diseases only infectious mononucleosis is subject to official statistical reporting in Russia that substantially complicates objective assessment of etiological structure, incidence rate, characteristics of developing epidemic process. Currently, the data on the genetic EBV heterogeneity, even at the level of the main types (EBV1 and EBV2), as well as HHV6A and HHV6B, prevalence and clinical importance are mainly limited to foreign research publications. Few publications assessing this issue are available in Russian scientific papers. At the same time, examining circulation of virus genetic types (variants) and use of such data in implementing epidemiological surveillance after some other infections have been commonly practiced. One of the key issues is the level of developed laboratory support for molecular genetic monitoring. The goal of the study was to improve methodological basics for differential detection of HHV6A/B and the major EBV types. There were used samples of peripheral blood leukocytes collected from children aged 1–15 years with acute (n = 50) and asymptomatic infectious mononucleosis (n = 29). The detection and quantification of EBV and HHV6 DNA was performed by using real-time PCR. For differential determination of EBV1/EBV2 and HHV6A/HHV6B, an optimized one-round PCR with electrophoretic agarose gel detection amplification products was used. The data from our own study showed that frequency of detected EBV and HHV6 DNA in acute infectious mononucleosis patients comprised 74 and 72% compared to control group reaching 35 and 74%, respectively. It was found that among the examined children of Nizhny Novgorod Region, EBV1 and HHV6B prevailed in the viral population that agrees with existing insights about their geographical distribution in the adjacent territories. EBV2 was found in a single sample only in the control group. HHV6A was not detected in any of the groups. The methodological approach optimized in this study allows to separately detect HHV6A/HHV6B and the main EBV types according to a unified laboratory protocol, whereas combining it with additional stage of DNA enrichment increases the diagnostic sensitivity of PCR analysis, minimizes proportion of discordant and false negative results. Such an integrated approach can be applied for diagnostic, epidemiological and research purposes.

Background and aim. Pseudomonas aeruginosa is considered as a notorious pathogen due to its multidrug resistance and life threatening infections. We investigated the relationship between type III secretion toxins, biofilm formation, and antibiotic resistance among clinical P. aeruginosa isolates. Methods. A total of 70 genetically distinct clinical P. aeruginosa isolates were characterized for antibiotic resistance by disk diffusion assay. Biofilm formation was evaluated by microtiter plate method and presence of four exo genes (exoS, exoU, exoT and exoY) was investigated by PCR. A p-value < 0.05 was regarded statistically significant. Results. The most effective antibiotics were Meropenem and Piperacillin. Multidrug resistance was more prevalent in the ciprofloxacin-resistant isolates than in the susceptible isolates. The most frequently identified exo was exoS (37.1%). Genotype exoS/exoT was found in 4 isolates, while genotype exoU/exoT was not found. Prevalence of exoS was generally higher in the susceptible isolates than in the resistant isolates. A significant association was found between the formation of strong biofilm and resistance to antibiotics (p < 0.05). Prevalence of exoY and exoU was higher in the non-strong biofilm producers compared to the strong biofilm producers. Conclusion. Our study revealed formation of strong biofilm along with antibiotic resistance and the presence of exo genes in P. aeruginosa isolates. Knowledge of virulence gene profiles and biofilm formation may be useful in deciding appropriate treatment.

Introduction. In spite of significant advances in medical care, neonatal sepsis remains an important risk factor for neonatal morbidity and mortality. Accordingly, the present study was conducted to compare the number of nucleated red blood cells per 100 white blood cells (NRBC/100WBC) in neonates with early onset sepsis and non-infectious neonates. Materials and methods. In this cross-sectional study of 154 neonates admitted to the NICU of Ghaem Hospital in Mashhad, Iran within the first three days of life, during 2014 to 2018, the characteristics of 44 neonates identified early onset sepsis (Case group) were compared with 110 non-infectious neonates (Control group). After the confirmation of sepsis in neonates based on positive blood culture and laboratory results, a researcher-made questionnaire containing neonatal characteristics (gestational age, weight, first minute Apgar scores, fifth minute Apgar score, duration of oxygen therapy, and mechanical ventilation duration) and neonatal laboratory profiles (routine blood culture, WBC, NRBC/100WBC, CRP, blood glucose, calcium and venous blood gas) was filled in. Results. The results of this study showed that the absolute number of NRBC/mm³ in control group was 56.07±86.65 and in case group was 592.70±1166.75 (p = 0.000). Also, the number of NRBC per 100 white blood cells in control group was 6.54±11.18 and in case group was 31.84±40.07 (p = 0.000). The absolute number of NRBC/mm³ for the detection of early onset sepsis had a good sensitivity (78%) and NRBC/100WBC was suitable specificity (68.2%). Conclusion. This study indicated that NRBC/100 WBC and absolute NRBC count/mm³ can be helpful in the diagnosis of early onset sepsis and have an acceptable sensitivity and specificity.

Several hypotheses regarding the pathophysiology of interstitial cystitis/bladder pain syndrome (IC/BPS) have been put forward, but no consensus has yet been reached. It is suggested that cytokines are involved in IC/BPS, as well as in inflammatory autoimmune diseases. Examining various biomarkers of bladder tissue, including mast cells is underway. However, few reports on a consistent immune activity profile are available. The aim of the study was to assess the cytokine profile, counts of mast cells and plasma cells as well as their relationship in patients with IC/BPS with and without Hunner’s lesions. 44 women with Hunner’s lesions (group I) and 82 women with non-ulcer type (group II) were examined. Patients were questioned on the Pelvic Pain Symptom Scale, Urinary Frequency Scale (PUF), Visual Analogue Scale (VAS) and O’Leary–Santa Interstitial Cystitis Symptom Index (ICSI). Cystoscopy and hydrodistension of the bladder were performed under general anesthesia. In biopsies of the bladder wall, the number of plasma cells and mast cells was assessed, in urine — IL-1β, IL-6, IL-8, TNFα. Statistical processing of the results was carried out using the Statistica 6 software (StatSoft, USA). The Student’s t-test, Spearman’s correlation coefficient were calculated. In patients with Hunner's ulcer, there was an insignificant increase in the scale indices and decreased magnitude for average volume, urination frequency, and maximum emptying volume. The maximum bladder capacity during hydrodistension was lowered by 42.01% (p <0.001). The level of IL-8 in group I was higher on average by 28.57% (p = 0.434) as compared with that in group II, IL-6 — by 13.46% (p = 0.638), TNFα — by 9.09% (p = 0.244) and IL-1β — by 4.13% (p = 0.859). The number of mast cells in patients of group I vs. group II was higher by 40.65% (p < 0.05). In group I, a marked relationship was found between count of mast cells and plasma cells (r = –0.555, p > 0.05), as well as between count of plasma cells and IL-6 level (r = –0.639, p < 0.05). In group II, a significant connection was found between count of mast cells and TNFα level (r = +0.562, p < 0.05). Patients with Hunner’s IC/BPS are characterized by severe inflammation, wherein mast cells are involved in induction. Determination of cytokines in urine can provide a non-invasive division of IC/BPS into ulcerative and non-ulcerative groups.

The aim of the study is to conduct a comparative analysis of percentages for peripheral blood effector lymphocyte subsets in patients with uveal melanoma manifested by recurrent and chronic herpesvirus infection. There were 141 subjects enrolled: 70 patients with uveal melanoma, 38 patients with corneal ulcers and involvement of the uveal tract as well as 33 healthy donors. Immunophenotyping was performed by using laser flow cytometry with panel of monoclonal antibodies to differentiate lymphocyte subpopulations. IgM and IgG antibodies to herpesvirus infections were determined by using enzyme-linked immunosorbent assay on an automatic ELISA analyzer Lazurit (USA) with diagnostic kits of CJSC “Vector-Best” (Koltsovo). The data obtained showed that the absolute number of blood lymphocytes (CD45⁺) in patients with uveal melanoma did not differ from those in healthy donors. In contrast, patients with corneal ulcers and involvement of the uveal tract had this parameter increased. A decreased relative and absolute count of T cells (CD3⁺) in uveal melanoma, but increased absolute CD3⁺ number in inflammation was observed. No difference in relative and absolute content of the CD3⁺CD4⁺ helper/inducer subpopulation in patients with recurrent herpesvirus infections was found. Corneal ulcers in cancer patients revealed significantly increased absolute level of CD3⁺CD4⁺ helpers/inductor cells. Chronic herpesvirus infection in uveal melanoma patients showed increased relative and absolute number of cytotoxic T lymphocytes (CD3⁺CD8⁺). Recurrent herpesvirus infection was featured with decreased relative number of T lymphocytes (CD3⁺CD8⁺), upon inflammation, there was noted increased absolute and decreased relative number compared with healthy subjects. Double positive T cells increased in tumor and inflammation. B lymphocytes (CD19⁺) increased in melanoma and inflammation. The relative number of blood natural killer cells (CD16⁺CD56⁺) in uveal melanoma increased upon recurrent infection. Inflammation was coupled to decreased relative level of natural killer cells (CD16⁺CD56⁺). Melanoma showed no changes in CD4⁺/ CD8⁺ ratio; upon inflammation, its increase was noted in acute and chronic herpesvirus infections (p < 0.05). The suppression of the immune system in uveal melanoma, restricting antiviral defense, was revealed. The data obtained seem to be important for development of personalized approaches to prognosis and treatment of patients with uveal melanoma.

The aim of the study was to investigate the phenotype of effector T lymphocytes in patients with chronic viral hepatitis C (CVHC) before and after of treatment with direct antiviral drugs depending on the genotype of the virus. 50 patients with CVHC without signs of liver cirrhosis were examined. The diagnosis was made on the basis of epidemiological and clinical laboratory data as recommended by the European Association for the Study of the Liver when specific serological markers of CHCV and RNA of hepatitis C virus (HCV) were detected (EASL, 2016). The determination of HCV RNA was carried out by the method of quantitative polymerase chain reaction in real time. The degree of liver fibrosis in patients with CVHC was assessed using ultrasound elastography. Patients were treated for 3 months with direct antiviral drugs according to the recommendations of the European Association for the Study of the Liver (2016). The control group included 46 practically healthy individuals with severe chronic diseases of various organs and systems excluded during a routine examination, no health complaints, having normal clinical and biochemical blood tests in the absence of markers for viral hepatitis B and C, antibodies to opisthorchis and denying history of alcohol abuse. The study of the subpopulation composition of helper and cytotoxic T lymphocytes was carried out by direct immunofluorescence of whole peripheral blood. We obtained a 100% sustained virological response in patients with 1, 2 and genotypes of CHCV without signs of liver cirrhosis when using therapy with Sofosbuvir (400 mg) and Daclatasvir (60 mg) for 12 weeks. It was found that in CVHC patients were found characteristic features in the phenotypic composition of effector T lymphocytes before and after treatment with direct antiviral drugs in depending on the genotype of HCV. Patients with HCV genotypes 1 and 3 had an increase in the content of terminal differentiated effector (TEMRA) T helpers and effector memory (EM). Only patients with HCV genotype 2 had a decrease in the level of EM T-helper cells in the blood. A decrease in the relative number of T helpers of central memory (СM) was independent of the HCV genotype. The level of effector subpopulations of cytotoxic T lymphocytes in patients with CVHC was consistent with or exceeded control levels in depending on the genotype of HCV. The level of all investigated subpopulations of effector cytotoxic T lymphocytes in patients with HCV genotype 1 was equal to the control values. The number of naïve cytotoxic T cells and CM in peripheral blood in patients with HCV genotype 2 was increased. The content of naïve cytotoxic T lymphocytes, CM and TEMRA in patients with genotype 3 HCV in the blood was increased. The highest viral load was detected in patients with CVHC with genotype 1 HCV. Liver fibrosis was most pronounced in patients with CVHC infection with HCV genotypes 2 and 3. After 3 months of treatment with direct antiviral drugs the patients with CVHC had a reduced content of CM T helpers regardless of the HCV genotype. In addition, patients with HCV genotypes 1 and 3 had a decrease in the number of naïve T helpers and patients with HCV genotypes 2 and 3 had a normalization of the content of naïve cytotoxic T lymphocytes.

Influenza is a serious public health problem. The ability of influenza virus to change upon replication is the most serious issue for practical medicine and virology, which can fundamentally alter virus biological properties, such as infectivity and virulence. The high mutational variability of influenza viruses can contribute to rapidly emerging drug resistance. Therefore, the study of antiviral drug sensitivity among influenza viruses is necessary to justify proper drug use for treatment and prevention of influenza infection. The aim of the study was to examine antiviral drug susceptibility of influenza A/H1N1 and B virus strains isolated from various regions of Kazakhstan in the years 2018–2019. Materials and methods. The susceptibility analysis of 20 strains of influenza A/H1N1 and B viruses was carried out by using chemotherapeutic agents including Remantadine, Tamiflu, Arbidol, and Ingavirin. Viruses were cultured in the allantoic cavity of developing 10-day-old chicken embryos for 48 hours at 36^оC. The hemagglutinating activity was determined according to the standard method on 96-well plates using 0.75% chicken red blood cell suspension; the infectivity was calculated by the Reed–Muench method. The susceptibility of virus strains to different concentrations of antiviral drugs was evaluated by the level of virus reproductive suppression of 100 lg EID₅₀/0.2 ml in chicken embryos. Statistical analysis was performed using Microsoft Office Excel 2010 software. Results. A study of susceptibility to chemotherapeutic agents demonstrated heterogeneity of influenza A and B virus population isolated in Kazakhstan during the 2018–2019 period. The susceptibility to tamiflu was found in all Kazakhstan strains of influenza A/H1N1 virus and three type B strains (inhibitory concentration was 0.44–25.38 μg/mL). The reproduction of most viruses was effectively inhibited by Tamiflu at a concentration of 0.68–3.23 μg/mL. The inhibitory concentration for three strains of A/H1N1 virus was 7.23–25.38 μg/mL. Remantadine inhibited reproduction of viruses at higher doses (12.60–25.55 μg /mL). All investigated viruses were resistant to Arbidol and Ingavirin. A single type B influenza virus strain was found to be weakly susceptible to Ingavirin. Conclusion. The heterogeneity of influenza virus population in susceptibility to antiviral drugs suggest a need for constant epidemiological surveillance in order to identify drug-resistant variants.

Smallpox eradication and lack of adequate animal model for smallpox infection underlies a necessity to assess immunogenic and protective properties of genetic engineering-created live attenuated smallpox vaccines in several animal models of orthopoxviral infections. Here we compared immunogenic and protective properties of the recombinant vaccinia virus (VACV) LIVP-GFP intradermally (i.d.) inoculated to mice, guinea pigs and rabbits. LIVP-GFP immunization in all animal species was applied at dose of 2 × 10⁴ or 2 × 10⁶ PFU. Control animals were injected with saline. Blood sampling was performed on day 28 after virus LIVP-GFP or saline inoculation. Blood samples were taken intravitally from the retro-orbital venous sinus in mice, heart in guinea pigs or marginal ear vein in rabbits. Serum samples were isolated by precipitating blood cells via centrifugation. The serum anti-VACV IgG titers were determined by ELISA. On day 30 post-immunization animals were intranasally challenged with lethal dose of host specific orthopoxvirus species. Mice were infected by cowpox virus (CPXV) strain GRI-90 at dose 68 LD₅₀, guinea pigs – by VACV GPA at dose 56 LD₅₀, rabbits — by VACV HB-92 at dose 100 LD₅₀. All animals in control group died afterwards, whereas all animals immunized by attenuated recombinant virus LIVP-GFP at dose 2 × 10⁶ PFU survived. In case of the LIVP-GFP immunization at dose 2 × 10⁴ PFU, 88% of mice, 67% of rabbits and 50% of guinea pigs survived after being challenged with species-specific CPXV, VACV HB-92, and VACV GPA. ELISA data for the blood serum samples revealed a correlation between level of VACV-specific antibodies and level of protection in animal species. Based on the data obtained, it could be concluded that all three “animal–orthopoxvirus” models allow to provide with a proper evaluation of immunogenicity and protectiveness for generated modern attenuated vaccines against smallpox and other orthopoxviral human infections. Upon that, it was shown that BALB/c mouse strain was the most convenient investigational host species.

Introduction. In connection with the Ebola epidemic in the West African countries, including the Republic of Guinea, a failure in implementing measles immunization program was noted. A proportion of measles seronegative subjects in 2017 was 52.4% of total examined individuals. In 2018, a high proportion of measles cases among children aged 1–5 years (61.6%) was identified. In order to stop the 2018 outbreak, the Supplemental Immunization Campaign was conducted in the Konakri and Nzerekore prefectures in the Republic of Guinea. The aim of this study was to examine the 2019–2020 measles epidemic situation and assess the measles population immunity in the Republic of Guinea. Materials and methods. Measles-specific antibodies were examined in 1697 blood serum samples collected from residents of different regions of the Republic of Guinea, aged from 7 months to 76 years, obtained in 2019–2020, and tested retrospectively. The ELISA test systems Anti-Measles Virus ELISA (IgM) Euroimmun and Anti-Measles Virus ELISA (IgG) Euroimmun (Germany) were used. The presence of serum IgM measles antibodies was considered as acute measles infection. Statistical analysis was performed using the software package Statistica 6.0. Results. Blood sera (n = 638) were tested for IgM-measles, and in 46.6% of cases the diagnosis was confirmed by laboratory tests. The biggest proportion of the total cases (61.6%) was found in children aged 1–4 years. The second most important age group was 5–9 years of age, the third is children under 1 year: 18.5% and 11.8% of the total number of patients, respectively. Measles infection was registered in vaccinated patients in 7.4% of the total number of laboratory-confirmed cases. 1059 subjects were examined for IgG measles antibody. The lowest seroprevalence rate was found among children under 4 years of age (47.8%). The highest (85.5%) was found among subjects of 40 years old and older. Conclusion. Measles in GR remains a poorly controlled infection. As in the previous years of observation (2017–2018), children under 5 years of age are the most vulnerable cohort of the population, despite the 2018 DI campaign conducted in a number of GR territories. More problems with the measles control in the Republic of Guinea are expected in the period from 2021, as along with the COVID-19 epidemic, Ebola is repeatedly registered in the country. The Republic of Guinea particularly requires assistance from the international community to implement the WHO measles elimination program on a global scale.

The study of the small intestinal microbiota in humans is complicated due to the low availability of biomaterial. Non-invasive methods of metabolomics and bioinformatic data analysis can expand our understanding of the small intestinal microbiota structure and its role in maintaining body homeostasis. Here we assess the trajectory of age-related changes in the small intestinal microbial community of healthy individuals in the context of metaorganism-wide interaction between cytokine and neuroendocrine systems, by using the methods of gas chromatography mass spectrometry of microbial markers (GCMS MM) and optimal scaling. 110 apparently healthy children, adults and elderly individuals were enrolled to the study. The main types of the small intestine microbiota (Bacteroidetes, Firmicutes, Actinobacteria, Proteobacteria, and Fusobacteria) were quantified in peripheral blood by the GCMS MM method. To construct age-related trajectories of changes in the small intestinal microbiota and parameters of cytokine and neuroendocrine systems, the optimal scaling technique based on the multivariate Gifi transformation (CATPCA method) was used. It was found, that the small intestinal bacterial community of both children and seniors contained significantly lowered total number of microorganisms due to the low number of bacteria of Firmicutes and Actinobacteria types along with high number of members of Proteobacteria and Fusobacteria types compared with adults. Assessment of the trajectory of age-associated changes in microbiota of the small intestine showed that 1) children have strong dynamic fluctuations in the number and links within microbial community along with formation of links between the main regulatory immune and neuroendocrine systems of the metaorganism, 2) adults display plasticity and consistency in functioning of immune and nervous systems that determine the state of dynamic balance of the small intestinal microbiota, 3) healthy aging is characterized by high degree of cooperation between the main members of the bacterial community, which ensures system stability at new level, as one of the mechanisms of host adaptation. Thus, using methods of GCMS MM and optimal scaling, allows us to expand our understanding about age-associated trajectory of changes in the small intestinal microbiota and its cooperation with immune and neuroendocrine systems within the metaorganism, which can be used to develop new methods of therapy for infectious and non-infectious diseases.