Vol 13, No 6 (2023)

Group A streptococcal infections dominate among invasive streptococcal infections, with the major causative agent, Streptococcus pyogenes, being quite stable in the environment and bearing a large number of chromosome encoded pathogenicity factors or transmitted by horizontal transfer through bacteriophages. Different genetic variants of S. pyogenes can have a different set of pathogenicity factors able to change during pathogen evolution and determine virulence level for specific isolate. With a short incubation period, the disease can proceed with developing invasive infection and toxic shock syndrome with unfavorable outcome within 7 days from disease onset. The purpose of this article is to increase the doctors’ alertness to early recognition and diagnosis, which directly affects adequate treatment in a timely manner and disease outcome. The data on streptococcal morbidity in Russia and worldwide, review of laboratory diagnostic methods and pathogen genetic typing are presented. The maximum number of cases of streptococcal septicemia in Russia was registered in 2022, which accounted for 69% of all cases during the 2014–2022 observation period. The article also describes two clinical cases of fulminant invasive group A streptococcal infection in children with symptoms of acute respiratory viral infections at the onset of the disease. The results of various laboratory diagnostics methods verifying the diagnosis are presented. The genetic characterization of microbial isolates was performed by deep DNA sequencing. In the biological material from patients (including autopsy in one case), S. pyogenes sequence type ST-28, serotypes emm-1.25 and emm-1.0 were identified. The increasing importance of invasive streptococcal infection for health care in Russia and other countries may be associated with a possible change in dominating S. pyogenes genetic variants. In this regard, the study on circulating S. pyogenes genotypes on an ongoing basis as part of surveillance of streptococcal infection and development of vaccine for specific prevention are required.

Classical reassortment in developing chicken eggs is a well-established technique for obtaining LAIV strains. Naturally generated reassortant vaccine strains are characterized by high reproductive capacity, genetically stable characteristics of temperature sensitivity and cold resistance, which correspond to the characteristics of the MDV involved in crossing with the epidemic virus. Along with antigenic relevance, natural reassortment ensures attenuation of vaccine strains, good reproduction capacity in upper respiratory tract cells and inability to reproduction in the lower respiratory tract. With classical reassortment, the speed and efficiency of obtaining vaccine reassortants largely depend on the properties of epidemic virus, and therefore cannot be stable. The potential of reverse genetics is attractive because it allows to obtain vaccine reassortants quickly and efficiently, reduce the likelihood of spontaneous mutations; however, the vaccine strain is deprived of the advantages of natural selection, in which the most viable clones are selected. This study presents the results of comparatively assessed A(H3N2) LAIVs obtained in parallel by classical reassortment and reverse genetics according to criteria confirming that vaccine strains inherit the necessary properties that guarantee their harmlessness and high reproduction in chicken embryos. Strains for LAIV obtained by both methods retained all attenuating mutations inherited from the MDV, were highly reproductive at the optimal temperature, with temperature sensitivity corresponded to the MDV. However, strains obtained by reverse genetics, was observed to have partial loss of cold resistance in comparison with that of the MDV and classical reassortants. Reduced cold adaptation may negatively affect vaccine effectiveness. It is important that after several additional passages in chicken embryos at low temperature, the cold resistance of the vaccine strain, assembled by reverse genetics, was increased. Credibly that cold resistance is a phenotypic trait, the degree of manifestation of which depends on the temperature conditions of virus multiplication. The selective factor of reduced incubation temperature is missing in reverse genetics. In order for the cold-adapted phenotype to be fully realized, additional passages at low temperature of RG-reassortants are necessary. Thus, the reverse genetics method using plasmid technology allows to effectively prepare reassortant strains for LAIV. An important stage in obtaining vaccine strains using genetic engineering techniques should be the control of their cold-adapted phenotype and its optimization by additional passages at low temperature.

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.

Introduction. Features of cytokine production in mononuclear cell cultures from Lyme borreliosis patients based on clinical data remained poorly studied. The study aim was to estimate the patterns of baseline and lipopolysaccharide-induced cytokine-secretory activity of peripheral blood mononuclear leukocytes from patients with erythema migrans form of acute Lyme borreliosis based on clinical parameters. Materials and methods. Groups of 22 and 12 patients with the diagnoses of mild or moderate severity of monoinfection and co-infection with tick-borne encephalitis of Lyme borreliosis with erythema migrans were examined twice: on week 1 after disease onset and day 14. The control group included 17 healthy donors. Basal and lipopolysaccharide-induced IL-6, IL-10, and TNFα secretion levels were assessed in mononuclear leukocyte culture supernatants applying enzyme immunoassay. Statistical analysis was performed by using the Mann–Whitney U-test, Wilcoxon test, and Spearman’s rank correlation. Results. The group of moderate severity patients was clinically distinguished by severer fever and intoxication manifestations. At the disease onset, the basal TNFα, IL-6 and IL-10 secretion levels in the moderate severity patient cultures were significantly higher than in those of the other groups. After antibiotics treatment, the baseline TNFα and IL-10 levels tended to decrease. At the onset, lipopolysaccharide-induced cultures from the moderate severity patients showed significantly suppressed TNFα production and increased IL-10 secretion as compared to the other groups. Lipopolysaccharide-induced IL-6 secretion in the moderate vs. mild severity group supernatants was significantly lower. In dynamics, the induced TNFα levels in the moderate severity patients were increased to the magnitude exceeding that in the controls. Positive correlations between the IL-6 and TNFα basal levels and maximum body temperature or the C-reactive protein serum concentrations were revealed in the patients. Induced TNFα levels showed negative correlations with fever levels or with IL-10 secretion. Conclusions. It was demonstrated that basal TNFα, IL-6 and IL-10 secretion levels in the mononuclear cell cultures of acute Lyme borreliosis patients increased with the increasing disease severity. Suppression of lipopolysaccharide-induced TNFα production in the moderate severity patient cultures was presumably associated with the regulatory cytokine IL-10 effects.

The use of long-term multicomponent antibiotic therapy is the most effective way to treat tuberculosis (TB). However, little is known about the effect of this chemotherapy on the human intestinal microflora. The purpose of this study was to analyze an effect of long-term antibiotic therapy on gut microbiome composition and metabolic profile in TB patients. We used deep sequencing of fecal samples from 23 treatment-naive TB patients to reconstruct the metabolic capacity and strain/species-level abundance in the gut microbiome. Two fecal samples were obtained from each patient: before and after treatment. We showed that TB treatment regimen does not disrupt the overall diversity of the gut microbiome but does have an impact on gut bacterial microbiome composition and metabolic profile. While taking first-line anti-tuberculosis drugs (isoniazid, rifampicin, ethambutol, pyrazinamide), TB patients showed an apparent increase in Actinobacteria abundance. Pairwise comparison of metagenomic data revealed 28 differentially represented bacterial taxa, of which three species Bacteroides cellulosilyticus, Enterocloster aldensis, Clostridium spiroforme were strongly enriched in TB patients post-chemotherapy, whereas 25 species were enriched in TB patients before treatment (Bifidobacterium catenulatum, Enterococcus faecium, Bacteroides salyersiae, Bacteroides xylanisolvens, Bacteroides eggerthii, Lachnospira eligens, Akkermansia muciniphila, Ruminococcus lactaris, etc.) (p < 0.05). The metabolic profile of the gut microbiome was characterized by increased metabolic processes aimed at the growth and division of microbial cells. Iron is the main limiting factor for growth and reproduction. In addition, it is important to note the prevalence of glycolysis and lactate fermentation as the major means for energy production by intestinal microbiota.

Introduction. Diarrheal diseases are a global public health issue and cause 15% of deaths in children under 5 years old, of which about 80% occur in the regions of Africa and Southeast Asia. According to the Global Enteric Multicentre Study (GEMS) conducted in a number of African countries, one of the leading pathogens of high risk of death in infants and young children is diarrheagenic E. coli (DEC). In recent decades, antimicrobial resistance (AMR) has become globally ubiquitous. The Republic of Guinea urgently needs large-scale studies devoted to assessing DEC distribution and antibiotic resistance. The purpose of the study is to assess the pattern of E. coli infections and to test the susceptibility to antibiotics in strains of diarrheagenic E. coli sampled from individuals residing in the Republic of Guinea.

Materials and methods. From 2019 to 2022, we studied 724 samples of faeces of patients with acute diarrhea, among them 72 (9.9%) children aged 1–5 years, 128 (17.7%) children aged 6–17 years, and 524 (72.4%) people aged 18 years and older; a method of polymerase chain reaction (PCR) was applied with the use of the AmpliSense^® Escherichioses-FL reagent kit to identify the genetic determinants of DEC: EPEC, EHEC, ETEC, EIEC, and EAgEC (Central Research Institute of Epidemiology of Rospotrebnadzor, Russia). Susceptibility to 15 antimicrobial agents was found by the disc-diffusion method using Mueller–Hinton agar (Russia) and Oxoid discs (UK). Results were interpreted according EUCAST criteria, versions 2019–2022 (https://www.eucast.org/ast_of_bacteria/previous_versions_of_documents).

Results. For the period from 2019 to 2022, the percentage of E. coli infections in the etiological pattern of acute intestinal infections amounted to 51.7%. In the age-related manner, DEC was significantly more common in young children aged 0–5 (96.9%, p < 0.05) as compared to school age children aged 6–17 (53.9%) and adults (45.6%). In all years of observation, EAgEC strains prevailed, accounting for 38.4%. Other DEC pathotypes, EPEC, ETEC, EIEC and STEC, accounted for 27.2%, 17.5%, 11.8%, and 5.1%, respectively. DEC strains are susceptible to meropenem, amikacin, and nitrofurantoin. The activity of other antibiotics ranged from 11.3% for ampicillin, 28.3% for trimethoprim-sulfamethoxazole, and 34.0% for tetracycline to 73.6% for cephalosporins, 84.0% for aminoglycosides, and 98.1% for fluorinated quinolones.

Conclusion. To reduce the burden of diarrheal diseases in the Republic of Guinea, it may be necessary to conduct targeted epidemiological and microbiological studies to identify DEC and monitor the development of antimicrobial resistance of E. coli infection pathogens in the population.

The aim of this study was to assess the prevalence of occult hepatitis B infection among blood donors in St. Petersburg, as well as to characterize the identified virus isolates. The study material was represented by 2800 blood plasma samples collected in 2019 from blood donors living in St. Petersburg. The ELISA study for HBV marker rate consisted of HBsAg, anti-HBs IgG, anti-HBcore IgG. HBV DNA was analyzed by nested PCR with real-time hybridization-fluorescence detection on three targets allowing to determine virus DNA at low viral load, including HBsAg-negative chronic hepatitis B. Hepatitis B serological markers were detected in 69.43% of those surveyed, HBsAg was found in 0.43% of individuals, and all of which donated blood first time. A significant excess of the anti-HBcore IgG antibodies occurrence among primary donors (15.1%) compared with repeated/regular donors (7.48%) was shown. The prevalence of virus DNA in the group was 3.14%, including 2.71% of cases in HBsAg-negative CHB. Based on phylogenetic analysis of 88 isolates, HBV subgenotypes were determined in the following order: D1 and D2, 40.91% each, D3 and A2, 9.09% each. While determining the serological subtype in detected isolates, the serotype ayw3 (52.27%) vs ayw2 (46.59%) and adw2 (10.23%) prevailed. Drug resistance mutations, including compensatory ones, were detected in six examined patients (6.82%). In all genotype D isolates, multiple amino acid substitutions were identified in the RT, SHB, MHB, LHB, and Core regions; mutations in the preCore region were detected in 21.59% samples. In the MHR of the HBV genotype D genome, twenty-six positions were identified in which amino acid substitutions occurred, and all isolates showed modifications at positions 113, 114, 131, 134, 159, 161, 168, in 76 — at position 122, in 68 — at position 127, in 36 — at position 118, in 24 — at position 128. In HBV A2 isolates, mutations T113S, S143T, Y161F were identified. Nine isolates in the preCore region showed a polymorphism including a stop codon W28*W; in five isolates the W28S substitution was shown in the same position, and the W28*S variant was found in one more sample. The high incidence of HBsAg-negative CHB cases among blood donors, as well as the predominance of HBV isolates that simultaneously carry mutations resulting in diagnostic failure of HBsAg tests and prophylactic failure of immunoglobulin or vaccines and virus reactivation, mutations that contribute to disease progression obviously pose a threat to health and require to be further examined.

Introduction. In Montenegro, vaccination against COVID-19 infection began with the use of Gam-COVID-Vac, which was not approved for emergency use before the end of clinical trials, by the Food and Drug Administration and the European Medicines Agency. Therefore, it is necessary to emphasize the adverse effects.

Materials and methods. For the purpose of this study, there were collected data from national adverse events reporting form for Gam-COVID-Vac obtained from the Health Institution Pharmacy of Montenegro — Montefarm, as the holder of permits for these vaccines. Results. For the period March 1, 2021 to February 13, 2022, after administration of 16 756 doses of vaccine Gam-COVID, a total of 220 case reports, or 716 adverse effects were recorded. The mean age of vaccinated individuals who reported adverse effects was 40.79±11.35 years. Totally, 79.55% females versus 20.45% males reported side effects post-vaccination. The most common adverse reaction was pyrexia (79.55%). Other very common adverse effects were as follows: injection site pain (38.18%), headache (33.18%), myalgia (32.27%), malaise (31.82%), fever (30.45%), arthralgia (22.73%) as well as swelling and redness at the site of application (15.91%). Less common adverse effects were nausea, pain in extremity, diarrhea, dizziness, fatigue, sore throat and labial herpes. Serious adverse effects were recorded in 8 cases including tinnitus, thrombophlebitis, hypotension, chest pain, palpitations and peripheral cyanosis related to specific comorbidities.

Conclusions. After the administration of Gam-COVID vaccine, the population in Montenegro experienced mild to moderate adverse effects, with rare serious transient adverse effects related to specific comorbidities. The data presented here on investigating Gam-COVID-Vac caccine verified good safety profile and high tolerability evidenced by the statistics analysis as lacked COVID-19-associated hospitalizations or deaths.

Introduction. Since May 2022, an unusually large number of new monkeypox infections-a previously rare viral zoonotic disease, mainly reported from central and western Africa has been reported globally, and the World Health Organization (WHO) declared a global health emergency in July 2022. We aimed to systematically review the monkeypox virus epidemiology, pathogenesis, transmission, presentations, and outcomes.

Materials and methods. Our aim is to systematically review the epidemiology, pathogenesis, manifestations, and outcomes of Monkeypox disease. We searched the keywords in the online databases of PubMed, Embase, Scopus, and Web of Science and investigated all English articles until December 2022. In order to ascertain the findings, this study adheres to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) checklist. In order to optimize the quality, this review study benefits from the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) checklist. To minimize any probable bias risk, we utilized the Newcastle-Ottawa Scale (NOS) risk assessment tool.

Results. The most prevalent symptoms were rash and fever. The infection was accompanied by different complications such as, but not limited to, encephalitis (mainly in children), septicemia, bacterial cellulitis, retropharyngeal and parapharyngeal abscesses, etc. A wide range of hospitalization from 3.7% to 100% has been reported. The mortality rate ranged from 0% to 23%, which mainly occurred in infants and children. High mortality of the monkeypox rate was reported among pregnant women. The mortality rate of monkeypox is lower among women and those who received the smallpox vaccine compared to men and those who did not receive the vaccine. A wide range of the overall second-rate attack was reported, which is more pronounced in unvaccinated patients.

Conclusion. In our systematic review of 35 studies on monkeypox, we cast light on the existing evidence on its epidemiology, pathogenesis, manifestation, and outcomes. Further studies are needed to elucidate the natural history of the disease in various patients’ population, as well as detailing the monkeypox attack rate.

Catalase is a heme-containing enzyme belonging to protection factors that destroys peroxide compounds. The presence of catalase activity is an important ability of microorganisms that allows them to be protected from unfavorable factors as well as adapt to macroorganism conditions. Catalase along with superoxide dismutase plays an important role in pathogen resistance to phagocyte oxygen-dependent bactericidal mechanisms. The aim of the study was to investigate microsymbiont catalase activity from female reproductive tract in normocenosis and candidiasis dysbiosis using the chronobiological approach. The study was conducted on clinical isolates, isolated from female reproductive tract microsymbiocenosis. The catalase activity was determined by spectrophotometry based on 24 hour-long hydrogen peroxide reduction with 3-hours interval in winter season. Dynamic hydrogen peroxide was assessed in 3–5 experiment replicates. In some Lactobacillus spp., catalase was found containing no heme group — pseudocatalase. Chronobiological approach allowed to reveal enzyme activity from all microsymbionts. The dominant and associative microbiota isolated from healthy females was characterized by circadian (24 hours) rhythms of catalase activity early in the morning — 5 a.m. (р < 0.05). Hydrogen peroxide decomposes spontaneously or via non-enzymatic catalysts, and microorganisms cope with this situation under such conditions. In microsymbionts characteristic of female reproductive tract dysbiosis, and usually found in large numbers along with decreased Lactobacillus spp. ultradian rhythms with 12- and 8-hour harmonics of catalase activity with acrophase were recorded in the morning (8 a.m.) and evening hours (8 p.m.). The minimum values of enzyme production in all cultures were recorded at 12 p.m. and 5 p.m. Therefore, the contribution of the rhythm of the studied parameter at varying degree of vaginal sterility reflects the adaptive pathogen capabilities to the conditions of existence and can be the basis for studying related regulatory mechanisms. Mesor and amplitude phase stability are universal rhythmometric parameters used to evaluate patient’s condition independent of species assignment.