Journal of microbiology, epidemiology and immunobiology

Introduction. Bst polymerase plays a key role in the rapid diagnosis of infectious diseases due to its unique biochemical properties and potential application in loop-mediated isothermal amplification (LAMP). Several analogs of Bst polymerase have been described in the literature; however, these enzymes have not been widely used in molecular diagnostics.

The aim of the study is to obtain recombinant Bst and Btlv polymerases with the Sso7d domain and to test new possibilities for their application.

Materials and methods. Expression constructs carrying the polymerase gene were obtained using standard genetic engineering methods. The target enzyme was produced in Escherichia coli cells. Purification was carried out using metal-affinity chromatography methods followed by dialysis and concentration. RNA-dependent DNA polymerase (reverse transcriptase) and DNA polymerase activities of the enzymes were determined using non-radioactive methods with fluorescent detection. The functional properties of the enzymes were assessed using the Amplisens SARS-CoV-2-IT reagent kit and a method designed for the detection of mumps virus RNA in biological material using the LAMP format combined with reverse transcription.

Results. In the E. coli-based expression system, the following recombinant chimeric enzymes with displacing activity have been obtained: Bst_Sso7d, Bst_Sso7d_mut4 and Btlv_Sso7d. The developed cultivation and purification protocols allow for the production of enzymes in soluble form with a yield of up to 25% of the collected cell mass. Functional testing showed that in LAMP, the chimeric polymerases demonstrated similar activity to Bst polymerase without the Sso7d domain. At the same time, the Btlv_Sso7d polymerase exhibited increased reverse transcriptase activity and resistance to inhibitors.

Conclusion. The obtained chimeric polymerase Btlv_Sso7d, due to its improved properties, can be used in reagent kits for the diagnosis of infectious diseases by the LAMP method when using nucleic acid extraction methods.

Introduction. To date, among intestinal infections, salmonellosis is a pressing health problem worldwide, including in the Russian Federation, causing acute infectious outbreaks of disease.

Objective of the study: to assess the impact of the COVID-19 pandemic on the epidemiological patterns and molecular biological characteristics of salmonella infection in Primorsky Krai for 2019–2023.

Materials and methods. A retrospective epidemiological analysis of salmonellosis incidence during the COVID-19 pandemic in the population of Primorsky Krai was conducted. Plasmid characteristics of non-typhoidal Salmonella strains and their serotype were established using alkaline lysis and serological typing methods.

Results. During the COVID-19 pandemic, both in the Russian Federation and in Primorsky Krai, the registration of salmonellosis was significantly lower than in the pre-pandemic period, and the seasonality of salmonellosis shifted to the autumn months. An inverse relationship of medium strength was revealed between the monthly incidence of salmonellosis and COVID-19. No significant changes in the plasmid spectrum and in the proportion of registration by serogroups and serotypes of Salmonella were observed. It was shown that salmonellosis infection in Primorsky Krai is mainly caused by S. enteritidis strains with a plasmid characteristic of 38 MDa, 38:1.4 MDa, 38:2.3 MDa, 38:4.4 MDa, rare variants of plasmid types were also detected, the influence of the COVID-19 pandemic on the emergence of new plasmid types has not been established.

Conclusion. The presented results of the study on the molecular biological characteristics of Salmonella circulating in Primorsky Krai during the COVID-19 pandemic will open up prospects for understanding the ecological and epidemiological patterns of the development of the epidemic process of salmonella infection.

Introduction. The characteristics of the HIV epidemic in Cuba are comparable to those of the HIV epidemic in Russia. Migration between Cuba and Russia can affect HIV-1 genetic diversity and drug resistance in Russia. The vulnerable group of men with risk sexual behavior including men who have sex with men (MSM) and commercial sex workers (CSW) deserves special attention. The aim of our study was the analysis of HIV-1 genetic variants and HIV-1 drug resistance in blood plasma samples obtained from MSM migrants from Cuba living in Moscow.

Materials and methods. A collection of blood plasma samples, epidemiological and clinical information was collected from MSM patients — migrants from Cuba, nucleotide sequences of the HIV-1 genome were obtained. HIV-1 genotyping, cluster analysis and analysis of drug resistance (DR) were carried out.

Results. Samples and epidemiological data obtained in 2022–2024 from 27 patients were analyzed. 24/27 patients (including 12/15 suspected of infection in Moscow and all 10 — in Cuba) harbored HIV-1 variants typical for Cuba, not Russia. This indicates that 88.89% of patients were infected by their fellow citizen. DR was detected in 9 patients (33.33%; 95% CI 15.55–51.11). The most common resistance was DR to efavirenz (EFV) and nevirapine (NVP), which was associated with K103N, Y181C and P225H mutations.

Conclusion. The migration factor should be taken into account in HIV-1 prevention and control of HIV-spreading programs in Russia, and the genetic characteristics of HIV-1 in migrants should be taken into account in effective therapy selecting.

Introduction. Monitoring the resistance of Streptococcus pneumoniae to antimicrobials is an important component of epidemiological surveillance of pneumococcal infection, addressing the challenges of improving therapeutic approaches and containing the spread of resistance at regional and national levels.

The aim of the study is to investigate pneumococcal antibiotic resistance in preschool children following immunization with a 13-valent conjugated vaccine within the national immunization schedule.

Materials and methods. From 2016 to 2022, 1250 healthy children under the age of 6 from organized groups were examined. Detection of nasopharyngeal carriage of pneumococcus was performed using the culture method, and serotype was determined by polymerase chain reaction. Antimicrobial susceptibility was investigated using the disk diffusion method, and the minimum inhibitory concentration of antibiotics in resistant isolates was determined using E-tests.

Results. 265 isolates of S. pneumoniae were studied, and resistance to antimicrobials was found in 36.6%. The proportion of resistant isolates was 33.8% in the early post-vaccination period (2016–2018) and 45.3% in the late period (2020–2022) (p = 0.097). Penicillin-resistant pneumococci were detected only in the early post-vaccination period, with a frequency of 11.4% (p = 0.005). The prevalence of macrolide-resistant isolates in the analyzed periods was 27.9% and 35.9% (p = 0.219), and those resistant to 3 or more classes of antibiotics were 23.4% and 17.2%, respectively (p = 0.297). The serotype composition of resistant S. pneumoniae has changed: vaccine serotypes 19F, 6A, 6B, etc. (83.8%) have been replaced by non-vaccine serotypes – 15AF, 23A, etc. The proportion of vaccine serotypes in the late post-vaccination period was only 10.3%.

Conclusion. Against the backdrop of child immunization against pneumococcal infection, there has been a decrease in the prevalence of pneumococci with reduced susceptibility to β-lactams. S. pneumoniae resistance to macrolides remains high (35.9%). Resistant pneumococcal isolates primarily belong to serogroup 15 (15AF) and serotype 23A, which are not included in the 13-valent conjugate vaccine.

Introduction. Helicobacter pylori infection, however, data on the mechanisms of metronidazole (MTZ) and levofloxacin (LVX) resistance in Russia remain scarce. and levofloxacin (LVX) in Russia.

The aim of the study is to identify the determinants of resistance in clinical isolates of H. pylori to MTZ and LVX using whole-genome sequencing data.

Materials and methods. A retrospective analysis of 43 H. pylori isolates obtained from adult patients (2014–2022) was conducted. Susceptibility to antibiotics was determined using the bacteriological disk diffusion method. Whole-genome sequencing of 43 H. pylori strains was performed using a DNBSEQ-G50 sequencer.

Results. The evaluation of the phenotypic drug susceptibility test results showed that 11 isolates were susceptible to MTZ (MTZ-S), 31 were susceptible to LVX (LVX-S), while 32 isolates were resistant to MTZ (MTZ-R), and 12 were resistant to LVX (LVX-R). To identify the association between phenotypic and genotypic resistance, an analysis of nucleotide substitutions in the gyrA, gyrB, rdxA, frxA, fdxB and fur genes was conducted. Of all the mutations identified in the gyrA and gyrB genes, only D91/GNY in the gyrA gene was associated with phenotypic resistance to LVX and was found in 4/12 (33.3%) of the isolates (p < 0.05). The combined mutation D91G/N/Y+N87K in the gyrA gene was detected in 6/12 (50.0%) of LVX-R isolates (p < 0.001). Point mutations in the rdxA gene were detected in 21.9% (7/32) of MTZ-R isolates, leading to a frameshift or premature termination of protein synthesis. None of the mutations in the frxA, fur and fdxB genes were associated with H. pylori resistance to MTZ.

Conclusion. Based on the results of whole-genome sequencing of Russian clinical isolates of H. pylori, the detection of the combined mutation D91G/N/Y+N87K in the gyrA gene can serve as a predictor of the phenotypic resistance of H. pylori to levofloxacin.

In recent years, significant progress has been made in the field of drug development, particularly due to the use of computer modeling methods. One of the key stages in the development of new antiviral drugs is testing the efficacy of promising candidates in in vitro experiments using target viruses. The application of new technologies for conducting primary screening with pseudotyped viruses simplifies research, increases its efficiency and ensures the biosafety of the conducted studies.

The aim of this review is to analyze previous studies that have demonstrated the successful use of pseudovirus technology for the search of new chemotherapeutic agents against a range of RNA-containing viruses.

The analysis involved the literature presented in the PubMed, Scopus, Elsevier, and Google Scholar databases as of March 1, 2025. For the search, the following keywords were used: pseudovirus, virus inhibition, antiviral drugs, RNA viruses.

Pseudotyped viruses are recombinant viral particles that have the core proteins of one virus and the surface proteins of another, studied virus. The advantages of pseudovirus technology are its safety, high level of reproducibility of results, and the possibility of standardization. The lentivirus-based system was one of the first to be developed and remains one of the most in-demand. Using pseudoviruses, candidate molecules for infections caused by RNA-containing viruses, such as HIV-1, hepatitis C virus, tick-borne encephalitis virus, avian influenza viruses, and SARS-CoV-2, have been selected and studied. Most of the selected drugs act at the initial stage of the virus entry into the target cell. The examples provided illustrate the significant contribution of pseudovirus technology in dealing with serious socially significant diseases caused by RNA-containing viruses.