Vol 68, No 2 (2023)

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Full Issue


Influence of siRNA complexes on the reproduction of influenza A virus (Orthomyxoviridae: Alphainfluenzavirus) in vivo

Pashkov E.A., Momot V.Y., Pak A.V., Samoilikov R.V., Pashkov G.A., Usatova G.N., Kravtsova E.O., Poddubikov A.V., Nagieva F.G., Sidorov A.V., Pashkov E.P., Svitich O.A., Zverev V.V.


Introduction. Influenza is one of the most pressing global health problems. Despite the wide range of available anti-influenza drugs, the viral drug resistance is an increasing concern and requires the search for new approaches to overcome it. A promising solution is the development of drugs with action that is based on the inhibition of the activity of cellular genes through RNA interference.

Aim. Evaluation in vivo of the preventive potential of miRNAs directed to the cellular genes FLT4, Nup98 and Nup205 against influenza infection.

Materials and methods. The A/California/7/09 strain of influenza virus (H1N1) and BALB/c mice were used in the study. The administration of siRNA and experimental infection of animals were performed intranasally. The results of the experiment were analyzed using molecular genetic and virological methods.

Results. The use of siRNA complexes Nup98.1 and Nup205.1 led to a significant decrease in viral reproduction and concentration of viral RNA on the 3rd day after infection. When two siRNA complexes (Nup98.1 and Nup205.1) were administered simultaneously, a significant decrease in viral titer and concentration of viral RNA was also noted compared with the control groups.

Conclusions. The use of siRNAs in vivo can lead to an antiviral effect when the activity of single or several cellular genes is suppressed. The results indicate that the use of siRNAs targeting the cellular genes whose expression products are involved in viral reproduction is one of the promising methods for the prevention and treatment of not only influenza, but also other respiratory infections.

Problems of Virology. 2023;68(2):95-104
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Evaluation of the dynamics of detection of viable SARS-CoV-2 (Coronaviridae: Betacoronavirus: Sarbecovirus) in biological samples obtained from patients with COVID-19 in a health care setting, as one of the indicators of the infectivity of the virus

Kuznetsova N.A., Ogarkova D.A., Gushchin V.A., Antipyat N.A., Bacalin V.V., Burgasova O.A., Vasilchenko L.A., Samkov A.A., Simakova Y.V., Divisenko E.V., Siniavin A.E., Tkachuk A.P., Kolobukhina L.V., Shidlovskaya E.V., Tyurin I.N., Kruzhkova I.S., Zlobin V.I., Nikiforova M.A., Odnoralov M.A., Gintsburg A.L.


Introduction. The study of the mechanisms of transmission of the SARS-CoV-2 virus is the basis for building a strategy for anti-epidemic measures in the context of the COVID-19 pandemic. Understanding in what time frame a patient can spread SARS-CoV-2 is just as important as knowing the transmission mechanisms themselves. This information is necessary to develop effective measures to prevent infection by breaking the chains of transmission of the virus.

The aim of the work – is to identify the infectious SARS-CoV-2 virus in patient samples in the course of the disease and to determine the duration of virus shedding in patients with varying severity of COVID-19.

Materials and methods. In patients included in the study, biomaterial (nasopharyngeal swabs) was subjected to analysis by quantitative RT-PCR and virological determination of infectivity of the virus.

Results. We have determined the timeframe of maintaining the infectivity of the virus in patients hospitalized with severe and moderate COVID-19. Based on the results of the study, we made an analysis of the relationship between the amount of detected SARS-CoV-2 RNA and the infectivity of the virus in vitro in patients with COVID-19. The median time of the infectious virus shedding was 8 days. In addition, a comparative analysis of different protocols for the detection of the viral RNA in relation to the identification of the infectious virus was carried out.

Conclusion. The obtained data make it possible to assess the dynamics of SARS-CoV-2 detection and viral load in patients with COVID-19 and indicate the significance of these parameters for the subsequent spread of the virus and the organization of preventive measures.

Problems of Virology. 2023;68(2):105-116
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The prevalence of IGM antibodies to Zika virus in pregnant women in Northern Nigeria

Adekola H.A., Ojo D.A., Balogun S.A., Dipeolu M.A., Mohammed M., Adejo D.S., Aliyu R.M., Abdullahi M.A., Madugu N.H.


Introduction. Zika virus (ZIKV) infection during pregnancy can result in severe outcomes for both the pregnant woman and the developing fetus.

The objective of this study was to investigate the prevalence of Zika virus infection among pregnant women who sought healthcare services at Ahmadu Bello University Teaching Hospital.

Materials and methods. Serum samples were collected and analyzed using Enzyme Linked Immunoassay and RT-qPCR methods, while a structured questionnaire was used to gather relevant information about the participants.

Results. The results showed that 53 out of the 180 pregnant women tested positive for Anti-Zika IgM antibodies, which represents a 29.4% prevalence rate. Subsequent RT-qPCR analysis found that only 6 out of the 53 positive samples contained Zika virus RNA. Fever and headache were the most commonly reported symptoms related to the infection.

Conclusion. These findings indicate a potential outbreak of Zika fever in Northern Nigeria emphasizing the importance for pregnant women to take precautions to avoid getting infected.

Problems of Virology. 2023;68(2):117-123
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Selection of conditions for effective inactivation of Pseudopestis avium virus (Paramyxoviridae: Orthoavulovirus: Avian orthoavulovirus 1) for the production of a Newcastle disease vaccine

Jekebekov K.K., Assanzhanova N.N., Nurpeisova A.S., Ryskeldinova S.Z., Absatova Z.S., Abay Z.S., Shayakhmetov Y.A., Omurtay A.D., Moldagulova S.U., Kalimolda E.Z., Sadikalieva S.O., Shorayeva K.A., Zakarya K.D.


Introduction. Newcastle disease (ND) is classified as especially dangerous pathogen. Its primary source is an infected or recovered bird. The virus shedding begins just in a day after infection, and virus remains in the body for another 2-4 months after the recovery. The complexity of the final elimination of the causative agent of the disease lies in its ability for long-term preservation in the external environment and the possibility of constant circulation in one complex between groups of birds of different sex and age. Therefore, the main element of protecting birds from ND is immunoprophylaxis that is based on vaccines containing an inactivated ND virus (NDV).

The aim of the work ‒ is to optimize the parameters of inactivation of the NDV actual strain H with formaldehyde at final concentrations of 0.01, 0.025, 0.05, and 0.1% under temperature conditions of 20 ± 2 and 37 ± 0.5 °C.

Materials and methods. We used a virus-containing suspension of the NDV strain H with an initial biological activity of 10.75 lg EID50/cm3 grown by cultivation in 10-day-old developing chick embryos.

Results. On the 16th day after the administration of the tested suspensions of NDV inactivated at different temperatures and concentrations of the inactivant , the geometric mean titers of antibodies to NDV in sera of vaccinated birds were at least 1 : 63 in the hemagglutination inhibition reaction, indicating that the studied inactivated suspensions were antigenically active.

Conclusion. The optimal parameters of the inactivation mode (final concentration, temperature and time of inactivation) of the NDV strain H were established. The inactivation process at 37 ± 0.5 °C with inactivant concentrations of 0.01, 0.025, 0.05, and 0.1% lasts up to 72, 22, 18, and 12 hours, respectively. The inactivation process at 20 ± 2 °C with inactivant concentrations of 0.05 and 0.1% lasts up to 22 and 18 hours, respectively.

Problems of Virology. 2023;68(2):124-131
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Antigenic and immunogenic activity of virus-like particles based on rabbit hemorrhagic disease virus (Caliciviridae: Lagovirus) genotypes GI1 and GI2 recombinant major capsid proteins

Mukhin A.N., Alekseev K.P., Yuzhakov A.G., Selezneva E.V., Moskvina A.S., Verkhovsky O.A., Aliper T.I.


Introduction. Rabbit hemorrhagic disease is an acute highly contagious infection associated with two genotypes of pathogenic Lagovirus. Antibodies to major capsid protein (Vp60) are protective.

The aim of the work ‒ is an evaluation of antigenic and immunogenic activity of virus-like particles (VLPs) based on recombinant major capsid proteins of both genotypes of rabbit hemorrhagic disease virus (RHDV) (recVP60-GI1 and recVP60-GI2).

Materials and methods. Baculovirus-expressed VLPs were evaluated using electron microscopy and administered to clinically healthy 1.5–3 month old rabbits in a dose of 50 µg. Rabbits were challenged with 103 LD50 of virulent strains “Voronezhsky-87” and “Tula” 21 days post immunization. Serum samples were tested for the presence of RHDV-specific antibodies.

Results. VLPs with hemagglutination activity forming VLP 30–40 nm in size were obtained in Hi-5 cell culture. Specific antibody titers in rabbits measured by ELISA were 1 : 200 to 1 : 800 on 21th day post immunization with VLPs. Immunogenic activity of recVP60-GI1 VLPs was 90 and 40%, while it was 30 and 100% for recVP60-GI2 VLPs after the challenge with RHDV genotypes 1 and 2 respectively. The immunogenicity of two VLPs in mixture reached 100%.

Discussion. VLPs possess hemagglutinating, antigenic and immunogenic activity, suggesting their use as components in substances designed for RHDV specific prophylaxis in rabbits. Results of the control challenge experiment demonstrated the need to include the antigens from both RHDV genotypes in the vaccine.

Conclusion. Recombinant proteins recVP60-GI1 and recVP60-GI2 form VLPs that possess hemagglutinating an antigenic activity, and provide 90–100% level of protection for animals challenged with RHDV GI1 and GI2 virulent strains.

Problems of Virology. 2023;68(2):132-141
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Circulation of bovine herpesvirus (Herpesviridae: Varicellovirus) and bovine viral diarrhea virus (Flaviviridae: Pestivirus) among wild artiodactyls of the Moscow region

Pchelnikov A.V., Yatsenyuk S.P., Krasnikova M.S.


Introduction. Pestiviruses and viruses of the Herpesviridae family are widely distributed among different species of ungulates, but the main information about these pathogens is related to their effect on farm animals. Data on detection of bovine viral diarrhea virus (BVDV) and bovine herpes virus (BoHV) in wild ungulates reported from different countries in recent years raises the question of the role of wild animals in the epidemiology of cattle diseases.

Aim of work. To study the prevalence of herpesviruses and pestiviruses in the population of wild artiodactyls of the Moscow region.

Materials and methods. Samples of parenchymal organs and mucosal swabs from 124 wild deer (moose and roe deer) shot during hunting seasons 2019–2022 in Moscow Region were examined by PCR, virological and serological methods for the presence of genetic material and antibodies to bovine infectious rhinotracheitis and viral diarrhea.

Results. BVDV RNA was found in a sample from one moose, BoHV DNA was detected in samples from three roe deer and two moose shot in the Moscow region. Seropositive animals were of different sex and age, the total BoHVs and BVDV seroprevalence rates in wild artiodactyls were 46 and 29%, respectively.

Conclusion. Wild ruminant artiodactyls of the Moscow Region can be a natural reservoir of BoHV-1, and this must be taken into account when planning and organizing measures to control the infectious bovine rhinotracheitis. Cases of BVDV infection in wild artiodactyls are less common, so more research is needed to definitively establish their role in the epidemiology of this disease in cattle.

Problems of Virology. 2023;68(2):142-151
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Antiviral activity of basidial fungus Inonotus obliquus aqueous extract against SARS-CоV-2 virus (Coronaviridae: Betacoronavirus: Sarbecovirus) in vivo in BALB/c mice model

Shipovalov A.V., Kudrov G.A., Kartashov M.Y., Drachkova I.A., Pyankov O.V., Omigov V.V., Taranov O.S., Teplyakova T.V.


Introduction. The COVID-19 pandemic combined with seasonal epidemics of respiratory viral diseases requires targeted antiviral prophylaxis with restorative and immunostimulant drugs. The compounds of natural origin are low-toxic, but active against several viruses at the same time. One of the most famous compounds is Inonotus obliquus aqueous extract. The fruit body of basidial fungus I. obliquus is called Chaga mushroom.

The aim of the work ‒ was to study the antiviral activity of I. obliquus aqueous extract against the SARS-CoV-2 virus in vivo.

Materials and methods. Antiviral activity of I. obliquus aqueous extract sample (#20-17) was analyzed against strain of SARS-CoV-2 Omicron ВА.5.2 virus. The experiments were carried out in BALB/c inbred mice. The SARS-CoV-2 viral load was measured using quantitative real-time PCR combined with reverse transcription. The severity of lung tissue damage was assessed by histological methods.

Results. The peak values of the viral load in murine lung tissues were determined 72 hours after intranasal inoculation at dose of 2,85 lg TCID50. The quantitative real-time PCR testing has shown a significant decrease in the viral load compared to the control group by 4,65 lg copies/ml and 5,72 lg copies/ml in the lung tissue and nasal cavity samples, respectively. Histological methods revealed that the decrease in the number and frequency of observed pathomorphological changes in murine lung tissues depended on the introduction of the compound under study.

Conclusion. The results obtained indicate the possibility of using basidial fungus Inonotus obliquus aqueous extract as a preventive agent against circulating variants of SARS-CoV-2 virus.

Problems of Virology. 2023;68(2):152-160
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Virus-like particles based on rotavarus A recombinant VP2/VP6 proteins for assessment the antibody immune response by ELISA

Filatov I.E., Tsibezov V.V., Balandina M.V., Norkina S.N., Latyshev O.E., Eliseeva O.V., Cherepushkin S.A., Verkhovsky O.A., Grebennikova T.V.


Introduction. Rotavirus infection is one of the main concerns in infectious pathology in humans, mammals and birds. Newborn piglets or rodents are usually being used as a laboratory model for the evaluation of immunogenicity and efficacy for all types of vaccines against rotavirus A (RVA), and the use of ELISA for the detection of virus-specific antibodies of specific isotype is an essential step of this evaluation.

Objective. Development of indirect solid-phase ELISA with VP2/VP6 rotavirus VLP as an antigen to detect and assess the distribution of RVA-specific IgG, IgM and IgA in the immune response to rotavirus A.

Materials and methods. VP2/VP6 rotavirus VLP production and purification, electron microscopy, PAGE, immunoblotting, ELISA, virus neutralization assay.

Results. The study presents the results of development of a recombinant baculovirus with RVA genes VP2-eGFP/VP6, assessment of its infectious activity and using it for VLP production. The morphology of the VP2/VP6 rotavirus VLPs was assessed, the structural composition was determined, and the high antigenic activity of the VLP was established. VLP-based ELISA assay was developed and here we report results for RVA-specific antibody detection in sera of different animals.

Conclusion. The developed ELISA based on VP2/VP6 rotavirus VLP as a universal antigen makes it possible to detect separately IgG, IgM and IgA antibodies to rotavirus A, outlining its scientific and practical importance for the evaluation of immunogenicity and efficacy of traditional vaccines against rotavirus A and those under development.

Problems of Virology. 2023;68(2):161-171
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Nikolai Nikolaevich Nosik (04/07/1932–03/19/2023)

Problems of Virology. 2023;68(2):172-172
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In memory of an outstanding virologist

Mikhailov M.I., Anjaparidze A.G., Mammadov M.K.
Problems of Virology. 2023;68(2):173-174
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