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Gene therapy is considered a promising approach to treating infections caused by human immunodeficiency virus (HIV). One strategy is to introduce antiviral genes into cells in order to impart resistance to HIV. In this work, the antiviral activity of new anti-HIV lentiviral vector pT has been studied. The vector carries a combination that consists of two identical artificial miRNA mic13lg and the TRIM5α-HRH gene. Two mic13lg microRNAs suppress the expression of the CCR5 gene, which encodes the HIV coreceptor and, thus, prevents the penetration of R5-tropic HIV strains into the cell. It has been shown that pT effectively inhibits the expression of CCR5 in both the HT1080 CCR5-EGFP model cell line and in human primary lymphocytes. The second line of protection against R5- and X4-tropic HIV is provided by the TRIM5α-HRH protein, which binds virus capsids after the virus enters the cell. Indeed, when infecting cells of the SupT1 line, which contains four copies of the vector per cell, with the X-4 tropic HIV, more than 1000-fold suppression of viral replication has been observed. The process of generation of the pT vector and conditions of transduction of CD4+ lymphocytes were optimized for testing the antiviral activity of the vector on primary human lymphocytes. As a result, the transduction efficiency for the pT vector was 28%. After infection with the R5-tropic strain of the virus, the survival of cells in the culture of lymphocytes with the vector was significantly higher than in the control. However, the complete suppression of HIV replication was not achieved, presumably due to the inadequate fraction of cells that carry the vector in culture. In the future, it is planned to find the best way to enrich the lymphocyte culture with modified cells to increase resistance to HIV. [ABSTRACT FROM AUTHOR]

Obtaining a pure recombinant Modified Vaccinia Ankara (MVA) virus is a multistage, time-consuming procedure. We describe a novel single-tube real-time PCR which enables determination of the amount of wild type and recombinant viruses and their ratio in plaques. Use of the real-time PCR significantly reduce the time and efforts needed to obtain purified recombinant MVA. The new approach has been applied to generate recombinant MVAs encoding different SARS-COV-2 antigens., (Copyright © 2020 Elsevier B.V. All rights reserved.)

To define frequencies of drug resistance mutations among HIV-1 variants circulating within the territory of Russia, subtype A HIV-1 nucleotide sequences encoding protease and reverse transcriptase were analyzed. The analysis was carried out in 141 antiretroviral-naive individuals. Low frequency (less than 1%) of primary drug resistance mutations was shown. However, high frequencies of secondary mutations V77I in protease and A62V in RT (67% H 63%, respectively) linked to each other in most cases were observed. The HIV-1 isolates bearing both substitutions (MutV77I/A62V) were also characterized by the presence of several synonymous mutations, suggesting common origin for these viruses. HIV Biochip Hybridization microarray and/or Restriction fragment-length polymorphism analyses were performed to characterize gene pol polymorphism in additional 178 subtype A HIV-1 isolates. Among total 319 samples studied, Mutv77IA62V variant accounted for 56%, and was found to predominate in Russia in terms of both its geographical distribution and number of cases caused. Moreover, these viruses were prevalent in the regions known to have highest incidence of HIV-1 infection (Irkutsk, Samara, and Moscow regions). In addition, three other variants were found: viruses not containing the substitutions V77I or A62V, and variants bearing only one of them. Evolutional relationships between all four HIV-1 variants, as well as potential impact of the gene pol polymorphism on HIV-1 replicative fitness and drug resistance development are discussed.