Introduction. The aim of the study was to use comparative analysis for assessing efficiency of detection and confirmation of dual HIV infection, using conventional population sequencing (PS) and next generation sequencing (NGS) for an HIV-1 pol gene fragment, which encompasses protease and partially reverse transcriptase (positions 2253–3368).Materials and methods. The study was performed on intersubtype dual HIV infection model samples containing viruses of HIV-1 subtype B, sub-subtype A6 and recombinant form CRF63_02A1. Viruses were mixed pairwise in proportions from 10 to 90% to obtain 3 groups of model samples: CRF63vsB, CRF63vsA6, and A6vsB. The nucleotide sequences obtained by using PS and NGS technologies having 5, 10, 15, and 20% sensitivity thresholds for minor virus variants (NGS5–NGS20, respectively) were used to estimate the number of degenerate nucleotides or the degenerate base (DB) count and the number of synonymous mutations (SM) or the SM count. The fragment of the studied region (positions 2725–2981) was used for the analysis of operational taxonomic units.Results. The application of NGS5 proved highly efficient for detection of dual HIV infection in the model samples. The statistically significant (p < 0.01) increase in DB and SM counts was demonstrated by NGS5 compared to PS. As a result, NGS5 helped detect dual HIV infection in 25 out of 27 model samples, while with PS it was detected only in 15 samples. The analysis of operational taxonomic units confirmed dual HIV infection in all the groups of model samples.Discussion. The efficiency of detection and confirmation of dual HIV infection depends both on the content of each virus in the sample and on genetic characteristics of these viruses. Conclusion. Using NGS genetic testing in routine practice will be instrumental for efficient identification of genetic characteristics of infectious agents and for thorough analysis of the epidemiological situation.

The aim of the study: to analyze the prevalence of resistance mutations to elsulfavirine and to evaluate the effectiveness of it among HIV-infected treatment-naïve patients in real clinical practice.Materials and methods. The study included 578 patients with HIV infection, which divided into 3 groups. The first group is 354 HIV-infected treatment-naïve patients for whom HIV-1 nucleotide sequences were obtained as part of routine drug resistance testing. The second study group included 111 HIV-infected treatment-naïve patients, tested for drug resistance before the antiretroviral therapy containing elsulfavirine. The third study group included 113 HIV-infected treatment-naïve patients, each of whom was assigned a treatment regimen containing elsulfavirine without prior drug resistance testing. The observation period for patients of the second and third groups who received treatment was 24 weeks. To assess the effectiveness of antiretroviral therapy in patients, viral load, CD4+ T-cell counts, and adherence to therapy were assessed. HIV-1 subtypes and drug resistance mutations were determined using the Stanford HIV Resistance Database (v. 8.9-1). To clarify the results of subtyping, phylogenetic analysis of nucleotide sequences was carried out using the MEGA program (v. 6.0).Results. The prevalence of mutations associated with decreased susceptibility to elsulfavirine among HIV-infected treatment-naïve patients was 1.7% and 4.5% for the first and second groups of patients, respectively. All of the patients have only single resistance mutations which, according to the results of preclinical studies, cannot cause drug resistance. The use of elsulfavirine in real clinical practice among treatment-naïve patients has demonstrated good virological and immunological efficacy of the drug. As a result of 24 weeks of therapy in patients of the second group, no treatment ineffectiveness, and the development of drug resistance were observed. Among the patients of the third group, 6 patients (5.3%) have the virological failure of therapy associated with the resistance to the used drugs. All patients with virological failure had a resistance mutation profile associated with a high level of drug resistance to one of the drugs in the treatment regimen, lamivudine. Additionally, 1 patient had a combination of mutations that reduce susceptibility to elsulfavirine, and 4 patients had mutations that can reduce susceptibility to elsulfavirine in combination with other mutations.Conclusion. The low prevalence of mutations associated with a decrease in susceptibility to elsulfavirine and the absence of combinations of mutations make it possible to predict the successful use of this drug for Russian treatment-naïve patients. Reported cases of virological failure of antiretroviral therapy are difficult to interpret in the context of elsulfavirine due to the lack of an exact list of mutations and their combinations, and associations with the degree of resistance to it. This study describes for the first time the mutation profiles in patients with virological failure of therapy containing elsulfavirine and demonstrates the necessity of the further study of drug resistance profile to drug in vitro and in vivo.

Objective. Accurate identification of recent HIV-1 infection cases will ensure a more effective and precise assessment of the dynamics of virus transmission, the time between infection and diagnosis, and the quality of screening and prevention programs. This study was undertaken to adjust the recent HIV-1 infection testing algorithm using a cohort of patients, in whom the time since infection was known. Materials and methods. We used blood plasma samples obtained from 264 HIV-infected patients with a known date of infection. All samples were analyzed using two serological assays aimed to differentiate between cases of recent and established HIV infection. Using the results of sequencing of the pol region, we calculated the proportion of variable positions in order to determine the duration of infection. To identify the cases of recent HIV infection, we evaluated different variants of a diagnostic algorithm that included a combination of serological tests, molecular genetic analysis of the viral genome, and other clinical and laboratory parameters. Results. The effectiveness of the DS-ELISA-HIV-AB-TERM (DS) assay for the detection of recent infection was higher than that of the Architect HIV Ag/Ab Combo assay (Abbott). The sensitivity and specificity of the DS assay were 94.4% and 96.7%, respectively. The sensitivity and specificity of the Abbott assay were 86.4% and 77,4%, respectively. The HIV-1 genome variability threshold of 0.33% allowed the differentiation between samples depending on the time since infection with a cut-off of 12 months: 82.1% of recent samples and 62.7% of established samples were correctly identified using this method. We analyzed the effectiveness of schemes of the algorithm for the detection of recent infection lasting no longer than 9 months. Conclusion. Our findings allow us to recommend the algorithm based on the Russian DS assay for the detection of recent HIV-1 cases in routine clinical practice. This algorithm will enable the detection of new HIV cases, thereby improving the disease control. Key words: HIV-1, HIV infection, genetic variability, time since infection, duration of infection, recent infection, early infection, seroconversion

Aim: to carry out the comparative analysis of Russian kits for HIV-1 viral load (VL) measuring: AmpliSens HIV-monitor-FRT, AmpliSens HIV-monitor-Duo-FL, RealBest RNA HIV quantitative, HIV-GEN quantitative.Materials and methods: we analyzed the collection of clinical blood plasma samples with VL 590–2,05×106 copies/ml, dilutions of HIV-1 2nd International Standard, as well as panels of subtypes and recombinant forms of HIV-1 group M. The convergence of the results was determined using the Spearman correlation analysis and the Bland-Altman method.Results. A high level of convergence of VL measuring results in clinical samples (n=76) was determined for all compared kits with the value of approximation reliability R of at least 0,85. The maximum difference between the median values of VL results (0,48 log, ≈3-fold) was obtained for HIVMonitor-FRT and RealBest quantitative, which did not exceed the acceptable discordance rate in two kits comparing. All dilutions of the 2nd international HIV-1 standard were successfully determined by all kits with middle error 0,16–0,34 lg copies/ml. All kits successfully revealed the most of HIV-1 subtypes and recombinants studied with middle deviation 0,02–0,56 log copies/ml. HIV-GEN quantitative demonstrated the highest accuracy, but didn’t reveal ADG-recombinant.

Objective. To develop and validate the method of dual HIV infection detection based on the results of Sanger sequencing and to evaluate of dual HIV infection frequency in Russia. Materials and methods. We carried out the sequencing of HIV-1 pol gene fragment in model samples of dual HIV infection, the calculation of degeneracy bases index (DB-index), and the index of synonymity (SM-index) and determination of their threshold values for dual HIV infection detection. We analyzed nucleotide sequences of the pol gene fragment in HIV-1 samples (n = 1561) taken from Russian patients. Results. The threshold values of 34 for the DB-index and 0.05 for the SM-index allow detecting effectively the dual HIV infection using reverse transcriptase (RT) and protease-reverse transcriptase (PR-RT) fragments of the pol gene correspondingly. At total 21 (1.35%) dual HIV-infected samples (18 of which were collected in 2014 and later) were revealed in the collection studied; 12 of them were obtained from patients residing in the Central Federal District. An inverse correlation between the likelihood of dual HIV infection detecting and the duration of the patient's infection was found. In total, 11 out of 21 samples with dual HIV infection had mutations associated with drug resistance; 7 of them (33.3%) had substitutions that were not associated with natural polymorphism of HIV-1. Conclusion. The problem of dual HIV infection in Russia is actual and requires further study. Key words: HIV, dual HIV infection, drug resistance, nucleotide sequence, DB-index, SM-index, PR-RT region

Aim: to analyze the prevalence, structure of drug resistance and drug resistance mutations in the protease and reverse transcriptase genes of HIV-1 among treatment naïve patients.Materials and methods. We analyzed protease and reverse transcriptase sequences from 1560 treatment naïve HIV-infected patients from all Federal Districts of the Russian Federation with the first positive immune blot during 1998–2017. Sequences were analyzed for the presence of drug resistance mutations and predicted drug resistance to antiretroviral drugs using two algorithms — Stanford HIVDR Database (HIVdb) and the 2009 SDRM list (CPR).Results. The prevalence of drug resistance mutations was 11,1%. More often the prevalence of drug resistance was found for non-nucleoside reverse transcriptase inhibitor drugs (rilpivirine, nevirapine, efavirenz). The prevalence of transmitted drug resistance associated with mutations from the SDRM list was 5,3%, which is classified by the WHO as a moderate level. However, it should be noted that since the large-scale use of antiretroviral drugs in the Russian Federation, there has been a trend towards a gradual increase in the level of the transmitted drug resistance, and in 2016 it has already reached 6,1%.Conclusion. The results demonstrate the need for regular surveillance of the prevalence of HIV drug resistance to antiretroviral drugs among treatment naïve patients in the Russian Federation.