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Introduction: Parenteral viral hepatitis (B, C, D) and HIV share modes of transmission and risk groups, in which the probability of infection with two or more of these viruses simultaneously is increased. Mutual worsening of the course of viral infections is important issue that occurs when HIV positive patients are coinfected with parenteral viral hepatitis. The aim of the study was to determine the prevalence of HCV, HBV and HDV in HIV positive patients in the Novosibirsk region and to give molecular genetic characteristics of their isolates., Materials and Methods: Total 185 blood samples were tested for the presence of total antibodies to HCV, HCV RNA, HBV DNA and HDV RNA. The identified isolates were genotyped by amplification of the NS5B gene fragment for HCV, the polymerase gene for HBV and whole genome for HDV., Results: The total antibodies to HCV were detected in 51.9% (95% CI: 44.758.9), HCV RNA was detected in 32.9% (95% CI: 26.639.5) of 185 studied samples. The distribution of HCV RNA positive cases completely repeated the distribution of HCV serological markers in different sex and age groups. The number of HCV infected among HIV positive patients increases with age. HCV subgenotypes distribution was as follows: 1b (52.5%), 3а (34.5%), 1а (11.5%), 2а (1.5%). 84.3% of detected HCV 1b isolates had C316N mutation associated with resistance to sofosbuvir and dasabuvir. The prevalence of HBV DNA in the studied samples was 15.2% (95% CI: 10.721.0). M204I mutation associated with resistance to lamivudine and telbivudine was identified in one HBV isolate. Two HDV isolates that belonged to genotype 1 were detected in HIV/HBV coinfected patients., Conclusion: The data obtained confirm the higher prevalence of infection with parenteral viral hepatitis among people living with HIV in the Novosibirsk region compared to the general population of that region. The genetic diversity of these viruses among HIV infected individuals is similar to that observed in the general population.

Hepatitis D (delta, δ) virus (HDV) was discovered more than 40 years ago, but the understanding of its origin and evolution is poor. This is mainly due to the lack, until recently, of data on the existence of any viruses similar to HDV. The discovery in recent years of sequences of new delta-like agents in a wide range of vertebrate (Vertebrata) and invertebrate (Invertebrata) species has facilitated a revision of views on the origin of HDV and contributed to understanding the place of this unique virus among other animals' viral agents. The purpose of this review is to analyze the latest published data on new delta-like agents and their biological characteristics.

Introduction: Hepatitis D (delta, 5) is caused by an RNA virus (hepatitis D virus, HDV) from genus Deltavirus, and is the most severe and difficult to treat disease among both viral hepatitis and infectious diseases in general. The development of HDV infection in the host organism is possible only in the presence of hepatitis B virus (HBV). Coinfection with HBV and HDV is associated with a more rapid progression of chronic viral hepatitis (CVH) to liver cirrhosis (LC) and an unfavorable outcome in comparison with HBV monoinfection. Data on the influence of clinical, biochemical and virological factors on the infectious process in patients with hepatitis D are limited due to the insufficient amount of research on this theme.The study aimed to determine demographic, clinical, biochemical, and virological factors influencing the course and progression of CVH D in patients followed during 10 years, residing in the territory of the Tuva Republic, one of the endemic regions of the Russian Federation., Material and Methods: Changes in clinical and laboratory parameters were analyzed in dynamics in 121 HDV infected patients with a different course of the disease, who were under observation from 2009 to 2019. Three groups of patients were identified: group 1 - 61 patients with disease progression of chronic hepatitis to LC (Child-Pugh class B-C), group 2 - 49 patients with non-progressive chronic hepatitis, and group 3 - 11 patients with slowly progressive LC (class A). Demographic data, the presence of detectable HBV DNA, indicators of the functional state of the liver: alanine aminotransferase (ALT/GPT), aspartate aminotransferase (AST/GOT), alkaline phosphatase (ALP), gamma-glutamyltransferase (GGT), and total bilirubin content were analyzed. The severity of hepatic encephalopathy was assessed by the duration of the numbers connection test (NCT)., Results: All patients belonged to the same ethnic group (Tuvinians), were infected with HDV genotype 1 and were positive for HDV RNA throughout the entire follow-up period. There were no significant differences in sex ratio and mean age at the time of inclusion in the study between the groups. In group 1, the average number of years from inclusion in the study to the formation of LC was 3.65 ± 2.3 years, years to the lethal outcome: 4.5 ± 3 years. Significantly higher levels of AST/GOT, ALP, GGT, total bilirubin (TB) and NCT grade were found in group 1 compared to group 2. ALT/GPT levels did not differ significantly in these groups. When comparing groups with disease progression and slowly progressive LC (groups 1 and 3), no significant differences were found in any of the clinical and biochemical parameters. ALT/GPT, GGT, TB and NCT values were significantly higher in patients with slowly progressive LC (group 3) compared to group 2. No differences in AST/GOT and ALP levels were found between these groups. Detectable HBV DNA was significantly more frequent in patients with progressive disease and with chronic viral hepatitis than in patients with slowly progressive LC. There were no significant differences in the frequency of HBV DNA detection in patients from groups 1 and 2., Conclusion: The results obtained on a relatively homogeneous cohort demonstrated that age and gender are not the factors influencing the progression of chronic viral hepatitis D to cirrhosis. The lack of detectable HBV DNA is associated with the slow progression of LC. The revealed differences in clinical and biochemical parameters reflect the degree of functional liver damage in chronic viral hepatitis D and HDV-associated cirrhosis.

Association of RNA molecules forming a two-component B:LS trans-analog of antigenomic HDV ribozyme was studied. From previously synthesized trans-ribozymes the B:LS ribozyme differs by length and sequence of its RNA molecules (33 and 34 bp, respectively), topology of functional parts and the absence of very short reaction product. The ribozyme displays a biphasic kinetics of self-cleavage similar to that of cis-ribozyme. Our original kinetic scheme for the B:LS trans-ribozyme self-cleavage (www.cardio.ru\labgen\RZ_e.html)describes a possible cause of biphasic nature of the reaction curve, namely, variation of the rate-limiting stage in the series of successive conformational transformations which coincide with the ribozyme self-cleavage. Interactions between the molecules involved in the reaction, i.e., "multimerization" of entire ribozyme and its components can be regarded as another cause of the biphasic kinetics. B:LS trans-ribozyme is a convenient model for the investigation of this process, since the binding of LS and B allows the formation of complexes with 1B:2LS or 2B:1LS stoichiometry and complexes with the cleavage products. We examined the factors determining dissociation-association of the ribozyme components using a series of electrophoreses under nondenaturing conditions. The possibility of interaction between cis- and transribozyme components was confirmed experimentally. In the presence of LS excess over B the ribozyme can form multimeres. These findings suggest the involvement of intermolecular interactions in native cis-ribozyme self-cleavage.

B : LS ribozyme, a trans-variant of naturally occurring HDV ribozyme, has been constructed. The ribozyme consists of a substrate-containing LS chain and an enzyme B chain and differs from previously constructed trans-ribozymes in the length and nucleotide sequence of its oligonucleotide chains (34 and 33 bp, respectively). The chains readily associate with each other at a room temperature while the LS cleavage reaction at this temperature is negligible slow, which allowed us to investigate the association of the intact chains. At the same time the self-cleavage rate constant for the trans-ribozyme B : LS at 50 degrees C is close to those for the previously studied permuted cis-ribozymes, especially LSB variant. In addition, the dependence on the reaction conditions (Mg2+ concentration, pH, temperature) of the trans-ribozyme was similar to that of cis-ribozyme. Similar to other trans-ribozymes, B : LS ribozyme demonstrates the ability for multiple use of the enzyme B-chain with an excess of the substrate LS chain. The kinetics model of self-cleavage reaction for B : LS is presented in http://www.cardio.ru/labgen/RZ_r.html. Taken together, our results show that the original trans-variant of HDV ribozyme can be used as a model for the investigation of self-cleavage process of HDV ribozymes.

Internal symmetry of the mirror type has been first found in molecules of RNA referred to as ribozymes. For identification and investigation of the internal symmetry in RNA primary structure, two methods were developed, dot matrix and scanning, respectively. The methods are based on a comparison of normal and reversible nucleotide sequences. The objects of our study was ribozymes from introns of Tetrahymena thermophila and hepatitis delta virus, and also a group of related ribozymes, possessing both cleavage and ligation activities. The centers of internal symmetry are mainly localized in the catalytic sites and other important regions of ribozyme molecules. A positive correlation was found between the conservativity and symmetry of the primary structure of ribozymes. However, this correlation was not so clear in comparison with the correlation in the case of proteins. As reported earlier (Shpakov, 1995, 2001), the internal symmetry was discovered in protein and DNA molecules. The obtained data enable us to consider the internal symmetry as a common characteristics of nucleotide and amino acid sequences of the biopolymers.

Nine peptides from immunodominant (53-68 and 65-80 aa) and hypervariable (201-213 aa) regions derived from delta antigen sequences corresponding to 3 HDV genotypes were synthesized. Type specificity of antibodies to the resultant peptides was evaluated by indirect enzyme immunoassay with sera from patients with hepatitis D and asymptomatic carriers of anti-delta antibodies. Analysis of the results showed that HDV circulating in the Central Volga region belongs to type I in the majority of cases. High heterogeneity and changeability of HDV genome during the period of transition from acute forms to asymptomatic carriership was revealed. Possibility of circulation of new hybrid HDV strains causing mixed infection in Russia is discussed.

The kinetic characteristics have been studied for noncircularly permuted variants of the human hepatitis delta virus (HDV) antigenomic ribozyme to find out the cause of the two-phase kinetics of the self-cleavage reaction. Different ways of reaction initiation, suboptimal conditions, and jumpwise changes of reaction conditions have been used, and the temperature dependences have been studied. A correlation has been shown between the apparent kinetic constant of the first reaction phase and the portion of the ribozyme molecules that self-cleaved during the first phase. Partial restoration of the initial reaction characteristics has been shown by the reinitiation of reaction being stopped after completing the first phase. On the basis of all the data obtained, a scheme of the self-cleavage reaction has been proposed including: (i) activation of the ribozyme with energy of 40-50 kcal/mol and a characteristic time of several deciminutes under optimal reaction conditions; (ii) fast and reversible reaction of the phosphodiester bond cleavage; (iii) reaction leading to isomerization of the 3',5'-phosphodiester bond to the 2',5' bond in the self-cleavage site with a characteristic activation time of tens of minutes; and (iv) practically irreversible conformational change leading to fixation of the cleavage by immobilization of the 5'-terminal nucleotide of the product in the center of the formed structure and displacement of the 3'-terminal nucleotide to the periphery. The latter process has a characteristic time of tens of minutes and a low activation energy.

Six synthetic peptides from immunodominant region 65-80 aa derived from the sequences of delta-antigen corresponding to 3 HDV genotypes were obtained. Their antigenic activities were assessed by indirect enzyme immunoassay with sera of patients with hepatitis D and of asymptomatic carriers of anti-delta antibodies. Differences in antigenic characteristics of full-size 65-80 and truncated 71-80 aa peptides corresponding to three HDV genotypes were revealed. The diagnostic significance of synthetic 65-80 aa fragments is shown.

Genome RNA of hepatitis Delta virus (HDV) was isolated from the blood sera of 4 independently infected patients. The genome fragment of 901-1265 was obtained for each isolate by the RT-PCR. The resultant sequences showed that all isolates belonged to HDV genotype I. Russian isolates were shown to be closely related to an Italian one (98.1-97.5% and 97.1-95.1% identity for the nucleic acid and amino acid levels, respectively); they may represent an extra subtype within genotype I. No appreciable differences between the Russian strains were found, which fact may reflect a possible homogeneity of HDV population in North-Western Russia.

A set of 5 peptides from the immunodominant region (65-80 aa) of delta-antigen was prepared by solid phase synthesis. Peptide 71-80 was synthesized in two variants with different amino acid residues in positions 73, 74, 76. Free peptides and their conjugates with bovine serum albumin were tested for antigenicity in ELISA. Correlation between the length of the peptide chain and its antigenic activity was noted. Peptides 65-80 and 69-80 demonstrated positive reaction with all sera from acute HDV patients. Both variants of peptide 71-80 reacted with 83% of positive sera. The smaller peptide 73-80 reacted strongly with one positive serum out of five used.