[Genetic diversity of group A rotavirus isolates found in Western Siberia in 2007-2011].

Genetic analysis of group A rotavirus recovered from fecal samples of children admitted to hospitals in Novosibirsk and Omsk during four epidemic seasons 2007, 2007/2008, 2009/2010, 2010/2011 was performed. A total of 1416 rotavirus isolates were genotyped using multiplex PCR. The isolates of the most common rotavirus genotypes G1P[8], G4P[8], G2P[4], G3P[8] co-circulated in Western Siberia during 2007-2011. In isolated cases G9P[8], G2P[8], G3P[9], and G4P[6] genotypes were detected. Change of dominant genotype from G1P[8] to G4P[8] occurred in 2008 in Omsk and in Novosibirsk in 2009 as well. Incidence and distribution of rotavirus genotypes differed and changed every epidemic season in both cities. The phylogenetic analysis based on VP4 (VP8*), VP7, and VP6 gene sequences showed that the majority of isolates from Novosibirsk and Omsk were clustered together and demonstrated high level homology with rotavirus isolates found in other regions of Eurasia. In addition, a rare P[8]b (OP354-like) subtype of the VP4 gene was identified in fourteen isolates (G9, G1, and G4) in Novosibirsk and in a single isolate Omsk08-381/G9P[8]b in Omsk. The results obtained in this study demonstrate the necessity of long-term monitoring of rotavirus isolates in Western Siberia. This is important for selection of rotavirus vaccine for immunization of infants, improvement of diagnostic kits and understanding of the epidemiology and the evolution of group A rotaviruses.

[Molecular genetic characteristics of astroviruses circulating in Novosibirsk].

A total of 1107 fecal samples from young children admitted to hospital for acute enteric infection in January to December 2007 were tested for astroviruses. Astroviruses were detected in 64 (5.8%) of the 1107 stool samples, only 50% of them were found as monoinfections. Astroviruses were recorded throughout the year; however, no seasonality for this infection could be ascertained. Cases of astrovirus infection were mainly observed in infants under one year of age (90%). Astroviruses were typed sequencing the ORF2 fragment; only HAstV-1 and HAstV-2 were found in Novosibirsk.

Characterization of the complete genome sequence of the recombinant norovirus GII.P16/GII.4_Sydney_2012 revealed in Russia.

Noroviruses (the Caliciviridae family) are a common cause of acute gastroenteritis in all age groups. These small non-envelope viruses with a single-stranded (+)RNA genome are characterized by high genetic variability. Continuous changes in the genetic diversity of co-circulating noroviruses and the emergence of new recombinant variants are observed worldwide. Recently, new recombinant noroviruses with a novel GII.P16 polymerase associated with different capsid proteins VP1 were reported. As a part of the surveillance study of sporadic cases of acute gastroenteritis in Novosibirsk, a total of 46 clinical samples from children with diarrhea were screened in 2016. Norovirus was detected in six samples from hospitalized children by RT-PCR. The identified noroviruses were classified as recombinant variants GII.P21/GII.3, GII. Pe/GII.4_Sydney_2012, and GII.P16/GII.4_Sydney_2012 by sequencing of the ORF1/ORF2 junction. In Novosibirsk, the first appearance of the new recombinant genotype GII.P16/ GII.4_Sydney_2012 was recorded in spring 2016. Before this study, only four complete genome sequences of the Russian GII.P16/GII.3 norovirus strains were available in the GenBank database. In this work, the complete genome sequence of the Russian strain Hu/GII.P16-GII.4/RUS/Novosibirsk/NS16-C38/2016 (GenBank KY210980) was determined. A comparison of the nucleotide and the deduced amino acid sequences showed a high homology of the Russian strain with GII.P16/GII.4_Sydney_2012 strains from other parts of the world. A comparative analysis showed that several unique substitutions occurred in the GII.P16 polymerase, N-terminal p48 protein, and minor capsid protein VP2 genes, while no unique changes in the capsid VP1 gene were observed. A functional significance of these changes suggests that a wide distribution of the strains with the novel GII.P16 polymerase may be associated both with several amino acid substitutions in the polymerase active center and with the insertion of glutamic acid or glycine in an N-terminal p48 protein that blocks the secretory immunity of intestinal epithelial cells. Further monitoring of genotypes will allow determining the distribution of norovirus recombinants with the polymerase GII.P16 in Russia.

[Rotaviruses in younger children in Novosibirsk in 2005-2007: detection and genotyping].

Examination of 1898 patients with acute enteric infection from March 2005 to February 2007 showed that group A rotaviruses were the most frequent cause (35%) of acute gastroenteritis among children under 3 years of age. Majority of cases of rotavirus infection was detected in infants under 1 year of age (71.8%). The peak of sporadic incidence was observed between February and May. High rate of mixed infection (45.6%) was observed - associations of rotaviruses with other viruses (noroviruses, astroviruses) and bacteria (Salmonella, Shigella, enteroinvasive Escherichia coli, Campylobacter, and opportunistic species) were detected. P- and G-genotypes of 337(50.8%) isolates of group A rotaviruses were determined by RT-PCR. The most prevalent strain was P[8]G1 (54.6%) followed by P[8]G3 (10.7%), P[8]G9 (8.6%), P[4]G2 (8.3%), and P[8]G4 (4.5%) genotypes.

[Construction and characterization of combinatorial naive phage library of human scFvs].

A combinatorial phage display library of human single-chain antibody fragments (scFv) was constructed on the basis of variable domains of heavy (Vh) and light (VI) genes cloned from the lymphocytes of six healthy donors. The size of the library was 2? 10(8) independent clones. Single-chain antibodies against recombinant human TNF?, vaccinia virus and virus-like particles formed by core protein of hepatitis B virus were selected from the library. Unique scFv sequences were identified using the HaeIII fingerprinting. The specificity of the selected clones was proved by the Western-blot analysis.

[Human recombinant antibodies to variola virus].

Eight specific antibodies to live variola virus (VV), Ind-3a strain, and 7 antibodies to VV, Butler strain, were selected from the synthetic combinatorial phage display library on single-chain (scFv) human antibodies. Indirect solid-phase enzyme immunoassay showed the ability of these antibodies to bind the VV strains Ind-3a, Butler, Brazil-131, Kuw-5, and Congo-2. Moreover, earlier selected human scFv antibodies were also tested in the reaction of binding to the above VV strains. The experiments could reveal the antibodies that bound alastrim strains more effectively that did other VV strains. The nucleotide sequences encoding for the selected scFv antibodies were determined.

[Human mini-antibodies to orthopoxviruses]. / Mini-antitela cheloveka protiv ortopoksvirusov.

A library of human scFv antibodies displayed on the surface of bacteriophages (MRC, Cambridge, England) was panned against the Elstree strain of vaccinia virus (VACV), which resulted in the phage repertoire enriched with clones positive to the strain. Individual clones from the repertoire were screened for binding, independently, to the vaccinia and ectromelia viruses; phage antibodies to the orthopoxviruses were selected. Ten unique antibodies were identified after their Vh- and Vl-genes were sequenced. All selected antibodies were assayed by ELISA for binding to the vaccinia, cowpox and ectromelia viruses. Furthermore, all selected antibodies were assayed for binding with major alastrim strains of the live variola virus. According to the results, the above phage antibodies recognized genus-specific epitopes, some of which differed in their conformation.