Search

Extensive spread of tick-borne diseases poses a significant problem for public health and the health of the population living in endemic areas.The aim of the study was to search, analyze genetic material and identify new viral agents of the Phenuiviridae family in taiga ticks collected in Asian regions of Russia using the method of high throughput sequencing.Materials and methods. The study involved 1460 taiga ticks collected in suburban areas of the Tomsk, Yekaterinburg and Primorsky Territory. The genetic material isolated from ticks was sequenced using Illumina technology followed by phylogenetic analysis.Results and discussion. Analysis of the sequencing results made it possible to detect extended nucleotide sequences of the L-gene fragment characteristic of the Phenuiviridae family viruses. We were able to identify 20 nucleotide sequences the length of 250 bp on average in homogenates of Ixodes persulcatus ticks. Eighteen isolates have been identified as members of the genus Uukuvirus and two isolates have been assigned to the genus Phlebovirus, Phenuiviridae family. Phylogenetic analysis has shown that all isolates of the genus Uukuvirus fall under the cluster of Tacheng tick virus 2 belonging to the species Tacheng uukuvirus. They form a separate phylogenetic group which is closely related to two Romanian variants of 2019. Tacheng tick virus 2 was detected in all three surveyed regions of the Asian part of Russia. Two Tomsk isolates of phlebovirus were classified as Sara tick phlebovirus and they clustered with two isolates of phleboviruses from Karelia. Thus, the genetic material of Tacheng tick virus 2 and Sara tick phlebovirus belonging to two genera of the family Phenuiviridae was found in I. persulcatus ticks collected in three geographically different regions of the Asian part of Russia.

The south of Western Siberia traditionally belongs to the regions with unfavorable epidemiological situation for tick-transmitted infections. In Novosibirsk and its suburbs, approximately 20 thousand people annually seek for medical care after tick bites. The purpose of this work was to determine human-attacking tick species composition in the territory of Novosibirsk and its suburbs by identifying nucleotide sequence of tick mitochondrial gene cytochrome oxidase COI fragment during the 2018 spring-summer season, determination of infections of individual ticks with viral and bacterial tick-borne infections, followed by genotyping identified pathogens by PCR methods, by determining nucleotide sequences and phylogenetic analysis. It is established that ticks of five species attack the humans: Ixodes persulcatus, Ixodes pavlovskyi, Dermacentor reticulatus, Dermacentor marginatus, Dermacentor nuttali in the Novosibirsk metropolis. The majority of tick attacks on humans are associated with I. pavlovskyi (43.6%) and D. reticulatus (41.2%) ticks. The genetic material of the tick-borne encephalitis virus was detected in 3.6%, borrelia 13.8% and Rickettsia 23.1% individual ticks collected from humans. Genotyping of the tick-borne encephalitis virus (TBEV) in the collected ticks showed the presence of TBEV Siberian and Far Eastern genotype in 83% and 17% cases, respectively. The Kemerovo virus could not be detected. Genetic material from Borrelia spp. and Rickettsia spp. has been detected in all five tick species attacking the local human population. The bulk of the identified Borrelia spp. isolates were genotyped as Borrelia garinii (86%) and 13% of the isolates as Borrelia afzelii species. The genetic material of Borrelia miyamotoi was found in one individual tick. Ticks of the genus Dermacentor have been found to be infected by Rickettsia raoultii, and ticks of the genus Ixodes were largely infected with Rickettsia tarasevichiae, and I. persulcatus tick was infected by Rickettsia helvetica in a single case. Thus, the human population of Novosibirsk and its suburbs is attacked by five species of ixodid ticks dominated by I. pavlovskyi and D. reticulatus ticks. The genetic material of the following tick-borne pathogens was found: Siberian and Far Eastern TBEV genotype, B. garinii, B. afzelii, B. miyamotoi, R. raoulti, R. tarasevichiae and R. helvetica in the individual ticks collected from humans.

Objective of the study was to assess the level of infection with Borrelia miyamotoi in ticks Ixodes persulcatus and Ixodes pavlovskyi collected in the south of Western Siberia.Materials and methods. 688 ticks I. persulcatus and I. pavlovskyi collected on the territory of Novosibirsk, Tomsk and Kemerovo Regions were examined. Borrelia DNA was detected by a two-round polymerase chain reaction with specifc primers to the omp66 gene region, followed by sequencing of the isolated fragments and phylogenetic analysis, including the sequences of prototype isolates published in the international GenBank database. Comparison and analysis of nucleotide sequences was carried out using the MEGA 7 software package. In parallel, the samples were examined for the presence of the agents of transmissible infections – West Nile fever and tick-borne encephalitis, anaplasmosis, ehrlichiosis, babesiosis, bartonellosis and candidate pathogens of tick-borne rickettsiosis through RT-PCR and PCR with species-specifc and genus-specifc primers.Results and discussion. The genetic material of B. miyamotoi was found in 2.2 % of I. persulcatus and I. pavlovskyi ticks in the Tomsk, Novosibirsk, and Kemerovo Regions and their appurtenance to the Asian genotype was determined. It was revealed that the degree of similarity of the omp66 gene fragments within the group of isolates of the Asian genotype was 100 %. No relation between B. miyamotoi and a specifc vector species was identifed.

Determined is the complete genome sequence of Kolarovo-2008 strain (Siberia subtype) of Tick-borne encephalitis virus (TBEV), isolated from a tick in the suburbs of the Tomsk city. Nucleotide sequence analysis testifies of the fact that the level of genetic differences within the Siberian subtype of TBEV amounts to 10 % of the nucleotide sequence and to 7 % of amino-acid sequence for certain virus genes. 3'-HTO of the genome of Siberian subtype has the highest rate of variability and the homology level ranging from 65 to 97 %. Kolarovo-2008 and Vasilchenko (isolated in Novosibirsk in 1969) strains have the highest level of genome homology. The level of dissimilarity between the two Tomsk strains is substantially higher: the total number of amino-acid substitutions in Tomsk Zausaev and Kolarovo-2008 strains equals to 124, and 3'HTO level of homology is 79 %. Identified genetic variability of the Siberian subtype of TBEV is of a great importance for further development and enhancement of tick-borne encephalitis virus diagnostics.