Igor V. Babkin

Evolutionary time-scale of primate bocaviruses

Human bocavirus (HBoV) is associated with acute gastroenteritis in humans, occurring mostly in young children and elderly people. Four bocavirus genotypes (HBoV1-HBoV4) have been found so far. Since there were no data on the contribution of HBoV to gastroenteritis in Russia, 1781 fecal samples collected from infants hospitalized with acute gastroenteritis in Novosibirsk, Russia during one year were tested for the presence of nucleic acids from HBoV and three major gastrointestinal viruses (rotavirus A, norovirus II, and astrovirus). HBoV was detected only in 1.9% of the samples: HBoV1 was detected in 0.6% and HBoV2, in 1.3%. Complete genome sequencing of three Novosibirsk isolates was performed. An evolutionary analysis of these sequences and the available sequences of human and great apes bocaviruses demonstrated that the current HBoV genotypes diverged comparatively recently, about 60-300years ago. The independent evolution of bocaviruses from chimpanzees and gorillas commenced at the same time period. This suggests that these isolates of great apes bocaviruses belong to separate genotypes within the species of human bocavirus, which is actually the primate bocavirus. The rate of mutation accumulation in the genome of primate bocaviruses has been estimated as approximately 9×10(-4)substitutions/site/year. It has been demonstrated that HBoV1 diverged from the ancestor common with chimpanzee bocavirus approximately 60-80years ago, while HBoV4 separated from great apes bocaviruses about 200-300years ago. The hypothesis postulating independent evolution of HBoV1 and HBoV4 genotypes from primate bocaviruses has been proposed.

A retrospective study of the orthopoxvirus molecular evolution.

The data on the structure of conserved genes of the Old and New World orthopoxviruses and unclassified Yoka poxvirus were used for a Bayesian dating of their independent evolution. This reconstruction estimates the time when an orthopoxvirus ancestor was transferred to the North American continent as approximately 50 thousand years ago (TYA) and allows for relation of this time interval with the global climate changes (with one of the short-term warmings during the Last Ice Age). The onset of the Yoka poxvirus evolution was assessed as approximately 90TYA. Availability of a large number of genome sequences of various cowpox virus strains provided for a comprehensive analysis of the orthopoxvirus evolutionary history. Such a study is especially topical in view of the postulated role of this virus in the evolution of various orthopoxviruses, namely, as an progenitor virus. The computations have demonstrated that the orthopoxviruses diverged from the ancestor virus to form the extant species about 10TYA, while the forbear of horsepox virus separated about 3TYA. An independent evolution of taterapox, camelpox, and variola viruses commenced approximately 3.5TYA. Study of the geographic distribution areas of the hosts of these three orthopoxviruses suggests the hypothesis on the region of their origin. It is likely that these viruses first emerged in Africa, in the region of the Horn of Africa, and that the introduction of camels to East Africa induced their divergent evolution.

High evolutionary rate of human astrovirus.

Human astrovirus is one of the etiological agents of acute gastroenteritis in humans, mostly in young children and elderly people. Complete genome sequencing of four human astrovirus strains isolated in Novosibirsk, Russia was performed. Analysis of these sequences and the sequences available in GenBank database has detected numerous potential recombination breakpoints. For the first time the rate of human astrovirus evolution was estimated based on the genome fragments without recombination breakpoints; the determined rate is typical of the RNA viruses with high evolutionary rate, amounting to approximately 3.7 × 10(-3) nucleotide substitutions per site per year, and for the synonymous changes, 2.8 × 10(-3) nucleotide substitutions per site per year.

The Origin of the Variola Virus

The question of the origin of smallpox, one of the major menaces to humankind, is a constant concern for the scientific community. Smallpox is caused by the agent referred to as the variola virus (VARV), which belongs to the genus Orthopoxvirus. In the last century, smallpox was declared eradicated from the human community; however, the mechanisms responsible for the emergence of new dangerous pathogens have yet to be unraveled. Evolutionary analyses of the molecular biological genomic data of various orthopoxviruses, involving a wide range of epidemiological and historical information about smallpox, have made it possible to date the emergence of VARV. Comparisons of the VARV genome to the genomes of the most closely related orthopoxviruses and the examination of the distribution their natural hosts’ ranges suggest that VARV emerged 3000 to 4000 years ago in the east of the African continent. The VARV evolution rate has been estimated to be approximately 2 × 10−6 substitutions/site/year for the central conserved genomic region and 4 × 10−6 substitutions/site/year for the synonymous substitutions in the genome. Presumably, the introduction of camels to Africa and the concurrent changes to the climate were the particular factors that triggered the divergent evolution of a cowpox-like ancestral virus and thereby led to the emergence of VARV.

Molecular Dating in the Evolution of Vertebrate Poxviruses

Objectives: The goal of this work was to study the evolutionary history of the vertebrate poxviruses using the Bayesian relaxed clock and a large set of highly conserved vitally important viral genes. Methods: Phylogenetic analysis was performed by the maximum likelihood method using the Paup program. The dating method of Bayes, realized in the Multidivtime, was made. Results: The rate of poxviral evolution is estimated as 0.5–7 × 10–6 nucleotide substitutions per site per year. We inferred that the modern viruses of the genus Avipoxvirus diverged from the ancestor nearly 249 ± 69 thousand years ago (Tya). The progenitor of the genus Orthopoxvirus separated approximately 166 ± 43 Tya. The separation of the forebear of the genus Leporipoxvirus took place about 137 ± 35 Tya. The next to diverge was the ancestor of the genus Yatapoxvirus. The progenitor of Capripoxvirus and Suipoxvirus diverged 111 ± 29 Tya. Conclusion: The evolutionary analysis based on the historical data and utilizing the Bayesian relaxed clock allowed us to determine the molecular evolution rates of the AT-rich genomes of the vertebrate poxviruses and assess the times of their emergences. Involvement of a large set of the conserved genes controlled by stabilizing selection allowed us to perform molecular dating of the vertebrate poxvirus history.

A study of the human bocavirus replicative genome structures.

The complete genomes of two human bocavirus 4 (HBoV4) isolates recovered in 2011 in Novosibirsk, Russia have been determined. A set of primers was designed based on the determined and previously published HBoV sequences; this primer pair was able to detect all possible HBoV replicative intermediates. This primer set was used to assay all HBoV genotypes and detected only those structures that correspond to an episomal form of this viral genome. Also, for the first time, head-to-tail nucleotide sequences have been determined for HBoV4. Secondary structures of the terminal noncoding regions (NCRs) of episomal forms have been computed for all HBoV genotypes, as well as for the canine bocavirus. Conserved secondary structures in episomal NCRs, which are likely to play an important part in the replication of bocaviruses, were found. NCR heterogeneity in the genomes of individual HBoV isolates has been shown for the first time.

Recombination in the evolution of human bocavirus

Whole genome sequencing of Novosibirsk human bocavirus (HBoV) isolates has detected an isolate that emerged via recombination between HBoV3 and HBoV4 genotypes. The recombination site is located between regions with abnormally low and abnormally high GC contents in the genome. This site is a bocavirus recombination hotspot and coincides with one of two parvovirus recombination hotspots. The Novosibirsk recombinant isolate, which is similar to a previously studied isolate from Thailand, utilizes the strategy of borrowing ORF3, which encodes structural proteins, of a rare genotype HBoV4. The role of recombination in HBoV evolution is discussed.

Recombination analysis based on the HAstV-2 and HAstV-4 complete genomes.

Complete genome sequences of previously unstudied human astrovirus subgenotypes - HAstV-2a and HAstV-2c - and two isolates of a rare genotype HAstV-4 have been determined. These isolates were recovered from fecal samples of young children hospitalized with acute intestinal infections in Novosibirsk (Russia). Three of the four sequenced isolates (HAstV-2a, HAstV-2c, and HAstV-4) are recombinants. It has been shown that all known HAstV-2 genomes have emerged via recombination; the HAstV-1 and HAstV-4 genotypes contain both recombinant and non-recombinant isolates; and all HAstV-3, HAstV-5, and HAstV-6 whole-genome sequences display no reliable signs of recombination. The average mutation accumulation rate has been determined based on an extended ORF2 fragment and amounts to 1.0×10(-3) substitutions per site per year. The evolutionary chronology of current HAstV genotypes has been reconstructed.

Detection and genetic characterization of a wide range of infectious agents in Ixodes pavlovskyi ticks in Western Siberia, Russia

BackgroundThe Ixodes pavlovskyi tick species, a member of the I. persulcatus/I. ricinus group, was discovered in the middle of the 20th century in the Russian Far East. Limited data have been reported on the detection of infectious agents in this tick species. The aim of this study was to investigate the prevalence and genetic variability of a wide range of infectious agents in I. pavlovskyi ticks collected in their traditional and recently invaded habitats, the Altai Mountains and Novosibirsk Province, respectively, which are both located within the Western Siberian part of the I. pavlovskyi distribution area.ResultsThis study reports the novel discovery of Borrelia bavariensis, Rickettsia helvetica, R. heilongjiangensis, R. raoultii, “Candidatus Rickettsia tarasevichiae”, Anaplasma phagocytophilum, Ehrlichia muris, “Candidatus Neoehrlichia mikurensis” and Babesia microti in I. pavlovskyi ticks. In addition, we confirmed the previous identification of B. afzelii, B. garinii and B. miyamotoi, as well as tick-borne encephalitis and Kemerovo viruses in this tick species. The prevalence and some genetic characteristics of all of the tested agents were compared with those found in I. persulcatus ticks that were collected at the same time in the same locations, where these tick species occur in sympatry. It was shown that the prevalence and genotypes of many of the identified pathogens did not significantly differ between I. pavlovskyi and I. persulcatus ticks. However, I. pavlovskyi ticks were significantly more often infected by B. garinii and less often by B. bavariensis, B. afzelii, “Ca. R. tarasevichiae”, and E. muris than I. persulcatus ticks in both studied regions. Moreover, new genetic variants of B. burgdorferi (sensu lato) and Rickettsia spp. as well as tick-borne encephalitis and Kemerovo viruses were found in both I. pavlovskyi and I. persulcatus ticks.ConclusionAlmost all pathogens that were previously detected in I. persulcatus ticks were identified in I. pavlovskyi ticks; however, the distribution of species belonging to the B. burgdorferi (sensu lato) complex, the genus Rickettsia, and the family Anaplasmataceae was different between the two tick species. Several new genetic variants of viral and bacterial agents were identified in I. pavlovskyi and I. persulcatus ticks.

Occurrence and genetic variability of Kemerovo virus in Ixodes ticks from different regions of Western Siberia, Russia and Kazakhstan.

Kemerovo virus (KEMV), a member of the Reoviridae family, Orbivirus genus, is transmitted by Ixodes ticks and can cause aseptic meningitis and meningoencephalitis. Recently, this virus was observed in certain provinces of European part of Russia, Ural, and Western and Eastern Siberia. However, the occurrence and genetic diversity of KEMV in Western Siberia remain poorly studied. Therefore, the aim of this work was to investigate the prevalence and genetic variability of KEMV in Ixodes ticks from Western Siberia. A total of 1958 Ixodes persulcatus, I. pavlovskyi ticks and their hybrids from Novosibirsk and Omsk provinces, Altai Republic (Russia) and East Kazakhstan province (Kazakhstan) were analyzed for the presence of KEMV and tick-borne encephalitis virus (TBEV) RNA. It was observed that the KEMV distribution area in Western Siberia was wider than originally thought and included Northern and Northeastern Altai in addition to the Omsk and Novosibirsk provinces. For the first time, this virus was found in Kazakhstan. The occurrence of KEMV was statistically lower than TBEV in most locations in Western Siberia. KEMV was found both in I. persulcatus and I. pavlovskyi ticks and in their hybrids. Notably, KEMV variants observed in the 2010s were genetically different from those isolated in the 1960s, which indicated the ongoing process of evolution of the Kemerovo virus group. Moreover, the possibility of reassortment for KEMV was demonstrated for the first time.