Yulia A. Vakulenko

Bovine viral diarrhea virus (BVDV) is a pathogen of globally significance in cattle that has two biotypes: non-cytopathogenic (non-cp) and cytopathogenic (cp). The cp biotype arises from the non-cp through genome rearrangements, which frequently involve the insertion of host cellular RNA sequences, often accompanied by viral genome duplications. Here, we performed a systematic analysis of insertions across all complete BVDV genomes available in GenBank. Despite a 10-fold increase in available sequences over the past 10 years (from 59 to 670), the repertoire of known rearrangements associated with the cp phenotype has expanded only modestly, with insertions occurring predominantly at five conserved genomic hotspots. Notably, independent acquisitions of similar insertions—such as DNAJC14 (Jiv), ubiquitin-like sequences, the NS4B-NS5A cleavage site, and the PYPDPQTLG motif—in phylogenetically unrelated virus lineages reflect a limited number of permissive sites. Our analysis further demonstrates that many insertions are mosaic and suggests multiple recombination events that are poorly compatible with template switching by viral polymerase. Mechanistically, we propose that non-replicative recombination provides the most consistent explanation for the observed insertion patterns, particularly common coupling of cellular RNA acquisitions with viral sequence duplications. Both the steric proximity of virus replication to the endoplasmic reticulum and the RNA end requirements (2′-3′ cyclic phosphate and 5′-OH) suggest that non-replicative recombination is likely mediated by the endoplasmic reticulum-associated IRE1 RNase and RtcB ligase, which are responsible for the alternative cytoplasmic splicing of cellular mRNA.

Human astroviruses are small, non-enveloped RNA viruses belonging to the family Astroviridae. Among the four species known to infect humans, the species Mamastrovirus hominis (the classical human astroviruses, formerly MAstV1) is associated with gastrointestinal illness worldwide, while three more recently identified species have been linked to lethal central nervous system infections. High substitution rates and recombination drive their rapid evolution, yet recombination patterns in classical human astroviruses remain poorly characterized. This study systematically analyzes patterns and temporal dynamics of natural recombination in classical human astroviruses. Publicly available genomes of classical human astroviruses were analyzed to identify recombination hotspots. Recombinant forms were defined as stable phylogenetic lineages unaffected by recombination, and their half-lives were estimated based on time-scaled phylogenies (BEAST2v2.7.7). Recombination in classical human astroviruses occurred most frequently at the ORF1b/ORF2 junction, but also within ORF1a, at the ORF1a/ORF1b junction, and within ORF2. Only the 3′-part of ORF1a and a fragment of ORF1b exhibited robust temporal signal, yielding substitution rates of 2.35 × 10⁻³ and 2.14 × 10⁻³ s/s/y, respectively. The half-lives of recombinant forms varied considerably by genomic region: longest for exchanges between the parts of ORF1a (21 years), intermediate for ORF1a/ORF1b recombinants (7–9 years), and shortest for ORF1ab/ORF2 recombinants (2.5–3.6 years). The estimated half-lives for recombinants align with those reported for human enteroviruses and noroviruses. These findings highlight the dynamics of the generation of astrovirus diversity and may inform advanced surveillance of emerging strains.

Foot-and-mouth disease virus (FMDV) is a highly contagious pathogen of cloven-hoofed livestock. Recombination is one of the mechanisms that contribute to genetic diversity of FMDV and facilitate the generation of new viral lineages, or recombinant forms. While the general patterns of recombination in FMDV are well-known, the temporal dynamics of this process remain unexplored. This study systematically analyzed recombination across 1485 publicly available complete genome sequences of FMDV, collected from 1934 to 2024. In addition to the well-known general recombination pattern with hotspots on the borders of the genome region that encodes capsid proteins VP2-VP3-VP1, we identified serotype-specific recombination patterns. A significant temporal signal required to analyze temporal dynamics was found in serotypes A, Asia1, O, and SAT1 in the VP2-VP3-VP1 genome region. To assess the lifetimes of FMDV recombinant forms, we compared these time-scaled phylogenetic trees with phylogenies for other genomic regions exchanged by recombination events. The median lifetimes of FMDV recombinant forms ranged from 2 to 18 years, depending on the serotype and the nonstructural genomic region involved in recombination. These timescales are comparable to human (+)RNA viruses, such as enteroviruses and caliciviruses. In distinct serotypes, recombination could be more frequent on the 5′ or 3′ border of the capsid-encoding genome region, without a uniform pattern.

Metaviromic studies of potential emerging infection reservoirs led to discovery of many novel viruses. Since metaviromes contain viruses from target host, its food or other sources, fast and robust approaches are needed to predict hosts of unknown viruses based on their genome data. Four machine learning algorithms (random forest, two gradient boosting machines, support vector machine) were used here to predict the hosts of RNA viruses that infect mammals, insects and plants. The prediction efficiency was largely dependent on the dataset composition. In the more challenging task of predicting hosts of unknown virus genera, median weighted F1-score of 0.79 was achieved using support vector machine and 4-mer frequencies, a notable improvement over baseline methods (median weighted F1-scores 0.68 for the homology-based tBLASTx and 0.72 for ML trained on mono-, di- and trinucleotide frequencies). More complicated features and feature combinations provided worse results. When predicting hosts of short virus sequence fragments quality decreased but using same-length fragments instead of full genomes for training consistently produced an improvement of prediction quality. Therefore, short k-mers carry sufficient information to predict hosts of novel RNA virus genera. This algorithm can be useful in rapid analysis of metaviromic data to highlight potential biological threats.

Mammalian glutamate dehydrogenase (GDH) is an indispensable metabolic enzyme. GDH duplication has led to the presence of two paralogs, GDH1 and GDH2, in apes. Multiple GDH pseudogenes are also present in the human genome. The novel GDH2, supposed to be a target of positive selection, differs from GDH1 in regulation and is believed to be tightly linked to brain development. Although the differences of modern human GDH2 from GDH2 of other apes have been studied, the evolution of ancient human GDH2 remains a blank space. The goal of this work was to elucidate GDH2 evolution in the genus Homo using the accumulated data on the ancient genomes with high coverage—three Neanderthal and one Denisovan genome. Such analysis clarifies the difference between GDH2 of the last common ancestor of humans and chimpanzees and all Homo to be in M468L substitution, localized in the regulatory “antenna” region of the protein. A few novel missense mutations have been found in Denisovan and Altai Neanderthal GDH2, namely R76H, present in both genomes, and Denisovan-specific T154P, I358L, and S498A substitutions. Another mutation, R352K, has likely occurred independently in modern humans and later Neanderthals. The potential impact of these mutations was estimated using GDH2 structural data and evidence from contemporary medical data. All substitutions are supposed to be benign, with only the S498A GDH2 substitution connected to Parkinson’s disease with late onset. Additionally, the ancient genomes were revealed to have all GDH pseudogenes present in modern humans, including the RNA-coding ones. The GLUD1P3 RNA expression was found to correlate negatively with GDH1 in human tissues. A possible regulatory role has been proposed, and the GLUD1P3 RNA sequence identity in all the studied human genomes suggests its conservation in the genus Homo.

Orthoflaviviruses are arthropod-borne viruses that are transmitted by mosquitoes or ticks and cause a range of significant human diseases. Among the most important tick-borne orthoflaviviruses (TBFVs) is tick-borne encephalitis virus (TBEV), which is endemic in Eurasia, and Powassan virus, which is endemic in Asia and North America. There is a significant controversy regarding species assignment in the tick-borne encephalitis virus complex due to the complex phylogenetic, serological, ecological, and pathogenetic properties of viruses. Comparing the rate of non-synonymous to synonymous substitutions (dN/dS) over the course of tick-borne orthoflavivirus diversification suggests that there is a very strong stabilizing selection (Nei-Gojobori dN/dS < 0.1) among tick-borne orthoflaviviruses that differ by less than 13.5% amino acid/21.4% nucleotide sequences, and discretely more rapid accumulation of non-synonymous substitutions (dN/dS > 0.13) among more divergent viruses that belong to distinct species. This pattern was similarly observed in genome regions encoding structural (E) and non-structural (NS3) proteins. Below this distance threshold, viruses appear fit and strongly tied to their ecological niche, whereas above the threshold, a greater degree of adaptation appears necessary. This species criterion suggests that all subtypes of TBEV, all related ovine/caprine encephalomyelitis viruses, and Omsk hemorrhagic fever virus (OHFV) together correspond to a single species. Within this species, viruses make up 11 subtypes that are reliably segregated by a 10% nucleotide distance cut-off suggested earlier for TBEV. The same 10% subtype cut-off suggests that Powassan virus includes two subtypes, Powassan and Deer Tick virus.

There are two paralogs of glutamate dehydrogenase (GDH) in humans encoded by the GLUD1 and GLUD2 genes as a result of a recent retroposition during the evolution of primates. The two human GDHs possess significantly different regulation by allosteric ligands, which is not fully characterized at the structural level. Recent advances in identification of the GDH ligand binding sites provide a deeper perspective on the significance of the accumulated substitutions within the two GDH paralogs. In this review, we describe the evolution of GLUD1 and GLUD2 after the duplication event in primates using the accumulated sequencing and structural data. A new gibbon GLUD2 sequence questions the indispensability of ancestral R496S and G509A mutations for GLUD2 irresponsiveness to GTP, providing an alternative with potentially similar regulatory features. The data of both GLUD1 and GLUD2 evolution not only confirm substitutions enhancing GLUD2 mitochondrial targeting, but also reveal a conserved mutation in ape GLUD1 mitochondrial targeting sequence that likely reduces its transport to mitochondria. Moreover, the information of GDH interactors, posttranslational modification and subcellular localization are provided for better understanding of the GDH mutations. Medically significant point mutations causing deregulation of GDH are considered from the structural and regulatory point of view.

The 2023 International Virus Bioinformatics Meeting was held in Valencia, Spain, from 24–26 May 2023, attracting approximately 180 participants worldwide. The primary objective of the conference was to establish a dynamic scientific environment conducive to discussion, collaboration, and the generation of novel research ideas. As the first in-person event following the SARS-CoV-2 pandemic, the meeting facilitated highly interactive exchanges among attendees. It served as a pivotal gathering for gaining insights into the current status of virus bioinformatics research and engaging with leading researchers and emerging scientists. The event comprised eight invited talks, 19 contributed talks, and 74 poster presentations across eleven sessions spanning three days. Topics covered included machine learning, bacteriophages, virus discovery, virus classification, virus visualization, viral infection, viromics, molecular epidemiology, phylodynamic analysis, RNA viruses, viral sequence analysis, viral surveillance, and metagenomics. This report provides rewritten abstracts of the presentations, a summary of the key research findings, and highlights shared during the meeting.

Noroviruses infect a wide range of mammals and are the major cause of gastroenteritis in humans. Recombination at the junction of ORF1 encoding nonstructural proteins and ORF2 encoding major capsid protein VP1 is a well-known feature of noroviruses. Using all available complete norovirus sequences, we systematically analyzed patterns of natural recombination in the genus Norovirus both throughout the genome and across the genogroups. Recombination events between nonstructural (ORF1) and structural genomic regions (ORF2 and ORF3) were found in all analyzed genogroups of noroviruses, although recombination was most prominent between members of GII, the most common genogroup that infects humans. The half-life times of recombinant forms (clades without evidence of recombination) of human GI and GII noroviruses were 10.4 and 8.4–11.3 years, respectively. There was evidence of many recent recombination events, and most noroviruses that differed by more than 18% of nucleotide sequence were recombinant relative to each other. However, there were no distinct recombination events between viruses that differed by over 42% in ORF2/3, consistent with the absence of systematic recombination between different genogroups. The few inter-genogroup recombination events most likely occurred between ancient viruses before they diverged into contemporary genogroups. The recombination events within ORF1 or between ORF2/3 were generally rare. Thus, noroviruses routinely exchange full structural and nonstructural blocks of the genome, providing a modular evolution.

Surveillance for acute flaccid paralysis syndrome (AFP) in children under 15 is the backbone of the Global Polio Eradication Initiative. Laboratory examination of stool samples from AFP cases allows the detection of, along with polioviruses, a variety of non-polio enteroviruses (NPEV). The etiological significance of these viruses in the occurrence of AFP cases has been definitively established only for enteroviruses A71 and D68. Enterovirus Coxsackie A2 (CVA2) is most often associated with vesicular pharyngitis and hand, foot and mouth disease. Among 7280 AFP cases registered in Russia over 20 years (2001–2020), CVA2 was isolated only from five cases. However, these included three children aged 3 to 4 years, without overt immune deficiency, immunized with 4–5 doses of poliovirus vaccine in accordance with the National Vaccination Schedule. The disease resulted in persistent residual paralysis. Clinical and laboratory data corresponded to poliomyelitis developing during poliovirus infection. These findings are compatible with CVA2 being the cause of AFP. Molecular analysis of CVA2 from these patients and a number of AFP cases in other countries did not reveal association with a specific phylogenetic group, suggesting that virus genetics is unlikely to explain the pathogenic profile. The overall results highlight the value of AFP surveillance not just for polio control but for studies of uncommon AFP agents.