Microevolution of tick-borne encephalitis virus in course of host alternation

AbstractTwo tick-borne encephalitis (TBE) virus variants were studied: mouse brain-adapted strain EK-328 and its derivate adapted to Hyalomma marginatum ticks. The tick-adapted virus exhibited small-plaque phenotype and slower replication in PEK cells, higher yield in ticks, decreased neuroinvasiveness in mice, increased binding to heparin-sepharose. A total of 15 nucleotide substitutions distinguished genomes of these variants, six substitutions resulted in protein sequence alterations, and two were in 5′NTR. Two amino acid substitutions in E protein were responsible for the observed phenotypic differences. Data obtained during reverse passaging of the tick-adapted virus in vivo and in vitro suggest that TBE virus exists as a heterogeneous population that contains virus variants most adapted to reproduction in either ticks or mammals. Host switch results in a change in the ratio of these variants in the population. Plaque purification of the tick-adapted virus resulted in the prompt emergence of new mutants with different virulence for mammals

Evolutionary origins of hepatitis A virus in small mammals

The origins of human hepatitis A virus (HAV) are unknown. We conducted a targeted search for HAV-related viruses in small mammals sampled globally and discovered highly diversified viruses in bats, rodents, hedgehogs, and shrews. We demonstrate that these viruses share unique biological features with HAV, including structural, genomic, antigenic, and pathogenic properties. We found evidence of major shifts of HAV-related viruses between mammalian hosts in the past, suggesting both an origin of this viral genus in small mammals and a zoonotic origin of human HAV. Our data show that risk assessments for emerging viruses can benefit greatly from the analysis of viral infection patterns that evolved within animal reservoirs

Evolutionary origins of hepatitis A virus in small mammals

Hepatitis A virus (HAV) is an ancient and ubiquitous human pathogen recovered previously only from primates. The sole species of the genus Hepatovirus, existing in both enveloped and nonenveloped forms, and with a capsid structure intermediate between that of insect viruses and mammalian picornaviruses, HAV is enigmatic in its origins. We conducted a targeted search for hepatoviruses in 15,987 specimens collected from 209 small mammal species globally and discovered highly diversified viruses in bats, rodents, hedgehogs, and shrews, which by pairwise sequence distance comprise 13 novel Hepatovirus species. Near-complete genomes from nine of these species show conservation of unique hepatovirus features, including predicted internal ribosome entry site structure, a truncated VP4 capsid protein lacking N-terminal myristoylation, a carboxyl-terminal pX extension of VP1, VP2 late domains involved in membrane envelopment, and a cis-acting replication element within the 3Dpol sequence. Antibodies in some bat sera immunoprecipitated and neutralized human HAV, suggesting conservation of critical antigenic determinants. Limited phylogenetic cosegregation among hepatoviruses and their hosts and recombination patterns are indicative of major hepatovirus host shifts in the past. Ancestral state reconstructions suggest a Hepatovirus origin in small insectivorous mammals and a rodent origin of human HAV. Patterns of infection in small mammals mimicked those of human HAV in hepatotropism, fecal shedding, acute nature, and extinction of the virus in a closed host population. The evolutionary conservation of hepatovirus structure and pathogenesis provide novel insight into the origins of HAV and highlight the utility of analyzing animal reservoirs for risk assessment of emerging viruses

Evidence for Novel Hepaciviruses in Rodents

Hepatitis C virus (HCV) is among the most relevant causes of liver cirrhosis and hepatocellular carcinoma. Research is complicated by a lack of accessible small animal models. The systematic investigation of viruses of small mammals could guide efforts to establish such models, while providing insight into viral evolutionary biology. We have assembled the so-far largest collection of small-mammal samples from around the world, qualified to be screened for bloodborne viruses, including sera and organs from 4,770 roden

Comprehensive Elucidation of the Role of L and 2A Security Proteins on Cell Death during EMCV Infection

The EMCV L and 2A proteins are virulence factors that counteract host cell defense mechanisms. Both L and 2A exhibit antiapoptotic properties, but the available data were obtained in different cell lines and under incomparable conditions. This study is aimed at checking the role of these proteins in the choice of cell death type in three different cell lines using three mutants of EMCV lacking functional L, 2A, and both proteins together. We have found that both L and 2A are non-essential for viral replication in HeLa, BHK, and RD cell lines, as evidenced by the viability of the virus in the absence of both functional proteins. L-deficient infection led to the apoptotic death of HeLa and RD cells, and the necrotic death of BHK cells. 2A-deficient infection induced apoptosis in BHK and RD cells. Infection of HeLa cells with the 2A-deficient mutant was finalized with exclusive caspase-dependent death with membrane permeabilization, morphologically similar to pyroptosis. We also demonstrated that inactivation of both proteins, along with caspase inhibition, delayed cell death progression. The results obtained demonstrate that proteins L and 2A play a critical role in choosing the path of cell death during infection, but the result of their influence depends on the properties of the host cells

Nonreplicative homologous RNA recombination: Promiscuous joining of RNA pieces?

Biologically important joining of RNA pieces in cells, as exemplified by splicing and some classes of RNA editing, is posttranscriptional, whereas in RNA viruses it is generally believed to occur during viral RNA polymerase-dependent RNA synthesis. Here, we demonstrate the assembly of precise genome of an RNA virus (poliovirus) from its cotransfected fragments, which does not require specific RNA sequences, takes place before generation of the viral RNA polymerase, and occurs in different ways: Apparently unrestricted ligation of the terminal nucleotides, joining of any one of the two entire fragments with the relevant internal nucleotide of its partner, or internal crossovers within the overlapping sequence. Incorporation of the entire 5′ or 3′ partners into the recombinant RNA is activated by the presence of terminal 3′-phosphate and 5′-OH, respectively. Such postreplicative reactions, fundamentally differing from the known site-specific and structurally demanding cellular RNA rearrangements, might contribute to the origin and evolution of RNA viruses and could generate new RNA species during all stages of biological evolution

Independent evolution of tetraloop in enterovirus oriL replicative element and its putative binding partners in virus protein 3C

Background Enteroviruses are small non-enveloped viruses with a (+) ssRNA genome with one open reading frame. Enterovirus protein 3C (or 3CD for some species) binds the replicative element oriL to initiate replication. The replication of enteroviruses features a low-fidelity process, which allows the virus to adapt to the changing environment on the one hand, and requires additional mechanisms to maintain the genome stability on the other. Structural disturbances in the apical region of oriL domain d can be compensated by amino acid substitutions in positions 154 or 156 of 3C (amino acid numeration corresponds to poliovirus 3C), thus suggesting the co-evolution of these interacting sequences in nature. The aim of this work was to understand co-evolution patterns of two interacting replication machinery elements in enteroviruses, the apical region of oriL domain d and its putative binding partners in the 3C protein. Methods To evaluate the variability of the domain d loop sequence we retrieved all available full enterovirus sequences (>6, 400 nucleotides), which were present in the NCBI database on February 2017 and analysed the variety and abundance of sequences in domain d of the replicative element oriL and in the protein 3C. Results A total of 2,842 full genome sequences was analysed. The majority of domain d apical loops were tetraloops, which belonged to consensus YNHG (Y = U/C, N = any nucleotide, H = A/C/U). The putative RNA-binding tripeptide 154–156 (Enterovirus C 3C protein numeration) was less diverse than the apical domain d loop region and, in contrast to it, was species-specific. Discussion Despite the suggestion that the RNA-binding tripeptide interacts with the apical region of domain d, they evolve independently in nature. Together, our data indicate the plastic evolution of both interplayers of 3C-oriL recognition