Here a virus, there a virus, everywhere the same virus?

There are an estimated 10(31) viruses on Earth, most of which are phages that infect bacteria. Metagenomic analyses have shown that environmental viral communities are incredibly diverse. There are an estimated 5000 viral genotypes in 200 liters of seawater and possibly a million different viral genotypes in one kilogram of marine sediment. By contrast, some culturing and molecular studies have found that viruses move between different biomes. Together, these findings suggest that viral diversity could be high on a local scale but relatively limited globally. Also, by moving between environments, viruses can facilitate horizontal gene transfer.     

Discovery of the first virus, the tobacco mosaic virus: 1892 or 1898?

Two scientists contributed to the discovery of the first virus, Tobacco mosaic virus. Ivanoski reported in 1892 that extracts from infected leaves were still infectious after filtration through a Chamberland filter-candle. Bacteria are retained by such filters, a new world was discovered: filterable pathogens. However, Ivanovski probably did not grasp the full meaning of his discovery. Beijerinck, in 1898, was the first to call 'virus', the incitant of the tobacco mosaic. He showed that the incitant was able to migrate in an agar gel, therefore being an infectious soluble agent, or a 'contagium vivum fluidum' and definitively not a 'contagium fixum' as would be a bacteria. Ivanovski and Beijerinck brought unequal but decisive and complementary contributions to the discovery of viruses. Since then, discoveries made on Tobacco mosaic virus have stood out as milestones of virology history.     

Founder virus population related to route of virus transmission: a determinant of intrahost human immunodeficiency virus type 1 evolution?

We and others have shown that in individual human immunodeficiency virus type 1 (HIV-1) infection, the adaptive evolution of HIV-1 is influenced by host immune competence. In this study, we tested the hypothesis that in addition to selective forces operating within the host, transmission bottlenecks have an impact on HIV-1 intrahost evolution. Therefore, we studied the intrahost evolution of the V3 region of the external glycoprotein gp120 of HIV-1 during the 3- and 5-year periods following seroconversion after parenteral versus sexual (male-to-male) transmission in 41 participants of the Amsterdam prospective cohorts of homosexual men (n = 31) and intravenous drug users (IVDUs; n = 10) who were AIDS free and had comparable numbers of CD4+ cells. We observed that HIV-1 strains in homosexual men accumulated over 5 years more nonsynonymous substitutions within the V3 loop than HIV-1 strains in IVDUs as a result of lower rates of nonsynonymous evolution in both the initial 3-year period from seroconversion and the following 2-year period as well as a larger proportion of nonsynonymous back substitutions in IVDUs. The mean numbers of synonymous substitutions did not differ between the two risk groups. Since HIV-1 strains in IVDUs could be distinguished from the viruses of homosexual men based on several nucleotide substitutions of which the most conserved is a synonymous substitution at the tip of the V3 loop (GGC pattern), we studied whether the founder virus population itself has an impact on the intrahost evolution of HIV-1. The mean number of nonsynonymous substitutions accumulated over 5 years within the V3 loop was lower in 10 IVDUs infected by the HIV-1 strains with the GGC signature than in 4 IVDUs infected by HIV-1 strains lacking this pattern, while the mean numbers of synonymous substitutions were similar in the two groups.     

Limits in virus filtration capability? Impact of virus quality and spike level on virus removal with xenotropic murine leukemia virus

Virus filtration (VF) is a key step in an overall viral clearance process since it has been demonstrated to effectively clear a wide range of mammalian viruses with a log reduction value (LRV) > 4. The potential to achieve higher LRV from virus retentive filters has historically been examined using bacteriophage surrogates, which commonly demonstrated a potential of > 9 LRV when using high titer spikes (e.g. 10¹⁰ PFU/mL). However, as the filter loading increases, one typically experiences significant decreases in performance and LRV. The 9 LRV value is markedly higher than the current expected range of 4‐5 LRV when utilizing mammalian retroviruses on virus removal filters (Miesegaes et al., Dev Biol (Basel) 2010;133:3‐101). Recent values have been reported in the literature (Stuckey et al., Biotech Progr 2014;30:79‐85) of LRV in excess of 6 for PPV and XMuLV although this result appears to be atypical. LRV for VF with therapeutic proteins could be limited by several factors including process limits (flux decay, load matrix), virus spike level and the analytical methods used for virus detection (i.e. the Limits of Quantitation), as well as the virus spike quality. Research was conducted using the Xenotropic‐Murine Leukemia Virus (XMuLV) for its direct relevance to the most commonly cited document, the International Conference of Harmonization (ICH) Q5A (International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use, Geneva, Switzerland, 1999) for viral safety evaluations. A unique aspect of this work is the independent evaluation of the impact of retrovirus quality and virus spike level on VF performance and LRV. The VF studies used XMuLV preparations purified by either ultracentrifugation (Ultra 1) or by chromatographic processes that yielded a more highly purified virus stock (Ultra 2). Two monoclonal antibodies (Mabs) with markedly different filtration characteristics and with similar levels of aggregate (<1.5%) were evaluated with the Ultra 1 and Ultra 2 virus preparations utilizing the Planova 20 N, a small virus removal filter. Impurities in the virus preparation ultimately limited filter loading as measured by determining the volumetric loading condition where 75% flux decay is observed versus initial conditions (V₇₅). This observation occurred with both Mabs with the difference in virus purity more pronounced when very high spike levels were used (>5 vol/vol %). Significant differences were seen for the process performance over a number of lots of the less‐pure Ultra 1 virus preparations. Experiments utilizing a developmental lot of the chromatographic purified XMuLV (Ultra 2 Development lot) that had elevated levels of host cell residuals (vs. the final Ultra 2 preparations) suggest that these contaminant residuals can impact virus filter fouling, even if the virus prep is essentially monodisperse. Process studies utilizing an Ultra 2 virus with substantially less host cell residuals and highly monodispersed virus particles demonstrated superior performance and an LRV in excess of 7.7 log₁₀. A model was constructed demonstrating the linear dependence of filtration flux versus filter loading which can be used to predict the V₇₅ for a range of virus spike levels conditions using this highly purified virus. Fine tuning the virus spike level with this model can ultimately maximize the LRV for the virus filter step, essentially adding the LRV equivalent of another process step (i.e. protein A or CEX chromatography). © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 31:135–144, 2015     

Isolation of recombinant field strains of Marek’s disease virus integrated with reticuloendotheliosis virus genome fragments

Two Marek's disease virus (MDV) field strains were isolated from chickens with tumors independently from Guangdong and Guangxi provinces, and it was confirmed that there were no co-infections with reticuloendotheliosis viruses (REV) in chicken embryo fibroblast cells (CEF) in indirect fluorescence antibody test (IFA) with REV-specific monoclonal antibodies. By dot blot hybridization and PCR of genomic DNA of MDV-infected CEF, it was indicated that LTR fragments of REV genome were integrated into genome of these two MDV field strains. To amplify and clone the integrated REV LTR with MDV sequence at the junction, 4 primers from REV LTR and 7 primers from MDV genome fragment with REV LTR insertion hot points were synthesized and 28 (4x7) pairs of primers (one from REV and another from MDV for each pair) were used in PCR while using the genomic DNA of both strains as the templates. The sequence data demonstrated that both recombinant field strains contained the same REV LTR inserted into MDV at the identical sites in US fragment of the genomes. From the above, it was speculated that both recombinant field MDVs were originated from a same recombinant virus and spread among chicken flocks in two provinces.     

Construction and immune efficacy of recombinant pseudorabies virus expressing PrM-E proteins of Japanese encephalitis virus genotype І

Background

Japanese encephalitis (JE) is an arboviral disease with high case fatality rates and neurologic or psychiatric sequelae among survivors in Asia, western Pacific countries and northern Australia. Japanese encephalitis virus (JEV) is the cause of JE and the emergence of genotype ? (GI) JEV has displaced genotype III (GIII) as the dominant strains circulating in some Asian regions. The currently available JE vaccines are safe and effective in preventing this disease, but they are developed based on the GIII JEV strains.

Methods

The recombinant virus PRV TK^?/gE^?/PrM-E⁺ which expressed the premembrane (prM) and envelope (E) proteins of JEV SX09S-01 strain (genotype I, GI) was constructed by homologous recombination between the genome of PRV TK^?/gE^?/LacZ⁺ digested with EcoRI and plasmid pIE-CAG-PrM-E-BGH. Expression of JEV PrM and E proteins was analyzed by Western blot analysis. Immune efficacy of PRV TK^?/gE^?/PrM-E⁺ was further evaluated in mouse model.

Results

A recombinant pseudorabies virus (PRV TK^?/gE^?/PrM-E⁺) was successfully constructed. Mice experiments showed that PRV TK^?/gE^?/PrM-E⁺ could induce a high level of ELISA antibodies against PRV and JEV, as well as high titer of PRV neutralizing antibodies. After challenge with 1?×?10⁷ PFU virulent JEV SX09S-01 strain, the time of death was delayed and the survival rate was improved in PRV TK^?/gE^?/PrM-E⁺ vaccinated mice.

Conclusions

PRV TK^?/gE^?/PrM-E⁺ is a potential vaccine candidate against PRV and JEV GI infection in the future.

     

Multi-year evolutionary dynamics of West Nile virus in suburban Chicago,USA, 2005–2007

West Nile virus has evolved in concert with its expansion across North America, but little is known about the evolutionary dynamics of the virus on local scales. We analysed viral nucleotide sequences from mosquitoes collected in 2005, 2006, and 2007 from a known transmission ‘hot spot’ in suburban Chicago, USA. Within this approximately 11 × 14 km area, the viral envelope gene has increased approximately 0.1% yr⁻¹ in nucleotide-level genetic diversity. In each year, viral diversity was higher in ‘residential’ sites characterized by dense housing than in more open ‘urban green space’ sites such as cemeteries and parks. Phylodynamic analyses showed an increase in incidence around 2005, consistent with a higher-than-average peak in mosquito and human infection rates that year. Analyses of times to most recent common ancestor suggest that WNV in 2005 and 2006 may have arisen predominantly from viruses present during 2004 and 2005, respectively, but that WNV in 2007 had an older common ancestor, perhaps indicating a predominantly mixed or exogenous origin. These results show that the population of WNV in suburban Chicago is an admixture of viruses that are both locally derived and introduced from elsewhere, containing evolutionary information aggregated across a breadth of spatial and temporal scales.     

Phylodynamics of hepatitis C virus subtype 2c in the province of Córdoba, Argentina

The Hepatitis C Virus Genotype 2 subtype 2c (HCV-2c) is detected as a low prevalence subtype in many countries, except in Southern Europe and Western Africa. The current epidemiology of HCV in Argentina, a low-prevalence country, shows the expected low prevalence for this subtype. However, this subtype is the most prevalent in the central province of Córdoba. Cruz del Eje (CdE), a small rural city of this province, shows a prevalence for HCV infections of 5%, being 90% of the samples classified as HCV-2c. In other locations of Córdoba Province (OLC) with lower prevalence for HCV, HCV-2c was recorded in about 50% of the samples. The phylogenetic analysis of samples from Córdoba Province consistently conformed a monophyletic group with HCV-2c sequences from all the countries where HCV-2c has been sequenced. The phylogeographic analysis showed an overall association between geographical traits and phylogeny, being these associations significant (α = 0.05) for Italy, France, Argentina (places other than Córdoba), Martinique, CdE and OLC. The coalescence analysis for samples from CdE, OLC and France yielded a Time for the Most Common Recent Ancestor of about 140 years, whereas its demographic reconstruction showed a “lag” phase in the viral population until 1880 and then an exponential growth until 1940. These results were also obtained when each geographical area was analyzed separately, suggesting that HCV-2c came into Córdoba province during the migration process, mainly from Europe, which is compatible with the history of Argentina of the early 20th century. This also suggests that the spread of HCV-2c occurred in Europe and South America almost simultaneously, possibly as a result of the advances in medicine technology of the first half of the 20th century.     

Vesicular stomatitis virus mRNA and inhibition of translation of cellular mRNA--is there a P function in vesicular stomatitis virus?

Infection of animal cells by vesicular stomatitis virus (VSV) results in inhibition of translation of cellular mRNA. We showed previously that, in BHK cells infected by the Glasgow isolate of VSV Indiana, this is due to competition during the initiation step of protein synthesis of viral and cellular mRNA for a constant, limiting number of ribosomes. We show here that infection of the same cells with the San Juan isolate of VSV resulted in a more rapid shutoff of host protein synthesis and that this was paralleled by a more rapid accumulation of viral mRNA. Extending our conclusion that shutoff is due to mRNA competition, we show further that the average size of polysomes translating viral and cellular mRNA was threefold smaller in cells infected by VSV San Juan than by VSV Glasgow, which, in turn, was about one-half that of uninfected cells. In all cases, cellular and viral mRNA's which encoded the same-sized polypeptides were found on the same-sized polysomes, a result indicating that the efficiency of translation of both types of mRNA's is about the same in the infected cell. Also, there was no preferential sequestration of viral or cellular mRNA's in ribonucleoprotein particles. Additional correlations between the levels of viral mRNA's and the inhibition of protein synthesis came from studies of three other wild-type VSV strains and also from studies with Vero and L cells. In particular, the rate of shutoff of L-cell protein synthesis after infection by any VSV isolate was slower than that in BHK cells, and this was correlated with a slower rate of accumulation of viral mRNA. VSV temperature-sensitive mutants which synthesized, at the nonper-missive temperature, no VSV mRNA failed to inhibit synthesis of cellular proteins. Stanners and co-workers (C. P. Stanners, A. M. Francoeur, and T. Lam, Cell 11:273-281, 1977) claimed that VSV mutant R1 inhibited synthesis of L cell protein synthesis less rapidly than did its parent wild-type strain HR. They concluded that this effect was due to a mutation in an unspecified VSV protein, “P.” We found, in both L and BHK cells, that R1 infection resulted in a slightly slower inhibition of cellular mRNA translation than did HR infection and that this was correlated with a slightly reduced accumulation of VSV mRNA. The level of VSV mRNA, rather than any specific VSV protein, appeared to be the key factor in determining the rate of shutoff of host protein synthesis.     

Yellow fever and dengue—the interactions of virus,vector and host in the re-emergence of epidemic disease

The incidence of the mosquito-borne flavivirus diseases, yellow fever, dengue and dengue hemorrhagic fever has increased dramatically in recent years. Both diseases are characterized by the emergence of explosive epidemics. Yellow fever outbreaks appear to have a periodicity dependent upon fluctuations in sylvatic (enzootic) transmission cycles and the ecological factors that influence these cycles. Spread of the virus from the sylvatic cycle to human settlements, ultimately with interhuman transmission by domestic Aedes aegypti is a repeating event in Africa, and presents a renewed threat in the Americas, where effective Ae. aegypti control collapsed in the 1970s. The incidence of dengue has also increased dramatically in recent years, with up to 80 million persons living in tropical regions of the world now affected annually—an attack rate of 4%. The severe form, dengue hemorrhagic fever (DHF) has become a leading health problem throughout Asia in the last 20 years and is emerging as an epidemic disease in the Americas. Unlike yellow fever, sylvatic dengue transmission cycles are not responsible for disease emergence. The major factors underlying dengue epidemics are changes in human ecology, increasing contact with Ae. aegypti, the co-circulation of multiple dengue serotypes, and a rising prevalence of immunity and immunopathological events that underlie the pathogenesis of DHF. In this review, the complex interplay of virus, host, vector, environment and weather in the ecology of yellow fever and dengue are explored.     

Design,synthesis, anti-TMV,fungicidal, and insecticidal activity evaluation of 1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid derivatives based on virus inhibitors of plant sources

By drawing the creation ideas of botanical pesticides, a series of tetrahydro-β-carboline-3-carboxylic acid derivatives were designed and synthesized, and first evaluated for their anti-TMV, fungicidal and insecticidal activities. Most of these derivatives exhibited good antiviral activity against TMV both in vitro and in vivo. Especially, the activities of compounds 8 and 15 in vivo were higher than that of ribavirin. The compound 8 exhibited more than 70% fungicidal activities against Cercospora arachidicola Hori, Alternaria solani, Bipolaris maydis, and Rhizoctonia solani at 50 mg/kg, compounds 16 and 20 exhibited more than 60% insecticidal activities against Mythimna separate and Ostrinia nubilalis.     

Epstein-Barr virus–specific cytokine-induced killer cells for treatment of Epstein-Barr virus–related malignant lymphoma

Background

Prolonged immunosuppression or delayed T-cell recovery may favor Epstein-Barr virus (EBV) infection or reactivation after allogeneic hematopoietic stem cell transplantation (HSCT), which can lead to post-transplant lymphoproliferative disease (PTLD) and high-grade malignant B-cell lymphoma. Cytokine-induced killer (CIK) cells with dual specific anti-tumor and virus-specific cellular immunity may be applied in this context.

Nuclear localization of Sindbis virus nonstructural protein nsP2

In early infection, approximately 10% of nonstructural protein nsP2 of Sindbis virus was transported into the nuclei of virus-infected BHK-21 cells. Nuclear asP2 was dominantly associated with nuclear matrix. During the course of infection, increasing amounts of nsP2 accumulated in the nuclear fraction. A prominent accumulation of nuclear nsP2 occurred early in infection, from 1 h to 3 h postinfection. Meanwhile. a weak NTPase activity was found to be associated with the immunocomplexed nsP2. Nuclear localization of nsP2 and its possible role were diseussed in relation to the inhibition of host macromolecular synthesis.     

Phylogenetic assessment of filoviruses: how many lineages of Marburg virus?

Filoviruses have to date been considered as consisting of one diverse genus (Ebola viruses) and one undifferentiated genus (Marburg virus). We reconsider this idea by means of detailed phylogenetic analyses of sequence data available for the Filoviridae: using coalescent simulations, we ascertain that two Marburg isolates (termed the "RAVN" strain) represent a quite-distinct lineage that should be considered in studies of biogeography and host associations, and may merit recognition at the level of species. In contrast, filovirus isolates recently obtained from bat tissues are not distinct from previously known strains, and should be considered as drawn from the same population. Implications for understanding the transmission geography and host associations of these viruses are discussed.