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PubMed1993

Dynamics of genome change in the E2/NS1 region of hepatitis C virus in vivo.

To substantiate and extend the quasispecies model of hepatitis C virus (HCV), we made a pairwise comparison in the nucleotide and deduced amino acid sequences for multiple recombinant clones of the E2/NS1 region, which derived from each blood sample taken from five patients of subtype II or subtype III infection at different stages. Sequence heterogeneity among the clones was generally high. The heterogeneity, however, changed temporally and appeared to be significantly lowered after interferon therapy. The temporal fluctuation involved selection of particular amino acids at particular positions, which had represented only a minor fraction or had been absent in the previous clones. Evolution of defective viruses was featured by most of the cases, the proportion of which also fluctuated temporally and was extremely high at a certain stage in one of the patients. A comparison was also made among the clones from different stages of each patient and revealed highly divergent clones. An extreme case of subtype III infection showed sequence differences well beyond those among chronologically and geographically different isolates of subtype I or subtype II, which were defined by the same calculation of available sequence data. These features of HCV genome suggest that the virus could circulate as an extremely heterogeneous population including defective viruses and that this heterogeneity lends itself to selection pressures including interferon therapy and host immune response. Our results also indicate an absolute need of population based approach in HCV genetics.

Aged↗

PubMed1999

Genomic nucleotide sequence of a foot-and-mouth disease virus clone and its persistent derivatives. Implications for the evolution of viral quasispecies during a persistent infection.

The consensus nucleotide sequence of the entire genome of foot-and-mouth disease virus (FMDV) (biological clone C-S8c1) has been completed, and compared with that of two persistent derivatives R99 and R146, rescued after 99 and 146 passages of the carrier BHK-21 cells. Consensus sequences were determined directly from supernatants of persistently infected cells, without intervening cytolytic amplification of the viruses. These genomic sequences have also been compared with that of FMDV R100, a virus that was also rescued from persistently infected cells, but that was subjected to cytolytic amplification prior to sequencing. Mutation frequencies for R99 and R146 relative to C-S8c1 were in the range of 2.8x10(-3) to 7.7x10(-3) substitutions per nucleotide for the 5'-UTR and the L-, P1-, P2- and P3-coding regions. No mutations were fixed in the polymerase (3D)-coding region. Striking contrasts were noted regarding the distribution of mutation types along the persistent genomes, notably the complete absence of transversion mutations within the 5'-UTR, compared with 53% transversions in the L- and P1-coding regions. The sequencing results presented here, combined with previous sequences of FMDV C-S8c1 genomes at the onset of persistence, provide evidence of sequence fluctuations with a non-linear accumulation of mutations during prolonged persistence, a hallmark of quasispecies dynamics.

3' Untranslated Regions↗

PubMed2000

Viral mutation accelerated by nitric oxide production during infection in vivo.

Nitric oxide (NO), superoxide (O(2)(-)), and their reaction product peroxynitrite (ONOO(-)) are generated in excess during a host's response against viral infection, and contribute to viral pathogenesis by promoting oxidative stress and tissue injury. Here we demonstrate that NO and peroxynitrite greatly accelerates the mutation of Sendai virus (SeV), a nonsegmented negative-strand RNA virus, by using green fluorescent protein (GFP) inserted into and expressed by a recombinant SeV (GFP-SeV) as an indicator for mutation. GFP-SeV mutation frequencies were much higher in the wild-type mice than in those lacking inducible NO synthase, suggesting that mutation of the virus in vivo is NO dependent. High levels of NO and NO-mediated oxidative stress were induced by GFP-SeV infection in the lung of the wild-type mice, but not in the iNOS-deficient mice, as evidenced by electron spin resonance spectroscopy and immunohistochemical analysis for nitrotyrosine formation as well as histopathological examination. Furthermore, peroxynitrite, an NO-derived reactive nitrogen intermediate, enhanced viral mutation in vitro. These results indicate that the oxidative stress induced by NO produced during the natural course of viral infection increases mutation, expands the quasispecies spectrum, and facilitates evolution of RNA viruses.

Animals↗

PubMed2004

RNA recombination in vivo in the absence of viral replication.

To study fundamental aspects of RNA recombination, an in vivo RNA recombination system was established. This system allowed the efficient generation of recombinant cytopathogenic pestiviruses after transfection of synthetic, nonreplicatable, subgenomic transcripts in cells infected with a replicating noncytopathogenic virus. Studies addressing the interplay between RNA recombination and replication revealed that cotransfection of noninfected cells with various pairs of nonreplicatable RNA derivatives also led to the emergence of recombinant viral genomes. Remarkably, homologous and nonhomologous recombination occurred between two overlapping transcripts, each lacking different essential parts of the viral RNA-dependent RNA polymerase (RdRp) gene. Apart from the generally accepted viral replicative copy choice recombination, our results prove the existence of a viral RdRp-independent mechanism of RNA recombination that occurs in vivo. It appears likely that such a mechanism not only contributes to the evolution of RNA viruses but also leads to the generation of recombinant cellular RNAs.

Amino Acid Sequence↗

PubMed1999

Hepatitis G virus infection markers (RNA and anti-E2 antibodies) in a multicenter cohort of hemophiliacs.

BACKGROUND AND OBJECTIVE: To investigate the prevalence and evolution of hepatitis G virus (HGV) infection in hemophiliacs and to correlate evolution of HGV infection markers with immunologic parameters in those patients co-infected with HIV. DESIGN AND METHODS: HGV RNA and anti-E2 antibodies were studied in 124 patients. Serial samples were drawn every 4 months from 1992 to 1996. Lymphocyte subsets including T-helper lymphocytes, T-suppressor lymphocytes, T-cytotoxic lymphocytes, activated T-lymphocytes and natural killer cells were analyzed. RESULTS: Prevalences were 22.6% for HGV RNA and 18.5% for anti-E2. Four patients had both HGV RNA and anti E2, so the overall prevalence of HGV infection in hemophiliacs was 37.9% (11.5% in 200 controls, p<0.0001). After a median follow-up of 36.6 months 20 patients remained HGV RNA positive, whereas HGV RNA had cleared in 8, with an actuarial probability of clearance at 36 months of 34.6%. Only 2 patients developed anti-E2 antibodies. Four patients cleared anti-E2, with an actuarial probability at 36 months of 24.8%. In patients with HIV infection, both lower CD4+ lymphocyte count (p=0.01) or higher CD8+ lymphocyte count (p=0.03) showed predictive value for probability of clearing HGV-RNA. CD4+/CD8+ ratio (p=0.002) was the only variable included in the best model for HGV-RNA disappearance. INTERPRETATION AND CONCLUSIONS: A more accurate estimation of the prevalence of HGV infection can be achieved with the determination of both HGV RNA and anti-E2. Anti-E2 response can be undetectable or transitory after disappearance of HGV-RNA, giving therefore rise to the possibility of underestimating HGV prevalence with currently diagnostic methods. In HIV-positive patients, cellular immune function seems to be involved in the resolution of HGV infection, following the significant correlation found between clearance of HGV-RNA and CD4+/CD8+ lymphocyte populations.

Adolescent↗

PubMed1984

Homologous sequences in non-structural proteins from cowpea mosaic virus and picornaviruses.

Computer analyses have revealed sequence homology between two non-structural proteins encoded by cowpea mosaic virus (CPMV), and corresponding proteins encoded by two picornaviruses, poliovirus and foot-and-mouth disease virus. A region of 535 amino acids in the 87-K polypeptide from CPMV was found to be homologous to the RNA-dependent RNA polymerases from both picornaviruses, the best matches being found where the picornaviral proteins most resemble each other. Additionally, the 58-K polypeptide from CPMV and polypeptide P2-X from poliovirus contain a conserved region of 143 amino acids. Based on the homology observed, a genetic map of the CPMV genome has been constructed in which the 87-K polypeptide represents the core polymerase domain of the CPMV replicase. These results have implications for the evolution of RNA viruses, and mechanisms are discussed which may explain the existence of homology between picornaviruses (animal viruses with single genomic RNAs) and comoviruses (plant viruses with two genomic RNAs).

Journal Article↗

PubMed1991

[The significance of the structural characteristics of the agent for the problems of immunization against blue tongue].

Bluetongue virus (BTV), an arthropod-borne virus, is transmitted primarily by biting midges of the genus Culicoides. Some insect species, which might serve as a potential vector, are prevalent in Central Europe. In sheep, bluetongue is acute and mortality is high, whereas in cattle, goats and most wild ruminants the infection is usually clinically inapparent. Viremia is of short duration in sheep, but cattle experience a prolonged viremia and provide a reservoir for the dissemination of BTV. At least 24 different BTV serotypes have been identified. Antigenic variations occur in the polypeptides of the outer viral capsid and the segmented nature of the viral genome provides the potential for evolution of the virus by a mechanism of reassortment. This renders the use of polyvalent vaccines inefficient and emphasizes the significance of import/export restrictions on ruminants from BTV endemic areas.

Animals↗

PubMed2007

Disease control and its selection for damaging plant virus strains in vegetatively propagated staple food crops; a theoretical assessment.

Viral diseases are a key constraint in the production of staple food crops in lesser developed countries. New and improved disease control methods are developed and implemented without consideration of the selective pressure they impose on the virus. In this paper, we analyse the evolution of within-plant virus titre as a response to the implementation of a range of disease control methods. We show that the development of new and improved disease control methods for viral diseases of vegetatively propagated staple food crops ought to take the evolutionary responses of the virus into consideration. Not doing so leads to a risk of failure, which can result in considerable economic losses and increased poverty. Specifically in vitro propagation, diagnostics and breeding methods carry a risk of failure due to the selection for virus strains that build up a high within-plant virus titre. For vegetatively propagated crops, sanitation by roguing has a low risk of failure owing to its combination of selecting for low virus titre strains as well as increasing healthy crop density.

Animals↗

PubMed2001

Phenotypic recombination between bean common mosaic and bean common mosaic necrosis potyviruses in vivo.

The US-5 strain of bean common mosaic virus (BCMV) and the NL-8 strain of bean common mosaic necrosis virus (BCMNV) are both seedborne potyviruses in common bean (Phaseolus vulgaris L). They have contrasting and highly stable biological characteristics which are genetically controlled. BCMV strain US-5 belongs to pathogenicity group IV. BCMNV strain NL-8 belongs to pathogenicity group III. The two strains have contrasting serological characteristics: NL-8 is serotype A; US-5 is serotype B. When these two strains were maintained separately or as a mixture for more than three years (39 serial transfers) or in more than 100 plants of either of two susceptible hosts, we were unable to isolate a single virus strain that exhibited mutant-like or recombinant-like characteristics. However, within 28 days (during the 1st passage) after these 2 strains were inoculated to opposite primary leaves of bean plants that were susceptible to one virus and resistant to the other, we were able to recover 17 strains that clearly possessed recombinations of various phenotypic characteristics from each of the two "parental" viruses. Three types of phenotypic characteristics were recombined singly or in combination during a single passage in vivo: 1) Biological characteristics known to be controlled by genes for pathogenicity; 2) Serotype; and 3) Temperature-induced hypersensitive vascular necrosis. Each of the phenotypic recombinant strains contained only pathogenicity genes or serological characteristics found in one or both parents. In no case did we isolate a strain that could be described as a random mutation or one that contained pathogenicity or serological characteristics which were not found in at least one parent strain. This is the first known demonstration of phenotypic recombinations between distinct potyviruses in vivo. Implications for the evolution of new virus strains through the use of resistant cultivars and its impact on breeding programs and bean seed production are discussed.

Antigens, Viral↗

PubMedDate not supplied

[Genomic S segment of Crimean-Congo hemorrhagic fever virus circulating in Russia and Bulgaria].

S-segment nucleotide sequences for two Crimean-Congo hemorrhagic fever (CCHF) virus strains isolated in the Rostov Region of Russia and in Bulgaria have been determined. Analysis of complete S-segment nucleotide sequences in the viral strains from different regions of the world has established that the CCHF virus strains isolated from ticks and human beings in different southern Russian regions in 1967 and 2000 are very closely genetically and they form an individual subgroup in the basic European genetic group. By the S-segment structure, the CCHF virus strain isolated in Bulgaria in 1978 belongs to the same genetic group as a representative of its second subgroup. Analysis of the S-segment 3'-noncoding region suggests that the CCHF virus circulating in Europe, Central Asia, and China may have originated from one global focus of infection, including several CCHF virus genovariants. During evolution, fragmental exchange apparently occurred in the S-segment 3'-noncoding region as a result of homological recombination.

3' Untranslated Regions↗

PubMed2004

West Nile virus and other zoonotic viruses in Russia: examples of emerging-reemerging situations.

Studies of the interactions of vertebrates, viruses and arthropod vectors of these viruses were monitored in terms of different ecological groups of viruses transmitted by mosquitoes and ticks in Northern Eurasia in an area encompassing more than 15 million km2. About 90 viruses were isolated, including 24 new to science. Newly recognized infections of vertebrates, including humans, were described. Many unusual epidemic situations were analysed. Permanent efforts were established to prevent bioterrorist activities and their consequences. Extensive epidemic outbreaks of West Nile fever (WNF; i.e., fever caused by West Nile virus) and Crimean-Congo hemorrhagic fever (CCHF) with unusual high mortality appeared in the last four years in southern Russia. We determined infection rates in humans, domestic and wild animals, mosquitoes and ticks from natural and synanthropic biocenoses [Editorial note: "synanthropic" means, roughly, all species living with (c.f. lice, fleas) or near people, such as in houses (c.f. house mice), parks (c.f. Rattus spp.), and the like, rather like "peridomestic", but not strictly so; "biocenosis" is the biome, the "totality of living populations in a particular habitat, which itself is only a part of the ecosystem".]. CCHF virus strains were phylogenetically similar to strains isolated in this area 35 years ago but different from Central-South-Asian and African strains. Before the outset of the current emergence of epidemic WNF, three genetic variants of this virus had been isolated in USSR, two African and one Indian. Phylogenetic analysis of complete genome sequences of epidemic strains demonstrated considerable similarity to strains from USA and Israel and differences from strains isolated in the same USSR areas 20-30 years before. In addition to strains of genotype 1, we isolated strains of second and third lineages and a strain of a fourth genetic variant. Nucleotide differences of these strains from all three genotypes was about 30%. The emerging WNF situation in Russia for the last 4 years probably has been the result of not only natural and social factors, but also to introduction of more virulent strains or by evolution of the virus.

Animals↗

PubMed2005

Evolution of nef variants in gut associated lymphoid tissue of rhesus macaques during primary simian immunodeficiency virus infection.

We utilized the simian immunodeficiency virus model of AIDS to examine evolution of nef gene in gut-associated lymphoid tissue (GALT) during primary and early asymptomatic stages of infection. Macaques were infected with a cloned virus, SIVmac239/nef-stop harboring a premature stop codon in the nef gene. Restoration of the nef open reading frame occurred in GALT early at 3 days post-infection. Analysis of nef sequences by phylogenetic tools showed that evolution of nef was neutral thereafter, as evidenced by the ratio of synonymous to nonsynonymous substitutions, a star pattern in unrooted trees and distribution of amino acid replacements fitting a simple Poisson process. Two regions encoding for a nuclear localization signal and a CTL epitope were conserved. Thus, GALT was a site for strong positive selection of functional nef during initial stages of infection. However, evolution of the nef gene thereafter was neutral during early asymptomatic stage of infection.

Amino Acid Substitution↗

PubMed2004

High recombination rate in herpes simplex virus type 1 natural populations suggests significant co-infection.

Herpes simplex virus type 1 (HSV-1) is highly prevalent in all human populations and has been presumed to evolve in a clonal manner because of a lack of evidence for significant levels of co-infection. Different HSV-1 populations have distinct distributions of strains and the long timescale evident from HSV-1 population diversity has led to the suggestion that studies of virus variability may yield information about host population history. In this sequencing study of three segments of the HSV-1 genome in population samples from the UK and Korea, evidence of recombination was widespread both at the level of reassortment between widely separated loci and within shorter contiguous sequences and the estimated rate of recombination was comparable to that of mutation. Since recombination requires the coexistence of two viral genomes, these results suggest that co-infection by genetically distinct strains may be a more important aspect of HSV-1 epidemiology than previously realized. With its capacity to make new combinations of variants available for selection, substantial recombination requires a radically revised model for the rate and mode of evolution of the virus.

Base Sequence↗

PubMed2006

Full-length sequence analysis of subacute sclerosing panencephalitis (SSPE) virus, a mutant of measles virus, isolated from brain tissues of a patient shortly after onset of SSPE.

Subacute sclerosing panencephalitis (SSPE) virus, a measles virus (MeV) mutant, was isolated from brain tissues of a patient shortly after the clinical onset, and the entire viral genome was sequenced. The virus, named SSPE-Kobe-1, formed syncytia on B95a and Vero/SLAM cells without producing cell-free infectious virus particles, which is characteristic of SSPE virus. Phylogenetic analysis classified SSPE-Kobe-1 into genotype D3. When compared with an MeV field isolate of the same genotype (Ich-B strain), SSPE-Kobe-1 exhibited mutation rates of 0.8-1.6% at the nucleotide level in each of the proteincoding regions of the viral genome. It is noteworthy that the mutation rate of the M gene (1.2%) of SSPE-Kobe-1 was considerably lower than for other SSPE virus strains reported so far, but that the majority of the mutations (75%) were the uridine-to-cytidine biased hypermutation characteristic of the SSPE virus M gene. At the amino acid level, the viral proteins, such as N, P, C, V, M, F, H and L proteins, had point-mutations on 3, 7, 1, 4, 3, 9, 8 and 14 residues, respectively, compared with the Ich-B strain. In addition, the F and H proteins had mutated C-termini due to single-point mutations near or at the stop codons. Two of the three mutations in the M protein were Leu-to-Pro mutations, which are likely to affect the conformation and, therefore, the function of the protein. Because of the relatively small number of mutations, SSPE-Kobe-1 would be a useful tool to study genetic evolution of SSPE virus.

Amino Acid Sequence↗

PubMed1988

Nucleotide sequence of the fusion and phosphoprotein genes of human respiratory syncytial (RS) virus Long strain: evidence of subtype genetic heterogeneity.

The nucleotide and deduced amino acid sequences of the fusion (F) and phosphoprotein (P) genes of the Long strain of human respiratory syncytial (RS) virus have been determined from cDNA copies cloned into pBSV9 shuttle vector. Comparison of these sequences with their counterparts of other strains reveals genetic heterogeneity within the same subtype. The percentage of nucleotide and amino acid changes occurring in both proteins is similar. Thus, the Long F and P proteins share 97.9% and 98.3% amino acid identity, respectively, with their homologs of the A2 strain. Nevertheless the F2 subunit of the fusion protein accumulates 3.1 times more amino acid changes than the F1 subunit. In addition, the percentage of nucleotide changes in the 3' extracistronic sequences is 6 times higher in the P than in the F gene. These results are discussed in terms of selective pressures operating in the evolution of RS virus in nature.

Amino Acid Sequence↗

PubMed2002

JC virus strains indigenous to northeastern Siberians and Canadian Inuits are unique but evolutionally related to those distributed throughout Europe and Mediterranean areas.

Human polyomavirus JC virus (JCV) isolates around the world are classified into more than 10 geographically distinct genotypes (designated as subtypes). Evolutionary relationships among JCV subtypes were recently examined, and the following pattern of JCV evolution was indicated. The ancestral JCV first divided into three superclusters, designated Types A, B, and C. A split in Type A generated two subtypes, EU-a and -b, containing mainly European and Mediterranean isolates. The split in Type B generated Af 2 (the major African subtype), Bl-c (a minor European subtype), and various Asian subtypes. Type C generated a single subtype (Afl), consisting of isolates derived from western Africa. In this study, JCV isolates prevalent among northeastern Siberians and Canadian Inuits were evaluated in the context of the above-described pattern of JCV evolution. The Siberian/Arctic JCV isolates were classified as belonging mainly to Type A, based on the result of a preliminary phylogenetic analysis. We then examined, using the whole-genome approach, the phylogenetic relationships among worldwide Type A isolates. In neighbor-joining and maximum-likelihood analyses, Type A JCVs worldwide consistently diverged into three subtypes, EU-a, -b, and -c, with high bootstrap probabilities. EU-c was constructed only by northeastern Siberian isolates, derived mainly from Nanais living in the lower Amur River region, and was shown to have been generated by the first split in Type A. Most Siberian/Arctic isolates derived from Chukchis, Koryaks, and Canadian Inuits formed a distinct cluster within the EU-a subtype, with a high bootstrap probability. Based on the present findings, we discuss ancient human migrations, accompanied by Type A JCVs, across Asia and to Arctic areas of North America.

Adult↗

PubMed1993

Topical treatment with BV-araU of immunosuppressed and immunocompetent shaved mice cutaneously infected with herpes simplex virus type 1.

Effect of topical treatments with BV-araU was tested in cutaneous infections of shaved Balb/c mice with herpes simplex virus type 1. Evolution of zosteriform skin lesions associated with infection with a low virulent KOS(S) strain was almost completely suppressed by treatments with 5% BV-araU cream given 4 times daily for 5 days starting 1 day after inoculation. This effect was equivalent to that of Zovirax Cream including 5% acyclovir. One percent BV-araU cream was also effective in inhibiting progression of symptoms, while 0.2% cream was not effective. Five percent BV-araU cream significantly suppressed progression of skin lesion even if initiation of treatment was delayed to 2 days after infection. However, the efficacy was diminished by further delay in starting treatment. The effect of BV-araU cream was also evident during infection of immunosuppressed mice. Virus titers in the skin tissue encompassing the inoculation site of mice decreased the day after the first treatment. In the lower flank site, virus replication was almost completely suppressed by the treatment beginning 1 day postinfection. Topical application of BV-araU may be useful therapy for HSV-1 infections in humans, including immunocompromised patients.

Administration, Topical↗

PubMedDate not supplied

[Molecular characterization of human influenza viruses--a look back on the last 10 years].

Influenza A (H3N2) viruses and influenza B viruses have caused more than 90% of influenza infections in Germany during the last then years. Continuous and extensive antigenic variation was evident for both the hemagglutinin (HA) and neuraminidase (NA) surface proteins of H3N2 and influenza B viruses. Molecular characterisation revealed an ongoing genetic drift even in years when the antigenic profiles of circulating strains were indistinguishable from those of the previous season. Retrospective phylogenetic studies showed that viruses similar to vaccine strains circulated one or two years before a given strain was recommended as vaccine strain. New drift variants of H3N2 viruses with significantly changed antigenic features appeared during the seasons 1997/1998 and 2002/2003. Most influenza seasons were characterised by a co-circulation of at least two different lineages of H3N2 viruses. Genetic reassortment between H3N2 viruses belonging to separate lineages caused the different evolutionary pathways of the HA and viruses was responsible for the occurrence of H1N2 viruses during the season 2001/02. This new subtype has been detected only sporadically in Germany. The evolution of influenza B viruses was characterised by the re-emergence of B/Victoria/2/87-lineage viruses and their co-circulation with viruses of the B/Yamagata/16/88-lineage. Reassortant B viruses possessing a Victoria/87-lineage HA and a Yamagata/88-like NA were predominant in Germany during 2002/03 and 2004/05.

Antigenic Variation↗