Clustering of classical swine fever virus isolates by codon pair bias

<p>Abstract</p> <p>Background</p> <p>The genetic code consists of non-random usage of synonymous codons for the same amino acids, termed codon bias or codon usage. Codon juxtaposition is also non-random, referred to as codon context bias or codon pair bias. The codon and codon pair bias vary among different organisms, as well as with viruses. Reasons for these differences are not completely understood. For classical swine fever virus (CSFV), it was suggested that the synonymous codon usage does not significantly influence virulence, but the relationship between variations in codon pair usage and CSFV virulence is unknown. Virulence can be related to the fitness of a virus: Differences in codon pair usage influence genome translation efficiency, which may in turn relate to the fitness of a virus. Accordingly, the potential of the codon pair bias for clustering CSFV isolates into classes of different virulence was investigated.</p> <p>Results</p> <p>The complete genomic sequences encoding the viral polyprotein of 52 different CSFV isolates were analyzed. This included 49 sequences from the GenBank database (NCBI) and three newly sequenced genomes. The codon usage did not differ among isolates of different virulence or genotype. In contrast, a clustering of isolates based on their codon pair bias was observed, clearly discriminating highly virulent isolates and vaccine strains on one side from moderately virulent strains on the other side. However, phylogenetic trees based on the codon pair bias and on the primary nucleotide sequence resulted in a very similar genotype distribution.</p> <p>Conclusion</p> <p>Clustering of CSFV genomes based on their codon pair bias correlate with the genotype rather than with the virulence of the isolates.</p

The viral RNase E(rns) prevents IFN type-I triggering by pestiviral single- and double-stranded RNAs

Interferon (IFN) type-I is of utmost importance in the innate antiviral defence of eukaryotic cells. The cells express intra- and extracellular receptors that monitor their surroundings for the presence of viral genomes. Bovine viral diarrhoea virus (BVDV), a Pestivirus of the family Flaviviridae, is able to prevent IFN synthesis induced by poly(IC), a synthetic dsRNA. The evasion of innate immunity might be a decisive ability of BVDV to establish persistent infection in its host. We report that ds- as well as ssRNA fragments of viral origin are able to trigger IFN synthesis, and that the viral envelope glycoprotein E(rns), that is also secreted from infected cells, is able to inhibit IFN expression induced by these extracellular viral RNAs. The RNase activity of E(rns) is required for this inhibition, and E(rns) degrades ds- and ssRNA at neutral pH. In addition, cells infected with a cytopathogenic strain of BVDV contain more dsRNA than cells infected with the homologous non-cytopathogenic strain, and the intracellular viral RNA was able to excite the IFN system in a 5'-triphosphate-, i.e. RIG-I-, independent manner. Functionally, E(rns) might represent a decoy receptor that binds and enzymatically degrades viral RNA that otherwise might activate the IFN defence by binding to Toll-like receptors of uninfected cells. Thus, the pestiviral RNase efficiently manipulates the host's self-nonself discrimination to successfully establish and maintain persistence and immunotolerance

Autocatalytic Cleavage within Classical Swine Fever Virus NS3 Leads to a Functional Separation of Protease and Helicase

International audienceClassical swine fever virus (CSFV) is a positive-stranded RNA virus belonging to the genus Pestivirus within the Flaviviridae family. Pivotal for processing of a large portion of the viral polyprotein is a serine protease activity within nonstructural protein 3 (NS3) that also harbors helicase and NTPase activities essential for RNA replication. In CSFV-infected cells, NS3 appears as two forms, a fully processed NS3 of 80 kDa and the precursor molecule NS2-3 of 120 kDa. Here we report the identification and mapping of additional autocatalytic intramolecular cleavages. One cleavable peptide bond occurs between Leu1781 and Met1782, giving rise to a helicase subunit of 55 kDa and, depending on the substrate, a NS2-3 fragment of 78 kDa (NS2-3p) or a NS3 protease subunit of 26 kDa (NS3p). In trans-cleavage assays using NS4-5 as a substrate, NS3p acts as a fully functional protease that is able to process the polyprotein. NS3p comprises the minimal essential protease, as deletion of Leu1781 results in inactivation. A second intramolecular cleavage was mapped to the Leu1748/Lys1749 peptide bond that yields a proteolytically inactive NS3 fragment. Deletion of either of the cleavage site residues resulted in a loss of RNA infectivity, indicating the functional importance of amino acid identity at the respective positions. Our data suggest that internal cleavage within the NS3 moiety is a common process that further extends the functional repertoires of the multifunctional NS2-3 or NS3 and represents another level of the complex polyprotein processing of Flaviviridae

Recombinational history and molecular evolution of western equine encephalomyelitis complex alphaviruses.

Western equine encephalomyelitis (WEE) virus (Togaviridae: Alphavirus) was shown previously to have arisen by recombination between eastern equine encephalomyelitis (EEE)- and Sindbis-like viruses (C. S. Hahn, S. Lustig, E. G. Strauss, and J. H. Strauss, Proc. Natl. Acad. Sci. USA 85:5997-6001, 1988). We have now examined the recombinational history and evolution of all viruses belonging to the WEE antigenic complex, including the Buggy Creek, Fort Morgan, Highlands J, Sindbis, Babanki, Ockelbo, Kyzylagach, Whataroa, and Aura viruses, using nucleotide sequences derived from representative strains. Two regions of the genome were examined: sequences of 477 nucleotides from the C terminus of the E1 envelope glycoprotein gene which in WEE virus was derived from the Sindbis-like virus parent, and 517 nucleotide sequences at the C terminus of the nsP4 gene which in WEE virus was derived from the EEE-like virus parent. Trees based on the E1 region indicated that all members of the WEE virus complex comprise a monophyletic group. Most closely related to WEE viruses are other New World members of the complex: the Highlands J, Buggy Creek, and Fort Morgan viruses. More distantly related WEE complex viruses included the Old World Sindbis, Babanki, Ockelbo, Kyzylagach, and Whataroa viruses, as well as the New World Aura virus. Detailed analyses of 38 strains of WEE virus revealed at least 4 major lineages; two were represented by isolates from Argentina, one was from Brazil, and a fourth contained isolates from many locations in South and North America as well as Cuba. Trees based on the nsP4 gene indicated that all New World WEE complex viruses except Aura virus are recombinants derived from EEE- and Sindbis-like virus ancestors. In contrast, the Old World members of the WEE complex, as well as Aura virus, did not appear to have recombinant genomes. Using an evolutionary rate estimate (2.8 x 10(-4) substitutions per nucleotide per year) obtained from E1-3' sequences of WEE viruses, we estimated that the recombination event occurred in the New World 1,300 to 1,900 years ago. This suggests that the alphaviruses originated in the New World a few thousand years ago