The nucleotide sequence and genome organization of <it>Plasmopara halstedii </it>virus

• Main Authors: Göpfert Jens C, Heller-Dohmen Marion, Pfannstiel Jens, Spring Otmar
• Format: Article
• Language: English
• Published: BMC 2011-03-01
• Series: Virology Journal
• Online Access: http://www.virologyj.com/content/8/1/123

<p>Abstract</p> <p>Background</p> <p>Only very few viruses of Oomycetes have been studied in detail. Isometric virions were found in different isolates of the oomycete <it>Plasmopara halstedii</it>, the downy mildew pathogen of sunflower. However, complete nucleotide sequences and data on the genome organization were lacking.</p> <p>Methods</p> <p>Viral RNA of different <it>P. halstedii </it>isolates was subjected to nucleotide sequencing and analysis of the viral genome. The N-terminal sequence of the viral coat protein was determined using Top-Down MALDI-TOF analysis.</p> <p>Results</p> <p>The complete nucleotide sequences of both single-stranded RNA segments (RNA1 and RNA2) were established. RNA1 consisted of 2793 nucleotides (nt) exclusive its 3' poly(A) tract and a single open-reading frame (ORF1) of 2745 nt. ORF1 was framed by a 5' untranslated region (5' UTR) of 18 nt and a 3' untranslated region (3' UTR) of 30 nt. ORF1 contained motifs of RNA-dependent RNA polymerases (RdRp) and showed similarities to RdRp of <it>Scleropthora macrospora </it>virus A (SmV A) and viruses within the Nodaviridae family. RNA2 consisted of 1526 nt exclusive its 3' poly(A) tract and a second ORF (ORF2) of 1128 nt. ORF2 coded for the single viral coat protein (CP) and was framed by a 5' UTR of 164 nt and a 3' UTR of 234 nt. The deduced amino acid sequence of ORF2 was verified by nano-LC-ESI-MS/MS experiments. Top-Down MALDI-TOF analysis revealed the N-terminal sequence of the CP. The N-terminal sequence represented a region within ORF2 suggesting a proteolytic processing of the CP <it>in vivo</it>. The CP showed similarities to CP of SmV A and viruses within the Tombusviridae family. Fragments of RNA1 (ca. 1.9 kb) and RNA2 (ca. 1.4 kb) were used to analyze the nucleotide sequence variation of virions in different <it>P. halstedii </it>isolates. Viral sequence variation was 0.3% or less regardless of their host's pathotypes, the geographical origin and the sensitivity towards the fungicide metalaxyl.</p> <p>Conclusions</p> <p>The results showed the presence of a single and new virus type in different <it>P. halstedii </it>isolates. Insignificant viral sequence variation indicated that the virus did not account for differences in pathogenicity of the oomycete <it>P. halstedii</it>.</p>