ABSTRACT
In this study, we determined the complete sequence of the genomic RNA of a Florida isolate of maracuja mosaic virus (MarMV-FL) and compared it to that of a Peru isolate of the virus (MarMV-P) and those of other known tobamoviruses. Complete sequence analysis revealed that the isolate should be considered a member of a new species and named passion fruit mosaic virus (PafMV). The genomic RNA of PafMV consists of 6,791 nucleotides and encodes four open reading frames (ORFs) coding for proteins of 125 kDa (1,101 aa), 184 kDa (1,612 aa), 34 kDa (311 aa) and 18 kDa (164 aa) in consecutive order from the 5' to the 3' end. The sequence homologies of the four ORFs of PafMV were from 78.8% to 81.6% to those of MarMV-P at the amino acid level. The sequence homologies of the four ORFs of PafMV ranged from 36.0% to 77.9% and from 21.7% to 81.6% to those of other tobamoviruses, at the nucleotide and amino acid level, respectively. Phylogenetic analysis revealed that these PafMV-encoded proteins are closely related to those of MarMV-P. In conclusion, the results indicate that PafMV and MarMV-P belong to different species within the genus Tobamovirus.
Subject(s)
Genome, Viral , Passiflora/virology , RNA, Viral/genetics , Tobamovirus/classification , Tobamovirus/genetics , Chromosome Mapping , Molecular Sequence Data , Mosaic Viruses/classification , Mosaic Viruses/genetics , Open Reading Frames , Phylogeny , Sequence Homology, Nucleic AcidABSTRACT
A full-length cDNA clone (pSP6PepMoV-Vb1) of the genomic RNA of a Korean isolate of Pepper mottle virus (PepMoV-Vb1) was constructed downstream of a bacteriophage SP6 RNA polymerase promoter in the plasmid. In vitro RNA transcripts generated from pSP6PepMoV-Vb1 corresponded to PepMoV-Vb1 RNA (9641nt) with an extra guanosine residue at the 5' terminus and a 15-nt, poly (A) tract at the 3' end. The RNAs synthesized from the pSP6PepMoV-Vb1 clone, by in vitro run-off transcription in the presence of the 5' cap analog m(7)GpppG, were highly infectious in Nicotiana benthamiana and Capsicum annuum cv. Early Calwonder. Visible symptoms appeared at 4-5 days post-inoculation, at essentially the same time as occurred on these host plant species inoculated with wild-type PepMoV-Vb. Symptoms induced by progeny virus of the transcripts were indistinguishable from wild-type PepMoV-Vb on their experimental and natural hosts. The gene encoding the green fluorescent protein (GFP), turboGFP, was inserted between the coding regions for NIb and CP in the pSP6PepMoV-Vb1 clone. RNA transcripts of the resulting GFP-tagged clone, designated SP6PepMoV-Vb1/GFP, were highly infectious and symptoms were not different from those induced by either transcripts of pSP6PepMoV-Vb1 or wild-type PepMoV-Vb. However, GFP expression could be detected earlier than virus-induced symptom in plants infected by SP6PepMoV-Vb1/GFP. This study is the first report of the construction of a biologically active, full-length cDNA copy of the Pepper mottle virus RNA genome and the stable expression of a foreign gene within the modified virus.
