title: A novel virus of the genus Cilevirus causing symptoms similar to citrus leprosis.
abstract: Citrus leprosis in Colombia was previously shown to be caused by cytoplasmic Citrus leprosis virus (CiLV-C). In 2011, enzyme-linked immunosorbent assay and reverse-transcription polymerase chain reaction (RT-PCR)-based diagnostic methods failed to identify CiLV-C from citrus samples with symptoms similar to citrus leprosis; however, virions similar to CiLV-C were observed in the cytoplasm of the symptomatic leaves by transmission electron microscopy. Furthermore, the causal organism was transmitted by the false spider mite, Brevipalpus phoenicis, to healthy citrus seedlings. A library of small RNAs was constructed from symptomatic leaves and used as the template for Illumina high-throughput parallel sequencing. The complete genome sequence and structure of a new bipartite RNA virus was determined. RNA1 (8,717 nucleotides [nt]) contained two open reading frames (ORFs). ORF1 encoded the replication module, consisting of five domains: namely, methyltransferase (MTR), cysteine protease-like, FtsJ-MTR, helicase (Hel), and RNA-dependent RNA polymerase (RdRp); whereas ORF2 encoded the putative coat protein. RNA2 (4,989 nt) contained five ORFs that encode the movement protein (MP) and four hypothetical proteins (p7, p15, p24, and p61). The structure of this virus genome resembled that of CiLV-C except that it contained a long 3' untranslated terminal region and an extra ORF (p7) in RNA2. Both the RNA1 and RNA2 of the new virus had only 58 and 50% nucleotide identities, respectively, with known CiLV-C sequences and, thus, it appears to be a novel virus infecting citrus. Phylogenetic analyses of the MTR, Hel, RdRp, and MP domains also indicated that the new virus was closely related to CiLV-C. We suggest that the virus be called Citrus leprosis virus cytoplasmic type 2 (CiLV-C2) and it should be unambiguously classified as a definitive member of the genus Cilevirus. A pair of CiLV-C2 genome-specific RT-PCR primers was designed and validated to detect its presence in citrus leprosis samples collected from the Casanare and Meta states in Colombia.
authors.affiliation: University of Florida, FL, USA.
authors.forename: Avijit
authors.initials: A
authors.lastname: Roy
authors.forename: Nandlal
authors.initials: N
authors.lastname: Choudhary
authors.forename: Leon M
authors.initials: LM
authors.lastname: Guillermo
authors.forename: Jonathan
authors.initials: J
authors.lastname: Shao
authors.forename: Ananthakrishnan
authors.initials: A
authors.lastname: Govindarajulu
authors.forename: Diann
authors.initials: D
authors.lastname: Achor
authors.forename: G
authors.initials: G
authors.lastname: Wei
authors.forename: D D
authors.initials: DD
authors.lastname: Picton
authors.forename: L
authors.initials: L
authors.lastname: Levy
authors.forename: M K
authors.initials: MK
authors.lastname: Nakhla
authors.forename: John S
authors.initials: JS
authors.lastname: Hartung
authors.forename: R H
authors.initials: RH
authors.lastname: Brlansky
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001096:Arachnid Vectors / Q000821:virology*; D002957:Citrus / Q000648:ultrastructure / Q000821:virology*; D003105:Colombia; D005638:Fruit; D015723:Gene Library; D016679:Genome, Viral / Q000235:genetics; D059014:High-Throughput Nucleotide Sequencing; D046529:Microscopy, Electron, Transmission; D008925:Mites / Q000821:virology*; D008969:Molecular Sequence Data; D010802:Phylogeny; D010935:Plant Diseases / Q000821:virology*; D018515:Plant Leaves / Q000821:virology; D012328:RNA Viruses / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D012367:RNA, Viral / Q000235:genetics; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics; D036226:Seedlings / Q000648:ultrastructure / Q000821:virology; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
chemical_list: D012367:RNA, Viral; D012324:RNA-Dependent RNA Polymerase