Virome Analysis of Aconitum carmichaelii Reveals Infection by Eleven Viruses, including Two Potentially New Species.

Aconitum carmichaelii is a herbaceous herb indigenous to China that has been cultivated for traditional medicine for centuries. Virus-like symptoms of A. carmichaelii plants were observed on leaves in some A. carmichaelii plantations in Zhanyi and Wuding Counties, Yunnan Province, southwest China. High-throughput sequencing (HTS) was performed on 28 symptomatic plants, and the results revealed infection with 11 viruses, including 2 novel viruses and 9 previously described viruses: Aconitum amalgavirus 1 (AcoAV-1), aconite virus A (AcVA), cucumber mosaic virus (CMV), currant latent virus (CuLV), apple stem grooving virus (ASGV), chilli veinal mottle virus (ChiVMV), tomato spotted wilt orthotospovirus (TSWV), tobacco vein distorting virus (TVDV), and potato leafroll virus (PLRV). Two novel viruses tentatively named Aconitum potyvirus 1 and Aconitum betapartitivirus 1, were supported by sequence and phylogenetic analysis results of their genomes. We proposed the names Potyvirus aconiti and Betapartitivirus aconiti. RT-PCR assays of 142 plants revealed the predominance and widespread distribution of CMV, AcVA, and AcoPV-1 in plantations. The detection of isolates of CuLV, ASGV, ChiVMV, TSWV, TVDV, and PLRV infections for the first time in A. carmichaelii expands their known host ranges.

Identification of a putative novel cholivirus in the transcriptome of Gymnema sylvestre R. Br.

Gymnema sylvestre is a tropical climber species that is widely used in traditional medicine since ages. In the present study, the transcriptome datasets of G. sylvestre available in public domain were screened for the presence of novel plant viral sequences and a putative novel virus tentatively named as Gymnema sylvestre virus 1 (GysV1) was identified. Coding-complete genome segments of GysV1 that are 6.35 kb (RNA1) and 3.98 kb (RNA2) long possessed a single large open reading frame coding for a polyprotein. BLASTp, sequence identity and phylogenetic analyses revealed the relatedness of GysV1 to the members of the subgenus Cholivirus (genus Sadwavirus; family Secoviridae; order Picornavirales). Based on the species demarcation criteria of the family Secoviridae, GysV1 can be regarded as a new cholivirus member.

Genomic characterization of a novel torradovirus infecting Arctium lappa L. in China.

Burdock (Arctium lappa L.) is not only a popular vegetable crop but also an important medicinal plant. In burdock plants with symptoms of leaf mosaic, a novel torradovirus tentatively named "burdock mosaic virus" (BdMV) was identified by high-throughput sequencing. The complete genomic sequence of BdMV was further determined using RT-PCR and the rapid amplification of cDNA ends (RACE) method. The genome is composed of two positive-sense single-stranded RNAs. RNA1 (6991 nt) encodes a polyprotein of 2186 aa, and RNA2 (4700 nt) encodes a protein of 201 aa and a polyprotein of 1212 aa that is predicted to be processed into one movement protein (MP) and three coat proteins (CPs). The Pro-Pol region of RNA1 and the CP region of RNA2 shared the highest amino acid sequence identity of 74.0% and 70.6%, respectively, with the corresponding sequences of lettuce necrotic leaf curl virus (LNLCV) isolate JG3. Phylogenetic analysis based on the amino acid sequences of the Pro-Pol and CP regions showed that BdMV clustered with other non-tomato-infecting torradoviruses. Taken together, these results suggest that BdMV is a new member of the genus Torradovirus.

Apple latent spherical virus (ALSV)-induced gene silencing in a medicinal plant, Lithospermum erythrorhizon.

Lithospermum erythrorhizon is a medicinal plant that produces shikonin, a red lipophilic naphthoquinone derivative that accumulates exclusively in roots. The biosynthetic steps required to complete the naphthalene ring of shikonin and its mechanism of secretion remain unclear. Multiple omics studies identified several candidate genes involved in shikonin production. The functions of these genes can be evaluated using virus-induced gene silencing (VIGS) systems, which have been shown advantageous in introducing iRNA genes into non-model plants. This study describes the development of a VIGS system using an apple latent spherical virus (ALSV) vector and a target gene, phytoene desaturase (LePDS1). Virus particles packaged in Nicotiana benthamiana were inoculated into L. erythrorhizon seedlings, yielding new leaves with albino phenotype but without disease symptoms. The levels of LePDS1 mRNAs were significantly lower in the albino plants than in mock control or escape plants. Virus-derived mRNA was detected in infected plants but not in escape and mock plants. Quantitative PCR and deep sequencing analysis indicated that transcription of another hypothetical PDS gene (LePDS2) also decreased in the defective leaves. Virus infection, however, had no effect on shikonin production. These results suggest that virus-based genetic transformation and the VIGS system silence target genes in soil-grown L. erythrorhizon.

Genomic sequence and host range studies reveal considerable variation within the species Arracacha virus B.

Arracacha virus B type (AVB-T) and oca (AVB-O) strains from arracacha (Arracacia xanthorrhiza) and oca (Oxalis tuberosa) samples collected in 1975 and two additional isolates obtained from arracacha (AVB-PX) and potato (AVB-6A) in Peru in 1976 and 1978, respectively, were studied. In its host responses and serological properties, AVB-PX most resembled AVB-T, whereas AVB-6A most resembled AVB-O. Complete genomic sequences of the RNA-1 and RNA-2 of each isolate were obtained following high-throughput sequencing of RNA extracts from isolates preserved for 38 (AVB-PX) or 32 (the other 3 isolates) years, and compared with a genomic sequence of AVB-O obtained previously (PV-0082). RNA-2 was unexpectedly divergent compared to RNA-1, with the nucleotide (nt) sequence identity of different AVB isolates varying by up to 76% (RNA-2) and 89% (RNA-1). The coat protein amino acid sequences were the most divergent, with AVB-O and AVB-6A having only 68% identity to AVB-T and AVB-PX. Since the RNA2 sequence differences between the two isolate groupings also coincided with host range, symptom, and serological differences, AVB demonstrates considerable intraspecific divergence.

Evidence for participation of RNA 1-encoded elicitor in Cowpea mosaic virus-mediated concurrent protection.

The cowpea (Vigna unguiculata) line Arlington, inoculated with Cowpea mosaic virus (CPMV), showed no symptoms, and no infectivity or accumulation of capsid antigen was detected at several days after inoculation. Coinoculation, but not sequential inoculation, of CPMV with similar concentrations of another Comovirus; Cowpea severe mosaic virus (CPSMV), resulted in reduced numbers of CPSMV-induced lesions. This apparent, CPMV-mediated reduction in number of CPSMV-induced infection centers was termed concurrent protection. We report results obtained by inoculating two nearly isogenic cowpea lines derived from a CPMV-susceptible cowpea crossed to Arlington, one line CPMV-susceptible and the other resistant. The CPMV virions B and M, encapsidating genomic RNAs 1 and 2, respectively, were extensively purified by gradient centrifugation. In the CPMV-resistant cowpea, either CPMV or CPMV B affected concurrent protection against CPSMV and against two distinct non-Comoviruses: Cherry leafroll virus and Southern bean mosaic virus. Adding CPMV M to the inoculum did not enhance CPMV-B-mediated protection. CPMV B was ineffective in protecting CPMV-susceptible cowpea. We postulate that CPMV-mediated concurrent protection is elicited in CPMV-resistant cowpea by a CPMV RNA-1-encoded factor and acts to reduce accumulation or spread of CPMV and certain coinoculated challenging viruses in or from the inoculated cell. Coinoculated CPMV did not protect CPMV-resistant cowpea against Tomato bushy stunt virus or Cucumber mosaic virus.