Abstract
Cigar tobacco (Nicotiana tabacum L.) has been recently introduced into China for various industrial applications. From March 2022, certain symptoms of begomovirus infection, including leaf curling and thickening of veins, were sporadically (disease incidence was approximately 0.2%) observed in several cigar tobacco plantations in numerous counties in Hainan Province, China (Figure 1A). These typical symptoms of begomovirus infection were similar to those caused by the sida leaf curl virus-Hainan (SiLCV-HN) begomovirus and its associated betasatellite, as reported in our previous study on cigar tobacco plants in the same region (Wang et al. 2022). In order to determine whether these symptoms were caused by SiLCV-HN or other begomoviruses, samples of leaves were collected from the diseased tobacco plants for DNA extraction, and the total DNA was extracted for viral metagenomics using an Illumina Sequencing platform at Tiangen Biotech, Beijing. A total of 65711396 filtered reads were obtained, of which 65362322 (99.47%) reads matched to the genome of tobacco. The remaining unmapped 349074 (0.53%) reads were analyzed by BLASTn against the virus Refseq Database of GenBank and subsequently assembled. A total of 8 (5+2+1) enriched contigs of the complete sequence of ludwigia yellow vein Vietnam virus (LuYVVNV) and 9 (8+1) contigs of ludwigia yellow vein virus-associated DNA beta (LuYVB) were finally obtained (Table 1). LuYVVNV belongs to the Begomovirus genus that infects various weeds, including Ludwigia octovalvis and Impatiens balsamina. As far as we know, it was reported earliest on weed in Vietnam (Ha et al. 2008). GenBank contains data pertaining to previously identified isolates of LuYVVNV, and the data revealed that the virus was discovered in Vietnam and the Yunnan province of China currently. However, there are no reports on the infection of crops by LuYVVNV to date. The findings of the present study indicated that LuYVVNV and LuYVB could be responsible for the aforementioned symptoms observed on cigar tobacco. The complete genomes of LuYVVNV and LuYVB were amplified using primer pairs designed based on sequence assembly for viral metagenomics (Table 2). Indeed, two DNA bands with length 2763 bp of LuYVVNV genome and 1348 bp of LuYVB were amplified from leaf samples of diseased tobacco (Figure 1B). The products of polymerase chain reaction (PCR) amplification were analyzed by Sanger sequencing, and the complete nucleotide sequences of LuYVVNV and its associated betasatellite were obtained. Analysis with the BLASTn tool of NCBI revealed that the genome sequence of LuYVVNV isolated from the Hainan province of China had the highest identity of 96.9% to a different isolate of LuYVVNV (GenBank accession number: MN210347.1). These two isolates belong to the same strain, according to the latest revision of Begomovirus taxonomy (Brown et al. 2015). The isolate of LuYVVNV identified in this study was designated as LuYVVNV, Hainan isolate (LuYVVNV-HN, GenBank accession number: OP948731). BLASTn analysis further revealed that the associated betasatellite had the highest sequence identity of 96.9% with an LuYVB (GenBank accession number: AJ965541.1) of a different viral isolate, according to the classification and nomenclature of DNA betasatellites of begomoviruses (Briddon et al. 2008). The sequence of LuYVB obtained herein was therefore designated as LuYVB, Hainan isolate (LuYVB-HN, GenBank accession number: OP948732). The pathogenicity of LuYVVNV-HN and LuYVB-HN was determined using infectious clones that were constructed by ligating two fragments of LuYVVNV-HN or LuYVB-HN to a binary pCAMBIA1300 expression vector, as previously described (Wang et al. 2019). Infectious clones of LuYVVNV-HN, LuYVB-HN, and LuYVVNV-HN plus LuYVB-HN were separately agroinfiltrated into N. benthamiana for determining viral pathogenicity. The typical symptoms of begomovirus infection were observed in N. benthamiana plants inoculated with LuYVVNV-HN alone or LuYVVNV-HN plus LuYVB-HN, and the emerging leaves were mildly or severely down-curled, respectively, at 7 days post inoculation (dpi), with 100% disease incidence (6/6) (Figure 1C). Positive PCR products of the AV1 gene of LuYVVNV-HN were obtained from N. benthamiana plants inoculated with LuYVVNV-HN alone or LuYVVNV-HN plus LuYVB-HN. The βC1 gene of LuYVB-HN was only obtained from N. benthamiana plants co-infected with LuYVVNV-HN and LuYVB-HN (Figure 1D). No symptoms of viral infection were observed in plants individually inoculated with LuYVB-HN, and the results of PCR were negative (Figure 1C and 1D). These findings indicated that the N. benthamiana plants had been successfully inoculated with LuYVVNV-HN, and that LuYVB-HN was incapable of causing infections on its own, but functioned as a helper and enhanced viral pathogenicity. This report is the first to identify isolates of LuYVVNV and LuYVB from cigar tobacco, which is an economically important crop plant. The findings provide insights into the epidemic threat of begomovirus reservoirs in weeds to crop plants, and emphasize the need for monitoring and controlling whitefly-transmitted viral diseases in tobacco plantations worldwide (Ye et al. 2021).
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Claims 11
Claim · model-written summary
Begomovirus-type symptoms (leaf curling and vein thickening) occurred sporadically in cigar tobacco plantations in Hainan Province, with about 0.2% disease incidence.
- Taxon
- cigar tobaccoe0
- Interactor
- begomovirus
Begomoviruse0pathogen - Outcome
- disease incidencee0
- Value
- approximately 0.2%e0
- Setting
- Hainan Province, Chinae0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“symptoms of begomovirus infection, including leaf curling and thickening of veins, were sporadically (disease incidence was approximately 0.2%) observed in several cigar tobacco plantations in numerous counties in Hainan Province, China”
Claim · model-written summary
Isolates of ludwigia yellow vein Vietnam virus and its associated betasatellite were identified from cigar tobacco for the first time.
Verbatim
e0“This report is the first to identify isolates of LuYVVNV and LuYVB from cigar tobacco”
Claim · model-written summary
The LuYVVNV genome from Hainan had 96.9% highest identity to another LuYVVNV isolate.
Not reportedTaxon
Verbatim
e0“genome sequence of LuYVVNV isolated from the Hainan province of China had the highest identity of 96.9%”
Claim · model-written summary
The associated betasatellite had 96.9% highest sequence identity with an LuYVB of a different isolate.
Not reportedTaxon
Verbatim
e0“the associated betasatellite had the highest sequence identity of 96.9% with an LuYVB”
Claim · model-written summary
Inoculation of N. benthamiana with LuYVVNV-HN alone caused mild down-curling of emerging leaves at 7 dpi, with 100% disease incidence (6/6).
- Taxon
- N. benthamiana
Nicotiana benthamianae1 - Treatment
- inoculated with LuYVVNV-HN alonee0
- Interactor
- LuYVVNV-HNe0pathogen
- Outcome
- mildly or severely down-curlede0
- Value
- 100% disease incidence (6/6)e0
- Qualifier
- 7 days post inoculatione0
Not reportedComparatorDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“were observed in N. benthamiana plants inoculated with LuYVVNV-HN alone or LuYVVNV-HN plus LuYVB-HN, and the emerging leaves were mildly or severely down-curled, respectively, at 7 days post inoculation (dpi), with 100% disease incidence (6/6)”
e1“agroinfiltrated into N. benthamiana”
Claim · model-written summary
Inoculation of N. benthamiana with LuYVVNV-HN plus LuYVB-HN caused severe down-curling of emerging leaves at 7 dpi, with 100% disease incidence (6/6).
- Taxon
- N. benthamianaNicotiana benthamianae1
- Treatment
- LuYVVNV-HN plus LuYVB-HNe0
- Interactor
- LuYVVNV-HNe0pathogen
- Second interactor
- LuYVB-HNe0pathogen
- Outcome
- mildly or severely down-curlede0
- Value
- 100% disease incidence (6/6)e0
- Qualifier
- 7 days post inoculatione0
Not reportedComparatorDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“were observed in N. benthamiana plants inoculated with LuYVVNV-HN alone or LuYVVNV-HN plus LuYVB-HN, and the emerging leaves were mildly or severely down-curled, respectively, at 7 days post inoculation (dpi), with 100% disease incidence (6/6)”
e1“agroinfiltrated into N. benthamiana”
Claim · model-written summary
AV1 gene PCR products of LuYVVNV-HN were obtained from N. benthamiana plants inoculated with LuYVVNV-HN alone or with LuYVVNV-HN plus LuYVB-HN.
- Taxon
- N. benthamianaNicotiana benthamianae0
- Interactor
- LuYVVNV-HNe0pathogen
- Gene
- AV1 genee0
- Outcome
- Positive PCR products of the AV1 genee0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Positive PCR products of the AV1 gene of LuYVVNV-HN were obtained from N. benthamiana plants inoculated with LuYVVNV-HN alone”
Claim · model-written summary
The betaC1 gene of LuYVB-HN was obtained only from N. benthamiana co-infected with LuYVVNV-HN and LuYVB-HN.
- Taxon
- N. benthamianaNicotiana benthamianae0
- Interactor
- LuYVB-HNe0pathogen
- Second interactor
- LuYVVNV-HNe0pathogen
- Gene
- βC1 genee0
- Outcome
- only obtainede0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“The βC1 gene of LuYVB-HN was only obtained from N. benthamiana plants co-infected with LuYVVNV-HN and LuYVB-HN”
Claim · model-written summary
N. benthamiana plants inoculated with LuYVB-HN alone showed no viral infection symptoms and negative PCR results.
- Taxon
- N. benthamianaNicotiana benthamianae1
- Treatment
- individually inoculated with LuYVB-HNe0
- Interactor
- LuYVB-HNe0pathogen
- Outcome
- symptoms of viral infectione0
Not reportedComparatorDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“No symptoms of viral infection were observed in plants individually inoculated with LuYVB-HN, and the results of PCR were negative”
e1“agroinfiltrated into N. benthamiana”
Claim · model-written summary
LuYVB-HN could not cause infection on its own but acted as a helper that enhanced viral pathogenicity.
- Taxon
- N. benthamianaNicotiana benthamianae0
- Interactor
- LuYVB-HNe0pathogen
- Second interactor
- LuYVVNV-HNe0pathogen
- Outcome
- viral pathogenicitye0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“that the N. benthamiana plants had been successfully inoculated with LuYVVNV-HN, and that LuYVB-HN was incapable of causing infections on its own, but functioned as a helper and enhanced viral pathogenicity”
Claim · model-written summary
Monitoring and controlling whitefly-transmitted viral diseases in tobacco plantations is needed.
- Taxon
- tobacco
Nicotiana tabacume0 - Outcome
- monitoring and controlling whitefly-transmitted viral diseasese0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“emphasize the need for monitoring and controlling whitefly-transmitted viral diseases in tobacco plantations worldwide”
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inoculation · disease · Nicotiana benthamianaPositive and negative results
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