Abstract
A cucumber necrosis tombusvirus (CNV) mutant which lacks the coding sequence for the coat protein protruding domain, PD(-), was constructed by site-directed mutagenesis of an infectious CNV cDNA clone, pK2/M5 (wild-type, 4701 nt). Transcripts of PD(-) were infectious in Nicotiana clevelandii, however, the resulting local lesions were significantly smaller than those on the corresponding leaves of plants inoculated with wild-type transcript. In addition, systemic symptoms took 8 to 12 days longer to develop than in wild type-inoculated plants. N. clevelandii inoculated with sap from systemically infected leaves of PD(-) transcript-inoculated plants showed local lesions the same as wild-type andthe delay in systemic symptom development was either reduced or, occasionally, lost. Systemic symptoms in both PD(-) transcript-and sap-inoculated N. clevelandii were either similar to thesystemic necrosis of wild-type or an oak-leaf necrosis was observed. High molecular weight RNA from mutant- and wild-type-infected plants was extracted and analyzed by Northern blotting. Full-length PD(-) RNA could be detected only rarely in RNA preparations from transcript-inoculated leaves; a further deleted, stable RNA species of approximately 3800 nucleotides was found in preparations from systemically infected leaves of PD(-) transcript-and sap-inoculated plants. CNV coat protein could not be detected by ELISA or ISEM in PD(-)-infected leaf material. The ca. 3.8 kbRNA, when cloned and sequenced, was found to have lost all but 74of the 1140 nucleotides of the CNV coat protein open reading frame. Transcripts from this coat-proteinless CNV cDNA clone produced symptoms on N. clevelandii that were similar to those induced inPD(-)-sap-inoculated plants; wild-type and oak-leaf necroses were observed and there was a trend for systemic symptoms to be delayed when compared with wild-type. It would appear that CNV is able to replicate and move systemically, in both transcript-inoculated and sap-inoculated N. clevelandii, in the absence of a functional coat protein. Additionally, mechanical transmission of this virus occurs in the absence of the coat protein, however, such transmission is less efficient when compared with wild-type infections. CNV and tomato bushy stunt tombus virus (TBSV) have the same genome organization and share high levels of nucleotide and amino acid sequence similarity throughout their genomes, yet differ biologically. CNV, for which the natural vector is the fungus Olpidium radicale, has been found only in greenhouse-grown cucumbers. TBSV, by contrast, has a relatively diverse natural host range and has no known vector. The role of the CNV coat protein in determining transmission by 0. radicale was examined by exchanging the coat protein genes contained in full-length cDNA clones of CNV and TBSV. Transcripts from the resulting chimeric clones (CNV with TBSV coat protein and vice versa) were infectious in N. clevelandii. Virus particles, produced in infected plants, were purified and tested for in vitro acquisition and transmission to roots of cucumber seedlings by zoospores of a cucumber strain of O. radicale. Particles produced by the TBSV chimeric clone encoding the CNV coat protein were transmitted, while those produced by the CNV chimeric clone encoding the TBSV coat protein were not. These results demonstrate that the coat protein gene of the CNV genome is specifically involved in fungus transmissibility.
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Claims 16
Claim · model-written summary
PD(-) transcripts produced significantly smaller local lesions than wild type in Nicotiana clevelandii.
- Taxon
- Nicotiana clevelandii
Nicotiana clevelandiie0 - Treatment
- PD(-)e0
- Outcome
- local lesionse0
- Direction
- smallere0
- Comparator
- wild-type transcripte0
Not reportedInteractor
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Transcripts of PD(-) were infectious in Nicotiana clevelandii, however, the resulting local lesions were significantly smaller than those on the corresponding leaves of plants inoculated with wild-type transcript.”
Claim · model-written summary
Systemic symptoms after PD(-) transcript inoculation developed 8 to 12 days later than after wild type inoculation.
- Taxon
- cucumber
Cucumis sativust0 - Treatment
- PD(-)e1
- Interactor
- cucumber necrosis virusCucumber necrosis virust0pathogen
- Outcome
- systemic symptomse0
- Direction
- longere0
- Value
- 8 to 12 dayse0
- Comparator
- wild type-inoculated plantse0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“cucumber necrosis virus”
e0“systemic symptoms took 8 to 12 days longer to develop than in wild type-inoculated plants.”
e1“Transcripts of PD(-) were infectious in Nicotiana clevelandii”
Claim · model-written summary
Sap from PD(-) transcript infected plants yielded wild-type sized lesions and reduced or lost delay of systemic symptoms in N. clevelandii.
- Taxon
- N. clevelandiiNicotiana clevelandiie0
- Treatment
- sap from systemically infected leaves of PD(-) transcript-inoculated plantse0
- Outcome
- local lesions the same as wild-type andthe delay in systemic symptom development was either reduced or, occasionally, loste0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“N. clevelandii inoculated with sap from systemically infected leaves of PD(-) transcript-inoculated plants showed local lesions the same as wild-type andthe delay in systemic symptom development was either reduced or, occasionally, lost.”
Claim · model-written summary
PD(-) transcript and sap inoculation produced either wild-type systemic necrosis or oak-leaf necrosis in N. clevelandii.
- Taxon
- N. clevelandiiNicotiana clevelandiie0
- Outcome
- systemic necrosis of wild-type or an oak-leaf necrosise0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Systemic symptoms in both PD(-) transcript-and sap-inoculated N. clevelandii were either similar to thesystemic necrosis of wild-type or an oak-leaf necrosis was observed.”
Claim · model-written summary
Full-length PD(-) RNA was only rarely detected in transcript-inoculated leaves.
- Taxon
- cucumberCucumis sativust0
- Interactor
- cucumber necrosis virusCucumber necrosis virust0pathogen
- Outcome
- Full-length PD(-) RNAe0
- Qualifier
- only rarelye0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“cucumber necrosis virus”
e0“Full-length PD(-) RNA could be detected only rarely in RNA preparations from transcript-inoculated leaves”
Claim · model-written summary
A stable further deleted RNA species of approximately 3800 nucleotides occurred in systemically infected leaves.
- Taxon
- cucumberCucumis sativust0
- Interactor
- cucumber necrosis virusCucumber necrosis virust0pathogen
- Outcome
- a further deleted, stable RNA speciese0
- Value
- approximately 3800 nucleotidese0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“cucumber necrosis virus”
e0“a further deleted, stable RNA species of approximately 3800 nucleotides was found in preparations from systemically infected leaves of PD(-) transcript-and sap-inoculated plants.”
Claim · model-written summary
CNV coat protein was undetectable in PD(-) infected leaves.
- Taxon
- cucumberCucumis sativust0
- Interactor
- cucumber necrosis virusCucumber necrosis virust0pathogen
- Outcome
- CNV coat protein could not be detectede0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“cucumber necrosis virus”
e0“CNV coat protein could not be detected by ELISA or ISEM in PD(-)-infected leaf material.”
Claim · model-written summary
Sequenced 3.8 kb RNA retained only 74 of 1140 nucleotides of the coat protein open reading frame.
- Taxon
- cucumberCucumis sativust0
- Interactor
- cucumber necrosis virusCucumber necrosis virust0pathogen
- Outcome
- CNV coat protein open reading framee0
- Value
- 74of the 1140 nucleotidese0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“cucumber necrosis virus”
e0“The ca. 3.8 kbRNA, when cloned and sequenced, was found to have lost all but 74of the 1140 nucleotides of the CNV coat protein open reading frame.”
Claim · model-written summary
Coat-proteinless CNV transcripts produced wild-type and oak-leaf necrosis in N. clevelandii, resembling PD(-) sap inoculation.
- Taxon
- N. clevelandiiNicotiana clevelandiie0
- Interactor
- CNVCucumber necrosis viruse0pathogen
- Outcome
- wild-type and oak-leaf necrosese0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Transcripts from this coat-proteinless CNV cDNA clone produced symptoms on N. clevelandii that were similar to those induced inPD(-)-sap-inoculated plants; wild-type and oak-leaf necroses were observed”
Claim · model-written summary
Coat-proteinless CNV tended to delay systemic symptoms compared with wild type.
- Taxon
- cucumberCucumis sativust0
- Treatment
- cucumber necrosis virust0
- Interactor
- cucumber necrosis virusCucumber necrosis virust0pathogen
- Outcome
- systemic symptomse0
- Direction
- delayede0
- Qualifier
- a trende0
- Comparator
- wild-typee0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“cucumber necrosis virus”
e0“there was a trend for systemic symptoms to be delayed when compared with wild-type.”
Claim · model-written summary
CNV replicated and moved systemically in inoculated N. clevelandii without functional coat protein.
- Taxon
- N. clevelandiiNicotiana clevelandiie0
- Interactor
- CNVCucumber necrosis viruse0pathogen
- Gene
- coat proteine0
- Outcome
- replicate and move systemicallye0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“It would appear that CNV is able to replicate and move systemically, in both transcript-inoculated and sap-inoculated N. clevelandii, in the absence of a functional coat protein.”
Claim · model-written summary
Mechanical transmission of CNV without coat protein was less efficient than wild type.
- Taxon
- cucumberCucumis sativust0
- Treatment
- absence of the coat proteine0
- Interactor
- cucumber necrosis virusCucumber necrosis virust0pathogen
- Outcome
- mechanical transmission of this viruse0
- Direction
- less efficiente0
- Comparator
- wild-type infectionse0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“cucumber necrosis virus”
e0“mechanical transmission of this virus occurs in the absence of the coat protein, however, such transmission is less efficient when compared with wild-type infections.”
Claim · model-written summary
Both reciprocal CNV and TBSV coat protein chimeras infected N. clevelandii.
- Taxon
- N. clevelandiiNicotiana clevelandiie0
- Interactor
- CNVCucumber necrosis viruse0pathogen
- Outcome
- were infectiouse0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Transcripts from the resulting chimeric clones (CNV with TBSV coat protein and vice versa) were infectious in N. clevelandii.”
Claim · model-written summary
The TBSV chimera carrying CNV coat protein was transmitted to cucumber seedling roots by fungal zoospores.
- Taxon
- cucumberCucumis sativuse0
- Interactor
- TBSVTomato bushy stunt viruse0pathogen
- Second interactor
- O. radicalee0other
- Gene
- CNV coat proteine0
- Outcome
- were transmittede0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“transmission to roots of cucumber seedlings by zoospores of a cucumber strain of O. radicale. Particles produced by the TBSV chimeric clone encoding the CNV coat protein were transmitted”
Claim · model-written summary
The CNV chimera carrying TBSV coat protein was not transmitted by fungal zoospores to cucumber seedling roots.
- Taxon
- cucumberCucumis sativuse0
- Interactor
- CNVCucumber necrosis viruse0pathogen
- Second interactor
- O. radicalee0other
- Gene
- TBSV coat proteine0
- Outcome
- those produced by the CNV chimeric clone encoding the TBSV coat protein were note0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“transmission to roots of cucumber seedlings by zoospores of a cucumber strain of O. radicale. Particles produced by the TBSV chimeric clone encoding the CNV coat protein were transmitted, while those produced by the CNV chimeric clone encoding the TBSV coat protein were not.”
Claim · model-written summary
The CNV coat protein gene specifically determines fungal transmissibility.
- Taxon
- cucumberCucumis sativust0
- Interactor
- cucumber necrosis virusCucumber necrosis virust0pathogen
- Gene
- coat protein gene of the CNV genomee0
- Outcome
- fungus transmissibilitye0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“cucumber necrosis virus”
e0“These results demonstrate that the coat protein gene of the CNV genome is specifically involved in fungus transmissibility.”