scholarly journals Deep Sequencing of Recombinant Virus Populations in Transgenic and Nontransgenic Plants Infected with Cucumber mosaic virus

2013 ◽  
Vol 26 (7) ◽  
pp. 801-811 ◽  
Author(s):  
Marco Morroni ◽  
Mireille Jacquemond ◽  
Mark Tepfer
Keyword(s):  
Transgenic Plants ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
De Novo ◽  
Recombinant Viruses ◽  
Viral Genomes ◽  
Large Deletions ◽  
Cp Gene ◽  
Viral Sequences ◽  
Virus Variability

Recombination is a major source of virus variability, and the question of whether novel recombinant viruses would emerge in transgenic plants expressing viral sequences has been a biosafety issue. We describe the results of pyrosequencing the recombinant viral RNAs appearing in transgenic plants expressing the coat protein (CP) gene and 3′ noncoding region of Cucumber mosaic virus RNA3, as well as in nontransgenic controls. The populations of recombinants in both transgenic and nontransgenic plants were similar to those previously described from Sanger sequencing but many more recombinant types were observed, including a novel class of large deletions removing all or nearly the entire CP gene. These results show that populations of recombinant viral genomes arising de novo can be characterized in detail by pyrosequencing, and confirm that the transgenic plants did not harbor novel recombinants of biosafety concern.

scholarly journals Evaluation of potential risks associated with recombination in transgenic plants expressing viral sequences

2008 ◽  
Vol 89 (1) ◽  
pp. 327-335 ◽  
Author(s):  
Camilla Turturo ◽  
Arianna Friscina ◽  
Stéphane Gaubert ◽  
Mireille Jacquemond ◽  
Jeremy R. Thompson ◽  
...  
Keyword(s):  
Transgenic Plants ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Viral Rna ◽  
Rna Recombination ◽  
Recombinant Viruses ◽  
Tomato Aspermy Virus ◽  
Potential Risks ◽  
Viral Sequences

Virus-resistant transgenic plants have been created primarily through the expression of viral sequences. It has been hypothesized that recombination between the viral transgene mRNA and the RNA of an infecting virus could generate novel viruses. As mRNA/viral RNA recombination can occur in virus-resistant transgenic plants, the key to testing this risk hypothesis is to compare the populations of recombinant viruses generated in transgenic and non-transgenic plants. This has been done with two cucumoviral systems, involving either two strains of cucumber mosaic virus (CMV), or CMV and the related tomato aspermy virus (TAV). Although the distribution of the sites of recombination in the CMV/CMV and TAV/CMV systems was quite different, equivalent populations of recombinant viruses were observed in both cases. These results constitute the first comparison of the populations of recombinants in transgenic and non-transgenic plants, and suggest that there is little risk of emergence of recombinant viruses in these plants, other than those that could emerge in non-transgenic plants.

Effectiveness of coat protein and defective replicase gene-mediated resistance against Australian isolates of cucumber mosaic virus

1998 ◽  
Vol 38 (4) ◽  
pp. 375 ◽  
Author(s):  
Z. Singh ◽  
M. G. K. Jones ◽  
R. A. C. Jones
Keyword(s):  
Coat Protein ◽  
Transgenic Plants ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Transgenic Tobacco ◽  
Extreme Resistance ◽  
Tobacco Plants ◽  
Systemic Movement ◽  
Cp Gene ◽  
Subgroup Ii

Summary. Transgenic tobacco (Nicotiana tabacum) plants of (i) cv. Samsun NN containing the cauliflower mosaic virus 35S constitutive promoter linked to a defective replicase (DR) gene derived from cucumber mosaic virus (CMV) subgroup I isolate Fny, and (ii) cv. Xanthi containing the CaMV 35S promoter linked to the coat protein (CP) gene of CMV subgroup I isolate C were tested for resistance to various Australian isolates of CMV. The tobacco plants were challenged with 3 CMV subgroup 1 isolates (BNRR, BMR and B6) using sap inoculation. When used to challenge non-transgenic tobacco plants with 5 subgroup II CMV isolates from lupins (LY, LCH, LAcc, LGu and LD), this inoculation method did not result in systemic infection so graft inoculation was used instead to challenge transgenic plants with these 5 isolates. When plants of the line with the DR gene were challenged with the 3 subgroup I isolates, extreme resistance was revealed as none showed symptoms and CMV was not detectable by ELISA. When the same 3 isolates were inoculated to the 3 lines with the CP gene, resistance was characterised by fewer plants becoming virus infected, delayed systemic movement and, in the plants that were infected, partial remission of symptoms plus somewhat decreased virus concentration. Challenge of transgenic plants with DR or CP with the 5 subgroup II isolates resulted in fewer plants becoming infected. Actual numbers of plants infected varied with line and subgroup II isolate and the DR gene was as effective as the CP gene at decreasing infection. With subgroup II isolate LY, infection was associated with remission of symptoms and with the other 4 isolates with delayed systemic movement. Thus the DR gene approach was more effective than the CP approach in obtaining extreme resistance against Australian subgroup I isolates of CMV. These results suggest that introducing a similar DR gene construct made from a subgroup II isolate from lupins into commercial lupin cultivars may be a suitable strategy for obtaining extreme resistance to subgroup II isolates from lupins.

scholarly journals Mutation and Recombination Frequencies Reveal a Biological Contrast within Strains of Cucumber Mosaic Virus

2015 ◽  
Vol 89 (13) ◽  
pp. 6817-6823 ◽  
Author(s):  
Justin S. Pita ◽  
Viktoriya Morris ◽  
Marilyn J. Roossinck
Keyword(s):  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Mutation Frequency ◽  
Recombination Frequency ◽  
In Planta ◽  
Emerging Viruses ◽  
Content Type ◽  
Viral Genomes ◽  
High Mutation Frequency ◽  
Major Player

ABSTRACTRecentin plantastudies have shown that strains Fny and LS ofCucumber mosaic virus(CMV) display differential genetic diversities, Fny and LS having higher and lower mutation frequencies, respectively (J. S. Pita and M. J. Roossinck, J Virol 87:790–797, 2012http://dx.doi.org/10.1128/JVI.01891-12). In this article, we show that these virus strains have differential recombination frequencies as well. However, the high-diversity Fny strain is a low-recombination virus, whereas the very-low-diversity LS strain is instead a high-recombination virus. Unlike the mutation frequency that was determined by both RNAs 1 and 2, the control elements of recombination frequency reside predominantly within RNA 2, specifically within the 2a gene.IMPORTANCERecombination is an important mechanism in virus evolution that can lead to increased or decreased variation and is a major player in virus speciation events that can lead to emerging viruses. Although viral genomes show very frequent evidence of recombination, details of the mechanism involved in these events are still poorly understood. We show here that the reciprocal effects of high mutation frequency and low recombination frequency (andvice versa) involve the RNA-dependent RNA polymerase of the virus, and we speculate that these evolutionary events are related to differences in processivity for two strains of the same virus.

Anti-sense RNAs of cucumber mosaic virus in transgenic plants assessed for control of the virus

1988 ◽  
Vol 11 (4) ◽  
pp. 463-471 ◽  
Author(s):  
M. Ali Rezaian ◽  
Ken G. M. Skene ◽  
Jeff G. Ellis
Keyword(s):  
Transgenic Plants ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus

scholarly journals The effect of variant interference on de novo assembly for viral deep sequencing

2019 ◽  
Author(s):  
Christina J. Castro ◽  
Rachel L. Marine ◽  
Edward Ramos ◽  
Terry Fei Fan Ng
Keyword(s):  
Deep Sequencing ◽  
De Novo ◽  
Gc Content ◽  
Read Length ◽  
Viral Genomes ◽  
Minor Variant ◽  
Main Driver ◽  
Next Generation Sequencing Ngs ◽  
Viral Sequences ◽  
Generation Sequencing

AbstractViruses have high mutation rates and generally exist as a mixture of variants in biological samples. Next-generation sequencing (NGS) approach has surpassed Sanger for generating long viral sequences, yet how variants affect NGS de novo assembly remains largely unexplored. Our results from >15,000 simulated experiments showed that presence of variants can turn an assembly of one genome into tens to thousands of contigs. This “variant interference” (VI) is highly consistent and reproducible by ten most used de novo assemblers, and occurs independent of genome length, read length, and GC content. The main driver of VI is pairwise identities between viral variants. These findings were further supported by in silico simulations, where selective removal of minor variant reads from clinical datasets allow the “rescue” of full viral genomes from fragmented contigs. These results call for careful interpretation of contigs and contig numbers from de novo assembly in viral deep sequencing.

scholarly journals Performance of Transgenic Tomatoes Expressing Cucumber Mosaic Virus CP Gene under Epidemic Conditions

HortScience ◽  
1998 ◽  
Vol 33 (6) ◽  
pp. 1032-1035 ◽  
Author(s):  
John F. Murphy ◽  
Edward J. Sikora ◽  
Bernard Sammons ◽  
Wojciech K. Kaniewski
Keyword(s):  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Capsid Protein ◽  
Field Trials ◽  
Enzyme Linked Immunosorbent Assay ◽  
Capsid Protein Gene ◽  
Cp Gene ◽  
Subgroup Ii ◽  
Transgenic Lines ◽  
Transgenic Tomatoes

Three processing tomato (Lycopersicon esculentum Mill.) lines engineered to express the cucumber mosaic virus (CMV) capsid protein (CP) gene were evaluated in the summers of 1995 and 1996 under high levels of naturally occurring CMV disease pressure. One tomato line expressed the capsid protein gene from a subgroup II isolate of CMV (line 11527), whereas two lines (12261 and 12295) expressed the capsid protein genes from a CMV subgroup I and a subgroup II isolate. Evaluation of CMV incidence based on symptomatic plants revealed that only 9% and 8% of the plants in line 11527 were infected in 1995 and 1996, respectively, 5 weeks after being transplanted. None of the plants in line 12261 developed symptoms in 1995, whereas 26% were symptomatic in 1996. There were no symptomatic plants in line 12295 in either the 1995 or the 1996 trial. In contrast to the CMV transgenic lines, 96% and 95% of the susceptible control plants were symptomatic by the 5-week rating period. CMV incidence in the CMV transgenic lines was much higher when infection was based on detection of virus by enzyme-linked immunosorbent assay (ELISA). This was particularly true in the 1996 trial where no less than 97% of the plants within a treatment were determined to be infected. Though a relatively high percentage of the transgenic plants were infected, the amount of CMV that accumulated in these plants was significantly less than in the susceptible controls, which may explain the occurrence of the attenuated symptoms. Despite CMV infection of the transgenic lines in the Alabama field trials, the performance of these lines could be of practical value to growers.

Virus resistance in transgenic plants that express cucumber mosaic virus satellite RNA

Nature ◽  
1987 ◽  
Vol 328 (6133) ◽  
pp. 799-802 ◽  
Author(s):  
Bryan D. Harrison ◽  
Michael A. Mayo ◽  
David C. Baulcombe
Keyword(s):  
Transgenic Plants ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Virus Resistance ◽  
Satellite Rna

scholarly journals Temporal and Spatial Appearance of Recombinant Viruses Formed Between Cauliflower Mosaic Virus (CaMV) and CaMV Sequences Present in Transgenic Nicotiana bigelovii

1998 ◽  
Vol 11 (4) ◽  
pp. 309-316 ◽  
Author(s):  
Lorant Király ◽  
June E. Bourque ◽  
James E. Schoelz
Keyword(s):  
Transgenic Plants ◽  
Mosaic Virus ◽  
Leaf Tissue ◽  
Cauliflower Mosaic Virus ◽  
Recombinant Viruses ◽  
Predominant Type ◽  
Nicotiana Bigelovii ◽  
Temporal And Spatial ◽  
Systemic Symptoms ◽  
Symptom Type

Cauliflower mosaic virus (CaMV) strain CM1841 is able to recombine with a CaMV transgene sequence present in Nicotiana bigelovii. In the present study we have characterized the temporal and spatial appearance of recombinant viruses formed between CM1841 and the transgene within individual transgenic plants. CM1841 infections were initiated by mechanical inoculation and by agro-inoculation to nontransformed N. bigelovii and transgenic N. bigelovii that expressed the gene VI product of CaMV strain D4. In agroinoculated transgenic plants, inoculated leaf tissue turned necrotic around the point of agroinocu-lation, while chlorotic lesions appeared in the leaves inoculated with CM1841 virions. The first systemic symptoms in both agroinoculated and mechanically inoculated transgenic N. bigelovii consisted of necrotic patches. The predominant type of virus recovered from the inoculated and first systemically infected leaves was the wild-type CM1841 rather than a recombinant. As the infection progressed in the transgenic plants, symptoms changed from necrosis in the lower leaves to a chlorotic mosaic in the upper leaves. This shift in symptom type was associated with the recovery of recombinant viruses, indicating that the recombinants predominated only in later stages of pathogenesis.

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