Cocksfoot Mottle Virus Research Articles

Sequencing and biological characterization of historical cynosurus mottle virus isolates from Germany.

We report sequencing of four historical cynosurus mottle virus (CnMoV) isolates, originating from different hosts and locations. The CnMoV genome, ranging from 4417 to 4419 nt, encodes five ORFs. It shares 48.1% nucleotide sequence identity with cocksfoot mottle virus and 69.8% with the recently discovered Poaceae Liege sobemovirus. Phylogenetic analysis supports classification within the genus Sobemovirus. Sequenced CnMoV isolates exhibit 96.4-99.9% identity. Nucleotide substitutions leading to amino acid changes showed no host associations. However, amino acid changes in the coat protein appear to be linked to differences in serological properties. Aphid transmission tests confirmed non-transmissibility, consistent with earlier observations for the English isolate.

First report of Cocksfoot mottle virus infecting wheat in Estonia

First report of <i>Cocksfoot mottle virus</i> infecting wheat in Estonia

Cocksfoot mottle virus (Cocksfoot mottle virus)

Cocksfoot mottle virus (Cocksfoot mottle virus)

Occurrence and High-Throughput Sequencing of Viruses in Ohio Wheat.

Ohio is a leading producer of soft red winter wheat in the United States. Many viruses impact wheat production, but there is a lack of contemporary information on the distribution and potential impact of wheat viruses in Ohio. To address this knowledge gap, we created a comprehensive dataset of viruses identified by high-throughput sequencing (HTS) and their incidence in field sites sampled across the state. Samples were collected from 103 field sites in surveys conducted in 2012, 2016, and 2017 and subjected to RNA HTS, reverse transcription (RT) PCR, or enzyme-linked immunosorbent assay to assess virus sequence diversity, prevalence, and incidence within fields. Partial and complete virus sequences were assembled and detection validated by RT-PCR. Assembled sequences were compared with previously known virus sequences, and novel sequences were validated by Sanger sequencing. The viruses detected most often included barley yellow dwarf virus (BYDV), cereal yellow dwarf virus (CYDV), wheat streak mosaic virus (WSMV), and wheat spindle streak mosaic virus (WSSMV). These viruses were detected at 67, 69, 55, and 28% of the field sites sampled, with mean incidences of 18, 19, 20, and 49%, respectively, within fields where they were detected. Brome mosaic virus (BMV) and cocksfoot mottle virus (CfMV) were also viruses of potential importance detected in Ohio, found in 26 and 17% of the field sites sampled, respectively. Based on results from logistic regression analyses, the presence of BYDV, CYDV, WSMV, and WSSMV was associated with the presence of volunteer wheat, BYDV and CfMV with monocots as the previous crop, and BMV with the presence of nearby corn fields (P < 0.10). For six viruses, there was evidence of spatial clustering in at least one field site and the variance of mean incidence was higher at the county level than at the regional spatial level. This finding suggests that county- and site-specific factors influenced the incidence and spatial pattern of some viruses. The results of this study provide a snapshot of viruses present in Ohio wheat and insights into their biology, potential risks to wheat production, and possible management strategies.

First Report of Cocksfoot Mottle Virus Infecting Dactylis glomerata in Forage Production Fields in California.

Orchardgrass (Dactylis glomerata L.) is a cool season perennial grass commonly used in pastures and also grown for hay and silage. A Sobemovirus infecting orchardgrass, cocksfoot mottle virus (CfMV), was originally described in 1963 in England (Serjeant 1964). The symptoms of CfMV are visible in spring with yellow streaking in mild cases, but these symptoms can progress into whitening of the leaves, followed by necrosis and plant death in severe cases (Serjeant 1964). Although beetles (Oulemma spp.) are thought to introduce this virus into pastures, mechanical transmission is believed to be responsible for the spread of this virus once it has been introduced into an area (Serjeant 1964). This virus is associated with stand decline in orchardgrass fields in Europe, Japan, New Zealand (Mahy and Van Regenmortel 2010), Canada (Bittman et al. 2006), and recently in U.S. seed production fields in Oregon, where 79% of the orchardgrass fields surveyed in 2016 were infected with CfMV (Gilmore et al. 2017). Subsequently, in early April of 2017, several forage growers in northern California that were experiencing extensive die-off in their fields requested testing for CfMV and other pathogens. Orchardgrass samples were tested for CfMV using double antibody sandwich ELISA, with antiserum developed to an isolate of CfMV from British Columbia, Canada (R. R. Martin). Ten samples, collected (into individual plastic bags) from areas of the field with die-off, were ELISA positive for CfMV, and one was also positive for cereal yellow dwarf virus (CYDV). Five healthy samples collected from this same field tested negative for CYDV and CfMV. Total RNA was extracted from four of the CfMV-ELISA-positive samples following the nucleic acid extraction protocol of Tzanetakis et al. (2007) and used for cDNA synthesis using the SuperScriptIII First-Strand Synthesis SuperMix kit (Invitrogen by Life Technologies, CA). PrimeSTAR HS DNA Polymerase (Takara Clonetech, Japan) was used for polymerase chain reaction with CfMV-specific primers CfCP-F1 (GATGGAGCCAGTCTCTCG) and CfCP-R2 (ATCCGTCAATCTTCAAGC) (Gilmore et al. 2017) using a BioRad thermal cycler with the following program: 98°C for 2 min; 40 cycles of 98°C for 15 s, 55°C for 5 s, and 72°C for 45 s; and 72°C for 7 min. The predominant band at ≈750 bp was reamplified and sequenced using CfCP-F1 and CfCP-R2 primers. A BLAST search indicated one sample with 99% nucleotide sequence identity to CfMV reported in the Willamette Valley, OR, and three samples that were most closely related (≈95 to 96% identity) to CfMV reported in Ohio. Sequences have been submitted to GenBank (accession nos. MG765457 to MG765460). To our knowledge, this is the first report of CfMV in orchardgrass forage production fields in California. Although infection with CfMV can be lethal (Serjeant 1964), it is also possible that the presence of CfMV in combination with the drier and hotter summers experienced in the last few years may have weakened the plants and increased susceptibility to other pathogens, leading to the devastation observed in orchardgrass fields in northern California. Growers are encouraged to look for symptoms of CfMV in early spring and prevent mechanical transmission of CfMV by harvesting clean fields first, or by cleaning machinery thoroughly between fields. Additionally, growers are encouraged to grow CfMV-resistant cultivars, if possible. Cultivars Cheam-VR, Haida-VR, and Chilliwack-VR are listed as resistant to CfMV.

Isolation and Characterization of Two Distinct Types of Unmodified Spherical Plant Sobemovirus-Like Particles for Diagnostic and Technical Uses.

Plant virus-like particles (VLPs) structurally resemble their progenitor viruses, but are noninfectious due to absence of viral nucleic acids. Since the 1980s, VLPs have been actively studied with the aim of constructing different nanomaterials, including immunologically active carriers for peptides and whole proteins and proteinaceous shells for the packaging of different ligands.The technological developments using VLPs require large amounts of purified particles. Here, we describe the laboratory process for isolation and purification of two unmodified plant VLPs, derived from two sobemoviruses, cocksfoot mottle virus (CfMV) and rice yellow mottle virus (RYMV), which is based on cultivation of recombinant Escherichia coli cells, VLP precipitation from bacterial extracts and ultracentrifugation. The suggested purification scheme allows the production of 4-45 mg of purified sobemoviral VLPs from a 1l bacterial culture, depending on the required purity level. Additionally, we provide short protocols for VLP characterization using SDS-PAGE, agarose gel electrophoresis, ultraviolet and mass spectrometry, dynamic light scattering, and electron microscopy.

First Report of Cocksfoot mottle virus Infecting Wheat (Triticum aestivum) in Ohio

Cocksfoot mottle virus (CfMV) was discovered in Ohio wheat during a 2016 field survey utilizing RNA-Seq to identify virus-like sequences. Virus sequences were confirmed by reverse transcriptase-polymerase chain reaction (RT-PCR) and Sanger sequencing, and CfMV was transmitted to orchardgrass and passaged to wheat to isolate virus. This is the first report of CfMV in Ohio, and the first report of CfMV naturally infecting wheat.

First Report of Cocksfoot mottle virus Infecting Dactylis glomerata in Poland

First Report of <i>Cocksfoot mottle virus</i> Infecting <i>Dactylis glomerata</i> in Poland

P1-independent replication and local movement of Rice yellow mottle virus in host and non-host plant species

Sobemovirus P1 protein, characterized previously as a suppressor of posttranscriptional gene silencing, is required for systemic virus spread and infection in plants. Mutations in the ORF1 initiation codon do not affect viral replication indicating P1 is not necessary for this process. Wild type, recombinant and P1 deletion mutants of Cocksfoot mottle virus and Rice yellow mottle virus were used to infect oat, rice, wheat, barley, Arabidopsis thaliana and Nicotiana benthamiana plants. Wild type RYMV, RYMV without P1 and RYMV with CfMV P1 were detected in inoculated leaves of all tested plant species. We found that RYMV does not need P1 for replication and for local movement neither in host nor non-host species tested in this study. However, it is crucial for successful systemic spread of the virus in its host plant rice. Moreover, adding CfMV P1 into RYMV genome did not help it to overcome restriction to the inoculated leaf.

First Report of Cocksfoot mottle virus Infecting Dactylis glomerata in Oregon and the United States

HomePlant DiseaseVol. 100, No. 5First Report of Cocksfoot mottle virus Infecting Dactylis glomerata in Oregon and the United States PreviousNext DISEASE NOTES OPENOpen Access licenseFirst Report of Cocksfoot mottle virus Infecting Dactylis glomerata in Oregon and the United StatesS. C. Alderman, R. C. Martin, B. S. Gilmore, R. R. Martin, G. D. Hoffman, C. S. Sullivan, and N. P. AndersonS. C. AldermanSearch for more papers by this author, R. C. MartinSearch for more papers by this author, B. S. GilmoreSearch for more papers by this author, R. R. MartinSearch for more papers by this author, G. D. HoffmanSearch for more papers by this author, C. S. SullivanSearch for more papers by this author, and N. P. AndersonSearch for more papers by this authorAffiliationsAuthors and Affiliations S. C. Alderman R. C. Martin B. S. Gilmore , USDA-ARS FSCRU, Corvallis, OR 97331 R. R. Martin , USDA-ARS HCRU, Corvallis, OR 97330 G. D. Hoffman , Department of Crop and Soil Science, Oregon State University, Corvallis, OR 97331 C. S. Sullivan , Linn County Extension, Tangent, OR 97389 N. P. Anderson , Yamhill County Extension, McMinnville, OR 97128. Published Online:3 Mar 2016https://doi.org/10.1094/PDIS-09-15-1017-PDNAboutSections ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinked InRedditEmailWechat Cocksfoot mottle virus (CfMV) is a mechanically and beetle-transmitted, non-seed-transmitted sobemovirus associated with orchardgrass (Dactylis glomerata L.) stand decline in Europe, Japan, New Zealand (Mahy and Van Regenmortel 2010), and Canada (Bittman et al. 2006). Additional hosts, reported from New Zealand (Delmiglio et al. 2010), include Festuca novae-zelandiae, Lolium spp., Poa anceps, Poa cita, Chionochloa rubra, and Dichelachne crinita. To determine if the virus is present in the United States, surveys for CfMV were conducted in 2014 and 2015 in orchardgrass seed production fields in the Willamette Valley in Oregon, where most of the orchardgrass seed produced in the United States is grown. During June, in each of 2014 and 2015, 18 orchardgrass fields were selected arbitrarily. Stand age of the seed production fields ranged from 2 to 24 years. Two of the fields were sampled in both years. Four samples (each containing 4 leaves, one from each of four plants) were collected along each of four transects in an M-shape pattern from each field. All plants sampled appeared healthy. In 2014, samples were placed in separate plastic bags and in 2015 samples were placed in deep well plates (VWR, International LLC, Radnor, PA). In each year, samples were transported over ice and stored at 5°C until processed. Samples were homogenized and tested for CfMV using DAS-ELISA, with antibodies derived from an isolate of CfMV from British Columbia, Canada. In 2014 and 2015, CfMV was detected in 61% and 72% of the fields, respectively. Symptoms were not present at the time of sampling. In 2015, eight ELISA positive samples were further examined with RT-PCR. RNA was extracted with the Direct-zol RNA MiniPrep kit (Zymo Research, Irvine, CA) following the manufacturer’s instructions and used for cDNA synthesis using the SuperScriptIII First-Strand Synthesis SuperMix kit (Life Technologies, ThermoFisher, Waltham, MA). PrimeSTAR HS DNA Polymerase (Takara Clonetech, Kyoto, Japan) was used following the manufacturer’s instructions with CfMV specific primers CfCP-F1 (GATGGAGCCAGTCTCTCG) and CfCP-R2 (ATCCGTCAATCTTCAAGC) (Delmiglio 2008) for PCR with the following program: 98°C for 2 min; 40 cycles of 98°C for 15 s, 55°C for 5 s, and 72°C for 45 s; followed by 72°C for 7 min. The predominant band at ∼750 bp was reamplified and sequenced using CfCP-F1 and CfCP-R1 primers. A BLAST search indicated 94.4 to 95.4% identity with CfMV isolates found in native and exotic grasses of New Zealand (Delmiglio 2008). Sequences were submitted to GenBank (Accession No. KT984653 to KT984660). To our knowledge, this is the first report of CfMV in orchardgrass seed production fields in Oregon and in the United States. Results of this study are significant given the severity of CfMV in Canada and elsewhere, and this could account for the winter die-out occurring in orchardgrass seed-production fields in Oregon. Additional studies will be needed to determine to what extent CfMV is associated with stand decline in Oregon and what vectors may be present.

Cocksfoot mottle virus coat protein is dispensable for the systemic infection

BackgroundThe Sobemovirus genome consists of polycistronic single-stranded positive-sense RNA. The first ORF encodes P1, a suppressor of RNA silencing required for virus movement. The coat protein (CP) is expressed from the 3′ proximal ORF3 via subgenomic RNA. In addition to its structural role, the CP of some sobemoviruses has been reported to be required for systemic movement and to interact with P1. The aim of this study was to analyse the role of Cocksfoot mottle virus (CfMV) CP in the suppression of RNA silencing and virus movement.MethodsAgrobacterium-mediated transient expression method was used for testing CfMV CP capacity to suppress RNA silencing. CP substitution and deletion mutants were generated to examine the role of this protein in CfMV infection, using three host plants (oat, barley and wheat). The viral movement was characterised with CfMV expressing EGFP fused to the C-terminus of CP.ResultsIn the current study we show that CfMV CP is an additional RNA silencing suppressor. Interestingly, we observed that all CP mutant viruses were able to infect the three tested host plants systemically, although usually with reduced accumulation. CfMV expressing EGFP was detected in epidermal and mesophyll cells of inoculated leaves. Although EGFP fluorescence was not detected in upper leaves, some plants displayed CfMV symptoms. Analysis of the upper leaves revealed that the viruses had lost the EGFP sequence and sometimes also most of the CP gene.ConclusionsThe present study demonstrates that CfMV CP suppresses RNA silencing but, surprisingly, is dispensable for systemic movement. Thus, CfMV does not move as virion in the tested host plants. The composition of the movement RNP complex remains to be elucidated.

Viruses of New Zealand pasture grasses and legumes: a review

This article reviews knowledge of 23 plant viruses infecting pasture grasses and legumes in New Zealand. The incidence, ecology and impact of each virus and prospects for control using natural or artificial resistance genes or by vector control is discussed. The most prevalent viruses are Alfalfa mosaic virus and White clover mosaic virus in pasture legumes and Cocksfoot mottle virus, Ryegrass mosaic virus and Barley yellow dwarf virus in pasture grasses. Lucerne Australian latent virus is restricted to the North Island and Red clover necrotic mosaic virus is largely restricted to the South Island. These patterns are likely to be dynamic with ongoing changes in weather patterns, land use, the spread of insect vectors and the continuing introduction of viruses and vectors. The existing and potential threats to 12 pasture species are tabulated and the knowledge gaps for each species highlighted. Control of vectors including aphids, eriophyid mites and soil-borne fungi is probably not economic per se but could be an additional benefit of integrated pest management in pasture and cropping systems. The most cost-effective and practical preventative measures are likely to be the use of virus-tested seed to establish new pastures and the incorporation of resistance genes by conventional breeding or by genetic engineering. Finally, recommendations are made for future research for New Zealand, which is also relevant to other temperate regions of the world.

The crystallographic structure of Panicum Mosaic Virus (PMV).

The structure of Panicum Mosaic Virus (PMV) was determined by X-ray diffraction analysis to 2.9Å resolution. The crystals were of pseudo symmetry F23; the true crystallographic unit cell was of space group P2(1) with a=411.7Å, b=403.9Å and c=412.5Å, with β=89.7°. The asymmetric unit was two entire T=3 virus particles, or 360 protein subunits. The structure was solved by conventional molecular replacement from two distant homologues, Cocksfoot Mottle Virus (CfMV) and Tobacco Necrosis Virus (TNV), of ∼20% sequence identity followed by phase extension. The model was initially refined with exact icosahedral constraints and then with icosahedral restraints. The virus has Ca(++) ions octahedrally coordinated by six aspartic acid residues on quasi threefold axes, which is completely different than for either CfMV or TNV. Amino terminal residues 1-53, 1-49 and 1-21 of the A, B and C subunits, respectively, and the four C-terminal residues (239-242) are not visible in electron density maps. The additional ordered residues of the C chain form a prominent "arm" that intertwines with symmetry equivalent "arms" at icosahedral threefold axes, as was seen in both CfMV and TNV. A 17 nucleotide hairpin segment of genomic RNA is icosahedrally ordered and bound at 60 equivalent sites at quasi twofold A-B subunit interfaces at the interior surface of the capsid. This segment of RNA may serve as a conformational switch for coat protein subunits, as has been proposed for similar RNA segments in other viruses.

Cocksfoot mottle sobemovirus establishes infection through the phloem

Cocksfoot mottle virus (CfMV) localization in oat plants was analyzed during three weeks post infection by immunohistochemical staining to follow its spread through different tissues. In early stages of infection, the virus was first detectable in phloem parenchyma and bundle sheath cells of inoculated leaves. Bundle sheath and phloem parenchyma were also the cell types where the virus was first detected in stems and systemic leaves of infected plants. In later stages of infection, CfMV spread also into the mesophyll surrounding vascular bundles and was seldom detected in xylem parenchyma of inoculated leaves. In systemic leaves, CfMV was not detected from xylem. Moreover, sometimes it was found from phloem only. In straw and roots, CfMV was detected both from phloem and xylem. According to our observations, CfMV predominantly moves through phloem, which makes the systemic movement of CfMV different from that of another monocot-infecting sobemovirus, Rice yellow mottle virus (RYMV).

Protein-RNA linkage and post-translational modifications of two sobemovirus VPgs

Sobemoviruses possess a viral genome-linked protein (VPg) attached to the 5' end of viral RNA. VPg is processed from the viral polyprotein. In the current study, Cocksfoot mottle virus (CfMV) and Rice yellow mottle virus (RYMV) VPgs were purified from virions and analysed by mass spectrometry. The cleavage sites in the polyprotein and thereof the termini of VPg were experimentally proven. The lengths of the mature VPgs were determined to be 78 and 79 aa residues, respectively. The amino acid residues covalently linked to RNA in the two VPgs were, surprisingly, not conserved; it is a tyrosine at position 5 of CfMV VPg and serine at position 1 of RYMV VPg. Phosphorylations were identified in CfMV and RYMV VPgs with two positionally similar locations T20/S14 and S71/S72, respectively. RYMV VPg contains an additional phosphorylation site at S41.

Incidence of cereal and pasture viruses in New Zealand's native grasses

This study provides evidence for frequent and multiple invasions of New Zealand's native grasses by exotic cereal and pasture viruses. Fifteen native and three exotic grasses from 29 North Island and six South Island sites were surveyed for the presence of viruses using enzyme‐linked immunosorbent assay (ELISA). Barley yellow dwarf virus and Cereal yellow dwarf virus (BYDV, CYDV: Luteoviridae) and Cocksfoot mottle virus (CoMV, Sobemovirus) are widespread throughout New Zealand. CoMV, previously considered to have a natural host range restricted to Dactylis and Triticum, was detected in Poa anceps, P. cita, Festuca novae‐zelandiae, and Chionochloa rubra. New virus host reports include BYDV‐PAV in Microlaena stipoides and Dichelachne crinita; BYDV‐MAV in P. cita, F. novae‐zelandiae and Hierochloe redolens; and CYDV‐RPV in P. cita and M. stipoides. Nominal logistic regression analyses showed a correlation between the presence of exotic grass species and virus incidence. Host range experiments for BYDV‐PAV and CoMV were performed with selected native and exotic grasses, and the results are discussed in context of the field‐survey findings.

Cocksfoot mottle sobemovirus coat protein contains two nuclear localization signals

Cocksfoot mottle virus (CfMV) coat protein (CP) localization was studied in plant and mammalian cells. Fusion of the full-length CP with enhanced green fluorescent protein (EGFP) localized to the cell nucleus whereas similar constructs lacking the first 33 N-terminal amino acids of CP localized to the cytoplasm. CP and EGFP fusions containing mutations in the arginine-rich motif of CP localized to the cytoplasm and to the nucleus in plant cells indicating the involvement of the motif in nuclear localization. In mammalian cells, mutations in the arginine-rich region were sufficient to completely abolish nuclear transport. The analysis of deletions of amino acid residues 1-11, 1-22, and 22-33 of CP demonstrated that there were two separate nuclear localization signals (NLS) within the N-terminus--a strong NLS1 in the arginine-rich region (residues 22-33) and a weaker NLS2 within residues 1-22. Analysis of point mutants revealed that the basic amino acid residues in the region of the two NLSs were individually not sufficient to direct CP to the nucleus. Additional microinjection studies with fluorescently labeled RNA and CP purified from CfMV particles demonstrated that the wild-type CP was capable of transporting the RNA to the nucleus. This feature was not sequence-specific in transient assays since both CfMV and GFP mRNA were transported to the cell nucleus by CfMV CP. Together the results suggest that the nucleus may be involved in CfMV infection.

Stem-loop structure of Cocksfoot mottle virus RNA is indispensable for programmed −1 ribosomal frameshifting

The −1 programmed ribosomal frameshifting (−1 PRF) mechanism utilized by many viruses is dependent on a heptanucleotide slippery sequence and a downstream secondary structure element. In the current study, the RNA structure downstream from the slippery site of cocksfoot mottle sobemovirus (CfMV) was proven to be a 12 bp stem-loop with a single bulge and a tetranucleotide loop. Several deletion and insertion mutants with altered stem-loop structures were tested in wheat germ extract (WGE) for frameshifting efficiency. The impact of the same mutations on virus infectivity was tested in oat plants. Mutations shortening or destabilizing the stem region reduced significantly but did not abolish −1 PRF in WGE. The same mutations proved to be deleterious for virus infection. However, extending the loop region to seven nucleotides had no significant effect on frameshifting efficiency in WGE and did not hamper virus replication in infected leaves. This is the first report about the experimentally proven RNA secondary structure directing −1 PRF of sobemoviruses.

Biological and molecular characterization of a putative new sobemovirus infecting Imperata cylindrica and maize in Africa

A new virus was isolated from both the grass Imperata cylindrica and maize plants that had yellow mottle symptoms in Burkina Faso, West Africa. The virus has isometric particles ca. 32 nm in diameter. The experimental host range was restricted to Rottboellia exaltata. Virions were isolated from leaves of systemically infected maize plants. Koch's postulates were completed by mechanically inoculating uninfected Imperata or maize with either purified virus or sap from infected Imperata plants. Virion preparations were used to produce a specific polyclonal antiserum, and an enzyme-linked immunosorbent assay test was set up. The full genome of the virus was sequenced, and it comprised 4,547 nucleotides. Phylogenetic studies indicated that the virus is closely related to rice yellow mottle virus, a sobemovirus that infects monocotyledons in Africa, and is more distantly related to cocksfoot mottle virus, another sobemovirus that infects monocotyledons. Although the virus can infect R. exaltata experimentally, it differs from Rottboellia yellow mottle virus, a member of a tentative species of the genus Sobemovirus that also infects monocotyledons in Africa. Particle morphology, serological properties, genomic organization, and phylogenetic analysis are all consistent with assignment of the new virus to the genus Sobemovirus. The name Imperata yellow mottle virus is proposed.

Phenotypes and Functional Effects Caused by Various Viral RNA Silencing Suppressors in Transgenic Nicotiana benthamiana and N. tabacum

RNA silencing suppressor genes derived from six virus genera were transformed into Nicotiana benthamiana and N. tabacum plants. These suppressors were P1 of Rice yellow mottle virus (RYMV), P1 of Cocksfoot mottle virus, P19 of Tomato bushy stunt virus, P25 of Potato virus X, HcPro of Potato virus Y (strain N), 2b of Cucumber mosaic virus (strain Kin), and AC2 of African cassava mosaic virus (ACMV). HcPro caused the most severe phenotypes in both Nicotiana spp. AC2 also produced severe effects in N. tabacum but a much milder phenotype in N. benthamiana, although both HcPro and AC2 affected the leaf tissues of the two Nicotiana spp. in similar ways, causing hyperplasia and hypoplasia, respectively. P1-RYMV caused high lethality in the N. benthamiana plants but only mild effects in the N. tabacum plants. Phenotypic alterations produced by the other transgenes were minor in both species. Interestingly, the suppressors had very different effects on crucifer-infecting Tobamovirus (crTMV) infections. AC2 enhanced both spread and brightness of the crTMV-green fluorescent protein (GFP) lesions, whereas 2b and both P1 suppressors enhanced spread but not brightness of these lesions. P19 promoted spread of the infection into new foci within the infiltrated leaf, whereas HcPro and P25 suppressed the spread of crTMV-GFP lesions.