Limited potexvirus diversity in eastern Gulf of Mexico seagrass meadows.

Turtlegrass virus X, which infects the seagrass Thalassia testudinum, is the only potexvirus known to infect marine flowering plants. We investigated potexvirus distribution in seagrasses using a degenerate reverse transcription polymerase chain reaction (RT-PCR) assay originally designed to capture potexvirus diversity in terrestrial plants. The assay, which implements Potex-5 and Potex-2RC primers, successfully amplified a 584 nt RNA-dependent RNA polymerase (RdRp) fragment from TVX-infected seagrasses. Following validation, we screened 74 opportunistically collected, apparently healthy seagrass samples for potexviruses using this RT-PCR assay. The survey examined the host species T. testudinum, Halodule wrightii, Halophila stipulacea, Syringodium filiforme, Ruppia maritima, and Zostera marina. Potexvirus PCR products were successfully generated only from T. testudinum samples and phylogenetic analysis of sequenced PCR products revealed five distinct TVX sequence variants. Although the RT-PCR assay revealed limited potexvirus diversity in seagrasses, the expanded geographic distribution of TVX shown here emphasizes the importance of future studies to investigate T. testudinum populations across its native range and understand how the observed fine-scale genetic diversity affects host-virus interactions.

Biological and molecular characterization of a new potexvirus isolated from Adenium obesum.

Adenium obesum plants showing virus-like symptoms were collected in several regions of Brazil. Mottling symptoms like those observed in symptomatic plants in the field were reproduced in mechanically inoculated A. obesum plants. This potexvirus was named "desert rose mottle virus" (DRMoV), and its genome sequence was first determined by high-throughput sequencing and then confirmed by Sanger sequencing. The complete genome of DRMoV is 6,781 nt in length, excluding the poly(A) tail, and five ORFs were predicted in order from 5' to 3': Rep-TGB1-TGB2-TGB3-CP. Phylogenetic analysis based on Rep amino acid sequences showed different clustering among potexviruses. These data suggest that RDMoV is a new member of the genus Potexvirus, and the binomial name "Potexvirus adenii" is proposed for its species.

An Unwanted Association: The Threat to Papaya Crops by a Novel Potexvirus in Northwest Argentina.

An emerging virus isolated from papaya (Carica papaya) crops in northwestern (NW) Argentina was sequenced and characterized using next-generation sequencing. The resulting genome is 6667-nt long and encodes five open reading frames in an arrangement typical of other potexviruses. This virus appears to be a novel member within the genus Potexvirus. Blast analysis of RNA-dependent RNA polymerase (RdRp) and coat protein (CP) genes showed the highest amino acid sequence identity (67% and 71%, respectively) with pitaya virus X. Based on nucleotide sequence similarity and phylogenetic analysis, the name papaya virus X is proposed for this newly characterized potexvirus that was mechanically transmitted to papaya plants causing chlorotic patches and severe mosaic symptoms. Papaya virus X (PapVX) was found only in the NW region of Argentina. This prevalence could be associated with a recent emergence or adaptation of this virus to papaya in NW Argentina.

Subcellular localization and interactions among TGB proteins of cowpea mild mottle virus.

Cowpea mild mottle virus (CPMMV) is a flexuous filamentous virus that belongs to the genus Carlavirus (family Betaflexiviridae). The CPMMV genome contains six open reading frames (ORFs), among which the triple gene block (TGB), encoded by ORFs 2 to 4, has been reported to encode movement proteins for different viruses. The subcellular localization of the TGB proteins of CPMMV isolate CPMMV:BR:MG:09:2 was analysed by transient expression of each protein fused to a fluorophore. Overall, the accumulation pattern and interactions among CPMMV TGB proteins (TGBp) were similar to those of their counterparts from the potex-like group. Considering these similarities, we evaluated the potential interactions between the TGB proteins of CPMMV and of potato virus X, which could complement cell-to-cell movement. The TGBp2 and TGBp3 of PVX had an effect on CPMMV TGBp1, directing it to the plasmodesmata, but the reverse was not true.

The Phylogeography of Potato Virus X Shows the Fingerprints of Its Human Vector.

Potato virus X (PVX) occurs worldwide and causes an important potato disease. Complete PVX genomes were obtained from 326 new isolates from Peru, which is within the potato crop's main domestication center, 10 from historical PVX isolates from the Andes (Bolivia, Peru) or Europe (UK), and three from Africa (Burundi). Concatenated open reading frames (ORFs) from these genomes plus 49 published genomic sequences were analyzed. Only 18 of them were recombinants, 17 of them Peruvian. A phylogeny of the non-recombinant sequences found two major (I, II) and five minor (I-1, I-2, II-1, II-2, II-3) phylogroups, which included 12 statistically supported clusters. Analysis of 488 coat protein (CP) gene sequences, including 128 published previously, gave a completely congruent phylogeny. Among the minor phylogroups, I-2 and II-3 only contained Andean isolates, I-1 and II-2 were of both Andean and other isolates, but all of the three II-1 isolates were European. I-1, I-2, II-1 and II-2 all contained biologically typed isolates. Population genetic and dating analyses indicated that PVX emerged after potato's domestication 9000 years ago and was transported to Europe after the 15th century. Major clusters A-D probably resulted from expansions that occurred soon after the potato late-blight pandemic of the mid-19th century. Genetic comparisons of the PVX populations of different Peruvian Departments found similarities between those linked by local transport of seed potato tubers for summer rain-watered highland crops, and those linked to winter-irrigated crops in nearby coastal Departments. Comparisons also showed that, although the Andean PVX population was diverse and evolving neutrally, its spread to Europe and then elsewhere involved population expansion. PVX forms a basal Potexvirus genus lineage but its immediate progenitor is unknown. Establishing whether PVX's entirely Andean phylogroups I-2 and II-3 and its Andean recombinants threaten potato production elsewhere requires future biological studies.

Potato virus X-mediated constitutive expression of Plutella xylostella PxSDF2L1 gene in Nicotiana benthamiana confers resistance to Phytophthora parasitica var. nicotianae.

BACKGROUND: The Plutella xylostella PxSDF2L1 gene was previously reported to enhance insect resistance to pathogen at high basal transcription rate. PxSDF2L1 shows similitude with the stromal cell-derived factor 2 (SDF2), an ER stress-induced chaperon protein that is highly conserved throughout animals and plants. The precise biological function of SDF2 is not clear, but its expression is required for innate immunity in plants. Here, we investigate whether a continuous expression of PxSDF2L1 in Nicotiana benthamiana can similarly confer resistance to plant pathogen, particularly, the black shank Phytophthora parasitica var. nicotianae. RESULTS: The N. benthamiana plants were inoculated with agrobacteria transformed with a PVX-based binary vector carrying the PxSDF2L1 gene; similar agroinoculation experiments with a PVX vector carrying the GFP gene were used for controls. In pot trials, agroinfected N. benthamiana plants constitutively expressing PxSDF2L1 showed a significant reduction of stem disease symptoms caused by the inoculation with P. parasitica, compared with controls. CONCLUSIONS: We confirm a role of PxSDF2L1 in resistance to black shank, with a potential application to engineering active resistance against this oomycete in the commercial N. tabacum species and propose its evaluation in other crop families and plant pathogens.

Molecular Viral Diagnosis and Sanitation of Yam Genetic Resources: Implications for Safe Yam Germplasm Exchange.

Yam (Dioscorea spp.) is an important crop in tropical and subtropical regions. Many viruses have been recently identified in yam, hampering genetic conservation and safe international exchanges of yam germplasm. We report on the implementation of reliable and cost-effective PCR-based detection tools targeting eight different yam-infecting viruses. Viral indexing of the in vitro yam collection maintained by the Biological Resources Center for Tropical Plants (BRC-TP) in Guadeloupe (French West Indies) unveiled a high prevalence of potyviruses, badnaviruses, Dioscorea mosaic associated virus (DMaV) and yam asymptomatic virus 1 (YaV1) and a high level of coinfections. Infected yam accessions were subjected to a combination of thermotherapy and meristem culture. Sanitation levels were monitored using PCR-based and high-throughput sequencing-based diagnosis, confirming the efficacy and reliability of PCR-based detection tools. Sanitation rates were highly variable depending on viruses. Sixteen accessions were successfully sanitized, paving the way to safe yam germplasm exchanges and the implementation of clean seed production programs worldwide.

Differential Accumulation of Innate- and Adaptive-Immune-Response-Derived Transcripts during Antagonism between Papaya Ringspot Virus and Papaya Mosaic Virus.

Papaya ringspot virus (PRSV), a common potyvirus infecting papaya plants worldwide, can lead to either antagonism or synergism in mixed infections with Papaya mosaic virus (PapMV), a potexvirus. These two unrelated viruses produce antagonism or synergism depending on their order of infection in the plant. When PRSV is inoculated first or at the same time as PapMV, the viral interaction is synergistic. However, an antagonistic response is observed when PapMV is inoculated before PRSV. In the antagonistic condition, PRSV is deterred from the plant and its drastic effects are overcome. Here, we examine differences in gene expression by high-throughput RNA sequencing, focused on immune system pathways. We present the transcriptomic expression of single and mixed inoculations of PRSV and PapMV leading to synergism and antagonism. Upregulation of dominant and hormone-mediated resistance transcripts suggests that the innate immune system participates in synergism. In antagonism, in addition to innate immunity, upregulation of RNA interference-mediated resistance transcripts suggests that adaptive immunity is involved.

Characterization of a not so new potexvirus from babaco (Vasconcellea x heilbornii).

A new member of the genus Potexvirus was fully sequenced and characterized. The virus was isolated from babaco (Vasconcellea x heilbornii), a natural hybrid native to Ecuador. The virus contains a 6,692 nt long genome organized in five open reading frames in an arrangement typical of other potexviruses. Sequence comparisons revealed close relatedness with Papaya mosaic virus (PapMV), Alternathera mosaic virus (AltMV) and Senna mosaic virus (SenMV), exhibiting nucleotide identities up to 67% for the polymerase (Pol) and 68% for the coat protein (CP), with deduced amino acid identities of 70% and 72% for the Pol and CP, respectively. The presence of an AlkB domain, in the polymerase region, was observed. Terminal nucleotide sequences were conserved across potexviruses with characteristic motifs and predicted secondary structures at the 3' UTR. Although serologically undistinguishable from PapMV and AltMV, the new virus showed differences in host range and symptom induction. The name babaco mosaic virus is proposed for this newly characterized Potexvirus. The complete genome sequence of the new virus has been deposited in NCBI GenBank under accession number MF978248.

Resolution of cassava-infecting alphaflexiviruses: Molecular and biological characterization of a novel group of potexviruses lacking the TGB3 gene.

Several potexviruses (Family Alphaflexiviridae) have been reported infecting cassava (Manihot esculenta Crantz) in the Americas. They were isolated from severely diseased plants during the last 30-40 years and include: Cassava common mosaic virus (CsCMV), Cassava Caribbean mosaic virus (CsCaMV), Cassava Colombian symptomless virus (CsCSV) and Cassava virus X (CsVX). However, their definitive classification as distinct species remains unresolved for several reasons, including the lack of sequence data and unavailability of samples from original isolates. This complicates disease diagnostics, cassava germplasm exchange certification, evaluation of virus cleaning protocols and epidemiological studies. Furthermore, a recently detected novel alphaflexivirus, indicates that cassava-infecting potexviruses may be more diverse. To solve the identity of these viruses, we started indexing samples from different parts of Colombia using different sets of PCR primers, antisera available and inoculation to indicator plants. Results show that there are three major phylogenetic groups of potexviruses infecting cassava, and they correspond to CsCMV, CsVX and the newly identified Cassava new alphaflexivirus (CsNAV). Bioassays and sequence analysis established that isolates of CsNAV and CsVX cause latent infections in different cassava landraces, they are not efficiently transmitted to the indicator plant Nicotiana benthamiana and they lack the gene 3 of the conserved potexviral 'triple gene block' (TGB). In contrast, all isolates of CsCMV (which have a characteristic potexvirus genome arrangement) caused Cassava Common Mosaic Disease (CCMD) in single infections and were efficiently transmitted to N. benthamiana. Although phylogenetic analysis of the replicase sequence placed CsNAV and CsVX as members of the Potexvirus genus, their distinct genome arrangement and biological characteristics suggest they can be considered as members of a separate taxonomic group.

Strain-dependent mutational effects for Pepino mosaic virus in a natural host.

BACKGROUND: Pepino mosaic virus (PepMV) is an emerging plant pathogen that infects tomatoes worldwide. Understanding the factors that influence its evolutionary success is essential for developing new control strategies that may be more robust against the evolution of new viral strains. One of these evolutionary factors is the distribution of mutational fitness effect (DMFE), that is, the fraction of mutations that are lethal, deleterious, neutral, and beneficial on a given viral strain and host species. The goal of this study was to characterize the DMFE of introduced nonsynonymous mutations on a mild isolate of PepMV from the Chilean 2 strain (PepMV-P22). Additionally, we also explored whether the fitness effect of a given mutation depends on the gene where it appears or on epistatic interactions with the genetic background. To address this latter possibility, a subset of mutations were also introduced in a mild isolate of the European strain (PepMV-P11) and the fitness of the resulting clones measured. RESULTS: A collection of 25 PepMV clones each containing a single nucleotide nonsynonymous substitution was created by site-directed mutagenesis and the fitness of each mutant was determined. PepMV-P22 genome showed a high degree of robustness against point mutations, with 80% of mutations being either neutral or even beneficial and only 20% being deleterious or lethal. We found that the effect of mutations strongly depended on the gene in which they were introduced. Mutations with the largest average beneficial effects were those affecting the RdRp gene, in contrast to mutations affecting TGB1 and CP genes, for which the average effects were deleterious. Moreover, significant epistatic interactions were observed between nonsynonymous mutations and the genetic background, meaning that the effect of a given nucleotide substitution on a particular genomic context cannot be predicted by knowing its effect in a different one. CONCLUSIONS: Our results indicated that PepMV genome has a surprisingly high robustness against mutations. We also found that fitness consequences of a given mutation differ between the two strains analyzed. This discovery suggests that the strength of selection, and thus the rates of evolution, vary among PepMV strains.

Biological and molecular characterization of a putative new potexvirus infecting Senna occidentalis.

In this work, we report the complete genome sequence of, production of polyclonal antibodies against, and development of biological assays for a putative new potexvirus, named senna mosaic virus (SenMV), found infecting Senna occidentalis in the state of São Paulo, Brazil. The complete genome sequence of SenMV comprises 6775 nucleotides excluding the poly(A) tail. The genome organization is similar to those of other potexviruses, with five open reading frames coding for RNA-dependent RNA polymerase (RdRp), the triple gene block (TGB 1, 2, and 3) proteins, and coat protein (CP). The virus was transmitted to S. occidentalis by mechanical inoculation and trimming scissors, but not by seeds.

Combining abilities for agronomic traits and marker-assisted selection for Potato virus X and Potato virus Y resistance.

Disease-resistant potato cultivars with good tuber appearance and desirable agronomic traits are essential for meeting the demands of producers and the market. Attaining these cultivars is the focus of potato breeding programs whose aim is to benefit the productive chain. The purpose of this study was to estimate combining abilities and evaluate potato clones based on tuber appearance, yield, and resistance to the PVY and PVX viruses. Crosses between four commercial cultivars of potato with good tuber appearance were performed, using eight clones with proven resistance to PVY and PVX from the breeding program of UFLA. The clones obtained were evaluated for agronomic traits, tuber appearance, and the presence of both Ryadg and Rx1 alleles, which confer extreme resistance to the PVY and PVX viruses, respectively. The independent culling level method was used to select genotypes of commercial interest, as well as to estimate the combining abilities of the parents. We identified clones carrying the Ryadg and Rx1 alleles with agronomic traits suitable for the fresh market and for processing. The BRS Ana cultivar and CMA-399 and CMA-385 clones showed positive effects on general combining ability (GCA) for tuber yield, while the Monalisa cultivar showed positive effects on GCA for general tuber appearance.

Ultrastructural insights into tomato infections caused by three different pathotypes of Pepino mosaic virus and immunolocalization of viral coat proteins.

This paper presents studies on an ultrastructural analysis of plant tissue infected with different pathotypes of Pepino mosaic virus (PepMV) and the immunolocalization of viral coat proteins. Because the PepMV virus replicates with a high mutation rate and exhibits significant genetic diversity, therefore, isolates of PepMV display a wide range of symptoms on infected plants. In this work, tomato plants of the Beta Lux cultivar were inoculated mechanically with three pathotypes representing the Chilean 2 (CH2) genotype: mild (PepMV-P22), necrotic (PepMV-P19) and yellowing (PepMV-P5-IY). The presence of viral particles in all infected plants in the different compartments of various cell types (i.e. spongy and palisade mesophyll, sieve elements and xylem vessels) was revealed via ultrastructural analyses. For the first time, it was possible to demonstrate the presence of crystalline inclusions, composed of virus-like particles. In the later stage of PepMV infection (14 dpi) various pathotype-dependent changes in the structure of the individual organelles (i.e. mitochondria, chloroplasts) were found. The strongest immunogold labeling of the viral coat proteins was also observed in plants infected by necrotic isolates. A large number of viral coat proteins were marked in the plant conductive elements, both xylem and phloem.

Complete genome sequence of an isolate of Potato virus X (PVX) infecting Cape gooseberry (Physalis peruviana) in Colombia.

Transcriptome analysis of a Cape gooseberry (Physalis peruviana) plant with leaf symptoms of a mild yellow mosaic typical of a viral disease revealed an infection with Potato virus X (PVX). The genome sequence of the PVX-Physalis isolate comprises 6435 nt and exhibits higher sequence similarity to members of the Eurasian group of PVX (~95 %) than to the American group (~77 %). Genome organization is similar to other PVX isolates with five open reading frames coding for proteins RdRp, TGBp1, TGBp2, TGBp3, and CP. 5' and 3' untranslated regions revealed all regulatory motifs typically found in PVX isolates. The PVX-Physalis genome is the only complete sequence available for a Potexvirus in Colombia and is a new addition to the restricted number of available sequences of PVX isolates infecting plant species different to potato.

Ecological and genetic determinants of Pepino Mosaic Virus emergence.

UNLABELLED: Virus emergence is a complex phenomenon, which generally involves spread to a new host from a wild host, followed by adaptation to the new host. Although viruses account for the largest fraction of emerging crop pathogens, knowledge about their emergence is incomplete. We address here the question of whether Pepino Mosaic Virus (PepMV) emergence as a major tomato pathogen worldwide could have involved spread from wild to cultivated plant species and host adaptation. For this, we surveyed natural populations of wild tomatoes in southern Peru for PepMV infection. PepMV incidence, genetic variation, population structure, and accumulation in various hosts were analyzed. PepMV incidence in wild tomatoes was high, and a strain not yet reported in domestic tomato was characterized. This strain had a wide host range within the Solanaceae, multiplying efficiently in most assayed Solanum species and being adapted to wild tomato hosts. Conversely, PepMV isolates from tomato crops showed evidence of adaptation to domestic tomato, possibly traded against adaptation to wild tomatoes. Phylogenetic reconstructions indicated that the most probable ancestral sequence came from a wild Solanum species. A high incidence of PepMV in wild tomato relatives would favor virus spread to crops and its efficient multiplication in different Solanum species, including tomato, allowing its establishment as an epidemic pathogen. Later, adaptation to tomato, traded off against adaptation to other Solanum species, would isolate tomato populations from those in other hosts. IMPORTANCE: Virus emergence is a complex phenomenon involving multiple ecological and genetic factors and is considered to involve three phases: virus encounter with the new host, virus adaptation to the new host, and changes in the epidemiological dynamics. We analyze here if this was the case in the recent emergence of Pepino Mosaic Virus (PepMV) in tomato crops worldwide. We characterized a new strain of PepMV infecting wild tomato populations in Peru. Comparison of this strain with PepMV isolates from tomato crops, plus phylogenetic reconstructions, supports a scenario in which PepMV would have spread to crops from wild tomato relatives, followed by adaptation to the new host and eventually leading to population isolation. Our data, which derive from the analysis of field isolates rather than from experimental evolution approaches, significantly contribute to understanding of plant virus emergence, which is necessary for its anticipation and prevention.

Identification of host proteins modulated by the virulence factor AC2 of Tomato chlorotic mottle virus in Nicotiana benthamiana.

Tomato, one of the most important crops cultivated worldwide, has been severely affected by begomoviruses such as the Tomato chlorotic mottle virus (ToCMoV). Virulence factor AC2 is considered crucial for a successful virus-plant interaction and is known to act as a transcriptional activator and in some begomoviruses to function as an RNA silencing suppressor factor. However, the exact functions of the AC2 protein of the begomovirus ToCMoV are not yet established. The aim of the present study was to identify differentially expressed proteins of the model plant Nicotiana benthamiana in response to the expression of the AC2 gene, isolated from ToCMoV. N. benthamiana plants were inoculated with Agrobacterium tumefaciens containing the viral vector Potato virus X (PVX) and with the PVX-AC2 construction. 2DE was performed and proteins were identified by MS. The results showed that the expression of ToCMoV AC2 alters the levels of several host proteins, which are important for normal plant development, causing an imbalance in cellular homeostasis. This study highlights the effect of AC2 in the modulation of plant defense processes by increasing the expression of several oxidative stress-related and pathogenesis-related proteins, as well as its role in modulating the proteome of the photosynthesis and energy production systems.

Deep sequencing of small RNAs in tomato for virus and viroid identification and strain differentiation.

Small RNAs (sRNA), including microRNAs (miRNA) and small interfering RNAs (siRNA), are produced abundantly in plants and animals and function in regulating gene expression or in defense against virus or viroid infection. Analysis of siRNA profiles upon virus infection in plant may allow for virus identification, strain differentiation, and de novo assembly of virus genomes. In the present study, four suspected virus-infected tomato samples collected in the U.S. and Mexico were used for sRNA library construction and deep sequencing. Each library generated between 5-7 million sRNA reads, of which more than 90% were from the tomato genome. Upon in-silico subtraction of the tomato sRNAs, the remaining highly enriched, virus-like siRNA pools were assembled with or without reference virus or viroid genomes. A complete genome was assembled for Potato spindle tuber viroid (PSTVd) using siRNA alone. In addition, a near complete virus genome (98%) also was assembled for Pepino mosaic virus (PepMV). A common mixed infection of two strains of PepMV (EU and US1), which shared 82% of genome nucleotide sequence identity, also could be differentially assembled into their respective genomes. Using de novo assembly, a novel potyvirus with less than 60% overall genome nucleotide sequence identity to other known viruses was discovered and its full genome sequence obtained. Taken together, these data suggest that the sRNA deep sequencing technology will likely become an efficient and powerful generic tool for virus identification in plants and animals.

Genetic structure of the aphid, Chaetosiphon fragaefolii, and its role as a vector of the Strawberry Yellow Edge Virus to a native strawberry, Fragaria chiloensis in Chile.

The monoecious anholocyclical aphid, Chaetosiphon fragaefolii (Cockerell) (Homoptera: Aphididae), was collected on a native strawberry, Fragaria chiloensis (L.) Duchesne (Rosales: Rosaceae) from different sites in Chile. The presence of this aphid was recorded during two consecutive years. F. chiloensis plants were collected from seven natural and cultivated growing areas in central and southern Chile. Aphids were genotyped by cross-species amplification of four microsatellite loci from other aphid species. In addition, the aphid borne virus Strawberry mild yellow edge virus was confirmed in F. chiloensis plants by double-antibody sandwich ELISA and RT-PCR. Genetic variability and structure of the aphid populations was assessed from the geo-referenced individuals through AMOVA and a Bayesian assignment test. The presence of C. fragaefolii, during the two-year study was detected in only four of the seven sites (Curepto, Contulmo, Chilián and Cucao). Genetic variation among these populations reached 19% of the total variance. When assigning the individuals to groups, these were separated in three genetic clusters geographically disjunct. Of the seven sampled sites, six were positive for the virus by RT-PCR, and five by double-antibody sandwich ELISA . The incidence of the virus ranged from 0-100%. Presence of the virus corresponded with the presence of the aphid in all but two sites (Chilian and Vilches). The greatest incidence of Strawberry mild yellow edge virus was related to the abundance of aphids. On the other hand, sequences of the coat protein gene of the different virus samples did not show correspondence with either the genetic groups of the aphids or the sampling sites. The genetic structure of aphids could suggest that dispersal is mainly through human activities, and the spread to natural areas has not yet occurred on a great scale.

Salicylic acid is involved in the Nb-mediated defense responses to Potato virus X in Solanum tuberosum.

To evaluate the role of salicylic acid (SA) in Nb-mediated hypersensitive resistance to Potato virus X (PVX) avirulent strain ROTH1 in Solanum tuberosum, we have constructed SA-deficient transgenic potato plant lines by overexpressing the bacterial enzyme salicylate hydroxylase (NahG), which degrades SA. Evaluation of these transgenic lines revealed hydrogen peroxide accumulation and spontaneous lesion formation in an age- and light-dependent manner. In concordance, NahG potato plants were more sensitive to treatment with methyl viologen, a reactive oxygen species-generating compound. In addition, when challenged with PVX ROTH1, NahG transgenic lines showed a decreased disease-resistance response to infection and were unable to induce systemic acquired resistance. However, the avirulent viral effector, the PVX 25-kDa protein, does induce expression of the pathogenesis-related gene PR-1a in NahG potato plants. Taken together, our data indicate that SA is involved in local and systemic defense responses mediated by the Nb gene in Solanum tuberosum. This is the first report to show that basal levels of SA correlate with hypersensitive resistance to PVX.