Inoculation Of Cotyledons Research Articles

Inter- and Intra-Pathogen Interactions Emanating from Coinfection with Different Fungal and Viral strains in Canola Cultivars with Differing Host Resistances.

Few recent investigations examine coinfection interactions between fungal and viral plant pathogens. Here, we investigated coinfections between Leptosphaeria maculans and turnip mosaic virus (TuMV) in canola (Brassica napus). Different combinations of L. maculans isolate P11 and resistance breaking isolates L. maculans UWA192 and TuMV 12.1, were inoculated to three cultivars with differing pathogen resistances/susceptibilities. They were inoculated first to entire or half cotyledons 10-12 days after emergence, and second to opposite entire or half cotyledons on the same day (day 0) or 3 or 7 days afterwards. The parameters measured were L. maculans cotyledon disease index (%CDI), and TuMV systemically infected leaf symptom intensity (SI) and virus concentration (VC). Except when both day 0 inoculations were with isolate UWA192, %CDI values were supressed strongly or only weakly when isolates P11 and/or UWA192 were inoculated to plants with L. maculans single gene resistance (SGR) or polygenic resistance, respectively. However, except when isolate P11 was inoculated first and UWA192 second, these values declined after inoculation day 0 when SGR was absent. TuMV infection suppressed %CDI values, although this decrease was usually smaller following day 0 half cotyledon inoculations. When TuMV temperature sensitive extreme resistance was present and both inoculations were with TuMV, SI and VC values diminished greatly. However, the extent of this decrease was reduced when second inoculations were with L. maculans. SI and VC values were also smaller when SGR was present and second inoculations were with L. maculans. When L. maculans resistance was lacking, SI and VC values were smaller when second inoculations to entire cotyledons were with L. maculans rather than TuMV. This also occurred after second half cotyledon inoculations with isolate P11 but not isolate UWA192. Therefore, diverse inter- or intra-pathogen interactions developed depending upon host resistance, isolate combination, cotyledon inoculation approach and second inoculation timing.

Arabinogalactan Protein-Like Proteins From Ulva lactuca Activate Immune Responses and Plant Resistance in an Oilseed Crop.

Natural compounds isolated from macroalgae are promising, ecofriendly, and multifunctional bioinoculants, which have been tested and used in agriculture. Ulvans, for instance, one of the major polysaccharides present in Ulva spp. cell walls, have been tested for their plant growth-promoting properties as well as their ability to activate plant immune defense, on a large variety of crops. Recently, we have characterized for the first time an arabinogalactan protein-like (AGP-like) from Ulva lactuca, which exhibits several features associated to land plant AGPs. In land plant, AGPs were shown to play a role in several plant biological functions, including cell morphogenesis, reproduction, and plant-microbe interactions. Thus, isolated AGP-like proteins may be good candidates for either the plant growth-promoting properties or the activation of plant immune defense. Here, we have isolated an AGP-like enriched fraction from Ulva lactuca and we have evaluated its ability to (i) protect oilseed rape (Brassica napus) cotyledons against Leptosphaeria maculans, and (ii) its ability to activate immune responses. Preventive application of the Ulva AGP-like enriched fraction on oilseed rape, followed by cotyledon inoculation with the fungal hemibiotroph L. maculans, resulted in a major reduction of infection propagation. The noticed reduction correlated with an accumulation of H2O2 in treated cotyledons and with the activation of SA and ET signaling pathways in oilseed rape cotyledons. In parallel, an ulvan was also isolated from Ulva lactuca. Preventive application of ulvan also enhanced plant resistance against L. maculans. Surprisingly, reduction of infection severity was only observed at high concentration of ulvan. Here, no such significant changes in gene expression and H2O2 production were observed. Together, this study indicates that U. lactuca AGP-like glycoproteins exhibit promising elicitor activity and that plant eliciting properties of Ulva extract, might result not only from an ulvan-originated eliciting activities, but also AGP-like originated.

Developing a Greenhouse Protocol for Evaluating Resistance to Corynespora cassiicola in Cotton (Gossypium hirsutum).

Target spot, caused by Corynespora cassiicola, has reemerged as a disease of economic importance in cotton (Gossypium hirsutum L.) in the southeastern United States. Although this pathogen affects other economically important crops, relatively little is known about C. cassiicola on cotton, especially with regard to conditions conducive for disease development and sources of genetic resistance. Therefore, to more efficiently screen cotton genotypes for resistance, a greenhouse protocol is needed. Optimum temperature and conducive leaf wetness duration were determined in growth chamber trials. Temperature range for disease onset and greatest lesion counts occurred between 20 and 28°C. Generally, with ≥24 h of leaf wetness at these temperatures, disease onset was noted as rapidly as 1 day after inoculation on a cultivar that was previously determined to be susceptible to target spot. A mist irrigation system was used to maintain prolonged periods of leaf wetness in the greenhouse. In greenhouse trials, inoculation of cotyledons with 4 × 104 conidia/ml allowed differentiation of five selected genotypes with disease reactions that reflected their field rankings. The current protocol will be useful for evaluating cotton breeding lines for resistance to target spot.

A gene-for-gene interaction involving a 'late' effector contributes to quantitative resistance to the stem canker disease in Brassica napus.

Summary The control of stem canker disease of Brassica napus (rapeseed), caused by the fungus Leptosphaeria maculans is based largely on plant genetic resistance: single‐gene specific resistance (Rlm genes) or quantitative, polygenic, adult‐stage resistance. Our working hypothesis was that quantitative resistance partly obeys the gene‐for‐gene model, with resistance genes ‘recognizing’ fungal effectors expressed during late systemic colonization.Five LmSTEE (stem‐expressed effector) genes were selected and placed under the control of the AvrLm4‐7 promoter, an effector gene highly expressed at the cotyledon stage of infection, for miniaturized cotyledon inoculation test screening of a gene pool of 204 rapeseed genotypes.We identified a rapeseed genotype, ‘Yudal’, expressing hypersensitive response to LmSTEE98. The LmSTEE98–RlmSTEE98 interaction was further validated by inactivation of the LmSTEE98 gene with a CRISPR‐Cas9 approach. Isolates with mutated versions of LmSTEE98 induced more severe stem symptoms than the wild‐type isolate in ‘Yudal’. This single‐gene resistance was mapped in a 0.6 cM interval of the ‘Darmor_bzh’ × ‘Yudal’ genetic map.One typical gene‐for‐gene interaction contributes partly to quantitative resistance when L. maculans colonizes the stems of rapeseed. With numerous other effectors specific to stem colonization, our study provides a new route for resistance gene discovery, elucidation of quantitative resistance mechanisms and selection for durable resistance.

Rapid detection of Leptosphaeria maculans avirulence gene AvrLm4-7 conferring the avirulence/virulence specificity on Brassica napus using a tetra-primer ARMS-PCR

Leptosphaeria maculans is the causal agent of blackleg disease in canola (Brassica napus), resulting in significant yield loss in canola fields worldwide. AvrLm4-7 is an avirulence effector gene in L. maculans, and a single nucleotide mutation at codon 358 is responsible for the absence of the AvrLm4 allele. A tetra-primer amplification refractory mutation system-PCR assay (ARMS-PCR) was developed to rapidly differentiate the AvrLm4AvrLm7 and avrLm4AvrLm7 genes of L. maculans isolates, which differ by a single point mutation. By this approach, we were able to amplify distinct PCR products to infer the gene of the tested isolates. These results were also confirmed through phenotyping, using the cotyledon inoculation test and two canola genotypes with the corresponding resistance genes. The tetra-primer ARMS-PCR assay developed in this study is a simple, rapid, and useful protocol to identify the AvrLm4-7 alleles in L. maculans isolates. This assay has potential applications in the selection of resistant canola cultivars as part of broader antifungal strategies.

Pathogenic Variability within Indian Alternaria brassicae Isolates Using Seed, Cotyledon and Leaf of Brassica Species

AbstractThirty Alternaria brassicae (Berk.) Sacc. isolates from diverse geographical locations of India were studied for pathogenic variability on seed, cotyledon and true leaves of Brassica species. Seed germination was reduced maximum by isolate BAB‐39 in Brassica juncea cultivar Varuna (22.1%), Brassica rapa var. Toria cultivar PT‐303 (12%), Brassica carinata cultivar Kiran (12%), Brassica napus cultivar GSL‐1 (11%) and tolerant source of B. juncea genotype PHR‐2 (7%), although least by isolate BAB‐49. Maximum lesion size on leaf was recorded in B. juncea cultivar Rohini (11.2, 16.5 and 16.8 mm) with isolates BAB‐09 (Pantnagar), BAB‐19 (Bharatpur) and BAB‐39 (Kangra), respectively, and categorized as highly virulent, while minimum lesion size of 3.2, 3.7 and 3.8 mm was observed with isolates BAB‐47 (Tonk), BAB 49 (Jobner) and BAB 04 (Kamroop), respectively, considered the weak isolates. On B. alba, BAB‐09, BAB‐19 and BAB‐39 isolates caused maximum lesion size of 3.7, 3.8 and 3.9 mm, respectively, even though it showed maximum tolerance. In both seed and cotyledon inoculation method, the per cent Alternaria blight severity above 80% was observed with isolate of BAB‐39 (91.5%), BAB‐19 (89.0%), BAB‐09 (85.5%) and least in isolate BAB‐49 (34.0%). Brassica seed, cotyledon and leaf showed the similar positive response for categorizing A. brassicae isolates as virulent and avirulent. This information could be used to the development and assessment of resistant brassica germplasm, especially with A. brassicae populations exhibiting increased virulence.

RAPID SCREENING TECHNIQUE FOR ALTERNARIA BLIGHT RESISTANCE IN INDIAN MUSTARD (Brassica juncea L.) USING COTYLEDONARY LEAF METHOD

Alternaria blight (AB) caused by Alternaria brassicae (Berk.) Sacc. is a devastating disease of oilseed Brassicas all over the world, and responsible for significant seed yield losses up to 47%. No reliable, resistant germplasm is available to develop AB resistant cultivars. Various screening techniques have been reported so far, but cotyledonary leaf method is not yet reported. Three methods were tested using one susceptible cultivar (Varuna): inoculation of seed, inoculation of cotyledons, and inoculation of both seed and cotyledons. Fungal conidia were inoculated directly onto the seedlings with 1.5x105, 2.5x105, 4x105 and 5x105 conidia ml-1 concentrations for standardization. Percentage AB severity increased with the increase in conidial concentration, therefore the highest concentration was used for final screening. Among the three screening methods, inoculation of both seed, and cotyledon method was found highly effective where mean AB severity on cotyledon was 84.6% in comparison to 49.3% in the inoculation of seed and 62.5% in the inoculation of cotyledon methods. The technique was validated by screening susceptible and putative tolerant genotypes. The severity of AB was 54% of susceptible cultivar and 16.4%-21.2% of tolerant genotypes. The conidia number per microscopic field was 21.5 in putative tolerant and 43.5 in susceptible genotypes. Thus, in vitro screening of AB using inoculation of both seed and cotyledon method was found most effective and could be used for rapid screening in early stages of plant growth. A new 0-7 rating scale was also devised to observe the AB pathogen interaction phenotype at the cotyledonary stage of oilseed Brassica.

Breakdown of Rlm3 resistance in the Brassica napus–Leptosphaeria maculans pathosystem in western Canada

Blackleg disease, caused by the fungal pathogen Leptosphaeria maculans, is a serious disease of Brassica napus. The disease is mainly controlled by genetic resistance and crop rotation. However, L. maculans has displayed a high evolutionary potential to overcome major resistance genes in B. napus. This study aimed to analyze the major-gene and adult-plant resistance (APR) of Canadian B. napus varieties/lines (accessions) and the avirulence allele frequency in L. maculans populations in western Canada. For resistance identification, a set of L. maculans isolates with known avirulence genes were used to characterize major resistance (R) genes in 104 Canadian B. napus accessions and 102 seed samples collected from growers’ fields; with 104 B. napus accessions further evaluated for APR under controlled conditions. In addition, avirulence genes of 300 L. maculans isolates collected from infected canola stubbles in growers’ fields were determined by cotyledon inoculation and gene-specific PCR assays. The results indicated that R genes were present in the majority of these B. napus accessions, with the Rlm3 gene being predominant while other R genes were rarely detected. APR was identified in more than 50 % of the accessions. Predominance of Rlm3 in 102 seed samples from growers’ fields suggested Rlm3-carrying B. napus varieties were currently widely used in western Canada. Avirulence allele frequency identification of field L. maculans isolates revealed the scarcity of the avirulence allele towards Rlm3, AvrLm3. This indicated the breakdown of Rlm3 resistance, which could be due to the over use of this single resistance gene in Canadian B. napus germplasm.

Pathotypic diversity of Hyaloperonospora brassicae collected from Brassica oleracea

Downy mildew caused by Hyaloperonospora brassicae is an economically destructive disease of brassica crops in many growing regions throughout the world. Specialised pathogenicity of downy mildews from different Brassica species and closely related ornamental or wild relatives has been described from host range studies. Pathotypic variation amongst Hyaloperonospora brassicae isolates from Brassica oleracea has also been described; however, a standard set of B. oleracea lines that could enable reproducible classification of H. brassicae pathotypes was poorly developed. For this purpose, we examined the use of eight genetically refined host lines derived from our previous collaborative work on downy mildew resistance as a differential set to characterise pathotypes in the European population of H. brassicae. Interaction phenotypes for each combination of isolate and host line were assessed following drop inoculation of cotyledons and a spectrum of seven phenotypes was observed based on the level of sporulation on cotyledons and visible host responses. Two host lines were resistant or moderately resistant to the entire collection of isolates, and another was universally susceptible. Five lines showed differential responses to the H. brassicae isolates. A minimum of six pathotypes and five major effect resistance genes are proposed to explain all of the observed interaction phenotypes. The B. oleracea lines from this study can be useful for monitoring pathotype frequencies in H. brassicae populations in the same or other vegetable growing regions, and to assess the potential durability of disease control from different combinations of the predicted downy mildew resistance genes.

Identification and mapping of a novel blackleg resistance locus LepR4 in the progenies from Brassica napus × B. rapa subsp. sylvestris

Blackleg, caused by Leptosphaeria maculans, is one of the most economically important diseases of Brassica napus worldwide. Two blackleg-resistant lines, 16S and 61446, were developed through interspecific hybridization between B. napus and B. rapa subsp. sylvestris and backcrossing to B. napus. Classical genetic analysis demonstrated that a single recessive gene in both lines conferred resistance to L. maculans and that the resistance alleles were allelic. Using BC(1) progeny derived from each resistant plant, this locus was mapped to B. napus linkage group N6 and was flanked by microsatellite markers sN2189b and sORH72a in an interval of about 10 cM, in a region equivalent to about 6 Mb of B. rapa DNA sequence. This new resistance gene locus was designated as LepR4. The two lines were evaluated for resistance to a wide range of L. maculans isolates using cotyledon inoculation tests under controlled environment conditions, and for stem canker resistance in blackleg field nurseries. Results indicated that line 16S, carrying LepR4a, was highly resistant to all isolates tested on cotyledons and had a high level of stem canker resistance under field conditions. Line 61446, carrying LepR4b, was only resistant to some of the isolates tested on cotyledons and was weakly resistant to stem canker under field conditions.

Evaluation of greenhouse inoculation techniques used to screen for Sclerotinia stem rot resistance in soybeans

Numerous inoculation methods have been used to screen soybean germplasm for resistance to Sclerotinia stem rot, caused by Sclerotinia sclerotiorum. This study was conducted to compare six inoculation methods viz. (i) spray mycelium, (ii) drop mycelium, (iii) cut stem, (iv) cotyledon inoculation, (v) straw inoculation and (vi) petiole. Four soybean cultivars were planted in 1–l pots in the greenhouse and grown to V3 (third trifoliate) growth stage. Overseeded pots were thinned to four plants which were inoculated using the respective methods and covered for nine days with transparent plastic bags. Plants were arranged in a randomized block design with eight replicates. The number of infected plants was counted and leaf lesion development and degree of plant wilting were scored using a 0 to 5 rating scale. The spray mycelium method yielded the highest incidence of wilting although a significant cultivar × inoculation technique (P<0.05) was recorded, particularly where host tissues were damaged prior to inoculation. Results suggest that cultivar responses are affected by the degree and area of tissue damage associated with the respective inoculation methods.

High Temperatures Activate Local Viral Multiplication and Cell-to-Cell Movement ofMelon necrotic spot virusbut Restrict Expression of Systemic Symptoms

The infection of melon plants by Melon necrotic spot virus (MNSV) and the development of necrotic disease symptoms are a seasonal occurrence in Japan, which take place between winter and early summer, but not during mid-summer. In this paper we investigate the effect of three different temperatures (15, 20, and 25 degrees C) on the local and systemic expression of MNSV in melon plants. Previously, the incidence of plants expressing systemic symptoms caused by MNSV and other viruses was found to be greater at temperatures less than 20 degrees C. In this study, our temperature-shift experiments support previous studies that found the expression of systemic symptoms increases as temperature falls from 25 to 20 degrees C and decreases as temperature rises from 20 to 25 degrees C. However, MNSV replication in melon cells and local viral movement within leaves following the inoculation of melon protoplasts or cotyledons were more frequent at 25 degrees C than at 15 or 20 degrees C.

Cucumber vein yellowing ipomovirus

<i>Cucumber vein yellowing ipomovirus</i>

Acquisition of an Agrobacterium Ri Plasmid and Pathogenicity by Other α- Proteobacteria in Cucumber and Tomato Crops Affected by Root Mat

Root mat of cucumbers and tomatoes has previously been shown to be caused by Agrobacterium radiobacter strains harboring a root-inducing Ri plasmid (pRi). Nine other pRi-harboring alpha-Proteobacteria have subsequently been isolated from root mat-infected crops. Fatty acid profiling and partial 16S rRNA sequence analysis identified three of these strains as being in the genus Ochrobactrum, five as being in the genus Rhizobium, and one as being in the genus Sinorhizobium: An in vitro pathogenicity test involving inoculation of cucumber cotyledons was developed. All pRi-harboring alpha-Proteobacteria induced typical root mat symptoms from the cotyledons. Average transformation rates for rhizogenic Ochrobactrum (46%) and Rhizobium (44%) strains were lower than those observed for rhizogenic A. radiobacter strains (64%). However, individual strains from these three genera all had transformation rates comparable to those observed from cotyledons inoculated with a rhizogenic Sinorhizobium strain (75%).

Identification and Cultivar Reaction to Three New Races of the Spinach Downy Mildew Pathogen from the United States and Europe.

Since 1996, commercial spinach cultivars with resistance to four previously described races of Peronospora farinosa f. sp. spinaciae (races 1, 2, 3, and 4) were observed with high incidences of downy mildew both in California and Europe. Isolates of P. farinosa f. sp. spinaciae collected in California between 1997 and 2001, Arizona in 1999, and a single isolate collected in the Netherlands in 1996 were examined for their disease reaction on differential spinach cultivars and a set of commercial spinach cultivars. Disease reactions on the differential cultivars indicated the occurrence of three new races of P. farinosa f. sp. spinaciae. Two newly identified races, designated race 5 (isolate CA1) and race 6 (isolate SP1), were detected in the United States. The isolate from the Netherlands also was distinct and designated race 7 (isolate JVN7). Some cultivars with resistance to races 1, 2, 3, and 4 were susceptible to race 5, whereas others were resistant, indicating that resistance to a given race may be governed by different genes (or alleles) depending on the source of resistance. A survey of races in California indicated that races 5 and 6 predominated. Although the majority of the cultivars examined were susceptible to race 6 based on the traditional qualitative cotyledon inoculation assay, significant quantitative differences in resistance to race 6 were observed using a true-leaf greenhouse screening procedure. Although more work is needed to confirm the results of the true-leaf assays, the quantitative resistance observed using this procedure appears to be race specific.

Inheritance of resistance to Leptosphaeria maculans in two accessions of Brassica napus

Inheritance of resistance to Leptosphaeria maculans was studied in Brassica napus accessions RB87-62 and DH88-752. The two resistant accessions and a susceptible B. napus cultivar, 'Westar', were crossed in a complete diallel, and their progenies were evaluated for disease reaction by cotyledon and stem inoculation with L. maculans, isolate Pl86-12, belonging to pathogenicity group 2 (PG2). For crosses between 'Westar' and either resistant parent, segregation of F2 populations fits a ratio of 3:1 (resistant to susceptible) both for cotyledon and adult-plant reactions, indicating a single-dominant-gene model. Analysis of the relationship between resistance of the cotyledons and that of adult plants showed that genes controlling these two traits were different but linked in RB87-62 and DH88-752. This was confirmed by the segregation data from F3 families. Segregation in the cross between the two resistant accessions demonstrated that resistances of the cotyledons and adult plants from the two different sources were nonallelic and independently assorting.

Localization of Zucchini yellow mosaic virus to the veinal regions and role of viral coat protein in veinal chlorosis conditioned by the zym potyvirus resistance locus in cucumber

The zym locus in cucumber (Cucumis sativus L.) is marked by multiple alleles conferring resistance to the potyvirus, Zucchini yellow mosaic virus (ZYMV). Cotyledon inoculation of greenhouse grown plants expressing the zymDina allele results in a distinct pattern of veinal chlorosis limited to a single systemic leaf; leaf-inoculated plants remain free of symptoms. Inoculation of cotyledons of plants of different growth stages indicated that the virus moved to the leaf that was newly emerging at the time of cotyledon infection where it then remained localized to the veinal regions for up to 30 days post-inoculation (d.p.i). Cotyledon removal experiments indicated that the inability of ZYMV to spread within the uninoculated leaf, or to infect inoculated leaves, was not due to the inability to replicate in leaves. Chimeric viruses generated from a strain of ZYMV that did not induce the veinal chlorosis response, indicated that the coat protein amino terminus, which has been shown to be involved in long distance movement, affected occurrence of veinal chlorosis. In the growth chamber, inoculation of the first leaf, but not subsequent leaves, gave a high percentage of plants expressing veinal chlorosis. Together, these observations suggest that resistance conferred by zymDina is developmentally regulated and occurs at the level of systemic movement.

A Monitoring System for Green Fluorescence Protein Gene-transformed Fusarium oxysporum in Melon Seedlings

Using melon seedlings at the cotyledon stage and genetically marked fungi, a system for monitoring pathogenic and nonpathogenic Fusarium oxysporum was devised in the present study. Protoplasts were prepared from three formae speciales (melonis, radicis-lycopersici and fragariae )of F. oxysporum and transformed with a synthetic gene for green fluorescence protein. Transformants were primarily isolated in the presence of hygromycin B and then screened by the emission of bright green fluorescence. Roots of melon seedlings were inoculated with fluorescing microconidia of these fungi, and fungal infection behavior was traced. Using fluorescence microscopy, we directly observed not only the fungus at the root surface, but also the mycelia elongating in the trachea of roots. Both pathogenic and nonpathogenic fungi germinated and hyphae elongated superficially on the surface of root. Only pathogenic fungi caused root necrosis at the inoculation site. Hyphae grew within the stem to induce constriction or cracking of lower hypocotyls, then causing wilting of the seedlings. Infection behavior of genetically marked pathogenic and nonpathogenic F. oxysporum could be successfully monitored after inoculation of cotyledons of seedlings.

Induced Resistance in Hypocotyl of Cucumber by Infection with Colletotrichum lagenarium in Leaves

Localized infection of cucumber leaves with Colletotrichum lagenarium induced resistance against infection after challenge inoculation with pathogenic fungi, including C. lagenarium and Pythium ultimum var. ultimum. Systemic acquired resistance in the hypocotyl was observed when challenge inoculation was made 4 to 7 days after the first inoculation of cotyledons. Seven days after the first inoculation of a primary leaf, induced resistance against the challenge inoculation in the hypocotyl was also observed. However, the hypocotyl did not develop induced resistance when plants were challenged by a wound inoculation with P. ultimum.

Induction of Cell Death in Transgenic Plants Expressing a Fungal Glucose Oxidase

Hydrogen peroxide (H2O2) has been implicated in the induction of plant defense genes and programmed cell death. Expression of a chimeric fungal glucose oxidase (GO) gene driven by a pathogen- and wound-inducible promoter was evaluated in transgenic tobacco and canola as a possible tool for engineering plant cell death and defense gene induction. Expression of this gene under the control of a peroxidase gene promoter resulted in the accumulation of relatively low levels of H2O2 in the young leaves of transgenic tobacco plants and this was not sufficient to cause any visible cell death and defense gene induction as measured by PR-1a mRNA induction. Older leaves of transgenic tobacco plants, however, exhibited visible necrotic lesions and constitutively expressed PR-1a mRNA when grown under high light conditions. Inoculation of cotyledons of control and transgenic canola with Leptosphaeria maculans resulted in rapid cotyledon senescence in the transgenic plants. Strong activators of the peroxidase promoter, i.e., wounding and inoculation of transgenic plants with Cercospora nicotianae, were not sufficient to trigger any additional visible cell death in transgenic tobacco plants, compared with controls. However, when exogenous glucose was supplied to transgenic tissue, massive cell death and PR-1a gene induction were observed in tobacco. Exogenously applied salicylic acid further increased the rate and extent of cell death. Our results suggest that efficacy of GO expression for the induction of cell death is restricted by glucose supply in the plants and are consistent with a role for salicylic acid in the potentiation of plant cell death by H2O2.