A Graft Mimic Strategy for Verticillium Resistance in Tomato

Grafting vegetables for disease resistance has increased greatly in popularity over the past 10 years. Verticillium wilt of tomato is commonly controlled through grafting of commercial varieties on resistant rootstocks expressing the Ve1 R-gene. To mimic the grafted plant, proteomic analyses in tomato were used to identify a suitable root-specific promoter (TMVi), which was used to express the Ve1-allele in susceptible Craigella (Cs) tomato plants. The results indicate that when infected with Verticillim dahliae, race 1, the transformed plants are comparable to resistant cultivars (Cr) or grafted plants.     

Artificial elevation of glutathione contents in salicylic acid‐deficient tobacco (Nicotiana tabacum cv. Xanthi NahG) reduces susceptibility to the powdery mildew pathogen Euoidium longipes

• The effects of elevated glutathione levels on defence responses to powdery mildew (Euoidium longipes) were investigated in a salicylic acid‐deficient tobacco (Nicotiana tabacum cv. Xanthi NahG) and wild‐type cv. Xanthi plants, where salicylic acid (SA) contents are normal.
• Aqueous solutions of reduced glutathione (GSH) and its synthetic precursor R‐2‐oxothiazolidine‐4‐carboxylic acid (OTC) were injected into leaves of tobacco plants 3 h before powdery mildew inoculation.
• SA‐deficient NahG tobacco was hyper‐susceptible to E. longipes, as judged by significantly more severe powdery mildew symptoms and enhanced pathogen accumulation. Strikingly, elevation of GSH levels in SA‐deficient NahG tobacco restored susceptibility to E. longipes to the extent seen in wild‐type plants (i.e. enhanced basal resistance). However, expression of the SA‐mediated pathogenesis‐related gene (NtPR‐1a) did not increase significantly in GSH or OTC‐pretreated and powdery mildew‐inoculated NahG tobacco, suggesting that the induction of this PR gene may not be directly involved in the defence responses induced by GSH.
• Our results demonstrate that artificial elevation of glutathione content can significantly reduce susceptibility to powdery mildew in SA‐deficient tobacco.

     

The oxalyl‐CoA synthetase‐regulated oxalate and its distinct effects on resistance to bacterial blight and aluminium toxicity in rice

• Oxalic acid is widely distributed in biological systems and known to play functional roles in plants. The gene AAE3 was recently identified to encode an oxalyl‐CoA synthetase (OCS) in Arabidopsis that catalyses the conversion of oxalate and CoA into oxalyl‐CoA. It will be particularly important to characterise the homologous gene in rice since rice is not only a monocotyledonous model plant, but also a staple food crop.
• Various enzymatic and biological methods have been used to characterise the homologous gene.
• We first defined that AAE3 in the rice genome (OsAAE3) also encodes an OCS enzyme. Its K_m for oxalate is 1.73 ± 0.12 mm , and V_m is 6824.9 ± 410.29 U·min^?1·mg protein^?1. Chemical modification and site‐directed mutagenesis analyses identified thiols as the active site residues for rice OCS catalysis, suggesting that the enzyme might be regulated by redox state. Subcellular localisation assay showed that the enzyme is located in the cytosol and predominantly distributed in leaf epidermal cells. As expected, oxalate levels increased when OCS was suppressed in RNAi transgenic plants. More interestingly, OCS‐suppressed plants were more susceptible to bacterial blight but more resistant to Al toxicity.
• The results demonstrate that the OsAAE3‐encoded protein also acts as an OCS in rice, and may play different roles in coping with stresses. These molecular, enzymatic and functional data provide first‐hand information to further clarify the function and mechanism of OCS in rice plants.

     

Further Investigation of the Specificity of Interactions Between Wild Lactuca spp. and Bremia lactucae Isolates from Lactuca serriola

Callus cultures derived from isogenic lines of the tomato cultivars Moneymaker and Craigella, resistant or susceptible to F. oxysporum f. sp. lycopersici, were inoculated with Fusarium oxysporum f. sp. lycopersici race 1. Fungal growth was restricted on callus derived from resistant plants, after inoculation with a conidial suspension, whereas callus derived from susceptible plants was totally overgrown by the fungus within 7 days. The concentration of the phytoalexin rishitin was significantly higher in the callus culture derived from a resistant tomato line compared with the callus culture from a susceptible line, 2 and 3 days after inoculation with mycelium. The results of the experiments were compared with experiments with whole plants. Rishitin production as well as growth of the fungus was comparable with responses in plant-fungus interaction. Therefore callus culture may be useful in studying the interaction between tomato plants and race 1 of F. oxysporum f. sp. lycopersici.     

Evaluation of Tomato Hybrids Carrying Ty‐1 and Ty‐2 Loci to Japanese Monopartite Begomovirus Species

Tobacco leaf curl Japan virus, Honeysuckle yellow vein mosaic virus and Tomato yellow leaf curl virus are three begomoviruses that infect tomato crops in Japan. Tomato infection by begomoviruses has increased in Japan after the development of a high level of resistance to certain insecticides in some populations of the vector B. tabaci biotypes ‘B and Q’. Ty‐1 and Ty‐2 homozygous tomato hybrids were evaluated for reaction to monopartite begomovirus species in Japan by Agrobacterium‐mediated inoculation. Test plants were evaluated by a disease assessment scale (DAS), varying from 1 = no symptoms to 4 = severe symptoms, and systemic infection was evaluated by polymerase chain reaction (PCR), using specific begomovirus primers for each virus. Ty‐1 hybrids showed tolerance to HYVMV and with a large number of plants being neither virus‐free nor symptom‐free. The response of Ty‐1 hybrids was also resistant to moderately resistant against TbLCJV. The response of Ty‐2 hybrids was resistant to highly resistant against the three monopartite begomoviruses, when compared with susceptible plants.     

Stripe Rust Resistance in Roegneria kamoji (Poaceae: Triticeae) and its Genetic Analysis

The Roegneria kamoji accession ZY 1007 was resistant to the mixed predominant races of Puccinia striiformis f.sp. tritici (Pst) in China based on field tests at adult‐plant stage. The seedling resistance evaluation of ZY 1007 showed that it was resistant to stripe rust physiological strains CYR29, CYR33 and PST‐V26, which were the predominant races of Pst in China. The female parent R. kamoji cv. Gansi No.1 (susceptible to Pst) was crossed with ZY 1007 (resistant to Pst). Parents, F₁ and F₂ populations were tested in a field inoculated with the mixed urediniospores. ZY 1007 and all the observed 11 F₁ hybrid plants were resistant, while plants of Gansi No.1 were susceptible. Among the 221 F₂ plants, 168 plants were resistant and 53 were susceptible, and the segregation of resistant and susceptible plants fits 3R:1S ratio (χ² = 0.074, P > 0.75). It confirmed that the resistance of stripe rust in ZY 1007 was controlled by a single dominant gene and temporarily designated as YrK1007.     

Behavior and biology of the tomato psyllid, Bactericerca cockerelli, in response to the Mi-1.2 gene

The Mi‐1.2 gene, identified from wild varieties of tomato, Solanum peruvianum (Mill) (Solanaceae), has been incorporated into near‐isogenic commercial varieties of tomato and has been shown to confer resistance to three different species of phloem feeders: aphids, whiteflies, and nematodes. The results presented here show that plants bearing Mi‐1.2 were also resistant to the tomato psyllid, Bactericerca [Paratrioza] cockerelli (Sulc) (Homoptera: Psyllidae), a serious pest of tomato, Solanum lycopersicon (Mill), in the western half of North America. In choice studies, tomato psyllids preferred to settle on plants that did not contain the gene [Moneymaker (mi‐1.2)] compared to near‐isogenic plants with the gene [Motelle (Mi‐1.2)]. As a result, total oviposition was higher on the susceptible variety, although no‐choice studies indicated that there were no differences in numbers of eggs laid by individual females on either variety. Survival from egg to adult was higher on plants lacking the gene compared to plants containing the gene. However, there were no differences in total development time of individuals reared from either variety. The results suggest that mechanisms of resistance to the tomato psyllid observed in plants bearing the Mi‐1.2 gene are distinct from the mechanisms of resistance to the three phloem feeders examined in other studies.     

The effects of rootstock and scion on tobacco mosaic virus infection in susceptible, tolerant and immune cultivars of tomato

Reciprocal grafts were made between tomato cultivars Potentate, susceptible, and Virocross, tolerant (heterozygous for resistance gene Tm-i) to tobacco mosaic virus (TMV) isolates of Pelham type o and between isogenic lines of cv. Craigella, susceptible and homozygous for gene Tm-i. The grafted plants were inoculated with a type o isolate; both scion and stock inoculation were studied in the former, scion inoculation only, in the latter. With scion inoculation the virus content of a tolerant scion was greater on a susceptible stock than on a tolerant one, but that of a susceptible scion was unaffected by the type of stock: in contrast, the virus content of a tolerant stock was unaffected by the type of scion but that of a susceptible stock was less with a tolerant than with a susceptible scion. With root inoculation the virus contents of both tolerant and susceptible scions were greater on a susceptible than on a tolerant stock. With cv. Craigella the genotype Tm-1/Tm-1 was found to be immune to the type o isolate used, but in grafts the leaves of Tm-1/Tm-1 scions became tolerant to leaf inoculation when on susceptible stocks and the virus entered the stock. Tm-1/Tm-1 stocks became infected when attached to infected, susceptible scions and did not affect the virus content of those scions. The results indicate that a susceptible healthy stock may change the reaction of a tolerant or immune scion to infection by a strain of TMV.     

The Occurrence of 1,3-{beta}-Glucanase in Healthy and Verticillium-infected, Resistant and Susceptible Tomato Plants

Leaf, stem, and root extracts of near-isogenic tomato plantscv. Craigella, resistant and susceptible to Verticillium albo-atrum,showed constitutive 1,3-ß-glucanase activity whichincreased following inoculation with the pathogen. Partiallypurified enzyme extracts were obtained by dialysing a 30–80%ammonium sulphate fraction of the tissue brei. The enzyme hadpH and temperature optima of 5?5 and 44 ?C respectively, withhigh activity between 50 and 60 ?C. The response to laminarinconcentration was linear between 1?2 and 7?5 mg ml^–1.Root inoculation of susceptible plants with 10⁶ propagules ml^–1V. albo-atrum led to a umform 300 per cent increase in all steminternodes except the terminal one, which was 500 per cent ofthe controls. No spatial relationship of enzyme activity tothe localization of fungus within the stem was apparent. Petioles,leaves, and roots of susceptible infected plants similarly showedan increase in activity but less than that in stems. Changedlevels of stern enzyme activity at different times after inoculationwere associated with reductions in the number of vessels containinghyphae. Extracts of plants of the resistant isoline showed increasedglucanase activity over controls, but this was substantiallylower than that in susceptible plants and was associated withthe greatly reduced mycelial colonization in resistant plants. It is concluded that single gene resistance in tomato to Verticilliumis not associated with innately higher levels of 1,3-ß-glucanasein healthy plants. The increased activity in infected plantsis proportional to the overall quantity of pathogen in the plantor of pathogenic metabolites.     

Do native and invasive herbivores have an effect on Brassica rapa pollination?

• Mutualistic (e.g. pollination) and antagonistic (e.g. herbivory) plant–insect interactions shape levels of plant fitness and can have interactive effects.
• By using experimental plots of Brassica rapa plants infested with generalist (Mamestra brassicae) and specialised (Pieris brassicae) native herbivores and with a generalist invasive (Spodoptera littoralis) herbivore, we estimated both pollen movement among treatments and the visiting behaviour of honeybees versus other wild pollinators.
• Overall, we found that herbivory has weak effects on plant pollen export, either in terms of inter‐treatment movements or of dispersion distance. Plants infested with the native specialised herbivore tend to export less pollen to other plants with the same treatment. Other wild pollinators preferentially visit non‐infested plants that differ from those of honeybees, which showed no preferences. Honeybees and other wild pollinators also showed different behaviours on plants infested with different herbivores, with the former tending to avoid revisiting the same treatment and the latter showing no avoidance behaviour. When taking into account the whole pollinator community, i.e. the interactive effects of honeybees and other wild pollinators, we found an increased avoidance of plants infested by the native specialised herbivore and a decreased avoidance of plants infested by the invasive herbivore.
• Taken together, our results suggest that herbivory may have an effect on B. rapa pollination, but this effect depends on the relative abundance of honeybees and other wild pollinators.

     

Overexpression of AtPAD4 in transgenic Brachypodium distachyon enhances resistance to Puccinia brachypodii

• Brachypodium distachyon (L.) has recently emerged as a model for temperate grasses for investigating the molecular basis of plant–pathogen interactions. Phytoalexin deficient 4 (PAD4) plays a regulatory role in mediating expression of genes involved in plant defence.
• In this research, we generated transgenic B. distachyon plants constitutively overexpressing AtPAD4. Two transgenic B. distachyon lines were verified using PCR and GUS phenotype.
• Constitutive expression of AtPAD4 in B. distachyon enhanced resistance to Puccinia brachypodii. P. brachypodii generated less urediniospores on transgenic than on wild‐type plants. AtPAD4 overexpression enhanced salicylic acid (SA) levels in B. distachyon‐infected tissues. qRT‐PCR showed that expression of pathogenesis‐related 1 (PR1) and other defence‐related genes were up‐regulated in transformed B. distachyon following infection with P. brachypodii.
• Our results indicate that AtPAD4 overexpression in B. distachyon plants led to SA accumulation and induced PR gene expression that reduced the rate of colonisation by P. brachypodii.

     

Defence‐Related Enzymes in Soybean Resistance to Target Spot

Target spot, caused by the fungus Corynespora cassiicola, has become a serious foliar disease in soybean production in the Brazilian Cerrado. Information in the literature regarding the biochemical defence responses of soybean to C. cassiicola infection is rare. Therefore, the objective of this study was to determine the biochemical features associated with soybean resistance to target spot. The activities of chitinases (CHI), β‐1‐3‐glucanases (GLU), phenylalanine ammonia‐lyases (PAL), peroxidases (POX), polyphenol oxidases (PPO) and lipoxygenases (LOX), as well as the concentrations of total soluble phenolics (TSP) and lignin‐thioglycolic acid (LTGA) derivatives, were determined in soybean leaves from both a resistant (FUNDACEP 59) and a susceptible (TMG 132) cultivar. The target spot severity, number of lesions per cm² of leaflet and area under the disease progress curve were significantly lower for plants from cv. FUNDACEP 59 compared to plants from cv. TMG 132. The GLU, CHI, PAL, POX and PPO activities and the concentration of LTGA derivatives increased significantly, whereas LOX activity decreased significantly on the leaves infected by C. cassiicola. Inoculated plants from cv. FUNDACEP 59 showed a higher PPO activity and concentrations of TSP and LTGA derivatives at 4 and 6 days after inoculation compared to plants from cv. TMG 132. In conclusion, the results of this study demonstrated that the defence‐related enzyme activities increased upon C. cassiicola infection, regardless of the basal level of resistance of the cultivar studied. The increases in PPO activity and concentrations of TSP and LTGA derivatives, but lower LOX activity, at early stages of C. cassiicola infection were highly associated with soybean resistance to target spot.     

Development of a CAPS marker for the verticillium wilt resistance in tomatoes

The nucleotide sequences of the Verticillium wilt resistance locus of resistant and susceptible tomato genotypes were cloned and analyzed. The nucleotide sequences displaying high degree of homology to the earlier cloned Ve1 and Ve2 genes were detected in the tomato forms susceptible to Verticillium wilt. The polymorphism at this locus between the resistant and susceptible plants was detected and used to elaborate a highly efficient CAPS marker.     

Hessian fly Avirulence gene loss‐of‐function defeats plant resistance without compromising the larva's ability to induce a gall tissue

Plant pathogen effectors encoded by Avirulence (Avr) genes benefit the pathogen by promoting colonization and benefit plants that have a matching resistance (R) gene by constituting a signal that triggers resistance. The Hessian fly, Mayetiola destructor (Say) (Diptera: Cecidomyiidae), resembles a plant pathogen in showing R/Avr interactions. Because of these interactions, a wheat plant with the H13 resistance gene can be resistant or susceptible depending on the genotype of the larva that attacks the plant, being resistant if attack comes from a larva with a functional vH13 gene, but susceptible if attack comes from a larva with a non‐functional vH13 gene. In this study we asked: does this susceptible interaction involving plants with H13 look like susceptible interactions with plants lacking H13? Possibly, the H13 plant attacked by a larva with a non‐functional vH13 is induced to partial rather than complete resistance. Or the larva, lacking its vH13‐encoded effector, is compromised in its ability to induce susceptibility, which includes forcing the plant to create a gall nutritive tissue. Responses of epidermal cells to larval attack were explored using imaging techniques (light microscopy, scanning and transmission electron microscopy). Whole‐organism responses were investigated by measuring the growth of plants and larvae. No evidence was found for partial resistance responses by H13 plants or for a compromise in the ability of vH13 loss‐of‐function larvae to induce susceptibility. It appears that disrupting vH13 function is sufficient for overcoming the induced resistance mediated by the H13 gene.