Extracellular DNA as an elicitor of broad-spectrum resistance in Arabidopsis thaliana.

Like in mammals, the plant immune system has evolved to perceive damage. Damaged-associated molecular patterns (DAMPs) are endogenous signals generated in wounded or infected tissue after pathogen or insect attack. Although extracellular DNA (eDNA) is a DAMP signal that induces immune responses, plant responses after eDNA perception remain largely unknown. Here, we report that signaling defenses but not direct defense responses are induced after eDNA applications enhancing broad-range plant protection. A screening of defense signaling and hormone biosynthesis marker genes revealed that OXI1, CML37 and MPK3 are relevant eDNA-Induced Resistance markers (eDNA-IR). Additionally, we observed that eDNA from several Arabidopsis ecotypes and other phylogenetically distant plants such as citrus, bean and, more surprisingly, a monocotyledonous plant such as maize upregulates eDNA-IR marker genes. Using 3,3'-Diaminobenzidine (DAB) and aniline blue staining methods, we observed that H2O2 but not callose was strongly accumulated following self-eDNA treatments. Finally, eDNA resulted in effective induced resistance in Arabidopsis against the pathogens Hyaloperonospora arabidopsidis, Pseudomonas syringae, and Botrytis cinerea and against aphid infestation, reducing the number of nymphs and moving forms. Hence, the unspecificity of DNA origin and the wide range of insects to which eDNA can protect opens many questions about the mechanisms behind eDNA-IR.

Molecular characterization for food safety assessment of a genetically modified late blight resistant potato: an unusual case.

Standard food safety assessments of genetically modified crops require a thorough molecular characterization of the novel DNA as inserted into the plant that is intended for commercialization, as well as a comparison of agronomic and nutritional characteristics of the genetically modified to the non-modified counterpart. These characterization data are used to identify any unintended changes in the inserted DNA or in the modified plant that would require assessment for safety in addition to the assessment of the intended modification. An unusual case of an unintended effect discovered from the molecular characterization of a genetically modified late blight resistant potato developed for growing in Bangladesh and Indonesia is presented here. Not only was a significant portion of the plasmid vector backbone DNA inserted into the plant along with the intended insertion of an R-gene for late blight resistance, but the inserted DNA was split into two separate fragments and inserted into two separate chromosomes. One fragment carries the R-gene and the other fragment carries the NPTII selectable marker gene and the plasmid backbone DNA. The implications of this for the food safety assessment of this late blight resistant potato are considered.

Plant immunity in natural populations and agricultural fields: Low presence of pathogenesis-related proteins in Solanum leaves.

Plant immunity has mainly been studied under controlled conditions, limiting our knowledge regarding the regulation of immunity under natural conditions where plants grow in association with multiple microorganisms. Plant pathology theory, based on laboratory data, predicts complex biochemical plant-pathogen interactions leading to coevolution of pathogen infectivity vs. plant recognition of microbes in multiple layers over time. However, plant immunity is currently not evaluated in relation to ecological time-scales and field conditions. Here we report status of immunity in plants without visible disease symptoms in wild populations of nightshades, Solanum dulcamara and Solanum nigrum, and in agricultural fields of potato, Solanum tuberosum. We analysed presence of pathogenesis-related proteins in over 500 asymptomatic leaf samples collected in the field in June, July and August over three years. Pathogenesis-related proteins were present in only one-third of the collected samples, suggesting low activity of the immune system. We could also detect an increase in pathogenesis-related proteins later in the growing season, particularly in S. tuberosum. Our findings, based on pathogenesis-related protein markers, indicate major gaps in our knowledge regarding the status and regulation of plant immunity under field conditions.

Ex situ conservation priorities for the wild relatives of potato (solanum L. Section petota).

Crop wild relatives have a long history of use in potato breeding, particularly for pest and disease resistance, and are expected to be increasingly used in the search for tolerance to biotic and abiotic stresses. Their current and future use in crop improvement depends on their availability in ex situ germplasm collections. As these plants are impacted in the wild by habitat destruction and climate change, actions to ensure their conservation ex situ become ever more urgent. We analyzed the state of ex situ conservation of 73 of the closest wild relatives of potato (Solanum section Petota) with the aim of establishing priorities for further collecting to fill important gaps in germplasm collections. A total of 32 species (43.8%), were assigned high priority for further collecting due to severe gaps in their ex situ collections. Such gaps are most pronounced in the geographic center of diversity of the wild relatives in Peru. A total of 20 and 18 species were assessed as medium and low priority for further collecting, respectively, with only three species determined to be sufficiently represented currently. Priorities for further collecting include: (i) species completely lacking representation in germplasm collections; (ii) other high priority taxa, with geographic emphasis on the center of species diversity; (iii) medium priority species. Such collecting efforts combined with further emphasis on improving ex situ conservation technologies and methods, performing genotypic and phenotypic characterization of wild relative diversity, monitoring wild populations in situ, and making conserved wild relatives and their associated data accessible to the global research community, represent key steps in ensuring the long-term availability of the wild genetic resources of this important crop.

Expressed sequence tags in cultivated peanut (Arachis hypogaea): discovery of genes in seed development and response to Ralstonia solanacearum challenge.

Although an important oil crop, peanut has only 162,030 expressed sequence tags (ESTs) publicly available, 86,943 of which are from cultivated plants. More ESTs from cultivated peanuts are needed for isolation of stress-resistant, tissue-specific and developmentally important genes. Here, we generated 63,234 ESTs from our 5 constructed peanut cDNA libraries of Ralstonia solanacearum challenged roots, R. solanacearum challenged leaves, and unchallenged cultured peanut roots, leaves and developing seeds. Among these ESTs, there were 14,547 unique sequences with 7,961 tentative consensus sequences and 6,586 singletons. Putative functions for 47.8 % of the sequences were identified, including transcription factors, tissue-specific genes, genes involved in fatty acid biosynthesis and oil formation regulation, and resistance gene analogue genes. Additionally, differentially expressed genes, including those involved in ethylene and jasmonic acid signal transduction pathways, from both peanut leaves and roots, were identified in R. solanacearum challenged samples. This large expression dataset from different peanut tissues will be a valuable source for marker development and gene expression analysis. It will also be helpful for finding candidate genes for fatty acid synthesis and oil formation regulation as well as for studying mechanisms of interactions between the peanut host and R. solanacearum pathogen.

Production of Phytophthora infestans-resistant potato (Solanum tuberosum) utilising Ensifer adhaerens OV14.

Based on the use of Agrobacterium tumefaciens-mediated transformation commodity crop improvement through genetic engineering is the fastest adopted crop technology in the world (James 2010). However, the complexity of the Agrobacterium patent landscape remains a challenge for non-patent holders who wish to generate novel varieties for a commercial purpose. The potential of non-Agrobacterium strains (Transbacter(™)) to modify a plant genome has previously been described. However, they are unlikely to be widely used without significant adjustments in transformation protocols in order to improve their gene transfer efficiencies. In this study we set out to identify alternative bacteria species that could (a) utilize vir genes for genetic transformation and (b) substitute for A. tumefaciens in existing transformation protocols, without a prerequisite for protocol modifications. To this end we isolated a collection (n=751) of plant-associated bacteria from the rhizosphere of commercially grown crops. Based on various screens, including plant transformation with the open-source vector pCAMBIA5105, we identified a strain of the bacterium Ensifer adhaerens with the capacity to transform both Arabidopsis thaliana (0.12%) and potato (mean transformation frequency 35.1%). Thereafter, Ensifer adhaerens was used to generate blight- (causative organism Phytophthora infestans) resistant potato using the Solanum bulbocastanum 'resistance to blight' (RB) gene. Resistant genotypes were confirmed by associated molecular analysis and resistant phenotypes demonstrated by the development of hypersensitive lesions on inoculated leaf tissue post-pathogen inoculation. These data confirm the potential of Ensifer-mediated transformation (EMT) as a novel platform for the high frequency generation of transgenic potato.

Prevalence of sensitisation to oilseed rape and maize pollens in France: a multi-center study carried out by the Allergo-Vigilance Network.

BACKGROUND: Oilseed rape and maize crops represent a large part of agriculture fields in European countries. OBJECTIVE: To establish the actual prevalence of sensitization to oilseed rape and maize pollen, and to determine if this is correlated to the amount of exposure as well as to the patient's history of atopy or asymptomatic atopy. METHODS: The study was conducted by 69 allergists belonging to the Allergo-Vigilance Network, in collaboration with the French Agency for Safety of food, and compiles the results of skin prick-tests using oilseed rape and maize pollens and seeds, as well as common aeroallergens. The patients were classified into 3 groups: nonatopic, asymptomatic atopy, and actual atopic diseases. RESULTS: Among the 5372 subjects studied (2515 children, 2857 adults), 62.3% had an atopic disease, 10.2% had an asymptomatic atopy, and 27.5% were non-atopic. The level of sensitization was higher in the subjects with atopic disease, as compared to those with asymptomatic atopy: oilseed rape pollen: 11.8% vs 8%, maize pollen, 26% vs 19%, oilseed rape seeds, 7.7% vs 6.9%, corn seeds: 8.3% vs 4.8% (p < 0.001). The rate of sensitization was significantly increased in those living in high crop density regions. The association of an atopic disease with a high rate of exposure yielded a higher rate of sensitization of 13.8% and 21.3% for rapeseed pollen, and 22.9% and 30.7% for maize pollen in both children and adults, respectively. CONCLUSIONS: The incidence of sensitisation to rapeseed and maize pollen is positively correlated to the level of exposure. This prevalence is higher in patients with actual atopic disease as compared to those with asymptomatic atopy. The frequency of sensitization confirms the allergenicity of these plants destined for food supply and demonstrates the importance of monitoring for respiratory allergies to these pollens, not only in workers exposed to these types of crops, but also in atopic patients living in regions that contain a high density of rapeseed and maize fields. Cross-reactivities between pollens and seeds could potentially elicit cross-reacting food allergies.

Field testing, gene flow assessment and pre-commercial studies on transgenic Solanum tuberosum spp. tuberosum (cv. Spunta) selected for PVY resistance in Argentina.

Solanum tuberosum ssp. tuberosum (cv. Spunta) was transformed with a chimeric transgene containing the Potato virus Y (PVY) coat protein (CP) sequence. Screening for PVY resistance under greenhouse conditions yielded over 100 independent candidate lines. Successive field testing of selected lines allowed the identification of two genetically stable PVY-resistant lines, SY230 and SY233, which were further evaluated in field trials at different potato-producing regions in Argentina. In total, more than 2,000 individuals from each line were tested along a 6-year period. While no or negligible PVY infection was observed in the transgenic lines, infection rates of control plants were consistently high and reached levels of up to 70-80%. Parallel field studies were performed in virus-free environments to assess the agronomical performance of the selected lines. Tubers collected from these assays exhibited agronomical traits and biochemical compositions indistinguishable from those of the non-transformed Spunta cultivar. In addition, an interspecific out-crossing trial to determine the magnitude of possible natural gene flow between transgenic line SY233 and its wild relative Solanum chacoense was performed. This trial yielded negative results, suggesting an extremely low probability for such an event to occur.

SolRgene: an online database to explore disease resistance genes in tuber-bearing Solanum species.

BACKGROUND: The cultivated potato (Solanum tuberosum L.) is an important food crop, but highly susceptible to many pathogens. The major threat to potato production is the Irish famine pathogen Phytophthora infestans, which causes the devastating late blight disease. Potato breeding makes use of germplasm from wild relatives (wild germplasm) to introduce resistances into cultivated potato. The Solanum section Petota comprises tuber-bearing species that are potential donors of new disease resistance genes. The aim of this study was to explore Solanum section Petota for resistance genes and generate a widely accessible resource that is useful for studying and implementing disease resistance in potato. DESCRIPTION: The SolRgene database contains data on resistance to P. infestans and presence of R genes and R gene homologues in Solanum section Petota. We have explored Solanum section Petota for resistance to late blight in high throughput disease tests under various laboratory conditions and in field trials. From resistant wild germplasm, segregating populations were generated and assessed for the presence of resistance genes. All these data have been entered into the SolRgene database. To facilitate genetic and resistance gene evolution studies, phylogenetic data of the entire SolRgene collection are included, as well as a tool for generating phylogenetic trees of selected groups of germplasm. Data from resistance gene allele-mining studies are incorporated, which enables detection of R gene homologs in related germplasm. Using these resources, various resistance genes have been detected and some of these have been cloned, whereas others are in the cloning pipeline. All this information is stored in the online SolRgene database, which allows users to query resistance data, sequences, passport data of the accessions, and phylogenic classifications. CONCLUSION: Solanum section Petota forms the basis of the SolRgene database, which contains a collection of resistance data of an unprecedented size and precision. Complemented with R gene sequence data and phylogenetic tools, SolRgene can be considered the primary resource for information on R genes from potato and wild tuber-bearing relatives.

Occupational IgE-mediated allergy after exposure to lupine seed flour.

The ingestion of lupine seed flour (LSF) has been reported as a cause of allergic reactions, particularly in patients sensitized to peanut, but there is little evidence of its allergenic potential after inhalation. We sought to evaluate the clinical and immunologic reactivity to lupine in employees working with this seed flour. An occupational history was obtained in 7 subjects (median age, 35 years) working with LSF at an agricultural research center. Three subjects (1, 6, and 7) reported work-related allergy symptoms immediately after being exposed to lupine. Skin prick test results with LSF extract were positive in these 3 patients with work-related symptoms. Moreover, lupine-specific IgE antibodies were detected in subjects 6 and 7. In subject 6, the controlled exposure to LSF elicited immediate naso-ocular symptoms without changes in FEV(1). In subject 7, a bronchial provocation with LSF extract elicited an immediate fall (25%) in FEV(1). Double-blinded, placebo-controlled LSF oral challenge results were positive in subjects 6 and 7. Immunologic reactivity to other legumes was detected in subjects 6 and 7, but specific inhalation testing and oral challenge results were negative. Thus, the inhalation of lupine flour could be an important cause of allergic sensitization in exposed workers and might give rise to occupational asthma and food allergy.