Nucleotide binding site-leucine rich repeat (NBS-LRR) proteins are encoded by a ubiquitous gene family in sunflower and frequently harbor disease resistance genes. We investigated NBS-LRR-encoding resistance gene candidates (RGCs) flanking the downy mildew resistance genes Pl ( 8 ) and Pl ( 14 ) and the rust resistance gene R ( Adv ), which map on the NBS-LRR clusters of linkage groups 1 and 13 in sunflower genome. We shotgun sequenced bacterial artificial chromosome (BAC) clones proximal to Pl ( 8 ), Pl ( 14 ) , and R ( Adv ) and identified seven novel non-Toll/interleukin-1 receptor (TIR)-like NBS-LRR RGCs, which clustered with previously identified RGCs of linkage group 13 but were phylogenetically distant from the TIR- and non-TIR-NBS-LRR-encoding superfamilies of sunflower. Six of the seven predicted RGCs have intact open reading frames and reside in genomic segments with abundant transposable elements. The genomic localization and sequence similarity of the novel non-TIR-like predicted RGCs suggests that they originated from tandem duplications. RGCs in the proximity of Pl ( 8 ) and R ( Adv ) were likely introgressed from silverleaf sunflower genome, where the RGC cluster of linkage group 13 is duplicated in two independent chromosomes that have different architecture and level of recombination from the respective common sunflower chromosomes.

The content of coumarins, as probable phytoalexins, was analysed in four sunflower genotypes that ranged in responses to head rot from highly susceptible to highly resistant. Low levels of all coumarins (scopolin, scopoletin and ayapin) were detected in the three most susceptible genotypes irrespective of time after inoculation. However, in the resistant genotype there was a clear time-dependent disease-induced increase of all coumarins that reached a maximum after 10-14 days. Detailed comparison of the most susceptible and the resistant genotype showed that in the resistant but not the susceptible, scopoletin peroxidase activity increased during the course of the experiment. Results confirmed a clear negative correlation between coumarin content and disease symptoms and in particular for scopolin. Furthermore we show for the first time that scopolin is inhibitory to Sclerotinia at similar doses to scopoletin. As scopolin is known to be less phytotoxic than ayapin and scopoletin, its accumulation may well confer head rot resistance with minimal plant damage and might be one of the bases for resistance to Sclerotinia.Abbreviations: EMT, esculetin methyl transferase; SGT, scopoletin glucosyl transferase; SP, scopoletin peroxidase

The establishment of Lotus tenuis can interfere with colonization by Carduus acanthoides during the early post‐burn recovery of Flooding Pampa grasslands. The purpose of this research was to determine the potential role of L. tenuis seeds as a source of allelopathic compounds involved in that interaction. Imbibed seeds of L. tenuis and aqueous leachates from them were bioassayed for their ability to inhibit germination and seedling growth of C. acanthoides, both on sterilized filter paper and on pasteurized soil as substrata. Germination and/or emergence of C. acanthoides were inhibited and root length was reduced on filter paper or soil, by both the presence of L. tenuis seeds and their leachate, at densities of L. tenuis near the maximum values observed in the field. Germination and seedling growth of C. acanthoides were less affected by the presence of L. tenuis seeds than by the addition of their leachate, and the presence of L. tenuis seeds or their leachate showed stronger effects on emergence of C. acanthoides from soil than on its germination on filter paper. Methods applied for leachate sterilization, ultrafiltration or autoclaving did not modify C. acanthoides responses. Neither the germination rate nor the root length of C. acanthoides seedlings were affected by solutions of polyethylene glycol with similar osmolarity to the leachates. We conclude that the release of inhibitory substances on to filter paper and into pot soil from imbibed L. tenuis seeds would be the mechanism responsible for the observed effects.

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Sunflower plants at I (vegetative), II (closed flower‐buds) and III (open flower‐buds) growth stages were experimentally inoculated with Sclerotinia sclerotiorum. At the 5th day after inoculation, all plants from I, III and 70 % of II (IIT) did not present wilt symptoms while 30 % of age II plants (IIs) presented severe wilt symptoms in all their leaves and had collapsed. Total soluble phenols were measured in the “lesion” (L) and its “healthy” (H) surrounding zones of all inoculated and control plants. A significant increase in phenols was found in the L and H zones of tolerant plants (I, IIt and III). The corresponding methanolic extracts showed a strong inhibitory effect on the mycelial growth (MG) of the fungus. On the other hand no statistically significant increase took place in the L zones from susceptible plants; their methanolic extracts did not inhibit MG. The chemical analysis of methanolic extracts showed that the isochlorogenic acid was the more inhibitory compound. It significantly accumulated in the L of tolerant plants but did not in those of susceptible ones.

The object of this work was a study of the relationship between the field reactions of different sunflower genotypes to basal stalk rot (in terms of severe (dead plants) and incipient wilting, and lesion length) and some biochemical (phenol concentration), morphological (plant height, and stem and flower-bud diameters) and anatomical (xylem and cortical indexes) characters of the host. Plants from 8 inbred lines at closed flower-bud stage were artificially inoculated with mycelium at the base of the stem. The percentage of dead plants for each inbred line and the lesion length and wilting range for individual plants after 7 days were recorded.A positive and highly significant correlation coefficient between the percentage of dead plants and lesion length was found for the three years of the study (r = 0.83; P < 0.01). A highly significant association between lesion length and wilting range for individual plants was always found (P = 0.00). Postinfectional phenol content exhibited a strong negative correlation with lesion length and the percentage of dead plants in all the experiments (P = 0.05). Association between postinfectional phenol content and wilting range for individual plants was significant for all the years studied (P < 0.05). No correlation between phenol levels in healthy plants of the different sunflower genotypes and their susceptibility was found. Morphological characters positively correlated with lesion length but only plant height exhibited significant values for the three years. Associations between wilting range and morphological characters for individual plants were significant for one of the two years analyzed (P < 0.05). Xylem index showed a negative correlation with lesion length which was significant one of the two years studied.The lesion length measure seems to be a simple and direct method for resistance screening before the flowering period. Although strong relations with postinfectional phenol levels were found, their determinations would be too much time consuming and not completely reliable. The relationships between other characters measured and disease resistance would indicate that physiological mechanisms could be related to resistance. 196

Sunflower head rot is a major disease caused by Sclerotinia sclerotiorum. Sunflower varieties which are tolerant to the fungus have been developed. The changes occurring in flower parts at different times after inoculation with pathogen ascospores were studied for two sunflower varieties (tolerant HA 302 and susceptible HA 89). In variety HA 302 there was cell collapse, changes in cell wall composition, and an increase in phenolic compounds in the tissues of corolla and style, which prevented the pathogen from advancing. This response was weaker in susceptible variety HA 89, and occurred only in the style, so did not stop the pathogen from developing and reaching the ovary. Phenolic compounds were found in HA 302 corolla and style tissues only when the pathogen was present, constituted an induced response that prevented further development of the fungus. Principal component analysis (PCA) showed that at the beginning of the infection there was no difference in behavior between the two varieties. The difference arose during the final observation times, when in variety HA 89, the pathogen colonized ovary, style and base of filaments and produced noticeable colonization of the corolla.

In a previous study, we observed that bract and corolla extracts from a Sclerotinia sclerotiorum-resistant sunflower contained high amounts of the known coumarins scopoletin, scopolin, and ayapin. There was a correlation between coumarin concentration and disease resistance. Thin layer chromatography showed higher concentrations of three other compounds in the resistant genotype when compared to the susceptible. A bioassay-directed purification that used column chromatography and HPLC allowed the isolation of a new compound, 3-acetyl-4-acetoxyacetophenone, and known compounds, demethoxyencecalin and 3-acetyl-4-hydroxyacetophenone. Structures were assigned from spectral data, and bioactivities were characterized by in vitro bioassays against S. sclerotiorum. The new compound, 3-acetyl-4-acetoxyacetophenone, had an antifungal activity similar to the coumarin ayapin, previously described as a potent Sclerotinia inhibitor. The speed and simplicity by which these compounds can be detected make them suitable for use in screening procedures that may identify genotypes with valuable levels of resistance. A screening of seven sunflower genotypes in a field experiment showed a correlation between these compounds and resistance to Sclerotinia.