Two programmes to investigate the inheritance of resistance to black stem in sunflowers were undertaken in a controlled-environment growth chamber. In the first, an experiment was performed using a randomized complete block design with 24 lines, six male-sterile (A lines), six maintainers (B lines), six restorers (R lines) and their six F 1 hybrids in six replications. Each treatment consisted of 12 seedlings. Twelve-day-old seedlings were inoculated with a suspension of pycniospores, and 7 days later the two cotyledon petioles of each seedling were scored on a 1±9 scale for the percentage of necrotic area. Some alloplasmic lines (which have the same nucleus and different cytoplasms) gave a significant cytoplasmic effect for improved partial resistance to the disease. Deviation of F 1 hybrids from the mean of the parent values was significant for partial resistance to Phoma macdonaldii in three of the six F 1 hybrids. Two further experiments with eight lines (resistant and susceptible) at the fifth leaf-pair and flowering stages were carried out under the same conditions. At both growth stages the previous classification of lines at the seedling stage was confirmed. In the second programme, five male-sterile sunflower lines were crossed with five fertility-restorers in a factorial mating design. The 10 inbred lines and their 25 F 1 hybrids were studied in two successive experiments under the same conditions and with the same experimental design and isolate of Phoma as in the seedling-stage experiment in the first programme. Analysis of variance showed that male-sterile and restorer lines possessed general combining abilities, and also that specific combining abilities of F 1 hybrids were significant. The estimates of general combining ability for partial resistance were significant in AS617A, AS618A and AS614R inbred lines. These lines are available for developing F 1 hybrids with improved resistance to Phoma in sunflower-breeding programmes.

Growth of 17 isolates of Phoma macdonaldii, the causal agent of sunflower black stem, was investigated for response to pH and temperature, and for morphology and asexual morphogenesis (pycnidiogenesis and pycnidium size). For all isolates, the optimum pH for growth was between 4 and 5, and the optimum temperature varied between 20 and 30ЊC and radial growth was slowest at 5 and 35ЊC. Significant differences in the number and size of pycnidia were observed between isolates. Pycniospore germination was investigated under various conditions in five isolates chosen for their geographical origins, pigmentation, optimum growth temperature and pycnidiogenesis. Increasing the concentration from 10 6 to 10 7 pycniospores per mL decreased the germination rate. The optimum temperature for pycniospore germination varied between 15 and 30ЊC, depending on the isolate, and the optimum and maximum pH values were 5 and 7, respectively. The optimum and minimum relative humidities allowing pycniospore germination were 100 and 95%, respectively. Pycniospore germination was photo-independent. An artificial inoculation method was developed and the aggressiveness of the pathogen was assessed on a susceptible sunflower cultivar, using a 1-9 scale that integrated the percentage of necrotic area on the cotyledon petiole at the stage when the first pair of leaves was fully developed. Significant differences in aggressiveness were observed among the 17 isolates. The parameters investigated clearly suggest the occurrence of a wide phenotypic variability in Phoma macdonaldii.

Summary• To identify the genes involved in the partial resistance of sunflower ( Helianthus annuus ) to the necrotrophic fungus Phoma macdonaldii , we developed a 1000-element cDNA microarray containing carefully chosen genes putatively involved in primary metabolic pathways, signal transduction and biotic stress responses.• A two-pass general linear model was used to normalize the data and then to detect differentially expressed genes. This method allowed us to identify 38 genes differentially expressed among genotypes, treatments and times, mainly belonging to plant defense, signaling pathways and amino acid metabolism.• Based on a set of genes whose differential expression was highly significant, we propose a model in which negative regulation of a dual-specificity MAPK phosphatase could be implicated in sunflower defense mechanisms against the pathogen. The resulting activation of the MAP kinase cascade could subsequently trigger defense responses (e.g. thaumatin biosynthesis and phenylalanine ammonia lyase activation), under the control of transcription factors belonging to MYB and WRKY families. Concurrently, the activation of protein phosphatase 2A (PP2A), which is implicated in cell death inhibition, could limit pathogen development.• The results reported here provide a valuable first step towards the understanding and analysis of the P. macdonaldii -sunflower interaction.