Nitrogen and water management can limit premature ripening of sunflower induced by Phoma macdonaldii

Premature ripening (PR) is one of the most important diseases of sunflower in France since the 90s. Previous results indicated that girdling canker of the stem base, caused by Phoma macdonaldii was its primary cause but elucidation of critical environmental factors involved is crucial for better control of the disease. A field study was conducted in three contrasting cropping seasons (2006–2008) and investigated the effect of N fertilization (0, 75 and 150 kg N ha−1) and water regime (rainfed, irrigated) on two cultivars with artificial inoculation (AI) and natural infection (NI). Disease assessment was recorded weekly to calculate the area under disease progress curve (AUDPC) and the final percentage of PR plants. Data showed that high levels of N fertilization led to significantly (P < 0.05) more PR than non-fertilization. Water deficit conditions were significantly (P < 0.05) involved in disease severity, and AUDPC and PR were increased when dry conditions were associated with high N supply. This was true for two cultivars which differed in their susceptibility to the disease but cv. Heliasol RM was significantly (P < 0.05) more affected than cv. Melody, partially resistant to PR. Despite contrasting weather patterns, these results demonstrated a clear role of crop management and environmental conditions on the incidence and severity of stem base attacks responsible for the PR syndrome. These findings suggest that sunflower crop husbandry should be adapted to minimize premature ripening induced by P. macdonaldii

Genotype-isolate interaction for resistance to black stem in sunflower (Helianthus annuus)

Two experiments were undertaken to determine the partial resistance of sunflower genotypes to seven isolates of Phoma macdonaldii. In the first experiment, 28 genotypes, including recombinant inbred lines and their parents, M6 mutant lines developed by gamma irradiation, and some genotypes from different geographical origins, were used. The experiment consisted of a split‐plot design with three replications, each with 12 seedlings per genotype per isolate, in controlled conditions. Seven days after inoculation, plantlets were scored on a 1–9 scale for percentage necrotic area. Highly significant differences were observed among genotypes, isolates and their interactions. The presence of a differential interaction between genotypes and P. macdonaldii isolates was confirmed in a second experiment using 12 genotypes representing large variability for partial resistance to P. macdonaldii isolates, as identified in the first experiment. Inbred lines B454/03, ENSAT‐B5 and LC1064C were the most susceptible sunflower genotypes, whereas two American lines SDR19 and SDR18 presented high partial resistance to all P. macdonaldii isolates studied. The least and most aggressive isolates were MA6 and MP3, respectively. Isolates interacted differentially with sunflower genotypes. This study identified two genotypes (AS613 and PAC2) presenting specific resistance to isolate MP8. The results also showed a wide range of isolate‐nonspecific partial resistances among the lines tested. The information presented here could assist sunflower breeders to choose parents of crosses for breeding of durable resistance to phoma black stem disease

Low plant density can reduce sunflower premature ripening caused by Phoma macdonaldii

In France, premature ripening (PR) is a widespread damage of a fungal disease of sunflower caused by Phoma macdonaldii. Previous results indicated that girdling canker at the stem base, caused by P. macdonaldii, was its primary cause. Previous studies have reported the influence of nitrogen and water supply on the incidence and severity of PR but an additional study was required to analyze the effect of plant density on the level of attack for a more comprehensive cultural control of PR. In a 2-year field study (2008 and 2009) in Toulouse (France), a susceptible cultivar (cv. Heliasol) artificially inoculated at star bud stage with P. macdonaldii was grown at three plant densities (4, 6.5 and 9 plants m−2) factorially combined with three N fertilization rates (0, 50/75 and 150 kg N ha−1) and two water regimes (irrigated and rainfed). P. macdonaldii symptoms were scored weekly to calculate the area under disease progress curve (AUDPC) and percentage of PR plants. Microclimatic conditions were monitored using thermo-hygrometers within the crop. The fraction of photosynthetically active radiation intercepted by the canopy (fPARi) and leaf area index (LAI) were measured at anthesis. Plant water status during the disease progression was characterized by crop simulation (SUNFLO) and N status at anthesis was assessed from shoot N content (Nm) analysis and N Nutrition Index (NNI) calculation. Increasing plant density resulted in a greater proportion of PR plants, and this proportion increased further when N was applied at 150 kg ha−1, the highest rate, and the crop was not irrigated. Despite differing canopy development, differences in microclimatic conditions between density levels were too small to explain the PR differences. However plant N concentration and diameter at stem base were closely related to PR incidence. Thin plants (grown at high density) with non-limiting N supply were the most susceptible to premature ripening. This study opens new avenues for the control of PR through crop management and emphasizes the key role of plant morphology in the development of the disease. Stem base diameter is a morphological trait that could be manipulated through crop management (plant density, N fertilization) and probably breeding in the future when developing integrated disease management systems in sunflower

Genetic control of partial resistance to ‘collar’ and ‘root’ isolates of Phoma macdonaldii in sunflower

Phoma macdonaldii is one of the most important pathogens of sunflower (Heliantus annuus) in France. In order to determine the inheritance of resistance to the disease, five sunflower genotypes with wide genetic variability for resistance to two ‘collar’ and two ‘root’ Phoma isolates were crossed in a diallel programme. Four separate experiments were undertaken under controlled conditions. In each one, the response of parental genotypes and their F1 hybrids were evaluated with one of the four Phoma isolates. Analysis of variance was performed to determine the effects of genotype on disease severity score when inoculated with ‘collar’ or ‘root’ Phoma isolates and showed significant variability among parents and F1 hybrids for disease severity score. Diallel analysis showed that general combining ability (GCA) and specific combining ability (SCA) effects for resistance to ‘collar’ and ‘root’ Phoma isolates were highly significant for each of the four isolates indicating that both kinds of gene effects were important in controlling the resistance. The GCA/SCA ratios were more than one for three out of four isolates showing that additive genetic effects were more important than non-additive effects for resistance to three of the studied Phoma isolates. Hence, conventional breeding methods could be recommended to achieve genetic improvement to such ‘collar’ and ‘root’ Phoma isolates

Use of GGEbiplot methodology and Griffing's diallel method for genetic analysis of partial resistance to phoma black stem disease in sunflower

The objectives of the present study were to estimate the general combining ability (GCA) and specific combining ability (SCA) for partial resistance to phoma black stem and to identify the most promising combination for the selection of improved breeding lines. The response of five parental genotypes and their F1 hybrids to a phoma black stem isolate (MA6) were evaluated in a diallel programme under controlled growth chamber conditions. Significant GCA and SCA indicate that both additive and non-additive gene effects contributed in the inheritance of partial resistance to phoma black stem, however, the Baker ratio showed that the additive genetic effects were more important than nonadditive ones. It is recommended that the GGEbiplot methodology could be an excellent tool for visualizing entry by tester (diallel) data. By using this technique to analyse black stem severity data, interaction among the sunflower genotypes in providing partial resistance to phoma black stem was clearly identified. Based on GGEbiplot presentation and Griffing's diallel analysis, the mutant line ‘M6-54-1’ showed the largest GCA, indicating contribution towards partial resistance, and the genotype B454/03 presented the smallest GCA, indicating contribution towards susceptibility. Our results show that the F1 hybrids ‘SDR18×B454/03’ and ‘M6-54-1×B454/03’ showing heterosis for partial resistance to phoma black stem come from the crosses between a susceptible genotype ‘B454/03’ and two partially resistant genotypes (SDR18 and M6-54-1), originated from different breeding programmes. We conclude therefore that these genotypes possess at least some different resistance genes, which were expressed in the hybrids and led to the observed effects

Quantitative trait loci associated with isolate specific and isolate nonspecific partial resistance to Phoma macdonaldii in sunflower

Black stem, caused by Phoma macdonaldii, is one of the most important diseases of sunflower in the world. Quantitative trait loci (QTLs) implicated in partial resistance to two single pycnidiospore isolates of P. macdonaldii (MP8 and MP10) were investigated using 99 recombinant inbred lines (RILs) from the cross between sunflower parental lines PAC2 and RHA266. The experimental design was a randomized complete block with three replications. High genetic variability and transgressive segregation were observed among RILs for partial resistance to P. macdonaldii isolates. QTL‐mapping was performed using a recently developed high‐density SSR/AFLP sunflower linkage map. A total of 10 QTLs were detected for black stem resistance. The phenotypic variance explained by each QTL (R2) was moderate, ranging from 6 to 20%. Four QTLs were common between two isolates on linkage group 5 and 15 whereas the others were specific for each isolate. Regarding isolate‐specific and isolate‐nonspecific QTLs detected for partial resistance, it is evident that both genetic effects control partial resistance to the disease isolates. This confirms the need to consider different isolates in the black stem resistance breeding programmes. The four SSR markers HA3700, SSU25, ORS1097 and ORS523_1 encompassing the QTLs for partial resistance to black stem isolates could be good candidates for marker assisted selection

Characterisation of sunflower root colonisation by Phoma macdonaldii

Phoma macdonaldii, the causal agent of black stem disease of sunflower (Helianthus annuus), also attacks roots and collars of the plants, resulting in early death. Totally resistant lines do not exist for infection of the aerial parts, but tolerant lines have been characterised. This paper presents a study on colonisation of a partially resistant and a susceptible sunflower line by P. macdonaldii. The fungus was transformed with a constitutively expressed reporter gene encoding the jellyfish green fluorescent protein via Agrobacterium tumefaciens, and colonisation of sunflower roots by this transformed strain was studied by various microscopy techniques including confocal and scanning electron microscopy. The results show that penetration of the fungus into the root occurred through natural fissures or through the epidermis and was similar in both lines. In contrast, the colonisation rate of the stele was reduced in the partially resistant line, and the morphology of the fungal hyphae was also affected. The effect on hyphal morphology was strongest in the stele, indicating a localised production of defence compounds in this line

Analyse et modélisation des effets des pratiques culturales sur les épidémies de mildiou de la pomme de terre. Adaptation du modèle SIPPOM (Simulator for Integrated Pathogen POpulation Management) au pathosystème

Le mildiou de la pomme de terre, causé par l'agent pathogène Phytophthora infestans est l'une des maladies les plus préjudiciables de la culture. Jusqu'à présent, la lutte chimique reste le moyen de contrôle le plus utilisé pour la maîtriser, classant la pomme de terre au premier rang en termes d'Indices de Fréquence de Traitement en grande culture. Par ailleurs, l'utilisation de variétés résistantes, comportant notamment des gènes de résistances spécifiques, a également démontré son efficacité pour limiter les dégâts engendrés par cette maladie. Mais leur efficacité est peu durable avec une durée moyenne de 4 ans avant l'apparition du phénomène de contournement par les isolats plus virulents. Il est donc nécessaire de développer des stratégies de contrôle de la maladie en combinant un ensemble d'approches génétiques, culturales, physiques, et chimiques afin de satisfaire au mieux les objectifs agronomiques, environnementaux et socio-économiques. Un modèle, nommé SIPPOM (Simulator for Integrated Pathogen POpulation Management), avait été développé dans le cas de la gestion durable du phoma du colza. Le présent travail a consisté à adapter la structure générique de SIPPOM au cas de la gestion intégrée du mildiou de la pomme de terre en développant des modules spécifiques à partir de modèles préexistants ou développés spécifiquement. Un modèle de culture (Spudgro), un modèle épidémiologique (Guntz-Divoux / Milsol), un modèle de nuisibilité (modèle de Shtienberg) et une fonction de dispersion (modèle de Scherm) ont été identifiés dans la littérature, adaptés et intégrés dans la structure générique de SIPPOM. Cet ensemble de modules a donné lieu à la réalisation du premier prototype opérationnel de la version informatisée de SIPPOM-de-terre sur la plate-forme de modélisation RECORD. Un nouveau modèle, appelé VOLPONE, a été développé pour représenter les dynamiques de repousses de pomme de terre dans une parcelle et sur un tas de déchets. Ce modèle permet de représenter les sources d'inoculum primaire à l'échelle du territoire sous l'influence du climat et des pratiques agricoles. La qualité prédictive du modèle de nuisibilité a été estimée à partir d'un jeu de données expérimentales générées au cours du travail de thèse et s'est montrée correcte. Des exemples de résultats issus de simulations réalisées à l'aide du premier prototype montrent la capacité du modèle, en fonction d'une combinaison de pratiques culturales et de conditions climatiques variables, à prédire la dynamique épidémique de la maladie, ainsi que les dégâts et dommages associés à l'échelle de la parcelle ou sur un parcellaire simple. Néanmoins, d'autres tâches restent à accomplir afin de disposer d'une version achevée de SIPPOM-de-terre sur la plate-forme RECORD. Le travail réalisé illustre la généricité du modèle d'origine SIPPOM et contribue à la production de connaissances et de références permettant l'optimisation d'outils existants tels Mileos®. Les avancées réalisées pourront contribuer à la conception de stratégies de gestion intégrée, collectives et durables pour cette maladie.Potato late blight caused by the pathogen Phytophthora infestans is one of the most dreaded diseases to this culture. So far, chemical control is the most widespread method used against the pathogen. In turn, the amount of fungicides used to control this disease makes potato the crop with the highest Treatment Frequency Index of all arable crops. The use of resistant varieties, including specific resistance genes, has also demonstrated an effective limitation of injuries that this disease can cause. Still, specific resistances lack durability as there is an average span of 4 years before the emergence of a resistance breakdown phenomenon by more virulent strains. It is therefore necessary to develop control strategies that combine a set of methods (genetic, cultural, physical and chemical) to meet agronomic, environmental and socio-economic criteria. A model named SIPPOM (Simulator for Integrated Pathogen POpulation Management), was developed to address these issues for the control of phoma stem canker on oilseed rape. The present work consisted in adapting the generic structure of SIPPOM for the integrated control of potato late blight by designing specific sub-models using existing models or specifically developed models. A crop model (Spudgro), an epidemiologic model (Guntz-Divoux/Milsol), a damage model (Shtienberg's model) and a dispersal function (Scherm's model) were selected in the literature, adapted, and embedded in SIPPOM's generic structure. This set of sub-models led to an operational prototype of SIPPOM-de-terre under the RECORD modelling platform. A new model, named VOLPONE, was designed to simulate potato volunteer dynamics in a field or on a waste pile. This model permits to simulate inoculum sources at the territory scale under the influence of climate and cropping practices. The predictive quality of the damage model was assessed with data generated in specific field experimentations and proved to be good. Simulation examples produced with the first prototype of SIPPOM-de-terre illustrate the capacity of the model to represent epidemiological dynamics at the field scale or at a small regional scale under the influence of cropping practices and climate. However, further work is required before to totally achieve the design and the implementation of SIPPOM-de-terre under the modelling platform RECORD. The conducted study illustrates the genericity of the model SIPPOM and produced knowledge, references and tools for the integrated management of the disease. The outputs of this work will help design integrated, collective and durable control strategies of potato late blight.TOULOUSE-ENSAT-Documentation (315552324) / SudocSudocFranceF

Phenotypic variability of Leptosphaeria lindquistii (anamorph: Phoma macdonaldii), a fungal pathogen of sunflower

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 30°C and radial growth was slowest at 5 and 35°C. 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 106 to 107 pycniospores per mL decreased the germination rate. The optimum temperature for pycniospore germination varied between 15 and 30°C, 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

Ultrastructural studies of the mode of penetration byPhoma macdonaldiiin sunflower seedlings

An ultrastructural investigation of the artificial inoculation of sunflower with Phoma macdonaldii conidia was undertaken using light, scanning, and transmission electron microscopy to elucidate the host-parasite relationship. The behavior of the conidia deposited on the cotyledon petiole was investigated at various time intervals after inoculation. Conidia adhesion and germination were observed first. The cotyledon petiole was invaded by the fungus directly through the cuticle and via stomata. Externally, the spore and germ tube were covered with a mucilaginous polysaccharide sheath of a cotton-like appearance and of variable thickness. At the time of penetration, the host cuticle was perforated mechanically. The cuticle was slightly depressed and no enzymatic alteration could be observed. The fungus did not form appressoria on the surface of the host tissues but developed an infection peg. As soon as the cuticle barrier was crossed, the fungus rapidly colonized the host parietal layer. In a first step, the plasmalemma of the host cell appeared to be stuck against the cell wall. As soon as the fungus passed through the epidermal cell wall to reach the host cytoplasm, the plasmalemma was disrupted, and the subsequent rapid breakdown of cell integrity favored the colonization of the tissues by the pathogen