Déterminisme génétique de la résistance du tournesol au Phoma

L'interaction "génotype-isolat" pour la résistance partielle au Phoma macdonaldii chez le tournesol a été déterminée en utilisant 28 génotypes inoculés par sept isolats de ce pathogène dans des conditions contrôlées. Deux lignées AS613 et PAC2 ont été isolées, qui présentent une résistance spécifique à l’isolat MP8. Nos résultats montrent également l’existence d’une résistance non-spécifique chez les génotypes testés. Pour déterminer l'hérédité de la résistance partielle à cette maladie, nous avons effectué des croisements entre cinq génotypes sélectionnés parmi les 28 de la première expérimentation présentant une variabilité importante pour la résistance partielle à des isolats du Phoma. Les génotypes parentaux et leurs hybrides F1 ont été inoculés par deux isolats sélectionnés (MP8 et PM10). Les aptitudes générales et spécifiques à la combinaison pour la résistance aux deux isolats sont significatives, indiquant que les effets additif et dominant des gènes sont importants dans le contrôle génétique de la résistance partielle au Phoma. Nous avons développé une nouvelle carte de liaison du tournesol en utilisant 100 individus F2 issus du croisement entre M6-54-1 et ENSAT-B4, identifiés par nos expérimentations précédentes. Les régions génomiques contrôlant la résistance partielle aux isolats de P. macdonaldi ont été identifiés. Un QTL localisé sur le groupe de liaison (LG) 8 est commun aux trois isolats testés. Des QTLs spécifiques sont également identifiés pour chaque isolat. Dans une autre expérimentation et en utilisant la carte récemment enrichie de notre laboratoire, des QTLs contrôlant la résistance partielle au Phoma ont été identifiés par l’étude d’une population de 99 LRs issue du croisement entre PAC2 et RHA266 inoculé par deux isolats (MP8 et MP10). Parmi les 10 QTLs détectés, quatre situés sur LG 5 et LG15 sont commun pour les deux isolats, les autres sont spécifiques de chaque isolat. La variabilité génétique de 60 mutants a été étudiée en utilisant les marqueurs AFLP. Des marqueurs potentiellement associés à la résistance partielle au Phoma ont été identifiés. le profil de 12 gènes potentiellement associés à la résistance partielle du tournesol à P. macdonaldii a été testé sur le mutant M6-54-1 et la lignée originelle AS613, inoculés par trois isolats de P. macdonaldii en utilisant la RT-PCR quantitative. Les gènes codant pour la phenylalanine ammonia-lyase 2 (PAL2) et la thaumatine ont montré des différences significatives au niveau de l'expression, entre les interactions compatibles et partiellement compatibles. Une corrélation directe entre les niveaux d'expression de gènes codant pour le facteur de transcription HD- Zip et la protéine phosphatase avec le symptôme de la maladie a été observée chez la lignée originelle et son mutant inoculé par l’isolat, MA6. ABSTRACT : Genotype-isolate interaction for resistance to Phoma black stem in sunflower were investigated using 28 genotypes, inoculated by seven isolates of Phoma macdonaldii in controlled conditions. Two specific resistant lines (AS613 and PAC2) to MP8 isolate were identified. Our results showed that there was also a wide range of isolate- onspecificpartial resistance among the genotypes tested. In order to determine the inheritance of partial resistance to the disease, five genotypes of 28 with wide range of genetic variability for partial resistance were crossed. Parental genotypes and their F1 hybrids were inoculated by one of the two selected isolates (MP8 and PM10). General and specific combining ability effects for resistance to Phoma black stem were highly significant for each of the two isolates, indicating that additive and dominant gene effects were important in controlling partial resistance to Phoma black stem. A new linkage map was developed using 100 F2 plants derived from the cross between M6- 54-1 and ENSAT-B4 selected according to the results of the above-mentioned experiments, and the gnomic regions controlling partial resistance to three P. macdonaldii were identified. One QTL located on linkage group (LG) 8 was common to the three isolates tested. Other QTLs were specific for each isolate separately. QTLs controlling quantitative resistance to Phoma black stem was also investigated using our new reference map (Poormohammad kiani et al., 2007) with 99 RILs derived from the cross between PAC2 and RHA266 inoculated by the two isolates (MP8 and MP10). Through ten detected QTLs, four of them were common between two isolates on LG5 and LG15 whereas the others were specific for each isolate. Genetic variability among a population of 60 mutant lines was studied using AFLP markers and putative molecular markers associated with partial resistance to Phoma black stem was identified. The profile of 12 genes potentially associated with partial resistance of sunflower to P. macdonaldii was investigated in resistant mutant line M6-54-1 and its original line AS613, which were selected according to the results of our previous experiments using quantitative real-time RT-PCR. Plants were inoculated by three P. macdonaldii isolates. Genes encoding phenylalanine ammonia-lyase 2 (PAL2) and thaumatin-like protein showed significant differences in expression between partial compatible and compatible interactions. A straightforward correlation between expression levels of genes encoding HD-Zip transcription factor and protein phosphatase with disease symptom was observed in original line and its mutant inoculated by MA6 isolat

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

Genotype-isolate interaction for resistance to Sclerotinia sclerotiorum in sunflower

The sunflower (Helianthus annuus L.) is one of the most important crops grown for edible oil. Sclerotinia sclerotiorum (Lib.) de Bary is a common and widespread pathogen of sunflower. In the present study the reaction of 35 genotypes, including recombinant inbred lines and their parents, M7 mutant lines developed by gamma irradiation, and some genotypes from different geographical origins, were evaluated against eight S. sclerotiorum isolates in controlled conditions. The proportion of the subsequent basal stem lesions was measured 3 days after inoculation. Highly significant differences were observed among sunflower genotypes and S. sclerotiorum isolates, with the isolates interacting differentially with sunflower genotypes. Two genotypes had high partial resistance to all S. sclerotiorum isolates, whereas others were susceptible to all isolates. Isolates of S. sclerotiorum differed in virulence to host genotypes. Some genotypes showed specific interactions with S. sclerotiorum isolates, being resistant to some isolates but susceptible to others. Recom­binant inbred lines used in this study showed different reactions to eight isolates of S. sclerotiorum when compared with their parental lines. The isolate-specific and isolate-nonspecific partial resistant genotypes identified in present experiments should be used in crossing programmes for breeding of durable resistance to Sclerotinia basal stem disease

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

Allelopathy of sage and white wormwood on purslane germination and seedling growth.

A bioassay run was carried out in the incubator to evaluate possible allelopathic effects of water extracts of sage and white wormwood on germination and seedling growth of purslane. Results showed that the type of extract and its concentration (0, 5, 10, 15 and 20%) significantly influenced final germination percentage, germination rate and index, root and shoot length, root/shoot ratio, fresh and dry weight of seedling of purslane. The interaction between these two experimental factors was always significant, producing different results according to the different combination levels. The statistical comparison of means indicated that the maximum germination percentage and germination rate (respectively 68% and 11.4% d-1) were obtained from the untreated control (0% extract), while the lowest values for the same two characters occurred with 15% of sage (respectively 45% and 6.4% d-1) and 20% of white wormwood (38% and 7.5% d-1). Concerning germination indexes, the highest (55) and the lowest (32) values were observed respectively on the control and with 15% of sage extract or 20% of white wormwood extract. The longest (4.5 cm) and the shortest (1.3 cm) root of purslane were obtained with 5% of white wormwood and 20% of sage extracts, respectively. Changes in shoot length with sage and white wormwood extracts were similar to those in root length, even though shoot length was less affected by the concentration of extracts. Root and shoot length changes brought to maximum (22.0) and minimum (8.9) values for the root/shoot length ratio, respectively with 5% of white wormwood and 20% of sage extracts. The maximum fresh (2.111 g) and dry (0.338 g) seedling weight of purslane were obtained from untreated control, producing the same seedling weight with 5% of sage, 5 and 10% of white wormwood extract. While the minimum fresh (0.692 g) and dry (0.111 g) seedling weight were obtained from 15% of white wormwood extract, so that there is no significant differences between this value and the seedling weight produced under 10, 15 and 20% of sage and 20% of white wormwood extract

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

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

Diallel Analysis of Yield and It’s related Traits in Sunflower (Helianthus annuus L.) under Well-watered and Water-stressed Conditions

Drought stress is one of the factors which influence sunflower production. Hence, breeding for tolerance to drought stress has become a major focus. In this paper, combining ability, gene action and genetic analysis of several characteristics were studied for five sunflower inbred lines and their ten hybrids (fifteen genotypes). The materials were evaluated in two separate experiments using a Randomized Complete Block Design (RCBD) with three replications in well-watered and waterstressed states and in field condition. Data were analyzed by Griffing’s diallel analysis model 1 (fixed effects) and method 2 (parents and crosses). Combined analysis of experiments revealed significant differences among genotypes for all studied traits. Combining ability analysis revealed that in well-watered conditions, general and specific combining abilities (GCA and SCA) effects were significant for all studied traits except for the leaf number. In water-stressed condition, GCA and SCA effects were significant for all studied traits except for the head and stem diameter. The Griffing diallel analysis showed that the genotype LR4 could be considered as the best combiner with the highest GCA effect for seed yield per plant, number of filled seed, stem diameter and head diameter in the sunflower breeding programs. In both water treatment conditions the cross ‘RHA266xLR4’ showed the highest positive SCA effects and the highest mean value for seed yield per plant

The resistance response of sunflower genotypes to black stem disease under controlled conditions

Phoma black stem, caused by Phoma macdonaldii, is one of the most important diseases of sunflower in the world. The sources of resistance to Phoma black stem were investigated. A total of 184 genotypes, including some recombinant inbred lines (RILs), several M6 mutant lines obtained by gamma irradiation of seed of the genotype AS 613, and other genotypes from different countries, were evaluated against an aggressive French isolate (MP6) in controlled conditions. The study was carried out in a randomized complete block design with three replicates. Each replicate consisted of 10–12 seedlings. Twenty μL of spore suspension (106 pycnidiospores mL-1) were deposited on the intersection of the cotyledon petiole and the hypocotyl of sunflower plantlets at the two-leaf stage. The percentage of the area exhibiting disease symptoms was scored on the two cotyledon petioles of each of the plantlets three, five and seven days after inoculation. The disease progress rate (rd), as the slope of the regression line for disease severity against time, was also calculated. Analysis of variance detected significant differences among sunflower genotypes for disease severity 7 days after inoculation,as well as for the disease progress rate. A strong correlation (r=0.96, P<0.01) was found between disease severity 7 days after inoculation and the disease progress rate. The inbred lines F1250/03 (origin: Hungary), M5-54-1, M6-862-1 (mutant lines), SDR 18 (origin: USA) and two wild Helianthus accessions, 1012 Nebraska and 211 Illinois, (wild type) were highly resistant to Phoma black stem. These findings will assist breeders in choosing parent plants for breeding durable resistance to Phoma black stem