Inheritance of resistance to sunflower downy mildew races 1, 2 and 3 in cultivated sunflower

Sunflower downy mildew (SDM) caused by Plasmopara halstedii, is a major disease of sunflower. Eleven resistance genes have been identified, but allelic relationships among these genes are not clear. This study examined the inheritance and allelic relationships of genes conferring resistance to SDM races 1, 2 and 3 (virulence phenotypes 100, 300 and 700, respectively) and confirmed a twelfth resistance gene. Three USDA Plant Introductions, AMES 3235, PI 497250, and PI 497938, and three released lines, RHA 266, RHA 274 and DM‐2 were studied. RHA 266 has only the Pl₁ gene for race 1 resistance. Digenic inheritance of resistance was found in AMES 3235, PI 497250, and RHA 274. These lines have the Pl₁ and Pl₁₂ genes, conferring resistance to race 1, and the Pl₂ and Pl₁₁ genes, conferring resistance to race 2. DM‐2 and PI 497938 have Pl₁₂ (but not Pl₁ for resistance to race 1, the Pl₁₂ gene (but not the Pl₂) for resistance to race 2, and Pl₅ for resistance to race 3. These resistance genes will serve as a foundation for future gene designations and genetic diversity studies of resistance to SDM.     

Development of co-dominant amplified polymorphic sequence markers for resistance of sunflower to downy mildew race 730

The inheritance of the reaction of sunflower to downy mildew was investigated using resistant and susceptible near isogenic lines (NILs) and their F₃ families. Resistance to race 730 was evaluated using the whole seedling inoculation technique. Seventy-three F₃ families were inoculated, among which 54 families were resistant and 19 susceptible, fitting a 3 : 1 segregation ratio. F₃ families were also studied using several PCR markers. Ten markers at the Pl6 locus, specific for the resistant line, also segregated in F₃ families with a 3 : 1 ratio. The same segregation ratio occurred for microsatellite haplotypes that resembled the resistant parent, and were amplified with ORS 166 and ORS 1043. The only common fragment that was observed between resistant and susceptible parental lines was one of the TIR-NBS-LRR resistance gene analogue markers, having a restriction site. Two co-dominant cleaved amplified polymorphic sequence (CAPS) markers were obtained. The mapping data indicate that several dominant markers and two CAPS markers, developed here, completely co-segregate with the Pl6 gene conferring resistance to race 730. CAPS markers will facilitate efficient marker-assisted selection for sunflower resistance to downy mildew race 730.     

Inheritance of resistance to two races of sunflower downy mildew (Plasmopara halstedii) in two Helianthus annuus L. lines

Sunflower downy mildew caused by Plasmopara halstedii is an important disease of sunflower capable of causing losses of more than 80% of production. Races 100, 300, 310, 330, 710, 703, 730 and770 of the fungus have been identified in Spain. Race 703, of high virulence, has been identified frequently in the northeast, while race 310 seems to occur over the south, the main sunflower growing region of the country. Oil sunflower lines RHA-274 and DM4 were studied for their resistance to races 310(RHA-274 and DM4) and 703 (DM4). In each cross, only one plant of the resistant parent was crossed to the inbred susceptible line HA-89 (or cmsHA-89).Plants from F₂ and backcross(BC₁F₁ to susceptible parent)generations were evaluated for fungal sporulation on true leaves and/or cotyledons. The resistant-to-susceptible ratios obtained in the F₂ and BC₁F₁ progenies from the crosses cmsHA-89 × RHA-274 and HA-89 × DM4suggested that one major gene in each line is responsible for resistance to race 703.The segregations of the progenies of the cross HA-89 × DM4 inoculated with race 703also fitted the ratios 1:1 and 3:1 (for BC₁F₁ and F₂, respectively)corresponding to control of resistance by a single dominant gene. In RHA-274, the gene for resistance to race 310 was designated Pl ₉, whereas Pl _v is tentatively proposed to designate the gene in DM4 responsible for resistance to races310 and 703. This revised version was published online in August 2006 with corrections to the Cover Date.     

Colocation of downy mildew (Plasmopara halstedii) resistance genes in sunflower (Helianthus annuus L.)

Summary The Pl6 locus in the inbred sunflower (Helianthus annuus L.) line HA335 giving resistance to French races of downy mildew (Plasmopara halstedii (Farl.) Berl. & de Toni. was localized by molecular techniques. A bulked segregant analysis was made on the F2 progeny from a cross between this line and H52, a downy mildew susceptible line. The resistance gene in HA335 was found to have the same linked RFLP marker loci as those determined for Pl1 (resistance to race 1 in the line RHA266) on linkage group 1 of the consensus RFLP map of the cultivated sunflower. Pl1 and Pl6 thus appear either to be allelic or closely linked. The implications for sunflower breeding are discussed.     

Inheritance of resistance to race F of broomrape in sunflower lines of different origins

A new race F of broomrape overcomes all known resistance genes in cultivated sunflower, but recently, sources of resistance against race F have been developed. The objective of the present research was to study the inheritance of resistance to race F in crosses between 12 resistant sunflower breeding lines, derived from three different sources of resistance, and the susceptible male‐sterile line P‐21. Parental lines and F₁, F₂, F₃ and BC₁ generations were evaluated for broomrape resistance. Segregations in the F₂ and BC₁ to resistant parent approached resistant to susceptible ratios of 1: 15 and 1: 3, respectively, in most of the crosses, suggesting a double dominant epistasis. However, segregations of 3: 13 and 1: 1 for F₂ and BC₁, respectively, indicating a dominant‐recessive epistasis, were also found. The F₃ data confirmed these results. Owing to the recessive nature of this resistance, it must be incorporated into both parental lines for developing resistant hybrid cultivars.     

Inheritance of quantitative resistance to downy mildew (Plasmopara halstedii) in sunflower (Helianthus annuus L.)

Quantitative resistance to sunflower downy mildew was studied on inbred lines and hybrids not carrying efficient major gene resistance, in field trials in one to four sites over 3 years. Hybrids from factorial crosses showed that inheritance is under additive control and comparison with reactions of parental inbred lines gave narrow sense heritabilities of 27–57%. Analysis of a polymorphic recombinant inbred line population without efficient major gene resistance indicated that two highly significant Quantitative Trait Loci (QTL) explained 42% of variation in field reaction to downy mildew. These QTL were mapped on linkage groups 8 and 10, and do not appear related to any of the known major resistance gene clusters. Possible bases of this type of resistance and its use in breeding are discussed.     

Genetic control of in vitro-organogenesis in recombinant inbred lines of sunflower (Helianthus annuus L.)

Ninety-three recombinant inbred lines (F₈) of sunflower developed by the single-seed descent method from the cross ‘PAC-2 × RHA-266’ were used to screen their regenerability by organogenesis. The experiment was designed in randomized complete blocks with three replications. Each replication consisted of 10 Petri dishes with four explants. Cotyledons were excised from 2-day-old seedlings. Each cotyledon was divided into two pieces (four explants), which were incubated in solid regeneration medium consisting of full-strength Murashige and Skoog medium modified by adding hormones. A high genotypic variability for organogenesis parameters between genotypes was observed in this study. The difference between all recombinant F₈ lines and their parents was not significant, showing that the 93 inbred lines used in this experiment are representative of the total possible recombinant lines from the cross ‘PAC-2 × RHA-266′. These F₈ lines were not consciously selected for any trait; therefore, they represent a random set of lines segregating for the organogenesis parameters, as well as for other traits which could be important for breeding.     

Inheritance of resistance to broomrape (Orobanche cumana Wallr.) race F in a sunflower line derived from wild sunflower species

Genetic resistance to broomrape (Orobanche cumana Wallr.) race F in sunflower line J1, derived from the wild perennial species Helianthusgrosseserratus Martens and Helianthus divaricatus L., has been reported to be controlled by dominant alleles at a single locus, Or6. However, deviations from this monogenic inheritance have been observed. The objective of the present study was to gain insight into the inheritance of resistance to broomrape race F in the sunflower line J1. F₁, F₂, F₃ and BC generations from crosses between J1 and three susceptible lines, P21, NR5 and HA821 were evaluated. F₁ hybrids showed both resistant (R) and moderately resistant (MR) plants, the latter having a maximum of five broomrape stalks per plant compared with >10 in the susceptible parents. This indicated incomplete dominance of the Or6 alleles. F₂ plants were classified as R, MR or susceptible (more than five broomrape stalks per plant). Three different segregation ratios were observed: 3 : 1, 13 : 3 and 15 : 1 (R + MR : S), suggesting the presence of a second gene, Or7, whose expression was influenced by the environment. A digenic model was confirmed, based on the evaluation of F_2:3 families.     

Inheritance of resistance to phomopsis (Diaporthe helianthi) in sunflower

The inheritance of resistance of sunflower to phomopsis (Diaporthe helianthi) was studied on 10 inbred lines representing a range of resistance and susceptibility levels, and 25 hybrids from a factorial cross of these lines. Analyses of observations over a 3-year period, of semi-natural infection (presence of stem lesions greater than 5 cm) on the hybrids indicated that additive gene control was predominant, with no significant interactions between parental effects. Correlations between infection of parental lines and the mean of their hybrids were not always significant, with certain inbreds which appeared quite susceptible giving hybrids with good levels of resistance. In order to predict hybrid values, it appears necessary to determine the general combining abilities of parental lines. Some inbreds gave very high levels of resistance, others gave very high levels of susceptibility. A test measuring the rate of extension of D. helianthi mycelium on leaves was significantly correlated with the results of natural infections. In particular, it permitted distinction of the most susceptible genotypes, and thus could be used in first generations of breeding to eliminate the most susceptible plants.     

Resistance to Sclerotinia sclerotiorum of 'high oleic' sunflower inbred lines

Cultivation of sunflower (Helianthus annuus L.) is strongly affected by Sclerotinia sclerotiorum. To identify new sources of genetic diversity for sunflower breeding 25 sunflower inbred lines were analysed using eight Amplified Fragment Length Polymorphism (AFLP) primer combinations and their genetic similarities (GS) were estimated. Data were used to develop a Unweighted Pair Group Method using Arithmetic Averages (UPGMA) dendrogram. GS values of 0.58‐0.98 were observed but with no separate groupings dependant on oil quality. The inbred lines were screened for their reaction to inoculation with Sclerotinia sclerotiorum (Lib.) de Bary. Sunflower heads were artificially inoculated with S. sclerotiorum in three environments. Head infection was monitored after 1 week (lesion length) and 2 weeks (head rot). The F5 generation of a cross between a resistant (SWS‐B‐04) and a susceptible inbred line (SWS‐B‐01) was also tested for sclerotinia reaction across three environments. Significant differences in sclerotinia resistance, moderate heritabilities and a high correlation between the two assessments were observed. Inbred lines with a high level of resistance could be identified. These lines can be used for further breeding to improve sunflower sclerotinia resistance and to develop superior new hybrids.     

Quantitative resistance to downy mildew (Plasmopara halstedii) in sunflower (Helianthus annuus)

Downy mildew of sunflower, caused by the Oomycete, Plasmopara halstedii is at present controlled by major resistance genes. However, the pathogen has shown a considerable capacity for changes in virulence and these resistance genes are overcome only a few years after they have been introduced into new sunflower varieties. This paper presents research for quantitative, non-race-specific resistance independent of major genes. The reaction of cultivated sunflower genotypes to field attack by downy mildew was studied over 4 years in several environments and in the presence of the two most common races in France: 703 and 710. An experimental protocol with pre-emergence irrigation was developed, making it possible to observe downy mildew reaction whatever the weather conditions. Significant levels of partial resistance were observed in about 50 inbred sunflower lines among the 800 observed. These results suggest that it should be possible to select for non-race-specific downy mildew resistance and to include it in modern varieties. However, since this non-specific resistance is partial, it may be necessary to combine it with major gene resistance. Possible strategies are discussed to obtain durable resistance to downy mildew.     

Screening for resistance to broomrape (Orobanche cernua) in cultivated sunflower

Racial evolution of sunflower broomrape (Orobanche cernua) has been very rapid in Spain during recent years, in which resistance has been overcome several times and there has been an important increase in areas infested with this parasitic angiosperm. In order to find resistance to a highly virulent population of sunflower broomrape that could be used directly in breeding programmes, three different sets of cultivated plant material composed of 429 entries were tested by artificial inoculation. All evaluated inbred lines from Moden, Canada, were fully susceptible. Out of the 240 P.I. accessions tested, only 10 segregated for resistance to broomrape, the rest being susceptible. From the 160 USDA breeding lines evaluated, 5% were resistant and 19% segregated for resistance to O. cernua. These lines traced back mainly to crosses of RHA 274 and RHA 801 with Russian, Turkish and Romanian hybrids. The origin of P.I. accessions that segregated for resistance were primarily derived from the former USSR and from Romania.     

Inheritance and linkage of a gene for resistance to race 4 of fusarium wilt and RAPD markers in chickpea

Several races of Fusarium oxysporum Schlechtend.:Fr f. sp. ciceris (Padwick) Matuo and K. Sato cause economic losses from wilting disease of chickpea ( Cicer arietinum L.). While the genetics of resistance to race 1 have been reported, little is known of the genetics of resistance to race 4. We undertook a study to determine the inheritance of resistance and identified random amplified polymorphic DNA markers (RAPDs) linked to the gene for resistance. For the investigation, we used 100 F₅ derived F₇ recombinant inbred lines (RILs) that had been developed from the cross of breeding lines C-104 x WR-315. Results indicated that resistance is controlled by a single recessive gene. The RAPD markers previously shown to amplify fragments linked to race 1 resistance also amplified fragments associated with race 4 resistance. The RAPD loci, CS-27₇₀₀, UBC-170₅₅₀ and the gene for resistance to race 4 segregated in 1:1 ratios expected for single genes. Both RAPD markers were located 9 map units from the race 4 resistance locus and were on the same side of the resistance gene. Our results indicated that the genes for resistance to race 1 and 4 are 5 map units apart. The need to determine the genomic locations of race specific resistance genes and the possibility that these genes are clustered to the same genomic region should be investigated. This revised version was published online in July 2006 with corrections to the Cover Date.     

Inheritance of reduced plant height in the sunflower line Dw 89

The objective of the present research was to study the inheritance of reduced plant height in the sunflower line Dw 89. Plants of the cytoplasmic male sterile version of this line, cmsDw 89 (mean plant height of 47.4 cm) were crossed with plants of the restorer line RHA 271 (mean of 120.9 cm). F₁ plants averaged 120.4 cm, which indicated dominance of standard over reduced plant height. F₂ plants followed a segregation pattern of 1 : 15 (reduced : normal height), suggesting that reduced plant height in Dw 89 is controlled by alleles at two loci, designated Dw1 and Dw2. Class assignment in the F₂ was confirmed through the evaluation of the F₃ generation. Backcrosses to Dw 89 segregated with 1 : 3 (reduced : normal height) ratios, which confirmed the digenic inheritance of the trait. The evaluation of plant height distributions in F₃ families suggested possible genetic interaction between the Dw1 and Dw2 loci.     

Optimal allocation of resources in evaluating current sunflower inbred lines for resistance to Sclerotinia

Sclerotinia sclerotiorum (Lib.) de Bary is a major limiting factor for sunflower (Helianthus annuus) production. This study was conducted to (1) evaluate a large number of current sunflower lines for their reaction to Sclerotinia, and (2) determine the optimum allocation of resources for their selection. A set of 85 inbred lines was screened under artificial leaf infection in three environments. The test revealed ample variation for all resistance traits in the current germplasm. A line selected from NDBLOS, an oilseed sunflower germplasm poo, and its derivatives were most resistant to Sclerotinia. Lines derived from interspecific crosses with the wild species Helianthus tuberosus and Helianthus argophyllus also showed satisfactory Sclerotinia resistance. The evaluation of five plants per plot in two replications and four environments appears to be a reasonable allocation of resources in order to optimize the selection response.     

Genetic variability of tocopherol composition in sunflower seeds as a basis of breeding for improved oil quality

The variability of seed tocopherol content in wild sunflower species, the expressivity of tph1 and tph2 mutations in different lines and the oxidative stability of sunflower oil with altered tocopherol and fatty acid composition were objectives of this research. Near-isogenic lines for three genes, i.e. Tph1, Tph2, and Ol, were developed and investigated. Tocopherol content was determined with TLC and HPLC, as well as fatty acid composition with GC of methyl esters. Rancimat tests were used to estimate the oxidative stability of the oil. The seed tocopherol composition of wild sunflower species was shown to be uniform with a prevailing content of the α-homologue (90-99%). The genetic background of different near-isogenic lines was found to influence expressivity of mutations for tocopherol composition. High content of strong antioxidants, such as β-, γ-, and δ-tocopherols increased oil oxidative stability of linoleic and oleic types of oil by 1.2–3.0 times. The breeding model of sunflower hybrids should include antioxidant and vitamin parameters balanced for oils of different applications.     

Reproductive behaviour and broomrape resistance in interspecific hybrids of sunflower

Interspecific hybrids and backcross generations between the wild perennial species Helianthus resinosus, Helianthus paucifiorus, Helianthus laevigatus, Helianthus nuttallii ssp. nuttallii T. & G. and Helianthus giganteus, resistant to broomrape (Orobanche cernua) and susceptible inbred lines were obtained to study crossability to cultivated sunflower and the transmission and expression of resistance to this parasitic weed. Conventional crosses with all the species tested were successful except for the crosses with diploid H. giganteus, for which embryo rescue techniques were needed to overcome hybrid incompatibility. Pollen viability and seed set were highest for F₁ hybrids with hexaploid species and lowest for those with the diploid H. giganteus. We evaluated F₁, BC₁F₁, some BC₂F₁ plants and the wild and cultivated parents. The wild species and interspecific hybrids were resistant to broomrape infection except for H. nuttallii, which showed segregation, indicating that the resistance is dominant. The crossability and resistance of F₁, and back-cross generations of species with different ploidy levels indicate that the transfer of broomrape resistance to cultivated sunflower is feasible.     

Inheritance and allelic relationships of fertility restoration genes for seven new sources of male-sterile cytoplasm in sunflower

The inheritance of fertility restoration of six mitomycin C and streptomycin‐induced cytoplasmic male‐sterile (cms) mutants and one cms line derived from Native American cultivar PI 432513 in sunflower was evaluated. These seven new cms sources were also compared with the commercially used cms PET1 (Helianthus petiolaris Nutt.) cytoplasm, using USDA inbred lines with restoration genes (Rf₁) specific for cms PET1 and new restoration lines identified for cms PI 432513. Restoration genes for cms PI 432513 were found in ‘Armavir’, VNIIMK, P21 and male‐fertile (MF) plants of PI 432513. F₂ and F₃ segregation ratios of crosses between cms PI 432513 and these restoration sources indicated a single dominant gene controlled fertility restoration. Progenies of cms PI 432513 testcrossed with F₁’s of half‐diallel crosses among the respective four homozygous restoration lines and RHA 274 suggested that the restoration genes of RHA 274, VNIIMK, P21 and PI 432513 were at the same locus. Restoration genes from VNIIMK, P21 and PI 432513 satisfactorily restored pollen stainability in the heterozygous condition. A very weak expression of the Rf gene in ‘Armavir’ was observed in the heterozygous condition. Fertility restoration capability of these genes for the six mutant cms HA 89 and cms HA 89 (in PET1 cytoplasm) was observed. The mutant cms HA 89 lines were also restored completely by RHA 266, RHA 274, RHA 280 and RHA 296, and F₂’s segregation ratios indicated single dominant gene control, implying a common cytoplasmic male sterility in all lines. F₁’s of half‐diallel crosses among RHA 266, RHA 273, RHA 274, RHA 280 and RHA 296 were testcrossed onto the cms lines, and their all MF progenies among lines, except RHA 280, confirmed that fertility restoration was controlled by a single Rf₁ gene locus. The restoration gene in confection line RHA 280, namely Rf₃, was at a different locus than Rf₁ and was equally capable of restoring all the cms lines. Cms HA 89 mutants and cms PI 432513 are in H. annuus cytoplasm, and are agronomically equal in hybrid performance to the cms PET1 used in commercial sunflower hybrids. These new cms lines will provide immediate alternative cms sources for reducing the genetic vulnerability resulting from the exclusive use of the single cms source PET1 in sunflower hybrid production.