Volume 37, Issue 61 -

Broomrape ( Orobanche cumana Wallr.) is a parasitic weed that causes economic damage in sunflower production in many countries, especially in Central and Eastern Europe, Spain, Turkey, Israel, Iran, Kazakhstan, and China. Genes for resistance to broomrape races A, B, C, D, and E are present in varietal populations of cultivated sunflower. Since broomrape is a highly variable parasitic weed, the breakdown of resistance is a frequent phenomenon, and multiple sources of resistance are needed to control the emerging races. Genes that confer resistance to races F, G, and H and others that have not been assigned a race designation have been identified in wild sunflower species and incorporated into hybrid sunflower through interspecific hybridization. The U.S. Department of Agriculture, Agricultural Research Service, National Plant Germplasm System wild sunflower collection contains 2,239 accessions with 1373 annual accessions represented by 14 species and 866 perennial accessions represented by 39 species. Sunflower germplasm evaluations for resistance to broomrape races have demonstrated that the Helianthus species constitute a substantial reservoir of genes conferring resistance to new virulence broomrape races. The resistance to broomrape, including immunity reported in seven annual and 32 perennial species, provides breeders a broad genetic base from which to search for resistance to existing and newly emerging races.

Wild Helianthus species possess valuable resistance genes for sunflower broomrape ( Orobanche cumana Wallr.), especially the 39 largely underutilized perennial species. Resistance to race F has been transferred into a cultivated background via bridging of interspecific amphiploids. More recently, a single dominant gene resistant to race G was identified in annual H. debilis ssp. tardiflorus and transferred into cultivated HA 89. Interspecific crosses between wild annual Helianthus species and cultivated lines are relatively easy compared to those involving wild perennial species, which were made easier only after the development of embryo rescue techniques. Interspecific amphiploids resulting from colchicine treatment of F 1 hybrids provide bridging materials for transferring genes without relying on embryo rescue. Among the diploid, tetraploid, and hexaploid perennial species, the speed of gene utilization follows the ploidy level of diploids, tetraploids, and hexaploids due to the time-consuming backcrosses required to eliminate the extra chromosomes in the latter two groups. In the development of pre-breeding materials, the retention rate of genetic material of the wild species is another concern with each additional backcross. For crosses involving tetraploid and hexaploid wild perennials, the use of 2 n =51 chromosome F 1 or BC 1 F 1 generation, as pollen source, could accelerate chromosome reduction to 2 n =34 in BC 1 F 1 or BC 2 F 1 , resulting in useful materials with fewer backcrosses for trait selection.

Much research has been conducted to identify sources of genetic resistance to sunflower broomrape ( Orobanche cumana Wallr.) and to study their mode of inheritance. However, studies on the parasite have been scarce. This manuscript reviews three genetic studies in sunflower broomrape. First, the inheritance of the absence of pigmentation in a natural mutant of this species with yellow plant color phenotype was studied. In a first stage, lines from the unpigmented mutant and a normally pigmented population were developed by several generations of self-pollination. Plants of both lines were crossed and the F 1 , F 2 , and F 3 generations were evaluated. The results indicated that plant pigmentation is controlled by a partially dominant allele at a single locus. Second, the unpigmented mutant was used to evaluate outcrossing potential of the species. Two experiments in which single unpigmented plants were surrounded by normally pigmented plants were conducted under pot and field conditions. The cross-fertilization rate was estimated as the percentage of F 1 hybrids in the progenies of unpigmented plants, which averaged 21.5% in the pot and 28.8% in the field experiment. The results indicated that, under the conditions of this study, the species was not strictly self-pollinated. Finally, the inheritance of avirulence was studied in crosses of plants from lines of O. cumana races E and F, developed by several generations of self-pollination. The F 1 and F 3 generations were evaluated on the differential line P-1380 carrying the race-E resistance gene Or 5 . The results suggested that race E avirulence and race F virulence on P-1380 are allelic and controlled by a single locus, which confirmed the gene-for-gene theory for the O. cumana –sunflower interaction.

Orobanche cumana is found in the Iberian Peninsula as an allochthonous species parasitizing exclusively sunflower, in contrast to the closely related species Orobanche cernua , which is an autochthonous species that only parasitizes wild Asteraceae hosts. Ten O. cumana populations were collected in the two traditional areas of sunflower broomrape occurrence, the Guadalquivir Valley, Southern Spain (six populations) and Cuenca province, Central Spain (four populations). Twelve O. cernua populations were collected on wild hosts across its natural distribution area in Southeastern Spain. Genetic relationships within and between both sets of populations were studied using a set of 50 robust and co-dominant SSR markers from O. cumana . The results supported the taxonomic separation of the two species and the existence of two distant genetic groups for O. cumana , one in Guadalquivir Valley and another one in Cuenca province. The inter- and intra-population variability was extremely low for O. cumana , whereas the overall genetic diversity was much higher for O. cernua . The genetic structure of O. cumana populations probably reflects a founder effect, with the two genetically distant groups deriving from separate introduction events. The high degree of genetic differentiation observed in O. cernua is mainly explained on the basis of restricted gene flow due to ecological barriers together with the occurrence of a predominantly self-pollinating mating system. Complementary diversity studies on both species in its current distribution area are required for understanding global genetic variability and evolutionary characteristics of the parasitism.

Orobanche cumana is an holoparasite and thus totally dependent on sunflower for fixed carbon. Initial stages of the infection occur in the first weeks after sowing and are critical for the establishment of a continuum between the host and the parasite vascular system. From that moment the parasite obtains its supply of water, mineral nutrients, and assimilates from the host plant. Alterations of plant metabolism can be detected using remote sensing techniques for detection of fluorescence emitted by plants. One of these indirect techniques is multicolour fluorescence imaging. In this work, we assessed the early infection of sunflower by O. cumana using multicolour fluorescence imaging and we inferred physiological processes affected in sunflower plants infected by the parasite. Ten germinated seeds of the inbred line NR5 were inoculated with population LP2013 of O. cumana . The same number of not inoculated seeds was used as control. Sunflower was planted in pots with soil mixture and grown in greenhouse at 12–22°C for 6 weeks. Multicolour fluorescence imaging was conducted 3, 4, and 5 weeks after inoculation. The two first pairs of fully expanded leaves of each sunflower plant were imaged, and, for each measure date, five fluorescence variables in inoculated plants were compared to those in the control. Three weeks after inoculation, when symptoms of infection were still not observed, decreased levels of blue and green fluorescence and increased far-red fluorescence were observed in leaves of the inoculated plants. At 4 and 5 weeks after inoculation, when inoculated plants displayed symptoms of infection by O. cumana , differences in fluorescence between inoculated plants and the controls were the same and statistically supported. These results are consistent with an increase in total chlorophyll content of sunflower plants infected by O. cumana , and a decrease in the accumulation of secondary metabolites, both related to the need of higher photosynthetic activity to supply the parasite with photosynthate. Biochemical mechanisms underlying alterations in photosynthesis must be further investigated. The results obtained showed that multicolour fluorescence imaging can be used to detect fluorescence differences in inoculated sunflower as early as 3 weeks after inoculation. Therefore, this technique can be used as a diagnostic tool for early detection of genotypes of sunflower which are susceptible or resistant to O. cumana .

Broomrape ( Orobanche cumana Wallr.) is an obligate parasite of higher plants, which affects sunflower in many countries, cultivating this crop. For the past decades, it is noted the formation of new highly virulent biotypes of broomrape and their spreading to other areas. In our work we studied the molecular genetic diversity of broomrape populations of O.cumana , parasitizing on sunflower in Russia, Romania, and Kazakhstan, by using codominant microsatellite markers. During cluster analysis, the broomrape populations are divided into two clusters, regardless of their racial composition. One cluster grouped 19 samples from Russia and Kazakhstan, and the other – 5 populations from Romania. The genetic distance between clusters according to Nei was 0.137. The analysis of molecular variance (AMOVA) revealed that 22% of genetic variability was due to differences among the gene pools and 78% was due to differences within the gene pools. Pairwise comparisons made using Wright’s statistics showed that the differences between these two gene pools are sufficient (Fst = 0.219) to state the existence of a small genetic differentiation between them. Descriptive population genetic statistics for each of the two pools showed that the broomrape populations from the former Soviet Union countries are characterized by a higher level of intrapopulation diversity than the populations from Romania. Molecular genetic differences between broomrape populations parasitizing on sunflower on the post-Soviet territory and in Romania were insignificant. Possible reasons for these results are being discussed.

Sclerotinia sclerotiorum (Lib.) de Bary is a yield-limiting factor and the major disease of sunflower ( Helianthus annuus L.) in the temperate regions of the world. In this study, we characterized resistance to S . sclerotiorum midstalk rot and morphological traits in a population derived from a cross between two Sclerotinia resistant lines. Phenotypic data for 114 F 3:4 lines and the two parents, NDBLOS sel and K04, were obtained under artificial infection in field experiments which yielded moderate to high heritabilities. Our results suggest that S. sclerotiorum resistance is highly quantitative and that different genomic regions may mediate the resistance in different tissues of the plant. We found transgressive segregation for all three resistance traits suggesting that both resistant parents carried complementary QTL. In addition, we investigated the segregation of two known QTL for midstalk resistance and found that one of them also acts as a major QTL in this cross between two resistant lines. Collectively, our results suggest that a QTL stacking approach is a promising way to increase resistance to S. sclerotiorum in elite sunflower germplasm.

Verticillium leaf mottle and wilt caused by the fungus Verticillium dahliae is a major disease of sunflower in Argentina and the USA. In the summer of 2013, an important outbreak of the disease occurred in one field in the Southwest of Spain. The determination of vegetative compatibility groups (VCGs) of V. dahliae is used in the genetic characterization of the fungus. Molecular markers have also been developed and applied for the analysis of V. dahliae , particularly for those isolates infecting artichoke, cotton and olive tree. The objective of this work was to determine the genetic and molecular features of V. dahliae of sunflower. Three V. dahliae isolates, one from Argentina collected in 2012 and two collected in Spain in 2013, were analysed. The VCGs were determined by complementation between nit mutants of the isolates from sunflower and the VCGs reference strains. Molecular characterization was conducted by polymerase chain reaction using primer pairs that are diagnostic of either V. dahliae species, defoliating or non-defoliating pathotypes, or VCGs. Complementation tests between nit mutants and reference strains clearly showed that the three isolates from sunflower belong to the VCG2B. The VCG2B has been identified in V. dahliae from crops as cotton, artichoke, eggplant, pepper and tomato among others. When molecularly analysed, the three V. dahliae isolates infecting sunflower had the same molecular pattern than the one found for non-defoliating isolates of V. dahliae pathogenic to artichoke or cotton. The results of this work show the closeness between non-defoliating isolates of V. dahliae infecting artichoke, cotton and sunflower and suggest that any of these three species can serve as carrier and source of inoculum for Verticillium outbreaks in them.

Currently, Orobanche cumana in the Russian Federation overcomes the influence of dominant genes of resistance in sunflower Or4 , Or5 , Or6 , Or7 , already known in European countries, and the combined impact of the two recessive genes or6or7 . The most virulent biotypes of parasite G and H are found in many regions of sunflower cultivation: the Rostov, Voronezh, Volgograd, Saratov, Orenburg, Stavropol and Krasnodar regions. The situation is especially unfavorable in the Rostov region where the race G became predominant in many populations of O. cumana . In the Krasnodar region broomrape began to spread in recent years (after a long absence) mainly in the northern regions (and adjacent), bordering with the Rostov region. Here the populations of O. cumana are a mixture of races of different virulence. The races D and E still often dominate. However, there are already the parasite populations, where the race G is predominant, and somewhere biotype H is already present. In a sample of seeds from Kazakhstan is dominating the low virulent race C, among which there is a small admixture of biotype G. This combination of low virulent race with a small amount of highly virulent specimens of biotype G indicates the natural origin of the latter, regardless of the influence of the breeding process of sunflower. Some deviations from the basic model of development of O. cumana in ontogenesis that contribute to increase and acceleration of seed preproduction of parasite’s specimens are described.

The genetic structure of F 2 sporophytic populations after F 1 sunflower pollen storage at low temperature has been studied. Freshly collected pollen was stored at the temperature of 3±1°C for a period of 7–8 days and used to self-pollinate the emasculated F 1 plants. F 2 seeds were germinated at 5±1°C, and then the percentage of seed germination was counted. Germinated and not germinated seeds were separately planted in the phytotron at an optimum temperature. Segregation ratios in F 2 populations for marker traits were analyzed at the stage of the second pair of true leaves. Pollen treatment compared with the control (fresh pollen) significantly changed in F 2 populations monogenic ratios for some marker traits. In some cross combinations, increase in the cold tolerance of F 2 populations was found. Obtained results show that pollen storage at low temperature selectively influences the male gametophytes of F 1 hybrids that change the genetic structure of F 2 populations.

Two field experiments were conducted on the Teaching and Research Farm of the Federal University of Agriculture, Abeokuta (7° 15′ N, 3° 25′ E, altitude 140 m above sea level) in south western Nigeria between June and November, 2012 to evaluate the performance of two sunflower varieties (SAMSUN-3 and Funtua) as affected by different sowing dates and organic fertilizer application. The experiment was laid out in split–split plot design with sowing dates as main plot, organic fertilizer as sub-plot and variety as sub-sub-plot, and replicated three times. Sowing date significantly ( P ≤0.05) increased and reduced number of phenological days to flowering of early and late sown sunflower, respectively. Funtua (a local adapted variety) matured 7 days later than SAMSUN-3 (recently released variety) when sown late. Application of organic fertilizer significantly ( P ≤0.05) increased plant height of early and late sown sunflower. Delay in sowing till first week in September significantly ( P ≤0.05) reduced head diameter, number of seeds per head and seed yield of late sown sunflower. Oil and protein contents of seeds from early and late sown sunflower plants were significantly ( P ≤0.05) increased by organic fertilizer application, except protein content of late sown sunflower. However, it significantly ( P ≤0.05) increased oil yield by 23% relative to the control in the late sown sunflower. Based on the comparatively high seed yield (850.45–1,525.78 kg/ha) recorded in our study, it is recommended that sunflower be sown in the forest–savanna transition zone in early July or second to third week in August.