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 F1, F2, and F3 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 F1 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 F1 and F3 generations were evaluated on the differential line P-1380 carrying the race-E resistance gene Or5. 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.The manuscript reviews research partially funded by
Fundación Ramón Areces, Madrid. The contribution of Dr. Enrique Quesada
Moraga, entomologist from the University of Córdoba, Spain, to taxonomic
classification of pollinators is gratefully acknowledged. R. Pineda-Martos was
the recipient of a PhD fellowship from the Spanish National Research Council
(CSIC) (JAEPre_08_00370)Peer Reviewe

Alonso, L. C.

Fernández Martínez, José María

Fernández-Escobar, J.

Pineda-Martos, Rocío

Pérez-Vich, Begoña

Rodríguez-Ojeda, M. I.

Velasco Varo, Leonardo

Digital.CSIC

Open AccessMaría I. Rodríguez-Ojeda, Rocío Pineda-Martos, Luis C. Alonso,José M. Fernández-Martínez, Leonardo Velasco*, JuanFernández-Escobar and Begoña Pérez-VichGenetic Studies in Sunflower BroomrapeAbstract: Much research has been conducted to identify sources of geneticresistance to sunflower broomrape (Orobanche cumana Wallr.) and to studytheir mode of inheritance. However, studies on the parasite have been scarce.This manuscript reviews three genetic studies in sunflower broomrape. First, theinheritance of the absence of pigmentation in a natural mutant of this specieswith yellow plant color phenotype was studied. In a first stage, lines from theunpigmented mutant and a normally pigmented population were developed byseveral generations of self-pollination. Plants of both lines were crossed and theF1, F2, and F3 generations were evaluated. The results indicated that plantpigmentation is controlled by a partially dominant allele at a single locus.Second, the unpigmented mutant was used to evaluate outcrossing potentialof the species. Two experiments in which single unpigmented plants weresurrounded by normally pigmented plants were conducted under pot and fieldconditions. The cross-fertilization rate was estimated as the percentage of F1hybrids in the progenies of unpigmented plants, which averaged 21.5% in thepot and 28.8% in the field experiment. The results indicated that, underthe conditions of this study, the species was not strictly self-pollinated.Finally, the inheritance of avirulence was studied in crosses of plants from*Corresponding author: Leonardo Velasco, Instituto de Agricultura Sostenible (IAS-CSIC),Alameda del Obispo s/n, 14004 Córdoba, Spain, E-mail: lvelasco@ias.csic.esMaría I. Rodríguez-Ojeda, Syngenta España S.A., Avenida San Francisco Javier, 24 7a planta,E-41018 Sevilla, Spain, E-mail: maribel.rodriguez@syngenta.comRocío Pineda-Martos, Instituto de Agricultura Sostenible (IAS-CSIC), Alameda del Obispo s/n,14004 Córdoba, Spain, E-mail: rpineda@ias.csic.esLuis C. Alonso, Syngenta España S.A., Avenida San Francisco Javier, 24 7a planta,E-41018 Sevilla, Spain, E-mail: luis_carlos.alonso@syngenta.comJosé M. Fernández-Martínez, Instituto de Agricultura Sostenible (IAS-CSIC), Alameda del Obispos/n, 14004 Córdoba, Spain, E-mail: jfernandezm@ias.csic.esJuan Fernández-Escobar, Syngenta España S.A., Avenida San Francisco Javier, 24 7a planta,E-41018 Sevilla, Spain, E-mail: juan.fernandez@syngenta.comBegoña Pérez-Vich, Instituto de Agricultura Sostenible (IAS-CSIC), Alameda del Obispo s/n,14004 Córdoba, Spain, E-mail: bperez@ias.csic.esHELIA 2014; 37(61): 151–159©2014, Rodríguez-Ojeda et al. This work is licensed under the Creative CommonsAttribution-NonCommercial-NoDerivatives 3.0 License.Brought to you by | CSIC - Unidad Coordinacion BibliotecasAuthenticatedDownload Date | 1/14/16 1:24 PMlines of O. cumana races E and F, developed by several generations ofself-pollination. The F1 and F3 generations were evaluated on the differentialline P-1380 carrying the race-E resistance gene Or5. The results suggestedthat race E avirulence and race F virulence on P-1380 are allelic and controlledby a single locus, which confirmed the gene-for-gene theory for the O. cumana–sunflower interaction.Key words: avirulence, cross-fertilization, genetic studies, hybridization, inheri-tance, virulenceDOI 10.1515/helia-2014-0038Received September 23, 2014; accepted October 13, 2014IntroductionThe O. cumana–sunflower parasitic system is characterized by a clear differen-tiation of physiological races of the parasite and a host–parasite interaction thatgenerally fits the gene-for-gene model. This contrasts with other crop–Orobanche associations, in which the genetic control of the avirulence of theparasite and the resistance of the crop plant are generally under quantitativegenetic control (Pérez-Vich et al., 2013). In recent years, increasingly virulentraces have appeared and spread into large areas of the main sunflower produ-cing countries in the Old World, jeopardizing sunflower production (Škorićet al., 2010; Antonova et al., 2013; Molinero-Ruiz et al., 2014). Understandingrace evolution in O. cumana requires basic studies on its mating system and theinheritance of avirulence/virulence mechanisms.The genus Orobanche encompasses species that are largely pollinated byinsects and species which are mainly self-pollinated (Musselman et al., 1981).Sunflower broomrape is considered a predominantly autogamous species onaccount of its flower morphology (Satovic et al., 2009) and population structure,generally characterized by marked among-population differences and low intra-population genetic diversity (Gagne et al., 1998; Pineda-Martos et al., 2013;Molinero-Ruiz et al., 2014).Even though other forms of genetic control have been reported, the occur-rence of resistance mechanisms to broomrape controlled by single dominantgenes is common in sunflower (Vrânceanu et al., 1980; Sukno et al., 1999;Velasco et al., 2012). A full demonstration of the occurrence of a gene-for-generelationship in the sunflower–O. cumana interaction requires studying theinheritance of avirulence in the parasite.152 M. I. Rodríguez-Ojeda et al.Brought to you by | CSIC - Unidad Coordinacion BibliotecasAuthenticatedDownload Date | 1/14/16 1:24 PMThis paper summarizes several recent genetic studies on O. cumana, inparticular the inheritance of a natural mutation resulting in absence of antho-cyanin pigmentation, the use of this mutation for estimating the rate of cross-pollination under experimental conditions, and the inheritance of avirulence incrosses between O. cumana plants of races E and F.Materials and methodsDetailed description of the materials and methods used in these genetic studiescan be found in the original papers reporting the inheritance of the unpigmentedplant trait (Rodríguez-Ojeda et al., 2011), the rate of cross-fertilization (Rodríguez-Ojeda et al., 2013a), and the inheritance of avirulence (Rodríguez-Ojeda et al.,2013b). In short, a spontaneous mutant lacking anthocyanin pigmentation wasidentified in a population collected in Central Spain (Figure 1). This mutant plantof O. cumana (S0 plant) was isolated before flowering with two microperforatedplastic bags 61 8 cm made up of Cryovac® SM570Y film (Sealed AirCorporation, Elmwood Park, NJ, USA) (Rodríguez-Ojeda et al., 2010). S1 seedswere used to inoculate eight plants of the susceptible sunflower cultivar DMM forconfirmation of the mutant phenotype in S1 plants. The S1 plants were bagged forself-fertilization as described above and the S2 seeds were bulked. This procedurewas repeated one more generation to obtain S3 seeds. This S2:3 O. cumana linewas named EK-A1.The inheritance of the trait was determined by studying the phenotypicsegregation of the F1, F2, and F3 generations from the cross of plants of themutant line EK-A1 with normally pigmented plants of the S2:3 line EK-12, derivedfrom a population collected in Écija (Spain). Crosses were made by removingwith forceps the corolla and the four stamens of the flower of the female plantapproximately 2 days before flower opening (Figure 2), followed by rubbingfresh anthers of the male plant onto the stigma of the emasculated flower. Inall cases, both female and male plants were bagged before flowering (Figure 3).The mutant trait was also used for estimating the rate of cross-fertilization takingadvantage of the intermediate phenotype of F1 hybrids (Figure 4). Individualmutant plants were surrounded by normally pigmented plants at a minimumdistance of around 15 cm under open pollination conditions in both pot and fieldexperiments. The rate of cross-fertilization was estimated as the percentage ofhybrid plants in the progenies of mutant plants.For the study of avirulence, lines of O. cumana races E and F were developedby selfing for three generations single plants parasitizing sunflower cultivarsGenetic Studies in Sunflower Broomrape 153Brought to you by | CSIC - Unidad Coordinacion BibliotecasAuthenticatedDownload Date | 1/14/16 1:24 PMresistant to races A through D (for race E O. cumana lines) or A through E (forrace F O. cumana lines). Confirmatory experiments showed that seeds of race Elines showed virulence on plants of sunflower cultivars Ritmo and S-1358,resistant to races A through D, and avirulence on plants of sunflower cultivarP-1380, resistant to race E (Vrânceanu et al., 1980). Similarly, seeds of race Flines showed virulence on plants of cultivar P-1380 and avirulence on plants ofcultivars I371B and P96, resistant to race F (Fernández-Martínez et al., 2004).Crosses between plants of race E and race F lines were made as described above.Figure 1: Mutant plant of Orobanche cumana lacking anthocyanin pigmentation (left) togetherwith a normally pigmented plant (right)154 M. I. Rodríguez-Ojeda et al.Brought to you by | CSIC - Unidad Coordinacion BibliotecasAuthenticatedDownload Date | 1/14/16 1:24 PMFigure 2: Manual emasculation of Orobanche cumanaFigure 3: Isolation of Orobanche cumana plants with microperforated plastic bagsGenetic Studies in Sunflower Broomrape 155Brought to you by | CSIC - Unidad Coordinacion BibliotecasAuthenticatedDownload Date | 1/14/16 1:24 PMThe study was based on the avirulence/virulence reactions observed for the O.cumana F1 and F3 generations on the race E-resistant differential sunflower lineP-1380 (Vrânceanu et al., 1980).Results and discussionIn the study of the inheritance of the unpigmented plant trait, F1 plants fromcrosses between unpigmented and normally pigmented plants showed an inter-mediate phenotype characterized by a greenish stem color, compared to ayellowish stem color in unpigmented plants and a bluish-violet stem color innormally pigmented plants. F2 plants segregated as 1 bluish violet:2 greenish:1yellowish stem color, suggesting that pigmentation of O. cumana plants iscontrolled by partially dominant alleles at a single locus. This monogenicinheritance was confirmed by studying the stem color of the F3 generation(Rodríguez-Ojeda et al., 2011).The intermediate phenotype of the F1 hybrid (Figure 4) allowed the use ofthe mutant trait as a morphological marker for the study of the rate of cross-fertilization in O. cumana. The results showed that 21.5% of the plants derivedfrom seeds produced by unpigmented plants in the pot experiment and 28.8% ofthe plants derived from seeds produced by unpigmented plants in the fieldexperiment had an intermediate phenotype, indicating the existence of cross-pollination (Rodríguez-Ojeda et al., 2013a). It is important to note that thesefigures were obtained under experimental conditions using individual mutantplants surrounded by many normally pigmented plants, but they do not neces-sarily reflect the actual cross-fertilization rate under specific natural conditions,which should be determined on a case-by-case basis. In any case, the resultsdemonstrated that the species is not strictly self-pollinated and some extent ofcross-pollination can be expected, which has important implications for under-standing population structure in O. cumana. We observed that cross-pollinationwas mainly conducted by small insects of the Hymenoptera, with a length fromaround 3 to 9 mm. Some of them were identified as belonging to the Halictidaefamily (Rodríguez-Ojeda et al., 2013a).The study of the inheritance of avirulence in crosses between plants of O.cumana lines of races E and F was conducted by evaluating the virulencereaction of the F1 and F3 generations on the sunflower differential line P-1380resistant to O. cumana race E but susceptible to race F. In all cases, O. cumana F1seeds did not parasitize sunflower P-1380 plants, which suggested dominance ofrace E avirulence over race F virulence. The evaluation of 387 F2:3 O. cumana156 M. I. Rodríguez-Ojeda et al.Brought to you by | CSIC - Unidad Coordinacion BibliotecasAuthenticatedDownload Date | 1/14/16 1:24 PMfamilies from the above-mentioned crosses showed complete absence of para-sitism on sunflower line P-1380 in one fourth of the F2:3 families, whereasthe remaining families were able to parasitize P-1380 plants. These resultssuggested that race E avirulence and race F virulence on P-1380 are allelic andcontrolled by a single locus, which confirmed the gene-for-gene theory for theO. cumana–sunflower interaction.Monogenic inheritance of avirulence/virulence in O. cumana is important forunderstanding race evolution in this parasitic weed, since new races may appearas the result of single mutation events. This is particularly relevant for anspecies in which single plants produce thousands of seeds. Recently, Pineda-Martos et al. (2013) and Molinero-Ruiz et al. (2014) identified two distinct geneFigure 4: F1 Orobanche cumana plant with intermediate pigmentation identified in the proge-nies of a mutant plant under open pollination conditionsGenetic Studies in Sunflower Broomrape 157Brought to you by | CSIC - Unidad Coordinacion BibliotecasAuthenticatedDownload Date | 1/14/16 1:24 PMpools of O. cumana in Spain, both of them containing low genetic diversity butpopulations classified as either race E or race F, suggesting that race F mutationsoccurred independently in both gene pools.Because of the simple inheritance of avirulence/virulence and the existenceof a relatively high rate of cross-fertilization in O. cumana, recombination ofdifferent virulence genes may play an important role in race evolution if popula-tions with different virulence genes get into contact. In this sense, Pineda-Martoset al. (2013) documented the presence of heterozygotes in a few Spanish popu-lations of O. cumana in which individuals of the two predominant gene poolswere present.Acknowledgments: The manuscript reviews research partially funded byFundación Ramón Areces, Madrid. The contribution of Dr. Enrique QuesadaMoraga, entomologist from the University of Córdoba, Spain, to taxonomicclassification of pollinators is gratefully acknowledged. R. Pineda-Martos wasthe recipient of a PhD fellowship from the Spanish National Research Council(CSIC) (JAEPre_08_00370). Technical support by Alberto Merino and PlácidaNieto is acknowledged.ReferencesAntonova, T.S., Araslanova, N.M., Strelnikov, E.A., Ramazanova, S.A., Guchetl, S.Z.,Chelyustnikova, T.A., 2013. Distribution of highly virulent races of sunflower broomrape(Orobanche cumana Wallr.) in the Southern regions of the Russian Federation. RussianAgricultural Sciences 39: 46–50.Fernández-Martínez, J.M., Pérez-Vich, B., Akhtouch, B., Velasco, L., Muñoz-Ruz, J., Melero-Vara,J.M. and Domínguez, J., 2004. Registration of four sunflower germplasm lines resistant torace F of broomrape. Crop Science 44: 1033–1034.Gagne, G., Roeckel-Drevet, P., Grezes-Besset, B., Shindrova, P., Ivanov, P., Grand-Ravel, C.,Vear, F., Tourvieille de Labrouhe, D., Charmet, G. and Nicolas, P., 1998. Study of thevariability and evolution of Orobanche cumana populations infesting sunflower in differentEuropean countries. Theoretical and Applied Genetics 96: 1216–1222.Molinero-Ruiz, M.L., García-Carneros, A.B., Collado-Romero, M., Raranciuc, S., Domínguez, J.and Melero-Vara, J.M., 2014. Pathogenic and molecular diversity in highly virulent popu-lations of the parasitic weed Orobanche cumana from Europe. Weed Research 54: 87–96.Musselman, L.J., Parker, C. and Dixon, N., 1981. Notes on the autogamy and flower structure inagronomically important species of Striga (Scrophulariaceae) and Orobanche(Orobanchaceae). Beiträge zur Biologie der Pflanzen 56: 329–343.Pineda-Martos, R., Velasco, L., Fernández-Escobar, J., Fernández-Martínez, J.M., Pérez-Vich, B.,2013. Genetic diversity of sunflower broomrape (Orobanche cumana) populations fromSpain. Weed Research 53: 279–289.158 M. I. Rodríguez-Ojeda et al.Brought to you by | CSIC - Unidad Coordinacion BibliotecasAuthenticatedDownload Date | 1/14/16 1:24 PMPérez-Vich, B., Velasco, L., Rich, P.J., Ejeta, G., 2013. Marker-assisted and physiology-basedbreeding for resistance to Orobanchaceae. In: Joel, D.M, Gressel, J., Musselman, L.J. (eds)Parasitic Orobanchaceae, Springer-Verlag, Berlin Heidelberg, pp. 369–391.Rodríguez-Ojeda, M.I., Fernández-Martínez, J.M., Velasco, L., Pérez-Vich, B., 2013a. Extent ofcross-fertilization in Orobanche cumana Wallr. Biologia Plantarum 57: 559–562.Rodríguez-Ojeda, M.I., Pineda-Martos, R., Alonso, L.C., Fernández-Escobar, J., Fernández-Martínez, J.M., Pérez-Vich, B., Velasco, L., 2013b. A dominant avirulence gene inOrobanche cumana triggers Or5 resistance in sunflower. Weed Research 53: 322–327.Rodríguez-Ojeda, M.I., Pérez-Vich, B., Alonso, L.C., Fernández-Escobar, J., 2010. The influenceof flowering plant isolation on seed production and seed quality in Orobanche cumana.Weed Research 50: 515–518.Rodríguez-Ojeda, M.I., Velasco, L., Alonso, L.C., Fernández-Escobar, J., Pérez-Vich, B., 2011.Inheritance of the unpigmented plant trait in Orobanche cumana. Weed Research 51:151–156.Satovic, Z., Joel, D.M., Rubiales, D., Cubero, J.I., Román, B., 2009. Population genetics in weedyspecies of Orobanche. Australasian Plant Pathology 38: 228–234.Škorić, D., Păcureanu-Joiţa, M., Sava, E., 2010. Sunflower breeding for resistance to broomrape(Orobanche cumana Wallr. Analele INCDA Fundulea 78: 63–79.Sukno, S., Melero-Vara, J.M., Fernández-Martínez, J.M., 1999. Inheritance of resistance toOrobanche cernua Loefl. in six sunflower lines. Crop Science 39: 674–678.Velasco, L., Pérez-Vich, B., Yassein, A.A.M., Jan, C.C., Fernández-Martínez, J.M., 2012.Inheritance of resistance to sunflower broomrape (Orobanche cumana Wallr.) in a inter-specific cross between Helianthus annuus and Helianthus debilis subsp. tardiflorus. PlantBreeding 121: 220–221.Vrânceanu, A.V., Tudor, V.A., Stoenescu, F., Pirvu, N., 1980. Virulence groups of Orobanchecumana Wallr., differential hosts and resistance source genes in sunflower. In: Proc 9thInternational Sunflower Conference, Torremolinos, Spain, International SunflowerAssociation, pp. 74–82.Genetic Studies in Sunflower Broomrape 159Brought to you by | CSIC - Unidad Coordinacion BibliotecasAuthenticatedDownload Date | 1/14/16 1:24 PM

Genetic studies in sunflower broomrape

Abstract

Similar works

Full text

Available Versions

Digital.CSIC