Characteristics of a <Emphasis Type="Italic">Plasmopara angustiterminalis</Emphasis> isolate from <Emphasis Type="Italic">Xanthium strumarium</Emphasis>

Leaves of Xanthium strumarium infected with downy mildew were collected in the vicinity of a sunflower field in southern Hungary in 2003. Based on phenotypic characteristics of sporangiophores, sporangia and oospores as well as host preference the pathogen was classified as Plasmopara angustiterminalis. Additional phenotypic characters were investigated such as the size of sporangia, the number of zoospores per sporangium and the time-course of their release. Infection studies revealed infectivity of the P. angustiterminalis isolate to both X. strumarium and Helianthus annuus. Inoculation of the sunflower inbred line, HA-335 with resistance to all known P. halstedii pathotypes, resulted in profuse sporulation on cotyledons and formation of oospores in the bases of hypocotyls. Infections of sunflower differential lines often led to damping-off. Molecular genetic analysis using simple sequence repeat primers and nuclear rDNA sequences revealed clear differences to Plasmopara halstedii, the downy mildew pathogen of sunflower.     

Host status effect of cowpea and sunflower on the populations ofMeloidogyne javanica andRotylenchulus reniformis

A highly susceptible cowpea,Vigna sinensis cv. Baladi plants were tested as trap plants for eitherMeloidogyne javanica orRotylenchulus reniformis under greenhouse conditions. The plants were gathered by cutting them above the surface of the soil or by uprooting them, 1/2, 1, 3, 6, 12, 24, 36 and 48 days after nematode inoculation. Both of the mentioned nematodes began to mature and lay eggs after the 12th day from their inoculation. Hence, it is advised to pull up cowpea plants from 3–12 days after nematode inoculation. After planting sunflower,Helianthus annus cv. Miak replacing cowpea, the nematode populations were higher, in most cases, on sunflower plants replacing cutting cowpea than those on sunflower replacing uprooted cowpea. The highest percentages of nematode reduction were 98.55 and 99.57 forM. javanica and 95.09 and 92.90% forR. reniformis on sunflower plants replacing cutting and uprooted cowpea plants after 12 days from nematode inoculation, respectively.M. javanica andR. reniformis decreased the length and weight of sunflower plants as affected by planting time and method of cowpea harvest. This method of nematode control is cheaper, easy and pollution free.     

Simulation study of the competitive ability of erect, semi-erect and prostrate cowpea (Vigna unguiculata) genotypes

Ecophysiological simulation models provide a quantitative method to predict the effects of management practices, plant characteristics and environmental factors on crop and weed growth and competition. The INTERCOM interplant competition model was parameterised, calibrated by monoculture data for three cowpea (Vigna unguiculata) genotypes that differed in growth habit, common sunflower (Helianthus annuus) and common purslane (Portulaca oleracea), and used to simulate competition of cowpea cover crops with sunflower or purslane. The simulation results were compared with observations from field competition experiments in 2003 and 2004. INTERCOM more accurately simulated actual field data for the competition of cowpea genotypes and sunflower than companion field experiments for the competition of cowpea and purslane. The validated simulation model of cowpea and sunflower at two densities was used to study the effects of cowpea growth habit on final biomass production of cowpea and sunflower. The model suggested that erect growth habit was more competitive than semi‐erect and prostrate growth habit, when cowpea genotypes were grown with sunflower. Cowpea leaf area distribution was important to higher cowpea biomass production, while cowpea height growth was important to reduce sunflower biomass. Our simulation approach is suggested as a method for crop breeders to gauge the likely success of selection for competitive crops before undertaking expensive long‐term breeding experiments.     

A dominant avirulence gene in Orobanche cumana triggers Or5 resistance in sunflower

Orobanche cumana is a weed that grows as a root parasite on sunflower. In general, the O. cumana–sunflower parasitic system is regarded to follow the gene‐for‐gene model, although this has never been demonstrated at the genetic level in O. cumana. The Or5 dominant gene in sunflower confers resistance to O. cumana race E, but not to race F. The objective of this research was to study the inheritance of avirulence/virulence in crosses between plants of O. cumana lines classified as races E and F. Four race E and three race F lines were developed, from which four race E × race F cross‐combinations were made, in three cases including reciprocals. In all cases, F₁ seeds did not have the ability to parasitise sunflower line P‐1380 carrying the Or5 gene, indicating dominance of race E avirulence allele(s). Five F₂ populations comprising a total of 387 F_2:3 families were evaluated on sunflower line P‐1380. In all cases, one‐fourth of the F_2:3 families did not possess the ability to parasitise P‐1380 plants, suggesting that race E avirulence and race F virulence on P‐1380 are allelic and controlled by a single locus. This study demonstrated the gene‐for‐gene interaction in the O. cumana–sunflower parasite system and provided useful information to identify genes involved in O. cumana virulence. The approach followed in this research can contribute to define precisely races of the parasite on the basis of the presence of avirulence genes.     

Methodology of virulence screening and race characterization of Plasmopara halstedii,and resistance evaluation in sunflower – a review

Sunflower downy mildew is a disease of high global economic impact as well as a causal agent that is extremely difficult to eradicate. During the past decades, several approaches for the determination of Plasmopara halstedii (Ph) races have been used worldwide and are discussed in this review. Procedures of isolation, cultivation and maintenance of Ph isolates, as well as the screening of sunflower for resistance, are also critically reviewed. The predominant, globally used resistance screening protocol is a ‘whole seedling immersion’ inoculation. ‘Soil drench’ inoculation allows more precise control of the number of Ph zoosporangia applied to a single sunflower seedling. A detached leaf assay has been described, but it has been used mainly for Ph subcultivation and fungicide tests. For race determination, a differential set consisting of nine sunflower genotypes has been used since 1988, coupled with a numerical triplet code for virulence phenotyping of Ph. The increasing variability in global Ph populations has demonstrated the inadequacy of the current set of differentials, and several researchers have proposed additional public lines as new differentials. Furthermore, bulk isolates may show different results in repeated tests, as Ph may contain genetically distinct zoospores within a single zoosporangium. For precise race determination, single zoosporangia or single zoospore isolates are advisable. However, due to low success of isolation, approximately 1–2%, this method cannot be applied in routine Ph race screening. Methods surveyed in this review have a broad spectrum of applications, including taxonomic studies.     

Cross-pathogenicity of <Emphasis Type="Italic">Verticillium dahliae</Emphasis> between potato and sunflower

This study examined cross-pathogenicity of the soilborne pathogen Verticillium dahliae between potato and sunflower. Four week-old potato and sunflower seedlings were inoculated with ten isolates from each of the two host species. Potato cultivars (Kennebec, susceptible, and Ranger Russet, moderately resistant) and sunflower hybrids (IS8048, susceptible, and 6946, moderately resistant) were assessed for disease severity and percent infection at 2 weeks, 3 weeks, 4 weeks, 5 weeks, and 6 weeks after inoculation (w.a.i), and for vascular discolouration at 6 w.a.i., using visual scales developed for each host species. The experiments were conducted in 2006 and repeated in 2007. Based on percent infection and disease severity, most V. dahliae isolates were highly aggressive on both host species. The tested isolates caused higher disease levels in the susceptible than in the moderately resistant phenotypes. They also caused more vascular discolouration in their original than in the alternative host. However, the isolates originating from sunflower caused less infection and disease severity on both hosts, compared to their potato counterparts. Cluster analysis based on all of the criteria used to assess pathogenicity led to three groups of isolates: (i) most V. dahliae potato isolates, which ranged with the highly aggressive control isolates, (ii) one V. dahliae sunflower isolate, which showed a similar pathogenicity level to the weakly-aggressive V. albo-atrum sub-group II control isolate, with no more symptoms than in the non-inoculated plants, and (iii) most V. dahliae sunflower isolates with mildly- to weakly-aggressive levels. Based on these results, V. dahliae cross-pathogenicity is very effective between potato and sunflower. Therefore, rotations involving these species should be avoided, especially where sunflower follows potato.     

Differentiation of two powdery mildews of sunflower (<Emphasis Type="Italic">Helianthus annuus</Emphasis>) by a PCR-mediated method based on ITS sequences

Powdery mildew can be found in most sunflower fields during the winter season in Taiwan and causes severe yellowing on the blade, petiole, stem, and calyx, as well as serious defoliation. Two types of powdery mildew fungi isolated from sunflower leaves showed variable status for fibrosin bodies. But only the cleistothecium of Podosphaera xanthii, one of the pathogens causing this disease, was observed on samples from Chungpu County at the beginning of 2005. With a species-specific primer pair, PN23/PN34, no specific PCR product was amplified from the pathogen’s genomic DNA. Based upon the ITS sequence of rDNA, three PCR primer sets (S1/S2, G1/G2, and L1/L2) specific to P. xanthii, Golovinomyces cichoracearum and Leveillula taurica, respectively, were designed to detect and differentiate pathogens causing powdery mildews on sunflower. Only the primer pairs S1/S2 and G1/G2 could amplify PCR products, with product sizes of 454 and 391 bp, respectively. Four samples of fungal DNA were subjected to a multiplex PCR amplification with primer pairs S1/S2 and G1/G2; P. xanthii and G. cichoracearum were successfully detected. These results suggest that the multiplex PCR method is a rapid, simple, and effective technique to detect and differentiate powdery mildews, for example P. xanthii and G. cichoracearum, found on sunflower. With morphologic characteristics, ITS sequence analysis and pathogenicity testing, P. xanthii and G. cichoracearum, the first case, are two powdery mildews on sunflower in Taiwan.     

Prohexadione-calcium induces sunflower (Helianthus annuus) resistance against the root parasitic weed Orobanche cumana

Prohexadione-calcium (PHDC) is a plant growth retardant that inhibits gibberellin biosynthesis and transiently alters the spectrum of flavonoids and their phenolic precursors. In this study, root chamber and pot experiments were conducted to evaluate the effects of PHDC on Orobanche cumana seed germination and induction of sunflower resistance to this root parasitic weed. Root chamber experiments indicated that PHDC did not interfere with the progress of Orobanche seed germination, but retarded O. cumana tubercle formation and development. In pot experiments, control of O. cumana by PHDC depended on its dose, method of application and sunflower cultivar. In sunflower cv. HA89, PHDC at 10 mg LS⁻¹ (litre soil) as two drenches or at 1 m m as two foliar sprays significantly reduced the total number and dry matter of O. cumana without influencing flower, leaf and root development. In sunflower cv. Albena, PHDC at 20 and 30 mg LS⁻¹, but not at 10 mg LS⁻¹, as two drenches significantly reduced O. cumana infection. Sunflower cv. HA89 was less susceptible to O. cumana and more inducible by PHDC than sunflower cv. Albena. The biochemical analysis of root extracts revealed a significantly higher accumulation of free phenolics in sunflower cv. HA89 after PHDC soil drench, but not in Albena. No significant lignification was observed after PHDC treatments. The results suggest that PHDC reduces O. cumana infection in sunflower by inducing host resistance, not by directly interfering with the parasite. Free phenolics play an important role in sunflower response to the parasite.     

Orobanche cumana race F: performance of resistant sunflower hybrids and aggressiveness of populations of the parasitic weed

The production of sunflower is threatened by Orobanche cumana (broomrape) infections in many countries of southern and eastern Europe. The most virulent race of this parasitic weed, race F, is widespread in Spain. Most of the current commercial hybrids of resistant sunflower do not completely prevent broomrape emergence in the field. The performance (broomrape incidence and severity, final yields and oil contents) of four resistant and two susceptible sunflower hybrids was studied in different years in several O. cumana – infested locations. O. cumana – sunflower interactions were characterised by low severity and intermediate incidence in the resistant hybrids; both depended on the environment. An increase in sunflower yield because of the use of resistant hybrids was most noticeable under heavy field infestations and under drought. Characterisation of 20 populations of O. cumana classified as race F in terms of the standardised area under the degree of attack progress curve (SAUDC) and O. cumana incidence (BI) suggested differential intrinsic pathogenicities. These results highlight the variability within race F of O. cumana , mirroring the environmentally dependent horizontal resistance in sunflower.     

Pathogenic diversity within field populations of Orobanche cumana and different reactions on sunflower genotypes

Different races of the parasitic Orobanche cumana (sunflower broomrape) have been reported in Spain, race F being the most virulent. Full resistance in sunflower to races A–E is achieved with each of the single major genes Or₁ to Or₅ respectively. However, parasitised hybrids allegedly resistant to race F were observed in early 2002. The purpose of this study was to verify broomrape incidences (BI) on resistant sunflower genotypes, to assess the mixture of races within field populations and to test for partial resistance to race F in the sunflower hybrids showing a low degree of attack (DA) by the weed. Tests were conducted under field conditions in two locations of southern Spain. While no significant differences were found for yield and BI between locations, the DA on the cultivars depended on the location. With high infection levels and significantly lower yield in susceptible controls, marked differences in BI and DA were found within resistant cultivars, but all of them showed similar crop yield. When artificially inoculated with several populations of race F, line P96 and mainly line L86, were consistently slightly infected, suggesting they were inbred lines responsible for horizontal resistance in infested fields. L86 was extremely susceptible to race E populations, which is unusual as sunflower resistance to one race provided resistance to all the previously described races of O. cumana. No different virulences were detected within two groups of subpopulations (races E and F) inoculated onto resistant sunflower genotypes. However, race F subpopulations showed significant differences in aggressiveness, which seems to be related to horizontal (multigenic) resistance of the crop to the parasitic weed.     

Indigenous highly virulent accessions of the sunflower root parasitic weed Orobanche cumana

Orobanche cumana (broomrape) is a severe constraint to sunflower production in southern and eastern Europe and the Middle East. Races A to E of this parasitic weed controlled by genes Or₁ to Or₅ have been described. A study of 38 seed accessions of O. cumana collected from different locations in Spain between 1983 and 2003 investigated the effect of long‐term storage in the laboratory on germination and infectivity and assessed their virulence on a number of sunflower cultivars. Only 18 infected the susceptible cultivar B117. Infectivity was maintained for up to 17 years of storage, but with a greatly decreased vigour as compared with that of recently collected seed. The 12 oldest viable accessions overcame the resistance of the gene Or₅ (in resistant line NR5). Seven out of them, in particular those collected in 1988 and 1989, were identified as race F. Three accessions were identified as race E allegedly holding components of higher virulence. Our results show evidence of the occurrence of race F prior to the use of sunflower hybrids resistant to race E, suggesting the former as indigenous to the country. This finding suggests the necessity of a continuous breeding of sunflower for resistance to O. cumana. The effectiveness and sustainability of genetic resistance must rely on the knowledge of the diverse virulence characteristics of O. cumana accessions.     

PCR-based detection of sunflower white blister rust (<Emphasis Type="Italic">Pustula helianthicola</Emphasis> C. Rost &; Thines) in soil samples and asymptomatic host tissue

Sequencing of partial cox2 (part of the mitochondrial cytochrome-c-oxydase (COX) gene) was performed with samples from the oomycete genus Pustula, the white blister rusts of Asteraceae and related families. Sequence comparison uncovered several single nucleotide polymorphisms (SNPs) between P. spinulosa and host specific strains of Pustula isolated from Senecio vulgaris, Tragopogon pratensis and cultivated sunflower, Helianthus annuus. Based on these differences, specific primers were designed for PCR-based detection of white blister rust strains pathogenic to sunflower. The specificity of the primers was confirmed by cross testing with DNA from various oomycetes occurring in the same locality. The limit of detection for DNA of P. helianthicola was 10 pg. This allowed detection with DNA from single sporangia and single oospores. The PCR-based experiments allowed detection of the presence of sunflower white blister rust in soil samples from fields on which infected plants had been cultivated several months before. Moreover, the molecular tools were successfully applied to trace the pathogen in asymptomatic tissue of infected plants, demonstrating the systemic nature of Pustula on sunflower.     

Estimating the potential development of Diaporthe helianthi epidemics in Italy*

Diaporthe helianthi is the causal agent of a severe sunflower disease but, in Italy, disease outbreaks are sporadic with no significant losses. The present work investigates the role of meteorological conditions on the potential development of D. helianthi epidemics in Italy, using the French model Asphodel, which simulates the effect of air temperature, relative humidity and rainfall on ascospore maturation and dispersal, infection establishment, disease onset and severity during the period of host susceptibility. Meteorological data measured in eight stations distributed from north to south Italy, over a 5‐year period (1995–99), was used as model input. Results showed that meteorological conditions in Italy are frequently favourable for D. helianthi infections on sunflower, and severe epidemics are possible. Therefore, climatic conditions are not a limiting factor for disease development in the Italian sunflower‐growing areas. The lack of disease epidemics in Italy may be related to differences in the pathogen populations compared with the French ones.     

干旱和盐胁迫对向日葵叶片蜡质积累的影响及蜡质烷烃合成基因的克隆

植物表皮蜡质在植物逆境胁迫响应中发挥重要作用。为了研究干旱和盐胁迫对向日葵叶片蜡质积累的影响,对三周龄向日葵分别进行干旱和盐胁迫处理,7 d后提取蜡质并利用气相色谱-质谱联用仪进行分析。结果表明：向日葵幼苗叶片蜡质主要由初级醇、烷烃和脂肪酸组成,其含量分别占蜡质总含量的79%、10%和9%。干旱和盐胁迫均没有改变向日葵幼苗叶片蜡质的组成,但是能够显著提高蜡质总含量,与对照相比,分别增加了8.8%和8.5%。所有蜡质组分中,烷烃含量变化最大,分别在干旱和盐胁迫后较对照增加了62.5%和47.0%,表明了向日葵幼苗叶片蜡质组分中,烷烃积极响应干旱和盐胁迫。为进一步研究参与向日葵蜡质烷烃合成的基因,在向日葵基因组中进行同源检索,共获得6个差异表达的候选基因,克隆了两个在叶片中表达水平较高的HaCER1-1和HaCER3-1。测序结果显示,HaCER1-1和HaCER3-1编码区长度分别为1 869 bp和1 674 bp,编码622个和557个氨基酸的蛋白质。利用qRT-PCR技术进一步分析了NaCl和PEG溶液处理下HaCER3-1的表达水平,结果显示,在处理后的12 h时,HaCER3-1在叶片中的表达分别增加了11倍和3.5倍,表明HaCER3-1的表达受到干旱和盐胁迫的诱导。本研究为解析向日葵蜡质响应渗透胁迫和烷烃合成机制奠定了分子基础。     

Spectral discrimination of Ridolfia segetum and sunflower as affected by phenological stage

Ridolfia segetum is an umbelliferous weed frequent and abundant in sunflower crops in the Mediterranean basin. Field research was conducted to evaluate the potential of hyperspectral and multispectral reflectance and five vegetation indices in the visible to near infrared spectral range, for discriminating bare soil, sunflower and R. segetum at different phenological stages. This was a preliminary step for mapping R. segetum patches in sunflower using remote sensing for herbicide application decisions. Reflectance data were collected at three sampling dates (mid‐May, mid‐June and mid‐July, corresponding to vegetative‐early reproductive, flowering and senescent phenological stages respectively) using a handheld field spectroradiometer. Differences observed in hyperspectral reflectance curves were statistically significant within and between crop and weed phenological stages depending on sampling date, which facilitates their discrimination. Statistically significant differences in the multispectral and vegetation indices analysis showed that it is also possible to distinguish any of the classes studied. Our study provides some information for constructing the spectral libraries of sunflower and R. segetum in which the different phenological stages co‐existing in the field were considered. Hyperspectral and multispectral results suggest that mapping R. segetum patches in sunflower is feasible using airborne hyperspectral sensors, and high‐resolution satellite imagery or aerial photography, respectively, taking into account specific timeframes.     

Pathogenic and molecular diversity in highly virulent populations of the parasitic weed Orobanche cumana (sunflower broomrape) from Europe

The parasitic weed Orobanche cumana (sunflower broomrape) constrains sunflower production in eastern and southern Europe and in the Middle East. Although genetic resistance is the most effective control method, new parasite races evolve overcoming sunflower resistance. In this work, highly virulent populations of O. cumana were analysed for pathogenicity and genetic diversity. The virulence of 11 populations from Hungary, Romania, Spain and Turkey was assessed and compared after infection of sunflower inbred lines to differentiate races of the parasite under glasshouse conditions. Molecular diversity among and within 27 parasite populations was studied by RAPD‐PCR, UPGMA and amova analyses. Highly virulent race F was identified in Hungary, Spain and Turkey. The most virulent race (G) was also found in Turkey. The molecular analysis among highly virulent populations of O. cumana identified four molecular clusters, respectively, grouping populations from Central Spain, Hungary, South Spain and Turkey. The genetic homogeneity within parasite populations was confirmed, since no molecular divergences were found within them. This work constitutes the first geographical study of O. cumana together with pathogenicity and molecular traits inherent to each geographical group, and provides useful information for possible phylogenetic analyses of O. cumana. In addition, molecular markers associated with geographical origin could be developed and used as diagnostic tools to track new broomrape introductions into areas free of virulent races where they might represent a threat to sunflower production.     

Development of a PCR test to detect the downy mildew causal agent Plasmopara halstedii in sunflower seeds

Plasmopara halstedii , the causal agent of downy mildew of sunflower, is an obligate parasite but viable sporangia and oospores of the pathogen may be found in a quiescent state in seeds of sunflower and therefore may be transported with sunflower seeds in international commercial exchanges. In order to prevent the spread of this pathogen, especially the introduction of potentially new races, an efficient method to analyse sunflower seed samples is required. In this study, a P. halstedii -specific polymerase chain reaction (PCR) test was developed based on the ribosomal large sub unit (LSU) DNA. The forward (PHAL-F) and reverse (PHAL-R) PCR primers were designed from two polymorphic regions of LSU. After screening 22 isolates of P. halstedii corresponding to different races and countries and 32 other oomycete, deuteromycete and ascomycete isolates, the PHAL-F/R primers amplified only a single PCR band of c. 310 bp from P. halstedii . The PHAL-F/R PCR test could detect as little as 3 pg of P. halstedii genomic DNA per 20  µ L reaction volume and enabled the direct detection of P. halstedii in 35 g sunflower seed samples without the need for any prior biological baiting step. An internal amplification control (IAC) was developed to help discriminate against false negative samples due to the potential presence of inhibitory compounds in DNA extracts. The test was successfully used on samples of naturally contaminated seeds. These new molecular tools should be of great interest for quarantine seed testing purposes.     

Interference of the agrestal Helianthus annuus biotype with sunflower growth

The Helianthus annuus taxon includes the wild (ruderal), weedy (agrestal) and volunteer biotypes, which may share space and time with the sunflower crop. The agrestal biotype is one of the most harmful weeds challenging the use of modern biotechnological tools in sunflower. The interference of the agrestal biotype was studied at a range of weed densities within a sunflower crop. The area of influence of one agrestal plant was estimated as a circular area of more than 2.5 m². The agrestal biotype at 10.7 plants m^?2 reduced the sunflower seed production by 66%, the seed weight by 41% and the yield by 80%. The agrestal biotype showed intermediate vegetative stage duration between sunflower and the ruderal biotype. The interference of the agrestal biotype would be due to its high initial growth rate, its larger leaf area and plant height. Some agrestal plants showed partial dehiscence at the sunflower harvest stage replenishing the soil seedbank with over 18 000 seeds m^?2. The sunflower harvest operation and grain storage might be affected due to a fraction of agrestal plants remaining alive and moist when the crop reached harvest maturity. The results support the recommendation of preventing the entry of the agrestal biotype into crop production fields.     

Root absorption and translocation of 5-substituted analogs of the imidazolinone herbicide,imazapyr

In preparation for assessing quantitative structure-activity relationships (QSAR) for root absorption and translocation of imidazolinones herbicides, 13 radiolabeled analogs of imazapyr (2-(4-isopropyl–4-methy1–5-oxo-2-imidazolin-2-yl) nicotinic acid) substituted in the 5-position of the pyridine ring were evaluated in corn (Zea mays L.) and sunflower (Helianthus annuus L.). The compounds (10 μm) were supplied to the roots through a hydroponic solution for 8 h and, following harvest, plant tissues were either combusted to measure total uptake of radiolabeled material or were extracted for determination of the extent of degradation of the parent compound. Root absorption in both species varied by two orders of magnitude among analogs while translocation, expressed as a percentage of absorption, varied by only three- or four-fold. Few differences were observed for translocation of radioactivity from sunflower stems to leaves. Although six of the analogs were partially metabolized in corn, little metabolism of the imidazolinone analogs occurred in sunflower. These data indicate that meaningful models of root absorption and subsequent translocation to shoots may be developed for 5-substituted analogs of imazapyr, particularly when applied to sunflower.