No abstract

No abstract

White rot caused by Sclerotinia sclerotiorum (SS) is one of the most devastating plant diseases of sunflower. Controlling this pathogen by available tools hardly result in acceptable control. The aim of this study was to elucidate the effects of plant resistance inducers, BTH (benzothiadiazole in Bion 50 WG) and arbuscular mycorrhizal fungi (AMF) on disease development of white rot in three sunflower genotypes. Defence responses were characterized by measuring the disease severity and identifying cellular/ histological reactions (e.g. autofluorescence) of host plants upon infection. Depending on the host genotype, a single application of inducers reduced disease symptoms. Histological examination of host responses revealed that BTH and/or AMF pre-treatments significantly impeded the development of pathogenic hyphae in Iregi szürke csíkos and P63LE13 sunflower plants and it was associated with intensive autofluorescence of cells. Both localized and systemic induction of resistance was observed. Importantly, the frequency of mycorrhization of hybrid P63LE13 and PR64H41 was significantly increased upon BTH treatment, so it had a positive effect on the formation of plant-mycorrhiza interactions in sunflower. To our knowledge, this is the first report on the additive effect of BTH on mycorrhization and the positive effect of these inducers against SS in sunflower.

In the present study, the mechanism of resistance to Plasmopara halstedii, the downy mildew pathogen of sunflower, triggered by three resistance-inducing chemicals, benzo (1,2,3) thiadiazole-7-carbothioic acid S-methyl ester (BTH), DL-b-amino butyric acid (BABA) and 2,6-dichloroisonicotinic acid (INA), was investigated in susceptible, completely resistant and partially resistant sunflower genotypes. By applying P. halstediispecific primers, no detectable pathogen marker transcript accumulation was found in the infected but completely resistant sunflower hypocotyls; however, pretreatments with either of the three resistance inducers decreased the transcript accumulation in both the infected susceptible and the partially resistant lines. Benzothiadiazole pretreatment before inoculation considerably enhanced enzyme activities in the infected susceptible and the completely resistant genotypes but not in partially resistant plants. Pretreatment with resistance inducers before inoculation increased glutathione S-transferase, defensin and catalase transcript levels in the susceptible but decreased in the partially resistant plants. Our results indicate that the resistance-inducing chemicals can improve resistance in all of the sunflower genotypes to downy mildew and increase enzyme activities of peroxidase and polyphenol oxidase, as well as accumulation of mRNAs of glutathione S-transferase, defensin and catalase. However, it is important to emphasize that activation of these defence-related proteins did not correlate with the degree of resistance, but rather with the amount of necrotic tissues.

Downy mildew of sunflower, caused by Plasmopara halstedii (Farlow) Berlese et de Toni, is an economically important disease in Hungary and much of Europe. The known pathotypes (races) of the pathogen influence the resistance genes (Pl genes) incorporated into new sunflower hybrids to manage the disease. There are at least 36 pathotypes of P. halstedii worldwide (3), but the number of races is increasing rapidly. In 2010, race 704 was identified in Hungary for the first time (2). Race 704 has been reported to confer virulence on Pl6, a broad spectrum resistance gene that is widely used in sunflower hybrids. This has coincided with a significant increase in disease severity since 2010 in the country. Our objectives are to continuously monitor this pathogen and identify pathotypes of P. halstedii. Because of the unfavorable weather conditions for downy mildew in 2013, samples were collected at a single site (Kunszentmárton, South Hungary) in the beginning of July from NK Neoma sunflower hybrids. Disease incidence (early and late primary infection) was as high as 40%. Systemically mildewed plants showed severe stunting and leaf chlorosis, signs and symptoms consistent with downy mildew. P. halstedii was identified microscopically. Examination of isolates was carried out using a set of sunflower differential lines based on the internationally standardized method for race identification of P. halstedii (1). Inoculum of the isolates was increased on a susceptible cultivar (cv. Iregi szürke csíkos) and tested by inoculating 3-day-old seedlings of sunflower differential lines. Inoculated seedlings were planted in trays in glasshouse. After 8 to 9 days, seedlings were sprayed with distilled water, covered with black plastic bags, and left overnight to induce sporulation. Disease incidence was determined by examining cotyledons at 9 days after inoculation for sporulation and true leaves on 12 to 13 days after inoculation for secondary symptoms, such as leaf chlorosis and stunting (1). While several differential lines showed no typical susceptible/resistant reactions, i.e., the infection was much lower than 100%, it was concluded that the isolates were mixtures of different P. halstedii pathotypes. To obtain single isolates, we collected zoosporangia from the differential lines in question separately, and then inoculated the seedlings of the same genotype and a uniformly susceptible line. A single isolate caused as high as 100% infection on HA-335, containing resistance gene Pl6. Subsequent evaluation of this isolate with the entire differential set resulted in an aggregate virulence phenotype of 714. As resistance gene Pl6 is incorporated to the majority of sunflower hybrids grown in Hungary, pathotypes virulent on this gene, such as 704 and 714, are likely to spread. This underscores the need to prove the resistance to these races in the newly registered hybrids and for further research to identify P. halstedii pathotypes. It is also important to establish the identity of this new pathotype by already discovered 714 pathotypes in other countries like France and Italy and to discover the real conditions of local evolving of new pathogens. To our knowledge, this is the first report of pathotype 714 of P. halstedii in both Hungary and Central Europe. References: (1) T. J. Gulya et al. Helia 14:11, 1991. (2) K. Rudolf et al. Növényvédelem 47:279, 2011. (3) F. Virányi and O. Spring. Eur. J. Plant Pathol. 129:207, 2011.

Downy mildew of sunflower caused by Plasmopara halstedii (Farlow) Berlese et de Toni is a devastating disease worldwide. The treatment of seeds with fungicides and the use of resistant cultivars are widely employed control measures against this oomycete. Effective protection, however, may be hindered by the high genetic variability of the pathogen. There are 14 pathotypes of P. halstedii in Europe and as many as six of these were identified in Hungary before 2010 (1,4). In 2010, a new race, 704, was isolated in the eastern region of Hungary (3). Although the new pathotype was identified in two sunflower fields (near Vészto and Kondoros), it was expected to spread all over the country because of a lack of resistance against this race. The aim of our study, therefore, was to monitor the distribution of pathotype 704 in Hungary. Infected sunflower plants (2 to 5 samples/site) showing typical symptoms of downy mildew (leaf chlorosis, severe stunting) from four different sites (Árpádhalom, Rákóczifalva, Tiszasüly, and Újiráz) in the eastern region of the country were collected in mid-June 2012. Examination of isolates was carried out using a set of sunflower differential lines based on the internationally standardized method for race identification of P. halstedii (2). Inoculum of 17 isolates was increased on a susceptible cultivar (cv. Iregi Szürke Csíkos). Leaves containing zoosporangia were washed off in distilled water. The concentration of spore suspension for each isolate was adjusted to 20,000 to 30,000 viable zoosporangia per ml using a hemacytometer. Pre-germinated seeds of sunflower differential lines (20 seeds/line) with an optimal radicle length were selected and placed in separate petri dishes. They were filled with freshly prepared zoosporangial suspension of the isolates and incubated in the dark at 16°C for 6 h. Inoculated seeds were planted in trays. After 8 to 9 days when the first true leaves were ~0.5 to 1 cm long, the trays containing the plants were covered with transparent plastic bags overnight. Distilled water was sprayed into the bags to ensure a humid environment for stimulating sporulation. First disease assessment was performed immediately after incubation based on the appearance of characteristic white sporulation on cotyledons. A second evaluation was made of true leaves of 21-day-old plants. Twelve out of 17 isolates were pathotype 704, infecting either one of two commercial sunflower hybrids (NK Neoma and NK Brio) or volunteer sunflower plants. The remaining five isolates were races 700, 710, and 730, which are known to be widespread in Hungary (1). The presence of race 704 was proven in all sampling sites representing the eastern part of the country. This finding underscores the need to develop and grow improved sunflower hybrids with effective genes against this pathotype. To our knowledge, this is the first report of the wider distribution of pathotype 704 of P. halstedii in both Hungary and Central Europe. References: (1) T. J. Gulya. Adv. Downy Mildew Res. 3:121, 2007. (2) T. J. Gulya et al. Helia 14:11, 1991. (3) K. Rudolf et al. Növényvédelem 47:279, 2011. (4) F. Virányi and S. Maširević. Helia 14:7, 1991.

Downy mildew of sunflower (Helianthus annuus L.) is caused by Plasmopara halstedii (Farl.) Berl. et de Toni, leading to significant losses in crop production worldwide. The number of new and more aggressive pathotypes has increased rapidly over the last 10 years in Europe (Virányi et al. 2015, Bán et al. 2018), therefore, constantly monitoring the distribution of races is an important task. As part of regular surveys in June 2019, severe downy mildew was identified in some regions, appearing as chlorotic lesions along the veins of the adaxial side and white sporulation on the abaxial side of the leaves of severely stunted hybrids containing PI6 and PI7 resistance genes. The identification of the pathogen was performed microscopically based on morphological characteristics (average size of sporangia: 28x20 µm). Disease incidence (the ratio of diseased plants) ranged between 10 and 30% per field in three regions and resulted in moderate yield loss. Isolates (defined as a lesion per leaf) were collected from 4 to 8 infected leaves of each hybrid by region and stored at -70°C. Two, one and one isolates of P. halstedii were selected and characterized from the southeastern (Békés County), northern (Nógrád County) and northeastern (Borsod-Abaúj-Zemplén County) regions of Hungary, respectively. The pathotype of the four isolates was determined using the international standardized nomenclature method reviewed by Trojanová et al. (2017), including nine sunflower differential inbred lines (HA-304, RHA-265, RHA-274, PMI-3, PM-17, 803-1, HAR-4, QHP2 and HA-335). Zoosporangia from frozen sunflower leaves were washed off into bidistilled water and the concentration was adjusted to 3.5x104 sporangia/ml using a hemocytometer. Three-day-old seedlings with a radical of 1 to 1.5 cm long were immersed in the sporangial suspension and kept at 16°C overnight (Cohen and Sackston 1973). Inoculated seedlings were planted into trays containing clear moistened perlite (d = 4 mm) and grown in a growth chamber with a photoperiod of 12 h. The experiment was carried out twice with each isolate using 15 seeds/differential line with two replicates. Bidistilled water was sprayed on the plants 9 days after inoculation, and then trays were covered with a black polyethylene bag and maintained at 19°C overnight to induce sporulation. The first disease assessment was done based on cotyledons bearing white sporulation. Next, a second evaluation was performed 21 days after inoculation assessing stunting of the plants, chlorotic lesions on true leaves and damping-off. All 4 isolates examined caused disease on differential lines HA-304, RHA-265, RHA-274, PMI-3, PM-17 and HA-335, whereas the other lines showed no symptoms and signs of sunflower downy mildew. As a result, it was concluded that the presence of P. halstedii pathotype 734 was confirmed. This pathotype is likely widespread in Hungary as it could be detected from three different regions. Moreover, the possibility that pathotype 734 is present in Hungary has been raised before (Iwebor et al. 2018). This pathotype is already widespread in the USA and Russia and is considered to be highly aggressive, since it was able to infect hybrids with resistance genes PI6 and PI7 (Iwebor et al. 2018, Spring 2019). To our knowledge, this is the first report of pathotype 734 of P. halstedii in Hungary and Central Europe. Continuous monitoring and incorporation of new resistance genes into sunflower hybrids are essential steps in the future to control P. halstedii.

Downy mildew of sunflower, caused by Plasmopara halstedii (Farl.) Berl. et de Toni, is a relevant disease of this crop. High virulent pathotypes have been identified in several countries, while there are few data on the spread of P. halstedii pathotypes in some important sunflower-growing areas of Europe. The goal of this study was to give up-to-date information on the pathotype structure of P. halstedii in Hungary and provide some actual data on the virulence phenotype of the pathogen for six European countries. Infected leaves of different sunflower hybrids and volunteers were collected in seven countries (Hungary, Bulgaria, Serbia, Turkey, Greece, Romania, and Italy) between 2012 and 2019. A universally accepted nomenclature was used with a standardized set of sunflower differential lines for pathotype characterization of isolates. The virulence pattern of the isolates was determined by a three-digit code (coded virulence formula, CVF). A total of 109 P. halstedii isolates were characterized. As a result of our survey, 18 new P. halstedii pathotypes were identified in Europe. Two out of the eighteen pathotypes were detected from the Asian part of Turkey. The detailed distribution of pathotypes in Hungary is also discussed.