Inoculation techniques and evaluation methodologies for Sclerotinia sclerotiorum head and stem rot in sunflower and soybean

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Bester, Marlese Christine
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University of the Free State
Sclerotinia stem and head rot on soybean and sunflower are caused by the fungal plant pathogen Sclerotinia sclerotiorum. This pathogen can be responsible for up to 75.0% yield loss. A major constraint in the effective control of Sclerotinia in sunflowers and soybeans is the absence of stable resistance. Cultivar selection as a management strategy remains the ultimate goal in reducing Sclerotinia outbreaks among South African producers. Quantification of cultivar responses to pathogen and environmental stimuli could assist the selection and breeding of cultivars with tolerant to high disease potentials. The production of S. sclerotiorum ascospores and sclerotia were optimised in the laboratory. Maize meal and soybean stubble as substrates for sclerotia production, produced the largest sclerotia with mean masses 0.0292 mg per sclerotium, while sunflower meal produced the lightest sclerotia with a mean mass of only 0.0099 mg per sclerotium. Several literature sources have stated that a pre-conditioning step of 4 - 10°C is needed in order for sclerotia to germinate carpogenically. Results of this study indicate that no pre-conditioning is needed for South African isolates of S. sclerotiorum to form apothecial stipes and that the optimum temperature for apothecia development is 16°C. However, results indicated that extending pre-conditioning of sclerotia at 4°C by 4 - 8 weeks, will produce a higher number of apothecia. Regression analysis indicated that apothecial stipes take approximately five days longer to develop for each week of pre-conditioning. Size of sclerotia also affected the number of apothecial stipes and sclerotia 27 mm in length produced up to 118 apothecial stipes, while the smallest sclerotia, 5 mm in length only produced 14 apothecial stipes. The viability of the pathogen in infected sunflower stems whether milled into a fine powder or shredded into pieces of various sizes, was evaluated every 30 days for 12 months under laboratory conditions. All the infected stubble sizes were able to produce both S. sclerotiorum mycelium and sclerotia on Potato Dextrose Agar after 12 months. After the optimisation of Sclerotinia inoculum, efficacy of inoculum source, application technique and timing on disease severity under greenhouse and field conditions were evaluated on both soybean and sunflower. The techniques used were liquid spray mycelium, milled grain mycelium, sclerotia planted adjacent to seedlings and an ascospore suspension. A head punch method, whereby a hole is punched into a flowering head and a S. sclerotiorum colonised sorghum grain is inserted, was also applied to sunflowers. Treatments were applied to sunflower at bud, flower and head- ripening growth stage while on soybean, the liquid spray mycelium, milled grain mycelium and ascospore suspension were applied at flowering stage. A significant difference was observed between both sunflower and soybean greenhouse and field trials. Overall the technique that yielded the highest levels of Sclerotinia disease was the milled grain mycelium. A negative correlation was observed between soybean greenhouse and field trials, while a poor correlation was observed between sunflower greenhouse and field trials. This illustrated the importance of using multiple environments when screening for Sclerotinia resistance, especially under field conditions, as well as the need to consider the high genotype x environment interactions that occur. Performing resistance evaluations in the greenhouse alone, will limit the value of screening techniques. Field evaluations of resistance in soybean and sunflower cultivars were carried out in Delmas and Greytown respectively, over several planting dates. The cultivars were inoculated with milled grain mycelium and liquid spray mycelium. Regression analyses were used to determine the response type as well as the relationship between observed stem or head rot incidence within a cultivar and disease potential. The disease potential was defined as the mean disease severity within a planting date over all the cultivars within a trial. The regression model Y=aXb was used where Y is the observed incidence within a cultivar, X is the disease potential and a and b are regression parameters. Three cultivar response types were identified based on the b parameter i.e. cultivars tolerant of increasing disease potential (b>1), cultivar intolerant to increasing disease potential (b<1) and cultivars showing a linear relationship with changing disease potential (b≈1). The Sclerotinia potential required to initiate disease onset and the rate of disease increase subsequent to onset were calculated by re- arrangement of model parameters. The soybean cultivar, PAN 1454, responded less rapidly to changing stem rot potentials, having an onset potential of 19.3% and a subsequent response rate of 0.5 per potential unit increase, making it more tolerant to increasing disease potentials, while P 64 T 39 had an onset potential of 1.0% and a response rate of 3.4 per potential unit increase, making it less tolerant of increasing Sclerotinia potential. The sunflower cultivar, P 65 LC 54, responded less rapidly to changing head rot potentials, having an onset potential of 18.9% and a response rate of 0.6 per potential unit, while P 65 LL 02 had an onset potential of 2.2% and a response rate of 1.9 per potential unit increase. The level of lignin, total proteins, chitinase and β-1, 3-glucanase in the leaves of soybean and sunflower cultivars collected at different growth stages, were evaluated to determine the correlation between these resistance components and Sclerotinia disease development. No correlation was recorded. It was concluded that the reaction of soybean and sunflower cultivars to changing disease potentials is an essential component of any resistance screening study and genotype x environment interactions must be quantified. The use of multiple criteria is essential if the behaviour of the pathosystem is to be understood and if stable resistance of commercial value is to be obtained.
Dissertation (M.Sc. (Plant Sciences))--University of the Free State, 2018, Sclerotinia sclerotiorum, Soybean, Sunflower, Inoculation techniques, Resistance screening