Novel approaches for the management of leaf and bud nematodes (Aphelenchoides spp) in hardy nursery stock

Leaf and bud nematodes (primarily Aphelenchoides ritzemabosi and A. fragariae) are widespread pathogens of ornamental plants grown in greenhouses and nurseries. This proposal aims to develop new approaches for the management of these nematodes in hardy nursery stock by evaluating individually, and in combination, products derived from plant extracts, currently approved pesticides, and ‘elicitors’ of natural defences in plants.

The approach to be undertaken will target three distinct phases of nematode infection: movement from soil to the plant; movement within the plant; and induction of plant resistance to infection and nematode multiplication within the plant.

Laboratory and glasshouse pot tests will initially evaluate the efficacy and persistence of a range of soil applied treatments for their contact toxicity to leaf and bud nematodes. Testing of products with systemic properties will be undertaken to determine whether soil or foliar applications can reduce nematode infestation of plants. Application of products and compounds that act as elicitors of systemic acquired resistance will be evaluated to determine whether they can induce natural defences and confer levels of resistance to nematodes, and how long this level of resistance lasts.   The mechanism underlying any induction of nematode resistance will be characterised and looked for in a range of hardy nursery stock species. Treatments that have shown promise from the glasshouse pot tests will be investigated further in similar pot tests with differing combinations of treatments and timings to refine and develop treatment programmes that will be evaluated in glasshouse trials and at commercial nurseries.

The overall aim of this project will be to develop new guidelines for the integrated management of foliar nematodes in hardy nursery stock utilising existing and novel methods.

Project aim(s):

The research in this proposal aims to target different aspects of the routes of foliar nematode invasion into plants (Fig. 1), and to characterise the defence mechanisms induced by elicitors against nematodes.

Project objectives:

Objective 1 – Nematode movement from soil into the plant. Products containing oxamyl (a standard nematicide treatment), fosthiazate (a standard nematicide treatment), spirotetramat, garlic extract, liquid mustard, furfural, neem and others (to be determined in discussion with HNS sector research manager) will initially be evaluated in the laboratory in a well plate bioassay to determine their contact mortality (after 24, 48 and 72 hours exposure) on leaf nematodes (Jagdale & Grewal, 2002). Based on the well plate bioassay results, promising candidate products will be applied as a drench to the soil at the base of plants in pots in glasshouse tests to determine their efficacy in preventing leaf nematode infection from soil. Nematodes will be extracted from infected leaf tissues by incubating in water for >48h at 25°C, and inoculated into the soil. Nematode populations in the soil and in leaves taken from the plants will be assessed prior to treatment, and at 15 day intervals up to 45 days post-treatment. Nematodes will be extracted from soil using the Baermann funnel method (Southey, 1986). Leaf symptoms will be characterised at each 15 day interval by calculating the percentage of affected leaf area, using a grid method (Fu et al, 2012). Further tests will be carried out on pot plants to determine any persistence in treatments against nematodes inoculated 25, 50 and 75 days post-treatment.

Objective 2 – Soil and foliar treatments with systemic activity to prevent nematode infection. Products that are reported to have systemic activity such as Movento (spirotetramat), NeemAzal (azadiractin A), Vydate (oxamyl) and others (to be determined in discussion with the HNS sector research manager) will be applied either as a soil treatment (NeemAzal, Vydate) or foliar treatment (Movento) to plants that have been exposed to soil containing leaf nematodes for a set period of time (to be determined from the results arising from untreated plants in Objective 1above) to simulate the early, mid and chronic stages of leaf nematode infection.
In a separate series of tests, leaf inoculations of nematodes will also be conducted on two arbitrarily selected leaves of a plant. One leaf not damaged prior to inoculation and the other damaged by one of two methods: with a scalpel, by making cuts in the upper side of the leaf, or with a needle, by making perforations scattered between the leaf veins (Fu et al., 2012). The nematode populations in the soil and in leaves taken from the plants will be assessed prior to treatment, and at 15 day intervals up to 45 days post-treatment. Further tests will be carried out on pot plants to determine any persistence in treatments against nematodes inoculated 25, 50 and 75 days post-treatment. Leaf symptoms will be characterised at each 15 day interval by calculating the percentage of affected leaf area, using a grid method (Fu et al, 2012). Photographs of each inoculated leaf will be taken and used to aid in the development of a rating key for leaf nematode damage.

Objective 3 – Foliar treatments with elicitors to induce nematode resistance. A range of commercially available products that claim to induce systemic acquired resistance will be applied to plants at a range of timings prior to the introduction of leaf nematodes to the soil or by leaf inoculation. These products will include Bion (acibenzolar-S-methyl), N-Hibit Gold (harpin), Elexa (chitosan), Synermix (seaweed extract) and others (to be determined in discussion with the HNS sector research manager). Assessment of leaf symptoms and nematode populations in the soil and in leaves taken from the plants will be assessed at 15 day intervals up to 45 days after introduction of nematodes to the soil, and by leaf inoculation as detailed above. Further tests will be carried out on pot plants to determine any persistence in treatments against nematodes inoculated 25, 50 and 75 days post-treatment.

Objective 4 – Developing a test to establish whether plants are likely to respond to elicitors. As there are a wide range of plants that can be considered as hardy nursery stock, as well as the varieties within specific plant species, it is not feasible to carry out trials on all species. However, it may be possible to develop a rapid assay that can identify the capability for different species of plants to express induced resistance when treated by elicitors. The leaves of the plants tested in Objective 3 that express induced resistance to nematodes will be analysed for changes in leaf chemistry to determine the mechanism underlying the resistance to leaf nematodes. Initially changes in levels of phenolic compounds and lignin in elicitor treated plants will be characterised to determine whether resistance to leaf nematodes can be correlated with changes in leaf phenolics or lignin content. The total phenolic content in methanolic extracts from leaves will be determined using a Folin–Ciocalteu reagent based spectrophotometric assay. There are various methods available to measure lignin content in foliage (Hatfield & Fukushima, 2005). Selected plant species will be treated with elicitors and nematode resistance assessed (as in Objective 3) and correlated with changes in leaf chemistry to validate the accuracy of the phenolic/lignin test in predicting expression of nematode resistance.

Objective 5 – Development of a nematode treatment programme. Treatments that have shown promise from the glasshouse pot tests outlined in Objectives 1-3 will be investigated further in similar pot tests with differing combinations of treatments and timings (e.g. a mix of elicitor treatments and/or soil treatments) to refine and develop treatment programmes that will be evaluated in glasshouse trials and at commercial nurseries.