Field trials

Using spreader plants

[PSD (Pesticides Safety Directorate, www.pesticides.gov.uk) Efficacy Guideline 530_ 04/07/01]

Considerations:

Artificial inoculation of spreader plants will initiate the epidemic and subsequent spread of the disease will be by natural means.

Care should be taken to ensure that trial plots are exposed to equal inoculum pressure.

The strains used to inoculate the spreader plants should reflect the natural Phytophthora infestans population.

Mist irrigation can be used to maintain favorable conditions for disease development, but care should be taken that these conditions mimic natural conditions.

Care should be taken to minimize the risk of infection spreading from trial plots to neighboring crops by desiccating infected plots as soon as practicable.

Direct inoculation

Forbes G. A., Trillos O., Turkensteen L. and Hidalgo O. 1993. Field inoculation of potatoes with Phytophthora infestans and its effect on the efficiency of selection for quantitative resistance in the plants. Fitopatología 28:117–120.

Inoculum preparation. Preparation of initial inoculum was as follows. Infected leaves taken from plants growing in the ICA station in Rionegro (Colombia) were washed and incubated at 16 +1°C overnight to induce sporulation. Sporangia were subsequently washed from the sporulating lesion in sterilized water at 15°C then repeatedly rinsed in a 15 micron filter to remove small extraneous matter including excess bacterial flora. The suspension of sporangia was them incubated a 5°C for about 2 hr to induce formation of zoospores. Zoospores were then separted from sporangia with a 15 micron filter.

The suspension of zoospores was used to produce the inoculum for the inoculation by seeding small amounts on 1 cm potato slices. Prior to slicing, potatoes were washed and surgace disinfected by dipping them in 90% alcohol and flaming. Potato slices were placed on a wire mesh in the bottom of a plastic tray, which could be hermetically sealed. A 10 micron drop of the zoospore suspension (approx. 20,000 spores/ml) was placed on each slice. No water or other source of humidity was placed in the plastic tray; sufficient moisture is supplied by the potato slices. Plastic trays were incubated at 16 +1°C for 5 days, after which the potato slices were covered with dense mycelial growth. The inoculum was prepared by submerging the slices in a bucket of well water (approx. 15°C). The inoculum was filtered through 4 layers of cheese cloth before use. Inoculum was prepared about 2 hr before inoculation to allow time to induce zoospore formation. This was achieved by placing small plastic bags with ice in the inoculum until the temperature lowered to about 9°C.

Three inoculum concentrations were initially used: low (1750 sporangia/cc), medium (2500 sporangia/cc), and high (7000 sporangia/cc).

Field inoculation. Plants were inoculated at dusk, about 7:00pm. Prior to inoculation, the field was sprayed for approximately 30 minutes with overhead irrigation. Inoculum was applied in each plot with a hand-held, manually pumped sprayer. The pressure developed in the pump is unknown and probably variable, but application rates were calibrated to approximately 20 ml per plant.

Niemira B A, Kirk W W, Douches D and Stein J M. 2001. Susceptibility of potato (Solanum tuberosum) foliage and tubers to US8 biotype of Phytophthora infestans. American Journal of Potato Research 78:319-322.

Field inoculation. Inoculation was timed for immediately before row closure. The volume of inoculum applied was calibrated to deliver approximately 1000 zoospores per plant based on the canopy density of a typical variety, e.g. Snowden. To promote disease, rainfall was supplemented with mist sprinkler irrigation to maintain high humidity within the canopy. Visual foliar disease ratings were taken every 5 to 7 days beginning immediately following inoculation until approximately 30 days after inoculation. The relative area under the disease progress curve  (RAUDPC) was calculated for each plot.