Tomato bioassays with pathogens and pests
Tomato seedlings were inoculated two weeks after transfer (24 h after second PA treatment) with 250 Meloidogyne incognita second-stage juveniles (J2s), unless otherwise mentioned. Roots were collected 28 days after inoculation (dai), washed and stained with 1:8-diluted Alcoferm Raspberry Red dye. After destaining in acidified glycerol, galls were counted under a stereo microscope. Rice - Meloidogyne graminicola assays were performed similarly, but plants were harvested 14 dai. N = 8, repeated at least twice.
Pseudomonas syringae DC3000 was grown on King’s B agar with 75 mg l-1 rifampicin and transferred to liquid King’s B (without rifampicin, 28°C). Cells were centrifuged, resuspended in 10 mM MgSO4 (OD600 = 0.15) and amended with 0.05% (v/v) Silwet 77. Tomato seedlings (two fully expanded leaves, 24 h after second PA treatment) were then dipped in the bacterial suspension. Seedlings were kept at 100% relative humidity (RH) 24 h before and after dip-inoculation. Five dai, leaves were detached and photographed and the number of visible specks per cm2of leaf area was counted. N = 12, repeated twice.
Bioassays with Botrytis cinerea R16 (Faretra & Pollastro 1991) were performed as in Audenaert et al. (2002) using tomato leaves from four-week old plants detached 24 h after the second PA treatment. Four dai, disease progression was monitored in a phenotyping robot as in Meng et al. (2020). N = 8 (8 plants, 32 leaflets), repeated twice.
Aculops lycopersici was cultured on tomato (S. lycopersicum ‘Moneymaker’, 28°C, 50% RH, 16:8 light/dark). Thirty adults were transferred to a 1 cm-diameter tomato leaf disk, which was placed on the second leaf of a tomato seedling (24 h after second PA treatment). Ten dai, mites were counted under a stereo microscope. N = 6, repeated twice.
Tetranychus urticae (London strain, tomato-adapted (Wybouwet al. 2015)) was expanded on common bean (Phaseolus vulgaris L. ‘Prelude’). Ten adult females were transferred to tomato plants (S. lycopersicum ‘Moneymaker’, 28°C, 50% RH, 16:8 light/dark) 24 h after second PA treatment, and ten dai offspring (mobile stages) were counted under a stereo microscope. N = 8, repeated twice.
Frankliniella occidentalis was reared on bean pods (28°C, 70% relative humidity, 14:10 light/dark). Ten synchronized adult females were transferred to three-week-old tomato seedlings (S. lycopersicum ‘Moneymaker’) in ventilated Plexiglass cylinders (90 x 35 mm). Thrips could oviposit for 24 h, after which they were removed. Only plants from which all ten adults could be recovered were retained. Four days after adult removal, offspring were counted under a stereo microscope. N = 15, repeated twice.