Figure 2. Summary of the high throughput screening method.

(A). A stock solution of B. xylophilus was prepared with a concentration of approximately 10,000 nematodes/ml. An aliquot of B. xylophilus was dispensed into 96-well plates with a concentration of approximately 2000 nematodes/100 µl per each sample. An aliquot (100 µl) of diluted stock solution (200 mM) of the testing compounds was added onto each well and the treated nematodes were grown at 25°C for 6 h. After incubation, 20 µl of the sample was taken into 1.5% agar plate and the number of dead worms counted for measurement of LD50 while sitting at 25°C. If necessary, the nematicidal activity of those selected drugs (e.g., HWY-4213) was verified using cotton ball analysis (CBA). (B). The CBA was carried out as described [9]. Briefly, B. xylophilus are reacted with the drugs contained in cotton boll tips for 16 to 20 h as described [9] and then the drug-treated B. xylophilus on a plate were treated with a mold used as a feed are cultured at 25°C for 5 days. After 5-day incubation, the drug effects are assessed by measuring the reproductive rates.