Comparison of calibration curves prepared by soil compaction and ball milling methods for direct quantification of the potato cyst nematode Globodera rostochiensis in soil

Abstract

A direct quantification method consisting of soil compaction and real-time PCR was applied to quantify the potato cyst nematode (PCN), Globodera rostochiensis, in soil. We also compared the efficacies of pre-treatment methods for disrupting nematode bodies and obtaining homogeneous soil samples between soil compaction and ball milling. PCN was detected at a density of five second-stage juveniles (J2)/20 g of soil and there were highly significant correlations (r2 > 0.8697) between the cycle threshold (Ct) values and the number of J2 inoculated in all the soils, irrespective of the pre-treatment methods. Calibration curves showing the relationship between Ct values and nematode density were almost identical for both pre-treatment methods in two soils, while the Ct values were consistently lower for the ball milling method in one soil, suggesting that DNA extraction efficiency might be better in the ball milling method. The calibration curves obtained with the ball milling method were nearly identical in three soils, but were different with the compaction method. These results suggest that the ball milling method is more suitable for direct quantification of nematodes than the compaction method.