In the interaction between Pinus thunbergii and Bursaphelenchus xylophilus, nitric oxide (NO) is an important signaling molecule involving in the early response of P. thunbergii to the invasion of B. xylophilus. However, it is unclear that whether the NO production by P. thunbergii is triggered by the invaded B. xylophilus or its secreted metabolites. In the present study, the P. thunbergii was inoculated with living B. xylophilus, its secretion, and the suspension of grinded B. xylophilus, respectively, and the nitric oxide synthase (NOS) activity and NO content in the P. thunbergii were detected at the early stage. In all treatments, the inoculated P. thunbergii appeared disease symptoms, and the NOS activity and NO content in the P. thunbergii inoculated with B. xylophilus secretion and grinded B. xylophilus suspension increased, suggesting that besides living B. xylophilus, its contents or secretion could also trigger the expression of NO response signal in P. thunbergii, inducing the downstream response and causing the disease development of P. thunbergi. With the increasing temperature at 15-25 degrees C, both the NOS activity and the NO content in inoculated P. thunbergii increased, and the disease symptoms appeared earlier. The same patterns of NOS activity, NO content, and disease symptoms were also observed under increasing drought stress. It was suggested that within a definite range, increased. temperature and drought stress could enhance the NO signal expression in inoculated P. thunbergii and accelerate its disease development, and thus, the disease development of inoculated P. thunbergii under high temperature and drought condition could be related to the enhancement of the NO response signal in the host.