Cold tolerance and silencing of three cold-tolerance genes of overwintering Chinese white pine larvae.

The values of physiological indices and the enzymes activities involved in the overwintering stage were studied in D. armandi larvae in each month from October 2014 to March 2015. The sorbitol, trehalose and glycerol values initially tended to increase as the ambient temperature decreased, before declining until the end of the winter. The activities of four enzymes (SOD, CAT, LDH and AchE) decreased, whereas POD, PK and MDH showed opposite trends in activity. Other enzyme activities (those of TPS, SDH and GLK) were low during the overwintering period and later increased and stabilized during spring. In this study, a polymerase chain reaction (PCR) genes of SDH, TPS and GLK was utilized to identify DarmSDH, DarmTPS and DarmGLK in D. armandi. They were found to be abundantly expressed during the overwintering stage by quantitative real-time PCR (qRT-PCR) analyses; by contrast, these three genes showed higher expression levels in December 2014 than in May 2015. The qRT-PCR results demonstrated that the reduction of mRNA expression levels was significant in DarmSDH-, DarmTPS- and DarmGLK-dsRNA-treated D. armandi compared with water-injected and non-injected controls. The mortality responses at low temperature were also increased in the dsRNA-treated D. armandi compared with the controls.