## 1 Introduction  
_Pinus densiflora_ is an evergreen conifer that grows in Northeast Asia, specifically Korea, China, Japan, and Russia. This pine tree is widely cultivated for timber plantations and landscaping. However, pine wilt disease (PWD) caused by pine wood nematode (PWN; _Bursaphelenchus xylophilus_) that extremely damages pine forests in Korea [1], Japan [2; 3], China [4], and Portugal, in Europe [5].  
Resins are generally a mixture of volatile and non-volatile compounds. It is a representative of a defense substance of conifer, and secreted by reacting to diseases, insects, and wounds [6; 7; 8]. PWN infection causes a series of physiological and biochemical changes to the pine trees. Since epithelial cells and parenchyma cells in the resin canal are used as food sources for PWNs, the epithelial cells in the pine tree are seriously destroyed, causing PWD symptoms' development [3]. In addition, PWN infection highly affects the resin secretion of the pine tree [9].  
Volatile compounds principally consist of a complex mixture of terpenoid compounds, and act as an important defense mechanism against biotic and abiotic stresses [10; 11; 12]. [PERSON] [13] reported that monoterpenes are significantly enhanced in xylem tissues long before the initiation of tracheid cavitation in pine tree after PWN infection, and eventually, the volatile terpen induced tracheid cavitation (embolism), which inhibits water transport from soil to leaves.  
3-Carene, a bicyclic monoterpene hydrocarbon consisting of a fused cyclohexene and cyclopropane rings, is a common substance in the conifer. Each conifer species has relatively different proportions of 3-carene, and the amount of 3-carene can vary drastically between the same species [14]. The amounts of 3-carene were associated with tolerance or susceptibility to insect and white pine weevil in Sitka spruce (_Picea sitchenisis_) [15; 16]. Besides, in Lodgepole pine (_Pinus contorta_), the high amounts of 3-carene correlated with the resistance to Douglas-fir pitch moths (_Synanthedon novaroensis_) [17]. Furthermore, in Scots pine (_Pinus sytbestris_), the high level of 3-carene was found to have a correlation with low larval survival of the Sawfly (_Diprion pini_) [18]. PWN infection provoked releases of pinene-type monoterpenes and expression of \(\alpha\)-pinene synthase, which were temporarily increased in _Pinus pinea_[19; 20]. In addition, a monoterpene, \(\beta\)-myrcene, appeared as the most attractive to propagative larvae of PWNs among the volatile monoterpenes present in pine, and it plays an important role in the dispersal of the of PWNs [21]. 3-Carene synthase genes were characterized enzymatically in some conifer species (_Picea sabies, Pica sitchenisis, P. bankiana,_ and _P. contorta_) [14; 22]. However, no investigation has been reported for characterization of 3-carene synthase gene and accumulation of 3-carene in _P. densiflora_ and _P. koraiensis_ against PWN infection.  
An early diagnosis of the PWN can be an important tool to reduce damage in the pine forest. In this work, we have investigated the changes of monoterpenoids of _P. densiflora_ and _P. koraiensis_ after PWN infection. 3-Carene was highly enhanced in resin and volatile compounds of _P. densiflora_ and _P. koraiensis_ after PWN inoculation. Additionally, phylogenetic analysis of 3-carene synthase (_Pd3-cars_) of _P. densiflora_, and functional characterization of _Pd3-cars_, were performed by tobacco transformation. Nematicidal and antifungal activities of 3-carene were also investigated.