## 3 Results
### Geraniol Synthase Also Existed in Other Insects  
To identify whether GES was conserved in insects, the GES activity was also determined in other insects including larvae of _Ectropis oblique_ and _Euproctis pseudoconsversa_, and adult of _Cylas fomicarius_. The activity was measured with crude enzyme extracted from different insects. The monoterpene synthase assay results showed that only geraniol was produced using GPP as a substrate (Figure 6). No significant difference was observed in linalool formation between enzyme reaction and GPP self-degradation control (Figure 6). These results suggested that GES was the monoterpene synthase that existed in these insects.  
Geraniol Had in Vitro Ability to Inhibit the Growth of Endobacterial Isolated from Empoasca (Matsumurasca) onnuki  
To isolate the endobacteria in _E. (M.) onukii_, the dissected insect pieces were incubated in LB plate until the bacteria colonies appeared. Then, the grown bacterial was further purified to obtain single colony. The bacteria DNA was extracted and the 16S rRNA gene was amplified and sequenced. For preliminary identification, the 16S rRNA sequences were analyzed with BLASTn in GenBank database. Isolation A showed similarity of 99.86% with uncultured _Acinetobacter_ sp. (AB908752). Isolation B showed similarity of 99.79% with _Pseudomonas fulva_ (F)972539). Isolation C showed similarity of 99.79% with _Acinetobacter jolnsoniii_ (CP037424). In the next steps, these three 16S rRNA sequences were aligned with other sequences of different species in GenBank database. The neighbor-joining phylogenetic tree was constructed. The similar results were found in phylogenetic analysis (Figure 7). These endobacteria were also found in other insects [26; 27; 28; 29]. _Acinetobacter_ group was reported to be the major endobacteria in other insects [26]. _Acinetobacter jolnsoniii_ is a common endosymbiont in insect [27; 28]. Therefore, in our present study, isolation C _Acinetobacter jolnsoniii_ was chosen to initially analyze the potential antibacterial activity of geraniol. The results showed that geraniol could inhibit the isolation C growth in vitro. The geraniol concentration of 500 ug/mL inhibited the isolation C growth in LB medium effectively according to the OD\({}_{600}\) analysis (Table 1).  
Figure 5: Functional characterization of _E. (M.) onukii_ terpene synthases expressed in insect cell Sf9. The formation of geraniol was identified by GC-MS. The formation of geraniol is not different between EoFPPS1 and empty vector or EoFPPS2 and empty vector. The experiments were performed in three replicates with similar results. Authentic standard: Geraniol authentic standard.  
Figure 6: Evaluation of terpene synthases in other insects. GPP reacted with crude enzymes extracted from different insects. _Ectropis oblique_, larvae; _Euroctis pseudoconsperse_, larvae; _Cylas fornicarius_, adult; _E. (M.) onukii_, adult. The controls were GPP or crude enzyme only. The formation of linalool in each enzyme activity came from the GPP self-degradation. 1. Linalool; 2. geraniol. GPP CK: GPP alone in the reaction buffer; each insect CK: Crude enzyme alone in the reaction buffer. The experiments were performed in three replicates with similar results.  
The isolated C colony _Acinetobacter johnsonii_ was used for antibacterial activity assay. The experiments were performed in three replicates.