# Proteomic Evaluation of Insecticidal Action of Phosphine on Green Peach Aphids, _Myzus persicae_
## 3 Results  
### Susceptibility of M. persicae under PH\({}_{3}\) Treatments and Biochemical Changes  
Lethal concentration values due to PH\({}_{3}\) treatment are displayed in Figure 1. There were no dramatic changes in LCT values between developmental stages of _M. persicae_. However, adults _M. persicae_ were slightly more susceptible to PH\({}_{3}\) fumigation than aphid nymphs.  
One of the known target sites of PH\({}_{3}\) fumigation in insect pests is COX; its biochemical activity was measured using a substrate, cytochrome c, after the PH\({}_{3}\) treatments, corresponding to the LCT\({}_{10}\) and LCT\({}_{50}\) values in comparison with that of the control (Figure 2A). Cytochrome c oxidase activities in _M. persicae_ nymphs were not affected by PH\({}_{3}\) treatment but decreased in _M. persicae_ adults as PH\({}_{3}\) concentrations increased. These changes were statistically different, but they did not increase by up to 2-fold. The effects of PH\({}_{3}\) fumigation on the biochemical activities of another well-known target of organophosphorus insecticides such as malathion and fenitrothion, i.e., AChE, was also measured in _M. persicae_ using acetylcholinesterasehiocholine iodide (ATChI). There was no effect of PH\({}_{3}\) treatments on AChE (Figure 2B) probably because its expression was not related to PH\({}_{3}\) toxicity in _M. persicae_.  
Expression levels of six genes in _M. persicae_: cytochrome P450 (CYP6 CY3) (_CYP6 CY3_), farnesyl diphosphate synthase 1 (_fps1_), acetylcholinesterasehiochinesterase (_ache_), voltage gated sodium channel subunit 1 (_para_), nicotinic acetylcholine receptor alpha 3 (_ni-acr_), and ecdysone receptor (_ecr_), were determined in response to PH\({}_{3}\) fumigation (Figure 3). In _M. persicae_ nymphs, _ni-acr_, _fps1_, _para_, and _ecr_ genes were upregulated after PH\({}_{3}\) treatments (Figure 3). Similarly, in adult _M. persicae_, _ni-acr_ and _ecr_ were upregulated, whereas _fps1_ gene was down-regulated after PH\({}_{3}\) treatments.  
The expression of the tested genes did not increase by 2-fold in comparison with that in the control, but their expressions were statistically different in the two concentrations.  
Figure 1: Effects of phosphine fumigation on the different developmental stages of _Myzus persicae._ Lethal concentration time (LCT) values are obtained using Equation (1). More details in mortality are shown in Figure S1 and Table S2.  
Figure 3: Gene expressions in the phosphine-treated _M. persica_ determined by RT-qPCR. Six gene primers are listed in Table S1. Different letters on the bars indicate statistical differences between phosphine-treated samples and the control (\(p<0.05\)). Statistic differences were tested using one-way ANOVA and Tukey’s test as a post-hoc test. Treated concentrations were equivalent to LC\({}_{10}\) (0.055 mg/L for nymphs or 0.056 mg/L for adults) and LC\({}_{50}\) (0.130 mg/L for nymphs or 0.120 mg/L for adults) values.  
Figure 2: Enzyme assays in the two different developmental stages of _Myzus persica_ (nymphs and adults) after the phosphine treatment. (**A**) Cytochrome c oxidase (COX); (**B**) Acetylcholinesterasees (AChE). The enzyme activities were expressed as unit/mg of the insect. Different letters on the bars indicate statistical differences between phosphine-treated samples and the control (\(p<0.05\)). Treated concentrations were equivalent to LC\({}_{10}\) (0.055 mg/L for nymphs or 0.056 mg/L for adults) and LC\({}_{50}\) (0.130 mg/L for nymphs or 0.120 mg/L for adults) values.