瓜實蠅對馬拉松的抗性研究

Study on malathion resistance in Bactrocera cucurbitae

鄭喬浦(Chiao-Pu Cheng)

指導教授 ： 許如君

國立臺灣大學/生物資源暨農學院/昆蟲學研究所/碩士(2012年)

https://doi.org/10.6342/NTU.2012.10840

摘要

瓜實蠅 (Bactrocera cucurbitae) 屬於雙翅目果實蠅科，被認為是台灣重要經濟害蟲之一。馬拉松 (malathion) 為一種有機磷類化合物，在台灣自 1957 年開始被推薦用來防治田間瓜實蠅族群，但在 2002 年研究發現台灣野生瓜實蠅族群對馬拉松已經產生抗藥性。為了瞭解瓜實蠅對馬拉松的抗藥性機制，本實驗擬建立起瓜實蠅對馬拉松之抗性族群，以可造成瓜實蠅族群 50-80 % 死亡率的馬拉松藥劑逐代篩選 2011 年採自屏東九如的瓜實蠅野生族群，此族群 F1子代對馬拉松的半數致死劑量為 613 ng/fly。經馬拉松藥劑篩選 5 代之後，發現對馬拉松抗性成長 3 倍，且相較於實驗室品系具 20.9 倍抗藥性。瓜實蠅對馬拉松之抗性成長速率為 1.89，高於東方果實蠅的 0.63。交互抗性實驗顯示馬拉松抗性族群對乃力松、芬殺松、第滅寧以及芬普尼皆具有低度到高度的交互抗性。本實驗藉由協力實驗、測試馬拉松抗性有關的酵素如酯酶 (esterases) 及多功能氧化酶 (mixed function oxidases) 的活性、標的酵素乙醯膽鹼酯酶活性以及乙醯膽鹼酯酶基因序列來比較瓜實蠅馬拉松抗性品系及實驗室品系之間的差異，探討瓜實蠅對馬拉松的抗藥性機制。協力實驗顯示協力劑 piperonyl butoxide (PBO) 及 triphenyl phosphate (TPP) 對瓜實蠅馬拉松抗性品系分別具 1.3 倍及 2.1 倍的協力效果。代謝酵素活性測試顯示馬拉松抗性品系之酯酶相較於實驗室品系，對 α-naphthyl acetate 及 β-naphthyl acetate具較高之 1.6-1.9 倍活性。但多功能氧化酶對7-ethoxycoumarin 則無顯著之活性差異。此外，馬拉松抗性品系的標的酵素乙醯膽鹼酯酶顯示出較低的活性及對氧化馬拉松及乙基巴拉松較不敏感。瓜實蠅的乙醯膽鹼酯酶基因之轉譯框架區具 2031 個鹼基，在部分實驗室品系及馬拉松抗性蟲的乙醯膽鹼酯酶基因序列上在發現前端具兩個胺基酸的缺失，此外也在胺基酸 421 的位置發現具有丙胺酸 (Alanine) 取代甘胺酸 (Glycine) 的情況產生。我們的結果顯示，增加的酯酶活性及乙醯膽鹼酯酶對馬拉松的不敏感性為瓜實蠅對馬拉松產生抗藥性的可能原因。

關鍵字

瓜實蠅 ； 馬拉松 ； 協力劑 ； 代謝酵素 ； 乙醯膽鹼酯酶

並列摘要

The melon fly, Bactrocera cucurbitae (Conquillett) (Diptera: Tephritidae) is considered one of the most economically damaging pests in Taiwan. Malathion is an organophosphorus compound and has been the recommended insecticide for melon fly control since 1957. In 2002, malathion resistance was found in wild melon fly populations in Taiwan. In order to understand the mechanisms of malathion resistance in the melon fly, we established our resistant strain in this study using malathion dosages that caused 50-80 percent mortality in melon fly populations collected from Pintung in 2011, the F1 generation’s LD50 value is 613 ng/fly to malathion. The resistance increased by 3-times after 5 generations of selection, resulting in 20.9-times more resistance than the laboratory strain. The resistance development rate was 1.89 higher than the values of B. dorsalis (0.63). Cross resistance experiments indicate malathion-resistant strain has low to high level cross resistance to naled, fenthion, deltamethrin and fipronil. The mechanism basis of resistance to malathion in the melon fly (resistant and laboratory) strains were examined by synergist tests, comparing the activities of metabolic enzymes known to be involved in malathion resistance, such as esterases (ESTs) and mix function oxidases (MFOs), the target enzyme acetylcholinesterase activity and ace gene molecular assay. Synergism tests indicated that piperonyl butoxide (PBO) showed 1.3-fold synergistic ratio and 2.1-fold triphenyl phosphate (TPP) on the malathion-resistant strain. Metabolic enzymes activity showed that the malathion-resistant strain exhibited higher activity, 1.6-1.9 fold in esterase (by α-naphthyl acetate and β-naphthyl acetate as the substrates), compared to the laboratory strain. However, no significant differences were found on the MFOs activity (by 7-ethoxycoumarin as the substrates) between two strains. Moreover, the target enzyme acetylcholinesterase (AChE) from the resistant strain showed reduced AChE activity and more insensitivity to inhibition of malaoxon and ethyl-paraoxon. The opening reading frame of B. cucurbitae ace gene has 2031 base pairs, two amino acid deletion and one point mutation G421A were found in ace gene of some laboratory and resistant individuals. Our results indicated that increased ESTs efficiency and AChE insensitivity to malathion were possible mechanisms involved in malathion resistance.

並列關鍵字

Bactrocera cucurbitae ； malathion ； synergists ； metabolic enzyme ； acetylcholinesterase

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