本文選題：溴氰菊酯 + 秀麗隱桿線蟲　； 參考：《湖南農(nóng)業(yè)大學(xué)》2015年碩士論文

【摘要】：溴氰菊酯是常見的一種人工合成的擬除蟲菊酯,具有藥效的高效性,結(jié)構(gòu)的穩(wěn)定性、作用對(duì)象的廣泛性等特征,其毒性主要有急性毒性、亞慢性毒性、致畸、致突變性,中毒者主要表現(xiàn)為痙攣、流涎、嘔吐、四肢不調(diào)等癥狀。擬除蟲菊酯類殺蟲劑對(duì)昆蟲等生物的主要作用靶標(biāo)為電控門鈉離子通道。在前工作中我們發(fā)現(xiàn)溴氰菊酯對(duì)秀麗隱桿線蟲有較高的殺蟲活性,但由于秀麗隱桿線蟲不具備鈉離子通道,溴氰菊酯對(duì)秀麗隱桿線蟲的作用機(jī)制如何?是否作用于鈣離子通道?為此,本研究溴氰菊酯對(duì)秀麗隱桿線蟲的毒性效應(yīng),并通過實(shí)時(shí)熒光定量PCR技術(shù)檢測了溴氰菊酯對(duì)秀麗隱桿線蟲編碼鈣離子通道α1亞基egl-19、cca-1、unc-2的表達(dá)量的影響。本研究首先研究探討了不同濃度溴氰菊酯暴露下秀麗隱桿線蟲的毒性效應(yīng)。將L4期秀麗隱桿線蟲于5、0.5和0.05mg·L-1溴氰菊酯中暴露24h,以溶劑和K-m溶液為對(duì)照,分別在體式顯微鏡下觀察其后代數(shù)目、懷卵數(shù)、排卵速率、頭部擺動(dòng)頻率、身體彎曲頻率及菌落接觸率,結(jié)果發(fā)現(xiàn)溴氰菊酯暴露24h后,其后代數(shù)目、懷卵數(shù)及30min排卵的速度均有降低,其頭部的擺動(dòng)頻率和身體的彎曲頻率與對(duì)照組相比明顯下降,并且與溴氰菊酯的濃度有密切關(guān)系,5mg·L-1暴露組與對(duì)照組相比秀麗隱桿線蟲與大腸桿菌菌落接觸率明顯下降。本研究還通過運(yùn)用熒光定量PCR技術(shù),研究了50、5、0.5和0.05mg·L-1溴氰菊酯處理秀麗隱桿線蟲3、6、12、24h,然后測試其編碼鈣離子通道的α1亞基egl-19、cca-1、 unc-2相對(duì)表達(dá)量的差異,結(jié)果表明不同濃度溴氰菊酯暴露秀麗隱桿線蟲后,可以導(dǎo)致3個(gè)基因表達(dá)量的不同程度的改變,其中egl-19在暴露12h時(shí),濃度為0.05 mg·L-1上調(diào)幅度最大,為12.5倍；cca-1在暴露24 h后,濃度為50mg·L-1時(shí),表達(dá)量最高,達(dá)13.6倍；unc-2在暴露24h后,濃度為0.05mg·L-1時(shí),表達(dá)量最高,達(dá)8.27倍。綜上所述,本研究通過溴氰菊酯對(duì)秀麗隱桿線蟲的毒性效應(yīng)研究,證明了溴氰菊酯對(duì)秀麗隱桿線蟲可造成后代數(shù)目減少、懷卵數(shù)減少及排卵速率降低的生殖中毒癥狀,還可導(dǎo)致頭部擺動(dòng)及身體彎曲頻率降低的運(yùn)動(dòng)障礙和覓食缺陷,并且推測這些缺陷可能與編碼鈣離子通道的α1亞基egl-19、cca-1、unc-2有一定的關(guān)系。本研究從生理和分子水平初步研究了溴氰菊酯對(duì)秀麗隱桿線蟲的作用位點(diǎn),為進(jìn)一步探究擬除蟲菊酯對(duì)秀麗隱桿線蟲具體靶標(biāo)位點(diǎn)和研發(fā)新型殺線蟲藥劑提供一定的科學(xué)依據(jù)。
[Abstract]:Deltamethrin is a kind of synthetic pyrethroid, which has the characteristics of high efficacy, stability of structure and extensive action object. Its toxicity mainly includes acute toxicity, subchronic toxicity, teratogenicity, mutagenicity, etc. The main manifestations of the poisoning were spasms, salivation, vomiting, limb disorders and other symptoms. The main target of pyrethroid insecticides on insects and other organisms is electronic gate sodium channel. In the previous work, we found that deltamethrin has higher insecticidal activity against Rhizoma elegans, but what is the action mechanism of deltamethrin on nematode because it does not have sodium channel? Does it act on calcium channels? Therefore, the toxic effect of deltamethrin on cryptospira elegans was studied, and the effect of deltamethrin on the expression of 偽 1 subunit egl-19cca-1 unc-2 was detected by real-time quantitative PCR. In this study, the toxic effects of different concentrations of deltamethrin on cryptospira elegans were studied. The L4 nematodes were exposed to 50.5 and 0.05mg L-1 deltamethrin for 24 h. The number of offspring, the number of eggs, ovulation rate and the frequency of head swinging were observed under asana microscope, respectively, using solvent and K-m solution as control. Body bending frequency and colony contact rate. The results showed that the number of offspring, the number of pregnant eggs and the speed of 30min ovulation were all decreased after deltamethrin exposure for 24 hours, and the frequency of head swinging and bending of body were obviously decreased compared with the control group. There was a close relationship between the concentration of deltamethrin and the exposure rate of Escherichia coli in 5 mg L-1 exposure group, compared with the control group, the contact rate between C. elegans and Escherichia coli colonies was significantly decreased. In this study, fluorescence quantitative PCR technique was used to study the relative expression of 偽 1 subunit egl-19 cca-1, a 偽 1 subunit encoding calcium channel, treated with 0.05mg L-1 deltamethrin for 24 h. The results showed that different concentrations of deltamethrin could lead to the changes of three gene expression levels in different degree after exposure to Deltamethrin. After 12h exposure, the concentration of egl-19 was 0.05mg L-1, and the up-regulation was 12.5-fold after exposure to Ca-1 for 24 h. When the concentration of 50mg L-1 was the highest, the expression of 13.6 times of unc-2 was the highest, and that of 0.05mg L-1 was 8.27 times. To sum up, the toxic effect of deltamethrin on nematode elegans was studied, which proved that deltamethrin could cause reproductive poisoning symptoms of reducing the number of offspring, reducing the number of pregnant eggs and decreasing the ovulation rate. It is also suggested that these defects may be related to the 偽 1 subunit egl-19cca-1 unc-2 which encodes calcium channel. In this study, the action sites of deltamethrin on Cryptoderma elegans were preliminarily studied at physiological and molecular levels, which provided a scientific basis for further exploring the specific target sites of pyrethroids against Cryptoderma elegans and for the development of new nematicidal agents.
【學(xué)位授予單位】：湖南農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：R114

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