本文選題：朱砂葉螨　切入點(diǎn)：丁氟螨酯　出處：《西南大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：朱砂葉螨(Tetranychus cinnabarinus)是一種重要的農(nóng)業(yè)害螨,可為害豇豆、茄子、花卉等百余種農(nóng)業(yè)、經(jīng)濟(jì)作物。該螨具有繁殖力強(qiáng)、世代周期短、活動(dòng)范圍小等生物學(xué)特點(diǎn),加之殺螨劑大量不科學(xué)的使用,使其抗藥性問(wèn)題尤為嚴(yán)重。丁氟螨酯是一種新型殺螨劑,在我國(guó)上市晚,還未廣泛使用。該藥劑殺螨活性高,對(duì)非靶標(biāo)生物安全,具有成為未來(lái)主流殺螨劑的潛力。本論文通過(guò)長(zhǎng)期在室內(nèi)進(jìn)行朱砂葉螨對(duì)丁氟螨酯的抗性篩選,分析了朱砂葉螨丁氟螨酯抗性品系的抗性遺傳方式、適合度、抗性穩(wěn)定性,并評(píng)估了丁氟螨酯的抗性風(fēng)險(xiǎn),研究結(jié)果為丁氟螨酯的合理使用以及抗性治理的有效開(kāi)展奠定了理論基礎(chǔ)。本文主要研究?jī)?nèi)容與結(jié)果如下:1.以敏感品系(SS)為源頭,室內(nèi)經(jīng)過(guò)97代篩選獲得丁氟螨酯抗性品系(CyR),其LC50值由2.19mg/L上升到58.80mg/L,相對(duì)于室內(nèi)敏感品系抗性倍數(shù)為26.85倍。2.利用丁氟螨酯抗性品系與敏感品系進(jìn)行雜交和回交,明確其抗藥性遺傳形式,結(jié)果表明:F1代正交(CyR_80♀×SS♂)及反交(SS♀×CyR_80♂)所得顯性度DRS和DSR分別為-0.24和-0.32,表明朱砂葉螨對(duì)丁氟螨酯的抗性為不完全隱性遺傳;t測(cè)驗(yàn)得出正反交所得D值的95%置信限重疊,不存在顯著性差異,表明抗性基因位于常染色體上;回交F2代期望值與實(shí)際LD-P曲線及χ2值存在明顯差異,表明抗性受多對(duì)等位基因控制。3.通過(guò)構(gòu)建抗性品系(CyR)與敏感品系(SS)的生命表,逐日統(tǒng)計(jì)其生長(zhǎng)發(fā)育情況,計(jì)算生物學(xué)參數(shù),發(fā)現(xiàn)CyR品系的生殖力及存活率均低于SS品系,相對(duì)適合度(RR/SS)Rf=0.881,表明CyR品系存在一定的抗性適合度代價(jià)。4.通過(guò)從CyR_85中分出一支亞系,去除其藥劑篩選壓,研究了朱砂葉螨對(duì)丁氟螨酯的抗性穩(wěn)定性。主要以下三方面來(lái)評(píng)價(jià):首先在藥劑敏感性方面,抗性亞系(CyR_D)在停止藥劑篩選后僅過(guò)了5代,其LC50從47.34mg/L下降到11.35mg/L,抗性倍數(shù)從21.6倍下降到5.2倍,12代后抗性倍數(shù)又下降至3.8倍,表明去除藥劑壓力后,CyR_D對(duì)丁氟螨酯的藥劑敏感性逐漸恢復(fù)。其次從生理生化方面,CyR_D在停止藥劑篩選的情況下僅過(guò)了5代,三大解毒酶GSTs、MFO、CarE酶活性均發(fā)生顯著性下降,12代后MFO和CarE酶活性下降到敏感水平。表明去除藥劑壓力后,CyR_D解毒酶活性會(huì)逐漸下降。分子生物學(xué)方面,結(jié)合高通量測(cè)序及qPCR驗(yàn)證的結(jié)果,鑒定出10條可能與丁氟螨酯抗性相關(guān)的解毒酶基因,在停止篩選后的不同階段測(cè)定其mRNA表達(dá)量,發(fā)現(xiàn)TCGST-M2、TCGST-M3、TCGST-M4,TCCarE-4、TCCarE-14,CYP389C11、CYP392D6、CYP385C2,8條原本顯著性上調(diào)表達(dá)的基因在停止篩選12代后表達(dá)量較原表達(dá)水顯著性下降,2條顯著性下調(diào)的基因TCCarE-12、TCCarE-23較原表達(dá)水平顯著上升,表明去除藥劑壓力后,CyR_D解毒酶基因mRNA表達(dá)量呈現(xiàn)不穩(wěn)定性,會(huì)逐漸變化至與敏感品系相當(dāng)?shù)乃�。藥劑敏感�?解毒酶活性及相關(guān)解毒酶基因mRNA表達(dá)量三方面的結(jié)果均表明朱砂葉螨對(duì)丁氟螨酯的抗藥性不穩(wěn)定。5.應(yīng)用數(shù)量遺傳學(xué)方法估算出朱砂葉螨丁氟螨酯抗性品系的抗性現(xiàn)實(shí)遺傳力h2=0.005,表明其對(duì)丁氟螨酯的抗性發(fā)展較為緩慢�？剐燥L(fēng)險(xiǎn)評(píng)估表明,在70%-90%致死率的藥劑篩選壓力下,朱砂葉螨對(duì)丁氟螨酯產(chǎn)生10倍抗性需13-36代。通過(guò)測(cè)定朱砂葉螨丁氟螨酯抗性品系與5種不同作用殺蟲(chóng)殺螨劑的交互抗性,發(fā)現(xiàn)丁氟螨酯與甲氰菊酯、阿維菌素、聯(lián)苯肼酯、炔螨特之間無(wú)交互抗性,而與噠螨靈會(huì)產(chǎn)生明顯的交互抗性,在農(nóng)業(yè)害螨防治中應(yīng)當(dāng)避免丁氟螨酯與噠螨靈的混用、輪用等。本論文研究結(jié)果表明:朱砂葉螨對(duì)丁氟螨酯的抗性發(fā)展較為緩慢,去除藥劑壓力后,已產(chǎn)生的抗藥性會(huì)迅速衰退;多基因、不完全隱性的抗性遺傳形式及適合度代價(jià)使得抗藥性基因在種群間傳播不利,抗性種群發(fā)展慢。綜上所述,丁氟螨酯在防治朱砂葉螨時(shí)抗性風(fēng)險(xiǎn)較小。
[Abstract]:Tetranychuscinnabarinus (Tetranychus cinnabarinus) is one of the most important agricultural pest mites, can infect cowpea, eggplant, flowers and other hundreds of agricultural crops. The mites have high fecundity, short generation cycle, the scope of activities of small biological characteristics, coupled with the use of a large number of acaricides is not scientific, the drug resistance problem is serious. Ding fluorine mites ester is a new acaricide, listed in late in China, is not widely used. The chemical acaricidal activity and high safety to non target organisms, has become the mainstream of acaricide potential. This paper through indoors in tetranychuscinnabarinus screening of resistance to fluorine Ding chlorfenson, analyzes the inheritance mode, tetranychuscinnabarinus Ding fluorine mites ester resistant strains for resistance, stability, and to assess the resistance risk of Ding fluorine mites ester, the results for the rational use of Ding Fu and chlorfenson resistance management has laid a theoretical development effect The foundation. In this paper, the main research contents and results are as follows: 1. sensitive strains (SS) as the source of indoor, after 97 generations of screened mite resistant strains of Ding fluorine ester (CyR), the LC50 value increased from 2.19mg/L to 58.80mg/L, compared to the resistance ratio indoor sensitive strains was 26.85 times.2. using fluorine Ding chlorfenson resistance and susceptible strains of hybridization and backcross, the resistance genetic form, the results showed that the F1 generation (CyR_80 * SS male female) and reciprocal (SS * CyR_80 male female) the dominant degree of DRS and DSR were -0.24 and -0.32, indicated that the resistance of tetranychuscinnabarinus Ding fluorine mites ester is not completely recessive; t tests out the reciprocal values of the 95% confidence limit of the D overlap, there is no significant difference, indicated that the resistance gene is located on chromosome; and the actual LD-P curve and 2 were significantly different backcross F2 generation expectations, indicated that the resistance by multiple alleles.3. controlled by structure Construction of resistant strain (CyR) and susceptible strain (SS) of the life table, the daily statistics of its growth and development, computational biology parameters, found CyR strains of fecundity and survival rate were lower than that of the SS strain, relative fitness (RR/SS) Rf=0.881, showed that the CyR strain had some resistance to fitness costs.4. from the CyR_85 in the separation of a sub line, remove the chemical screening pressure, study the stability of resistance to Ding Zhu Shaye mite fluorine mites ester. Mainly the following three aspects to evaluate: first in the susceptibility to insecticides, resistance subline (CyR_D) in chemical screening after only 5 generations, the LC50 from 47.34mg/L down to 11.35mg/L, the resistance ratio from 21.6 times down to 5.2 times, after 12 generations of resistance ratio decreased to 3.8 times, showed that the removal agent pressure, drug sensitivity of CyR_D Ding Fu mite ester gradually resume. Secondly from the physiological and biochemical aspects of CyR_D in the case of only stop drug screening After 5 generations, three GSTs MFO, CarE enzyme and enzyme activity were significantly decreased after 12 generations of MFO and CarE activity declined to sensitive level. After that the removal agent pressure, detoxification enzyme activity of CyR_D decreased. The molecular biology, combined with high-throughput sequencing and qPCR verification results, identification 10 may be associated with resistance to fluoride Ding chlorfenson detoxification enzyme gene in different stages, stop after screening to determine the expression of mRNA, TCGST-M3, TCGST-M4, TCGST-M2, TCCarE-4, TCCarE-14, CYP389C11, CYP392D6, CYP385C2,8 had a significant increase in the expression of genes to stop screening than the original expression significantly the reduced expression of TCCarE-12 gene after 12 generations, 2 significantly reduced, compared with the original TCCarE-23 expression level increased significantly, suggesting that removal of chemical pressure, the expression of CyR_D gene mRNA detoxifying enzymes is not stable, will gradually change to sensitive strains The level of mRNA gene. Drug sensitivity, detoxification enzyme activity and detoxification expression in three aspects of the results showed that the mite resistance to mite ester unstable fluorine Ding.5. application method of quantitative genetics to estimate the resistance of reality tetranychuscinnabarinus Ding fluoride resistant strains of chlorfenson genetic h2=0.005, showed that the the Ding fluorine mites ester resistance development is relatively slow. Assessment showed the risk of resistance in 70%-90%, mortality rate of chemical screening under pressure, tetranychuscinnabarinus produces 10 times the 13-36 generation of fluoride resistant mites. Through the determination of ester Ding Ding tetranychuscinnabarinus fluoride resistant strains of mite ester and 5 kinds of different insecticidal acaricide cross resistance, Ding found fluorine mites ester and fenpropathrin, abamectin, bifenazate, no cross resistance between propargite, and pyridaben will produce significant cross resistance in agriculture, control of pest mite should avoid mixing Ding fluorine mites ester and pyridaben, Wheel etc. in this paper, the results show that the development of tetranychuscinnabarinus resistant Ding ester fluorine mites is slow, removal agent pressure, has produced resistance will rapidly decline; multi gene, genetic resistance is not completely recessive and the fitness costs of resistance gene in population spread bad resistance the population development is slow. To sum up, Ding fluorine mite ester resistance in control of t.cinnabarinus when the risk is small.

【學(xué)位授予單位】：西南大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：S433.7

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