寄主植物对小菜蛾田间种群抗药性衰退及其酯酶活性的影响

通过毒力测定和生化分析,研究小菜蛾[Plutella xylostella(L.)]田间种群对12种杀虫剂的抗性稳定性,探讨花椰菜(Brassica oleraceavar.botrytisDC)、芥兰(B.alboglabraBailey)、上海青[B.campestrisL.ssp.Chinensis(L.)M ak ino]、甘蓝(B.oleraceaKale)等4种不同寄主植物对抗药性衰退的影响,以及在此过程中小菜蛾幼虫乙酰胆碱酯酶和羧酸酯酶的活力变化.结果表明,在脱离选择压力的条件下,用花椰菜饲养4代后,小菜蛾对氯氰菊酯、杀虫双、灭多威、杀灭菊酯、毒死蜱、辛硫磷、甲胺磷抗性水平较快下降,抗性衰退比达1.65-3.68,而对阿维菌素、氟虫腈、虫螨腈、氟啶脲和刺糖菌素的敏感性变化相对较小,抗性衰退比为0.90-1.19.小菜蛾取食不同寄主植物4代后对甲胺磷敏感性变化显著,以取食上海青的小菜蛾种群最为敏感,其次为取食芥兰、甘蓝和花椰菜的种群,F1代与F5代的LC50比值分别为3.68、4.43、5.18、7.70.衰退过程中,不同种群乙酰胆碱酯酶和羧酸酯酶活力均有所下降,F5代各种群间乙酰胆碱酯酶比活力差异不显著,而羧酸酯酶比活力间则有显著差异,比活力大小顺序依次为花椰菜、芥蓝、甘监、上海青的小菜蛾种群,F1代与F5代活力比值分别为1.72、1.90、2.70、5.14;F1种群的米氏常数(Km)值明显高于F5种群,为F5种群的1.61-2.10倍,而F5代各种群变化不大,F1代种群的最大反应速率Vm ax与F5各种群相比没有明显差异.     

A Study on the Activity of Carboxylesterase and the Differential Expression of Its Gene in the Midguts of Bombyx mori Resistant to BmDNV-Z

This study was to discuss the relationship among the change in the activity of Bombyx mori carboxylesterase （BmCarE） in the midguts, the differential expression of BmCarE gene （bmcare） in the midguts, and the ability of Bombyx mori resistant to densonucleosis virus （BmDNV）, and to elucidate the molecular mechanism of resistance to BmDNV-Z. With two silkworm strains, HUABA, which is susceptible to BmDNV-Z, and BC8 （a near isogenic line of HUABA）, which is completely resistant to the same virus, as materials, the activity of BmCarE in the midgut was determined by Bio-Tek Synergy, and the differential expression of bmcare between the two strains was investigated by real-time fluorescence quantitative PCR, both at 12, 36, and 72 h post oral inoculation of the two strains with virus （hereafter referred as inoculation）. While the activity of BmCarE in the midguts of BC8 inoculation group at 12 h post inoculation was higher than that in the BC8 control group, the HUABA inoculated group, and the HUABA control group by 3.28, 2.26, and 3.02 times, respectively, with the difference being highly significant （P 〈 0.01）, there was no statistical difference among the other groups. The relative expression level of bmcare in the midguts of BC8 inoculation group at 12 h post inoculation was higher than that in the BC8 control group, the HUABA inoculation group, and the HUABA control group by 17.714, 21.76, and 15.09 times, respectively, with the difference being highly significant （P 〈 0.01）, and there was no statistical difference among other groups. The elevation of BmCarE activity and expression level of bmcare in the resistant strain at 12 h post inoculation may relate to the resistant gene （nsd/nsd） and the stimulation of BmDNV-Z. The molecular basis for the elevation of BmCarE activity in the resistant strain BC8 may be the change in the expression level of bmcare.     

不同棉花品种对棉蚜羧酸酯酶活性影响的研究

对取食不同棉花品种后棉蚜羧酸酯酶比活力测定结果表明：不同棉花品种对棉蚜羧酸酯酶活性影响较大，并且能影响棉蚜羧酸酯酶对底物的活性。棉蚜取食高抗品种后，总的羧酸酯酶活力提高，并且酶活力与单宁的含量呈不显著的正相关（ｒ＝０．７８７）。这种相关性表明棉花中的单宁的含量可能是影响棉蚜外源解毒酶活性提高的一个因素，同时棉蚜羧酯酯酶的活性也是衡量棉花抗性的一个指标。     

The effect of long-term exposure to phosphamidon on the ruminal microorganisms and circulating carboxylesterase ofBubalus bubalis

The effect of long-term exposure to the organophosphate insecticide phosphamidon on the ruminal microorganisms and serum carboxylesterase of buffalo calves was investigated. Oral administration of phosphamidon in doses of 0.25 and 0.5 mg/kg per day for 120 days caused significant inactivation of carboxylesterase activity (16–32%) without eliciting any intoxicating signs apart from mild intermittent diarrhoea in the animals receiving the higher dose. The higher dose also produced a significant reduction in the total number of rumen protozoa (16–24%). However, the insecticide had no discernible effect on the total bacterial count or pH of the rumen liquor.     

Kinetic efficiency of mutant carboxylesterases implicated in organophosphate insecticide resistance

Resistance to organophosphorus (OP) insecticides in Lucilia cuprina arises from two mutations in carboxylesterase E3 that enable it to hydrolyse the phosphate ester of various organophosphates, plus the carboxlyester in the leaving group in the case of malathion. These mutations are not found naturally in the orthologous EST23 enzyme in Drosophila melanogaster. We have introduced the two mutations (G137D and W251L) into cloned genes encoding E3 and EST23 from susceptible L. cuprina and D. melanogaster and expressed them in vitro with the baculovirus system. The ability of the resultant enzymes to hydrolyse the phosphate ester of diethyl and dimethyl organophosphates was studied by a novel fluorometric assay, which also provided a sensitive titration technique for the molar amount of esterase regardless of its ability to hydrolyse the fluorogenic substrate used. Malathion carboxylesterase activity was also measured. The G137D mutation markedly enhanced (>30-fold) hydrolysis of both classes of phosphate ester by E3 but only had a similar effect on the hydrolysis of dimethyl organophosphate in EST23. Introduction of the W251L mutation into either gene enhanced dimethyl (23-30-fold) more than diethyl (6-10-fold) organophosphate hydrolysis and slightly improved (2-4-fold) malathion carboxylesterase activity, but only at high substrate concentration.     

杀虫剂亚致死剂量对小菜蛾羧酸酯酶的影响

就阿维菌素和高效氯氰菊酯亚致死剂量对小菜蛾Plutella xylostella Linneae羧酸酯酶(CarE)活性的影响进行了研究。用亚致死剂量的阿维菌素和高效氯氰菊酯分别处理阿维菌素敏感和抗性小菜蛾,使敏感品系CarE比活力上升,抗性品系CarE比活力下降。酶动力学研究表明,两种药剂亚致死剂量处理对小菜蛾CarE与底物α-NA亲和力的影响存在差异。在敏感品系中,阿维菌素和高效氯氰菊酯处理前后CarE的Km值无明显变化;抗性品系中,用阿维菌素亚致死剂量处理后,小菜蛾体内CarE的Km值显著高于对照,即其对α-NA的亲和力比对照组明显降低,高效氯氰菊酯处理组CarE的Km值与对照相比无明显变化。     

棉铃虫不同龄期幼虫羧酸酯酶、谷胱甘肽转移酶、乙酰胆碱酯酶研究

作者报道了棉铃虫抗性种群(HJ-R)和相对敏感种群(HD-S)幼虫4个发育阶段羧酸酯酶、谷胱甘肽转移酶、乙酰胆碱酯酶的特性。随虫龄的增长,两个种群单头幼虫羧酸酯酶、谷胱甘肽转移酶的酶活力急剧增长;HJ-R种群与HD-S种群谷胱甘肽转移酶酶活力比值逐渐增大。HJ-R与HD-S种群间乙酰胆碱酯酶对杀虫剂的敏感性没有显著差异,但两种群幼虫从4日龄发育到3龄时,乙酰胆碱酯酶发生突变,对杀虫剂的敏感性显著降低。     

海南3地区瓜绢螟种群对3种药剂的敏感性及其酶系比活性的测定

采用FAO推荐的微量点滴法,测定了海南澄迈、儋州、乐东3地区瓜绢螟种群对辛硫磷、三氟氯氰菊酯和阿维菌素等药剂的敏感性;并用分光光度计终止法对3地区瓜绢螟种群的乙酰胆碱酯酶、羧酸酯酶和多功能氧化酶的比活性进行了测定和比较。结果表明,3地区瓜绢螟种群对辛硫磷、三氟氯氰菊酯和阿维菌素等药剂的敏感性已产生差异,其变化与乙酰胆碱酯酶、羧酸酯酶和多功能氧化酶的比活性的关系表现为:3地区瓜绢螟种群对三氟氯氰菊酯不敏感性的强弱顺序与瓜绢螟的羧酸酯酶比活性在3地区的高低顺序是一致的,澄迈、儋州、乐东地区瓜绢螟对三氟氯氰菊酯的LD50和羧酸酯酶比活性分别为5.088 4、3.045 4、3.382 9 μg/g和22.329 9、8.085 6、17.621 5 μmoL/(min·mg)。3地区瓜绢螟种群对辛硫磷不敏感性的强弱顺序与瓜绢螟的乙酰胆碱酯酶和多功能氧化酶的比活性在3地区的高低顺序一致,澄迈、儋州、乐东瓜绢螟对辛硫磷的LD50分别为12.543 7、7.598 6、3.811 2 μg/g,其乙酰胆碱酯酶、多功能氧化酶比活性分别为14.660 5、8.578 3、5.3804 μmoL/(min·mg)和0.168 8、0.112 1、0.294 2 μmoL/(min·mg)。但3地区瓜绢螟种群对阿维菌素的不敏感性变化与所测定的3种酶系的比活性变化趋势不一致,澄迈、儋州、乐东瓜绢螟对阿维菌素     

敌百虫对荔蝽脂肪体酯酶及其超微结构的影响

本文研究了荔枝蝽象(Tessarotoma papillosa Drury)在不同季节期间脂肪体酯酶活力及其超微结构的特点,并探讨了敌百虫对它们的影响。结果表明:1.碱性 磷酸酯酶在新羽化成虫中活力较高,为1.83微克分子/分钟,到越冬期最低,仅0.30微克分子/分钟,在生殖期活力为0.67微克分子/分钟。用敌百虫处理后对酶活力有诱导作用。2.酸性 磷酸酯酶的变化与碱性磷酸酯酶不同,从新羽化成虫经越冬到生殖期,酶活力从高逐渐降低,分别为5.7微克分子/分钟,5.47微克分子/分钟和4.8微克分子/分钟。敌百虫处理后其酶活力有所降低。3.羧酸酯酶的活力变化与碱性磷酸酯酶相似,新羽化成虫为1.48毫克分子/分钟,越冬虫为0.42毫克分子/分钟,生殖期为0.72毫克分子/分钟。敌百虫对此酶也有诱导作用。4.新羽化成虫的脂肪体亚细胞结构以液泡为主,越冬期以脂肪、精原颗粒为多,到生殖期线粒体、粗面内质网大量出现。未观察到敌百虫处理后的生理病变。本文还讨论了有关自然抗药性机理的一些问题。     

辛硫磷亚致死剂量对桑粉虱解毒酶活性的影响

【目的】研究辛硫磷对桑粉虱成虫的毒力及其亚致死剂量作用下桑粉虱成虫谷胱甘肽S-转移酶（GST）、羧酸酯酶（CarE）、乙酰胆碱酯酶（AchE）的活性变化,为桑粉虱抗药性研究及杀虫剂合理使用提供依据。【方法】采用闪烁管药膜法测定辛硫磷对桑粉虱亚致死剂量LC10、LC30、LC50,玻璃均浆器冰浴提取经过处理的桑粉虱酶液,用BIO-RAD全波长微孔板酶标读数仪测定GST、CarE、AchE。【结果】辛硫磷对桑粉虱成虫亚致死剂量LC10、LC30、LC50分别为0.0090、0.0899、0.4427μg/mL,随着辛硫磷亚致死剂量浓度增大,桑粉虱成虫GST活性显著增大、CarE及AchE活性显著下降。【结论】GST在桑粉虱对辛硫磷的解毒代谢中发挥作用。