The metabolism of p,p′-DDT by the feral pigeon (Columba livia)

Male feral pigeons were dosed with ring-labeled $\text{[}{}^{14}\text{C}\text{]p,p′-}\text{DDT}$ and the tissues and droppings analyzed for total ¹⁴C, extractable ¹⁴C, and metabolites. Only 16% of an intraperitoneal dose of 1.5–2.2 mg kg^?1 was voided in the droppings over 28 days; the rate of loss reached a maximum on the 14th day and then fell quickly away. The rate of removal of ¹⁴C in droppings was low in comparison to that found in the rat and the Japanese quail. When pigeons were dosed with 32–38 mg kg^?1 DDT per bird, and killed after 77 days, 5.4% of the dose was eliminated in droppings and 87% was recovered in the body. The tissues and droppings from this experiment were analyzed for DDT and its metabolites. Of the ¹⁴C remaining in tissues 88% was accounted for as the apolar compounds DDE, DDT, and DDD. Approximately half of the ¹⁴C in droppings was present as DDE, DDT, and DDD, whereas 27–35% was apparently in conjugated form, extractable from aqueous solutions by ethyl acetate after prolonged acid hydrolysis. Two polar metabolites were isolated from the acid-released material. One was p,p′-DDA; the other was extractable from aqueous solution at pH 8 and was tentatively identified as a monohydroxy derivative of p,p′-DDT. DDE accounted for 93% of the ¹⁴C present as metabolites in tissues and droppings, clearly indicating the importance of this intermediate in this study. The metabolism of DDT in the feral pigeon is discussed in relation to its metabolism by other species.     

The postmortem reductive dechlorination of pp′DDT in avian tissues

Pigeons, Bengalese finches, and Japanese quail were dosed with pure pp′DDT, and their tissues were examined for residues immediately after death, or following storage. DDD residues in livers extracted immediately after death were always low (<5% DDT concentration) and sometimes undetectable. Reductive dechlorination of DDT to DDD occurred postmortem, relatively repidly at 20°C (90% after 8 hr), but more slowly at ?10 and ?20°C. The conversion could not be attributed to bacterial action and was evidently due to processes operating within the cell under anaerobic conditions. During storage at ?12 to ?15°C, reductive dechlorination occurred in brain and in embryos, but did not occur to any significant extent in depot fat or in eggs without embryos. These findings raise questions about the importance of DDD as an in vivo metabolite in birds and other vertebrates. The interpretation of DDD residues in tissues from laboratory and field studies is discussed.     

Cuticular penetration of β-cypermethrin in insecticide-susceptible and resistant strains of Bactrocera dorsalis

Semi-thin sectioning and transmission electron microscope techniques were employed to investigate the cuticle thickness, integument structure, and fat body of larvae from susceptible and resistant strains of Bactrocera dorsalis. The results showed that the cuticle of β-cypermethrin-resistant strains (25.96 ± 1.00 μm) was thicker than that of susceptible strains (19.36 ± 0.82 μm). The number of chitin layers in the endocuticle of β-cypermethrin-resistant strains (98.00 ± 3.61 layers) was more than that in susceptible strains (75.67 ± 2.40 layers). Compared with susceptible strains, the laminated structure of the chitin layers in the endocuticle of resistant strains revealed higher density and more distinctive structure, and the interspace of epidermal cells was thicker. Fat body in the resistant insects contained more fat granules than those in susceptible insects. Moreover, HPLC analysis showed that the cuticular penetration of β-cypermethrin into larvae of resistant strains was slower than that of susceptible strains. In addition, the metabolism of β-cypermethrin in resistant strains was faster than that in susceptible strains, indicating that the resistant strains could enhance detoxification metabolism. These results indicated that cuticle thickness, fat body, laminated structure of the chitin layers, and interspace of epidermal cells might be correlated with cuticular penetration between susceptible and resistant strains, suggesting that the resistant strains could decrease the rate of penetration of insecticide into the internal cavity.     

Investigation of insecticidal activity of rye α-amylase inhibitor gene expressed in transgenic tobacco (Nicotiana tabacum) toward cotton boll weevil (Anthonomus grandis)

Innumerable proteinaceous α-amylase inhibitors have been isolated and identified from different plant species. Among them, an α-amylase inhibitor gene with bioinsecticidal potential toward Anthonomus grandis (cotton boll weevil) was previously identified in rye seeds (Secale cereale). This cereal inhibitor was expressed in tobacco plants (Nicotiana tabacum) under control of phytohemaglutinin promoter by using Agrobacterium tumefasciens - mediated transformation. Presence of αBIII-rye gene and further protein expression were confirmed by PCR and Western blot analysis, respectively. Immunological assays indicated that the recombinant inhibitor was expressed in concentration range from 0.1% to 0.28% (w:w) of the total protein in tobacco seeds of R₀ plants. From 14 independent transformants, five plants with expression levels between 0.20% and 0.28% in seeds were in vitro assayed against A. grandis amylolytic enzymes causing clear inhibition. Moreover, bioassays using transgenic seed flour mixture for artificial diet produced 74% mortality in A. grandis first larval instar. These data suggest that rye inhibitor could be a promising biotechnological tool for produce transgenic cotton plants with an increased resistance to cotton boll weevil. Moreover, αBIII-rye gene should be considered a potential compound for a pyramiding strategy aiming to delay insect-resistance.     

α -三联噻吩对红火蚁Solenopsis invicta觅食行为和触角功能的影响

通过研究光活化杀虫活性成分 α -三联噻吩( α -T)对红火蚁 Solenopsis invicta Buren工蚁觅食行为和触角识别功能的影响,探讨了光活化成分用于控制红火蚁的可行性。用Potter喷雾法将100 μg/mL的 α -T丙酮水溶液喷施于红火蚁中型工蚁体表,再经紫外(UVA)光照射30 min,在处理后5、10、15、20和30 min时,试蚁对火腿肠的识别率分别为20.00%、25.00%、38.33%、40.00% 和41.67%,对杀虫剂饵料的识别率分别为3.33%、0、0、3.33%和0,聚集率分别为17.50%、38.33%、 32.50%、45.00%和48.33%;经100 μg/mL的 α -T涂抹试蚁触角再经紫外光照处理30 min后,触角对丙酮反应的电位值为-0.177 mV,可正常行走的工蚁比率为75.00%;以100 μg/mL的 α -T涂抹试蚁触角并经紫外光照射30 min后10 h,红火蚁的死亡率为27.50%;上述结果均与 α -T黑暗处理、对照(CK)光照处理和CK黑暗处理结果差异显著。 α -T对红火蚁工蚁食物识别和探路行走能力均具有良好的光活化抑制作用,以其防治红火蚁具有良好可行性。     

华北大黑鳃金龟Holotrichia oblita中肠羧酸酯酶基因 HoCL1的克隆与序列分析

利用粉纹夜蛾(Trichoplusia ni)围食膜蛋白多克隆抗体,从已构建的华北大黑鳃金龟 Holotrichia oblita 中肠cDNA表达文库中筛选得到1个编码羧酸酯酶的cDNA克隆 HoCL1 ,其开放阅读框(ORF)长1 599 bp,编码532个氨基酸,推导的蛋白质分子质量为59.5 kDa,等电点(p I)为4.5。 HoCL1蛋白具有羧酸酯酶的保守结构域:1个二硫键形成的位点和1个丝氨酸活性中心,三联体催化活性中心位于Ser207、Asp333和His422上,不含有氮联糖基化位点和氧联糖基化位点,只含有3 个半胱氨酸残基。依据氨基酸序列同源性分析和保守结构域分析,HoCL1属于B类酯酶,与赤拟谷盗 Tribolium castaneum 羧酸酯酶相似性最高,为35.2%。通过与其他昆虫羧酸酯酶序列比对及构建系统发育树,发现HoCL1羧基端的氨基酸序列保守性低,但靠近N端的活性中心处的氨基酸序列则高度保守,可与赤拟谷盗、异色瓢虫 Harmonia axyridis 羧酸酯酶聚类在一起。羧酸酯酶 HoCL1 基因的克隆鉴定为进一步研究该基因在华北大黑鳃金龟体内的表达及功能奠定了基础。     

苹果炭疽叶枯病病原Glomerella cingulata及其侵染过程

为明确苹果炭疽叶枯病病原菌围小丛壳Glomerella cingulata的侵染致病特征,在分离获得该病原菌的基础上,采用形态学观察、ITS序列分析和致病性测定对其进行了鉴定,并利用光学和扫描电子显微镜对病原菌在嘎啦苹果叶片上的侵染过程进行了研究.结果表明,在陕西咸阳地区分离获得的9株病原菌均为围小丛壳G.cingulata.25 ℃下接种9 h后,分生孢子中间产生隔膜,双胞化,并萌发产生芽管和附着胞;24 h后分生孢子的2个细胞均可萌发并形成芽管及附着胞,部分芽管顶端可产生次级分生孢子;48 h后次级分生孢子萌发形成附着胞;72 h后,附着胞下形成的侵染钉可直接入侵寄主,在表皮细胞内形成初生菌丝和次生菌丝,此时叶片表面已出现褐色斑点.接种7 d后叶片病斑处出现分生孢子盘和子囊壳.表明陕西省近年出现的苹果炭疽叶枯病病原菌为围小丛壳G.cingulata,该病菌在嘎啦叶片上的一些特殊侵染行为可能是导致该病害易在短时间内暴发的重要原因.     

白背黄花稔Sida rhombifolia的生物活性及化学成分研究

用石油醚和乙酸乙酯分别对白背黄花稔 Sida rhombifolia L.地上部分的乙醇提取物进行萃取,得不同萃取相;测定了乙醇提取物及不同萃取相对白纹伊蚊 Aedes albopictus Skuse 4龄幼虫和福寿螺 Ampullaria gigas Spix的毒杀活性。结果表明,乙醇提取物和乙酸乙酯相对白纹伊蚊4龄幼虫的杀虫活性明显,而石油醚相和乙醇提取物对福寿螺的杀螺活性较好。对乙酸乙酯相进一步分离得到11个化合物,经核磁共振氢谱(1H NMR)和碳谱(13C NMR)鉴定其分别为已知化合物:间-羟基苯甲酸(1)、棕榈酸1-甘油酯(2)、丁香酸(3)、没食子酸(4)、 β -谷甾醇(5)、豆甾醇(6)、东莨菪内酯(7)、山奈酚(8) 、5-羟基-7,4'-二甲氧基二氢黄酮(9)、3- O - β -D-葡萄糖-(6- O -对羟基反式香豆酰基)-山奈素苷(10)和 β -谷甾醇-3- O -葡萄糖苷(11)。其中化合物 2 对白纹伊蚊4龄幼虫具有较好的杀虫活性,24 h和48 h的LC50值分别为28.66和13.50 μg/mL。     

棉铃虫Nanchung基因的全长cDNA克隆及序列分析

昆虫的Nanchtmg(Nan)基因是瞬时感受器电位香草素受体亚家族(transient receptor potential vanilloid,TRPV)通道蛋白的编码基因.有研究证实,Nan基因的缺失会造成昆虫听觉的完全丧失[1].目前,仅少数昆虫的Nan基因全序列被克隆[2-3].作者利用反转录-多聚酶链式反应(RT-PCR)和cDNA末端快速扩增技术(RACE)对棉铃虫Nan基因完整的cDNA进行了克隆、序列分析和同源性比较.     

小麦抗白粉病基因Pm2的SSR标记筛选

为筛选与小麦抗白粉病基因Pm2紧密连锁的分子标记,将感病品种Chancellor与Pm2的近等基因系杂交,获得F1、F2分离群体,采用分离群体分组法对Pm2进行了微卫星(microsatellite,又称simple sequence repeats,SSR)标记分析.结果表明,定位于小麦5D染色体上的71对SSR引物中有12对引物能在Pm2的近等基因系、Chancellor间稳定地揭示出多态性差异,7对引物Xcfd189、Xcfd29、Xcfd8、Xcfd102、Xcfd7、Xcfd57和Xgwm190分别能在抗病、感病池间和F2分离群体的抗病、感病单株间稳定地扩增出特异性产物.7对引物所扩增的特异谱带分别为:Xcfd189360、Xcfd29190、Xcfd8160、Xcfd102250、Xcfd7200、Xcfd57245和Xgwm190210,它们与Pm2基因间的遗传距离分别为0、1.5、2.3、5.4、10.2、31.5和54.3 cM,其中标记Xcfd189360与Pm2共分离,标记Xcfd29190、Xcfd8160和Xcfd102250与Pm2紧密连锁,可用于Pm2的标记辅助选择.     

阿维菌素对黄胸散白蚁Reticulitermes speratus的毒效

在室内采用毒土法、滤纸法和毒土柱法测定了0.2%阿维菌素乳油对黄胸散白蚁的毒杀效果。 结果表明,阿维菌素对黄胸散白蚁的触杀作用速度较慢,在处理的96 h内,随时间推移毒性逐渐增强;黄胸散白蚁接触或取食药剂96 h后,其LC50值均低于20 mg/L;在供试条件下,用2.5 mg/L以上的浓度处理土壤,即能阻止黄胸散白蚁在土柱中的穿透,显示阿维菌素是一种值得进一步研究的白蚁预防药剂。     

转cry1Ab/cry2Aj玉米对亚洲玉米螟的抗性评价

为探讨转cry1Ab/cry2Aj玉米对亚洲玉米螟Ostrinia furnacalis(Guenée)的抗性,采用室内生测和田间人工接虫鉴定方法评价了6个转cry1Ab/cry2Aj玉米品系对亚洲玉米螟的杀虫效果,并利用酶联免疫吸附法(ELISA)测定了Cry1Ab蛋白在各品系主要组织的表达水平.结果显示,亚洲玉米螟幼虫取食转基因玉米各品系雄穗、苞叶、花丝和雌穗48 h后死亡率在90％左右,取食心叶的死亡率在70％左右,96h后大部分幼虫死亡,而取食非转基因玉米的死亡率为3.3％～8.7％;春播和夏播田转基因玉米叶片、茎秆、雌穗基本没有被害,而对照玉米叶片、茎秆、雌穗被害严重,春播和夏播玉米平均每株分别有2.9、2.3头幼虫和蛹,2.8、3.4个蛀孔,隧道长度为7.5、14.2 cm,雌穗被害级别为5.3和5.1;6个转基因玉米品系中Cry1Ab蛋白在心叶、雄穗、花丝、苞叶和雌穗中稳定表达.综合评价认为,转基因玉米各品系在整个生育期内对亚洲玉米螟有很好的抗性,其中N50品系抗虫效果最佳,可以作为抗虫转多基因玉米育种的备选材料.     

转cry1Ac玉米对亚洲玉米螟的抗性评价

目标性状的有效性评价是转Bt基因抗虫玉米研发和安全性评价研究的重要内容之一。采用酶联免疫吸附剂测定（ELISA）、室内生测和田间人工接虫鉴定研究了转cry1Ac玉米纯合群体BT-X和分离群体BT-38、BT-181、BT-43、BT-105等转化事件Cry1Ac蛋白的表达量以及对亚洲玉米螟Ostrinia furnacalis（Guenée）的室内杀虫和田间抗虫效果。转cry1Ac玉米鲜组织中Cry1Ac蛋白含量在44.07～438.00 ng/g之间,不同转化事件及不同组织之间差异显著,其中BT-X鲜花丝中Bt蛋白表达量显著低于未展开心叶。室内生测试验中除BT-43与对照没有显著差异外,取食其它转基因玉米心叶的亚洲玉米螟初孵幼虫6或7天的存活率为0～15.3%,显著低于非转基因对照;取食夏播BT-X鲜花丝6天的幼虫存活率为62.5%,显著低于Bt11及非转基因对照。田间心叶期接虫鉴定显示BT-X高抗亚洲玉米螟。说明转cry1Ac玉米BT-X、BT-181、BT-38和BT-105对亚洲玉米螟有很高的杀虫作用,BT-X田间抗虫效果良好。     

不同抗逆诱导剂对水稻细胞PBZ 1 基因表达的影响

以水稻品种中花8号的悬浮细胞为试材,分别研究了真菌激发子和盐激发子对用稻瘟灵(IPT)和脱落酸(ABA)预处理24 h的水稻悬浮细胞中PBZ 1 基因表达的影响。结果发现:单独用IPT、ABA处理可以诱导细胞中PBZ 1 基因表达;只用真菌激发子和盐激发子处理的细胞中也可以检测到PBZ 1 基因的表达,但两种激发子对PBZ 1 基因表达的影响不同,在真菌激发子处理后的0~3 h之间PBZ 1 基因表达逐渐增强,而在盐激发子处理后的0~3 h之间则呈现逐渐减弱的趋势,到第3 h就检测不到了;用IPT预处理24 h后,再分别用真菌激发子和盐激发子诱导,水稻细胞中PBZ 1 基因表达强度都显著高于单独用IPT处理的细胞,而且在处理后的第0.5~3 h期间一直保持较强的表达;ABA＋盐激发子处理使细胞中PBZ 1 基因表达推迟且短暂,只在处理后第1~2 h之间可检测到,ABA＋真菌激发子处理使细胞中PBZ 1 基因在0.5~3 h期间一直保持较强的表达,明显强于单独用ABA处理的细胞。可见,IPT和ABA都可以诱导抗病基因PBZ 1 的表达,PBZ 1 基因也可由盐激发子诱导表达,说明PBZ 1 基因对除病害以外的非生物逆境也有积极的作用。     

南方根结线虫Col基因分子克隆及其VIGS效应分析

为了明确线虫角质层胶原蛋白基因(Col)与根结线虫病害的关系,利用从线虫基因组中预测的Col设计特异引物,克隆了南方根结线虫Meloidogyne incognitacol基因MiCol.该基因完整编码区长903bp,编码300个氨基酸.克隆的MiCol与预测的MiCol基因序列一致性高达99.67％,氨基酸序列一致性高达100％.利用病毒介导的基因沉默技术,将其导入番茄植株并接种南方根结线虫,60 d后,沉默载体pTV-MiColi处理番茄植株的根结数比空载体对照及清水对照分别减少49.4％和49.2％,表明MiCol基因沉默显著降低了南方根结线虫的侵染数量.     

转cry1Ab基因玉米对粘虫的抗性评价

粘虫Mythimna separata(Walker)是我国玉米上的重要害虫。转Bt基因玉米为防治粘虫提供了新的途径。从室内生测和田间抗性鉴定分别研究了转cry1Ab基因抗虫玉米YieldGard对粘虫的杀虫效果。用4~7叶期转cry1Ab基因玉米心叶室内饲养1~3龄粘虫幼虫3、7和10天的死亡率分别为39.0%~92.0%、96.0%~98.0%和97.0%~100%,第14天全部死亡,而对照的死亡率分别为3.0%~8.0%、6.0%~20.0%和11.0%~28.0%,差异极显著。取食转cry1Ab基因玉米的幼虫不能正常生长发育,存活10天的幼虫其体重仅为取食对照玉米的9.0%。田间玉米4~5叶期,分别接1~6龄粘虫幼虫于转cry1Ab基因和非转基因玉米心叶。转cry1Ab基因玉米上各龄期幼虫平均存活率仅为0.0~6.7%,极显著低于非转基因玉米的8.3%~80.0%。转cry1Ab基因玉米的危害级别0.1~0.7,显著小于非转基因玉米的2.1~7.3。结果说明转cry1Ab基因玉米YieldGard对粘虫各龄幼虫有很高的杀虫作用和田间控制效果,同时能有效保护玉米免受危害。     

鬼臼毒素4β含硫衍生物的 合成及其拒食活性

为研究鬼臼毒素类化合物结构与杀虫活性的关系,在前期研究基础上设计合成了3个4β-硫酯-4-脱氧鬼臼毒素、2个4β-硫醚-4-脱氧鬼臼毒素及4β-巯基-4-脱氧鬼臼毒素共6个未见文献报道的化合物。其结构经IR、MS、1H NMR和元素分析确证。采用小叶碟添加法测试了化合物对3龄粘虫Mythimna separate幼虫的拒食活性,结果表明:4β-巯基-4-脱氧鬼臼毒素的活性最高,24和48 h 的AFC50(拒食中浓度)分别为164.6 和248.1 mg/L,分别是母体化合物鬼臼毒素活性的5.8和 3.0倍。引入硫醚及硫酯后活性均降低。     

Wolbachia共生对丽蚜小蜂生殖和适应性的影响

通过喂食合有盐酸四环素的蜂蜜水抑制丽蚜小蜂体内Wolbachia的活性,并研究以烟粉虱为寄主时Wolbachia内共生菌对丽蚜小蜂生殖力、寿命和后代性别的影响.结果表明:抗生素处理后丽蚜小蜂产生雄性后代,雄性后代所占比例与抗生素浓度呈显著正相关(r=0.322,P<0.01).在1mg/mL抗生素浓度下丽蚜小蜂产生的寄生蛹数(黑蛹数)为68.1±8.9,较对照的49.1±4.5显著增加;而雌成蜂寿命与对照无显著差异(抗生素处理15.56 4-1.55,对照12.50±0.88),且从第4天开始产生的后代全部为雄性.说明Wolbachia的共生对丽蚜小蜂的生殖力具有一定的负面影响,Wolba-chia调控宿主生殖的强度与其密度有关.