Immunocytochemical localization of the Bacillus sphaericus binary toxin components in Culex quinquefasciatus (Diptera: Culicidae) larvae midgut     

Maria Helena N.L. Silva-Filha  Christina A. Peixoto 《Pesticide biochemistry and physiology》2003,77(3):138-146

This study describes the immunocytochemical localization of the Bacillus sphaericus 2362 binary toxin components, BinA and BinB, in Culex quinquefasciatus larvae that had been intoxicated with this entomopathogen. Ultrathin sections of C. quinquefasciatus larvae midgut embedded in the hydrophilic resin L.R. White were incubated with the antibodies anti-BinA or anti-BinB and then revealed with goat anti-rabbit IgG coupled to gold particles. Immunocytochemical detection demonstrated the presence of specific labeling in ultra-thin sections that had been incubated with the BinA antiserum. Gold particles were detected on the apical areas of cell membranes and inside the epithelial cell cytoplasm, particularly the mitochondria, of cells from the gastric caeca and posterior stomach in larvae exposed during 2 or 24 h to the entomopathogen. A similar labeling pattern was observed in ultrathin sections from both midgut regions when incubated with BinB antiserum.  相似文献   

致倦库蚊天蚕素CecB2基因的原核表达及蛋白纯化     

王吉平  张堤  张春林  赵文静  张晶  翟素珍 《动物医学进展》2016,(11):31-35

构建致倦库蚊(贵阳株)天蚕素B2(CecB2)基因原核表达载体,并原核表达获得重组蛋白。定向克隆CecB2成熟肽序列至原核表达载体pET32a(+)上,将成功构建的pET32a-CecB2重组表达质粒转化大肠埃希菌Rosetta中经IPTG诱导表达,对IPTG诱导浓度和诱导时间优化后表达所得目的蛋白采用镍离子亲和层析纯化,SDS-PAGE和Western blot检测鉴定表达蛋白。结果表明,成功构建原核表达载体pET32a-CecB2,IPTG浓度和时间优化结果为IPTG浓度为0.05mmol/L,诱导时间为3h。SDS-PAGE检测获得大小约25ku的可溶性纯化蛋白,Western blot鉴定纯化蛋白可与鼠抗His-tag单克隆抗体发生抗原抗体结合反应。说明所构建原核表达载体pET32a-CecB2能在大肠埃希菌中可溶表达,为进一步研究其生物学功能奠定了基础。  相似文献   

稻田放养满江红对蚊虫孳生影响的初步观察     

胡玉凤  李红梅  郭石山  许荣满  陆宝麟 《中国生物防治》1989,(3)

实验室笼内研究表明,容器水面覆盖细绿萍可阻止库蚊在其中产卵,但对中华按蚊产卵无显著作用。1987年在湖南桂东县对放养细绿萍的稻田观察结果,库蚊(主要是三带喙库蚊)幼虫密度明显比对照田为低,而且随着覆盖面积比例扩大而下降率愈大,当覆盖面积达到95％时,平均下降率达78.96％,但对按蚊幼虫密度下降率甚微,这与实验室观察结果相符。在桂东县,养萍稻田每亩可收鲜萍1500-2000公斤,可用作禽畜饲料或其他作物基肥。养萍稻田也可使稻谷增产8％左右。由于养萍既可减少库蚊幼虫孳生,也有较好的经济效益,是在稻田蚊虫综合治理中值得采用的方法之一。  相似文献   

Synthesis and insecticidal activities of pyridine ring derivatives of podophyllotoxin     

Xudong Di  Yanqing Liu  Hang Xiao  Rong Gao 《Pesticide biochemistry and physiology》2007,89(1):81-87

In an attempt to find the biorational pesticides, we synthesized 12 pyridinyl derivatives of podophyllotoxin (PPT) and 4′-demthylepipodophylltoxin (4′-DMEP) in this study. Their structures and the α/β substitution at C-4 were confirmed by ¹H NMR, IR, MS spectral analyses and elemental analysis. The insecticidal activities were tested against fifth-instar larvae of Pieris rapae and the third-instar larvae of Cullex pipiens pallens at concentrations of 250 and 10 μg ml⁻¹. Four derivatives of PPT, 4.1, 4.2, 4.3 and 4.5, showed higher insecticidal activities against P. rapae than PPT, while three derivatives of PPT, 4.4, 4.5 and 4.6, displayed higher mosquito larvicidal activity than PPT, with LC₅₀ values of 1.66, 3.96 and 1.54 mg l⁻¹, respectively. Interestingly, we also found that the pyridine ring derivatives of PPT showed delayed insecticidal activity, which is different from traditional neurotic insecticides. The results suggest that 4′-OCH₃ in the PPT derivatives is essential to keep the insecticidal activity and the insecticidal activities of pyridine ring derivatives of PPT are higher than that of the derivatives of 4′-DMEP, supporting PPT has the potential to be a lead structure of semi-synthetic insecticides.  相似文献   

West Nile virus in North America: perspectives on epidemiology and intervention     

Reisen W  Brault AC 《Pest management science》2007,63(7):641-646

West Nile virus (WNV) invaded New York in 1999 and rapidly swept across the North American continent to the West Coast, north into southern Canada and south into Latin America, with minimal genetic change. Regional epidemics in equines and humans typically have included a year of viral introduction with minimal activity, successful overwintering, explosive amplification to epidemic levels the following year and then rapid subsidence. Overwintering possibly included long-term mosquito or avian infections, continued low-level transmission at southern latitudes and dispersal by south-north migrants. Explosive amplification has been associated with infections in several corvid species and other urban birds that produce elevated viremias capable of efficiently infecting even moderately susceptible mosquito species. Intervention has included mass vaccination of equines, and proactive and reactive mosquito control. Proactive mosquito control in areas with established infrastructure has been successful in reducing case incidence.  相似文献   

淡色库蚊XND-P450基因cDNA全长克隆及表达     

郑文亮  李边征华  刘应保 《西北农林科技大学学报(自然科学版)》2014,42(5):211-216

【目的】克隆淡色库蚊氯菊酯抗性相关XND-P450基因,并对其进行生物信息学分析,研究其在敏感品系和抗性品系淡色库蚊中表达量的差异,为阐明XND-P450的功能和抗性机制奠定基础。【方法】依据淡色库蚊抗性品系与敏感品系差异表达的EST片段设计PCR引物,采用RACE技术克隆淡色库蚊氯菊酯抗性相关基因XND-P450全长cDNA序列,并对其进行生物信息学分析;采用半定量PCR技术,对XND-P450基因在敏感品系和抗性品系淡色库蚊中的表达量差异进行检测。【结果】经过克隆获得长度为1 679bp的淡色库蚊氯菊酯抗性相关基因XND-P450全长cDNA,编码523个氨基酸。生物信息学分析表明,该基因与致倦库蚊、埃及伊蚊CYP450基因的同源性在70%以上,其编码蛋白相对分子质量为33 244.9u,理论等电点为8.10,属于不稳定的亲水蛋白,氨基酸序列与致倦库蚊及冈比亚按蚊CYP450蛋白的同源性在40%以上。半定量PCR电泳分析显示,抗性品系淡色库蚊中的XND-P450基因表达量较敏感品系高。【结论】克隆了淡色库蚊氯菊酯抗性相关XND-P450基因,该基因在抗性品系淡色库蚊中的表达量高于敏感品系,推测其与氯菊酯抗性相关。  相似文献   

淡色库蚊氯菊酯抗性相关基因PR-XP1 cDNA的克隆与序列分析          下载免费PDF全文

段晓雷  温雪梅  刘虎岐 《西北农林科技大学学报(自然科学版)》2011,39(11):194-200

【目的】克隆淡色库蚊一个全新的氯菊酯抗性相关基因PR-XP1的全长序列,分析、预测其编码蛋白的结构与功能,为阐明该基因的抗性机制及研制新型卫生杀虫剂奠定基础。【方法】依据抗性与敏感品系库蚊差异表达的EST片段（GenBank号:EC093826.1）,设计特异性PCR引物,采用RACE技术,克隆淡色库蚊氯菊酯抗性相关基因PR-XP1的全长cDNA序列,运用生物信息学软件对其同源性与系统进化进行分析,并对PR-XP1编码蛋白的结构与功能进行预测。【结果】获得了全长为2 004bp的淡色库蚊氯菊酯抗性相关基因PR-XP1,该基因具有完整的CDS区,编码249个氨基酸。同源性比对与系统进化分析表明,PR-XP1基因核苷酸序列与致倦库蚊（GenBank号:XM₀01862469.1）、埃及伊蚊（GenBank号:XM₀01658032.1）和冈比亚按蚊（GenBank号:BX021208.1）相关基因的同源性分别为95%,83%和70%;其编码蛋白的氨基酸序列与致倦库蚊、埃及伊蚊、西方蜜蜂、入侵红火蚁、佛罗里达弓背蚁、印度跳蚁等昆虫中"假设性蛋白"的氨基酸序列同源性均达46%以上。结构分析显示,PR-XP1基因编码蛋白可能为具有2个跨膜螺旋的膜蛋白,跨膜区形成了一个特异性的"U"形结构;该基因还具有1个微弱的信号肽位点和24个磷酸化位点。【结论】克隆得到了一个全新的淡色库蚊氯菊酯抗性相关的"保守性假设蛋白"基因PR-XP1,推测其功能与氯菊酯抗性相关。  相似文献   

Repellency and toxicity of aromatic plant extracts against the mosquito Culex pipiens molestus (Diptera: Culicidae)   总被引：2，自引：0，他引：2  

Traboulsi AF  El-Haj S  Tueni M  Taoubi K  Nader NA  Mrad A 《Pest management science》2005,61(6):597-604

The insecticidal activities of essential oil extracts from leaves, flowers and roots of aromatic plants against fourth-instar larvae of the mosquito Culex pipiens molestus Forskal were determined. Extracts of Foeniculum vulgare Mill were the most toxic, followed by those of Ferula hermonis Boiss, Citrus sinensis Osbeck, Pinus pinea L, Laurus nobilis L and Eucalyptus spp with LC50 values of 24.5, 44.0, 60.0, 75.0, 117.0 and 120.0 mg litre(-1), respectively. Combination tests between the LC50 and the maximum sub-lethal concentration (MSLC) were determined. Over 20 major components were identified in extracts from each plant species tested. Five essential oils and nine pure components were studied for their repellency against mosquito bites. Terpineol and 1,8-cineole were the most effective against Culex pipiens molestus bites offering complete protection for 1.6 and 2 h, respectively.  相似文献   

Cloning and characterization of two glutathione S-transferases from pyrethroid-resistant Culex pipiens     

Samra AI  Kamita SG  Yao HW  Cornel AJ  Hammock BD 《Pest management science》2012,68(5):764-772

BACKGROUND: The Marin strain of Culex pipiens Say is a pyrethroid‐resistant population that was collected in Marin County, California, in 2001 and subsequently maintained in the laboratory under regular permethrin exposure. RESULTS: In this study, two cDNAs, CpGSTd1 and CpGSTd2, encoding glutathione S‐transferase (GST) were cloned from Cx. pipiens Marin. Phylogenetic analysis of the deduced amino acid sequences, CpGSTD1 and CpGSTD2, of these genes indicated that they belong to the Delta class of insect GSTs. The nucleotide and deduced amino acid sequences of CpGSTd1 and CpGSTd2 were 59 and 48% identical respectively. CpGSTD1 and CpGSTD2 were expressed in Escherichia coli and purified by affinity chromatography. The recombinant GSTs exhibited unique selectivity towards the general GST substrates 1‐chloro‐2,4‐dinitrobenzene (CDNB) and 1,2‐dichloro‐4‐nitrobenzene (DCNB), and also differed in their sensitivity to known inhibitors of GSTs. CpGSTD1 exhibited peroxidase activity with cumene hydroperoxide, while CpGSTD2 appeared to lack this activity. CpGSTD1 was able to metabolize 1,1,1‐trichloro‐2,2‐bis(4‐chlorophenyl)ethane (DDT), while DDT metabolism by CpGSTD2 was not detectable. CpGSTD1 and CpGSTD2 showed no detectable metabolism of permethrin. Gene expression of CpGSTd1 and CpGSTd2 in Marin mosquitoes was elevated about twofold in comparison with that found in a pyrethroid‐sensitive mosquito strain. CONCLUSION: The results indicate that CpGSTD1 and CpGSTD2 have unique biochemical characteristics, but they do not appear to play major roles in permethrin resistance in Marin mosquitoes. Copyright © 2012 Society of Chemical Industry  相似文献   

不同方法提取淡色库蚊总RNA的定量比较分析     

温雪梅  段晓雷  刘虎岐 《西北农业学报》2012,21(2):99-102

采用异硫氰酸胍法、Trizol法、Trizol改良法、CTAB法及Trizol+ CTAB 5种方法提取的淡色库蚊总RNA,以期得到最优方法.提取总RNA并用电泳及紫外分光光度计检测,经过Dnase I消化后反转录成cDNA,以此为模板,用CYP6F1引物作定量表达分析.结果表明,与其他几种方法相比,CTAB法提取的效果最好,提取的样品纯度高,完整性好,杂质较少,DNA残留少,降解不明显.CTAB法提取的RNA可以作为eDNA合成和基因克隆的模板,能够满足后续研究的需要.  相似文献