氟啶脲防治烟草甲的研究

本文通过室内及现场试验,研究氟啶脲防治烟草甲的效果。试验结果表明,烟草甲人工饲料中氟啶脲的浓度为4mg/kg时,烟草甲2龄幼虫的校对死亡率可达97.33%;对库存烟叶,当烟叶中氟啶脲的浓度为8～16mg/kg时,对烟草甲防治效果达到100%,并有24个月的防治作用。     

Digestive proteinases in Lasioderma serricorne (Coleoptera: Anobiidae)

The cigarette beetle, Lasioderma serricorne (Fabricius), is a common pest of stored foods. A study of digestive proteinases in L. serricorne was performed to identify potential targets for proteinaceous biopesticides, such as proteinase inhibitors. Optimal casein hydrolysis by luminal proteinases of L. serricorne was in pH 8.5-9.0 buffers, although the pH of luminal contents was slightly acidic. Results from substrate and inhibitor analyses indicated that the primary digestive proteinases were serine proteinases. The most effective inhibitors of caseinolytic hydrolysis were from soybean (both Bowman Birk and Kunitz), with some inhibition by chymostatin, N-tosyl-L-phenylalanine chloromethyl ketone, and leupeptin. Casein zymogram analysis identified at least eight proteolytic activities. Activity blot analyses indicated one major proteinase activity that hydrolysed the trypsin substrate N-alpha-benzoyl-L-arginine rho-nitroanilide, and three major proteinase activities that hydrolysed the chymotrypsin substrate N-succinyl ala-ala-pro-phe rho-nitroanilide. The absence of cysteine, aspartic, and metallo proteinases in L. serricorne digestion was evidenced by the lack of activation by thiol reagents, alkaline pH optima, and the results from class-specific proteinase inhibitors. The data suggest that protein digestion in L. serricorne is primarily dependent on trypsin- and chymotrypsin-like proteinases.     

Assessment of the efficacy of Japanese Bacillus thuringiensis isolates against the cigarette beetle,Lasioderma serricorne (Coleoptera: Anobiidae)

A total of 2,652 Japanese isolates of Bacillus thuringiensis, belonging to at least 54 H serogroups, were examined for assessment of the toxicity against the cigarette beetle, Lasioderma serricorne (Coleoptera: Anobiidae). When tested with spore/parasporal inclusion mixtures, strong larvicidal activities were associated with 28 isolates (1.1%). Serologically, these toxic isolates fell into 4 known H serovars: thuringiensis (9 isolates), kurstaki (2), kenyae (2), and darmstadiensis (15). Purified parasporal inclusions of the 10 selected isolates exhibited no larvicidal activity, while the supernatants of liquid cultures showed larvicidal and/or growth inhibitory effects. The activities were fully retained after heat treatment at 100 degrees C for 10 min. Overall results suggest that beta-exotoxin (or thuringiensin)-related substances are responsible for the toxicity of the present B. thuringiensis isolates against the cigarette beetle.     

Susceptibility of Lasioderma serricorne (Coleoptera: Anobiidae) life stages to elevated temperatures used during structural heat treatments

Heat treatment of food-processing facilities involves using elevated temperatures (50-60 degrees C for 24-36 h) for management of stored-product insects. Heat treatment is a viable alternative to the fumigant methyl bromide, which is phased out in the United States as of 2005 because of its adverse effects on the stratospheric ozone. Very little is known about responses of the cigarette beetle, Lasioderma serricorne (F.) (Coleoptera: Anobiidae), a pest associated with food-processing facilities, to elevated temperatures. Responses of L. serricorne life stages to elevated temperatures were evaluated to identify the most heat-tolerant stage. Exposure of eggs, young larvae, old larvae, and adults during heat treatment of a food-processing facility did not clearly show a life stage to be heat tolerant. In the laboratory, exposure of eggs, young larvae, old larvae, pupae, and adults at fixed times to 46, 50, and 54 degrees C and 22% RH indicated eggs to be the most heat-tolerant stage. Time-mortality responses at each of these three temperatures showed that the time for 99% mortality (LT99) based on egg hatchability and egg-to-adult emergence was not significantly different from one another at each temperature. Egg hatchability alone can be used to determine susceptibility to elevated temperatures between 46 and 54 degrees C. The LT99 based on egg hatchability and egg-to-adult emergence at 46 degrees C was 605 and 598 min, respectively, and it decreased to 190 and 166 min at 50 degrees C and 39 and 38 min at 54 degrees C. An exponential decay equation best described LT99 as a function of temperature for pooled data based on egg hatchability and egg-to-adult emergence. Our results suggest that during structural heat treatments eggs should be used in bioassays for gauging heat treatment effectiveness, because treatments aimed at controlling eggs should be able to control all other L. serricorne life stages.     

烟草甲谷胱甘肽S-转移酶基因LsGSTe1的表达及其与甲酸乙酯耐受性的关系

【目的】探究烟草甲Lasioderma serricorne谷胱甘肽S-转移酶(glutathione S-transferase, GST)基因LsGSTe1的分子特性和生物学功能。【方法】在烟草甲转录组数据的基础上,利用RT-PCR技术扩增LsGSTe1基因,并进行生物信息学分析;采用qPCR技术检测LsGSTe1在烟草甲不同发育阶段(低龄幼虫、高龄幼虫、蛹、低龄成虫、高龄成虫)及高龄幼虫不同组织(表皮、中肠、脂肪体、马氏管)中的表达水平,以及在甲酸乙酯熏蒸胁迫后的5龄幼虫中的表达变化。进一步采用RNAi技术沉默烟草甲5龄幼虫LsGSTe1基因,通过生物测定分析烟草甲对熏蒸剂甲酸乙酯的敏感性变化。【结果】获得LsGSTe1基因的全长cDNA序列(GenBank登录号:MN480468),开放阅读框长684 bp,编码227个氨基酸,N端和C端均存在催化保守位点。系统发育分析表明该基因属于GSTs的Epsilon家族。qPCR结果表明,LsGSTe1在烟草甲不同发育阶段均有表达,且在高龄幼虫期的表达量较高;表达部位主要在幼虫脂肪体,其次为中肠和表皮,而在马氏管中的表达量最低。LC     

The contribution of symbiotic yeast to toxin resistance of the cigarette beetle (Lasioderma serricorne)

Representative plant allelochemicals were tested for toxicity to larvae of the cigarette beetle Lasioderma serricorne (F.) that were either untreated, treated with fungicides, or treated to render them free of symbiotic yeast (aposymbiotic) through surface sterilization of eggs. Insects rendered symbiont-free had higher mortality and/or developmental rates than controls when fed diets containing flavone, resorcinol or tannic acid. Fungicides reduced symbiont populations and/or caused morphological abnormalities. Two hydrolytic enzymes, which made up a significant protion of mycetome hydrolytic activity, were absent when symbionts were not present, as indicated by gel electrophoresis. This information indicates symbionts do contribute to the survival of their host by detoxifying toxins.

---

Résumé La toxicité relative de substances allélochimiques végétales typiques a été examinée sur des larves de L. serricorne F. traitées aux fongicides et rendues aposymbiotiques par stérilisation superficielle des ufs. La toxicité de la nicotine est relativement inaffectée chez les larves aposymbiotiques; cependant les toxicités des resorcinol, flavone et acide tannique sont significativement augmentées chez les insectes ayant reçu un traitement pour éliminer leurs symbiontes. L'électrophorèse sur gel a montré que quelques enzymes responsables de l'hydrolyse du 1-naphthyl acétate, réaction enzymatique de la détoxification enzymatique, sont absentes chez les insectes aposymbiotiques. L'acide sorbique a réduit significativement l'effectif de symbiontes, tandis que le bénomyl a réduit leur gamme provoquant la multiplication anormale d'autres espèces. Ces résultats montrent que les symbiontes contribuent aux possibilités de détoxification de l'insecte. L'élimination des symbiontes peut être une technique efficace pour lutter contre les insectes qui en contiennent, puisqu'elle les priverait de leur contribution à l'alimentation et à la détoxification. Cette stratégie est vraisemblablement compatible avec des programmes de lutte intégrée, puisque les prédateurs et parasites ne contiennent pas de symbiontes.

     

Mating disruption of Lasioderma serricorne (Coleoptera: Anobiidae) in stored product habitats using the synthetic pheromone serricornin

Cigarette beetles, Lasioderma serricorne (F.) (Coleoptera: Anobiidae), cause significant damage to the multibillion dollar food and tobacco industries worldwide each year. A non‐insecticidal alternative to manage L. serricorne is the application of mating disruption, in which high levels of synthetic sex pheromone are released to create an atmosphere that results in males failing to mate females, thereby causing population suppression or extinction. The reported work used synthetic serricornin, the predominant sex pheromone of L. serricorne, in mating disruption trials conducted in selected food‐ and feed‐processing facilities in South Carolina during 2010 and 2011. Mills subjected to mating disruption trials were monitored using oviposition cups filled with larval food and pheromone traps for males that contained monitoring lures. Immediately after deployment of mating disruption dispensers, trap captures declined significantly and indicated a reduction in population levels, that is, there was ‘trap shutdown’. A significant reduction was observed in numbers of adult beetles caught in the traps 8 weeks before and 8 weeks after treatment in both years. Beetle numbers from pheromone traps in untreated buildings remained at similar levels or increased after the time of mating disruption deployment in treated buildings. The numbers of adults that emerged from oviposition food cups were generally low and varied irregularly in treated and untreated buildings and were determined to be of little value for assessing treatment effects on reproduction. These initial field studies in the USA suggest that release of the synthetic sex pheromone of L. serricorne for mating disruption can significantly inhibit proper orientation behaviour of male L. serricorne to females and may lead to pest population decline from mating disruption.     

烟草甲体内共生菌的初步研究

烟草甲Lasioderma serricorne（Fabricius）体内有类似酵母的胞内共生物YLS,其对烟草甲有重要的营养和解毒作用。该文对YLS的细胞和菌落形态进行了观察,并通过菌落测定、液体培养和细胞质量测定等方法,以探明YLS的最适生长温度。结果表明：28～35C比较适宜YLS的生长,35C为最佳生长温度,而36～48h是研究YLS的最佳时间。     

Susceptibility to insecticides of Lasioderma serricorne (F.) (Coleoptera anobiidae) grown on vitamin-deficient diets.

Variations in the susceptibility to pyrethrum and chlordane (two contact insecticides) of Lasioderma serricorne grown on different vitamin-deficient diets were studied and reasons for such variation were proposed.     

Structure of worldwide populations of Lasioderma serricorne (Coleoptera: Anobiidae) as revealed by amplified fragment length polymorphism profiles

The cigarette beetle Lasioderma serricorne (Fabricius) is the most widespread and destructive pest of stored tobacco. The capability to differentiate between populations from different geographic origins would enable researchers to better understand how insect dispersal through transportation affects the infestation of stored tobacco. Using amplified fragment length polymorphism (AFLP), DNA polymorphisms were assessed in 16 populations of L. serricorne collected from 15 countries. The dendrograms constructed from profile distance matrices revealed well-supported colony clusters. There was no clear clustering as a function of the geographic origin of the samples. The results suggest extensive insect dispersal among geographical regions due to movement of infested commodities worldwide. This first AFLP population study of a stored-product insect demonstrates the potential of AFLP for distinguishing L. serricorne populations.     

Die Rolle der hefeartigen Symbionten von Lasioderma serricorne F. (Coleoptera,Anobiidae) im Proteinmetabolismus ihrer Wirte

Zusammenfassung 1. Aufzuchtversuche an Lasioderma serricorne in Diäten mit 17 Protein- und 2 Hefepräparaten als Proteinquellen zeigten die Überlegenheit normaler Larven über aposymbiontische. 2. Entsprechende Befunde erzielten wir mit Peptonpräparaten und Caseinhydrolysaten; zum Teil vermochten sich aposymbiontisohe Larven nicht zu entwickeln. 3. In Diäten mit Gelatine wuchsen normale Larven langsam, aposymbiontische überhaupt nicht. Tryptophan verbesserte das Wachstum der ersteren und ermöglichte letzteren eine langsame Entwicklung; Histidin und Methionin blieben praktisch wirkungslos. 4. In einer Diät mit Caseinhydrolysat als Proteinquelle und Zusatz von Tryptophan in verschiedenen Konzentrationen lag das Optimum für normale Larven bei 0,4%, das für aposymbiontische bei 0,8%; 0,2 und 0,1% wurden von letzteren nicht vertragen, während normale Tiere auch in tryptophanfreien Diäten eine langsame Entwicklung zeigten. 5. Die Möglichkeit, ein künstlich hergestelltes Aminosäuregemisch als Proteinquelle zu bieten, wurde geprüft. 6. Ein Mangel nicht essentieller Aminosäuren blieb bei normalen und aposymbiontisohen Larven ohne Wirkung. Das Fehlen der essentiellen war für letztere letal, die Entwicklung normaler Tiere war mehr oder weniger stark verzögert. 7. Der Zusatz eines mit Methanol und Wasser erschöpfend extrahierten Hefepräparates hatte in Diäten mit Aminosäuregemischen als Proteinquellen Wuchsstoffwirkung; eine Zufütterung durch Hitze abgetöteter Symbionten war — in geringerem Maße — ebenfalls wirksam. 8. Die Symbionten sind in der Lage, Sulfate und Cystin als S-Quellen zur Methioninsynthese heranzuziehen, der Bedarf der Wirte wird jedoch nur teilweise gedeckt. 9. Die Befunde werden im Hinblick auf die Funktion der Symbionten diskutiert.

---

The role of yeastlike symbiotes of Lasioderma serricorne F. (Coleoptera, Anobiidae) in the protein metabolism of their hostsI. Growth of normal and aposymbiotic larvae in diets with proteins, protein derivates and amino acid mixtures as nitrogen sources

Summary 1. Rearing experiments with Lasioderma serricorne in diets containing 17 proteins and 2 yeast preparations as protein sources proved the superiority of normal larvae to aposymbiotic ones. 2. Similar results were obtained with peptones and casein hydrolysates; in some of these diets aposymbiotic larvae were unable to develop. 3. In diets with gelatine as protein source the development of normal larvae was greatly retarded, that of aposymbiotic ceased completely. An addition of tryptophane enabled the latter to develop slowly and improved growth of the former; histidine and methionine were ineffective. 4. In a diet containing a casein hydrolysate as protein source the optimum concentration of added tryptophane was found to be 0,4% for normal, 0,8% for aposymbiotic larvae. Concentrations of 0.2% and less were insufficient for the latter, while normal ones were able to grow slowly also in tryptophane-free diets. 5. Growth conditions in a diet containing an artificial mixture of 19 amino acids were studied. 6. A lack of single not essential amino acids did not influence both normal and aposymbiotic larvae. That of essential ones was letal for the latter and delayed growth of the former to different degrees. The symbiotic fungi must therefore be able to furnish amino acids to their hosts. 7. A yeast preparation thoroughly extracted with methanol and water showed a growth promoting effect in diets containing complete amino acid mixtures as protein sources; an addition of artificially cultivated symbiotic fungi killed by heat showed the same effect to a lower degree. 8. The symbiotes are able to use anorganic and cystine sulphur for methionine synthesis. This amino acid is furnished to the hosts in a degree not fully covering their requirement. 9. The results are discussed with regard to the function of symbiotic organisms in phytophagous insects.

---

---

Herrn Prof. Dr. H. Kühlwein, Botanisches Institut der Universität Karlsruhe, und Herrn Prof. Dr. K. Steffen, Pharmakognostisches Institut der Technischen Universität Braunschweig, sei für stetes Interesse und großzügige Förderung unserer Untersuchungen bestens gedankt. Der Deutschen Forschungsgemeinschaft danke ich für finanzielle Unterstützung.     

温度对党参害虫烟草甲实验种群生长发育的影响

以中药材党参(Codonopsis pilosula)为寄主食料,在人工气候箱内,研究了温度对烟草甲(Lasioderma serricorne)卵、幼虫和蛹的生长和发育的影响;计算了烟草甲各虫态的发育起点温度和有效积温;通过Logistic模型及"王-兰-丁"模型对温度与烟草甲各虫态之间的关系进行拟合,优化拟合温度与烟草卵和幼虫发育关系的Logistic方程6个,温度与烟草甲幼虫发育关系的"王-兰-丁"方程5个。结果表明,烟草甲卵、幼虫、蛹及卵-蛹期的发育起点温度分别为16.51℃、11.07℃、11.78℃和12.20℃,其卵、幼虫、蛹及卵-蛹期的发育有效积温分别为89.59、669.53、137.12和912.72日度;温度的升高有利于烟草甲的各虫态的生长和发育;"王-兰-丁"模型拟合对研究较宽温度幅度的昆虫发育与温度之间的关系拟合效果优于Logistic模型。本研究结果可为危害储藏期中药材的烟草甲种群数量控制技术提供技术支撑。     

烟草甲细胞色素P450还原酶基因的分子特征与表达模式分析

烟草甲Lasioderma serricorne是一种重要的仓储害虫,长期化学防治导致烟草甲已对多种传统熏蒸剂产生抗性,但其对新型熏蒸剂甲酸乙酯仍处于敏感水平。因此明确其体内细胞色素P450还原酶（cytochrome P450 reductase, CPR）对甲酸乙酯的代谢解毒功能,对开展该药剂的抗性监测及延缓抗性的发生发展具有重要意义。本研究旨在克隆烟草甲LsCPR基因,分析其分子特征和表达特性,为进一步明确其在烟草甲对甲酸乙酯解毒代谢过程中的作用奠定基础。利用RT-PCR技术克隆烟草甲LsCPR基因的开放阅读框（open reading frame, ORF）;利用生物信息学软件分析LsCPR编码蛋白的结构、特征和系统进化关系。采用实时定量PCR技术检测LsCPR在烟草甲不同发育阶段（低龄幼虫、高龄幼虫、蛹、低龄成虫、高龄成虫）、幼虫不同组织（表皮、肠道、脂肪体和马氏管）以及甲酸乙酯熏蒸胁迫后的表达模式。烟草甲LsCPR基因的ORF为2 046 bp（GenBank登录号：MZ423209）,编码681个氨基酸,具有典型的昆虫CPR基因FMN区域、NADPH区域和FAD等保守结构域。系统发育分析表明,烟草甲LsCPR与鞘翅目昆虫聚为一支,且与赤拟谷盗Tribolium castaneum CPR亲缘关系最近。LsCPR在烟草甲不同发育阶段均有表达,在高龄幼虫期的表达水平较高;在幼虫体内的表达部位主要在肠道,其次为脂肪体和马氏管,而在表皮的表达水平最低。LC10、LC30和LC50 3种浓度的甲酸乙酯处理24 h后,烟草甲LsCPR表达量随着胁迫浓度升高而上调且显著高于对照;甲酸乙酯LC50处理烟草甲不同时间（3、6、12、24和48 h）后,LsCPR基因上调表达,24 h时达到表达高峰。推测LsCPR是参与烟草甲代谢甲酸乙酯的候选基因。     

温度对烟草甲生长发育的影响

在２０－３２．５℃范围内,烟草甲卵期、幼虫期、肾期、成虫产卵前期及全世代的发育速率与温度呈逻辑斯蒂曲线关系,但高温（３５℃以上）对烟草甲的生长发育有明显的抑制作用。烟草甲各虫态的发育起点温度和有效积温分别为卵：１５．４８６６℃,３５．９８９２日度;幼虫：１０．３５５３℃,４２２．０４６２日度;蛹：１５．４２３９℃,７２．３８０８日度;产卵前期：１４．７００７℃,２０．８４２７日度;全世代：１２．５     

Larval movement ofLasioderma serricorne (F.) (Coleoptera: Anobiidae) in a plane

The mean maximum change in position of larvae of Lasioderma serricorne (F.) at 30°C, 60% RH increased by 0.290 cm/day, their mean position at the end of a 24 hour period increased by 0.245 cm/day. The linear distance from the point of origin was equal to approximately one third of the total distance moved. Either density or high numbers increased the rate of movement.     

食料对烟草甲共生菌数量和生长发育的影响

烟草甲Lasioderma serricorne(Fabricius)消化道内含有一种类酵母共生菌,其在烟草甲的营养需求和对外源物质的解毒过程中有重要作用。以麦粒、烟叶、麦片、甘草和面粉为食料饲养烟草甲,研究了食料对烟草甲共生菌数量和发育历期的影响,发现:烟草甲5龄幼虫体内的共生菌数量最多,蛹次之;食料对共生菌的数量有显著的影响,尤其在5龄幼虫期;麦粒饲养的烟草甲发育历期最短、体内的共生菌数量最多,而甘草饲养的烟草甲发育历期最长、体内的共生菌数量最少;共生菌的数量与烟草甲的生长发育历期之间存在着明显的负相关关系。