Regulation of sex pheromone biosynthesis in the oriental tobacco budworm,Helicoverpa assulta (Lepidoptera: Noctuidae)

The five components, Z9-16:Ald, 16:Ald, Z11-16:Ald, Z9-16:Ac and Z11-16:Ac, of the sex pheromone in Helicoverpa assulta were mostly detected during the scotophase, with their titer peaking at the 4th hour during the scotophase under a 15L/9D regime. They were not detected during the photophase, but were produced during the photophase when decapitated females were injected with extracts of virgin female (FHE), male heads (MHE), homogenates of the brain-suboesophageal ganglion complex (Br-SOG), or synthetic Hez-PBAN. Production of Z9-16:Ald increased during the first 45min after FHE injection and then declined to a very low level after 2h during the photophase. Synthetic Hez-PBAN stimulated the sex pheromone glands for at least 2h and the effect was more or less proportional to the concentration of the peptide. From the present results, we suggest the following: PBAN is released continuously into the haemolymph to stimulate pheromone biosynthesis at least during the first half of the scotophase, PBAN is synthesized and accumulated independent of photoperiod or sex, and the release starts just prior (about 1h) to the beginning of the scotophase.     

Effects of gamma radiation on different developmental stages of the oriental tobacco budworm,Helicoverpa assulta (Lepidoptera: Noctuidae)

Ionizing radiation is increasingly used as an alternative to post‐harvest crop fumigation by methyl bromide. We studied the effects of gamma irradiation on Helicoverpa assulta (Lepidoptera: Noctuidae) at different stages of development to determine the minimal dose for the prevention of normal emergence of adults. We selected five doses of gamma rays (100, 200, 300, 400 and 500 Gy) based on preliminary experiments and irradiated eggs, larvae, pupae and adults. A dose of 100 Gy to eggs allowed 21.83% of larvae to pupate, but these all died during the pupal stage. A dose of 100 Gy to last‐instar larvae caused larval or pupal death, or the emergence of abnormal adults; no normal adults developed. Irradiation of pupae with doses of 300 Gy and above resulted either in their death or emergence of abnormal adults; however, after 100 or 200 Gy, normal adults emerged and F₁ eggs were produced, but no eggs hatched. Following irradiation of adults, eggs were produced at all doses, although the numbers were significantly decreased compared to untreated controls (P < 0.05; 69.45–125.50 vs. 475.05 eggs per female); however, none of the eggs hatched. As prevention of normal emergence is a key outcome for measuring the effectiveness of radiation, then the 100 Gy dose was effective for irradiation of eggs and larvae, and 300 Gy for pupae.     

Effects of X‐ray irradiation on development and reproduction of the oriental tobacco budworm,Helicoverpa assulta (Lepidoptera: Noctuidae)

All stages of the life cycle of Helicoverpa assulta were irradiated with X‐rays to determine the inhibitory dose for development and reproduction to serve as a quarantine treatment. The 100‐Gy dose was effective for irradiation of eggs and larvae, and the 200‐Gy dose was effective for pupae and mixed‐sex adults. When either adult males or females were irradiated, however, 500 Gy was required to prevent the F1 eggs from hatching, and thus single‐sex adults required much higher doses of X‐ray irradiation. To gather confirmatory data applicable to phytosanitary quarantine regulations, pupae—the immature stage most resistant to X‐ray irradiation—were placed inside paprika in boxes for exportation and were irradiated with 300 Gy as a small‐scale confirmatory test. The dose given to 1,007 individual pupae resulted in 12.62 % survival, and 1.79 % of pupae emerged as normal adults; however, these adults produced only a few eggs that did not hatch, suggesting that a minimum dose of 300 Gy of X‐ray irradiation will provide quarantine security for immature H. assulta in paprika exports.     

Biology and life table studies of the oriental tobacco budworm,Helicoverpa assulta (Lepidoptera: Noctuidae), influenced by different larval diets

Development, survivorship, pupal weight, oviposition, and life table parameters of the oriental tobacco budworm, Helicoverpa assulta Guenée, were evaluated in the laboratory on an artificial diet, pepper (Capsicum frutescens L.), and tobacco (Nicotiana tobacum L.). We found that the average developmental time of immature stages was longest on tobacco (36.2 d), intermediate on pepper (34.4 d), and shortest on artificial diet (33.5 d). Immature survival from egg to pupa varied from 31% on tobacco, 43% on pepper, and 74% on artificial diet. Pupal weight ranged from 197.4 mg/pupa on tobacco, 233.1 mg/pupa on pepper and 253.4 mg/pupa on artificial diet. The average numbers of eggs laid by adults reared as larvae on the artificial diet, pepper, or tobacco were 614, 421 and 334 eggs/female, respectively. Numbers of remaining eggs in ovaries of the adult females reared as larvae on the artificial diet, pepper, or tobacco were 16, 26, and 42 eggs/female, respectively. The longevity of adult females developed from larvae reared on the three diets was not significantly different, whereas the longevity of male adults from the larvae reared on artificial diet was longer (16.8 d) than that for males reared on tobacco (13.8 d) and pepper (13.3 d). The intrinsic, finite, gross, and net rates of increase were highest for females reared as larvae on artificial diet, lowest for females emerging from larvae reared on tobacco, and intermediate for females emerging from larvae reared on pepper. Generation times and doubling time of H. assulta were shortest for larvae fed artificial diet, intermediate from larvae reared on pepper, and longest from larvae reared on tobacco. We concluded that the artificial diet was the most suitable larval diet of H. assulta followed by pepper, and tobacco.     

Effect of a non‐host plant Phaseolus vulgaris on larval performance and oviposition of the oriental tobacco budworm Helicoverpa assulta (Lepidoptera: Noctuidae)

The oriental tobacco budworm, Helicoverpa assulta, is a specialist herbivore feeding on a few plants of the Solanaceae family including tobacco. Larval performance and adult oviposition of H. assulta were investigated in a non‐host plant, Phaseolus vulgaris (Fabaceae) in comparison with two solanaceous host plants, Nicotiana tabacum and Datura stramonium. Larvae provided with the P. vulgaris leaf died off at day 15, whereas 50% and 40% of larval populations fed on the leaves of N. tabacum and D. stramonium, respectively, survived at day 15. Larval growth upon feeding showed significant difference between the non‐host plant (P. vulgaris) and the host plants (N. tabacum and D. stramonium), but it was not significantly different between the two host plants. In the no‐choice experiment of oviposition, gravid females laid more eggs in N. tabacum and D. stramonium than in P. vulgaris. When the most likely acceptable host plant, N. tabacum, and the non‐host plant, P. vulgaris, were subjected to the choice experiment of oviposition, H. assulta females preferred to lay eggs in N. tabacum, where eggs were continuously laid during the whole experiment period. However, eggs in P. vulgaris were hardly detected throughout the period. This study showed that the non‐host plant, P. vulgaris, had a negative influence on the larval performance and adult oviposition of H. assulta, implying neonate stage is critical for larval survivorship, and ovipositional preference by the female is highly specialized to host plants. Further investigation is required to identify non‐host factors, which could be applied to the development of alternative pest management strategy against H. assulta.     

Electrophysiological and behavioral responses of Helicoverpa assulta (Lepidoptera: Noctuidae) to tobacco volatiles

The Oriental tobacco budworm moth, Helicoverpa assulta (Guenée) (Lepidoptera: Noctuidae), is a serious pest on tobacco in China. The flowering stage of the host plant is one of the most attractive stages to H. assulta for feeding and oviposition. Nine electrophysiologically active compounds in tobacco headspace at flower stage were detected by gas chromatography?Celectroantennographic detection (GC?CEAD). These compounds were subsequently identified by gas chromatography-mass spectrometry (GC?CMS) as (E)-??-ocimene, octanal, (Z)-3-hexenyl acetate, (Z)-3-hexen-1-ol, nonanal, (Z)-3-hexenyl-2-methyl butyrate, decanal, linalool, and (E)-??-caryophyllene. The synthetic blend containing nine of the above compounds attracted mated H. assulta females from a distance by upwind oriented flight. Selected subtraction assays showed that the 4-component mixture of (E)-??-ocimene, (Z)-3-hexenyl acetate, nonanal, and (E)-??-caryophyllene elicited equivalent levels of attraction as the 9-component mixture. The removal of any of the four compounds from the 4-component blend resulted in a significant decrease in female upwind flight behavior.     

烟青虫对三种食料植物的选择性及适应性

采用笼罩植株接虫产卵试验和皿内食料吸引选择试验,测定烟青虫HelicoverpaassultaGuen?e对3种食料植物产卵选择性和嗜食性;通过单头饲养试验,测定6龄幼虫对3种食料植物的转化和利用率。结果表明:烟青虫成虫最喜好在番茄上产卵,其次为烟草,在辣椒上产卵最少。初孵幼虫喜食嫩烟叶,选食辣椒和番茄嫩叶的幼虫数较少,其取食量大小顺序为烟草>辣椒>番茄。6龄幼虫能有效利用和转化烟草、辣椒,而对番茄果实的利用和转化效率最低。烟青虫对食料植物的产卵选择和嗜食性差异,是其综合防治中合理进行作物布局,减轻危害的重要依据。     

用性信息素诱剂防治烟青虫的效果

本文研究了两个烟青虫性信息素诱剂配方对烟青虫Helicoverpaassulta (Guen e)成虫的诱杀效果及诱杀成虫后田间烟青虫幼虫的虫口减退率和有虫株减退率。 1型诱芯配比为Z9 1 6 :Ald与Z1 1 1 6 :Ald 1 0 0∶9 5 ;Ⅱ型诱芯配比为 1 6 :Ald和Z9 1 6 :Ald与Z1 1 1 6 :Ald( 1 9 3∶1 0 0∶70 )。结果表明 :烟青虫性信息素诱剂Ⅰ ,Ⅱ型诱芯对烟青虫成虫都有很强的诱杀效果 ,在长达 85d的时间内 ,5个Ⅰ型诱芯共诱杀了4 0 4头雄性成虫 ,5个Ⅱ型诱芯共诱杀 4 1 9头 ,二者没有明显的差异。诱蛾地烟青虫幼虫的虫口密度和有虫株率都有明显的降低 ,Ⅰ、Ⅱ型诱芯使虫口密度和有虫株率的降低幅度均在 6 6 6 7%～ 90 91 %之间。性信息素诱剂使用方便、无毒、不污染环境 ,对烟叶不产生药害 ,防治效果良好 ,具有良好的应用前景。     

(Z)-9-tetradecenal: a potent inhibitor of pheromone-mediated communication in the oriental tobacco budworm moth,Helicoverpa assulta

The behavioural significance of (Z)-9-tetradecenal to male H. assulta was tested by comparing the number of moths attracted to lures containing a standard synthetic female sex pheromone with lures in which (Z)-9-tetradecenal was also added. The standard pheromone mixture used contained 1000 g (Z)-9-hexadecenal, 50 g (Z)-11-hexadecenal, 300 g (Z)-9-hexadecenyl acetate and 15 g (Z)-11-hexadecenyl acetate impregnated on rubber septa. Addition of (Z)-9-tetradecenal to the standard pheromone was shown to significantly reduce the caught of male H. assulta when added in amounts greater than 10 g or 1% of the major pheromone component in both field and net-house experiments. The reduction in catch was found to be dependent on the quantity of (Z)-9-tetradecenal added to the standard pheromone. The implications of these results on conspecific and inter-specific pheromone-mediated communication in H. assulta and related sympatric heliothine species is discussed.Abbreviations Z9-16:AL (Z)-9-hexadecenal - Z11-16:AL (Z)-11-hexadecenal - Z9-16:AC (Z)-9-hexadecenyl acetate - Z11-16:AC (Z)-11-hexadecenyl acetate - Z9-14:AL (Z)-9-tetradecenal - Z9-16:OH (Z)-9-hexadecen-1-ol - Z11-16:OH (Z)-11-hexadecen-1-ol - RH relative humidity     

温周期对不同地理种群棉铃虫幼虫发育及蛹滞育的影响

为了探明不同地理种群棉铃虫Helicoverpa armigera (Hiibner)对温周期的反应,本研究系统调查了棉铃虫广东广州种群(23.08°N,113.14°E)、江西水修种群(29.04°N,115.82°E)、山东泰安种群(36.15°N,116.59°E)和辽宁喀佐种群(41.34°N,120.27°E...     

烟青虫成虫脑结构解剖和三维模型构建

【目的】解剖分析烟青虫 Helicoverpa assulta 成虫脑的结构,并构建脑三维结构数字化模型。【方法】利用神经突触蛋白抗体,对烟青虫成虫脑进行免疫组织化学染色标记,利用共聚焦激光扫描显微镜获得脑扫描数码图像,并结合三维图像分析软件对烟青虫脑结构进行识别分析,构建三维模型。【结果】突触蛋白抗体免疫染色将烟青虫脑和颚神经节的神经髓区域清晰标记出来。烟青虫成虫脑与颚神经节愈合而成为一体,中间具有一个孔洞,为食道穿过的通道。脑主要包括前脑、中脑和后脑3部分。依据染色标记结果识别和构建了至少16个脑神经髓结构。这些神经髓包括边界清晰的视叶、前视结节、蕈形体、中央复合体和触角叶及其亚结构。除此之外,还包括围绕这些神经髓的其他前脑神经髓区域,但这部分前脑神经髓内部边界模糊,不容易细分,而将其与颚神经节区域作为一个整体标记为中间脑,占脑总神经髓的55.05%。【结论】识别出烟青虫脑的主要功能结构区域,并成功构建了三维模型。该研究结果为进一步研究烟青虫脑接收、处理和整合感觉信息及调控行为的机制奠定了解剖学基础,并为研究烟青虫或其他昆虫脑结构发育、变异和重塑提供结构形态和体积大小依据。     

A simple and reliable method for discriminating between Helicoverpa armigera and Helicoverpa assulta (Lepidoptera: Noctuidae)

Abstract The cotton bollworm Helicoverpa armigera and the oriental tobacco budworm H. assulta are sibling species, both being important agricultural pests. Morphologically, the two insects are almost indistinguishable at the egg, larval and pupal stages. One of the big challenges in the study of these insects, in particular in integrated pest management, is a timely and dependable identification of these insects at their early stages of development. Here, we report a H. armigera‐specific nuclear DNA marker, and demonstrate that it can be employed to reliably discriminate between H. armigera and H. assulta by simple polymerase chain reaction amplification experiment.     

烟实夜蛾脂肪酸结合蛋白基因的克隆、序列分析与表达

应用RT-PCR技术,从烟实夜蛾Helicoverpa assulta幼虫脂肪体组织和血细胞总RNA中反转录扩增脂肪酸结合蛋白(fatty-acid binding protein,FABP)基因的cDNA片段,克隆到原核表达载体pGEX-4T-2上,转化大肠杆菌BL21(DE3),用IPTG进行诱导表达并进行检测。结果表明：扩增得到的片段全长399 bp(GenBank登录号为DQ299942),编码132个氨基酸残基,预测分子量15.0 kD,等点电5.83。FABP融合了GST。原核表达后经电泳检测到约41 kD大小的外源蛋白,Western blot检测表明是目的蛋白。     

烟夜蛾性肽受体基因的克隆与表达模式分析

【目的】克隆烟夜蛾Helicoverpa assulta (Guenée)性肽受体基因并分析其表达模式, 为深入研究性肽与交配后反应的关系奠定基础。【方法】采用RT-PCR方法, 从烟夜蛾雌蛾性信息素腺体中得到性肽受体基因cDNA全序列。利用荧光定量PCR方法, 分析该基因的表达模式。【结果】序列分析结果显示, 烟夜蛾性肽受体基因cDNA全长2 048 bp, 命名为HassSPR（GenBank登录号: AFH53182.1）。该基因的开放阅读框长1 275 bp, 编码424个氨基酸残基, 序列中含有7个跨膜域结构, 预测分子量和等电点分别为48.6 kDa和9.25。序列比对分析表明, HassSPR与近缘种棉铃虫H. armigera和其他蛾类性肽受体的氨基酸序列一致性分别达98.35%和超过84%, 与已经报道的其他昆虫的性肽受体的氨基酸序列一致性也在64%以上。不同组织表达分析表明, HassSPR在测定的1日龄雌蛾不同组织中均有表达, 以在脑中的表达量最高。时序表达分析表明, 在羽化前1 天至羽化后6日龄雌蛾的信息素腺体中均有表达, 以3日龄表达量最高。雌蛾交配后, HassSPR在性信息素腺体和脑中的表达量显著上调, 而在交配囊和卵巢中的表达量显著下调。【结论】从烟夜蛾雌蛾性信息素腺体中克隆得到性肽受体基因HassSPR, 其表达模式提示该基因的表达水平与雌蛾的生殖生理和生殖行为有关。     

表皮碳氢化合物分析用于棉铃虫与烟青虫幼虫分类鉴别

本文利用ＧＣ－ＭＳ技术分析了棉铃虫与烟青虫幼虫的表皮碳氢化合物,并与亚洲玉米螟幼虫表皮碳氢化合物进行了比较。结果表明,棉铃虫与烟青虫幼虫表皮碳氢化合物的组分与含量均有显著差异,与玉米螟相比,这二者之间的差异明显小于它们与玉米螟的差异,其表现与三者之间自然的系统发育关系相一致。     

Mating Effect on Sex Pheromone Production of the Oriental tobacco budworm,Helicoverpa assulta

This study was undertaken to clarify the suppression phenomenon of sex pheromone production after mating and its relationship to the physiological mechanism in adult females of Helicoverpa assulta, and determine the mating factor from males causing depletion of sex pheromonc production. Sex pheromone production of H. assulta females was mostly terminated in 3 hours after mating. Mated females maintained with a low titer of sex pheromone until 3 days when it started to increase again, which showed a characteristic of species mating more than once. The mated female again produced pheromone upon injection of pheromone biosynthesis activating neuropeptide (PBAN) or extracts of brain-suboesophageal ganglion complexes (Br-Sg) of mated female, which were shown similar pheromonotropic activities as compared with virgin females. These results indicated that the mating did not inhibit the receptivity of pheromone gland itself and PBAN biosynthesis in suboesophageal ganglion of the mated females. And it seems to support that the depletion of sex pheromone production is responsible for blocking of PBAN release from head. To investigate the mating factor from adult males, when extracts of reproductive organs of male were injected into hemocoel of virgin females evoking depletion of sex pheromone production as shown in mated female. The results suggest that a chemical substance(s) from the male reproductive organs could be responsible for the loss of sex pheromone biosynthesis in H. assulta.     

基于支持向量回归的棉铃虫蛹发育历期估测

温度与发育速率关系模拟是昆虫学研究的一个重要内容, 传统基于经验风险最小的非线性参数模型(Logan模型、Lactin模型和王氏模型)存在诸多弊端。本文基于结构风险最小的改进支持向量回归（SVR）研究温度与棉铃虫Helicoverpa armigera蛹发育历期关系。结果表明: 与传统非线性模型相比, SVR模型性能优异; 基于全部92个样本, SVR模型拟合和留一法预测的决定系数R²分别为0.998和0.996, 估测的蛹期三基点温度更可信。从全部样本中依温度均匀选取部分样本实施独立预测, 当训练集为20个样本时, SVR模型独立预测的R²为0.981, 优于传统非线性模型中独立预测最佳的Lactin模型（R²=0.958）; 当训练集进一步减少到12个样本时, SVR模型的R²仅降低到0.964, 而传统非线性模型均已不适用。结果提示SVR模型在小样本情况下较传统非线性模型优势明显, 在昆虫发育历期估测建模中有应用前景。     

Hybridization between Helicoverpa armigera and Helicoverpa assulta (Lepidoptera: Noctuidae): development and morphological characterization of F1 hybrids

Reciprocal hybridizations between Helicoverpa armigera (Hübner) and Helicoverpa assulta (Guenée) were studied. The cross between females of H. armigera and males of H. assulta yielded only fertile males and sterile individuals lacking an aedeagus, valva or ostium bursae. A total of 492 larvae of the F1 generation were obtained and 374 of these completed larval development and pupated. Only 203 pupae were morphologically normal males, the remaining 171 pupae were malformed. Larvae and pupae that gave rise to morphologically abnormal adults exhibited longer development times. Sterility was not only associated with malformed external sex organs, but also a range of abnormalities of the internal reproductive system: (i) loss of internal reproductive organs, (ii) with one to three copies of an undeveloped bursa copulatrix; or (iii) with one or two undeveloped testes. Normal male hybrid adults showed higher flight activity in comparison with males of both species. In contrast, the cross between females of H. assulta and males of H. armigera yielded morphologically normal offspring (80 males and 83 females). The interaction of the Z-chromosome from H. assulta with autosomes from H. armigera might result in morphological abnormalities found in hybrids and backcrosses, and maternal-zygotic incompatibilities might contribute to sex bias attributed to hybrid inviability.     

烟夜蛾精氨酸激酶基因的克隆及mRNA表达分析

为了深入了解精氨酸激酶基因的作用和寻求害虫防治新的分子靶标, 本研究采用RT-PCR和RACE技术, 从烟夜蛾Helicoverpa assulta脂肪体中克隆了精氨酸激酶cDNA序列, 命名为HassAK（GenBank登录号: HQ336337）, 并采用荧光定量PCR测定了HassAK基因在不同发育阶段（4龄幼虫第1天到化蛹第1天）、 不同组织（头部、 中肠、 脂肪体、 体壁和腹足）和不同温度条件下的表达情况。测序和序列分析结果表明, HassAK基因阅读框架全长1 068 bp, 编码355个氨基酸残基, 预测蛋白质分子量和等电点分别为40.0 kD和5.76。氨基酸序列分析表明, 该序列具有精氨酸激酶典型的酶活性部位、 酶活性中心位点和能形成离子偶结构的保守区。序列比对结果表明, HassAK与其他昆虫AK的氨基酸序列具有70%以上的一致性。荧光定量分析结果显示, HassAK基因在幼虫头部、 中肠、 脂肪体、 体壁和腹足均可表达, 其中以腹足和中肠内的表达水平较高。时序表达分析表明, 预蛹期HassAK基因的表达量达到高峰。此外, 高温和低温均诱导HassAK基因的表达, 说明该基因可能参与昆虫抵御外界不良环境。