转基因在苹果蠹蛾不育昆虫释放技术中的应用

使用不育昆虫释放技术是一项新兴的苹果蠹蛾Cydia pomonella(L.)防治方法,使用转基因得到不育雄蛾具有比传统辐射方法更多的优点。转基因技术通过使用基于piggyBac等转座子的质粒载体,并插入显性条件致死基因以培育遗传性别品系,同时插入荧光蛋白等标记基因来显示转基因的效果;在人工控制的发育条件,建立成熟的稳定苹果蠹蛾品系,用以最终的田间释放以达到防治的目的。     

苹果蠹蛾头部感器的电镜扫描结构

本文使用扫描电镜系统观察并描述了苹果蠹蛾Cydia pomonella(L.)成虫及幼虫触角上及口器上的感器。主要研究结果如下:1)苹果蠹蛾成虫触角背面密布鳞片,感器很少,腹面和侧面鳞片稀疏,具大量感器;2)触角上的感器大部分分布于鞭节各节,少部分分布于柄节和梗节;3)雄虫触角上着生10种感器,雌虫触角上着生9种感器,与雄虫相比,雌虫缺少鳞形感器;4)雄虫口器具鳞形感器和刺形感器,雌虫口器仅具刺形感器;5)幼虫触角3节,基节无感器,第2节具2刺形感器且其分布位置存在个体差异,端节端部具呈三角状排列的3个栓锥感器;6)幼虫口器具一定数量和不同形态的感器。     

苹果蠹蛾老熟幼虫诱杀技术

本文对简易杀虫诱集带诱杀苹果蠹蛾Cydia pomonella(L.)老熟幼虫的效果进行了研究。发现在编织袋,旧衣物,瓦楞纸3种材料中,旧衣物对苹果蠹蛾的老熟幼虫捕获效果最好,其诱捕量占总诱捕量的68.42%;在诱集带中添加的农药中(包括乐斯本、敌敌畏、白僵菌、灭幼脲3号、高效氯氰菊酯等),乐斯本的杀虫效果最好,幼虫死亡率达到98.20%;在不同浓度的乐斯本溶液中,500倍乐斯本杀虫效果最好,幼虫死亡率达到91.61%,且对幼虫没有驱避作用;500倍乐斯本+50倍白僵菌药杀虫诱集带有效期长达40多天,37d后对幼虫的杀伤效果依然达到96%以上,作为老熟幼虫的防治措施十分理想。     

Overcoming the resistance of codling moth against conventional Cydia pomonella granulovirus (CpGV-M) by a new isolate CpGV-I12

Recently, codling moth (CM, Cydia pomonella L.) populations with a significantly reduced susceptibility to C. pomonella granulovirus (CpGV) products have been observed in Germany. A novel CpGV isolate, designated CpGV-I12, is able to overcome the CpGV resistance. CpGV-I12 originated from Iran and showed superior efficacy in laboratory bioassays against a resistant CM strain (CpR), which has a 100-fold reduced susceptibility to commercially used isolate CpGV-M. Determination of the median lethal concentration (LC₅₀) indicated that CpGV-I12 is nearly as efficient in resistant CpR as CpGV-M in a susceptible CM strain (CpS). Beyond, CpGV-I12 caused superior mortality in CpS. Infection experiments showed that the resistance breaking effect can be observed in all instars of CpR. CpGV-I12 is a promising alternative control agent of CM in orchards where conventional CpGV products fail. In addition, we demonstrate in bioassays with recombinant expressed Cry1Ab that cross-resistance to CpGV and Bacillus thuringiensis products is not likely.     

Wastewater sludges as novel growth substrates for rearing codling moth larvae

Codling moth (CM), Cydia pomonella (L) is a pest of numerous fruit species and is also used as a host for baculovirus (granulovirus) and hence the high rate production of CM is important for their biological control. In this research, different wastewater residues from cheese industry, starch industry and municipal wastes (eight different types of larval diets) were investigated as potential substitutes for protein ingredient (principal component) of insect diet. CM larvae were produced with 65–75% survival rate and were healthy as observed under the microscope. Hence, use of cheaper alternatives in the diet derived from agriculture and/or municipal wastes was feasible. Moreover, the preparation steps involved, sterilization, mixing and homogenization and hence the simplicity of these steps makes it practical to scale-up the diet to industrial scale in future. There was no contamination all during the process development. Dryness of all larval diets increased with time, however, even after 26 days, the dryness was ≤18% which is an acceptable norm. Weight, length and diameter of CM larvae was higher when 50% of protein was replaced by cheese industry sludge and corresponded to 145 ± 5, 174 ± 5, and 103 ± 3% increase, respectively. Cheese industry sludge gave highest fecundity of 340 eggs/adult/g of larval diet and the values were higher for other alternative larval diets too. Hence, CM insect diet can be developed using alternative protein substitutes (wastewater residues) is feasible, by adopting simplified scheme with scope for industrial scale-up.     

库尔勒香梨上苹果蠹蛾发生规律及其性信息素迷向防控效果

【目的】苹果蠹蛾是我国进境检疫性有害生物。库尔勒香梨是苹果蠹蛾在新疆果树种植区的主要寄主。研究苹果蠹蛾性信息素迷向防控技术在新疆库尔勒香梨产区的应用效果,可为该害虫的绿色防控提供参考。【方法】通过持续监测与调查,掌握苹果蠹蛾在库尔勒香梨上的发生规律和危害特点,并通过定期调查诱蛾量和蛀果率,比较微胶囊迷向剂、迷向丝、常规化学农药对苹果蠹蛾的防治效果。【结果】库尔勒香梨产区苹果蠹蛾一年发生2~3代,以老熟幼虫越冬,且死亡率较低。4月中旬越冬代羽化出土,5月开始为害;第1代成虫高峰期为7月上中旬,第2代成虫高峰期为8月下旬至9月中旬。微胶囊迷向剂、迷向丝、常规化学农药处理区每周的平均诱蛾量分别为2.85、2.98、4.95头·台~(-1),平均蛀果率分别为2.25%、0.8%、0.99%,与空白对照区的差异均达极显著水平。【结论】使用性信息迷向剂能够降低苹果蠹蛾第1代、第2代幼虫及成虫的数量。     

Development of 2‐phenylethanol plus acetic acid lures to monitor obliquebanded leafroller (Lepidoptera: Tortricidae) under mating disruption

We evaluated the effectiveness of 2‐phenylethanol (PET) in combination with acetic acid (AA) as a binary lure for monitoring male and female obliquebanded leafroller, Choristoneura rosaceana (Harris). Studies were conducted in apple, Malus domestica Borkhausen, orchards treated with or without sex pheromone dispensers for mating disruption (MD). Open polypropylene vials, closed membrane cups, and rubber septa loaded with AA and/or PET in varying amounts were first evaluated in a series of trapping experiments. Membrane cups loaded with 800 mg of PET were as effective as 10‐mg septa, but longer lasting, and were comparable to the open vials. A membrane cup AA lure was effective in tests, but further work is needed to increase its release rate and extend its activity. Catches of codling moth, Cydia pomonella (L.), and C. rosaceana were unaffected by combining PET with (E,E)‐8,10‐dodecadien‐1‐ol, the sex pheromone of codling moth, pear ester, (E,Z)‐2,4‐ethyl‐decadienoate and AA lures. Adding (E)‐4,8‐dimethyl‐1,3,7‐nonatriene to this blend to enhance codling moth catch significantly reduced catches of C. rosaceana. PET + AA was a more attractive binary lure than AA plus phenylacetonitrile (PAN) for C. rosaceana. The addition of PET or PAN to traps already baited with the sex pheromone of C. rosaceana significantly reduced male catches. Traps baited with PET + AA placed in blocks not treated with MD caught significantly fewer C. rosaceana than traps baited with sex pheromone. In comparison, sex pheromone‐baited traps in MD blocks caught ≤1 male moth per season which was significantly lower than total moth (>10) or female moth (≥3) catch in these blocks with PET + AA. A high proportion (>70%) of trapped females were mated in both untreated and MD‐treated orchards. Further refinement of this binary, bisexual lure using membrane cup technology may allow the establishment of action thresholds and improve management timings for C. rosaceana.     

苹果蠹蛾热激蛋白Hsp90基因的克隆及热胁迫下的表达分析

世界检疫性害虫苹果蠹蛾Cydia pomonella是一种温度耐受可塑性很高的物种。本研究针对温度波动可能导致其耐热性增强的科学问题, 采用生测法鉴定了苹果蠹蛾实验种群的高温耐受阈值, 采用同源克隆、 RACE和实时荧光定量PCR (RT-qPCR)等方法研究了苹果蠹蛾热激蛋白Hsp90基因的应激表达对耐热性的重要作用。高温耐受阈值研究结果表明, 苹果蠹蛾实验种群的死亡率随温度的升高和时间的延长显著性升高, 1-5龄幼虫分别经50℃和52℃高温处理2, 5和10 min后, 3龄幼虫耐热性最差, 5龄幼虫最强。50℃和52℃分别处理10 min和5 min均可导致1-4龄幼虫全部死亡, 而5龄幼虫在这两种处理下仍有25.0%和11.1%的存活率。以35℃处理的5龄雌幼虫为材料克隆苹果蠹蛾Hsp90基因全长cDNA, 结果显示该基因全长为2 470 bp, 完整开放阅读框为2 148 bp, 共编码716个氨基酸, 预测分子量为82.07 kDa, 命名为Cphsp90 (GenBank登录号JN624775)。该基因编码的氨基酸序列与亚洲玉米螟Ostrinia furnacalis和甘蓝夜蛾Mamestra brassicae等昆虫的Hsp90的氨基酸序列一致性高达96%, 表明了Hsp90家族的保守特性。Cphsp90 mRNA的相对表达量在32~44℃高温胁迫下随温度的升高而显著增高, 证实Cphsp90是诱导型热激基因, 且mRNA相对表达量与胁迫程度正相关。Cphsp90基因的表达还具有组织特异性, 35℃处理幼虫的表皮中Cphsp90相对表达量显著高于血淋巴、 脂肪体和中肠, 应激响应最为活跃。与未经温热预处理的昆虫相比, 35℃温热预处理3 h后的5龄幼虫在40, 45和50℃更高的温度胁迫下, Cphsp90 mRNA达到最高表达量所需要的胁迫温度有所提升, 由未经预热处理的40℃处理10 min提高到45℃处理10 min, 这与温热预处理会增强5龄幼虫耐热性的现象相符, 表明Cphsp90基因的响应表达在苹果蠹蛾耐热性及其可塑性过程中发挥重要的作用。     

Enzymatic diagnosis of resistance to deltamethrin in diapausing larvae of the codling moth,Cydia pomonella (L.)

The resistance to deltamethrin was evaluated in diapausing larvae of 14 field populations of the codling moth (Cydia pomonella L.) From the main French orchard areas using biological assays. Resistance to deltamethrin was compared to mixed-function-oxidase (mfo) activity measured at the individual level through ethoxycoumarin-O-deethylase (ECOD) activity using a fluorescence microplate reader. The larvae were collected in corrugated paper band traps in the autumn of 1995. Analysis was previously performed on two laboratory strains, one susceptible and one resistant to deltamethrin, in order to characterize the changes in resistance during diapause development. Resistance to deltamethrin as well as the ECOD activity were stable during the diapause, and ECOD activity was always significantly lower in the susceptible strain than in the resistant one. The ECOD activity was significantly correlated to the frequency of resistant moths in the field populations. This strongly suggested the involvement of the mfo system in the resistance to deltamethrin in these populations. The intra-strain variabilities in ECOD activity of both laboratory and field resistant insects indicated that other resistance mechanisms might also be involved. Further investigations on these mechanisms are needed in order to develop complete diagnostic methods and to define suitable control strategies against each resistant population. Arch. Insect Biochem. Physiol. 39:55–64, 1998. © 1998 Wiley-Liss, Inc.     

Development of a direct test of baculovirus resistance in wild codling moth populations

The occurrence of resistance of codling moth (CM, Cydia pomonella L.) to Cydia pomonella granulovirus (CpGV) used as biological control agent revealed the need for fast and reliable resistance monitoring methods. Here, we describe the development of a laboratory resistance test that is directly performed on larvae extracted from infested apples. This test is based on a 14‐day bioassay at a discriminative CpGV concentration of 2 × 10⁵ occlusion bodies/ml diet and can be applied to L1–L4 larvae. Information on virus resistance can be obtained within <4 weeks. In a survey, CM larvae were isolated and tested from 6698 apples from 10 different orchards in Germany, Austria, Switzerland, Italy and the Netherlands. We identified seven CM populations resistant or partly resistant to CpGV isolates. Although some of the orchards were treated with commercial CpGV products, this method allowed us to obtain reliable information about the resistance status of the examined populations.