A Role for Ecdysteroids in the Induction and Maintenance of the Pharate First Instar Diapause of the Gypsy Moth,Lymantria dispar

Several lines of evidence suggest a novel regulatory mechanism for diapause regulation in the gypsy moth. We propose that ecdysteroids play a role in the induction and maintenance of the pharate first instar larval diapause in this species. A 55 kDa gut protein that is indicative of diapause is expressed in intact and neck-ligated pharate larvae but is not expressed when a ligature is placed posterior to the prothorax, site of the prothoracic gland. Guts cultured in vitro for 12 h cease to synthesize the 55 kDa protein, but synthesis of the protein resumes if the culture medium is enriched with a prothorax extract from pharate larvae or a prothoracic gland extract from fifth instar larvae. Injection of 20-hydroxyecdysone or the ecdysteroid agonist, RH-5992, into isolated abdomens stimulates synthesis of the diapause-specific 55 kDa protein, suggesting that the essential factor from the prothorax is an ecdysteroid. KK-42, an imidazole derivative known to inhibit ecdysteroid biosynthesis, averts diapause when applied to prediapausing pharate first instar larvae, but this effect can be countered by application of 20-hydroxyecdysone or RH-5992, i.e. KK-42 treated pharate larvae that are exposed to an ecdysteroid or RH-5992 readily enter diapause. A chilling period (120 days at 5 degrees C) is normally adequate to prompt an immediate termination of diapause when pharate larvae are transferred to 25 degrees C, but if such larvae are held in hanging drop cultures with ecdysteroids they fail to terminate diapause. Together, these results suggest that ecdysteroids are essential for the induction and maintenance of diapause and imply that a drop in the ecdysteroid titer is essential for diapause termination. Copyright 1997 Elsevier Science Ltd. All rights reserved     

Stage-dependent effects of 20-hydroxyecdysone on DNA synthesis of corpus allatum cells in the silkworm,Bombyx mori

DNA synthesis in cells of the corpus allata (CA) of the silkworm, Bombyx mori, was studied immunocytochemically after in vivo labeling with 5-bromo-2'-deoxyuridine (BrdU); developmental changes during the 3rd, 4th, and last larval instars and effects of 20-hydroxyecdysone treatment were examined. During both the 3rd and 4th larval instars, the number of DNA-synthesizing cells fluctuated, and relatively low levels were observed during the middle stages. On day 0 of the last larval instar, the number of DNA-synthesizing cells per gland was 9.2, which then increased on day 1 and remained at levels ranging from 12.9 and 16.9 cells per gland. A major peak level (28 BrdU-labeled cells per gland) occurred on day 8, two days after larvae entered the wandering stage. When last instar larvae were fed 20-hydroxyecdysone-supplemented mulberry leaves starting on day 0 or 1, the number of DNA-synthesizing cells dramatically decreased to very low levels and these low levels were maintained throughout the remainder of the instar. However, no effect was observed when last instar larvae were fed 20-hydroxyecdysone-supplemented mulberry leaves starting on day 3, indicating the stage-specific action of 20-hydroxyecdysone. The mechanism by which 20-hydroxyecdysone treatment inhibits DNA synthesis of CA cells was further examined by using continuous in vitro BrdU labeling for a 2-day incubation. It was found that the decrease in responsiveness of DNA synthesis of CA cells of 20-hydroxyecdysone-treated larvae to stimulation by growth factors from hemolymph may have been, at least in part, responsible for the indirect inhibitory effects of 20-hydroxyecdysone.     

外源激素处理后家蚕吡哆醛激酶和磷酸吡哆醇氧化酶转录水平的变化

【目的】研究家蚕 Bombyx mori 经蜕皮激素(20-hydroxyecdysone, 20-E)和保幼激素类似物(juvenile hormone analogue, JHA)处理后引起吡哆醛激酶(pyridoxal kinase, PLK)和磷酸吡哆醇氧化酶(pyridoxine-5′-phosphate oxidase, PNPO)的转录水平变化,为进一步研究激素对蚕体营养代谢等工作奠定基础。【方法】以20-E和JHA分别喂食不同发育时期(5龄第1, 3和5天)的家蚕幼虫,以喂食蒸馏水的家蚕为对照,采用实时荧光定量PCR(real-time quantitative PCR)方法在处理后24 和48 h对各组幼虫后部丝腺中PLP合成酶PLK和PNPO的转录水平进行分析。【结果】5龄第1天幼虫经20-E处理24和48 h后,PLK和PNPO的转录水平出现上调且与对照的差异达到极显著 (P＜0.01);5龄第3天幼虫经20-E处理,PLK的转录水平在48 h出现下调且与对照的差异达到显著(P＜0.05),PNPO的转录水平在24 和48 h均出现上调且与对照的差异达到极显著 (P＜0.01);5龄第5天幼虫经20-E处理后PLK和PNPO的转录水平无变化。5龄第1天幼虫经JHA处理后PLK和PNPO的转录水平未受到影响;5龄第3天幼虫经JHA处理后,PLK的转录水平在48 h出现显著下调且与对照的差异达到显著(P＜0.05),PNPO的转录水平在24和48 h后均出现显著下调且与对照的差异达到极显著(P＜0.05);5龄第5天幼虫经JHA处理24和48 h后,PLK和PNPO的转录水平出现下调且与对照的差异达到极显著 (P＜0.01)。【结论】20-E和JHA显著影响家蚕5龄幼虫PLK和PNPO的转录水平,20-E提高5龄前期家蚕PLK和PNPO的转录水平,JHA降低5龄后期它们的转录水平,为深入研究激素对VB₆的调控奠定基础。     

In vivo production of recombinant protein by a baculovirus vector inoculated perorally to the prefinal instar larvae of Bombyx mori L. (Lep., Bombycidae) aided by an optical brightener, Tinopal UNPA-GX

Abstract: Budded particles of nucleopolyhedrovirus (NPV) were found to infect perorally the 4th (prefinal) instar larvae of Bombyx mori L. that were treated by an optical brightener, Tinopal UNPA-GX (Tinopal). Host larvae were fed a diet containing 0.3% (w/w) Tinopal on day 1 in the 4th instar and then fed a diet contaminated by budded particles of NPV (1.0 × 10⁶ TCID₅₀ U/larva) that was pathogenic to B . mori (BmNPV) on day 2 (inoculation schedule 1). Another set of host larvae was fed a diet containing BmNPV budded particles (2.5 × 10⁶ TCID₅₀ U/larva) together with 0.3% (w/w) Tinopal on day 1 in the 4th instar (inoculation schedule 2). Host larvae treated by both schedules died of viral infection. The operation of schedule 2 is simpler than that of schedule 1, although the former required higher doses of the virus for satisfactory infection. We inoculated a baculovirus vector carrying human serum albumin (HSA) gene into 4th instar B . mori larvae by schedule 1. Recombinant HSA was detected in the homogenate of host larvae 4 days after inoculation. The peroral inoculation of BmNPV budded particles aided by Tinopal may thus lead to industrial pharmaceutical production using a baculovirus vector for large numbers of insect hosts.     

An in vitro study on regulation of prothoracic gland activity in the early last-larval instar of the silkworm Bombyx mori

The endocrine mechanisms that regulate prothoracic gland (PG) activity in early stages of final larval instar of the silkworm Bombyx mori were investigated using a newly developed long-term cultivation system of the gland. The PGs dissected from day-0 fifth instar larvae did not secrete detectable amounts of ecdysone for the first 24 h in culture but started secretion within the next 2 days. The amount of secreted ecdysone increased day by day. When day-0 PGs were co-cultivated with corpora allata, however, they remained inactive for at least 8 days. PGs dissected from 1-day younger larvae (day-3 fourth instar larvae) secreted ecdysone for the first 24 h but stopped secretion for the next 24 h, followed by recovery of ecdysone secretory activity. By contrast, PGs from day-1 fourth instar larvae remained active throughout a cultivation period without any sign of inactivation. However, when the same glands were exposed to a high titer of 20-hydroxyecdysone for the second 24h in culture, they gradually lost their activity. These results indicate that PGs of fourth instar larvae are inactivated by ecdysteroid through a negative feedback mechanism and that thus inactivated PGs spontaneously recover ecdysone secretory activity in the early fifth instar unless inhibited by juvenile hormone.     

Developmental arrest induced by juvenile hormone in larvae of Bombyx mori

Injection of the juvenile hormone analog (JHA) methoprene into day 3, fifthinstar larvae of Bombyx mori induced developmental arrest. Feeding activity declined, and the larvae remained as larvae for more than 2 weeks, after which they died. After JHA injection, the hemolymph ecdysteroid titer was low, and the prothoracic glands were almost inactive for 7 days. During this period, prothoracic glands were stimulated by prothoracicotropic hormone (PTTH) in vitro, indicating that JHA did not inhibit the competence of the glands to respond to PTTH. When brain-corpora cardiaca-corpora allata complexes were removed from intact fifth-instar larvae on day 4, the prothoracic glands became autonomously active and produced enough ecdysone for pupation. When PTTH injections were given to larvae previously injected with JHA (7 days before), the larvae recovered feeding activity, purged their guts, and pupated. Injections of 20-hydroxyecdysone into larvae that had been injected with JHA 7 days earlier induced larval molting. These results suggest that JHA affects both the brain and the prothoracic gland.     

保幼激素类似物及蜕皮甾类对亚洲玉米螟幼虫酚氧化酶活性的影响

分别用1 μg/头、0.1 μg/头和0.01 μg/头浓度的保幼激素类似物methoprene（蒙五一五）体外处理亚洲玉米螟5龄幼虫,测定幼虫体壁组织、血清和血细胞溶离物中酚氧化酶的活性。结果表明： 1 μg/头 methoprene处理组和0.1 μg/头处理组幼虫体壁组织中酚氧化酶活性与对照组相比有显著提高（P<0.01）,血清和血细胞溶离物中酚氧化酶活性也显著上升（P<0.01）。将含有20-羟基蜕皮酮的人工饲料饲喂亚洲玉米螟5龄幼虫,处理组幼虫体壁组织的酚氧化酶活性下降(P<0.05),血清和血细胞溶离物中的酚氧化酶活性均低于对照组 （P<0.01）。这些结果表明methoprene可以诱导亚洲玉米螟5龄幼虫体内酚氧化酶活性的上升,而20-羟基蜕皮酮则抑制了酚氧化酶的活性。     

转cry1Ab/cry1Ac基因水稻对大螟幼虫体内三种保护酶活性的影响

为阐明转cry1Ab/cry1Ac基因水稻对大螟Sesamia inferens (Walker)作用的生理生化机制, 本研究用转cry1Ab/cry1Ac基因水稻茎秆饲喂大螟3龄和5龄幼虫, 采用酶活性测定方法研究了取食转Bt水稻对大螟幼虫体内3种保护酶SOD(superoxide dismutase)、 CAT(catalase)和POD(peroxidase)活性的影响。结果表明, 大螟3龄幼虫在取食转基因水稻24 h后SOD活性与对照相比提高了43.44%, 48 h后降至最低值; 在取食24 h后POD值达到最高值, 其酶活性比对照升高了29.22%, 最终在取食48 h后降至最低值, 并显著低于对照; 在取食转基因水稻4 h后, CAT活性升高了30.33%, 在取食48 h后, 与对照相比, CAT活性降低了27.01%; 5龄幼虫取食4 h后SOD活性显著高于对照水平, 36 h后降至最低值, 与对照相比, 活性下降了31.62%; 在取食8 h后POD活性达到最高值, 与对照相比, 升高了73.20%, 36 h后酶活性降至最低值; 在取食之初4 h CAT活性达到最高值, 与对照相比, 其值升高了75.73%, 在取食48 h后, 其活性与对照相比减少了7.55%。3龄幼虫与5龄幼虫相比, 对Bt的抗性水平较低, 自身防卫能力较差。结果说明, 在取食初期, 试虫体内保护酶活性升高, 以抵御Bt毒蛋白对虫体的伤害作用, 随着取食时间的延长, 保护酶活性迅速降低, 从而干扰虫体正常的代谢过程, 导致虫体出现中毒症状, 致使昆虫死亡。     

桃蛀螟CYP6AE76基因参与对Cry1Ab蛋白的解毒作用

【目的】明确桃蛀螟Conogethes punctiferalis幼虫取食Cry1Ab蛋白后体内CYP6AE76的过表达及对Cry1Ab蛋白有解毒作用。【方法】分析桃蛀螟CYP6AE76序列特征;利用RT-qPCR检测CYP6AE76在不同发育阶段(1-5龄幼虫)和4龄幼虫不同组织(头、中肠、血淋巴和脂肪体)以及4龄幼虫取食含有Cry1Ab蛋白(LC₅₀=1.08 ng/cm2)的人工饲料3 d存活的幼虫中肠和血淋巴中的表达量;利用RNAi饲喂法沉默桃蛀螟4 龄幼虫CYP6AE76后检测中肠中CYP6AE76的表达量,并统计120 h后幼虫体重并计算幼虫存活率;利用RNAi饲喂法沉默桃蛀螟初孵幼虫CYP6AE76后饲喂含1.08 ng/cm² Cry1Ab蛋白的饲料,7 d后统计幼虫体重并计算幼虫存活率。【结果】桃蛀螟CYP6AE76基因开放阅读框长1 572 bp,编码524个氨基酸,分子量约为60.34 kD,属于CYP6家族基因。发育表达谱结果表明, CYP6AE76基因在桃蛀螟整个幼虫阶段均有表达且在1龄幼虫期表达量最高,随着幼虫龄期增大而表达量降低;组织表达谱结果表明,CYP6AE76在4龄幼虫中肠中表达量最高。4龄幼虫取食含有Cry1Ab蛋白(1.08 ng/cm²)的人工饲料后,CYP6AE76在中肠和血淋巴中的表达量相比对照显著上调。通过RNAi沉默CYP6AE76后,桃蛀螟初孵幼虫再取食含有Cry1Ab蛋白的人工饲料后体重显著降低。【结论】CYP6AE76可能参与对桃蛀螟幼虫摄入的Cry1Ab蛋白的解毒。     

Bt玉米(MG95)对粘虫的抗性和拒食作用

在室内以我国自主培育的转Bt基因玉米 (简称 :Bt玉米 )为材料 ,测定了Bt玉米对不同龄期 ( 1～ 4龄 )粘虫PseudaletiaseparataWalker的影响。测定指标包括 :幼虫体重、幼虫发育历期、幼虫死亡率、化蛹率、蛹重、蛹历期、羽化率、产卵量、成虫历期、卵孵化率。结果显示 :用Bt玉米喂养的 1～ 4龄幼虫在处理 6d及 1～ 3龄幼虫在处理 9d时的体重与对照相比均差异显著 ;除 1龄幼虫外 ,2～ 4龄幼虫处理 6d及 2～ 3龄幼虫处理 9d后的死亡率差异不显著 ;用Bt玉米喂养的各龄幼虫发育历期延长 ,与对照相比差异显著 ;1、2龄幼虫的化蛹率与对照相比有显著差异 ,3、4龄则差异不显著 ;1～ 4龄幼虫的蛹重、蛹历期及羽化率均差异显著 ;成虫历期仅 1龄幼虫差异显著 ;与对照相比 ,各龄粘虫的产卵数、卵孵化率均无显著差异。结果表明 ,1龄幼虫对Bt玉米最为敏感。另外 ,拒食实验结果显示 ,在实验开始后的 8,1 6,2 4,3 2及 48h ,非选择性测定的拒食率为 63 2 0 ,64 60 ,48 99,2 3 3 5和 0 5 4% ,选择性测定的拒食率为98 3 3 ,82 43 ,71 5 4,40 1 6和 0 2 7% ,总体趋势是拒食率随时间延长而降低 ,48h后Bt玉米对粘虫幼虫无拒食活性。     

双条杉天牛幼虫龄数的划分(鞘翅目:天牛科)

双条杉天牛 Semanotus bifasciatus Motschulsky 是危害我国侧柏 Platycladus orientalis 和圆柏 Sabina chinensis 的重要钻蛀性害虫,其幼虫龄数及最佳分龄指标的确定是进一步研究其生物学特性、发生规律的基础.本研究通过室内木段饲养法获取不同发育阶段的双条...     

Regulation of prothoracic gland ecdysteroidogenic activity leading to pupal metamorphosis

The prothoracic glands of early last (fifth) instar larvae of the silkworm are inactive with regard to ecdysteroidogenesis and unresponsive to prothoracicotropic hormone (PTTH) [J. Insect Physiol. 31 (1985) 455]. In an attempt to elucidate the hormonal mechanisms that cause the inactivity, we compared the effects of PTTH, dibutyryl cyclic AMP (dbcAMP), a cAMP phosphodiesterase inhibitor (IBMX), juvenile hormone analogue (JHA) and 20-hydroxyecdysone (20E) on secretory activity of the third, fourth and fifth instar glands. Among the factors examined, feedback inhibition by 20E was indicated to be the most likely factor. Inhibition was moderate in the third and early fourth instars while 20E strongly inhibited the glands of middle fourth instar larvae. The inhibitory effect of 20E was reduced by removal of the brain and corpora allata. Once the glands were suppressed by 20E to the degree of exhibiting neither secretory activity nor responsiveness to PTTH, dbcAMP or IBMX did not elicit ecdysone secretion at all. Thus the feedback inhibition may shut down ecdysteroidogenesis although it is obscure whether it affects the intracellular transductory cascade from the PTTH receptor through cAMP. Taken together, this evidence suggests that inactivity of the gland in the early fifth instar is brought about by feedback inhibition of the glands by 20E occurring in the late fourth instar, and that this inactivity is maintained by the juvenile hormone found in the early fifth instar.     

Plasmatocyte sensitivity to plasmatocyte spreading peptide (PSP) fluctuates with the larval molting cycle

Plasmatocyte spreading peptide (PSP) is a cytokine from the moth Pseudoplusia includens that activates a class of hemocytes called plasmatocytes to bind and spread on foreign surfaces. Previous structure-function studies on PSP used plasmatocytes collected from P. includens larvae that were in the late stages of the last (fifth) instar. Here, we report that plasmatocyte sensitivity to PSP varied significantly during the fourth and fifth instar. PSP weakly activated plasmatocytes early in the instar when hemolymph juvenile hormone (JH) titers were relatively high and ecdysteroid titers were low, but strongly activated plasmatocytes late in the instar after JH titers declined and ecdysteroid titers rose. In contrast, plasmatocytes did not vary in their response to plasma, which contains other factors besides PSP that affect plasmatocyte function. In vitro assays indicated that 20-hydroxyecdysone (20E) dose-dependently synergized PSP activity, whereas the JH analog methoprene antagonized PSP activity. Methoprene had no effect on adhesion and spreading of granular cells, but plasmatocytes from larvae topically treated with methoprene exhibited a reduction in sensitivity to PSP. Collectively, these results indicate that plasmatocyte sensitivity to PSP fluctuates in relation to the molting cycle, and that PSP activity is affected by juvenoids and ecdysone.     

The effect of the juvenile hormone analog, fenoxycarb, on ecdysone receptor B1 expression in the midgut of Bombyx mori during larval-pupal metamorphosis

The Bombyx mori (Lepidoptera: Bombycidae) midgut undergoes remodeling during the larval-pupal metamorphosis. All metamorphic events in insects are controlled by mainly two hormones: 20-hydroxyecdysone (20E) and juvenile hormone (JH). Fenoxycarb, O-ethyl N-(2-(4-phenoxyphenoxy)-ethyl) carbamate, has been shown to be one of the most potent juvenile hormone analogs against a variety of insect species. In this study, the effect of fenoxycarb on EcR-B1 protein expression in the midgut of Bombyx mori during the remodeling processwas investigated. Fenoxycarb was topically treated to the beginning of the fifth instar Bombyx larvae. Its application prolonged the last instar and prevented metamorphic events. Analyses were performed from day 6 of the fifth instar to 24 hr after pupation in controls and to day 14 of the fifth instar in the fenoxycarb treated group. According to our results, the presence of EcR-B1 in the midguts of the fenoxycarb treated group during the feeding period suggested that EcR-B1 was involved in the functioning of larval cells and during this period fenoxycarb did not affect EcR-B1 status. Immediately after termination of the feeding stage, the amount of EcR-B1 protein increased, which indicated that it may strengthen the ecdysone signal for commitment of remodeling process. In the fenoxycarb treated group, its upregulation was delayed, which may be related to the inhibition of ecdysone secretion from the prothoracic gland.     

具保幼激素活性昆虫生长调节剂与昆虫摄食行为反应的关系(英文)

本文以烯虫酯、双氧威、ＮＣ－１７０、ＮＣ－１８４和ＮＣ－１９６等具保幼激素活性的昆虫生长调节剂（ＩＧＲｓ）为药剂,以丝绵木金星尺蠖 Ｃａｌｏｓｐｉｌｏｓ ｓｕｓｐｅｃｔａ４龄幼虫和亚洲玉米螟 Ｏｓｔｒｉｎｉａ　ｆｕｒｎａｃａｌｉｓ５龄幼虫为试材,采用点滴法和混药饲喂法,结合非选择性饲料柱称重法,研究了ＩＧＲｓ与昆虫摄食行为之间的关系。结果表明,烯虫酯、双氧威、ＮＣ－１７０、ＮＣ－１８４、ＮＣ－１９６和川楝素对亚洲玉米螟５龄幼虫的非选择性拒食率依次为－３７．７、１４．６、－１２．３、１０．９、１４．７和６２．２％,烯虫酯刺激两种试虫取食,而双氧威则有一定的拒食作用,不同药剂表现出的影响不尽一致。因此,外源具保幼激素活性的活性物质对昆虫摄食行为反应的干扰破坏,可能是ＩＧＲs作用机理中不可忽视的重要方面之一。对饲料柱称重法和虫粪称重法进行了比较,证实两种方法结果相近,趋势一致,均可用于昆虫摄食行为的生测。     

THE RELATIONSHIP BETWEEN INSECT GROWTH REGULATORS WITH JUVENILE HORMONE ACTIVITY AND INSECT FEEDING BEHAVIOURAL RESPONSE

Abstract  Five insect growth regulators (IGRs) with juvenile hormone (JH) activity including fenoxy-carb, methoprene, NC-170, NC-184 and NC-196 were selected to study on the relationship between IGRs and feeding behaviour of the fourth instar larvae of Calospilos suspecta (Warren) and the fifth instar larvae of Ostrinia furnacalis Guenee, by topical and dietary applications as well as "diet column" methods. The results indicated that the antifeeding indices of methoprene, fenoxycarb, NC-170, NC-184, NC-196 and toosendanin against the fifth instar larvae of Ostrinia furnacalis in no-choice test were -37. 7, 14. 6, -12. 3, 10. 9, 14. 7 and 62. 2% respectively. Methoprene could stimulate the two species of insects to feed, whereas fenoxycarb had some antifeeding activity, and the effects of all the tested chemicals were different. Therefore, disturbances of IGRs with JH activity on insect feeding behavioural response may be one of the mechanisms of action not to be ignored. Moreover, by comparing "diet column" method with "faeces weight" method, it was suggested that the two methods might be applied in bioassay of antifeeding behaviour.     

Analysis of precocious metamorphosis induced by application of an imidazole derivative to early third instar larvae of the silkworm,Bombyx mori

When an imidazole derivative (KK-42) was applied to day 1 third instar larvae of the silkworm, Bombyx mori, 100% underwent precocious metamorphosis at the end of the fourth instar. Thus, the fourth instar becomes the last instar in these KK-42–treated larvae. The endocrine systems underlying the precocious metamorphosis were analyzed in the present study. Hydroprene application during the prolonged third instar after KK-42 treatment can prevent precocious metamorphosis, and the results showed dose-dependent and stage-specific effects. From analysis of the developmental changes in ecdysteroid levels in both KK-42–treated larvae and KK-42– and hydroprene-treated larvae, we conclude that changes in JH levels during the third larval instar can modify the secretion pattern of prothoracic glands and that during the next larval instar, very low ecdysteroid levels during the early stages of the presumptive last (fourth) larval instar are directly related to precocious metamorphosis. Arch. Insect Biochem. Physiol. 36:349–361, 1997. © 1997 Wiley-Liss, Inc.     

Growth and silk formation of silkworm larvae influenced by phytoecdysones

The fourth and fifth instar larvae of the silkworm were reared on artificial diets containing ponasterone A, ecdysterone, and inokosterone. The growth of the larvae and their silk glands, fibroin-synthesizing activity, and silk formation have been investigated. With a diet containing ponasterone A, the fourth instar larvae grew slowly and only a few larvae could ecdyse, while the growth of the fifth instar larvae was disturbed and they died with a darkening of the skin. Ponasterone A also inhibited the growth of the silk glands during the fifth instar. In contrast, the other two phytoecdysones did not greatly influence larval growth. The fourth instar larvae grew rapidly and their ecdysis was advanced with a diet which contained 10 μg of inokosterone/1 g of dry diet. The diet which contained 5 μg of ecdysterone or 10 μg of inokosterone/1 g of dry diet accelerated maturation, while that containing 10 or 20 μg of ecdysterone, or 40 μg of inokosterone, delayed maturation of the fifth instar larvae.Only phytoecdysones caused a decrease in growth of the silk glands in the early half of the instar, and a large amount of phytoecdysones accelerated their growth during the last part of the fifth instar. The fibroin-synthesizing activity was levelled up by feeding ecdysterone and inokosterone, and inokosterone appreciably stimulated activity. Assay of in vitro fibroin synthesis showed that ponasterone A competed with ecdysterone in a stimulative action. Silk formation was much lower in larvae fed the diet containing 5 μg of ecdysterone or 10 μg of inokosterone/1 g of dry diet and was far greater in larvae fed the diet containing 40 μg of inokosterone than in the controls.     

Manipulation of fifth-instar host (Manduca sexta) ecdysteroid levels by the parasitoid wasp Cotesia congregata

Although 5th (last) instar parasitized Manduca sexta larvae undergo developmental arrest and do not wander, they exhibit a small hemolymph ecdysteroid peak (300-400pg/&mgr;l) which begins one day prior to the parasitoid's molt to the 3rd (last) instar and concomitant emergence from the host. Ecdysteroids present in this peak were 20-hydroxyecdysone, 20,26-dihydroxyecdysone and one or more very polar ecdysteroids, as well as small amounts of 26-hydroxyecdysone and ecdysone. In parasitized day-1 and -2 5th instars ligated just behind the 1st abdominal proleg, hemolymph ecdysteroid levels increased in both anterior and posterior portions (100-500pg/&mgr;l), while in unparasitized larvae, hormone levels only increased in the anterior portion (100-350pg/&mgr;l). Thus, the ecdysteroid peak observed in host 5th instars was probably produced, at least in part, by the parasitoids. It may serve to promote Cotesia congregata's molt from the second to the third instar and/or to facilitate parasitoid emergence from the host. In parasitized day-1 and -2 5th instars ligated between the last thoracic and 1st abdominal segments, hemolymph ecdysteroid titers reached much higher levels (500-3500pg/&mgr;l) in the anterior portion (no parasitoids present) than in the posterior portion (150-450pg/&mgr;l). Therefore, it appears that the parasitoid's regulation of hemolymph ecdysteroid titers occurs at two levels. First, parasitization neutralizes the host's ability to maintain its normal hemolymph ecdysteroid levels. Second, in a separate action, the parasitoid manipulates the ecdysteroid-producing machinery so that hemolymph levels are maintained at the 200-400pg/&mgr;l characteristic of day 3-4 hosts. This is the first report of a parasitoid's ability to interfere with the normal inhibitory mechanisms which prevent prothoracic gland production of ecdysteroid at inappropriate periods of insect growth and development.