不同食物对甜菜夜蛾生长发育参数的影响

选择上海地区近年甜菜夜蛾Spodoptera exigua(Hübner)发生比较严重的甘蓝、豇豆、大葱、芦笋4种寄主植物,在温度(27±1)℃,相对湿度70%±5%,光照L:D=14:10的条件下研究了不同寄主植物对甜菜夜蛾发育特征的影响。结果表明,不同寄主植物饲喂甜菜夜蛾,甜菜夜蛾幼虫发育历期、存活率、蛹重、成虫寿命、净生殖力和内禀增长率等参数在寄主植物间有显著差异。用甘蓝饲喂的甜菜夜蛾雌虫产卵量明显高于取食大葱和芦笋的。同样,取食甘蓝的甜菜夜蛾蛹重、净生殖力和种群内禀增长率等参数明显高于取食大葱和芦笋的,说明甘蓝与其他寄主植物相比提供了较好质量的食物。     

Sterol/steroid metabolism and absorption in a generalist and specialist caterpillar: Effects of dietary sterol/steroid structure,mixture and ratio

Insects cannot synthesize sterols de novo, so they typically require a dietary source. Cholesterol is the dominant sterol in most insects, but because plants contain only small amounts of cholesterol, plant-feeding insects generate most of their cholesterol by metabolizing plant sterols. Plants almost always contain mixtures of different sterols, but some are not readily metabolized to cholesterol. Here we explore, in two separate experiments, how dietary phytosterols and phytosteroids, in different mixtures, ratios, and amounts, affect insect herbivore sterol/steroid metabolism and absorption; we use two caterpillars species – one a generalist (Heliothis virescens), the other a specialist (Manduca sexta). In our first experiment caterpillars were reared on two tobacco lines – one expressing a typical phystosterol profile, the other expressing high amounts/ratios of stanols and 3-ketosteroids. Caterpillars reared on the control tobacco contained mostly cholesterol, but those reared on the modified tobacco had reduced amounts of cholesterol, and lower total sterol/steroid body profiles. In our second experiment, caterpillars were reared on artificial diets containing known amounts of cholesterol, stigmasterol, cholestanol and/or cholestanone, either singly or in various combinations and ratios. Cholesterol and stigmasterol-reared moths were mostly cholesterol, while cholestanol-reared moths were mostly cholestanol. Moth tissue cholesterol concentration tended to decrease as the ratio of dietary cholestanol and/or cholestanone increased. In both moths cholestanone was metabolized into cholestanol and epicholestanol. Interestingly, M. sexta generated much more cholestanol than epicholestanol, while H. virescens did the opposite. Finally, total tissue steroid levels were significantly reduced in moths reared on diets containing very high levels of cholestanol. We discuss how dietary sterol/steroid structural differences are important with respect to sterol/steroid metabolism and uptake, including species-specific differences.     

Effects of Transgenic Petunia Expressing Bacillus thuringiensis Toxin on Selected Lepidopteran Pests

Toxicity of 11 transgenic petunia lines expressing the CryIA (c) insecticidal crystal protein of Bacillus thuringiensis subsp . kurstaki was investigated using lepidopteran neonates Spodoptera exigua, Trichoplusia ni and Manduca sexta. Mortality of S. exigua, T. ni and M. sexta varied within and among transgenic petunia lines . Bioassay results demonstrated that levels of CryIA (c) expression obtained in 7 out of the 11 transgenic petunia lines produced at least 50% mortality in S. exigua and T. ni, and all 11 transgenic lines produced more than 80% mortality in M. sexta. Demographic analysis of the biological impact of transgenic petunia on S. exigua revealed that sub - lethal feeding on transgenic petunia significantly reduced larval weight and prolonged larval and pupal development times . Continuous feeding on transgenic petunia significantly reduced lifetime fecundity , egg hatch and longevity in female and male moths . Compared with insects fed continuously on non - transgenic petunia , lifetime fecundity and net reproductive rate were reduced by 58 and 84% in insects fed continuously on transgenic petunia respectively . Mean generation time was 8 days longer for insects fed continuously on transgenic petunia than for insects fed on non - transgenic petunia . Ovipositional attractiveness of transgenic petunia to S. exigua with respect to non - transgenic tomato or lettuce plants was similar , suggesting that petunia / tomato and petunia / lettuce may not be effective trap - cropping combinations . The potential and implications of using transgenic petunia as trap plants interplanted with crop plants for management of lepidopteran pests in the field are discussed .     

幼虫密度对甜菜夜蛾生长发育与繁殖的影响

为了研究甜菜夜蛾Spodoptera exigua(Hübner)幼虫的密度对其发育及繁殖的影响,本实验观察了5种幼虫密度下（1,5,10,20,30头/瓶）,幼虫发育和成虫繁殖情况。结果表明：幼虫和蛹历期、存活率和蛹重均差异显著。幼虫和蛹历期均以20头/瓶的最短,1头/瓶的最长,其余随幼虫密度增加而延长;幼虫至蛹存活率以10头/瓶的最高,其余随幼虫密度增加而降低;1头/瓶的蛹最重,显著高于其他密度的,其余随幼虫密度增加而下降。尽管密度间成虫羽化率和产卵前期均无显著差异,但成虫产卵量、寿命和畸形率差异显著。1头/瓶的产卵量最多,其次为10头/瓶的,其余随幼虫密度增加而减少,30头/瓶的产卵量显著少于其他密度的; 密度在1～20头/瓶范围内,雌蛾寿命均较短,显著短于30头/瓶的,而雄蛾寿命以5头/瓶的最短,显著短于其他密度的（10头/瓶除外）,10头/瓶的次之,其余密度间差异不显著;不同幼虫密度下羽化的成虫畸形率差异显著,10头/瓶的最低,其余随幼虫密度增加而升高; 生命表结果表明甜菜夜蛾在10头/瓶下世代存活率和种群增长指数均最高,幼虫密度过低或过高均不利于种群增长;世代存活率（S）和种群增长指数（I）与幼虫密度之间的关系均呈抛物线关系：S =－0.2087x²+2.5694x+211.52 (R²=0.88),I=－0.0552x²+0.9166x+54.168 (R²=0.95)。结果提示幼虫密度影响甜菜夜蛾种群动态的重要生态因子之一。     

Enzymatic decomposition of elicitors of plant volatiles in Heliothis virescens and Helicoverpa zea

Feeding by larvae of Heliothis virescens induces cotton, corn and tobacco plants to release blends of volatile organic compounds that differ in constituent proportions from blends released when Helicoverpa zea larvae feed on the same plant species. The same elicitors (and analogs) of plant biosynthesis and release of volatiles, originally identified in oral secretions of Spodoptera exigua larvae, were also found in oral secretions of H. virescens and H. zea. However, relative amounts of these compounds, particularly N-(17-hydroxylinolenoyl)-L-glutamine (volicitin), 17-hydroxylinolenic acid, and N-linolenoyl-L-glutamine, varied among batches of oral secretions, more so in H. virescens than in H. zea. This variation was due to cleavage of the amide bond of the fatty acid-amino acid conjugates by an enzyme, or enzymes, originating in the midgut. The enzymatic activity in guts of H. virescens was significantly greater than that found in guts of H. zea. Furthermore, H. zea frass contains N-linolenoyl-L-glutamine in more than 0.1% wet weight, while this conjugate comprises only 0.003% wet weight in H. virescens frass. These results indicated that physiological differences between these two species affect the proportions of volicitin and its analogs in the caterpillars. Whether this causes different proportions of volatiles to be released by plants damaged by each caterpillar species is yet to be determined.     

Pinellia ternata agglutinin expression in chloroplasts confers broad spectrum resistance against aphid, whitefly, Lepidopteran insects, bacterial and viral pathogens

Broad spectrum protection against different insects and pathogens requires multigene engineering. However, such broad spectrum protection against biotic stress is provided by a single protein in some medicinal plants. Therefore, tobacco chloroplasts were transformed with the agglutinin gene from Pinellia ternata (pta), a widely cultivated Chinese medicinal herb. Pinellia ternata agglutinin (PTA) was expressed up to 9.2% of total soluble protein in mature leaves. Purified PTA showed similar hemagglutination activity as snowdrop lectin. Artificial diet with purified PTA from transplastomic plants showed marked and broad insecticidal activity. In planta bioassays conducted with T0 or T1 generation PTA lines showed that the growth of aphid Myzus persicae (Sulzer) was reduced by 89%-92% when compared with untransformed (UT) plants. Similarly, the larval survival and total population of whitefly (Bemisia tabaci) on transplastomic lines were reduced by 91%-93% when compared with UT plants. This is indeed the first report of lectin controlling whitefly infestation. When transplastomic PTA leaves were fed to corn earworm (Helicoverpa zea), tobacco budworm (Heliothis virescens) or the beet armyworm (spodoptera exigua), 100% mortality was observed against all these three insects. In planta bioassays revealed Erwinia population to be 10,000-fold higher in control than in PTA lines. Similar results were observed with tobacco mosaic virus (TMV) challenge. Therefore, broad spectrum resistance to homopteran (sap-sucking), Lepidopteran insects as well as anti-bacterial or anti-viral activity observed in PTA lines provides a new option to engineer protection against biotic stress by hyper-expression of an unique protein that is naturally present in a medicinal plant.     

利用茭白、荸荠及水稻饲养二化螟的技术研究

分别用水稻苗、茭白、水稻 +荸荠 3种方法 ,喂饲二化螟Chilosupperssalis初孵幼虫 ,比较室内 3种饲养方法对二化螟生长发育和繁殖力的影响 ,从而为二化螟的实验室饲养提供方法和依据。其中 ,在整个幼虫期用茭白饲养的二化螟幼虫发育历期和化蛹历期最短 ,分别为 ( 1 7 8± 0 5 )d和 ( 9 8± 0 8)d ,而用水稻苗饲养的二化螟幼虫发育历期和化蛹历期最长 ,分别为 ( 3 0 4± 1 2 )d和 ( 2 0 0± 1 6)d。水稻苗喂饲二化螟的雌雄蛹的比例较低 ,仅为 0 44;荸荠有利于雌性个体的分化 ,用水稻 +荸荠相结合喂饲二化螟的雌雄蛹比例为 1 1 8。用荸荠喂饲的蛹最重 (雌和雄蛹重分别为 71和 5 1mg 头 )。 3种饲养方法对二化螟的产卵率和孵化率无显著影响 ,但是 ,用水稻饲养二化螟蛹的羽化率比其它 2种饲养方法显著降低。由此可见 ,用茭白、水稻苗 +荸荠饲养方法进行二化螟饲养是切实可行的 ,它们有利于二化螟的生长发育。     

不同石榴品种对井上蛀果斑螟生长发育和繁殖的影响

井上蛀果斑螟是云南石榴上的一种主要蛀果害虫.为明确石榴不同品种对该虫生长发育和繁殖的影响,在温度(25±1) ℃、光周期15 L∶9 D、相对湿度(70±10)%条件下,系统观察了井上蛀果斑螟在甜绿籽、厚皮甜沙籽、甜光颜、酸绿籽和酸沙籽5个品种上的发育历期、存活率和产卵量等生物学特性,并组建了其实验种群生命表.结果表明: 井上蛀果斑螟在5个石榴品种上的卵期、幼虫期、蛹期、雄成虫寿命、全世代历期及卵孵化率均无显著差异;预蛹期、雌成虫寿命、幼虫存活率、化蛹率、羽化率、总成活率、单雌产卵量及雌雄蛹质量存在显著差异,除幼虫存活率外,均以酸绿籽种群较高,厚皮甜沙籽种群次之,二者间差异不显著.生命表参数表明,净增殖率、内禀增长率和周限增长率均为酸绿籽种群显著高于甜绿籽、甜光颜和酸沙籽种群,但与厚皮甜沙籽种群差异不显著;平均世代周期各品种间无显著差异;种群加倍时间为酸绿籽种群显著小于其余4个品种.5个石榴品种中,酸绿籽和厚皮甜沙籽较适宜井上蛀果斑螟的生长发育和繁殖.     

Dietary sterols/steroids and the generalist caterpillar Helicoverpa zea: Physiology,biochemistry and midgut gene expression

Sterols are essential nutrients for insects because, in contrast to mammals, no insect (or arthropod for that matter) can synthesize sterols de novo. Plant-feeding insects typically generate their sterols, commonly cholesterol, by metabolizing phytosterols. However, not all phytosterols are readily converted to cholesterol. In this study we examined, using artificial diets containing single sterols/steroids, how typical (cholesterol and stigmasterol) and atypical (cholestanol and cholestanone) sterols/steroids affect the performance of a generalist caterpillar (Helicoverpa zea). We also performed sterols/steroids analyses, using GC/MS techniques, to explore the metabolic fate of these different dietary sterols/steroids. Finally, we used a microarray approach to measure, and compare, midgut gene expression patterns that arise as a function of dietary sterols/steroids. In general, H. zea performed best on the cholesterol and stigmasterol diets, with cholesterol as the dominant tissue sterol on these two treatments. Compared to the cholesterol and stigmasterol diets, performance was reduced on the cholestanol and cholestanone diets; on these latter treatments stanols were the dominant tissue sterol. Finally, midgut gene expression patterns differed as a function of dietary sterol/steroid; using the cholesterol treatment as a reference, gene expression differences were smallest on stigmasterol, intermediate on cholestanol, and greatest on cholestanone. Inspection of our data revealed two broad insights. First, they identify a number of genes potentially involved in sterol/steroid metabolism and absorption. Second, they provide unique mechanistic insights into how variation in dietary sterol/steroid structure can affect insect herbivores.     

Larval and pupal induction and N-terminal amino acid sequence of lysozyme from Heliothis virescens

Fifth instar larvae and prepupae of Heliothis virescens (tobacco budworm) were injected with live Enterobacter cloacae and bled at different times after vaccination. Immune pupal hemolymph showed a 54 times increase in lysozyme activity when compared with normal larval hemolymph, and an 11 times increase of lysozyme activity when compared with immune larval hemolymph. Lysozyme activity of the normal pupal hemolymph increased as greatly as did lysozyme activity of the immune larval hemolymph after metamorphosis. The pupal immune response with regard to lysozyme was much greater than the larval immune response in H. virescens. Lysozyme was purified by heat treatment at 100 degrees C and a chromatography series that included reverse-phase HPLC. The molecular mass of H. virescens lysozyme was approximately 16 kDa by SDS-PAGE which is greater than other insect lysozymes and chicken lysozyme. Amino acid sequence of the N-terminus showed that H. virescens lysozyme is 82% homologous with lysozyme of Manduca sexta and Galleria mellonella. CNBr cleavage of H. virescens lysozyme produced 11 and 6 kDa peptide fragments indicating that one methionine was present, which was also supported by amino acid analysis. However, methionine was located at the carboxyl terminal side rather than the N-terminal side as judged by the N-terminal sequences of each peptide fragment. The residue 22 in most lepidopteran lysozymes is methionine, whereas H. virescens lysozyme had a leucine at residue 22. There was an amino acid deletion near the carboxyl terminal side of H. virescens lysozyme as also found in Trichoplusia ni.     

Functional analysis of two chitinase genes during the pupation and eclosion stages of the beet armyworm Spodoptera exigua by RNA interference

Insect chitinases are a multigene family that is encoded by a rather large and diverse group of genes. The main function of chitinases is to digest the chitin contained in tissues such as the cuticles and gut lining during molting. In this study, we examined the role of a chitinase (SeChi) and a bacterial type chitinase (SeChi-h) during the pupation and eclosion stages of Spodoptera exigua. First, efficient silencing of the SeChi and SeChi-h genes through specific double-stranded RNA (dsRNA) injection led to a significant reduction in the mRNA levels of SeChi and SeChi-h. Additionally, different phenotypic defects were observed at the pupal and adult stages after injection of the SeChi and SeChi-h dsRNAs. After injecting SeChi dsRNA in the pupal stage, the cuticle of the head split open and the pupal cuticle was visible under the old larval cuticle. However, after injecting the SeChi-h dsRNA, animals died without exhibiting any special phenotypes. At the adult death stage, animals injected with dsSeChi could not shed their pupal shell completely, and their old cuticles remained attached to their head or chest. However, the main lethal phenotype was that insects did not emerge after dsSeChi-h injection. Additionally, the average survival rates of S. exigua were 52.02% and 40.38% at the pupal and adult stages, respectively, after injection with SeChi dsRNA. For the insects injected with SeChi-h dsRNA, the survival rates were 72.38% and 48.52%, respectively. These results suggest that SeChi and SeChi-h may have different biologic functions during the pupal-adult molting.     

Effects of larval starvation and adult diet-derived amino acids on reproduction in a fruit-feeding butterfly

Availability of adequate nutrition is among the most important factors affecting growth, development, and reproduction in animals. In holometabolous insects, diets and nutritional needs change between life stages, with larval storage, and adult feeding and reproduction being linked to one another. In several butterfly species, adult feeding is of fundamental importance to realize the full reproductive potential, primarily due to a prominent role of adult diet-derived carbohydrates. In contrast, the role of adult diet-derived amino acids is still under debate, despite the fact that butterflies were often found to preferentially feed on amino acid-rich substrates. Recently it was found that amino acids from adult income could compensate for adverse effects of larval food quality. In our study on the tropical butterfly Bicyclus anynana (Butler) (Lepidoptera: Nymphalidae), we used larval starvation to investigate corresponding effects on female reproductive output. Short periods of larval starvation prolonged development time and reduced larval survival, larval growth rate, pupal mass, and egg size. Regardless of the degree of larval starvation, access to amino acids in the adult diet increased egg size, whereas egg number remained largely unaffected. Thus, although there was some evidence for adult diet-derived amino acids being overall beneficial to reproduction, there was no indication that they can compensate for larval starvation.     

Hexameric storage proteins during metamorphosis and egg production in the diamondback moth, Plutella xylostella (Lepidoptera)

As in many Lepidoptera, Plutella xylostella adults do not feed on protein and females must use accumulated reserves to supply vitellogenin synthesis. Storage proteins were quantified in females and males from the late larval stage through day 4 of adult life. The level of storage protein peaked in the early pupal stage, with females having about twice as much as males. In males, the level fell through pupal development and dropped to a trace by one day after eclosion. In females, level of storage proteins fell until eclosion, and then rose dramatically within four hours after the molt to about 2/3 of the original peak level. This post-eclosion increase, which has not been reported previously in insects, suggests that adult females synthesize hexamerins to resequester amino acids. Subsequently, the level of storage proteins fell as vitellogenin appeared and eggs were laid. The ability to synthesize and sequester amino acids as storage proteins during the adult stage has wide-ranging implication for protein management in insects, particularly those that are long-lived and have flexible schedules of reproduction.     

Viresin. A novel antibacterial protein from immune hemolymph of Heliothis virescens pupae.

Immune hemolymph was collected from fifth instar larvae and 1-day-old pupae of Heliothis virescens after injection of prepupae with live Enterobacter cloacae. Induction of antibacterial activity against Escherichia coli K12 D31 was 7.5 times greater in pupal than in larval immune hemolymph. Lysozyme activity of immune pupal hemolymph against Micrococcus lysodeikticus was 11 times greater when compared with lysozyme activity of immune larval hemolymph. Early pupal immune response with regard to antibacterial activity was much greater than larval immune response in H. virescens. Normal pupal hemolymph showed an increase in antibacterial activity and lysozyme that was induced during metamorphosis. Antibacterial protein was isolated together with lysozyme by gel filtration chromatography and then separated from lysozyme by sequential electrophoresis with a native acid gel and SDS gel. Molecular mass of antibacterial protein was estimated to be 12 kDa. The N-terminal amino acid sequence of 12-kDa protein was different from those of antibacterial molecules found in other insects and has not been identified before. A sample containing 12-kDa protein was negative for immunoblotting with anti-synthetic cecropin B antibody. We have named the novel 12-kDa antibacterial protein viresin. Viresin showed antibacterial activity against several Gram-negative bacteria including E. cloacae but not against Gram-positive bacteria.     

Effect of modified meridic diet on the development and growth of tomato fruitworm Helicoverpa armigera (Lepidoptera: Noctuidae)

The efficacy of one new modified and two old meridic diets on Helicoverpa armigera Hübner (Lepidoptera: Noctuidae) for rearing six successive generations was studied. Duration of larval development for insects fed on the modified diet was considerably shortened as most of them went through only five stadia before pupation, while the per cent pupation and per cent eclosion were relatively higher than on other diets. The lowest pupal mortality (6.33 ± 0.13%) was recorded in the F₁ generation reared on the modified diet, whereas the highest pupal mortality (19.49 ± 0.15%) was observed in insects reared on a natural diet in the F₆ generation. Blending of chickpea Cicer arietinum L. and red kidney bean Phaseolus vulgaris L. flours with tomato paste proved highly favorable for adult reproduction. These results suggest that the vitality of the tomato fruitworm did not decline obviously after rearing on a modified diet for several generations.     

Egg maturation, egg resorption and the costliness of transient egg limitation in insects

Although there is widespread agreement that the cost of oviposition underlies selective oviposition in insects, there is no consensus regarding which factors mediate the cost of oviposition. Models have suggested that egg costs are often paramount in those insects that do not continue to mature eggs during the adult stage (pro-ovigenic insects). Here we address the hypothesis that egg costs are generally less significant in synovigenic insects, which can replenish oocyte supplies through continuous egg maturation. A dynamic optimization model based on the biology of a highly synovigenic parasitoid, Aphytis aonidiae, suggests that the maximum rate of egg maturation is insufficient to balance the depletion of eggs when opportunities to oviposit are abundant. Transient egg limitation therefore occurs, which imposes opportunity costs on reproducing females. Thus, whereas the most fundamental constraint acting on the lifetime reproductive success of pro-ovigenic species is the fixed total number of eggs that they carry at eclosion, the most fundamental constraint acting on a synovigenic species is the maximum rate of oocyte maturation. Furthermore, the ability of synovigenic species to reverse the flow of nutrients from the soma to oocytes (i.e. egg resorption) has a dramatic influence on the cost of oviposition. Whereas females in hostrich environments may experience oviposition-mediated egg limitation, females in host-poor environments may experience oosorption-mediated egg limitation. Both forms of egg limitation are costly. Contrary to initial expectations, the flexibility of resource allocation that typifies synovigenic reproduction actually appears to broaden the range of conditions under which costly egg limitation occurs. Egg costs appear to be fundamental in mediating the trade-off between current and future reproduction, and therefore are an important factor favouring selective insect oviposition.     

Developmental and hormonal regulation of midgut remodeling in a lepidopteran insect, Heliothis virescens

Midgut tissue undergoes remodeling during metamorphosis in insects belonging to orders Lepidoptera and Diptera. We investigated the developmental and hormonal regulation of these remodeling events in lepidopteran insect, Heliothis virescens. In H. virescens, programmed cell death (PCD) of larval midgut cells as well as proliferation and differentiation of imaginal cells began at 108 h after ecdysis to the final larval instar (AEFL) and proceeded through the pupal stages. Expression patterns of pro- cell death factors (caspase-1 and ICE) and anti-cell death factor, Inhibitor of Apoptosis (IAP) were studied in midguts during last larval and pupal stages. IAP, Caspase-1 and ICE mRNAs showed peaks at 48 h AEFL, 96 h AEFL and in newly formed pupae, respectively. Immunohistochemical analysis substantiated high caspase-3 activity in midgut at 108 h AEFL. Application of methoprene, a juvenile hormone analog (JHA) blocked PCD by maintaining high levels of IAP, downregulating the expression of caspase-1, ICE and inhibiting an increase in caspase-3 protein levels in midgut tissue. Also, the differentiation of imaginal cells was impaired by methoprene treatment. These studies demonstrate that presence of JHA during final instar larvae affects both midgut remodeling and larval-pupal metamorphosis leading to larval/pupal deformities in lepidopteran insects, a mechanism that is different from that in mosquito, Ae. aegypti where JHA uncouples midgut remodeling from metamorphosis.     

Life history of Spodoptera exigua (Lepidoptera: Noctuidae) on various host plants

The incubation period of Spodoptera exigua (Hübner) was not influenced by the host plant, whereas larval development time and pupal period were affected. Larval development time was longest on shallot and lady's finger, followed by cabbage and long bean. Larvae did not develop beyond the first instar when fed on chilli. The pupal period was longer on lady's finger than on cabbage, shallot and long bean. Overall, adult longevity was not influenced by the host plant but there was a difference between female and male longevity among the host plants. Survival of S. exigua was affected by the host plant at the larval stage. The number of larval instars varied between 5 and 8 within and between the studied host plants. Long bean was found to be the most suitable host plant and provide the best food quality for S. exigua compared to the other host plants, as it allowed faster development, fewer larval instars and a higher survival rate.     

Molecular cloning and insecticidal effect of Inga laurina trypsin inhibitor on Diatraea saccharalis and Heliothis virescens

Native Inga laurina (Fabaceae) trypsin inhibitor (ILTI) was tested for anti-insect activity against Diatraea saccharalis and Heliothis virescens larvae. The addition of 0.1% ILTI to the diet of D. saccharalis did not alter larval survival but decreased larval weight by 51%. The H. virescens larvae that were fed a diet containing 0.5% ILTI showed an 84% decrease in weight. ILTI was not digested by the midgut proteinases of either species of larvae. The trypsin levels were reduced by 55.3% in the feces of D. saccharalis and increased by 24.1% in the feces of H. virescens. The trypsin activity in both species fed with ILTI was sensitive to the inhibitor, suggesting that no novel proteinase resistant to ILTI was induced. Additionally, ILTI exhibited inhibitory activity against the proteinases present in the larval midgut of different species of Lepidoptera. The organization of the ilti gene was elucidated by analyzing its corresponding genomic sequence. The recombinant ILTI protein (reILTI) was expressed and purified, and its efficacy was evaluated. Both native ILTI and reILTI exhibited a similar strong inhibitory effect on bovine trypsin activity. These results suggest that ILTI presents insecticidal properties against both insects and may thus be a useful tool in the genetic engineering of plants.