The production of azadirachtin by in-vitro tissue cultures of neem,Azadirachta indica

Callus produced from leaves of a Ghanaian strain of the neem tree, Azadirachta indica A. Juss has been shown to produce the natural insecticide azadirachtin when grown in a defined medium. The azadirachtin was isolated by standard procedures of solvent partition and column chromatography monitored by supercritical fluid chromatography. Biological activity was monitored with antifeedant tests using the desert locust (Schistocerca gregaria Forsk.). The azadirachtin was identified by chromatography on three independent chromatographic systems (SFC, HPLC & TLC) and two thin-layer colour tests. It has 100% antifeedant activity at < 0.04mg litre^?1. The yield of azadirachtin was 0.0007% based on dry weight of callus.     

印楝素对马铃薯甲虫的控制作用

为探讨植物源杀虫剂印楝素对马铃薯甲虫的防治作用,采用叶碟法测定印楝素乳油对其成虫的拒食作用,并通过药效试验测定印楝素乳油对马铃薯甲虫的控制效果。结果显示,印楝素对马铃薯甲虫成虫有较强的非选择性拒食作用,且拒食率随取食时间的增加而升高。在取食36 h后,浓度为6、7.5、10、15和30 mg/L印楝素的拒食率分别为55.5%、59.1%、53.4%、76.1%和63.3%,拒食率与浓度无显著相关性;但印楝素对马铃薯甲虫的选择性拒食率较低,各浓度处理间均无显著差异,取食36 h时,选择拒食率约20%;在田间10 mg/L印楝素溶液施药1 d和3 d后,虫口减退率分别为26.52%和50.76%,显著高于对照组,施药9 d后,马铃薯甲虫的校正防治效果为48.86%。表明印楝素对马铃薯甲虫有一定的控制效果。     

Bioactive compounds from neem tissue cultures and screening against insects

Hairy root cultures have been derived from neem (Azadirachta indica A Juss, Family Meliacceae) using Agrobacterium rhizogenes and have been studied for the production of compounds with antifeedant effects on insects. Six-week-old hairy root cultures were extracted, and HPLC yielded fractions ranging from polar to non-polar compounds. High antifeedancy levels against the desert locust were observed in fractions(F) 2, 3 and 4 whilst F1 and F5 were not significantly antifeedant. Interestingly F3 did not contain any of the well-known neem chemicals while F2 contained azadirachtin and 3-tigloylazadirachtol and F4 nimbin and salannin. © 1999 Society of Chemical Industry     

Effects of a commercial neem (<Emphasis Type="Italic">Azadirachta indica</Emphasis>) extract on<Emphasis Type="Italic">Streblote panda</Emphasis> larvae

Biological effects of a commercial formulation of azadirachtin, Align^TM, were assessed onStreblote panda Hübner [1820] (Lasiocampidae) first instars by means of choice and nochoice bioassays. Survival, development, and food consumption were recorded. The product tested did not show clear antifeedant effects, but produced dose-dependent larval mortality. All treatments resulted in significant inhibition of growth, presumably through endocrine regulatory effects. At higher doses assayed the product appears to have direct toxic effects. Our results suggest that Align^TM may be used in IPM programs forS. panda and should be evaluated for efficacy under field conditions. http://www.phytoparasitica.org posting July 10, 2003.     

The antifeedant effect of neem seed kernel extract and azadirachtin on nymphs ofEyprepocnemis plorans

Acridids belonging to different species and families exhibit large differences in their response to neem components. In this context the antifeedant effect of a methanolic neem seed kernel extract (NSKE) and of azadirachtin (AZA) on fourth-instar nymphs of the acrididEyprepocnemis plorans Charpentier (Saltatoria:Acrididae) was investigated. Nymphs were offered either saccharose-impregnated filter paper disks or leaves of broad beans, treated with neem components. The amount of substrate consumed was determined by weighing the filter paper or by measuring the leaf area. On filter paper both NSKE and AZA were highly active down to the 10⁻⁴% treatment. In the leaf treatment, however, AZA was definitely more active than NSKE, with 100% deterrence at 10⁻⁴% and 10⁻²%, respectively. The methanolic NSKE was somewhat more active than the commercial preparation ‘Neemark’.     

Lethal and sublethal effects of the neem insecticide formulation, ‘Margosan-O’, on the pea aphid

The effects of ‘Margosan-O’ (MO) on the pea aphid, Acyrthosiphon pisum (Harris), were determined. MO significantly reduced population increase of A. pisum in a concentration-dependent manner. At a concentration equivalent to 100 mg litre^?1 of azadirachtin, population increase was c. 3.5 times lower than the control. In more detailed studies, MO significantly reduced the number of molts, longevity, and fecundity of A. pisum that had been reared on treated broad bean. Viciafaba L., plants. MO also reduced the longevity and fecundity of young adult A. pisum exposed to MO-treated broad bean. MO was slow-acting against A. pisum. Mortality caused by MO stabilised seven days after newborn A. pisum were exposed to treated broad bean and 10 days for adults. The seven day LC₅₀ for individuals exposed from birth was 27.50 mg azadirachtin liter^?1 while the 10 day LC₅₀ for adults was 53.32 mg liter^?1. Contrary to previous studies suggesting that neem insecticides are not contact toxicants, we found that MO applied topically to adult A. pisum caused effects similar to those found in individuals that fed upon treated plants. However, MO was slower-acting when applied topically. Mortality in adult A. pisum caused by topically applied MO stabilised 17 days after treatment with a resultant LD₅₀of 2.91 μg azadirachtin g^?1.     

Effect of Clerodane-Type Diterpenoids Isolated from Salvia spp. on the Feeding Behaviour of Spodoptera littoralis

Twenty-nine clerodane-type diterpenoids isolated from species of Salvia growing in Mexico, or analogues of them, were tested for antifeedant activity against Spodoptera littoralis larvae using behavioural and electrophysiological bioassays. Eight of the compounds showed potent antifeedant activity in choice and no-choice bioassays and these compounds also stimulated dose-dependent responses from neurones in the lateral styloconic sensilla of S. littoralis.     

Bicyclophosphorothionate antagonists exhibiting selectivity for housefly GABA receptors

2,6,7-Trioxa-1-phosphabicyclo[2.2.2]octane 1-sulfides (bicyclophosphorothionates) with various C_1–4 alkyl groups at the 3- and 4-positions were synthesized and tested for their ability to compete with [³H]4′-ethynyl-4-n-propylbicycloorthobenzoate (EBOB), a non-competitive antagonist of γ-aminobutyric acid (GABA) receptors, for specific binding to rat-brain and housefly-head membranes, and for their insecticidal activity against houseflies. Among the 3,4-substituted analogues, 20 compounds were selectively active for housefly GABA receptors versus rat GABA receptors. The 3-alkyl groups of C₃ length and the 4-alkyl groups of C₄ length were tolerated in housefly receptors, whereas such bulky substituents were deleterious in rat receptors. The 4-isobutyl-3-isopropyl analogue was the most potent in housefly receptors (IC₅₀ = 45.2 nM ), and tert-butylbicyclophosphorothionate (TBPS), with the 4-tert-butyl group and no 3-substituent, was the most potent in rat receptors (IC₅₀ = 62.2 nM ). Their receptor selectivities (rat IC₅₀/housefly IC₅₀) were 52 and 0.038, respectively. The insecticidal activity (LD₅₀) of 20 active analogues was well correlated with their potency (IC₅₀) in inhibiting [³H]EBOB binding to housefly-head membranes (r = 0.93). The results obtained in the present study indicate that the introduction of appropriate alkyl groups into the 3- and 4-positions of bicyclophosphorothionate leads to non-competitive antagonists with increased affinity and selectivity for housefly ionotropic GABA receptors versus rat GABA_A receptors. © 1999 Society of Chemical Industry     

The toxicities of some analogues of dieldrin,endosulfan and isobenzan to bloodsucking diptera,especially tsetse flies

The dieldrin analogue, 1, 8, 9, 10, 11, 11-hexachloro-4, 5-exo-epoxy-2, 3-7, 6-endo-tricyclo-[6.2.1.0^{2, 7}]undec-9-ene (HEOM), and the isomeric 3, 4-exo-epoxy-and 3, 6-endo-epoxy compounds (HCE and ODA, respectively), incorporating structural modifications of the non-chlorinated ring system, were tested against adult mosquitoes (mainly Anopheles stephensi List.), tsetse flies (mainly Glossina austeni Newst.) and stable flies, Stomoxys calcitrans (L.). The order of toxicity was ODA > HCE > HEOM. In further tests of residual activity against A. stephensi, ODA was better than HCE but neither was sufficiently persistent to be a promising mosquito toxicant. Analogues of dieldrin (HEOD), endosulfan and isobenzan with reduced numbers of chlorine atoms were tested against G. austeni. In the pentachloro-analogues of dieldrin and α-endosulfan, the anti-bridge chlorine was more important than the syn-chlorine for toxicity. The general effect of reductive dechlorination was to reduce toxicity, but the effect was small for some endosulfan analogues and in one case there was an increase in toxicity. In the main, toxicities of the endosulfan analogues compared favourably with those of the dieldrin analogues. However, toxicity fell drastically following replacement of the second ethylenic chlorine in isobenzan. The effect of these structural changes on toxicity evidently varied from one series to another and was influenced in an unpredictable manner by the rest of the molecule.     

Effects of azadirachtin on Tetranychus urticae (Acari: Tetranychidae) and its compatibility with predatory mites (Acari: Phytoseiidae) on strawberry

BACKGROUND: The spider mite, Tetranychus urticae, is the major strawberry pest in Brazil. The main strategies for its control comprise synthetic acaricides and predatory mites. The recent register of a commercial formula of azadirachtin (Azamax^® 12 g L^?1) can be viable for control of T. urticae. In this work, the effects of azadirachtin on T. urticae and its compatibility with predatory mites Neoseiulus californicus and Phytoseiulus macropilis in the strawberry crop were evaluated. RESULTS: Azadirachtin was efficient against T. urticae, with a mortality rate similar to that of abamectin. In addition, the azadirachtin showed lower biological persistence (7 days) than abamectin (21 days). Azadirachtin did not cause significant mortality of adult predatory mites (N. californicus and P. macropilis), but it did reduce fecundity by 50%. However, egg viability of the azadirachtin treatments was similar to that of the control (>80% viability). The use of azadirachtin and predatory mites is a valuable tool for controlling T. urticae in strawberry crop. CONCLUSIONS: Azadirachtin provided effective control of T. urticae and is compatible with the predatory mites N. californicus and P. macropilis. It is an excellent tool to be incorporated into integrated pest management for strawberry crop in Brazil. Copyright © 2012 Society of Chemical Industry     

2-arylalkanoates,a new group of synthetic pyrethroid esters not containing cyclopropanecarboxylates

Although structure modifications of natural pyrethrin constituents have disclosed a variety of potent synthetic analogues, all known examples are cyclopropanecarboxylate esters, a grouping that appeared to be essential for insecticidal activity. Some new substituted 2-phenylalkanoates, whose biological activities are of a similar nature and potency to those of conventional pyrethroids, are now reported. 5-Benzyl-3-furylmethyl and 3-phenoxybenzyl 3-methyl-2-phenylbutyrates and their analogues are potent insecticides. Activity is increased on the introduction of appropriate groups into the 3 and/or 4-positions of the aryl ring and the (S)-2-phenylalkanoates are far more active than their (R)-enantiomorphs. Structure/activity relationships are compared with those for conventional pyrethroids. Some of the new series compare favourably with typical insecticides in tests against Musca domestica, Spodoptera litura and Plutella xylostella.     

Biological activity of neem seed kernel extracts and synthetic azadirachtin against larvae of Plutella xylostella L

The activity of two neem extracts, AZT and NEEM-AZAL (containing 30 and 3 mg azadirachtin ml^?1 respectively) and synthetic azadirachtin (AZ) against second-instar larvae (L2) of Plutella xylostella L. was examined using leafdip bioassays. On Chinese cabbage, AZ was significantly (P <0.05) less toxic (3 to 4-fold; LC₅₀ 0.54 μg AZ ml^?1) than either neem extract against a laboratory strain of P. xylostella (FS). The LC₅₀ values for AZT against the FS and another laboratory strain (Wellcome) were not significantly different on Chinese cabbage. The activity of AZT against the FS and Wellcome strains was similar on Chinese cabbage and Brussels sprout. AZT was significantly less toxic (3-fold) on Brussels sprout against an acylurea-resistant field strain (Sawi) when compared with the FS strain on Chinese cabbage. Larval mortality (at day 13) was found to increase with increasing exposure time of P. xylostella (FS) larvae to AZT-treated Chinese cabbage, although there was little difference in mortality between 48 and 120 h exposure. When AZT, NEEM-AZAL and AZ were applied at a dose (1 μg AZ ml^?1) which gave end-point mortalities between 50 and 90% (at day 13), all treatments delayed the development of a proportion of surviving larvae but no morphogenetic abnormalities were observed in larvae which reached pupation. Evidence for antifeedant (reduced weight gain) and repellant effects (choicechamber) for AZT were observed with L2 P. xylostella (Wellcome) on Chinese cabbage. AZT was also shown to have ovicidal activity against P. xylostella (Wellcome) at relatively high dose ranges (10-1000 μg AZ ml^?1) as well as some contact activity (FS strain) in topical bioassays. In residual bioassays on glass with adults of the hymenopteran endo-larval parasitoid of P. xylostella, Diadegma semiclausum (Ichneumonidae), AZT showed little or no activity at rates up to 1000 μg AZ ml^?1. In medium-volume (MV, 200 litre ha^?1) and ultra-low-volume (c. 1 litre ha^?1) spray bioassays on Brussels sprout, AZT gave 16-92% and 88-100% mortality respectively (Wellcome strain) at rates approximating to 1-20 g AZ ha^?1. The residual activity of AZT and NEEM-AZAL against P. xylostella (FS) on Brussels sprout (MV spray) was observed to decrease appreciably after three days, the decline in activity being particularly marked for NEEM-AZAL.     

Effect of azadirachtin-derived decalin (perhydronaphthalene) and dihydrofuranacetal (furo[2, 3-fo]pyran) fragments on the feeding behaviour of Spodoptera littoralis

Decalin and dihydrofuranacetal fragments related to those in azadirachtin exhibited antifeedant activity against larvae of the African leaf-worm Spodoptera littoralis. Boisd. All the decalin fragments tested were methoxy (C11) derivatives of azadirachtin. The most active decalin fragment had a ketone substitution at C7. Overall, the compounds were more active when tested in combinations of one decalin fragment and one dihydrofuranacetal fragment than when tested singly. Although some of these combinations did show significant levels of antifeedant activity, sometimes coupled with a synergistic effect, they were not as active as either azadirachtin or dihydroazadirachtin.     

Effect of azadirachtin applied systemically through roots of plants on the mortality,development and fecundity of the cabbage aphid (<Emphasis Type="Italic">Brevicoryne brassicae</Emphasis>)

The effect of low concentrations of azadirachtin applied systemically through root tissues of rape plants (Brassica napus subsp.napus) on the mortality, period of development, longevity and fecundity of the cabbage aphid (Brevicoryne brassicae L.) (Sternorrhyncha: Aphididae) was studied. The concentrations of azadirachtin A tested were: 0.25, 0.05, 0.005, 0.0005 and 0.00005 mg ml⁻¹. The mortality of cabbage aphid nymphs, particularly during the period of ecdysis, increased significantly with increasing concentrations. Azadirachtin had no effect on the length of the development period of the respective immature stages of cabbage aphids. The average longevity of the cabbage aphids decreased with increasing azadirachtin concentration and time spent feeding. The fecundity of aphids decreased after the application of azadirachtin. http://www.phytoparasitica.org posting May 6, 2004.     

嗜线虫致病杆菌对小菜蛾的致死和亚致死效应室内评价

嗜线虫致病杆菌是与昆虫病原线虫共生的细菌,对小菜蛾等多种害虫具有广谱的杀虫活性。为了深入了解该菌的杀虫活性,在室内测定了嗜线虫致病杆菌HB310 ( 简称 Xn HB310)菌株对小菜蛾幼虫的致死和亚致死效应以及持效期。结果表明, Xn HB310菌液对小菜蛾2龄和3龄幼虫72 h的LC50值分别是3.32×104 和8.56×104 CFU/mL,其活性组分主要存在于菌株细胞和经50 kDa超滤膜超滤后的上清液中。此外, Xn HB310菌液对小菜蛾幼虫还表现出明显的拒食活性,其2龄和3龄幼虫的拒食中浓度(AFC50)分别为4.03×105和8.48×105 CFU/mL。将2.24×106 CFU/mL菌液喷施到甘蓝苗上,其在室内的持效期为3~4 d。用亚致死剂量(LC30和LC50)的菌液饲喂小菜蛾2龄幼虫,存活幼虫的生长发育指标、蛹重、雌虫寿命以及成虫产卵量均明显下降,推测这可能导致小菜蛾下一代种群数量减少。研究结果表明,嗜线虫致病杆菌HB310菌株具有开发成为杀虫剂的潜力。     

Antifeedant activity of fruit and seed extracts ofMelia azedarach andazadirachta indica on larvae ofSesamia nonagrioides

Methanolic extracts of seeds and fruits of the chinaberry tree,Melia azedarach L. (Meliaceae), showed strong antifeedant activity against 2nd instar larvae ofSesamia nonagrioides Lefèbvre (Lepidoptera: Noctuidae), a very serious pest of maize(Zea mays L.) in Mediterranean countries. Extracts were applied in an artificial diet at concentrations of 1000 and 2000 ppm. The parameters used to evaluate the activity were larval growth rates; quantity of food ingested; phagodepression/phagostimulation index; quantity of frass produced; quantity of material ingested; duration of larval development; and cumulative mortality. Seed extract showed high bioactivity at both doses, while fruit extract proved to be less active, and only at the higher dose used (2000 ppm) did it display a slight antifeedant activity. The activity of theM. azedarach seed extract at the higher dose (2000 ppm) was comparable to that of pure azadirachtin applied at a dose of 1.25 ppm, or to ‘Mubel’, a commercial extract ofAzadirachta indica A. Juss. (Meliaceae), applied at a dose of 75 ppm.     

Laboratory studies with antifeedants against larvae of Boarmia (Ascotis) selenaria on alfalfa

The antifeedant effect of five organotin compounds (fentin acetate, fentin hydroxide, fentin chloride, cyhexatin and fenbutatin oxide) and of three other compounds known to possess antifeeding activity (guazatine triacetate, 4′-chloroacetophenone isobutyrylhydrazone and thiram) was investigated by feeding treated alfalfa to larvae of Boarmia selenaria. The three fentins and cyhexatin were highly active at 0.1 g litre^?1, the order of activity at this concentration being fentin chloride > fentin hydroxide > fentin acetate ? cyhexatin, whereas the other compounds were practically inactive at either this or a somewhat higher concentration (0.5 g litre^?1). Guazatine triacetate had a concentration-independent antifeedant effect at much higher concentrations (5–20 g litre^?1).     

Effect of uneven application of azadirachtin on reproductive and anti-feedant behaviour of Rhyzopertha dominica (Coleoptera: Bostrichidae)

The effects of uneven application of an azadirachtin-enriched neem extract to wheat grain on the reproduction and feeding behaviour in Rhyzopertha dominica (F) were investigated. The evenness of distribution amongst individual wheat grains during grain treatment was not important in ensuring its effectiveness as an anti-feedant or as an insect growth regulator, provided that, overall, the grain was treated with an effective level of azadirachtin. The effect of azadirachtin on reproduction was not improved when the diluent application rates were increased from 0.5 ml kg⁻¹ to 5 ml kg⁻¹. Treating 10% of the grain was shown to provide the same level of protection as treating 100% of the grain. The anti-feedant effect of azadirachtin on the beetle was also unaffected by unevenness of treatment, provided that 50% of the grain was treated. The fact that azadirachtin remains effective even when application to grain is uneven may be advantageous in field situations where uniform distribution of insecticides is difficult to achieve. © 1999 Society of Chemical Industry     

Behandlung von Reis mit Neem und Diatomeenerde zur Bek?mpfung der vorratssch?dlichen Coleopteren Sitophilus oryzae und Tribolium castaneum

The commercial available diatomaceous earth Fossil Shield® and the neem product Neem Azal-T/S® are tested as single treatments and in combination as controls for the stored product pests Sitophilus oryzae (L.) and Tribolium castaneum (Herbst). The diatomaceous earth, applied in concentrations of 0.5 g, 1 g and 2 g kg^-1 rice, reduced numbers of surviving beetles significantly (over a period of 3 weeks). A single treatment with the neem product, in concentrations of 0.01 g, 0.1 g, 0.2 g and 1 g azadirachtin kg^-1 rice, increased the mortality rate for both species significantly. The combination of neem and diatomaceous earth (1 g diatomaceous earth with 0.2 g or 1 g azadirachtin kg^-1 rice) was more effective than the single treatment in reducing numbers of surviving beetles.     

Behandlung von Reis mit Neem und Diatomeenerde zur Bek?mpfung der vorratssch?dlichen Coleopteren Sitophilus oryzae und Tribolium castaneum

The commercial available diatomaceous earth Fossil Shield® and the neem product Neem Azal-T/S® are tested as single treatments and in combination as controls for the stored product pests Sitophilus oryzae (L.) and Tribolium castaneum (Herbst). The diatomaceous earth, applied in concentrations of 0.5 g, 1 g and 2 g kg^-1 rice, reduced numbers of surviving beetles significantly (over a period of 3 weeks). A single treatment with the neem product, in concentrations of 0.01 g, 0.1 g, 0.2 g and 1 g azadirachtin kg^-1 rice, increased the mortality rate for both species significantly. The combination of neem and diatomaceous earth (1 g diatomaceous earth with 0.2 g or 1 g azadirachtin kg^-1 rice) was more effective than the single treatment in reducing numbers of surviving beetles.