Azadirachtin Research Articles

Acute and Sublethal Effects of a Neem Insecticide on the Commercial Biological Control Agents Phytoseiulus persimilis and Amblyseius cucumeris (Acari: Phytoseiidae) and Aphidoletes aphidimyza (Diptera: Cecidomyiidae)

We tested the effects of a neem insecticide, containing azadirachtin (AZA) as the active ingredient, on fecundity, adult and egg mortality, and preference for neem-treated or untreated foliage on 2 predacious mite species, Phytoseiulus persimilis Athias-Henriot, and Amblyseius cucumeris (Oudemans). We also tested the effects of neem on egg, larval, and adult survival,, and on ovipositional preference for neem treated or untreated foliage, on the aphid predator Aphidoletes aphidimyza (Rondani). We found no significant differences in egg or adult mortality between treated and untreated mites for either species. For A. cucumeris , untreated females that contacted leaf disks and prey treated with 60 ppm AZA laid fewer egg after 48 h than females on untreated disks. In leaf-disk choice tests, significantly more P persimilis were found on untreated disks than those treated with 60 ppm after 24 h, and significantly more A. cucumeris were found on untreated disks than those treated with 60 ppm after 48 h. However, the number of eggs laid on treated versus untreated disks in these choice tests were not significantly different. For A. aphidimyza we found no significant differences in the percentage eclosion between neem-treated and untreated eggs. However, larval mortality ranged from 30 to 100% for 1st instars treated with neem, and adult emergence of treated larvae was significantly lower for 1st- and 3rd-instar A. aphidimyza compared with untreated controls. First and 3rd instars that acquired neem from contact with treated foliage or ingestion of treated aphids had greater mortality than larvae that were sprayed directly with neem solutions. No significant differences in the sex ratio of adults of the surviving neem treated and control larvae occurred. Adult A. aphidimyza exposed to 60 ppm AZA residues for 48 h in residual contact tests had significantly greater mortality (13%) compared with controls (5%). No significant differences in the number of eggs laid on treated and untreated mustard cabbage plants were apparent. We compared our results with previously published data on the effect of neem on beneficial arthropods.

Root uptake, translocation, accumulation and dissipation of the botanical insecticide, azadirachtin, in young spruce trees

Abstract A formulation containing the biopesticide azadirachtin‐A (AZ‐A), extracted from the seeds of neem tree, Azadirachta indica A. Juss, was applied (10 μg/mL) in a nutrient solution (200 mL) to the roots of young spruce trees [Picea glauca (Moench) Voss]. The uptake, translocation, persistence and dissipation of AZ‐A in spruce were studied. The insecticide was taken up by the root system, translocated via xylem vessels and accumulated in the photosynthate sinks (areas of new growth, especially shoots), confirming that AZ‐A is systemic. The peak AZ‐A concentrations (μg/g, fresh weight) in the shoots (5.16 ± 0.73) and needles (2.56 ± 0.31) occurred at 8 and 15 d after treatment, respectively. The AZ‐A concentrations in bark and wood during this period were very low, however they were high in the root samples. The dissipation of AZ‐A from the shoots was rapid compared to other matrices. The initial (0‐d) and final (20‐d) AZ‐A concentrations (μg/mL) in the nutrient solution were 8.99 ± 1.24 and 3.24 + 0....

Photostabilization of the botanical insecticide azadirachtin in the presence of lecithin as UV protectant

Abstract The phytochemical insecticide, azadirachtin (AZ), undergoes UV‐induced photodegradation. Using the isomer AZ‐A as a standard, its photochemical stability was studied with and without adding lecithin surfactant as a UV protectant. Standard solutions of pure AZ‐A and Margosan‐O® were prepared in methanol‐hexane with (AZ‐A:lecithin, 1:2 by weight) and without lecithin, applied separately onto glass plates and maple (Acer L.) foliage and exposed to radiant energy under controlled conditions. Noticeable photostabilization of AZ‐A was achieved in the samples containing lecithin compared to AZ‐A samples without the lecithin additive. First‐order kinetic evaluation of the data showed that the DTy50 (half‐life) and C (rate constant) values for AZ‐A with and without lecithin on glass plates were 5.68 d and 0.122, and 5.42 d and 0.128, respectively. The corresponding values for the Margosan‐0 formulation were 7.37 d and 0.094, and 6.24 d and 0.111. The DT50 and C values for the pure AZ‐A on maple foliage wi...

Azadirachtin biopesticide: A review of studies conducted on its analytical chemistry, environmental behaviour and biological effects

Abstract This paper provides a brief review of studies conducted in our laboratory on the analytical chemistry, environmental behaviour and biological effects of azadirachtin‐A, a neem‐based pesticide. Azadirachtin‐A (AZ‐A) was isolated from samples of neem seed kernels, leaves, bark, root and stem, obtained from Kanthayapalayam, South India. The extracts were subjected to column cleanup and AZ‐A concentration was quantified by using a high‐performance liquid chromatographic (HPLC) method. A flow diagram is given describing the procedures involved. AZ‐A was also isolated from four commercial formulations and from several forestry substrates, and its concentration in each sample was quantified by HPLC. To investigate the environmental behaviour of AZ‐A, five studies were conducted: (i) persistence of AZ‐A on balsam fir and red oak foliage, (ii) dissipation of AZ‐A in forest nursery soils, (iii) leaching in sandy loam forest soil, (iv) adsorption and desorption of AZ‐A in sandy loam soil, and (v) rate of hy...

Azadirachtin: Potential Use for Controlling Lepidopterous Insects and Increasing Marketability of Cabbage

Azadirachtin (ATI), an insect growth regulator derived from extracts of neem (Azadirachta indica A. Juss) seed, was evaluated for the control of cabbage looper (Trichoplusia ni Hübner), diamondback moth (Plutella xylostella L.), and silverleaf whitefly (Bemisia argentifolii Bellows and Perring) in cabbage (Brassica oleracea L. Capitata Group) grown in southwestern Texas. In Fall 1992, ATI was tested with the a.i. at 0, 22, 33 and 44 g·ha–1. In 1993, ATI was evaluated at 33 g·ha–1 and in combination with M-Pede (1%, v/v), an organic insecticide based on potassium salts of fatty acids at 49%. Two commercial (Align and Neemix) and one experimental hydrogenated (LDF) ATI formulations were evaluated at 11 g·ha–1 in 1994. Insect populations were monitored weekly before and after treatment application. Plant damage was evaluated immediately before harvest, and marketable yields were determined. In 1992, large (>6 mm long) and total cabbage looper counts were reduced by ATI compared with the nontreated control. Insect mortality was similar for all ATI rates tested in 1992. In 1993, ATI at 33 g·ha–1 + M-Pede reduced the number of cabbage looper and diamondback moth larvae. ATI efficacy against cabbage looper and diamondback moth was enhanced when crop oil (polyol fatty acid esters with polyethoxylated derivatives) was tank-mixed with Align or LDF formulations in 1994. ATI did not reduce the number of silverleaf whitefly nymphs compared to the control. In all seasons, ATI-treated plants had lower insect-induced plant damage and higher marketable head weights than the nontreated control. Using ATI on lepidopterous pests appears to be beneficial for integrated pest management strategies.

Uptake, translocation, persistence and fate of azadirachtin in aspen plants ( Populus tremuloides Michx.) and its effect on pestiferous two-spotted spider mite ( Tetranychus urticae Koch)

A commercial neem formulation containing azadirachtin-A (AZ-A) was applied to the soil around the root system of potted aspen ( Populus tremuloides Michx.) plants. The uptake, translocation, persistence and dissipation of the chemical in the plants were studied. The effect of foliar residues of AZ-A on two-spotted spider mite ( Tetranychus urticae Koch) populations was also evaluated. The compound was taken up by the root system within 3 h and translocated in the stem and foliage within 3 days, confirming that AZ-A is systemic. The peak concentrations (μg/g, fresh weight) of AZ-A occurred at 10 days post-treatment, and were distributed in roots, stem and foliage in the ratio of 8.1:1.0:2.3, respectively. The rate of dissipation of AZ-A from the matrices was moderately rapid, and the residual concentrations on the last day of sampling (50 days post-treatment) in roots, stem and foliage were in the ratio 2.7:1.0:1.2, respectively. Control of mites by AZ-A residues in foliage was statistically significant, and the bioactivity declined within 30 days. The final residue of AZ-A in the soil after 50 days was about 25% of the initial value, with a half-life of dissipation of about 26 days.

Sorptive behavior of the neem-based biopesticide, azadirachtin, in sandy loam forest soil

Abstract Adsorption‐desorption of azadirachtin‐A (AZ‐A) was studied at pH 4.0 and 7.0 in sterilized and unsterilized sandy loam forest soil, by batch equilibrium method, using RH‐9999®, an experimental wettable powder (WP) formulation containing 20 % of AZ‐A. Data generated were fitted to linear, Freundlich and Langmuir adsorption isotherms. Adsorption of AZ‐A to the soil was not high and decreased with increasing pH. Differences observed in the adsorption between sterilized and unsterilized soil were attributed to alterations in soil porosity and aggregation of soil particles. The KD and KF parameters, and the Freundlich exponential term (1/n) were low, indicating that the interactions between the soil particles and AZ‐A molecules were weak due to physical forces and H‐bonding. Desorption studies showed that the adsorbed AZ‐A was not firmly retained by the soil participates and was reversible. If microbial degradation is not high, desorbed AZ‐A leaching to lower soil horizons is possible. The soil parameters, KOC and KOM, were calculated using the KD and the measured percent OC and OM values. Also, KOC was computed by using the KOM and the Van Bemmelen factor, and the two KOC values were compared.

Effects of pure and formulated azadirachtin, a neem-based biopesticide, on the phytophagous spider mite, Tetranychus urticae koch

Abstract Pure azadirachtin‐A (AZ‐A) and four neem‐based formulations (RH, MO, PT and AT) containing the insecticide isomer were tested for their repellency, toxicity and oviposition deterrence against the phytophagous two‐spotted spider mite (Tetranychus urticae Koch). Mites were placed on treated and untreated aspen (Populus tremuloides Michx.) leaf discs and their fecundity, feeding rate and mortality were assessed. Results indicated that the effects of AZ‐A on T. urticae varied with formulation type and AZ‐A concentration. Significant reductions in feeding and oviposition, which correlated with AZ‐A concentrations, were recorded concomitantly with a significant increase in repellency. Leaf discs treated with increasing concentrations of the formulations showed increased mortality of T. urticae, reduction in the total number of eggs laid, reduced % of eggs hatched and reduced survival of emerged mites. The deterrent and biological effects decreased in the order AT>PT>MO>RH>AZ‐A.

Insect growth regulating effects of neem extract and azadirachtin on aphids

Neem, Azadirachta indica (A. Juss.), seed oil (NSO) applied to leaf discs at a concentration of 1.0% resulted in 94% to 100% mortality of second instar nymphs of currant-lettuce aphid, Nasonovia ribis-nigri (Mosley), and green peach aphid, Myzus persicae (Sulzer), after nine days. The equivalent amount of pure azadirachtin (AZA) (ã40 ppm), the principle active ingredient of neem, was as effective as NSO. The survival of adult aphids was unaffected by NSO or AZA, but the survival of offspring from treated adult M. persicae and N. ribis-nigri was reduced significantly. The lethal concentration of AZA resulting in 50% mortality of second instar nymphs of nine species of aphids ranged from 2.4 ppm for M. persicae on pepper to 635.0 ppm for the strawberry aphid, Chaetosiphon fragaefolii (Cockerell), on strawberry. For M. persicae, the growth regulating effect of AZA was influenced by the host plant and the nymphal instar treated.

The effects of azadirachtin A on the morphology of the ring complex of Lucilia cuprina (Wied) larvae (Diptera: Insecta)

This study investigated the effect of azadirachtin A (AZA) on the ultrastructure of the endocrine glands (the prothoracic gland, the corpus allatum, and the corpus cardiacum) responsible for controlling moulting and ecdysis in Lucilia cuprina larvae. All constituent endocrine glands within the ring complex exhibited ultrastructural change after exposure of larvae to AZ A in the diet. The effects on the nucleus were the most noticeable and included: (1) crenulation of nuclear shape, (2) clumping of heterochromatin, and (3) pyknosis. The degenerative changes to the nuclei appeared to precede changes within the cytoplasm. It was concluded that the degeneration of all three endocrine glands within the ring complex would contribute to a generalised disruption of neuroendocrine function.

Inhibition by azadirachtin of phospholipid transfer from lipophorin to the oocytes in Rhodnius prolixus

AbstractThe growth of oocytes was severely reduced in female Rhodnius prolixus treated with azadirachtin A (AZA). When administered in a blood meal, AZA (1 μg/ml) inhibited phospholipid transfer to the ovaries without altering the availability of yolk protein in the hemolymph or its uptake by 1 mm oocytes. The lipid composition of lipophorin, its concentration in the hemolymph, and its density were normal in AZA‐treated females. Partial inhibition of phospholipid transfer was observed when lipophorin from AZA‐treated females was injected into normal females, or when lipophorin from normal females was injected into AZA‐treated females. The two effects were additive, so that the transfer of phospholipids from the lipophorin of AZA‐treated females to the oocytes of AZA‐treated females was nearly eliminated. In combination, these effects of AZA on phospholipid transfer to the oocytes limit the capacity of AZA‐treated females to produce eggs. © 1994 Wiley‐Liss, Inc.

High Performance Liquid Chromatographic Determination of Azadirachtin in Conifer and Deciduous Foliage, Forest Soils, Leaf Litter and Stream Water

Abstract A high performance liquid chromatographic (HPLC) method for the determination of azadirachtin (AZ-A) in conifer and deciduous foliage, forest soil, litter and stream water is described. AZ-A was extracted from the solid matrices by aqueous methanol, concentrated, partitioned with hexane and re-extracted into dichloromethane layer. After evaporation of the dichloromethane layer, the residues were dissolved in ethyl acetate, purified on a Florisil® mini-column, eluted with ethyl acetate and analyzed using a reversed-phase C-18 column, with UV detection at 210 nm and acetonitrile/water gradient system. Analysis of AZ-A in the stream water required only the steps from dichloromethane extraction onwards. Limits of detection were 0.2 μg/g for foliage and soil. 0.25 μg/g for litter and 5.0 μg/L for water. Overall mean recoveries from terrestrial samples fortified with AZ-A at 0.50 μg/g to 10.0 μg/g and stream water at 10.0 μg/L to 200 μg/L were > 80% with good reproducibility. The applicability of the method was demonstrated by studying the recover) levels of AZ-A in samples fortified with standard Margosan-O.

Corpus allatum activity and the effects of juvenile hormone on oocyte development in azadirachtin-induced over-aged nymphs ofLocusta migratoria(L.)

Summary The corpora allata (CA) of azadirachtin (AZA)-induced over-aged 5th instar female nymphs of Locusta migratoria were relatively inactive during the first 20 days in the instar. They afterwards produced juvenile hormone (JH) III in vitro, coinciding with limited vitellogenic oocyte development. The basal rate of JH production and the length of the vitellogenic oocytes were lower in the over-aged nymphs, but the farnesoic acid (FA)-stimulated rate of JH production and volume of CA were higher than in normal adult females. Injection of massive doses of methoprene into the over-aged nymphs slightly enhanced vitellogenic oocyte development, which still remained incomplete. Chemical allatectomy by precocene III prevented, and injection of JH III or of methoprene restored, the limited vitellogenic oocyte development in the over-aged nymphs. Therefore, some factor other than JH shortage—possibly inadequate competence of the ovaries/ovarioles, would seem to limit vitellogenic oocyte development in over-aged nymphs.

High performance liquid chromatographic method for the analysis of azadirachtin in two commercial formulations and neem oil

Abstract An improved high performance liquid Chromatographic method is reported to determine azadirachtin (AZ) content in neem formulations and neem oil. Aqueous methanol solutions of samples were extracted sequentially with hexane and dichloromethane. The dichloromethane layer, containing AZ, was taken to dryness and reconstituted in ethyl acetate. It was purified by Florisil® column cleanup, eluted with ethyl acetate and injected into the liquid Chromatograph. Separation was performed with a reversed‐phase Spherisorb C‐18 ODS 5 μm column with an acetonitrile/water gradient system. The eluent was monitored with UV detection at 210 nm. The method was found to be sufficiently rapid and reproducible for use in the analysis of AZ in neem formulations and neem oil. Limits of quantification and detection were respectively 6 μg/g and 3 μg/g.

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

AbstractThe 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.

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’.

Endocrine manipulations and ontogenesis of male sexual behaviour in Locusta: studies on azadirachtin‐induced over‐aged nymphs

ABSTRACT. Dose‐response relations of the moult‐inhibitory and lethal effects of azadirachtin (AZA) injected into fifth and fourth instar hoppers of Locusta migratoria migratorioides are presented. The optimal dose for obtaining maximum number of long‐surviving over‐aged nymphs is 1.6 and 1.0 μg of AZA per insect for young fifth and fourth instar nymphs, respectively. Over‐aged male nymphs are capable of exhibiting sexual behaviour. The mating behaviour of fifth instar over‐aged male nymphs, and of chemically allatectomized but otherwise similar over‐aged male nymphs (obtained by combined precocene treatment and injection of AZA), with and without injections of exogenous Juvenile Hormone III (JH), is investigated quantitatively; the results are compared with those obtained for normal, chemically allatectomized, and chemically allatectomized plus JH‐injected adult males. Over‐aged fifth instar male nymphs show a low intensity of mating behaviour and in chemically allatectomized ones this intensity is even lower. Injections of JH intensify the mating behaviour of chemically allatectomized fifth instar over‐aged male nymphs in a dose‐dependent way, but less markedly so than in chemically allatectomized adult males. We conclude that the intensity of male mating behaviour of the over‐aged nymphs is subnormal and their response to the mating behaviour intensifying effect of the JH is weaker than that of adult males. The relations between endocrine factors and ethological ontogenesis of male sexual behaviour in locusts are discussed.

Endocrine manipulations, juvenile hormone and ontogenesis of male sexual behaviour in locusts

By injection of azadirachtin (AZA) to 5th instar hoppers of Locusta migratoria migratorioides, “over-aged” nymphs were produced, some of which survived for long periods without moulting to adults. Such over-aged male nymphs exhibited mating behaviour belatedly, though at a lower intensity than normal adults. Juvenile hormone (JH) markedly intensifies male sexual behaviour in crowded adult (6th instar) Locusta; thus, the lower intensity of sexual behaviour of the over-aged nymphs might have been caused by a low endogenous JH titre, or by a weaker response to the hormone. To clarify this point, chemically allatectomized over-aged 5th instar nymphs were produced by combined treatment with precocene and AZA. These exhibited a very low intensity of mating behaviour. Injections of exogenous JH III intensified the mating behaviour of the chemically allatectomized over-aged 5th instar male nymphs in a dose-dependent way, but the effect was weaker than in chemically allatectomized adult males. AZA-induced over-aged 4th instar male Locusta hoppers also exhibited mating behaviour. By injection of AZA, 5th instar over-aged hoppers of Schistocerca gregaria were also produced. Males showed mating behaviour belatedly, despite the fact that JH completely controls male sexual behaviour in this species. The intensity of this behaviour was again lower than in adults. Interrelations between integumental morphogenesis, “ethogenesis” of male sexual behaviour, and endocrine factors in locusts are discussed.