The relationship between the chemical composition of three essential oils and their insecticidal activity against Acanthoscelides obtectus (Say)

The chemical composition of the essential oils isolated from various parts of three Greek aromatic plants (Lavandula hybrida Rev, Rosmarinus officinalis L and Eucalyptus globulus Labill) collected at different seasons was determined by GC/MS analysis. The insecticidal action of these oils and of their main constituents on Acanthoscelides obtectus (Say) adults was evaluated and their LC50 values were estimated. All essential oils tested exhibited strong activity against A. obtectus adults, with varying LC50 values depending on insect sex and the composition of the essential oils. A correlation between total oxygenated monoterpenoid content and activity was observed, with oxygenated compounds exhibiting higher activity than hydrocarbons. Among the main constituents, only linalyl and terpinyl acetate were not active against A. obtectus, while all the others exhibited insecticidal activity against both male and female adults, with LC50 values ranging from 0.8 to 47.1 mg litre(-1) air. An attempt to correlate the insecticidal activity to the monoterpenoid's structure is presented, and the difference in sensitivity between male and female individuals is also explored.     

Characterisation of midgut digestive proteases from the maize stem borer Busseola fusca

BACKGROUND: Insect damage is a major constraint on maize production. Control of Busseola fusca (Fuller) in sub‐Saharan Africa is relatively ineffective; the major larval digestive enzymes were characterised with a view to developing future control strategies. RESULTS: Using BODIPY‐FL Casein, maximal activity was at pH 9.5, with six protease forms visualised by gelatin‐PAGE. Synthetic substrates and diagnostic inhibitors demonstrated the presence of serine proteases. Chymostatin was a potent inhibitor of general proteolysis (90%), providing strong evidence for the presence of chymotrypsin; it also caused significant inhibition (>95%) with SA₂PFpNA as substrate. The I₅₀ values for chymostatin with casein and SA₂PFpNA were 0.0075 µM and 0.06 µM respectively. Z‐Phe‐Arg‐pNA activity was inhibited by chymostatin and TLCK (50 and 30% respectively), suggesting the presence of trypsin‐like activity. BApNA hydrolysis was also strongly inhibited by chymostatin and TLCK (92 and 75%), suggesting trypsin activity, while SBBI, PMSF, pepstatin and E‐64 had no significant effect. Interestingly, SBBI (I₅₀ = 0.39 µM ) and SBTI both inhibited general proteolysis by approximately 70%, suggesting that SBBI's dual inhibitory role makes this inhibitor a potentially useful candidate for expression in maize for control of B. fusca. CONCLUSION: These results provide a basis for the rational design of insect‐resistant transgenic maize expressing protease inhibitors. Copyright © 2008 Society of Chemical Industry     

The effects of phytochemical pesticides on the growth of cultured invertebrate and vertebrate cells

A range of cultured cells of invertebrate and vertebrate origin was grown in the presence of a number of phytochemical pesticides to test the effect of the latter on cell proliferation. The main observation was that azadirachtin was a potent inhibitor of insect cell replication, with an EC50 of 1.5 x 10(10) M against Spodoptera cells and of 6.3 x 10(9) M against Aedes albopictus cells, whilst affecting mammalian cells only at high concentrations (> 10(-4) M). As expected, the other phytochemical pesticides, except for rotenone, had little effect on the growth of the cultured cells. Rotenone was highly effective in inhibiting the growth of insect cells (EC50:10(-8) M) but slightly less toxic towards mammalian cells (EC50:2 x 10(-7) M). Neem terpenoids other than azadirachtin and those very similar in structure significantly inhibited growth of the cell cultures, but to a lesser degree. The major neem seed terpenoids, nimbin and salannin, for example, inhibited insect cell growth by 23% and 15%, respectively.     

Assessment of the suitability of Tinopal as an enhancing adjuvant in formulations of the insect pathogenic fungus Beauveria bassiana (Bals.) Vuillemin

BACKGROUND: Biopesticides based on Beauveria bassiana (Bals.) Vuillemin hold great promise for the management of a wide range of insect pests. The conidia in the biopesticide formulation require an adjuvant to protect them from photoinactivation by sunlight. The suitability of Tinopal, an optical brightener used as sunscreen for baculovirus formulations, for use with B. bassiana was assessed. The aim was to study the effect of Tinopal on the growth and photoprotection of B. bassiana, and its effect on the susceptibility of insects to B. bassiana. RESULTS: Tinopal was found to have no adverse effect on the growth of B. bassiana. It was found to confer total protection (approximately 95% conidial germination at 10 g Tinopal L(-1)) from sunlight up to 3 h of exposure, and a better survival rate than controls even up to 4 h. Helicoverpa armigera Hübner larvae fed on diet with 5 g kg(-1) Tinopal were found to have reduced growth. The duration of the larval stage increased by 3-4 days in 1 and 5 g kg(-1) Tinopal treatments. Among the moths that emerged from larvae fed on diet with 5 g kg(-1) Tinopal, a significantly high number were malformed compared with controls. The larvae that were fed diet with Tinopal showed quicker and higher mortality and required a lower effective lethal dose (LC(50)) than the controls. Tinopal was found to have a synergistic effect with B. bassiana in causing insect mortality. CONCLUSIONS: Tinopal was found to be a suitable adjuvant for B. bassiana-based biopesticide formulations. It conferred tolerance to sunlight and caused stress in the insect, leading to a synergistic effect with B. bassiana.     

Proteolytic enzymes secreted by larval stage of the parasitic nematode Trichinella spiralis

Excretory/secretory products (ES), collected from in vitro cultures of muscle larvae (L1) of Trichinella spiralis (Owen, 1835) were examined for the presence of proteolytic enzymes. Several discrete proteinases in the size range of 25-55 kDa were identified by substrate gel electrophoresis and were characterised according to pH optima, substrate specificity and inhibitor sensitivity using azocasein assay. Serine, cysteine and metalloproteinases active at pH 5-7 were identified. The serine proteinases were found to predominate and some of them were found to be specific for the larval stage of the parasite. The results from the substrate analysis indicated the presence of collagenolytic and elastolytic activities. The proteinase activity was inhibited by IgG isolated from T. spiralis-infected mice, an observation of relevance to understanding host/parasite interactions and, ultimately, the development of anti-Trichinella vaccine.     

Cell surfaces in plant micro-organism interactions. VIII. Increased proteinase inhibitor activity in melon plants in response to infection by Colletotrichum lagenarium or to treatment with an elicitor fraction from this fungus

Proteinase inhibitor activity had increased sharply in melon seedlings infected by Colletotrichum lagenarium 3 days after inoculation. The activity was associated with heat stable proteins and was effective against the protease produced by the fungus as well as against trypsin. Treatment of healthy melon leaves with an elicitor of ethylene isolated from the pathogen, resulted in a three-fold increase in proteinase inhibitor activity after 24 h. Ethylene production increased early in elicitor-treated leaves and may be involved in the elicitation of proteinase inhibitor activity. In the presence of aminoethoxyvinylglycine, an inhibitor of ethylene biosynthesis, both elicitor-induced ethylene and, to a lesser extent, elicitor-induced proteinase inhibitor activity were inhibited. In contrast, 1-aminocyclopropane-1-carboxylic acid, the direct precursor of ethylene, triggers proteinase inhibitor activity. It is concluded that ethylene is involved in the elicitation of proteinase inhibitor activity, but its exact role remains to be defined.     

Testing of an integrated regime for effective and sustainable control of invasive Crofton weed (Ageratina adenophora) comprising the use of natural inhibitor species,activated charcoal,and fungicide

Crofton weed is a major invasive species in China. It exhibits superior growth characteristics and can outcompete with native species via allolepathic effects and modulation of the soil fungal microbiome. The simple removal of invading plants will not ensure restoration of the habitat due to the persistence of allelochemicals and viable seeds in the surrounding soil. An orthogonal experimental design was employed to evaluate the effects of three control factors (A, powdered natural inhibitor species to retard growth; B, activated charcoal to absorb allelochemicals; and C, fungicide to reduce fungal modulation effects), applied at three levels, on the growth and competitive ability of Crofton weed against two native species, in a pot‐culture experiment. All treatments reduced all measured growth parameters (P < 0.05) except for a specific leaf area, when compared with control plants. Furthermore, the competitive capacity of Crofton weed was decreased in the treatments while that of the native species was improved. Application to soil of the powdered natural inhibitor species and of activated charcoal significantly inhibited plant growth and competitive ability of Crofton weed (P < 0.05). Application of fungicide was less effective, but significantly reduced the specific leaf area of Crofton weed plants (P < 0.05). The specific combination of factors producing the greatest decrease in plant growth and competitive ability (compared with the control) included the addition of Delavaya toxocarpa powder (37.5 g per kg soil), addition of activated charcoal to soil at a ratio of 1:3 (v/v) (62.5 g per kg soil), and application of fungicide (Thiophanate‐Methyl) (0.28 g per kg soil).     

Enhancement of fungicidal and insecticidal activity by reductive alkylation of chitosan

A series of N-alkyl chitosan (NAC) derivatives were synthesized using a reductive alkylation reaction to examine their fungicidal and insecticidal activity. The chemical structures were characterized by IR and (1)H NMR spectroscopy, and the degree of substitution (DS) ranged from 0.02 to 0.37. Their fungicidal activity was evaluated against the grey mould Botrytis cinerea Pers ex Fr (Leotiales: Sclerotiniaceae) and the rice leaf blast pathogen Pyricularia grisea Sacc [Teleomorph: Magnaporthe grisea (Hebert) Barr] by a radial growth bioassay. It was of interest that most of the NAC derivatives were more active against both fungi than chitosan itself. The most active derivative was N-(2,2-diphenylethyl)chitosan with EC50 values of 0.031 and 0.23 g L(-1) against B. cinerea and P. grisea respectively. In addition, some derivatives, at higher concentrations up to 1.0 g L(-1), inhibited the mycelial growth and spore formation of P. grisea. Bioassays against larvae of the cotton leafworm, Spodoptera littoralis (Boisd.) (Lepidoptera: Noctuidae) with the NAC derivatives at a rate of 5.0 g kg(-1) in artificial diet demonstrated that N-(3-phenylbutyl)chitosan was the most active compound. In addition, N-propylchitosan, N-undecanylchitosan and N-(3-phenylpropyl)chitosan derivatives strongly inhibited larval weight gain in S. littoralis, with respective reductions of 76, 66 and 65% after 4 days of feeding on treated diet.     

Interaction of Cry1Ac toxin (Bacillus thuringiensis) and proteinase inhibitors on the growth, development, and midgut proteinase activities of the bollworm, Helicoverpa zea

Potential resistance development to Bt cotton in certain lepidopterans has prompted research to develop strategies that will preserve this environmental-friendly biotechnology. Proteinase inhibitors are potential candidates for enhancing Bt toxicity against lepidopteran pests and for expanding the spectrum of control for other insects. Interactions of Bt toxin from Bacillus thuringiensis and proteinase inhibitors were investigated by monitoring growth, development, and gut proteinase activities of the bollworm, Helicoverpa zea. Several proteinase inhibitors were combined with Bt protoxin Cry1Ac in artificial diet and fed to newly molted 3rd-instar bollworm larvae to determine effects on larval body weight and length, pupation progress, and mortality rate. Major midgut proteinase activities, including caseinase, tryptic, and chymotrypsin activities, were examined after treatment. A concentration of Bt at a level causing minimal mortality (<10%), was mixed with the following proteinase inhibitors: benzamidine, phenylmethylsulfonyl fluoride (PMSF), and N-α-tosyl-l-lysine chloromethyl ketone (TLCK). When compared with controls, the synergistic effect of Bt toxin and proteinase inhibitors caused significant decreases in mean larval weight and length over time. Midgut samples tested against the substrates azocasein, α-benzoyl-dl-arginine-p-nitroanilide (BApNA), and N-succinyl-alanine-alanine-proline-phenylalanine-p-nitroanilide (SAAPFpNA) showed significant decreases in the protease activity of larvae fed Bt plus inhibitor versus control. Interaction of Bt and proteinase inhibitors significantly retarded larval growth and resulted in developmental delay and up to 20% mortality.     

Interaction of proteinase inhibitors with Cry1Ac toxicity and the presence of 15 chymotrypsin cDNAs in the midgut of the tobacco budworm, Heliothis virescens (F.) (Lepidoptera: Noctuidae)

BACKGROUND: The potential development of resistance to Bacillus thuringiensis (Bt) cotton and surging of non‐targeted insects is a major risk in the durability of Bt plant technology. Midgut proteinases are involved in Bt activation and degradation. Proteinase inhibitors may be used to control a wide range of insects and delay Bt resistance development. Proactive action to examine proteinase inhibitors for synergistic interaction with Bt toxin and cloning of proteinase cDNAs for RNAi is necessary to make transgenic cotton more versatile and durable. RESULTS: A sublethal dose (15 ppb) of Cry1Ac, 0.5% benzamidine and 0.02% phenylmethylsulfonyl fluoride significantly suppressed midgut azocaseinase, tryptic and chymotryptic activities, and resulted in reductions in larval and pupal length and mass of Heliothis virescens. The combination of proteinase inhibitor and Bt suppressed 20–37% more larval body mass and 26–80% more enzymatic activities than the inhibitor only or Bt only. To facilitate knockdown‐resistance‐related proteinase genes, 15 midgut chymotrypsin cDNAs were sequenced. Most predicted chymotrypsins contained the conserved N‐termini IVGG, three catalytic center residues (His, Asp and Ser), substrate specificity determinant (Ser or Gly) and cysteines for disulfide bridges. These putative chymotrypsins were separated into three distinct groups, indicating the diverse proteinases evolved in this polyphagous insect. CONCLUSION: H. virescens has evolved diverse midgut proteinase genes. Proteinase inhibitors have potential insecticidal activity, and the interaction of Bt with proteinase inhibitors is desirable for enhancing Bt toxicity and delaying resistance development. Intensive sequencing of chymotrypsin cDNAs will facilitate future functional examinations of individual roles in Bt toxicity and resistance development and facilitate targeted control using RNAi and/or proteinase inhibitors. Copyright © 2012 Society of Chemical Industry     

Production of a Mating Inhibitor by Ustilago hordei

ABSTRACT Ustilago hordei, the cause of barley covered smut, was found to produce a factor that inhibited its own mating. The mating inhibition factor (MIF) specifically inhibited mating of U. hordei and other Ustilago spp., but not teliospore germination or sporidial growth. MIF did prevent teliospore germination of Tilletia caries and T. contraversa. MIF was found at low levels in culture supernatants of either mating type of U. hordei grown separately, but at higher levels when both mating types were grown together, in the supernatants of MAT-1 mating type cells transformed with the MAT-1 pheromone gene mfa1 and of MAT-2 cells transformed with either mfa1 or the MAT-1 pheromone receptor gene pra1. Diploid cells produced no detectable inhibitor, nor did MAT-1 cells with a disrupted mating type locus that deleted both mfa1 and pra1. MIF production was restored when mfa1, but not pra1, was added back to the MAT-1Delta cells. MIF activity was altered by protease treatment. Highly purified MIF from MAT-1 cells contained cysteine methyl ester, farnesyl cysteine, farnesyl cysteine methyl ester, and a dodecapeptide with a mass consistent with that of MAT-1 pheromone lacking the terminal cysteine. Since smut fungi must first mate to become pathogenic, mating inhibition has the potential to be an effective method of disease control for these pathogens.     

伯氏嗜线虫致病杆菌蛋白酶抑制剂Xbpi-1对豌豆蚜Acyrthosiphon pisum Harris生物活性的研究

蛋白酶抑制剂是一类能够抑制蛋白水解酶活性的小分子蛋白质,在昆虫肠道内通过抑制蛋白酶的活性,可以干扰昆虫的正常生长和发育。本文利用体外重组表达的蛋白酶抑制剂Xbpi-1,研究了Xbpi-1对豌豆蚜Acyrthosiphon pisum Harris的杀虫活性及其生化机制。结果表明,取食含蛋白酶抑制剂Xbpi-1浓度为50、200、400、800μg·mL^-1人工饲料的豌豆蚜,96h后的校正死亡率分别为2．1％、9-4％、19．7％和50．9％,虫重分别减少12_3％、20．3％、25．7％和33．7％。生化测定结果表明,4种浓度处理的豌豆蚜体内总蛋白酶、氨肽酶、胰蛋白酶、胰凝乳蛋白酶的活性均有显著降低,浓度为800μ·mL^-1时,其酶活性分别降低了29．6％、26．1％、23．5％和23．8％。本研究结果说明,蛋白酶抑制剂Xbpi-1对豌豆蚜具有较好的生物活性,为进一步开发应用于豌豆蚜等刺吸式害虫的防治提供了理论依据。     

Laboratory and field evaluations of rhodojaponin-III against the imported cabbage worm Pieris rapae (L.) (Lepidoptera: Pieridae)

The activity of rhodojaponin-III (R-III), a grayanoid diterpene compound isolated from Rhododendron molle G. Don flowers, was determined under laboratory and field conditions as an antifeedant, stomach poison, contact toxicant and insect growth inhibitor against Pieris rapae (L.) larvae. The median antifeedant concentration (AFC(50)) values in no-choice leaf disc tests were 1.16 and 15.85 mg L(-1) at 24 h after treatment when tested against third and fifth instars respectively. The median lethal concentration (LC(50)) values in leaf disc tests were 2.84 and 9.53 mg L(-1) at 96 h after treatment against third and fifth instars respectively. R-III showed an almost 30 times higher contact toxicity against third instars than for fifth instars, and the median lethal dose (LD(50)) values for topical application were 1.18 and 34.09 mg kg(-1) at 72 h after treatment respectively. R-III disrupted the development of larvae to pupae or adults with median concentration for inhibiting growth (IC(50)) values of only 1.36 mg L(-1) for third instars and 11.28 mg L(-1) for fifth instars. In field trials, a greater than 80% reduction in the adjusted larval numbers was obtained against P. rapae 14 days after treatment when Rhodo 0.1% EC, a commercial botanical insecticide based on R-III, was applied at both 937.5 and 625 mL ha(-1). These results suggest that further research to develop R-III, and extracts from R. molle, as biorational pesticides or as lead compounds for integrated pest management deserve consideration.     

Persistence and residual activity of an organophosphate, pirimiphos-methyl, and three IGRs, hexaflumuron, teflubenzuron and pyriproxyfen, against the cowpea weevil, Callosobruchus maculatus (Coleoptera: Bruchidae)

Three insect growth regulators (IGR), the chitin synthesis inhibitors (CSI) teflubenzuron and hexaflumuron and the juvenile hormone mimic (JHM) pyriproxyfen, as well as the organophosphate (OP) pirimiphos-methyl, were evaluated for their activity against the cowpea weevil, Callosobruchus maculatus (F), in cowpea seeds stored for up to 8 months post-treatment. The initial activity data showed that, based on LC50 level, teflubenzuron had strong ovicidal activity (LC50 = 0.056 mg kg(-1)) followed by pirimiphos-methyl (1.82 mg kg(-1)) and pyriproxyfen (91.9 mg kg(-1)). The residual activity data showed that none of the IGRs tested had strong activity when applied at 200 mg kg(-1) in reducing the oviposition rates of C maculatus at various storage intervals up to 8 months post-treatment. However, teflubenzuron reduced adult emergence (F1 progeny), achieving control ranging from 96.2% at 1 month to 94.3% at 8 months. Hexaflumuron showed a similar trend in its residual activity, ranging between 93.8% control at 1 month to 88.2% control at 8 months post-treatment. However, pyriproxyfen was more active than the CSIs tested and caused complete suppression (100% control) of adult emergence at all storage intervals. Unlike the IGRs tested, pirimiphos-methyl applied at 25 mg kg(-1) was more effective in reducing oviposition rates of C maculatus up to 8 months post-treatment. A strong reduction of adult emergence was also observed at various bimonthly intervals (98.6% control at 1 month to 91.6% control at 8 months post-treatment). The persistence of hexaflumuron and pirimiphos-methyl in cowpea seeds was also studied over a period of 8 months. The loss of hexaflumuron residue in treated cowpeas (200 mg kg(-1)) was very slow during the first month post-treatment (4.43%). At the end of 8 months, the residue level had declined significantly to 46.4% of the initial applied rate. The loss of pirimiphos-methyl residue in treated cowpeas (25 mg kg(-1)) was relatively high during the first month post-treatment (36.7%) and increased to 81.6% after 8 months.     

Characterization of a Hydrophobic Amylase Inhibitor from Corn (Zea mays) Seeds with Activity Against Amylase from Fusarium verticillioides

ABSTRACT A hydrophobic 19.7-kDa amylase inhibitor (AI) was purified from corn kernels by 95% ethanol extraction and anionic exchange chromatography. The AI has an isoelectric point of 3.6 and was very stable at different pH values and high temperatures, maintaining 47.6% activity after heating to 94 degrees C for 60 min. Amino acid analysis indicated high valine, leucine, glycine, alanine, and glutamic acid/glutamine content, and especially high valine content (41.2 mol%). This inhibitor is not a glycoprotein. It required 30-min preincubation to maximize complex enzyme-inhibitor formation when the amylase from Fusarium verticillioides was tested. The optimal pH of interaction was 6.5. It showed broad-spectrum activity including the following amylases: human saliva, porcine pancreas, F. verticillioides, as well as those from some insects of agricultural importance (Acanthoscelides obtectus, Zabrotes subfasciatus, Sitophilus zeamais, and Prostephanus truncatus). This novel hydrophobic protein not only inhibited the amylase from F. verticillioides but also decreased the conidia germination. Thus, this protein represents an approach to decrease the production of fumonisin in corn, either by using it as a molecular marker to detect fungal resistance or through genetic engineering.     

安全剂AD-67保护玉米免受单嘧磺隆药害的作用及其机理

采用生物测定法研究了安全剂AD-67对除草剂单嘧磺隆造成玉米伤害的保护作用,并对其作用机理进行了初步探讨。结果表明:单嘧磺隆在浓度0.025～0.050mg/kg时,对玉米的生长有较为强烈的抑制作用;当AD-67的浓度在0.5～10mg/kg时,能不同程度地缓解单嘧磺隆对玉米的药害。当单嘧磺隆毒土浓度为0.025mg/kg,使用1mg/kgAD-67浸种处理,玉米的主根长和株高分别是对照的107.3%和100.82%;AD-67浸种处理的玉米,幼苗中谷胱甘肽(GSH)含量和乙酰乳酸合成酶(ALS)活性分别是对照的109.76%和113.33%。     

Effect of chitin-synthesis inhibitors on stored-product beetles

The fecundity of four important coleopteran stored-grain pests, Rhyzopertha dominica (Fab.), Sitophilus oryzae (L.), Oryzaephilus surinamensis (L.) and Tribolium castaneum (Herbst.) was assessed after exposure to wheat treated with five benzoylphenyl ureas (BPUs), members of the chitin-synthesis inhibitor group of insect growth regulators (IGRs). Chlorfluazuron was found to control all four species more effectively than triflumuron, teflubenzuron, flufenoxuron or diflubenzuron. Two weeks of exposure of parents to 0.5 mg kg ?1 of all the BPUs, except diflubenzuron, inhibited almost all F1 production and all the F2 production of each species. The fecundities of T. castaneum and O. surinamensis recovered almost to their untreated levels after two weeks of exposure to 1 mg kg ?1 of BPUs followed by two weeks on untreated wheat, but the effect of the exposure persisted for at least two weeks on untreated wheat in adult S. oryzae and to a lesser extent in R. dominica. The mean F1 response of the four species after two weeks of exposure to four doses showed clearly that the four newer BPUs were all similar in their overall effectiveness and significantly more effective than diflubenzuron, although diflubenzuron was equally effective against S. oryzae. At 0.1 mg kg ?1 triflumuron was less effective against O. surinamensis, while teflubenzuron and flufenoxuron were not very effective against the rice weevil, S. oryzae. Chlorfluazuron provided the most effective control of all four stored-product pest species, including the weevil, S. oryzae. The BPUs show particular potential for use in the stored-product industry because low dosages will effectively control the rice weevil in addition to the other major pest species.     

蛋白酶抑制剂在害虫防治中的研究与应用

蛋白酶抑制剂是一种抑制蛋白酶分解活性的蛋白质,对许多昆虫的生长发育有明显的抑制作用。蛋白酶抑制剂扰乱昆虫正常代谢,最终导致昆虫发育不正常甚至死亡,在害虫防治中有良好的应用前景。本文分别对蛋白酶抑制剂种类、分布及抗虫性进行了介绍,同时探讨了蛋白酶抑制剂的基因转化及应用前景。     

Effects on litter-dwelling earthworms and microbial decomposition of soil-applied imidacloprid for control of wood-boring insects

BACKGROUND: Imidacloprid is an effective, systemic insecticide for the control of wood-boring insect pests in trees. Systemic applications to trees are often made by soil injections or drenches, and the resulting imidacloprid concentrations in soil or litter may pose a risk of harm to natural decomposer organisms. The authors tested effects of imidacloprid on survival and weight gain or loss of the earthworms Eisenia fetida (Savigny) and Dendrobaena octaedra (Savigny), on leaf consumption rates and cocoon production by D. octaedra and on microbial decomposition activity in laboratory microcosms containing natural forest litter. RESULTS: Dendrobaena octaedra was the most sensitive of the two earthworm species, with an LC(50) of 5.7 mg kg(-1), an LC(10) of about 2 mg kg(-1) and significant weight losses among survivors at 3 mg kg(-1). Weight losses resulted from a physiological effect rather than from feeding inhibition. There were no effects on cocoon production among survivors at 3 mg kg(-1). The LC(50) for E. fetida was 25 mg kg(-1), with significant weight losses at 14 mg kg(-1). There were no significant effects on microbial decomposition of leaf material at the maximum test concentration of 1400 mg kg(-1).CONCLUSION: The results indicate that, when imidacloprid is applied as a systemic insecticide to the soil around trees, it is likely to cause adverse effects on litter-dwelling earthworms if concentrations in the litter reach or exceed about 3 mg kg(-1).     

Corn Seed Proteins Inhibitory to Aspergillus flavus and Aflatoxin Biosynthesis

ABSTRACT This study reports the presence of two fractions from corn seeds inhibitory to aflatoxin formation. Using a sensitive laboratory assay that can measure both inhibition of fungal growth and inhibition of aflatoxin biosynthesis, we examined aqueous extracts from seeds of Tex6, a corn inbred shown to be highly resistant to aflatoxin accumulation in field and laboratory evaluations. In these extracts, we identified two biologically active fractions. One inhibited growth of Aspergillus flavus and, thus, aflatoxin accumulation, and the other inhibited aflatoxin formation with little effect on fungal growth. The compounds responsible for these activities appear to be proteaceous, as they are water soluble, heat labile, and sensitive to proteinase K treatment. The compounds were partially purified by ultrafiltration and chromatography. The estimated molecular mass of the growth inhibitor is approximately 28 kDa, and that of the aflatoxin biosynthesis inhibitor appears to be greater than 100 kDa. Partially purified preparations of the growth inhibitor and aflatoxin biosynthesis inhibitor cause 50% inhibition at 26 and 75 mug of protein/ml, respectively. The presence of these compounds in Tex6 may explain its resistance to aflatoxin accumulation.