Screening of insecticidal activity of plant essential oils and extract-based formulations against four agricultural insect pests and their risk assessment

Frankliniella occidentalis (Pergande), Myzus persicae (Sulzer), Plutella xylostella (L), and Spodoptera exigua (Hübner) are notorious agricultural insect pests worldwide. Due to the rapid development of resistance to the conventional insecticides, the number of available insecticides might be reduced in a near future, and botanical insecticides can be considered as one of the potential candidates to solve this problem. In the present study, test formulations were prepared using plant essential oils and extracts as main active ingredients, and their insecticidal activity and phytotoxicity were examined. Among the combinations tested, two oil-in-water formulations of NRS-13 containing thyme white oil and NRS-24 containing basil and rosemary oils were selected based on their notable insecticidal activity and low phytotoxicity. Chemical analyses on the active essential oils, the quantitative analysis of the major components in the formulations, insecticidal activities under pot and field conditions, their acute toxicity on mammals, and ecotoxicity on fish were also examined. The NRS-13 (BaechooSafer^TM) and NRS-24 (JindiOut^TM) formulations showed species-specific insecticidal activity against P. xylostella and M. persicae, respectively. In addition, both formulations produced low acute oral and dermal toxicity, skin and eye irritation, and fish toxicities, which suggests those formulations based on essential oils could be considered as insecticides for pest control in small-scale environments in organic farming.     

Comparative bioassay of different isolates of Bacillus thuringiensis subsp. kurstaki on the third larval instars of diamondback moth,Plutella xylostella (L.) (Lep.: Plutellidae)

The diamondback moth, Plutella xylostella (L.) (Lepidoptera: Plutellidae) is one of the most important pests of cruciferous plants throughout the world. In recent years, this insect has been a serious pest for cabbage fields in Tehran province. Resistance of P. xylostella to all main groups of insecticides has been recorded and it is ranked in the 20 most resistant pest species reported up to now. According to many researchers, to eliminate the problem of pest resistance to chemical pesticides, an integrated pest management programme should be used. In line with this, the uses of microbial control agents (MCAs) are discussed. The bacterium, Bacillus thuringiensis (Bt) is one of microbial control agents of pests. It is characterised by its ability to produce proteic crystalline inclusions during sporulation. Cry1 protein has insecticidal activity and is highly specific to certain insects and not toxic to unrelated insects, plants or vertebrates. In this work, the pathogenicity of some Bt isolates, including Dipel, 20, 29, 79 and 87, was tested against P. xylostella and the lethal concentrations (LC₅₀) of their crystal proteins to P. xylostella third larval instar was determined. The experiment was designed in factorial in randomised complete design with 5 treatments (different concentrations including 10⁴, 10⁵, 10⁶, 10⁷, 10⁸ CFU/ml and 5 replications and with 10 third larval instars. Spore–crystal complex was applied to the surface of natural diets (cabbage leaves) and the mortality of P. xylostella larvae was assessed 120?h after exposure of Bt toxin in each treatment. Results showed that percentage of survival was significantly higher for control treatment. Results also showed that after 5?days, LC₅₀ for isolates of Dipel, 20, 29, 79 and 87 were equal to 1?×?10⁶, 1?×?10⁵, 5?×?10⁵, 4?×?10⁵ and 1?×?10⁴ CFU/ml, respectively. LT₅₀ were equal to 93.71, 48.04, 71, 40.49 and 75.28?h. Of and most the percentage larval mortality relate to attendance 87 and also at least percentage mortality is related to the groom Dipel.     

Plant-mediated vulnerability of an insect herbivore to Bacillus thuringiensis in a plant-herbivore-pathogen system

Laboratory studies were performed to explore the effects of host-plant quality on the vulnerability of Plutella xylostella to Bacillus thuringiensis. P. xylostella were kept on different host plants, including Brassica pekinensis (Chinese cabbage) cv. Hero, Brassica oleracea var. botrytis (cauliflower) cv. Royal, and B. oleracea var. capitata (common cabbage) cv. Globe Master (white cabbage) and cv. Red Dynasty (red cabbage) for at least two generations. These host plants are considered as the high (Chinese cabbage), intermediate (cauliflower and white cabbage) and low-quality (red cabbage) hosts for P. xylostella. The vulnerability of the pest larvae was then tested using two formulation of B. thuringiensis var. kurstaki, including Biolarv^® and Biolep^®. The results demonstrated that the susceptibility of P. xylostella to B. thuringiensis was influenced by host-plant quality. Indeed, B. thuringiensis acted better on the pest fed on the low-quality host plant compared with that on the high-quality host plant. The interaction between the pathogen and plant quality/resistance resulted in more mortality of the pest larvae, implying a synergistic effect. From a pest management viewpoint, these findings may be promising for the integration of the pathogen and the low-quality/partially resistant host plants against P. xylostella in field studies.     

Juvenile hormone antagonistic activity of secondary metabolites from Streptomyces lactacystinicus and their insecticidal activity against Plutella xylostella

Due to their specificity to target insects and low toxicity to non-target organisms, insect growth regulators (IGRs) have been promising alternatives to neurotoxic insecticides. Actinobacteria produce a wide range of secondary metabolites with insecticidal and insect growth regulatory activities. In this study, the culture media of 25 actinobacteria isolates showing high juvenile hormone antagonist (JHAN) activity were assessed for their insecticidal activity to identify novel IGR compounds toxic to Plutella xylostella. Among them, four isolates exhibited high insecticidal activity against 3rd instar larvae of P. xylostella. Two isolates of IMBL-1412 and IMBL-1823 showing relatively high insecticidal activities (greater than90% mortality) were identified as Streptomyces lactacystinicus based on colony color on various International Streptomyces Project (ISP) media and nucleotide sequences of the 16S rRNA gene. The ethyl acetate fractions of both isolates showed high JHAN and insecticidal activities against P. xylostella larvae at a concentration of 100 ppm when the culture media of these two isolates were extracted sequentially using hexane, ethyl acetate, and butyl alcohol. These results suggested that secondary metabolites of these actinobacterial isolates could be efficiently applied to develop novel IGR insecticides for the control of P. xylostella.     

Effects of nitrogen and foliar biomass on population parameters of cabbage insects

The effects of different nitrogen (N) fertilization rates (0, 45, 90, and 168 kg N/ha), plant nitrogen concentration, and plant biomass on abundance and population growth of diamondback moth, Plutella xylostella (L.), cabbage looper, Trichoplusia ni (Hübner), cabbage budworm, Hellula phidilealis (Walker), imported cabbageworm, Artogeia rapae (L.), and cross-striped cabbageworm, Evergestis rimosalis (Guenée), were investigated in Homestead and Sanford, Florida in 1987. The effects of these factors on the parasitization of P. xylostella were also examined. In Homestead, abundance of most insect pests and parasitized P. xylostella increased with an increase in the level of N applied and with an increase in plant biomass. Similar results were found in Sanford, although results were not consistently significant. Abundance of most insect pests was significantly positively correlated with plant N concentration. Multiple regression analyses indicated that foliar biomass was significantly more important than N fertilization rate and subsequent plant N concentration at predicting abundance of insect pests and parasitized P. xylostella on cabbage.     

Design,synthesis and insecticidal evaluation of aryloxy dihalopropene derivatives

Plutella xylostella (P. xylostella) is a highly migratory, cosmopolitan species and one of the most important pest of cruciferous crops worldwide. Pyridalyl as a novel class of insecticides has good efficacy against P. xylostella. On the basis of the commercial insecticide pyridalyl, a series of new aryloxy dihalopropene derivatives were designed and synthesized by using Intermediate Derivatization Methods. Their chemical structures were confirmed by ¹H NMR, high-resolution mass spectrum (HRMS), and single-crystal X-ray diffraction analysis. The insecticidal activities of the new compounds against P. xylostella were evaluated. The results of bioassays indicated that most of the compounds showed moderate to high activities at the tested concentration, especially compounds 10e and 10g displayed more than 75% insecticidal activity against P. xylostella at 6.25 mg/L, while pyridalyl showed 50% insecticidal activity at the same concentration. The field trials result of the insecticidal activities showed that compound 10e as a 10% emulsifiable concentrate (EC) was effective in the control of P. xylostella at 75–150 g a.i./ha, and the mortality of P. xylostella for treatment with compound 10e at 75 g a.i./ha was equivalent to pyridalyl at 105 g a.i./ha.     

Insecticidal Activity of Bacillus thuringiensis Cry1Bh1 against Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae) and Other Lepidopteran Pests

Bacillus thuringiensis is an important source of insect resistance traits in commercial crops. In an effort to prolong B. thuringiensis trait durability, insect resistance management programs often include combinations of insecticidal proteins that are not cross resistant or have demonstrable differences in their site of action as a means to mitigate the development of resistant insect populations. In this report, we describe the activity spectrum of a novel B. thuringiensis Cry protein, Cry1Bh1, against several lepidopteran pests, including laboratory-selected B. thuringiensis-resistant strains of Ostrinia nubilalis and Heliothis virescens and progeny of field-evolved B. thuringiensis-resistant strains of Plutella xylostella and Spodoptera frugiperda. Cry1Bh1 is active against susceptible and B. thuringiensis-resistant colonies of O. nubilalis, P. xylostella, and H. virescens in laboratory diet-based assays, implying a lack of cross-resistance in these insects. However, Cry1Bh1 is not active against susceptible or Cry1F-resistant S. frugiperda. Further, Cry1Bh1 does not compete with Cry1Fa or Cry1Ab for O. nubilalis midgut brush border membrane binding sites. Cry1Bh1-expressing corn, while not completely resistant to insect damage, provided significantly better leaf protection against Cry1Fa-resistant O. nubilalis than did Cry1Fa-expressing hybrid corn. The lack of cross-resistance with Cry1Ab and Cry1Fa along with independent membrane binding sites in O. nubilalis makes Cry1Bh1 a candidate to further optimize for in-plant resistance to this pest.     

Evaluating the phototactic behavior responses of the diamondback moth,Plutella xylostella,to some different wavelength LED lights in laboratory and field

Light traps equipped with light emitting diodes (LEDs) have been applied to manage some phototactic insect pests. The diamondback moth, Plutella xylostella, is a cosmopolitan insect pest to be seriously harmful to many cruciferous plants. The present research focused on evaluating the phototactic behavior responses of the moths to several wavelengths and photon flux densities of LED lights under laboratory and field conditions. The results from the laboratory showed that the highest phototactic behavior responses of P. xylostella moths were recorded for UV (380 nm) LED light under 1.5 µmol m⁻² s⁻¹ and 2.5 µmol m⁻² s⁻¹. The moths were more attracted to light traps equipped with 4 LEDs and 6 LEDs of 380 nm, respectively, between 20:00 and 22:00 than the other groups and night times in the field. The finding from the field was consistent with the results from the laboratory. We found that the 380 nm LED light results in the strongest attraction rate of the moths by 92.4 % and the moths caught in light trap with the UV LEDs was 1.62 times more than that with a black light. These data clearly demonstrate that P. xylostella moths have a high sensitivity to 380 nm, therefore, a 380 nm LED light trap could be useful for monitoring and controlling the moths.     

A stochastic model for predicting the stage emergence of Plutella xylostella under field conditions

The diamondback moth, Plutella xylostella, is a worldwide pest of brassicas, and its biology and ecology have been extensively studied over recent years. Despite the importance of mathematical models to the management of insect pests, no stochastic model has been developed to date for P. xylostella. In this context, the study aimed to develop a stochastic model capable of describing the stage emergence of P. xylostella under field conditions. The stochastic model was developed using simple nonlinear functions based on the laboratory data on development times under constant temperatures. Comparison between estimated and observed cumulative proportions of egg hatch, pupation and adult emergence recorded in the field in Southern Brazil shows that the model accurately describes the stage emergence of P. xylostella. The developed model shows potential to estimate the stage emergence of P. xylostella under field conditions, and can add significant advances to the management of this pest.     

Ser-Substituted Mutations of Cys Residues in Bacillus thuringiensis Vip3Aa7 Exert a Negative Effect on its Insecticidal Activity

Vegetative insecticidal proteins (VIPs), which were produced by Bacillus thuringiensis during its vegetative growth stage, display a broad insecticidal spectrum to Lepidoptera larvae. Sequence alignment of the Vip3A-type indicates that three cysteine residues were conserved in Vip3A-type proteins. To determine whether these conserved cysteine residues contributed to the insecticidal activity, the three residues were respectively substituted with serine in the Vip3Aa7 protein by site-directed mutagenesis. Bioassays using the third instar larvae of Plutella xylostella showed that the toxicity of C401S and C507S mutants were completely abolished. To find out the inactivity reason of mutants, three mutants and the wild-type Vip3Aa7 were treated with trypsin. The results indicated that the C507S mutant was rapidly cleaved and resulted in decrease of the 62?kDa toxic core fragment. These results indicated that the replacement of the Cys⁵⁰⁷ with a Ser⁵⁰⁷ caused decrease in C507S resistance against trypsin degradation. It is suggesting a possible association between insecticidal activity and trypsin sensitivity of Vip3A proteins. This study serves a guideline for the study of Vip3A protein structure and active mechanism.     

Characterization of Chimeric Bacillus thuringiensis Vip3 Toxins

Bacillus thuringiensis vegetative insecticidal proteins (Vip) are potential alternatives for B. thuringiensis endotoxins that are currently utilized in commercial transgenic insect-resistant crops. Screening a large number of B. thuringiensis isolates resulted in the cloning of vip3Ac1. Vip3Ac1 showed high insecticidal activity against the fall armyworm Spodoptera frugiperda and the cotton bollworm Helicoverpa zea but very low activity against the silkworm Bombyx mori. The host specificity of this Vip3 toxin was altered by sequence swapping with a previously identified toxin, Vip3Aa1. While both Vip3Aa1 and Vip3Ac1 showed no detectable toxicity against the European corn borer Ostrinia nubilalis, the chimeric protein Vip3AcAa, consisting of the N-terminal region of Vip3Ac1 and the C-terminal region of Vip3Aa1, became insecticidal to the European corn borer. In addition, the chimeric Vip3AcAa had increased toxicity to the fall armyworm. Furthermore, both Vip3Ac1 and Vip3AcAa are highly insecticidal to a strain of cabbage looper (Trichoplusia ni) that is highly resistant to the B. thuringiensis endotoxin Cry1Ac, thus experimentally showing for the first time the lack of cross-resistance between B. thuringiensis Cry1A proteins and Vip3A toxins. The results in this study demonstrated that vip3Ac1 and its chimeric vip3 genes can be excellent candidates for engineering a new generation of transgenic plants for insect pest control.     

Engineering of benzylglucosinolate in tobacco provides proof-of-concept for dead-end trap crops genetically modified to attract Plutella xylostella (diamondback moth)

Glucosinolates are biologically active natural products characteristic of crucifers, including oilseed rape, cabbage vegetables and the model plant Arabidopsis thaliana. Crucifer‐specialist insect herbivores, like the economically important pest Plutella xylostella (diamondback moth), frequently use glucosinolates as oviposition stimuli. This suggests that the transfer of a glucosinolate biosynthetic pathway to a non‐crucifer would stimulate oviposition on an otherwise non‐attractive plant. Here, we demonstrate that stable genetic transfer of the six‐step benzylglucosinolate pathway from A. thaliana to Nicotiana tabacum (tobacco) results in the production of benzylglucosinolate without causing morphological alterations. Benzylglucosinolate‐producing tobacco plants were more attractive for oviposition by female P. xylostella moths than wild‐type tobacco plants. As newly hatched P. xylostella larvae were unable to survive on tobacco, these results represent a proof‐of‐concept strategy for rendering non‐host plants attractive for oviposition by specialist herbivores with the long‐term goal of generating efficient dead‐end trap crops for agriculturally important pests.     

Tri-Trophic Insecticidal Effects of African Plants against Cabbage Pests

Botanical insecticides are increasingly attracting research attention as they offer novel modes of action that may provide effective control of pests that have already developed resistance to conventional insecticides. They potentially offer cost-effective pest control to smallholder farmers in developing countries if highly active extracts can be prepared simply from readily available plants. Field cage and open field experiments were conducted to evaluate the insecticidal potential of nine common Ghanaian plants: goat weed, Ageratum conyzoides (Asteraceae), Siam weed, Chromolaena odorata (Asteraceae), Cinderella weed, Synedrella nodiflora (Asteraceae), chili pepper, Capsicum frutescens (Solanaceae), tobacco, Nicotiana tabacum (Solanaceae) cassia, Cassia sophera (Leguminosae), physic nut, Jatropha curcas (Euphorbiaceae), castor oil plant, Ricinus communis (Euphorbiaceae) and basil, Ocimum gratissimum (Lamiaceae). In field cage experiments, simple detergent and water extracts of all botanical treatments gave control of cabbage aphid, Brevicoryne brassicae and diamondback moth, Plutella xylostella, equivalent to the synthetic insecticide Attack^® (emamectin benzoate) and superior to water or detergent solution. In open field experiments in the major and minor rainy seasons using a sub-set of plant extracts (A. conyzoides, C. odorata, S. nodiflora, N. tabacum and R. communis), all controlled B. brassicae and P. xylostella more effectively than water control and comparably with or better than Attack^®. Botanical and water control treatments were more benign to third trophic level predators than Attack^®. Effects cascaded to the first trophic level with all botanical treatments giving cabbage head weights, comparable to Attack^® in the minor season. In the major season, R. communis and A conyzoides treatment gave lower head yields than Attack^® but the remaining botanicals were equivalent or superior to this synthetic insecticide. Simply-prepared extracts from readily-available Ghanaian plants give beneficial, tri-trophic benefits and merit further research as an inexpensive plant protection strategy for smallholder farmers in West Africa.     

Improvement of Bioinsecticides Production by Sporeless <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis> Strains in Response to Various Stresses in Low Cost Medium

The use of bioinsecticides, particularly those produced by sporeless Bacillus thuringiensis strains, has been shown to be a good alternative in pest management. Two types of sporeless mutants were distinguished. The asporogenic mutants which completely lack spores produce a regular bipyramidal crystal inclusion. The oligosporogenic mutants kept the ability to produce insecticidal crystal proteins. However, sporulation in such mutants was not totally blocked and very few of them could still produce spores. In order to improve bioinsecticides production, adaptation of sporeless strains to heat shock and osmotic stress was investigated. Delta-endotoxin production by 78% of sporeless mutants was significantly improved by osmotic stress with an overproduction of about 17%, compared to the wild strain BNS3. However, toxin production was improved by only 21% of mutants after heat shock, in low cost medium. The statistical analysis proved that delta-endotoxin production, cell growth, and spore formation of asporogenic and oligosporogenic mutants depended on the type of applied stress. Each strain has an important potential when applying the adequate stress. Moreover, adaptation of sporeless mutants to NaCl may allow the substitution of all minerals of the medium by diluted sea water which appeared to be a good alternative for the economic production of bioinsecticides at industrial scale which is of great importance from the practical point of view.     

Influence of crucifer cropping system on the parasitism ofPlutella xylostella (Lep., Yponomeutidae) byCotesia plutellae (Hym., Braconidae) andDiadegma semiclausum (Hym., Ichneumonidae)

Field experiments were conducted to study the influence of cabbage monoculture and mixed cropping on the parasitism of diamondback moth,Plutella xylostella (L.), a destructive pest of all crucifers, by 2 larval parasites,Diadegma semiclausum Hellén andCotesia plutellae Kurdjumov. There was no significant difference in parasitism by either species whether cabbage was planted in insecticide-free monoculture or in mixed cropping with 8 noncrucifers which were sprayed twice a week with chemical insecticides mevinphos, methamidophos and permethrin. Population ofP. xylostella increased as the cabbage plants grew older. Parasitism byC. plutellae was higher soon after cabbage transplanting but decreased as the plants grew older. Parasitism byD. semiclausum was very low soon after cabbage planting but increased as the plants grew older. A significant negative correlation was found betwen parasitism byC. plutellae andD. semiclausum. In a caged field study where only one parasite species was used in an individual cage, parasitism ofP. xylostella by both species decreased as theP. xylostella population increased. This is believed to be due to the absence of competition between the two parasites inside the cage. There was no relationship between host-plant age and parasitism ofP. xylostella larvae by either parasite species.     

Genetic control of Plutella xylostella in omics era

Diamondback moth, Plutella xylostella (L.), is a specialist pest on cruciferous crops of economic importance. The large‐scale use of chemical insecticides for the control of this insect pest has caused a number of challenges to agro‐ecosystems. With the advent of the omics era, genetic pest management strategies are becoming increasingly feasible and show a powerful potential for pest control. Here, we review strategies for using transgenic plants and sterile insect techniques for genetic pest management and introduce the major advances in the control of P. xylostella using a female‐specific RIDL (release of insects carrying a dominant lethal gene) strategy. Further, the advantages of gene drive developed in combination with sex determination and CRISPR/Cas9 systems are addressed, and the corresponding prospects and implementation issues are discussed. It is predictable that under the policy and regulation of professional committees, the genetic pest control strategy, especially for gene drive, will open a new avenue to sustainable pest management not only for P. xylostella but also for other insect pests.     

Insecticidal activity of spore-free mutants of Bacillus thuringiensis against the Indian meal moth and almond moth

Three oligosporogenic mutants of Bacillus thuringiensis were assayed for toxicity against larvae of the Indian meal moth, Plodia interpunctella, and the almond moth, Ephestia cautella. The results were compared with insecticidal activity obtained from the parent strain (HD-1) and two standard B. thuringiensis formulations (HD-1-S-1971 and HD-1-S-1980) against the same insect species. The toxicity of the sporeless mutant preparations was significantly diminished against the Indian meal moth (10- to 26-fold increase in LC₅₀) but exceeded the toxicity of the standards against the almond moth. The toxicities of the B. thuringiensis preparations toward the Indian meal moth were consistent with the number of spores in the test samples, but spores did not contribute to toxicity to E. cautella larvae. A rationale for basing dosage on soluble protein was demonstrated for use in situations where spores are not a contributing factor in toxicity.     

Susceptibility of major lepidopterans to δ-endotoxins and a formulation of Bacillus thuringiensis (B.t.) on vegetable brassicas in Taiwan

Baseline susceptibility of Plutella xylostella, Crocidolomia binotalis and Hellula undalis to Bacillus thuringiensis (B.t.) δ-endotoxins (Cry1Aa, Cry1Ab, Cry1Ac, Cry1Ca) and a formulation (Xentari®) was assessed using a leaf-dip bioassay method. The toxins Cry1Ac, Cry1Aa and Cry1Ca were equally toxic to P. xylostella. Crocidolomia binotalis was highly susceptible to all Cry1A toxins and it was least sensitive to Cry1Ca. Conversely, H. undalis was highly sensitive to Cry1Ca, but less susceptible to Cry1A toxins. Hence, the susceptibility of H. undalis to Xentari®, a formulation containing Cry1C as a major toxin, was significantly higher than C. binotalis or P. xylostella.     

Site-Specific Cassette Exchange Systems in the Aedes aegypti Mosquito and the Plutella xylostella Moth

Genetically engineered insects are being evaluated as potential tools to decrease the economic and public health burden of mosquitoes and agricultural pest insects. Here we describe a new tool for the reliable and targeted genome manipulation of pest insects for research and field release using recombinase mediated cassette exchange (RMCE) mechanisms. We successfully demonstrated the established ΦC31-RMCE method in the yellow fever mosquito, Aedes aegypti, which is the first report of RMCE in mosquitoes. A new variant of this RMCE system, called iRMCE, combines the ΦC31-att integration system and Cre or FLP-mediated excision to remove extraneous sequences introduced as part of the site-specific integration process. Complete iRMCE was achieved in two important insect pests, Aedes aegypti and the diamondback moth, Plutella xylostella, demonstrating the transferability of the system across a wide phylogenetic range of insect pests.