Biofabricated nanoparticles: Their delivery and utility in Plutella xylostella management

The diamondback moth (DBM), Plutella xylostella (L.) (Lepidoptera: Plutellidae) is one of the most serious cosmopolitan pests of cruciferous plants. It causes severe damage to crop quality and production, with estimated global losses of 20-40% per year. Consumer preference regarding pest-free and uncontaminated crop, unavailability of highly competitive natural enemies as well as migratory nature of DBM has obligated for the mandatory use of insecticides. The over reliance and indiscriminate use of chemical insecticides to control DBM has lead to development of resistance against almost all the modern insecticides. In addition, it has also developed resistance against most of the Bt strain. To overcome the problem of resistance, there is an urgent need for new and highly efficient method to control DBM which must be eco-friendly, cost effective and has minimum impact on the environment. One of such alternatives is nanotechnology which fulfil all the criteria to become an ideal insecticide or delivery system for insect pest management especially DBM. Nanotechnology offers great improvement in the field of pesticides as it is less toxic, shelf-life enhancement and higher water solubility. Loading of Indoxacarb on nanoparticles are reported to be highly effective they suppressed the activity of detoxification enzymes such as GST, CarE, and P450. Green synthesis of AgNPs, Single walled carbon nano tube (SWCNT) also exhibits larvicidal and pupicidal effect. These are also reported to adversely affect food consumption, growth, pupation and fecundity of P. xylostella.These nanoparticles have been found to be more stable, also the controlled release of active ingredient for a long time and provides target specific control of P. xylostella for solving adverse situations ofthe crop fields like less food security, lesser food productivity and environmental imbalance.

Function and Characterization of the Argonaute2 Gene in the RNA Interference Pathway in the Diamondback Moth

Arch BiochemMol Biol2019;10(4):035-051DOI: 10.26502/abmb.007Archives of Biochemistry and Molecular BiologyVol. 10 No. 4 -December2019. 36AbstractArgonaute2 (AGO2) is a core catalytic component of the RNA-induces silencing complex (RISC) that binds to small guide RNAs containing small interfering RNA (siRNA) and microRNA (miRNA). The guide RNA leads RISC to the complementary mRNA for gene suppression. We cloned the full length cDNA (2193 bp) of the Ago2gene (PxAgo2) from diamondback moth (DBM, Plutella xylostella). The predicted PxAgo2 protein hadan83 kDa molecular weight withatheoretical isoelectric point of 9.39. The phylogenetic tree showed a high similarity of PxAgo2with Bombyx moriAgo2 (BmAgo2). Western blot and RT-qPCR analyses showed a clear increase in the PxAgo2 mRNA and protein expression levels in the egg, 4thinstar larva, pre-pupa, pupa and adult. The double-stranded RNA-mediated RNAi of PxAgo2in DBM larvae was found 3 h after dsRNA injection,and the knockdown level was increased over time up to 36 h. PxAgo2silencing recovered the expression of PxBurs-?(Bursicon-? inDBM) to the normal expression level, which was suppressed bydsPxBurs-?in a DBM cell line. The overexpression of PxAgo2fundamentally enhanced the PxBurs-? silencing efficiency in DBM cells. Our findings reveal that PxAgo2is involved in the dsRNA-regulated gene silencing mechanism and performs a crucial function in the RNAi process ofDBM

Toxicity of soursop extracts to diamondback moth / Toxicidade de extratos de graviola sobre a traça-das-crucíferas

This study aimed to evaluate the effects of three solvent extractors (water, ethanol and hexane) of grounded seeds of soursop, Annona muricata L. (Annonaceae), in the mortality, biology and oviposition of Plutella xylostella L. (Lepidoptera: Plutellidae). The results showed that the LC50 and LC99 were 0.0133 and 0.084%, 0.025% and 0.196%, 2.33 and 35.22% for the ethanolic, hexanic and aqueous extracts, respectively. The organic extracts affected only the larval phase and reduced viability in more than 60%, but did not affect pupal stage of the remaining larvae. Furthermore, the ethanolic extract at lethal concentraction also affected negatively the embryonic phase. The results lead to the conclusion that the ethanolic extract of soursop grounded seeds is a viable alternative to control diamondback moth on vegetables.

O presente trabalho teve como objetivo avaliar o efeito de extratos de sementes de graviola, Annona muricata L. (Annonaceae) obtidos com diferentes solventes sobre Plutella xylostella L. (Lepidoptera: Plutellidae). Foram estimadas as concentrações letais de três solventes extratores (água, hexano e etanol) e seus efeitos na biologia e oviposição. Os valores estimados das concentrações letais foram de 0,013 e 0,084%; 0,025 e 0,196%; 2,33 e 35,22%, para as CL50 e CL99 do extrato etanólico, hexânico e aquoso, respectivamente. Os extratos orgânicos afetaram apenas a fase larval e reduziu a viabilidade em mais de 60%, mas não afetou a fase pupal das lagartas remanescentes. Além disso, o extrato etanólico na concentração letal se mostrou eficiente afetando negativamente a fase embrionária. Conclui-se que o extrato etanólico da graviola é uma alternativa viável no controle da traça.

Detection of carboxylesterase and esterase activity in culturable gut bacterial flora isolated from diamondback moth, Plutella xylostella (Linnaeus), from India and its possible role in indoxacarb degradation

Abstract Diamondback moth (DBM), Plutella xylostella (Linnaeus), is a notorious pest of brassica crops worldwide and is resistant to all groups of insecticides. The insect system harbors diverse groups of microbiota, which in turn helps in enzymatic degradation of xenobiotic-like insecticides. The present study aimed to determine the diversity of gut microflora in DBM, quantify esterase activity and elucidate their possible role in degradation of indoxacarb. We screened 11 geographic populations of DBM in India and analyzed them for bacterial diversity. The culturable gut bacterial flora underwent molecular characterization with 16S rRNA. We obtained 25 bacterial isolates from larvae (n = 13) and adults (n = 12) of DBM. In larval gut isolates, gammaproteobacteria was the most abundant (76%), followed by bacilli (15.4%). Molecular characterization placed adult gut bacterial strains into three major classes based on abundance: gammaproteobacteria (66%), bacilli (16.7%) and flavobacteria (16.7%). Esterase activity from 19 gut bacterial isolates ranged from 0.072 to 2.32 µmol/min/mg protein. Esterase bands were observed in 15 bacterial strains and the banding pattern differed in Bacillus cereus – KC985225 and Pantoea agglomerans – KC985229. The bands were characterized as carboxylesterase with profenofos used as an inhibitor. Minimal media study showed that B. cereus degraded indoxacarb up to 20%, so it could use indoxacarb for metabolism and growth. Furthermore, esterase activity was greater with minimal media than control media: 1.87 versus 0.26 µmol/min/mg protein. Apart from the insect esterases, bacterial carboxylesterase may aid in the degradation of insecticides in DBM.

Preparation and in vitro evaluation of antibacterial peptides from Plutella xylostella poly(lactic-co-glycolic acid) nanoparticles / 中草药

Objective: To prepare the antibacterial peptides from Plutella xylostella-loaded nanoparticles based on poly(lactic-co- glycolic acid) (CA-PLGA-NPs) and evaluate its physicochemical property and in vitro release. Methods: CA-PLGA-NPs were prepared by S/W/O/W double emulsion method combined with high-pressure homogenization. The morphology, particle size, polydispersion index (PDI), drug loading, encapsulation efficiency (EE), and in vitro release of the nanoparticles were studied. Results: CA-PLGA-NPs were spherical or similarly spherical, and the average particle size, PDI, drug loading, and EE were (358.76 ± 22.51) nm, 0.168 1 ± 0.012 2, (10.50 ± 0.28)%, and (60.92 ± 1.58)%, respectively. And burst phenomenon was not significant. The drug delivery stable phase was 2-10 d. Conclusion: S/W/O/W double emulsion method combined with high pressure homogenization method is suitable for the preparation of CA-PLGA-NPs, and provides a pharmaceutical basis for CA administration.

Consumo foliar da traça-das-crucíferas em couve e brócolis tratados com sinigrina / Leaf area consumed by diamondback moth feeding on kale and brocolis treated with sinigrin

A estimativa da área foliar, em função da sua redução causada pelo consumo por insetos filófagos, é uma metodologia básica em várias áreas da Entomologia, como, por exemplo, em estudos sobre resistência de plantas, entomologia econômica e ecologia nutricional. Um dos fatores que pode interferir na quantidade de área foliar consumida pelo fitófago é a presença de certos químicos nas folhas, como a sinigrina em brassicáceas. Dessa forma, foram aplicadas em folhas de couve e brócolis diversas concentrações de sinigrina (0,2, 0,4, 0,8, 1,6, e 3,2 mg/mL em solução a 5% de Tween20(r)), medindo-se o consumo da área foliar por lagartas de P. xylostella, o qual foi aferido pela porcentagem de redução da área, diferença de peso e escala visual de notas. Os resultados mostraram que as concentrações 0,2, 0,4 e 3,2 mg/mL de sinigrina aumentam o consumo em folhas de couve e diminuem em folhas de brócolis por lagartas de P. xylostella. As metodologias utilizadas proporcionam resultados similares, sendo as estimativas de peso fresco e nota visual mais práticas.(AU)

Leaf area estimation, based on their reduction due to consumption by phytophagous insects, is a basic methodology in several areas of Entomology, for instance, in studies about plant resistance, economic entomology and nutritional ecology. One of the factors that can affect the amount of leaf area consumed by insects is the presence of certain chemicals in the leaves, such as sinigrin in Brassicaceae. Thus, several concentrations of sinigrin were applied on the leaves of cabbage and broccoli (0.2, 0.4, 0.8, 1.6, and 3.2 mg/mL in 5% Tween20(r) solution) in order to measure leaf area consumption by larvae of P. xylostella, estimated by the percentage of area reduction, weight loss and visual scale of notes. The results show that the concentrations of 0.2, 0.4 and 3.2 mg/mL of sinigrin increase larvae consumption of P. xylostella on kale leaves and decrease it on broccoli leaves. The methodologies used provide similar results, and fresh weight measurements and visual grades are more practical.(AU)

RP-HPLC determination of pxCECAl from Plutella xylostella L / 中国药学杂志

OBJECTIVE: To establish a method for determination of cecropin from Plutella xylostella (pxCECAl) by RP-HPLC. METHODS: An Agilent Alltima C18 column (4.6 mm × 250 mm, 5 μm) was adopted, and the mobile phase was composed of metha-nol-acetonitrile-0.05% TFA (2.5:2.5:95) at a flow rate of 1.0 mL · min-1. The detection wavelength was set at 275 nm and the column temperature was 25°C. RESULTS: The linear range of the calibration curve for cecropin was 0.08-0.800 μg (r=0.9994, n=6). The average recovery was 99.69%, and RSD was 1.20% (n=6), respectively. CONCLUSION: The method is rapid, simple, and accurate with good reproducibility. It can be used for quantitative analysis and quality control of cecropin from Plutella xylostella.

Efeito da idade, fotoperíodo e disponibilidade de hospedeiro no comportamento de parasitismo de Oomyzus sokolowskii Kurdjumov (Hymenoptera: Eulophidae) / Effect of age, photoperiod and host availability on the parasitism behavior of Oomyzus sokolowskii Kurdjumov (Hymenoptera: Eulophidae)

Plutella xylostella (L.) é uma praga de difícil controle em virtude de seu alto potencial reprodutivo e de causar danos, e ampla distribuição geográfica, além de apresentar resistência a inseticidas. No entanto, as fases de larva e pupa dessa praga são naturalmente parasitadas por Oomyzus sokolowskii (Kurdjumov), favorecendo o incremento do parasitismo natural. Este estudo investigou o efeito da idade do parasitóide, disponibilidade de hospedeiro e fotoperíodo no comportamento reprodutivo de O. sokolowskii. O número de encontros entre parasitóide e hospedeiro e a taxa de parasitismo aumentaram com a idade das fêmeas até 96h. O parasitismo foi maior quando fêmeas do parasitóide receberam diariamente um número constante de hospedeiros comparado à disponibilidade randômica (13,3 versus 8,9 lagartas). A atividade de parasitismo ocorreu até 20 dias de idade e com longevidade semelhante entre os tratamentos (constante = 33,5 d; randômico = 34,7 d). O número de descendentes por fêmea, no entanto, decresceu com a sua idade, sendo o mesmo observado para o número de parasitóides emergidos por lagarta parasitada. A exposição contínua do hospedeiro à luz ou ao escuro por 12h não afetou a taxa de parasitismo; porém, parasitóides apresentaram maior atividade após 3h de exposição à luz. Assim, futuros estudos e possível utilização de O. sokolowskii, em campo, devem considerar a liberações de parasitóides com idade entre 48h e 72h e, preferencialmente, na madrugada para maiores chances de sucesso de parasitismo.

The high reproduction rate, potential to cause damage, wide geographic distribution and resistance to insecticides of Plutella xylostella (L.) makes difficult its efficient control. However, larvae and pupae of this pest are naturally parasitized by Oomyzus sokolowskii (Kurdjumov), providing opportunities to improve the natural parasitism. This study investigated the effects of the age of adult parasitoids, host availability and time of exposure on O. sokolowskii parasitism behavior. The number of larvae encounters by parasitoid females and the parasitism rate increased with parasitoid age up to 96h. The parasitization was higher when wasps received a constant number of hosts daily in comparison with a random number (13.3 versus 8.9 larvae parasitized). Female parasitization activity was maintained up to the age of 20 days in both treatments and exhibited similar longevity (constant host = 33.5 d; and random host = 34.7 d). The progeny produced per female and the number of parasitoids emerged per host significantly decreased as wasps aged. There was no significant effect of the light regime (12h darkness or 12h light exposure) on the parasitization, although parasitoid was more active after 3h of light exposure. Therefore, further studies on field application of O. sokolowskii should consider the release of 48h- to 72h-old parasitoids at dawn as a way to increase the success of host parasitization.

Ação de extratos de plantas na oviposição e na mortalidade da traça-das-crucíferas / Action of plants extracts on oviposition and on mortality of diamondback moth

A traça-das-crucíferas, Plutella xylostella (L.) (Lepidoptera: Yponomeutidae), é uma das principais espécies de inseto-praga de plantas da família Brassicaceae no Brasil e no mundo. A utilização de extratos de plantas inseticidas no controle desse inseto é uma alternativa promissora. O presente trabalho teve como objetivo avaliar, em laboratório, o efeito de extratos de plantas na oviposição e na mortalidade de P. xylostella. Para análise do efeito sobre a oviposição, pedaços de folhas de couve foram expostos, em gaiola, aos seguintes tratamentos: extratos aquosos (10 por cento p/v) de folha de cinamomo (Melia azedarach), de ramo de cinamomo, de pó-de-fumo (Nicotiana tabacum), além de DalNeem (produto comercial à base de Azadirachta indica), a 10 por cento v/v. No interior da gaiola, foram liberados 50 adultos de P. xylostella e, após 48h, foi contado o número de ovos por folha, em sete repetições. Um segundo ensaio constou da exposição de três larvas aos extratos referidos anteriormente em tubos de vidro com um pedaço de folha de couve previamente tratado. Foram utilizados três tubos por tratamento, com quatro repetições. O número de larvas mortas foi avaliado diariamente. Os extratos aquosos de folha de cinamomo, de ramo de cinamomo e de pó-de-fumo, além de DalNeem, reduzem a oviposição de P. xylostella e a utilização de extrato de pó-de-fumo, de DalNeem e de ramo de cinamomo resultam em controle eficiente de larvas de P. xylostella a partir do segundo, do quinto e do sexto dias após a aplicação, respectivamente.

The diamondback moth, Plutella xylostella (L.) (Lepidoptera, Yponomeutidae), is one of the most important insect pest species of the Brassicaceae family plants in Brazil and in the world. The use of insecticidal plants extracts in the control of this insect is a promising alternative. Therefore, this research goal is to evaluate, in laboratory, the plants extracts effect on the oviposition and mortality of P. xylostella. In order to analyze the effect on the oviposition, kale leaf pieces were treated, in cage, as following: aqueous extracts (10 percent p/v) of leaf of chinaberry (Melia azedarach), of branch of chinaberry, of tobacco powder (Nicotiana tabacum), besides DalNeem (commercial product, based on Azadirachta indica), at 10 percent v/v. Inside the cage, 50 adults of P. xylostella were put and, 48h later, the egg number per leaf was counted, in seven repetitions. A second assay was consisted of the exposition of three larvae to the same cited extracts, in glass tube with a kale leaf piece previously treated. Three glass tubes were used per treatment, with four repetitions. The number of death larvae was evaluated daily. The aqueous extracts of chinaberry leaf, chinaberry branch and tobacco powder, besides DalNeem, reduce the oviposition of P. xylostella and the use of tobacco powder extract, DalNeem and chinaberry branch extract result in efficient control of P. xylostella larvae after the second, fifth and sixth days after application, respectively.

Interação tritrófica e influência de produtos químicos e vegetais no complexo: brássicas x traça-das-crucíferas x parasitóides de ovos / Tritrofic interaction and influence of insecticides and plant products on the complex: brassica diamondback moth egg parasitoids

Objetivou-se avaliar a interação tritrófica no complexo hospedeiro-vegetal (brássicas) x praga/hospedeiro-natural Plutella xylostella Linnaeus x inseto-entomófago (parasitóides - Trichogrmma pretiosum Riley e T. exiguum Pinto & Platner), associada a alguns produtos químicos e vegetais com efeito inseticida, utilizando-se os cultivares de repolho verde - Chato de quintal e híbrido Midori; roxo - Roxo precoce e Híbrido roxo - TPC00682; e couve manteiga - Geórgia e hibrido Geórgia HS20, pulverizadas com os inseticidas: lufenuron (2,52 ml/100L) e deltametrina (32 ml/100L), os produtos vegetais óleo de nim a 0,16 por cento e extrato pirolenhoso a 3,0 por cento, controle (água). Foi avaliada a interação das cultivares com os compostos por meio da exposição de lagartas recém-eclodidas aos produtos, avaliando-se os insetos nas fases de desenvolvimento até a emergência dos adultos. Para avaliar o efeito desses compostos sobre os parasitóides, foram empregados ovos de uma geração F2 de P. xylostella oriunda de lagartas alimentadas com folhas de brássicas, pulverizadas com esses produtos. A associação de produtos químicos ou vegetais, com efeito inseticida, com as cultivares de brássicas permitiu o manejo mais eficaz, especialmente na interação extrato pirolenhoso x a cultivar de repolho Chato de quintal. Observa-se que a interação entre as cultivares e os produtos pode ser prejudicial à atuação do parasitóide Trichogramma, sendo necessária uma avaliação criteriosa para minimizar o efeito sobre inimigos naturais.

The aim of this work was to evaluate the tritrofic interaction in brassica complex: host-vegetable (brassica) vs. pest/natural-host (Plutella xylostella) vs. entomophagous-insect (parasitoid - Trichogramma pretiosum and Trichogramma exiguum), combined with chemical and vegetable products with insecticide action. The cultivar used were: green cabbage - Chato de quintal and Midori hybrid; purple cabbage - Roxo Precoce and purple hybrid - TPC00682; kale - Georgia and Georgia hybrid HS20. They were sprayed using the insecticides - lufenuron (2,52 ml/100L) and deltamethrin (32 ml/100L); the vegetal products - neen oil (0,16 percent) and pyroligneous extract (3,0 percent); and water (check). The cultivars interaction was evaluated with the products, by larvae exposition to the treatments, being analyzed the different phases of development through adults emergence. To evaluate the effect of these products on the parasitoids, P. xylostella, eggs from F2 generation were used. Eggs of this pest were collected from adults fed with brassica leaves that were sprayed with the mentioned products. The insecticides association and vegetable products combined with brassica cultivars became more effective to manage the pest control, especially in the pyroligneous extract x Chato de Quintal interaction. The interaction between the cultivars and products may be harmful to Trichogramma performance being necessary a discerning evaluation to minimize the effect on natural enemies.

Effect of the age of Trichogramma exiguum and diamondback moth embryonic development on the biological characteristics of the parasitoid - DOI: 10.4025/actascibiolsci.v29i2.522 / Efeito da idade de Trichogramma exiguum e do desenvolvimento embrionário da Traça-das-Crucíferas sobre as características biológicas do parasitóide - DOI: 10.4025/actascibiolsci.v29i2.522

The objective of this work was to evaluate the influence of the embryonic development of eggs from the P. xylostella host on Trichogramma exiguum females of different ages. Fifty females either just-emerged or 24, 48, 72 and 96 hours after emergence of this parasitoid were divided into five groups. Each female was offered a recipient with 30 P. xylostella eggs, which were 1-, 2- and 3-days-old. The greatest rates of parasitism were observed on eggs with 3 days of embryonic development, with exception of those 72-hours-old. Viability was not influenced by parasitoid age or host embryonic stage. Sex ratio was similar in all three embryonic host stages for T. exiguum females that were 24-, 48- and 72-hours-old. While evaluating parasitoid progeny obtained from eggs with 3, 2 and 1 days of embryonic development, it was verified that the highest values of longevity were obtained when the females used were just-emerged, or 48- and 72-hours-old, respectively. These results indicate the importance of the age of the parasitoid, as well as the host embryonic development period, in order to maintain quality of T. exiguum at laboratory conditions, in mass culture, as well as in field releases for biological pest control.

Este trabalho teve como objetivo avaliar a influência do desenvolvimento embrionário dos ovos do hospedeiro P. xylostella sobre fêmeas de Trichogramma exiguum, com diferentes idades. Para tanto, cinquenta fêmeas recém-emergidas ou com 24, 48, 72, e 96 horas após a emergência deste parasitóide foram divididas em cinco lotes. Para cada fêmea, foi oferecida uma cartela, contendo 30 ovos de P. xylostella com um, dois e três dias de idade. As maiores taxas de parasitismo foram observadas em ovos de três dias de desenvolvimento embrionário, independente da idade do parasitóide, com exceção daqueles com 72 horas de idade. A viabilidade não foi influenciada pela idade do parasitóide e nem pela fase embrionária do hospedeiro. A razão sexual foi semelhante nas três fases embrionárias do hospedeiro para fêmeas de T. exiguum com 24, 48 e 72 horas de idade. Ao avaliar os descendentes do parasitóide provenientes de ovos com três, dois e um dia de desenvolvimento embrionário, verificou-se que os maiores valores de longevidade foram obtidos quando foram usadas fêmeas recém-emergidas ou com 48 e 72 horas de vida, respectivamente. Estes resultados indicam a importância da idade do parasitóide, bem como o perído de desenvolvimento embrionário do hospedeiro para manter a qualidade de T. exiguum em laboratório, em uma criação massal, e também nas liberações de campo pa

Performance of an oligophagous insect in relation to the age of the host plant / Performance de um inseto oligófago em relação à idade da planta hospedeira

The performance of monophagous and oligophagous herbivore insects should be negatively influenced by the ageing of their host plants. Plutella xylostella L. (Lepidoptera: Yponomeutidae), a specialist in Cruciferae, was reared in the laboratory on detached cabbage leaves (Brassica oleraceae var. capitata). We used fourth expanded leaves from differently aged plants, and leaves from three strata of the mature plant. Plant ageing increased pre-imaginal mortality and reduced larval developing rate, pupae weight, and fecundity. The insect net reproductive rate (Ro) and intrinsic rate of population growth (r) decreased as plant aged. No leaf category of mature plants was qualitatively superior to leaves from younger plants. If there exist a positive correlation between oviposition preference and offspring performance, in field conditions, a decreasing temporal trend in the intensity of attack along the host plant development can be expected.

A performance de insetos folívoros monófagos e oligófagos deveria ser afetada negativamente pelo envelhecimento das suas plantas hospedeiras. Plutella xylostella L. (Lepidoptera: Yponomeutidae), uma especialista em Cruciferae, foi criada em laboratório com folhas destacadas de repolho (Brassica oleraceae var. capitata). Foram utilizadas a quarta folha expandida de plantas de diferentes idades mais as folhas dos três estratos da planta madura. O envelhecimento da planta aumentou a mortalidade pre-imaginal e reduziu a taxa de desenvolvimento das larvas, o peso das pupas e a fecundidade dos adultos. A taxa reprodutiva líquida (Ro) e a taxa intrínseca de crescimento populacional (r) diminuíram com o envelhecimento da planta. Nenhuma das categorias de folhas da planta madura mostrou-se qualitativamente superior às folhas de plantas mais jovens. Se em condições de campo existir uma correlação positiva entre preferencia de oviposição e performance da prole, espera-se uma tendência temporal decrescente na intensidade de ataque de larvas ao longo do desenvolvimento da planta.