Formation characteristics of long-term memory in Bactrocera dorsalis.

Studies on insects have contributed significantly to a better understanding of learning and memory, which is a necessary cognitive capability for all animals. Although the formation of memory has been studied in some model insects, more evidence is required to clarify the characteristics of memory formation, especially long-term memory (LTM), which is important for reliably storing information. Here, we explored this question by examining Bactrocera dorsalis, an agricultural pest with excellent learning abilities. Using the classical conditioning paradigm of the olfactory proboscis extension reflex (PER), we found that paired conditioning with multiple trials (>3) spaced with an intertrial interval (≥10 min) resulted in stable memory that lasted for at least 3 d. Furthermore, even a single conditioning trial was sufficient for the formation of a 2-d memory. With the injection of protein inhibitors, protein-synthesis-dependent memory was confirmed to start 4 h after training, and its dependence on translation and transcription differed. Moreover, the results revealed that the dependence of memory on protein translation exhibited a time-window effect (4-6 h). Our findings provide an integrated view of LTM in insects, suggesting common mechanisms in LTM formation that play a key role in the biological basis of memory.

Insecticidal and P450 mediate metabolism of fluralaner against red imported fire ant, Solenopsis invicta (Hymenoptera: Formicidae).

The red imported fire ant (Solenopsis invicta), a worldwide invasive and polyphagous pest, and often nests in residential areas. Finding an alternative pesticide that is both effective on S. invicta and environmentally friendly is urgent and crucial. Fluralaner, a novel isoxazoline insecticide, has been proven to possess selective toxicity for insects versus mammals and has been safe for mammals and non-target organisms, suggesting its potential in pest management. However, little toxicity information is available for the controlment of S. invicta. In this article, we studied the toxicity of fluralaner against S. invicta systematically, and the roles of metabolism-related enzymes in the metabolism process of fluralaner. The toxicity results showed that the topical application and feeding application were all effective for S. invicta. Moreover, fluralaner can be transmitted among workers by contacting and feeding which leads to a toxic reaction among nestmates. By exploring the biochemistry change, we found cytochrome P450 monooxygenase (P450) may be involved in the detoxification of fluralaner as well as carboxylesterase (CarE), but not glutathione S-transferase (GST). Synergism assays gave solid evidence in which piperonyl butoxide, an activity inhibitor of P450, increased the toxicity of fluralaner to S. invicta. Importantly, with the RNAi treatment, four of S.invicta P450 genes were significantly inhibited and showed more sensitivity to fluralaner at LC50 concentration. Our result indicated that fluralaner could be a potential alternative pesticide in S. invicta control. And CYP9AS16, CYP6AS161, CYP6SQ20, and CYP336A45 genes were closely associated with the metabolism process of fluralaner.

Octopaminergic neurons function in appetitive but not aversive olfactory learning and memory in Bactrocera dorsalis.

The biogenic amine octopamine (OA, invertebrate counterpart of noradrenaline) plays critical roles in the regulation of olfactory behavior. Historically, OA has been thought to mediate appetitive but not aversive learning in honeybees, fruit flies (Drosophila), and crickets. However, this viewpoint has recently been challenged because OA activity through a ß-adrenergic-like receptor drives both appetitive and aversive learning. Here, we explored the roles of OA neurons in olfactory learning and memory retrieval in Bactrocera dorsalis. We trained flies to associate an orange odor with a sucrose reward or to associate methyl eugenol, a male lure, with N,N-diethyl-3-methyl benzoyl amide (DEET) punishment. We then treated flies with OA receptor antagonists before appetitive or aversive conditioning and a memory retention test. Injection of OA receptor antagonist mianserin or epinastine into the abdomen of flies led to impaired of appetitive learning and memory retention with a sucrose reward, while aversive learning and memory retention with DEET punishment remained intact. Our results suggest that the OA signaling participates in appetitive but not aversive learning and memory retrieval in B. dorsalis through OA receptors.

Interaction of fipronil and the red imported fire ant (Solenopsis invicta): Toxicity differences and detoxification responses.

Fipronil (FIP), a phenyl-pryazole pesticide, has been widely used for crop protection due to its broad insecticidal spectrum, especially for urban insect management. FIP also serves as the active ingredient of major baits used for the control of the red imported fire ant (RIFA; Solenopsis invicta). Although a vast majority of laboratory-based research has been performed using worker ants as a model, limited information is available regarding the toxicity of FIP in individuals from different castes and developmental stages. In this study, we investigated the interaction between FIP and this important pest, including FIP toxicity and transformation, RIFA enzyme activity and responses to FIP exposure. The topical and feeding toxicity of FIP in five adult castes, worker larvae and worker pupae were determined and compared. Topical toxicity assays showed that there were significant differences in FIP toxicity among adult workers (LD50 = 1.17 µg/g), larvae (LD50 = 1891.00 µg/g) and pupae (LD50 = 23981.00 µg/g). Although, no obvious differences in topical toxicity were observed among the adult castes, the differences in feeding toxicity were significant. For example, the LC50 value for the workers was 3.96-fold lower than that for soldiers at 24 h, and the LC50 value was slightly lower for male alates than for female alates at day 3 and day 4, respectively. The activities of detoxification enzymes in individuals of different castes and developmental stages were investigated with or without FIP treatment. Cytochrome P450 activity was approximately 24-fold higher in larvae than in workers, and adult workers exhibited 4-fold higher FIP-induced cytochrome P450 activity than individuals from other adult castes. In addition, in vitro experiments demonstrated that FIP was transformed into FIP-sulfone, and this process may be primarily mediated by RIFA P450(s).

Classical olfactory conditioning in the oriental fruit fly, Bactrocera dorsalis.

The oriental fruit fly, Bactrocera dorsalis, is a serious pest of fruits and vegetables. Methyl eugenol (ME), a male attractant, is used to against this fly by mass trapping. Control effect may be influenced by learning, which could modify the olfactory response of the fly to this attractant. To collect the behavioral evidence, studies on the capability of this fly for olfactory learning are necessary. We investigated olfactory learning in male flies with a classical olfactory conditioning procedure using restrained individuals under laboratory conditions. The acquisition of the proboscis extension reflex was used as the criterion for conditioning. A high conditioned response level was found in oriental fruit flies when an odor was presented in paired association with a sucrose reward but not when the odor and sucrose were presented unpaired. We also found that the conditioning performance was influenced by the odor concentration, intertrial interval, and starvation time. A slight sensitization elicited by imbibing sucrose was observed. These results indicate that oriental fruit flies have a high capacity to form an olfactory memory as a result of classical conditioning.

Effects of periplocoside X on midgut cells and digestive enzymes activity of the soldiers of red imported fire ant.

The pathological effects of ingested periplocoside X, an insecticidal component isolated from the root of Periploca sepium Bunge, on the midgut epithelial cells of the soldiers of red imported fire ant were studied and the symptom was described. The results showed that periplocoside X could induce a severe, time-dependent cytotoxicity in the midgut epithelial cells. An optical microscopy showed that epithelial cells swelled firstly and then lysed. Transmission electron microscopy (TEM) showed that numerous swollen lysosomes were appeared, microvilli were disrupted and sloughed off, and the numbers of the rough endoplasmic reticulum and the mitochondria decreased sharply in earlier stage. Numerous vacuoles were observed in the later stage. Finally, periplocoside X resulted in cell death by cytolysis. Assay of main three digestive enzymes activity indicated that amylase activity was significantly inhibited, but no significant changes were seen for lipase activity and total protease activity. So it is suggested that periplocoside X induced mainly to organic damage of midgut epithelium cells of insect. In all, insect midgut is one of targets for periplocoside X.

Bioactivity of Indonesian mahogany, Toona sureni (Blume) (Meliaceae), against the red flour beetle, Tribolium castaneum (Coleoptera, Tenebrionidae) / Bioatividade do mogno da Indonésia, Toona sureni (Blume) (Meliaceae), contra o besouro-das-farinhas, Tribolium castaneum (Coleoptera, Tenebrionidae)

Bioactivity of Indonesian mahogany, Toona sureni (Blume) (Meliaceae), against the red flour beetle, Tribolium castaneum (Coleoptera, Tenebrionidae). The insecticidal activity of Toona sureni (Blume) Merr. was evaluated considering repellency, mortality and progeny production of F1 adults of Tribolium castaneum (Herbst, 1797) (Coleoptera, Tenebrionidae). Dried extract of seeds of T. sureni was dissolved in acetone to prepare solution of various concentrations (0.5, 1.0, 2.5 and 5.0%). To test for repellency, the insects were exposed to treated filter paper. Mortality of larvae, pupae and adults was evaluated by the treatment of spraying the insects with different concentrations of T. sureni extract. Residual effect of the extract was also evaluated considering the production of progeny of F1 adults. The highest repellency (93.30%) of T. castaneum occurred at the highest concentration (5.0% suspension of T. sureni); while the lowest (0.0%) repellency occurred at 0.5% suspension after 1 day of treatment. The highest mortality against adults (86.71%), larvae (88.32%) and pupae (85%) occurred at 5% suspension at 8 days after application. There was a negative correlation between the concentrations of T. sureni and the production of F1 adult's progeny of T. castaneum. The highest number of progeny (147) of T. castaneum occurred in the control at 7 days after treatment; and the lowest number of progeny (43) occurred at 5.0% concentration in 1 day after treatment. The results show that T. sureni is toxic to T. castaneum and has the potential to control all stages of this insect in stored wheat.

Bioatividade do mogno da Indonésia, Toona sureni (Blume) (Meliaceae), contra o besouro-das-farinhas, Tribolium castaneum (Coleoptera, Tenebrionidae). A atividade inseticida de Toona sureni (Blume) Merr. foi avaliada considerando repelência, mortalidade e a produção de progênie de adultos F1 de Tribolium castaneum (Herbst, 1797) (Coleoptera, Tenebrionidae). Extrato seco de sementes de T. sureni foi dissolvido em acetona, para preparar soluções de várias concentrações (0,5; 1,0; 2,5 e 5,0%). Para testar a repelência, os insetos foram expostos a papel de filtro tratado. A mortalidade de larvas, pupas e adultos foi avaliada pulverizando os insetos com diferentes concentrações do extrato de T. sureni. O efeito residual do extrato também foi avaliado pela produção de progênie dos adultos F1. A maior repelência (93,30%) de T. castaneum ocorreu na maior concentração (5,0% suspensão de T. sureni); enquanto que a mais baixa repelência (0%) ocorreu na suspensão de 0,5% em um dia após o tratamento. A mortalidade mais elevada de adultos (86,71%), larvas (88,32%) e de pupas (85%) ocorreu com a suspensão a 5%, aos oito dias após a aplicação. Houve uma correlação negativa entre a concentração das suspensões de T. sureni e a produção de progênie de T. castaneum. O maior número de progênie (147) foi produzido na testemunha aos sete dias após o tratamento; enquanto o menor número de progênie (43) foi na suspensão de 5,0% no primeiro dia após o tratamento. Os resultados demostram que T. sureni é tóxico para T. castaneum e tem potencial para ser usado no controle de todos os estágios deste inseto em trigo armazenado.

Chemical constituents from the roots of Periploca sepium with insecticidal activity.

Five compounds were isolated from the root powder of Periploca sepium. By mainly HR-ESI-MS, (1)H, (13)C, and 2D NMR spectral data, they were characterized as periplocoside X (1), oligasaccharide A (2), periplocoside A (3), periplocoside E (4), and periplocoside N (5), respectively. Compounds 1-5 were found to possess insecticidal activities against the red imported fire ant. Among the compounds, periplocoside X showed significant activity with LD(50) values of 748.99, 116.62, 2169.58, and 3079.33mg/l against soldiers, workers, males, and alate females of red imported fire ant, respectively.

[Modification of the analytical method for rotenoids in plants].

Rotenoids are the active ingredients of some botanical insecticides and prospective candidates as anticancer agents. The proper isolation and determination of rotenoids in plants is of great importance for their further research and development. However, the HPLC method available for this purpose was developed particularly for the detection and determination of rotenone, so it appears to be unsuitable for the analysis of other rotenoids such as deguelin, elliptone and their analogues. By checking the UV spectra, it has been found that four types of UV absorption patterns occurred among the major rotenoids isolated from the roots of Derris elliptica and leaves of Tephrosia vogelii, and that the detection wavelength at 240 nm is more adequate for the analysis of a complex of rotenoids than at 280 nm-300 nm, which is used for rotenone detection. The extraction of rotenoids from plants is conveniently carried out by CHCl3-MeOH(9:1, V/V) and the purification can be accomplished by filtration of the crude residue through a C18 reversed-phase cartridge. Rotenoids can be isocratically eluted by MeOH-H2O(66:34, V/V). The results showed that rotenone, deguelin, elliptone, and their 12a-hydroxy- and 6a,12a-dehydro-analogs can be easily detected by the modified method, along with a satisfactory peak separation. The rotenoid components might be characterized by their retention times and relative retention times based on rotenone, which were at a range of 3.26 min-39.42 min and 40.4%-489.1% respectively.