Behavioral and metabolic effects of sublethal doses of two insecticides, chlorpyrifos and methomyl, in the Egyptian cotton leafworm, Spodoptera littoralis (Boisduval) (Lepidoptera: Noctuidae)

International audienceInsecticides have long been used as the main method in limiting agricultural pests, but their widespread use has resulted in environmental pollution, development of resistances, and biodiversity reduction. The effects of insecticides at low residual doses on both the targeted crop pest species and beneficial insects have become a major concern. In particular, these low doses can induce unexpected positive (hormetic) effects on pest insects, such as surges in population growth exceeding what would have been observed without pesticide application. Methomyl and chlorpyrifos are two insecticides commonly used to control the population levels of the cotton leafworm Spodoptera littoralis, a major pest moth. The aim of the present study was to examine the effects of sublethal doses of these two pesticides, known to present a residual activity and persistence in the environment, on the moth physiology. Using a metabolomic approach, we showed that sublethal doses of methomyl and chlorpyrifos have a systemic effect on the treated insects. We also demonstrated a behavioral disruption of S. littoralis larvae exposed to sublethal doses of methomyl, whereas no effects were observed for the same doses of chlorpyrifos. Interestingly, we highlighted that sublethal doses of both pesticides did not induce a change in acetylcholinesterase activity in head of exposed larva

Is the rapid post-mating inhibition of pheromone response triggered by ecdysteroids or other factors from the sex accessory glands in the male moth Agrotis ipsilon?

In many animals, male copulation is dependent on the detection and processing of female-produced sex pheromones, which is generally followed by a sexual refractory post-ejaculatory interval (PEI). In the male moth, Agrotis ipsilon, this PEI is characterized by a transient post-mating inhibition of behavioral and central nervous responses to sex pheromone, which prevents males from re-mating until they have refilled their reproductive tracts for a potential new ejaculate. However, the timing and possible factors inducing this rapid olfactory switch-off are still unknown. Here, we determined the initial time delay and duration of the PEI. Moreover, we tested the hypothesis that the brain, the testis and/or the sex accessory glands (SAGs) could produce a factor inducing the PEI. Lastly, we investigated the possible involvement of ecdysteroids, hormones essential for development and reproduction in insects, in this olfactory plasticity. Using brain and SAG cross-injections in virgin and newly-mated males, surgical treatments, wind tunnel behavioral experiments and EIA quantifications of ecdysteroids, we show that the PEI starts very shortly after the onset of copulation, and that SAGs contain a factor, which is produced/accumulated after copulation to induce the PEI. Moreover, SAGs were found to be the main source of ecdysteroids, whose concentration decreased after mating, whereas it increased in the haemolymph. 20-Hydroxyecdysone (20E) was identified as the major ecdysteroid in SAGs of A. ipsilon males. Finally, 20E injections did not reduce the behavioral pheromone response of virgin males. Altogether our data indicate that 20E is probably not involved in the PEI

Isolation and Maintenance-Free Culture of Contractile Myotubes from Manduca sexta Embryos

Skeletal muscle tissue engineering has the potential to treat tissue loss and degenerative diseases. However, these systems are also applicable for a variety of devices where actuation is needed, such as microelectromechanical systems (MEMS) and robotics. Most current efforts to generate muscle bioactuators are focused on using mammalian cells, which require exacting conditions for survival and function. In contrast, invertebrate cells are more environmentally robust, metabolically adaptable and relatively autonomous. Our hypothesis is that the use of invertebrate muscle cells will obviate many of the limitations encountered when mammalian cells are used for bioactuation. We focus on the tobacco hornworm, Manduca sexta, due to its easy availability, large size and well-characterized muscle contractile properties. Using isolated embryonic cells, we have developed culture conditions to grow and characterize contractile M. sexta muscles. The insect hormone 20-hydroxyecdysone was used to induce differentiation in the system, resulting in cells that stained positive for myosin, contract spontaneously for the duration of the culture, and do not require media changes over periods of more than a month. These cells proliferate under normal conditions, but the application of juvenile hormone induced further proliferation and inhibited differentiation. Cellular metabolism under normal and low glucose conditions was compared for C2C12 mouse and M. sexta myoblast cells. While differentiated C2C12 cells consumed glucose and produced lactate over one week as expected, M. sexta muscle did not consume significant glucose, and lactate production exceeded mammalian muscle production on a per cell basis. Contractile properties were evaluated using index of movement analysis, which demonstrated the potential of these cells to perform mechanical work. The ability of cultured M. sexta muscle to continuously function at ambient conditions without medium replenishment, combined with the interesting metabolic properties, suggests that this cell source is a promising candidate for further investigation toward bioactuator applications

Establishment of a New Cell Line from Lepidopteran Epidermis and Hormonal Regulation on the Genes

When an insect molts, old cuticle on the outside of the integument is shed by apolysis and a new cuticle is formed under the old one. This process is completed by the epidermal cells which are controlled by 20-hydroxyecdysone (20E) and juvenile hormone. To understand the molecular mechanisms of integument remolding and hormonal regulation on the gene expression, an epidermal cell line from the 5th instar larval integument of Helicoverpa armigera was established and named HaEpi. The cell line has been cultured continuously for 82 passages beginning on June 30, 2005 until now. Cell doubling time was 64 h. The chromosomes were granular and the chromosome mode was from 70 to 76. Collagenase I was used to detach the cells from the flask bottom. Non-self pathogen AcMNPV induced the cells to apoptosis. The cell line was proved to be an epidermal cell line based on its unique gene expression pattern. It responded to 20E and the non-steroidal ecdysone agonist RH-2485. Its gene expression could be knocked down using RNA interference. Various genes in the cell line were investigated based on their response to 20E. This new cell line represents a platform for investigating the 20E signaling transduction pathway, the immune response mechanism in lepidopteran epidermis and interactions of the genes

Effects of phthalate and bisphenol plasticizers on the activity of glycolytic enzymes of the moth Spodoptera littoralis

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Comparative physiological plasticity to desiccation in distinct populations of the malarial mosquito Anopheles coluzzii

International audienceBackgroundIn West Africa, populations of the malaria vector mosquito, Anopheles coluzzii, are seasonally exposed to strong desiccating conditions during the dry season. Their dynamics strictly follows the pace of the availability of suitable larval development sites (water collections). Accordingly, mosquitoes can reproduce all year long where permanent breeding is possible, or stop reproduction and virtually disappear at the onset of the dry season when surface water dries up, like observed in temporary habitats of dry savannah areas. This highlights the strong adaptive abilities of this mosquito species, which relies at least in part, upon physiological and molecular mechanisms of specific signatures.MethodsHere, we analysed a range of physiological and molecular responses expressed by geographically different populations of An. coluzzii inhabiting permanent and temporary breeding sites from the north and the south-west of Burkina Faso. Four mosquito colonies, namely (i) Oursi, built from females breeding in permanent habitats of the north; (ii) Déou, from temporary northern habitats; (iii) Soumousso from south-western temporary breeding sites; and (iv) Bama, from permanent habitats of the same south-western zone, were reared in climatic chambers under contrasted environmental conditions, mimicking temperature, relative humidity and light regimen occurring in northern Burkina Faso. Female mosquitoes were analysed for the seasonal variation in their amounts of proteins, triglycerides and free-circulating metabolites. The expression level of genes coding for the adipokinetic (AKH-I) and the AKH/corazonin-related peptides (ACP) were also assessed and compared among populations and environmental conditions.ResultsOur analysis did not reveal an apparent pattern of physiological and molecular variations strictly correlated with either the larval ecotype or the geographical origin of the mosquitoes. However, specific distinct responses were observed among populations, suggesting that dry season survival may rely on more complex ecological parameters at a micro-habitat scale. Interestingly, the physiological and molecular data support the hypothesis that different aestivation abilities exist among populations of An. coluzzii inhabiting contrasted ecological settings. In particular, the striking metabotypes differentiation and the AKH mRNA expression level observed in females from temporary northern populations may suggest the existence of a “strong” aestivation strategy in these specimens.ConclusionOur work provides insights into the physiological and molecular basis of dry and rainy season responses in An. coluzzii, and highlights the important diversity of the mechanisms involved. Such results represent key data for understanding the ecophysiological mechanisms underpinning the strong adaptive potential of this malaria vector species, which undoubtedly contributes to the spreading of mosquito distribution areas in space and time

Distinct physiological, biochemical and morphometric adjustments in the malaria vectors Anopheles gambiae and A. coluzzii as means to survive dry season conditions in Burkina Faso

International audienceAestivation and dispersive migration are the two strategies evoked in the literature to explain the way in which malaria vectorsandsurvive the harsh climatic conditions of the dry season in sub-Saharan Africa. However, the physiological mechanisms regulating these two strategies are unknown. In the present study, mosquito species were exposed to controlled environmental conditions mimicking the rainy and dry seasons of south western Burkina Faso. Survival strategies were studied through morphometric (wing length), ecophysiological (respiratory gas exchanges), biochemical (cuticular hydrocarbons composition) and molecular (AKH mRNA expression levels) parameters, variations of which are usually considered to be hallmarks of aestivation and dispersion mechanisms in various insects. Our results showed that ecophysiological and morphometric adjustments are made in both species to prevent water losses during the dry season. However, the usual metabolic rate modifications expected as signatures of aestivation and migration were not observed, highlighting specific and original physiological mechanisms sustaining survival in malaria mosquitoes during the dry season. Differences in epicuticular hydrocarbon composition and AKH levels of expression were found between the permanent and temporarypopulations, illustrating the great phenotypic plasticity of this mosquito species. Altogether, our work underlines the diverse and complex pattern of changes occurring in the two mosquito species and at the population level to cope with the dry season and highlights potential targets of future control tools

Additional file 1: Table S1. of Comparative physiological plasticity to desiccation in distinct populations of the malarial mosquito Anopheles coluzzii

Nucleotide sequences of the primers used in qRT-PCR reactions for the amplification of Actin, Rpl13, Rpl7, Rpl5, NADPH, hsp83, h3a, and 18s, AKH-I, ACP in An. gambiae. (DOCX 9 kb

Additional file 3: of Comparative physiological plasticity to desiccation in distinct populations of the malarial mosquito Anopheles coluzzii

Supplementary dataset 1. Raw data for protein, triglyceride and metabolomics assays. (XLSX 51 kb

Additional file 2: Table S2. of Comparative physiological plasticity to desiccation in distinct populations of the malarial mosquito Anopheles coluzzii

List of the 36 metabolites detected in females of Anopheles coluzzii. (DOCX 9 kb