Effect of the Insecticide Chlorpyrifos on Behavioral and Metabolic Aspects of the Spider Polybetes pythagoricus.

The toxicity of pesticides to organisms depends on the total amount of chemical exposure. Toxicity can be minimized if the organism recognizes the pesticide and alters its behavior. Furthermore, the physical barrier of cuticular hydrocarbons can prevent the entrance of the pesticide into the organism. Finally, if the pesticide enters the body, the organism experiences physiological changes favoring detoxification and the maintenance of homeostasis. We analyzed the behavioral and metabolic response of the spider Polybetes pythagoricus at different times of exposure to the organophosphate pesticide chlorpyrifos. First we observed that the individuals are capable of recognizing and avoiding surfaces treated with pesticides based on a behavioral analysis. Subsequently, we characterized cuticular hydrocarbons as a possible barrier against pesticides. Then we observed that the pesticide provoked histological damage, mainly at the level of the midgut diverticula. Finally, we analyzed the activity of several of the spider's enzymes linked to oxidative stress after exposure to chlorpyrifos for different lengths of time (6, 24, and 48 h). We observed that catalase activity was high at the start, whereas the activity of superoxide dismutase and glutathione S-transferase changed significantly at 48 h. Lipid peroxidation became high at 6 h, but decreased at 48 h. In conclusion, although P. pythagoricus can avoid contact with chlorpyrifos, this pesticide causes activation of the antioxidant system when it enters the body. Our results make a significant contribution to the ecotoxicology of spiders. Environ Toxicol Chem 2023;42:1293-1308. © 2023 SETAC.

Epicuticular hydrocarbons of the redbanded stink bug Piezodorus guildinii (Heteroptera: Pentatomidae): sexual dimorphism and alterations in insects collected in insecticide-treated soybean crops.

BACKGROUND: The redbanded stink bug Piezodorus guildinii (Heteroptera: Pentatomidae) is one of the most important species affecting soybean crops in southern South America. Capillary gas chromatography coupled to mass spectrometry was used to characterize the epicuticular hydrocarbon profiles of field-collected insects, and to identify differences in their composition between fifth-instar nymphs and adults, males and females, and between bugs collected in insecticide-treated and insecticide-free soybean crops. RESULTS: Straight chain saturated n-C27 and n-C29, and monomethyl and dimethyl chains of C31 and C33 were the most abundant compounds. A group of volatile hydrocarbons with n-C13 and n-C15 as the predominant compounds were also detected. The hydrocarbon pattern was different between nymphs and adults, either males or females. Heneicosene was almost exclusively detected in adult males and was the most important component to differentiate between both sexes, followed by tricosadiene. The total hydrocarbon amount was significantly higher in nymphs, males and females collected in insecticide-treated fields compared with insects obtained from untreated fields. CONCLUSION: Differences were found in the epicuticular hydrocarbon pattern among nymphs and adults, as well as sexual dimorphism in adult stink bugs. Interestingly, an alteration was also found in the hydrocarbon profile of insects collected in insecticide-treated soybean crops and its relevance is discussed within a pest management context.

Biochemical and morphological features of the uropygial gland of the Chilean Flamingo and their functional implications.

Flamingos inhabit specialized habitats and breed in large colonies, building their nests on islands that limit the access of terrestrial predators. Many aspects of their uropygial gland are still unknown. The uropygial gland, a sebaceous organ exclusive to birds, shares some histological features among species such as the presence of a capsule, adenomers with stratified epithelium and secondary and primary chambers. We found that the uropygial gland of the Chilean Flamingo (Phoenicopterus chilensis) displays most of these characteristics but lacks a primary storage chamber. This absence may be an adaptation to their aquatic environment. The uropygial secretion of this species has a variety of glycoconjugates while its lipid moiety is largely dominated by waxes and minor amounts of triacylglycerols and fatty acids. Mass spectrometry analysis of the preen wax showed branched fatty acids of varied chain length and unbranched fatty alcohols, resulting in a complex mixture of wax esters and no differences between sexes were observed. The glycoconjugates present in the preen secretion could play a role as antimicrobial molecules, as suggested for other bird species, while the absence of diester waxes in flamingos might be related with their nesting habits and limited exposure to predation. Our results were evaluated according to physiological and ecological aspects of the flamingo's biology.

Mosquitoes cloak their legs to resist insecticides.

Malaria incidence has halved since the year 2000, with 80% of the reduction attributable to the use of insecticides. However, insecticide resistance is now widespread, is rapidly increasing in spectrum and intensity across Africa, and may be contributing to the increase of malaria incidence in 2018. The role of detoxification enzymes and target site mutations has been documented in the major malaria vector Anopheles gambiae; however, the emergence of striking resistant phenotypes suggests the occurrence of additional mechanisms. By comparing legs, the most relevant insect tissue for insecticide uptake, we show that resistant mosquitoes largely remodel their leg cuticles via enhanced deposition of cuticular proteins and chitin, corroborating a leg-thickening phenotype. Moreover, we show that resistant female mosquitoes seal their leg cuticles with higher total and different relative amounts of cuticular hydrocarbons, compared with susceptible ones. The structural and functional alterations in Anopheles female mosquito legs are associated with a reduced uptake of insecticides, substantially contributing to the resistance phenotype.

Cytochrome P450 associated with insecticide resistance catalyzes cuticular hydrocarbon production in Anopheles gambiae.

The role of cuticle changes in insecticide resistance in the major malaria vector Anopheles gambiae was assessed. The rate of internalization of (14)C deltamethrin was significantly slower in a resistant strain than in a susceptible strain. Topical application of an acetone insecticide formulation to circumvent lipid-based uptake barriers decreased the resistance ratio by ∼50%. Cuticle analysis by electron microscopy and characterization of lipid extracts indicated that resistant mosquitoes had a thicker epicuticular layer and a significant increase in cuticular hydrocarbon (CHC) content (∼29%). However, the CHC profile and relative distribution were similar in resistant and susceptible insects. The cellular localization and in vitro activity of two P450 enzymes, CYP4G16 and CYP4G17, whose genes are frequently overexpressed in resistant Anopheles mosquitoes, were analyzed. These enzymes are potential orthologs of the CYP4G1/2 enzymes that catalyze the final step of CHC biosynthesis in Drosophila and Musca domestica, respectively. Immunostaining indicated that both CYP4G16 and CYP4G17 are highly abundant in oenocytes, the insect cell type thought to secrete hydrocarbons. However, an intriguing difference was indicated; CYP4G17 occurs throughout the cell, as expected for a microsomal P450, but CYP4G16 localizes to the periphery of the cell and lies on the cytoplasmic side of the cell membrane, a unique position for a P450 enzyme. CYP4G16 and CYP4G17 were functionally expressed in insect cells. CYP4G16 produced hydrocarbons from a C18 aldehyde substrate and thus has bona fide decarbonylase activity similar to that of dmCYP4G1/2. The data support the hypothesis that the coevolution of multiple mechanisms, including cuticular barriers, has occurred in highly pyrethroid-resistant An gambiae.

Biological Control of the Chagas Disease Vector Triatoma infestans with the Entomopathogenic Fungus Beauveria bassiana Combined with an Aggregation Cue: Field, Laboratory and Mathematical Modeling Assessment.

BACKGROUND: Current Chagas disease vector control strategies, based on chemical insecticide spraying, are growingly threatened by the emergence of pyrethroid-resistant Triatoma infestans populations in the Gran Chaco region of South America. METHODOLOGY AND FINDINGS: We have already shown that the entomopathogenic fungus Beauveria bassiana has the ability to breach the insect cuticle and is effective both against pyrethroid-susceptible and pyrethroid-resistant T. infestans, in laboratory as well as field assays. It is also known that T. infestans cuticle lipids play a major role as contact aggregation pheromones. We estimated the effectiveness of pheromone-based infection boxes containing B. bassiana spores to kill indoor bugs, and its effect on the vector population dynamics. Laboratory assays were performed to estimate the effect of fungal infection on female reproductive parameters. The effect of insect exuviae as an aggregation signal in the performance of the infection boxes was estimated both in the laboratory and in the field. We developed a stage-specific matrix model of T. infestans to describe the fungal infection effects on insect population dynamics, and to analyze the performance of the biopesticide device in vector biological control. CONCLUSIONS: The pheromone-containing infective box is a promising new tool against indoor populations of this Chagas disease vector, with the number of boxes per house being the main driver of the reduction of the total domestic bug population. This ecologically safe approach is the first proven alternative to chemical insecticides in the control of T. infestans. The advantageous reduction in vector population by delayed-action fungal biopesticides in a contained environment is here shown supported by mathematical modeling.

In vitro comparative analysis of antiproliferative activity of essential oil from mandarin peel and its principal component limonene.

The effects of the essential oil of mandarin peel (Corrientes, Argentina) and limonene (its major component) were studied on two human tumour cell lines growth (lung adenocarcinoma A549 and hepatocarcinoma HepG2). The essential oil was obtained by cold press and its composition was investigated by gas chromatography (GC) and GC/mass spectrometry (MS) analysis. The antiproliferative effect was studied using an MTT assay. Both mandarin essential oil and limonene tested showed a strong dose-dependent effect on the growth inhibition of these cell lines. The essential oil was more effective in A549 than in HepG2 cells and more effective than limonene in both the cases. It is likely that minor components and limonene of the oil could exert additive or synergistic effects. Hence, mandarin essential oil could lead to the development of anti-tumour agent or complementary and alternative medicines for the treatment of diverse cancers.

epicuticle lipids mediate mate recognition in Triatoma infestans.

Epicuticular lipids are contact cues in intraspecific chemical communication in insects, both for aggregation and sexual behavior. Triatomine bugs are vectors of the parasite Trypanosoma cruzi, the cause of Chagas disease. In Triatoma infestans, the major epicuticular lipids are hydrocarbons, fatty alcohols, and free and esterified fatty acids. Previously, we found that epicuticular lipid extracts, or selected fatty acid components, trigger aggregation and arrestment behavior in this bug. Using headspace solid phase microextraction, we found no sexual dimorphism in epicuticular hydrocarbons, but found female-specific fatty alcohols (eicosanol and docosanol). The role of epicuticular lipids in T. infestans copulation behavior was tested by observing male responses to live or various treatments of freeze-killed females. We report that hexane-soluble contact cues on females trigger copulation by males. Freeze-killed intact females were attractive to males, but no response was observed when males were exposed to hexane-washed females. Responses were partially recovered when epicuticular extract was applied to the dorsal surface of dead, hexane-washed females. One female equivalent of docosanol, evoked similar responses.

Volatile secretions and epicuticular hydrocarbons of the beetle Ulomoides dermestoides.

Many species of tenebrionids produce and secrete a defensive volatile blend containing mainly benzoquinones and alkenes. In this study we characterized the volatile organic compounds (VOC) of the beetle Ulomoides dermestoides (Coleoptera: Tenebrionidae). Solid phase microextraction (SPME) coupled to capillary gas chromatography-mass spectrometry (CGC-MS) analysis was used to identify methyl-1,4-benzoquinone (MBQ), ethyl-1,4-benzoquinone (EBQ), 1-tridecene (C(13:1)), and 1-pentadecene (C(15:1)), representing more than 90% of the volatile blend. We also used CGC-MS to analyze the epicuticular hydrocarbons of U. dermestoides. Saturated, unsaturated, and branched structures with chain lengths ranging from 13 to 43 carbons were detected. n-pentacosane (C(25:0)) and 9,11-pentacosadiene (9,11-C(25:2)) were the most abundant components, representing more than 40% of the cuticular hydrocarbons.

Control of pyrethroid-resistant Chagas disease vectors with entomopathogenic fungi.

BACKGROUND: Triatoma infestans-mediated transmission of Tripanosoma cruzi, the causative agent of Chagas disease, remains as a major health issue in southern South America. Key factors of T. infestans prevalence in specific areas of the geographic Gran Chaco region-which extends through northern Argentina, Bolivia, and Paraguay-are both recurrent reinfestations after insecticide spraying and emerging pyrethroid-resistance over the past ten years. Among alternative control tools, the pathogenicity of entomopathogenic fungi against triatomines is already known; furthermore, these fungi have the ability to fully degrade hydrocarbons from T. infestans cuticle and to utilize them as fuel and for incorporation into cellular components. METHODOLOGY AND FINDINGS: Here we provide evidence of resistance-related cuticle differences; capillary gas chromatography coupled to mass spectrometry analyses revealed that pyrethroid-resistant bugs have significantly larger amounts of surface hydrocarbons, peaking 56.2+/-6.4% higher than susceptible specimens. Also, a thicker cuticle was detected by scanning electron microscopy (32.1+/-5.9 microm and 17.8+/-5.4 microm for pyrethroid-resistant and pyrethroid-susceptible, respectively). In laboratory bioassays, we showed that the virulence of the entomopathogenic fungi Beauveria bassiana against T. infestans was significantly enhanced after fungal adaptation to grow on a medium containing insect-like hydrocarbons as the carbon source, regardless of bug susceptibility to pyrethroids. We designed an attraction-infection trap based on manipulating T. infestans behavior in order to facilitate close contact with B. bassiana. Field assays performed in rural village houses infested with pyrethroid-resistant insects showed 52.4% bug mortality. Using available mathematical models, we predicted that further fungal applications could eventually halt infection transmission. CONCLUSIONS: This low cost, low tech, ecologically friendly methodology could help in controlling the spread of pyrethroid-resistant bugs.

Epicuticular lipids induce aggregation in Chagas disease vectors.

BACKGROUND: The triatomine bugs are vectors of the protozoan parasite Trypanosoma cruzi, the causative agent of Chagas disease. Aggregation behavior plays an important role in their survival by facilitating the location of refuges and cohesion of aggregates, helping to keep them safely assembled into shelters during daylight time, when they are vulnerable to predators. There are evidences that aggregation is mediated by thigmotaxis, by volatile cues from their faeces, and by hexane-extractable contact chemoreceptive signals from their cuticle surface. The epicuticular lipids of Triatoma infestans include a complex mixture of hydrocarbons, free and esterified fatty acids, alcohols, and sterols. RESULTS: We analyzed the response of T. infestans fifth instar nymphs after exposure to different amounts either of total epicuticular lipid extracts or individual lipid fractions. Assays were performed in a circular arena, employing a binary choice test with filter papers acting as aggregation attractive sites; papers were either impregnated with a hexane-extract of the total lipids, or lipid fraction; or with the solvent. Insects were significantly aggregated around papers impregnated with the epicuticular lipid extracts. Among the lipid fractions separately tested, only the free fatty acid fraction promoted significant bug aggregation. We also investigated the response to different amounts of selected fatty acid components of this fraction; receptiveness varied with the fatty acid chain length. No response was elicited by hexadecanoic acid (C16:0), the major fatty acid component. Octadecanoic acid (C18:0) showed a significant assembling effect in the concentration range tested (0.1 to 2 insect equivalents). The very long chain hexacosanoic acid (C26:0) was significantly attractant at low doses (

Cuticular hydrocarbons of Chagas disease vectors in Mexico.

Capillary gas-liquid chromatography was used to analyse the cuticular hydrocarbons of three triatomine species, Triatoma dimidiata, T. barberi and Dipetalogaster maxima, domestic vectors of Chagas disease in Mexico. Mixtures of saturated hydrocarbons of straight and methyl-branched chains were characteristic of the three species, but quantitatively different. Major methylbranched components mostly corresponded to different saturated isomers of monomethyl, dimethyl and trimethyl branched hydrocarbons ranging from 29 to 39 carbon backbones. Sex-dependent, quantitative differences in certain hydrocarbons were apparent in T. dimidiata.

Cuticular hydrocarbons of Chagas disease vectors in Mexico

Capillary gas-liquid chromatography was used to analyse the cuticular hydrocarbons of three triatomine species, Triatoma dimidiata, T. barberi and Dipetalogaster maxima, domestic vectors of Chagas disease in Mexico. Mixtures of saturated hydrocarbons of straight and methyl-branched chains were characteristic of the three species, but quantitatively different. Major methylbranched components mostly corresponded to different saturated isomers of monomethyl, dimethyl and trimethyl branched hydrocarbons ranging from 29 to 39 carbon backbones. Sex-dependant, quantitative differences in certain hydrocarbons were apparent in T. dimidiata