Repellent activity of essential oils to the Lone Star tick, Amblyomma americanum.

BACKGROUND: The Lone Star tick, Amblyomma americanum is important to human health because of a variety of pathogenic organisms transmitted to humans during feeding events, which underscores the need to identify novel approaches to prevent tick bites. Thus, the goal of this study was to test natural and synthetic molecules for repellent activity against ticks in spatial, contact and human fingertip bioassays. METHODS: The efficacy of essential oils and naturally derived compounds as repellents to Am. americanum nymphs was compared in three different bioassays: contact, spatial and fingertip repellent bioassays. RESULTS: Concentration response curves after contact exposure to 1R-trans-chrysanthemic acid (TCA) indicated a 5.6 µg/cm2 concentration required to repel 50% of ticks (RC50), which was five- and sevenfold more active than DEET and nootkatone, respectively. For contact repellency, the rank order of repellency at 50 µg/cm2 for natural oils was clove > geranium > oregano > cedarwood > thyme > amyris > patchouli > citronella > juniper berry > peppermint > cassia. For spatial bioassays, TCA was approximately twofold more active than DEET and nootkatone at 50 µg/cm2 but was not significantly different at 10 µg/cm2. In spatial assays, thyme and cassia were the most active compounds tested with 100% and 80% ticks repelled within 15 min of exposure respectively and was approximately twofold more effective than DEET at the same concentration. To translate these non-host assays to efficacy when used on the human host, we quantified repellency using a finger-climbing assay. TCA, nootkatone and DEET were equally effective in the fingertip assay, and patchouli oil was the only natural oil that significantly repelled ticks. CONCLUSIONS: The differences in repellent potency based on the assay type suggests that the ability to discover active tick repellents suitable for development may be more complicated than with other arthropod species; furthermore, the field delivery mechanism must be considered early in development to ensure translation to field efficacy. TCA, which is naturally derived, is a promising candidate for a tick repellent that has comparable repellency to commercialized tick repellents.

Point Protection with Transfluthrin against Musca domestica L. in a Semi-Field Enclosure.

House flies are notoriously difficult to control, owing to their tendency to live in close relationships with humans and their livestock, and their rapid development of resistance to chemical controls. With this in mind, we explored an alternative chemical control, a spatial repellent to deter Musca domestica L. from points we wanted to protect (i.e., a baited trap). Our results demonstrated that the synthetic spatial repellent, transfluthrin, is effective in preventing M. domestica adults from entering protected traps for both a susceptible strain (CAR21) and a field-acquired permethrin-resistant strain (WHF; 24 h LD50 resistance ratio of 150), comprising 22% and 28% of the total number of flies collected, respectively. These results are promising and demonstrate that transfluthrin can be an effective spatial repellent to protect points of interest where needed.

Repellency and Toxicity of Vapor-Active Benzaldehydes against Aedes aegypti.

Chemical screening efforts recently found that 3-phenoxybenzaldehyde, a breakdown product of alpha-cyano pyrethroids, was a potent spatial repellent against Aedes aegypti mosquitoes in a glass tube repellency assay. In order to characterize this molecule further and identify structure-activity relationships, a set of 12 benzaldehyde analogues were screened for their repellency and toxicity in vapor phase exposures at 100 µg/cm2. Dose-response analyses were performed for the most active compounds in order to better characterize their repellent potency and toxicity compared to those of other commercially available toxicants. The three most toxic compounds (LC50 values) were 3-chlorobenzaldehyde (CBA) (37 µg/cm2), biphenyl-3-carboxaldehyde (BCA) (48 µg/cm2), and 3-vinylbenzaldehyde (66 µg/cm2), which makes them less toxic than bioallethrin (6.1 µg/cm2) but more toxic than sandalwood oil (77 µg/cm2), a repellent/toxic plant essential oil. The most repellent analogues with EC50 values below 30 µg/cm2 were 3-phenoxybenzaldehyde (6.3 µg/cm2), isophthalaldehyde (23 µg/cm2), BCA (17 µg/cm2), and CBA (22 µg/cm2), which makes them about as active as N,N-diethyl-3-methylbenzamide (25.4 µg/cm2). We further investigated the activity of a select group of these benzaldehydes to block the firing of the central nervous system of A. aegypti larvae. Compounds most capable of repelling and killing mosquitoes in the vapor phase were also those most capable of blocking nerve firing in the larval mosquito nervous system. The results demonstrate that benzaldehyde analogues are viable candidate repellent and insecticidal molecules and may lead to the development of future repellent and vapor toxic vector control tools.

Fir (Abies balsamea) (Pinales: Pinaceae) needle essential oil enhances the knockdown activity of select insecticides.

Because of the increased interest in plant essential oils (PEO) for both home pest control and personal bite protection, the ability of fir needle (Abies balsamea) oil to synergize the 1-h knockdown and 24-h toxicity of 9 different synthetic insecticides was evaluated. Fir needle oil strongly synergized knockdown of the neonicotinoids, clothianidin, and thiamethoxam (between 16- and 24-fold), as well as natural pyrethrins (12-fold), but had less effect with organophosphates and fipronil. For 24-h mortality, only pirimiphos-methyl was strongly synergized by fir needle oil pretreatment (18-fold). Chemical analysis and testing identified delta-3-carene is the most bioactive constituent, producing synergism similar to that of the whole oil. In fact, this constituent synergized the 24-h mortality of clothianidin to a higher degree than fir needle oil itself (4.9-fold vs. 2.4-fold). Synergism is unlikely to be mediated by effects on the nervous system, as fir needle oil caused no change in mosquito central nervous system firing at 100 ppm and did not synergize an inactive concentration of natural pyrethrins (10 nM). To better understand fir needle oil effects, we evaluated the ability of pretreatment with this oil to impact Aedes aegypti monooxygenase degradation of a model substrate, 7-ethoxycoumarin. Interestingly, both fir needle oil and delta-3-carene caused a significant increase in metabolic degradation of 7-ethoxycoumarin, perhaps indicating they upregulate oxidative metabolic processes. Such an action would explain why fir needle oil enhances knockdown, but not 24-h mortality for most of the insecticides studied here, whereas increased bioactivation would explain the synergism of pirimiphos-methyl toxicity.

Structural Exploration of Novel Pyrethroid Esters and Amides for Repellent and Insecticidal Activity against Mosquitoes.

The emergence of pyrethroid-resistant mosquitoes is a worldwide problem that necessitates further research into the development of new repellents and insecticides. This study explored the modification of existing pyrethroid acids to identify structural motifs that might not be affected by kdr active site mutations that elicit pyrethroid resistance. Because synthetic pyrethroids almost always contain activity-dependent chiral centers, we chose to focus our efforts on exploring alkoxy moieties of esters obtained with 1R-trans-permethrinic and related acids, which we showed in previous studies to have repellent and/or repellent synergistic properties. To this end, compounds were synthesized and screened for spatially acting repellency and insecticidal activity against the susceptible, Orlando, and pyrethroid-resistant, Puerto Rico, strains of Aedes aegypti mosquito. Screening utilized a high-throughput benchtop glass tube assay, and the compounds screened included a mixture of branched, unbranched, aliphatic, halogenated, cyclic, non-cyclic, and heteroatom-containing esters. Structure-activity relationships indicate that n-propyl, n-butyl, n-pentyl, cyclobutyl, and cyclopentyl substituents exhibited the most promising repellent activity with minimal kdr cross resistance. Preliminary testing showed that these small alcohol esters can be synergistic with phenyl amides and pyrethroid acids. Further derivatization of pyrethroid acids offer an interesting route to future active compounds, and while mosquitoes were the focus of this work, pyrethroid acids and esters have potential for use in reducing pest populations and damage in cropping systems as well.

Insecticidal and Synergistic Potential of Three Monoterpenoids against the Yellow Fever Mosquito, Aedes aegypti (Diptera: Culicidae), and the House Fly, Musca domestica (Diptera: Muscidae).

As resistance to the limited number of insecticides available for medical and veterinary pests becomes more widespread, there is an urgent need for new insecticides and synergists on the market. To address this need, we conducted a study to assess the toxicity of three monoterpenoids-carvone, menthone, and fenchone-in comparison to permethrin and methomyl against adults of two common pests: the yellow fever mosquito (Aedes aegypti) and the house fly (Musca domestica). We also examined the potential for these monoterpenoids to enhance the effectiveness of permethrin and methomyl when used together. Finally, we evaluated the ability of each monoterpenoid to inhibit acetylcholinesterase, comparing them to methomyl. While all three monoterpenoids performed relatively poorly as topical insecticides (LD50 > 4000 ng/mg on M. domestica; >6000 ng/mg on Ae. aegypti), they synergized both permethrin and methomyl as well as or better than piperonyl butoxide (PBO). Carvone and menthone yielded synergistic co-toxicity factors (23 and 29, respectively), which were each higher than PBO at 24 h. Currently, the mechanism of action is unknown. During preliminary testing, symptoms of acetylcholinesterase inhibition were identified, prompting further testing. Acetylcholinesterase inhibition did not appear to explain the toxic or synergistic effects of the three monoterpenoids, with IC50 values greater than 1 mM for all, compared to the 2.5 and 1.7 µM for methomyl on Aedes aegypti and Musca domestica, respectively. This study provides valuable monoterpenoid toxicity and synergism data on two pestiferous insects and highlights the potential for these chemistries in future pest control formulations.

Sodium channel-directed alkaloids synergize the mosquitocidal and neurophysiological effects of natural pyrethrins.

We explored the potential of two sodium channel activators, veratrine and aconitine, as both insecticides and synergists of natural pyrethrins (NP) on Aedes aegypti adults and larvae. Aconitine was more toxic than veratrine, with an LD50 of 157 ng/mg compared to 376 ng/mg, on the pyrethroid-susceptible Orlando strain, but only aconitine showed significant resistance in the pyrethroid-resistant Puerto Rico strain (RR = 14.6 in topical application and 8.8 in larval bioassay). When applied in mixtures with piperonyl butoxide (PBO) and NP, large synergism values were obtained on the Orlando strain. Aconitine + PBO mixture synergized NP 21.8-fold via topical adult application and 10.2-fold in larval bioassays, whereas veratrine + PBO synergized NP 5.3-fold via topical application and 30.5-fold in larval bioassays. Less synergism of NP was observed on the resistant Puerto Rico strain, with acontine + PBO synergizing NP only 4.1-fold in topical application (8-fold in larval bioassays) and veratrine + PBO synergizing NP 9.5-fold in topical application (13.3-fold in larval bioassays). When alkaloids were applied directly to the mosquito larval nervous system, veratrine was nearly equipotent on both strains, while aconitine was less active on pyrethroid-resistant nerve preparations (no block at 10 µM compared to block at 1 µM on the susceptible strain). The nerve blocking effect of NP was significantly synergized by both compounds on the pyrethroid-susceptible strain by about 10-fold, however only veratrine synergized NP block on the pyrethroid-resistant strain, also showing 10-fold synergism). These results highlight the potential of site II sodium channel activators as insecticides and their ability to synergize pyrethroids, which may extend the commercial lifetime of these chemistries so essential to public health vector control.

Mode of action and toxicological effects of the sesquiterpenoid, nootkatone, in insects.

Nootkatone, a sesquiterpenoid isolated from Alaskan yellow cedar (Cupressus nootkatensis), is known to possess insect repellent and acaricidal properties and has recently been registered for commercial use by the Environmental Protection Agency. Previous studies failed to elucidate the mechanism of action of nootkatone, but we found a molecular overlay of picrotoxinin and nootkatone indicated a high degree of structural and electrostatic similarity. We therefore tested the hypothesis that nootkatone was a GABA-gated chloride channel antagonist, similar to picrotoxinin. The KD50 and LD50 of nootkatone on the insecticide-susceptible strain of Drosophila melanogaster (CSOR) showed resistance ratios of 8 and 11, respectively, compared to the cyclodiene-resistant strain of RDL1675, indicating significant cross-resistance. Nootkatone reversed GABA-mediated block of the larval CSOR central nervous system; nerve firing of 78 ± 17% of baseline in the CSOR strain was significantly different from 24 ± 11% of baseline firing in the RDL1675 strain (p = 0.035). This finding indicated that the resistance was expressed within the nervous system. Patch clamp recordings on D. melanogaster central neurons mirrored extracellular recordings where nootkatone inhibited GABA-stimulated currents by 44 ± 9% at 100 µM, whereas chloride current was inhibited 4.5-fold less at 100 µM in RDL1675. Taken together, these data suggest nootkatone toxicity in D. melanogaster is mediated through GABA receptor antagonism.

Nutritional status significantly affects toxicological endpoints in the CDC bottle bioassay.

BACKGROUND: The CDC Bottle Bioassay serves as an inexpensive and effective way to screen field-caught mosquitoes against a wide variety of insecticidal active ingredients and commercial formulations, with the goal of detecting resistant individuals. For this study, we assessed how sucrose-water (10% w/v) feeding status impacted the response of Aedes aegypti mosquitoes to select insecticides. RESULTS: Starvation for 24 or 48 h decreased permethrin and malathion mean survival time by about 40%, with little difference in the two starvation times. Similar findings were also observed in a pyrethroid-resistant Puerto Rico strain challenged with permethrin, but these effects were less pronounced. To test the impact of mosquito weight, we measured weight under different 48-h nutritional conditions and found that sugar-water-fed and sugar-only-fed individuals were approximately the same weight (ANOVA, Bonferroni post-test P value >0.999) and that individuals fed water only were considerably lighter than either sugar-water-fed (ANOVA, Bonferroni post-test P value = 0.034) or sugar-only-fed individuals (ANOVA, Bonferroni post-test P value = 0.027) but equal in weight to starved mosquitoes (ANOVA, Bonferroni post-test P value >0.99). Of the nutritional challenges, water-only-fed mosquitoes were the most insecticide tolerant (ANOVA, Bonferroni post-test P values to all other treatments <0.01). CONCLUSIONS: The results indicate insect nutritional status is an important experimental variable, particularly the hydration status of mosquitoes shortly before insecticide exposure. Moreover, as significant differences were observed between starved and component-fed (water, sugar, or sugar and water) mosquitoes in a pyrethroid-resistant strain, some caution is appropriate when interpreting resistance/susceptibility diagnoses with this bioassay. © 2021 Society of Chemical Industry. This article has been contributed to by US Government employees and their work is in the public domain in the USA.

Recording central neurophysiological output from mosquito larvae for neuropharmacological and insecticide resistance studies.

Resistance to currently utilized chemical insecticidal agents represents a significant threat to public health and food security worldwide. Better understanding the neurophysiological effects of available and candidate insecticidal molecules is valuable for characterizing the mechanisms of insecticide resistance, as well as the design and study of novel control chemistries. In this paper, we describe a method of recording nerve firing from the central nervous system of Aedes aegypti fourth instar larvae. In short, mosquito larvae were immobilized by placing small pins through the head and siphon of the larvae in a wax dish, ventral side down. A single, longitudinal, dorsal incision from the distal abdomen to the pronotum of the larva was made, the alimentary canal removed, and the ventral nerve cord severed between the second and third abdominal ganglia. A recording suction electrode was connected directly to axons within the severed end of the connective in a novel way to record nerve firing in the ventral nerve cord at a high signal-to-noise ratio with conventional electrophysiological equipment. Using this novel method, we report the effects of four neuroactive compounds using this method: octopamine, pilocarpine, nicotine, and γ-aminobutyric acid (GABA). The utility of this recording technique for elucidating target site mechanisms involved in insecticide resistance is demonstrated with p,p'-dichlorodiphenyltrichlorethane (DDT) and its difluoro analog (DFDT).

Co-Toxicity Factor Analysis Reveals Numerous Plant Essential Oils Are Synergists of Natural Pyrethrins against Aedes aegypti Mosquitoes.

With insecticide-resistant mosquito populations becoming an ever-growing concern, new vector control technologies are needed. With the lack of new chemical classes of insecticides to control mosquito populations, the development of novel synergists may improve the performance of available insecticides. We screened a set of 20 plant essential oils alone and in combination with natural pyrethrins against Aedes aegypti (Orlando) female adult mosquitoes to assess their ability to synergize this natural insecticide. A co-toxicity factor analysis was used to identify whether plant oils modulated the toxicity of natural pyrethrins antagonistically, additively, or synergistically. Both knockdown at 1 h and mortality at 24 h were monitored. A majority of oils increased the toxicity of natural pyrethrins, either via an additive or synergistic profile. Many oils produced synergism at 2 µg/insect, whereas others were synergistic only at the higher dose of 10 µg/insect. Amyris, cardamom, cedarwood, and nutmeg East Indies (E.I.) oils were the most active oils for increasing the mortality of natural pyrethrins at 24 h with co-toxicity factors greater than 50 at either or both doses. A number of oils also synergized the 1 h knockdown of natural pyrethrins. Of these, fir needle oil and cypress oils were the most successful at improving the speed-of-action of natural pyrethrins at both doses, with co-toxicity factors of 130 and 62, respectively. To further assess the co-toxicity factor method, we applied selected plant essential oils with variable doses of natural pyrethrins to calculate synergism ratios. Only the oils that produced synergistic co-toxicity factors produced statistically significant synergism ratios. This analysis demonstrated that the degree of co-toxicity factor correlated well with the degree of synergism ratio observed (Pearson correlation coefficient r = 0.94 at 2 µg/insect; r = 0.64 at 10 µg/insect) and that the co-toxicity factor is a useful tool in screening for synergistic activity.

Vapor phase repellency and insecticidal activity of pyridinyl amides against anopheline mosquitoes.

It is important to identify repellents that can provide reliable protection from arthropod biting and prevent arthropod-borne diseases, such as malaria. In the present study, the spatial repellent activity and toxicity of two novel pyridinyl amides (1 and 2) were evaluated against Anopheles albimanus, Anopheles quadrimaculatus, and Anopheles gambiae. In vapor repellency bioassays, compound 2 was generally more effective than DEET and 2-undecanone, while compound 1 was about as active as these standards. Overall, transfluthrin was the most active compound for inducing anopheline mosquito repellency, knockdown, and lethality. Although they were not the most active repellents, the two experimental amides produced the largest electroantennographic responses in female antennae. They also displayed modest toxicity to anopheline mosquitoes. Significant synergism of repellency was observed for the mixture of a pyrethroid-derived acid and the repellent 2-undecanone against anopheline mosquitoes, similar to that observed previously in Aedes aegypti. Overall, this study provides insight for further synthesis of alternative amide compounds for use as spatial treatments.

Bioactivities and modes of action of VUAA1.

BACKGROUND: The compound 2-((4-ethyl-5-(pyridin-3-yl)-4H-1,2,4-triazol-3-yl)thio)-N-(4-ethylphenyl) acetamide (VUAA1) is reported to be an odorant receptor co-receptor (Orco) agonist in insects with potential use as an insect repellent. For this study, the biological activity of VUAA1 was investigated in several bioassays with Aedes aegypti, including adult contact, spatial repellency, and larval repellency assays, as well as topical, injection, and feeding toxicity assays. Neurophysiological action was further explored by analysis of fruit fly central nervous system firing, cockroach axon recordings, patch clamp analysis of Kv2 potassium channel, and acetylcholinesterase inhibition studies. Finally, the metabolic impact on the toxicity of VUAA1 was explored by applying it in combination with established metabolic synergists. RESULTS: In repellency and bite protection screens, VUAA1 showed little activity against adult mosquitoes, apparently due to its low volatility, since its effectiveness was increased by heating or mixing with transfluthrin acid and citronella oil. It did produce measurable repellency of mosquito larvae that was more potent than N,N-diethyl-m-toluamide (DEET). Overall, VUAA1 showed low acute toxicity to both insects and mice, and it was weakly synergized by triphenyl phosphate. There was no observed cross-resistance in a pyrethroid-resistant strain of Anopheles gambiae. VUAA1 showed a two-phase effect on the central nervous system, with neuroexcitation at 1 µmol L-1 and an inhibitory effect at 100 µmol L-1 that may relate to block of Kv2 potassium channels. CONCLUSIONS: VUAA1 presented low toxicity, similar to other insect repellents. Its limited solubility, low volatility, and resulting poor adult repellency without additional adjuvants may restrict the utility of VUAA1 in typical public health applications. © 2020 Society of Chemical Industry.

Structure-Activity Relationship Analysis of Potential New Vapor-Active Insect Repellents.

A total of 115 aryl amides were synthesized and screened for vapor repellency against the Orlando (OR) strain of Aedes aegypti mosquitoes. Of these compounds, 29 had 1 h repellency EC50 values comparable to or better than N,N-diethyl-meta-toluamide (DEET, 1 h EC50 value of 35 µg/cm2), with 2,2,3,3,3-pentafluoro-N-(4-fluorophenyl)propenamide (53) and 2,2,3,3,4,4,4-heptafluoro-N-(3,4,5-trifluorophenyl)butanamide (101) exhibiting the most potent EC50 values of 4.5 and 2.9 µg/cm2, respectively. The cross-resistance of select, highly potent, derivatives against the pyrethroid-resistant Puerto Rico (PR) strain of A. aegypti was also investigated, and little to no resistance was observed. When synergized with 1R-trans-permethrinic acid (TFA), compound 101 had a 1 h EC50 value 6 times lower than metofluthrin against OR and 40 times lower against PR mosquitoes. Additionally, preliminary mammalian oral toxicity was screened for compounds 69 and 101, and both exhibited LD50 values of >2000 mg/kg. The structure-activity relationship analysis, which guided the synthesis of these derivatives, is given, and key trends are highlighted to inform future analogue design.

Structure and Properties of Polyamide Fabrics with Insect-Repellent Functionality by Electrospinning and Oxygen Plasma-Treated Surface Coating.

The need for light-weight and high-strength insect-repellant fabrics is of critical importance to the cessation of viral diseases. The goal of the study is to investigate the structure and properties of insect-repellent polyamide fabrics for use in protective garments to guard against mosquitos. Permethrin was applied to the polyamide fabrics through incorporation into the nylon 6 polymer solution during electrospinning and dip coating onto the control untreated and oxygen plasma-treated polyamide fabrics: electropun nylon 6 nanofiber nonwovens, commercially available nylon 6 warp knit tricot, and nylon 66 double weft, knit interlock fabrics. The incorporation of permethrin into the polymer solution before the formation of fibers demonstrated the most efficient way to apply permethrin to the fiber/fabric systems. The plasma treatment significantly increased the amount of permethrin on the surface of the fabrics. All permethrin-containing polyamide fabrics showed excellent fastness of the insecticide to light. The electrospun nylon 6 nonwovens demonstrated the best fastness to washing among the plasma-treated electrospun nylon 6, nylon 66 double weft knit, and nylon 6 warp-knit tricot. All permethrin-treated fabrics were repellent and caused higher percentage of mosquito escape compared to the control untreated fabrics.

Mosquitocidal activity of p,p'-difluoro-diphenyl-trichloroethane (DFDT).

New insecticides are urgently needed for the control of arthropod vectors of public health diseases. As resistance to many insecticides used for the control of public health pests is ubiquitous, all available chemistries should be evaluated for their potential to effectively control both insecticide-susceptible and insecticide-resistant strains of mosquitoes. This study aimed to evaluate p-p'-difluoro-diphenyl-trichloroethane (DFDT) as a mosquito control technology and relate its activity to that of DDT. We found that topical DFDT was significantly less toxic than DDT to both pyrethroid-susceptible and pyrethroid-resistant strains of Anopheles gambiae and Aedes aegypti. Direct nervous system recording from Drosophila melanogaster CNS demonstrated that DFDT is approximately 10-times less potent than DDT at blocking nerve firing, which may explain its relatively lower toxicity. DFDT was shown to be at least 4500 times more vapor-active than DDT, with an LC50 in a vapor toxicity screening assay of 2.2 µg/cm2. Resistance to DFDT was assessed in two mosquito strains that possess target-site mutations in the voltage-gated sodium channel and upregulated metabolic activity. Resistance ratios for Akdr (An. gambiae) and Puerto Rico (Ae. aegypti) strains were 9.2 and 12.2, respectively. Overall, this study demonstrates that DFDT is unlikely to be a viable public health vector control insecticide.

Biodistribution of degradable polyanhydride particles in Aedes aegypti tissues.

Insecticide resistance poses a significant threat to the control of arthropods that transmit disease agents. Nanoparticle carriers offer exciting opportunities to expand the armamentarium of insecticides available for public health and other pests. Most chemical insecticides are delivered by contact or feeding, and from there must penetrate various biological membranes to reach target organs and kill the pest organism. Nanoparticles have been shown to improve bioactive compound navigation of such barriers in vertebrates, but have not been well-explored in arthropods. In this study, we explored the potential of polyanhydride micro- and nanoparticles (250 nm- 3 µm), labeled with rhodamine B to associate with and/or transit across insect biological barriers, including the cuticle, epithelium, midgut and ovaries, in female Ae. aeygpti mosquitoes. Mosquitoes were exposed using conditions to mimic surface contact with a residual spray or paint, topical exposure to mimic contact with aerosolized insecticide, or per os in a sugar meal. In surface contact experiments, microparticles were sometimes observed in association with the exterior of the insect cuticle. Nanoparticles were more uniformly distributed across exterior tissues and present at higher concentrations. Furthermore, by surface contact, topical exposure, or per os, particles were detected in internal organs. In every experiment, amphiphilic polyanhydride nanoparticles associated with internal tissues to a higher degree than hydrophobic nanoparticles. In vitro, nanoparticles associated with Aedes aegypti Aag2 cells within two hours of exposure, and particles were evident in the cytoplasm. Further studies demonstrated that particle uptake is dependent on caveolae-mediated endocytosis. The propensity of these nanoparticles to cross biological barriers including the cuticle, to localize in target tissue sites of interest, and to reach the cytoplasm of cells, provides great promise for targeted delivery of insecticidal candidates that cannot otherwise reach these cellular and subcellular locations.

Evaluation of pyrethroids and organophosphates in insecticide mixtures for management of western corn rootworm larvae.

BACKGROUND: The western corn rootworm is an economically important pest of corn. Management tactics include pyrethroid and organophosphate insecticides, which may be applied as a mixture to protect corn roots. The goal of our study was to characterize the effects of pyrethroids and organophosphates alone and in combination on larval corn rootworm mortality and injury to corn roots. We evaluated two insecticide combinations: tebupirimphos with ß-cyfluthrin and chlorethoxyfos with bifenthrin. Using a soil-based, laboratory bioassay, we exposed larvae to five concentrations of the pyrethroid alone, the organophosphate alone, the combined formulation, and a water control. We calculated LC50 values and co-toxicity factors to determine synergism or antagonism between organophosphates and pyrethroids. We also measured adult emergence and root injury in a field experiment that tested tebupirimphos alone, ß-cyfluthrin alone, the combined formulation, and an untreated control. RESULTS: Bioassay results indicated antagonism between the pyrethroid and organophosphate at most concentrations for both insecticide combinations. In the field experiment, tebupirimphos alone or in combination with ß-cyfluthrin significantly reduced adult emergence and root injury compared to the untreated controls, but ß-cyfluthrin alone did not differ from the untreated control for either metric. CONCLUSIONS: These results suggest that, at the concentrations tested, the pyrethroid component of pyrethroid-organophosphate mixtures may not contribute to a reduction of rootworm emergence or root injury. While these pyrethroids may confer a management benefit for other pests, such as seedcorn maggot, the concentrations of pyrethroids present in current formulations of these mixtures are likely too low for effective rootworm management. © 2020 Society of Chemical Industry.

Pyrethroid-Derived Acids and Alcohols: Bioactivity and Synergistic Effects on Mosquito Repellency and Toxicity.

Pyrethroids are one of the most commonly used classes of insecticides, and their acid and alcohol components are esterase degradation products, usually considered to be biologically inactive. In this study, it was found that several pyrethroid acids had a spatial repellent activity that was greater than DEET, often more active than the parent pyrethroids, and showed little cross resistance in a pyrethroid-resistant Puerto Rico strain of Aedes aegypti mosquitoes. Further investigation revealed that the acids can synergize not only contact repellent standards but also other pyrethroid components as well as the parent pyrethroids themselves. Synergism by the pyrethroid acids is expressed as both increased spatial repellency and vapor toxicity as well as human bite protection. Electrophysiological studies confirmed that pyrethroid acids (100 µM) had no effect on neuronal discharge in larval Drosophila melanogaster CNS and were detected by electroantennography, and there was little resistance to olfactory sensing of these acids in antennae from Puerto Rico strain mosquitoes carrying kdr mutations. Thus, the data suggest that the pyrethroid acids have a different mode of action than the parent pyrethroids, unrelated to the voltage-sensitive sodium channel. The results highlight the potential of pyrethroid acids to be useful in future repellent formulations.

Reduced effectiveness of repellents in a pyrethroid-resistant strain of Aedes aegypti (Diptera: culicidae) and its correlation with olfactory sensitivity.

BACKGROUND: The mosquito, Aedes aegypti (Diptera: Culicidae), is a vector of dengue fever, zika, chikungunya, and yellow fever viruses, and in many areas possesses significant levels of resistance to pyrethroids. Behavioral performance was assessed in 15, 30, and 60 min exposures in a high throughput vapor phase spatial repellency assay to three contact repellent standards: N,N-diethyl-3-methylbenzamide (DEET), ethyl 3-[acetyl(butyl)amino] propanoate (IR3535), and 2-undecanone, as well as pyrethrum extract, transfluthrin, and metofluthrin in susceptible (Orlando) and a pyrethroid-resistant Puerto Rico strain of Aedes aegypti. Additionally, electroantennographic studies were used to investigate the antennal sensitivities to these compounds in both strains. RESULTS: Resistance was found to all tested insect repellents in the Puerto Rico strain of Ae. aegypti. Resistance ratios at the different time points were about 2 for DEET, 3 for 2-undecanone, and 12 for IR3535. Resistance was also observed to pyrethrum extract (∼9-fold), transfluthrin (∼5-fold), and metofluthrin (∼48-fold) in repellent behavioral response. Electrophysiological analysis found decreased antennal sensitivity to all repellents tested, consistent with their behavioral effects. CONCLUSION: The reduced sensitivity to these repellents may represent a fitness cost arising from the kdr mutation present in Puerto Rico Aedes aegypti. This work highlights the need for understanding collateral effects from the evolution of pesticide resistance in mosquitoes, and the importance of finding alternative strategies to control resistance development. © 2019 Society of Chemical Industry.