Beauveria bassiana Culturing and Harvesting for Bioassays With House Flies.

The entomopathogenic fungus Beauveria bassiana (Balsamo) Vuillemin (Hypocreales: Cordycipitaceae) has been widely studied against a wide range of arthropod pests, including many of medical and veterinary importance. New investigators must sort through a wide array of published methods for the production, harvest, storage, and bioassay methods for this pathogen. Simplified methods for production of conidia using Sabouraud dextrose agar with yeast (SDYA) plates and two conidial harvesting methods are described. Dry harvesting yields conidia that are ready to incorporate into dusts and food baits, but the fungal product includes mycelial debris that can hamper quantification and introduces variable amounts of unwanted bulk. Wet harvesting with filtration produces a cleaner product that is immediately ready for testing in liquid formulations. Examples of bioassays with house flies are presented that include conidia applied topically to the dorsal thorax for dose-mortality assays and conidial suspensions applied to filter paper disks for concentration mortality assays.

Nontarget effects of aerial mosquito adulticiding with water-based unsynergized pyrethroids on honey bees and other beneficial insects in an agricultural ecosystem of north Greece.

We assessed the nontarget effects of ultra-low-volume (ULV) aerial adulticiding with two new water-based, unsynergized pyrethroid formulations, Aqua-K-Othrine (FFAST antievaporant technology, 2% deltamethrin) and Pesguard S102 (10% d-phenothrin). A helicopter with GPS navigation technology was used. One application rate was tested per formulation that corresponded to 1.00 g (AI)/ha of deltamethrin and 7.50 g (AI)/ha of d-phenothrin. Three beneficial nontarget organisms were used: honey bees (domesticated hives), family Apidae (Apis mellifera L.); mealybug destroyers, family Coccinellidae (Cryptolaemus montrouzieri Mulsant); and green lacewings, family Chrysopidae (Chrysoperla carnea (Stephens)). No significant nontarget mortalities were observed. No bees exhibited signs of sublethal exposure to insecticides. Beehives exposed to the insecticidal applications remained healthy and productive, performed as well as the control hives and increased in weight (25-30%), in adult bee population (14-18%), and in brood population (15-19%).

Droplet sampling of an oil-based and two water-based antievaporant ultra-low volume insecticide formulations using Teflon- and magnesium oxide-coated slides.

We estimated the diameters below which 50% and 90% of the volume of droplets exist (Dv50 and Dv90, respectively) of 1 oil-based (Permanone 30-30) and 2 water-based (AquaReslin, Aqua-K-Othrine) antievaporant aerosols (with the Film Forming Aqueous Spray Technology [FFAST]) using Teflon- and magnesium oxide (MgO)-coated slides and determined whether the aging of the droplets on the slides (up to 60 min) exhibited any significant effect on Dv50 and Dv90 calculations. There were no significant differences in either Dv50 or Dv90 estimates on MgO-coated slides at 0 min and 60 min for all 3 products tested. On Teflon-coated slides, the only product that showed significant difference between 0 min and 60 min in both Dv50 and Dv90 estimates was Aqua-K-Othrine, perhaps due to a difference in formulation components. Specifically, both values corresponding to Dv50 and Dv90 at 60 min decreased by approximately 50% when compared to the values at 0 min. For the other 2 products, AquaReslin and Permanone, aging of droplets on Teflon up to 60 min did not have any significant effect on Dv50 and Dv90 values. To further investigate the behavior of Aqua-K-Othrine droplets on Teflon-coated slides we observed the droplets immediately after spraying and at 10-min intervals under different conditions of temperature and humidity. The majority of the shrinkage occurred within the 1st 10 min after impaction on the slides under all conditions tested. So in most field situations where slides are read several hours or days after collection, this shrinkage would not be observed. The MgO-coated slides should be the preferred field method for sampling droplets of Aqua-K-Othirne with the FFAST antievaporant technology.

Kneallhazia carolinensae sp. nov., a microsporidian pathogen of the thief ant, Solenopsis carolinensis.

A new species of microsporidia is described from adults of the thief ant, Solenopsis carolinensis, collected in Florida, USA. Morphological and genetic characterization of this new species showed that it is most closely related to the genus Kneallhazia and is therefore formally designated, Kneallhazia carolinensae sp. nov. Masses of ovoid, binucleate spores were localized to fat body of adult workers and measured 6.2±0.1×3.1±0.1 µm (fresh) and 6.0±0.1×3.4±0.1 µm (fixed). These spores were in direct contact with the cell cytoplasm and contained an isofilar polar filament with 12-15 coils. Blastn analysis revealed that the K. carolinensae 16S rDNA sequence exhibited 91% identity with the 16S rDNA gene of K. solenopsae. The morphological and sequence data support the conclusion that K. carolinensae is a novel microsporidian species distinct from K. solenopsae.

Efficacy of aerial ultra-low volume applications of two novel water-based formulations of unsynergized pyrethroids against riceland mosquitoes in Greece.

We assessed the efficacy of ultra-low volume aerial adulticiding with 2 new water-based, unsynergized formulations of Aqua-K-Othrin (2% deltamethrin) and Pesguard S102 (10% d-phenothrin) against the riceland mosquitoes of Greece. A helicopter with Global Positioning System (GPS) navigation, real-time weather recording, and spray dispersal modeling (AgDISP) was utilized to accurately treat the experimental blocks by adjusting spray line positions to changing meteorological conditions. Two application rates were applied per formulation that corresponded to 0.75 and 1.00 g AI/ha of deltamethrin and 7.50 and 10.00 g AI/ha of d-phenothrin. The mosquitoes used for the trials were the main nuisance species found in rice field areas of Thessaloniki, which were primarily Aedes caspius, followed by Culex modestus and Anopheles sacharovi. Overall mean mortality of caged mosquitoes was 69.2% and 64.8% for deltamethrin and d-phenothrin, respectively. Mean population decrease in wild mosquito populations within the treatment areas was 76.5% and 78% for deltamethrin and d-phenothrin, respectively. The AgDISP dispersal model, coupled with GPS navigation and real-time weather recording, enabled accurate placement of the spray cloud such that the majority of the treatment area received sufficiently high droplet densities to result in uniform caged-mosquito mortality across all sampling sites.

Effect of population structure and size on aggregation behavior of Cimex lectularius (Hemiptera: Cimicidae).

The bed bug, Cimex lectularius L. (Hemiptera: Cimicidae), occurs in aggregations until the conditions are no longer beneficial, leading to dispersal. Active and passive bed bug dispersal causes migrations from main aggregations either within a room, from room to room within a building, or from building to building. Because bed bug movement is an important factor in the spread of infestations, we wanted to determine how population structure and size affect bed bug aggregations. Engorged bed bugs were placed in glass petri dish arenas at varying densities, sex ratios, and population compositions. Nymphs had a high tendency to aggregate, varying between 94 and 98%, and therefore were not the likely dispersal stage of the bed bug. At densities of 10 and 40 adults at a 1:1 sex ratio, there were significantly more lone females than lone males. When the population composition was varied, the percentage of lone females was significantly higher than that of males and nymphs at population compositions of 40 and 80% adults. When the sex ratio of adults was varied, there were significantly more lone females than males in arenas with 20, 50, and 80% males. Females, being found away from aggregations significantly more often than any other life stage, are potentially the dispersal stage of the bed bug. Active female dispersal away from main aggregations can potentially lead to treatment failures and should be taken into account when using control methods.

Toxicities of 31 volatile low molecular weight compounds against Aedes aegypti and Culex quinquefasciatus.

This research studied 31 volatile compounds for indoor control of the medically important mosquitoes Aedes aegypti L. and Culex quinquefasciatus Say. Only adult female mosquitoes were tested. The test compounds were from six families that included five heterobicyclics, eight formate esters, formic acid (a hydrolyzed metabolite of formate esters), eight acetate esters, four propionate esters, three butyrate esters, and two valerate esters. Also, the organophosphate compound dichlorvos (DDVP) was tested as a positive control. Cx. quinquefasciatus was generally more susceptible than Ae. aegypti. Cx. quinquefasciatus was most susceptible to a subset of heterobicyclics and formate esters (rank: n-butyl formate > hexyl formate = dihydrobenzofuran = menthofuran = heptyl formate = ethyl formate). Ae. aegypti was most susceptible to a subset of formate esters (rank: methyl > n-butyl > propyl = ethyl = hexyl). The most active materials against both species had LC50s of 0.4-1 mg active ingredient per 0.5 liter of air volume (0.8-2 mg/liter), which is 50- to 60-fold less toxic than dichlorvos (an organophosphate insecticide that is being phased out from indoor use). In relation to Drosophila melanogaster Meigen, both mosquito species were generally more susceptible to formate esters but more tolerant of heterobicyclics. Generally, the most toxic compound against all dipterans tested to date is n-butyl formate, whereas menthofuran is additionally toxic against Cx. quinquefasciatus and D. melanogaster. Finally, the toxicity differences between species point to the potential for differential toxicity among mosquito general/species, suggesting that further studies of a number of mosquito species might be warranted.

Vapor toxicity of three prototype volatile insecticidal compounds to house fly (Diptera: Muscidae).

Vapor toxicities of two prototype formate esters, ethylene glycol di-formate (EGDF) and heptyl formate, and the prototype heterobicyclic menthofuran were determined against laboratory-reared adult house flies, Musca domestica L. (Diptera: Muscidae). Toxicities were compared between house flies and other dipteran species tested previously. The organophosphate fumigant insecticide dichlorvos (DDVP) was used as a positive control. Although less toxic than DDVP, all three prototype compounds were toxic to house flies. Most toxic was menthofuran (LC50 = 3.70 mg/liter air), followed by EGDF (LC50 = 9.27 mg/liter air) and heptyl formate (LC50 = 32.62 mg/liter air). The toxicity of menthofuran when applied after piperonyl butoxide increased 1.5-fold. The toxicities of heptyl formate and EGDF when applied after S,S,S-tributyl phosphorotrithioate decreased by 1.5- and 2-folds, respectively. These results are slightly different from results obtained for other dipterans, further supporting previous hypotheses that toxicity may have some species specificity. Ceramic porous rods impregnated with heptyl formate were used to evaluate the effectiveness of controlled vapor release for a single dose of heptyl formate against house flies. Controlled vapor release ofheptyl formate can be used successfully to provide prolonged house fly mortality. Even though the prototype compounds did not exhibit toxicity as high as that of DDVP, their toxicities to dipterans, along with their reduced mammalian toxicities relative to DDVP, make them potential DDVP replacement candidates.

Lethal effects of heat and use of localized heat treatment for control of bed bug infestations.

Bed bugs, Cimex lectularius L., hide in cracks and crevices in furniture and are difficult to control. The bed bug thermal death kinetics were examined to develop a heat treatment method to eliminate bed bug infestations in room contents. High temperatures caused temporary immobilization (knockdown) of bed bugs even with exposures that did not have lethal effects. Exposure of bed bug adults to 39 degrees C for 240 min caused no mortality; however, as temperatures increased from 41 to 49 degrees C, exposure times that caused 100% mortality decreased. The temperature difference to provide a 10-fold change in the mortality was estimated at 4 degrees C, and the estimated activation energy (EA) was between 484 and 488.3 kJ/mol. This demonstrates that bed bugs are not more resistant or susceptible to changes in temperature than other tested insects and that the temperatures needed to kill bed bugs are relatively low. In room treatment tests, heat treatment times varied from 2 to 7 h with complete mortality of exposed bed bugs within the treatment envelope created by surrounding the treated furniture with polystyrene sheathing boards. Containment and circulation of heat around the treated material were crucial factors in an efficient heat treatment for bed bug control. The room floor material greatly affected containment of the heat. The tested method for limited heat treatment of furniture and other room contents required equipment costing less than US$400 and provided opportunity for residual pesticide application around the room with minimal disruption in use of treated room.

Comparative Quantification of Trail-Following Behavior in Pest Ants.

A comparison of trail-following movement parameters of six major urban pest ants, Nylanderia fulva (Forel) (Hymenoptera: Formicidae), Pheidole megacephala, Linepithema humile (Mayr), Solenopsis invicta Buren, Paratrechina longicornis (Forel), and Technomyrmex albipes (Smith) demonstrated several differences in velocity of movement, amplitude of the deviations from a marked trail, percent fidelity to the trail, number of curves per unit of trail, and trail-following accuracy. Paratrechina longicornis and N. fulva had the largest deviations from the marked trails and moved three times faster (25-30 mm/s) along the trail than the slowest ant, S. invicta (< 10 mm/s), with other ants following between these extremes. Species differences in relation to going toward or returning from food were observed in a few cases, especially with Pa. longicornis for which velocity, amplitude, and trail fidelity differed between the foraging and return trails. Quantification of ant trail-following movement parameters can be useful in understanding the mechanics of ant movement and may be important in testing specific strategies and products that disrupt trail-following behavior.

Factors Affecting Short-Range Host-Seeking for the Yellow Fever Mosquito (Diptera: Culicidae).

Understanding short-range cues (e.g., host odorants, heat, moisture) of host-seeking female Aedes aegypti L. is very important for attempts to reduce mosquito bites, to complement current control strategies, and to develop potential spatial repellents. We investigated behavior under semi-field conditions utilizing a new portable uni-port taxis box with a caged chicken host. The combined influences of airflow regimes (0, 1.5, 3, 5, and 6 m/s), distance from host odor (10, 50, 100 cm), host-odor confinement (partial confinement/unconfined), and foraging periodicity (day/evening) were studied. Statistical regression analysis was used to delineate the significant factors that predict upwind flight behavior and short-range source location. Almost 15% of host-seeking Ae. aegypti were activated by an unconfined chicken odor in still air. This was double the number of attracted mosquitoes to confined host odor. The maximum behavioral response was reported with airflow of 5 m/s during daytime (76.7% ± 2.85) at a distance of 10 (70.7% ± 2.47) and 50 cm (56.7% ± 8.88). However, airflow of 6 m/s activated host-seeking orientation during evening assays. The host-seeking response between indoor and outdoor experiments was not significantly different and demonstrated the reliability of the portable taxis box in evaluating mosquito short-range behavioral response toward hosts.

Mosquitocidal Chips Containing the Insect Growth Regulator Pyriproxyfen for Control of Aedes aegypti (Diptera: Culicidae).

Aedes aegypti were exposed to water treated with mosquitocidal chips containing the insecticide pyriproxyfen in a polymer formulation. Chips were tested under different conditions; different water volumes, in containers made of different material, and in water with different levels of organic matter. Treated chips caused 100% mortality of Ae. aegypti during their pupal stage independent of size or type of container, and the mount of organic matter contained in the water to which the mosquito larvae were exposed. When mosquitocidal chips were used in >25% of the oviposition containers within their cages, there was a significant control of the mosquito populations. Mosquitocidal chips worked in different environments, caused significant mosquito population decreases, and were effective in controlling Ae. aegypti.

Sticky Trap Design Considerations for Entrapping Bed Bugs.

Little evidence has been presented on the usefulness of sticky traps for monitoring bed bugs, Cimex lectularius. We examined how the surface roughness around the adhesive of a sticky trap affects both bed bug behavior and adhesive entrapment. In the first assay, bed bugs were placed onto acetate paper discs with different roughness averages (Ra). Each disc was surrounded by sticky trap adhesive and number of captured bed bugs were recorded. The second assay was set up similarly to the first assay except that the outer portion of the acetate disc had a different Ra than the center. In the third assay, bed bugs were placed into circular acetate arenas where they were surrounded by different Ra treatments. The number of times the bed bugs contacted the Ra treatment but did not cross onto the treatment was recorded. Results of these assays showed that as the acetate surfaces got smoother (lower Ra), bed bugs were more likely to get trapped in sticky trap adhesives but also less likely to travel across the smoother surfaces they encountered. A sticky trap design with a smooth plastic film around the adhesive was tested in the field to see if it could capture bed bugs in apartments with known bed bug activity. This trap was not only able to capture bed bugs but was also able to detect unknown German cockroach, Blattela germanica, infestations. Sticky trap designs with smooth surfaces around an adhesive could be used to monitor not only bed bugs but also German cockroaches.

A Low-Cost, Passive Release Device for the Surveillance and Control of Mosquitoes.

Mosquitoes continue to be a major threat to global health, and the ability to reliably monitor, catch, and kill mosquitoes via passive traps is of great importance. Global, low-cost, and easy-to-use outdoor devices are needed to augment existing efforts in mosquito control that combat the spread of disease, such as Zika. Thus, we have developed a modular, portable, non-powered (passive), self-contained, and field-deployable device suitable for releasing volatiles with a wide range of applications such as attracting, repelling, and killing mosquitoes. This unique device relies on a novel nested wick and two-reservoir design that achieves a constant release of volatiles over several hundred hours. Devices loaded with one of either two compounds, geraniol or 1-methylpiperazine (MP), were tested in a controlled environment (32 °C and 70% relative humidity), and both compounds achieved a constant release from our devices at a rate of 2.4 mg/h and 47 mg/h, respectively. The liquid payload can be volatile attractants or repellants as well as mosquitocide-containing feeding solutions for capture and surveillance. This low-cost device can be utilized for both civilian and military mosquito control purposes, but it will be particularly important for protecting those in economically repressed environments, such as sub-Saharan Africa and Central and South America.

Susceptibility of Aedes aegypti, Culex quinquefasciatus Say, and Anopheles quadrimaculatus Say to 19 pesticides with different modes of action.

To access the relative potency of pesticides to control adult mosquitoes, 19 pesticides with various modes of action were evaluated against Aedes aegypti, Culex quinquefasciatus Say, and Anopheles quadrimaculatus Say. On the basis of 24-h LD50 values after topical application, the only pesticide that had higher activity than permethrin was fipronil, with LD50 values lower than permethrin for 107-, 4,849-, and 2-fold against Ae. aegypti, Cx. quinquefasciatus Say, and An. quadrimaculatus Say, respectively. Abamectin, imidacloprid, spinosad, diazinon, and carbaryl showed slightly lower activity than permethrin (<20-fold). However, bifenazate showed very low activity against the three mosquito species tested, with LD50 values higher than permthrin for >1000-fold. On the basis of 24-h LD50 values, Cx. quinquefasciatus was the least susceptible species to nine pesticides tested (DNOC, azocyclotin, chlorfenapyr, carbaryl, spinosad, imidaclorid, diazinon, abamectin, and permethrin) , whereas Ae. aegypti was the least susceptible species to six pesticides tested (dicofol, amitraz, propargite, hydramethylnon, cyhexatin, and diafenthiuron), and An. quadrimaculatus was the least susceptible species to four pesticides tested (bifenazate, pyridaben, indoxacarb, and fipronil). Our results revealed that different species of mosquitoes had different susceptibility to pesticides, showing the need to select the most efficacious compounds for the least susceptible mosquito species to achieve successful mosquito control.

Beauveria bassiana: endophytic colonization and plant disease control.

Seed application of Beauveria bassiana 11-98 resulted in endophytic colonization of tomato and cotton seedlings and protection against plant pathogenic Rhizoctonia solani and Pythium myriotylum. Both pathogens cause damping off of seedlings and root rot of older plants. The degree of disease control achieved depended upon the population density of B. bassiana conidia on seed. Using standard plating techniques onto selective medium, endophytic 11-98 was recovered from surface-sterilized roots, stems, and leaves of tomato, cotton, and snap bean seedlings grown from seed treated with B. bassiana 11-98. As the rate of conidia applied to seed increased, the proportion of plant tissues from which B. bassiana 11-98 was recovered increased. For rapid detection of B. bassiana 11-98 in cotton tissues, we developed new ITS primers that produce a PCR product for B. bassiana 11-98, but not for cotton. In cotton samples containing DNA from B. bassiana11-98, the fungus was detected at DNA ratios of 1:1000; B. bassiana 11-98 was detected also in seedlings grown from seed treated with B. bassiana 11-98. Using SEM, hyphae of B. bassiana11-98 were observed penetrating epithelial cells of cotton and ramifying through palisade parenchyma and mesophyll leaf tissues. B. bassiana11-98 induced systemic resistance in cotton against Xanthomonas axonopodis pv. malvacearum (bacterial blight). In parasitism assays, hyphae of B. bassiana 11-98 were observed coiling around hyphae of Pythium myriotylum.

Expressed sequence tags from the red imported fire ant, Solenopsis invicta: annotation and utilization for discovery of viruses.

An expression library was created and 2304 clones sequenced from a monogyne colony of Solenopsis invicta. The primary intention of the project was to utilize homologous gene identification to facilitate discovery of viruses infecting this ant pest that could potentially be used in pest management. Additional genes were identified from the ant host and associated pathogens that serve as an important resource for studying these organisms. After assembly and removal of mitochondrial and poor quality sequences, 1054 unique sequences were yielded and deposited into the GenBank database under Accession Nos. EH412746 through EH413799. At least nine expressed sequence tags (ESTs) were identified as possessing microsatellite motifs and 15 ESTs exhibited significant homology with microsporidian genes. These sequences most likely originated from Thelohania solenopsae, a well-characterized microsporidian that infects S. invicta. Six ESTs exhibited significant homology with single-stranded RNA viruses (3B4, 3F6, 11F1, 12G12, 14D5, and 24C10). Subsequent analysis of these putative viral ESTs revealed that 3B4 was most likely a ribosomal gene of S. invicta, 11F1 was a single-stranded RNA (ssRNA) virus contaminant introduced into the colony from the cricket food source, 12G12 appeared to be a plant-infecting tenuivirus also introduced into the colony as a field contaminant, and 3F6, 14D5, and 24C10 were all from a unique ssRNA virus found to infect S. invicta. The sequencing project illustrates the utility of this method for discovery of viruses and pathogens that may otherwise go undiscovered.

Relative repellency and lethality of the neonicotinoids thiamethoxam and acetamiprid and an acetamiprid/bifenthrin combination to Reticulitermes flavipes termites.

Field-collected Reticulitermes flavipes (Kollar) termites were placed in bioassay tubes containing soil treated with one of three termiticides: thiamethoxam, acetamiprid, or a combination of acetamiprid + bifenthrin. In the bioassay tubes, treated soil was placed in a layer centered within untreated sand between two 1.5-cm agar plugs. All termiticides were tested at concentrations of 0.1, 1, 10, and 100 ppm with narrow (1 mm), medium (5 mm), and broad (50 mm) thicknesses of treated soil. Soil penetration and termite mortality were measured after 7 d, and repellency was assessed. Thiamethoxam treatments allowed the greatest soil penetration, whereas acetamiprid + bifenthrin treatments were the most inhibitory to soil penetration. Thiamethoxam treatments also caused consistently greater termite mortality than acetamiprid treatments. These data indicated that acetamiprid prevented soil penetration by termites more than thiamethoxam, although both were less repellent compared with bifenthrin alone, which causes little termite mortality at the tested doses. When there was direct contact of treated soil with the agar plugs in broad treatments, the combination of acetamiprid + bifenthrin was more toxic to R. flavipes termites than either acetamiprid or thiamethoxam alone. The combination acetamiprid + bifenthrin termiticide may be effective in keeping termites away from the treated soil, because of the combined effects of acetamiprid and bifenthrin.

Ability of bed bug-detecting canines to locate live bed bugs and viable bed bug eggs.

The bed bug, Cimex lectularius L., like other bed bug species, is difficult to visually locate because it is cryptic. Detector dogs are useful for locating bed bugs because they use olfaction rather than vision. Dogs were trained to detect the bed bug (as few as one adult male or female) and viable bed bug eggs (five, collected 5-6 d after feeding) by using a modified food and verbal reward system. Their efficacy was tested with bed bugs and viable bed bug eggs placed in vented polyvinyl chloride containers. Dogs were able to discriminate bed bugs from Camponotus floridanus Buckley, Blattella germanica (L.), and Reticulitermes flavipes (Kollar), with a 97.5% positive indication rate (correct indication of bed bugs when present) and 0% false positives (incorrect indication of bed bugs when not present). Dogs also were able to discriminate live bed bugs and viable bed bug eggs from dead bed bugs, cast skins, and feces, with a 95% positive indication rate and a 3% false positive rate on bed bug feces. In a controlled experiment in hotel rooms, dogs were 98% accurate in locating live bed bugs. A pseudoscent prepared from pentane extraction of bed bugs was recognized by trained dogs as bed bug scent (100% indication). The pseudoscent could be used to facilitate detector dog training and quality assurance programs. If trained properly, dogs can be used effectively to locate live bed bugs and viable bed bug eggs.

Structure-activity relationships of 33 piperidines as toxicants against female adults of Aedes aegypti (Diptera: Culicidae).

Aedes aegypti (L.) (Diptera: Culicidae) is the primary vector of both dengue and yellow fever. Use of insecticides is one of the primary ways to control this medically important insect pest. However, few new insecticides have been developed for mosquito control in recent years. As a part of our collaborative effort to search for new insecticides to control mosquitoes, piperidine was used as base compound for further optimization. Herein, we report the structure-activity relationships of 33 piperidines against adult female Ae. aegypti. On the basis of 24-h LD50 values after topical application, the most toxic compound was 2-ethyl-piperidine, with an LD50 as low as 0.8 microg per mosquito. The toxicities of piperidine derivatives were significantly decreased when a benzyl moiety was attached to the carbon of the piperidine ring, with an LD50 value as high as 29.2 microg per mosquito. The toxicity order of three moieties attached to the carbon of the piperidine ring was ethyl- > methyl- > benzyl-derivatives. When the same moiety was attached to the piperidine ring, the carbon position to which the moiety was attached conferred different toxicity and the toxicity order was second carbon > third carbon > fourth carbon. Together, these preliminary results may be useful in guiding further piperidine ring modifications in the development of potential new insecticides.