A methodology based on insecticide impregnated filter paper for monitoring resistance to deltamethrin in Triatoma infestans field populations.

The domiciliary presence of Triatoma infestans (Klug) (Hemiptera: Reduviidae) after control interventions was reported in recent years. Toxicological studies showed high levels of resistance to pyrethroids suggesting resistance as one of the main causes of deficient control. The aim of the present study was to develop a protocol to test resistance to deltamethrin in T. infestans collected from the field by discriminate concentration. To evaluate field insects, the effect of age (early vs. later) and nutritional state (starved vs. fed) on the deltamethrin susceptibility of each developmental stage was studied. Topical and insecticide impregnated paper bioassays were used. Using the impregnated paper, the susceptibility to deltamethrin was not affected by the age of the stadium and the nutritional states, and varied with the post-exposure time and with the different developmental stages. A discriminant concentration of deltamethrin (0.36% w/v) impregnated in filter paper was established for all developmental stages. Finally, the methodology and the discriminant concentration were evaluated in the laboratory showing high sensitivity in the discrimination of resistance. The present study developed a methodology of exposure to insecticide impregnated papers and proposes a protocol to test T. infestans in field populations with the aim to detect early evolution of resistance to deltamethrin.

The Botanical Monoterpenes Linalool and Eugenol Flush-Out Nymphs of Triatoma infestans (Hemiptera: Reduviidae).

Monoterpenes are the main components of essential oils. Some members of this chemical family present insecticidal activity. Triatoma infestans (Klug) is the main vector of Chagas disease in Argentina, Bolivia, Paraguay, and Perú. The objective of this work was to evaluate the effect of six monoterpenes (1,8-cineole, eugenol, linalool, menthol, α-terpineol, and thymol) on the locomotor and flushing out activity of T. infestans. A video tracking technique was used to evaluate the locomotor activity of nymphs exposed to different concentrations of these chemicals applied as films on filter paper. Papers treated with acetone alone were used as negative controls, while solutions of tetramethrin were applied as positive controls. Only linalool and menthol produced hyperactivation. Flushing out was assessed under laboratory conditions using a standardized aerosolization method. All monoterpenes were applied at 1.5 g/m3. 1,8-Cineole, α-terpineol, and thymol flushed out 10% or less nymphs. The average flushing out produced by eugenol was 36.7%. Values of median flushing out time (FT50) could only be calculated for linalool and menthol (16.67 and 42.98 min, respectively). The FT50 value for the positive control tetramethrin (applied at 0.006 g/m3) was 8.29 min. Following these results, the flushing out activity of a mixture of linalool and eugenol was evaluated. The FT50 of this 2:1 linalool:eugenol mixture was 40.73 min. Finally, flushing out assays performed in semifield conditions showed similar results to those obtained at the laboratory.

Behavioral and toxicological responses of Rhodnius prolixus and Triatoma infestans (Hemiptera: Reduviidae) to 10 monoterpene alcohols.

The effect on locomotor activity, the repellency, and the knock-down produced by 10 monoterpene alcohols were evaluated on first-instar nymphs of Rhodnius prolixus and Triatoma infestans, vectors of Chagas disease. A video tracking technique was used to evaluate locomotor activity and repellency by exposure to papers impregnated with monoterpenes. Eugenol on R. prolixus and (S)-cis-verbenol on T. infestans did not modify the locomotor activity. The remaining monoterpenes produced hyperactivity on both species, although the concentration required was at least a 1,000 times higher than that of deltamethrin (positive control). Carvacrol, eugenol, and geraniol resulted as repellent as N,N-diethyl-m-toluamide (positive control) for both species. A similar result was observed for almost every monoterpene on T. infestans. Knock-down effect was evaluated by exposing the nymphs in closed recipients. The order of increasing toxicity on R. prolixus was (KT50 values in min): geraniol (213.7) < alpha-terpineol (164.5) < linalool (124.2) < carvacrol (111.6) < eugenol (89.8) < thymol (78.9), and on T. infestans: alpha-terpineol (289.8) < eugenol (221.3) < carvacrol (164.2) < linalool (154.9) < thymol (96.7). All monoterpenes were less toxic than the positive control, dichlorvos (3.6 min for R. prolixus and 3.9 min for T. infestans). After 7 h of exposure, (-)-carveol, citronellol, and menthol (on both species) and geraniol (on T. infestans) produced < 50% of knock-down. After these results, it is worthwhile to explore more deeply the potential of these compounds as tools for controlling Chagas disease vectors.

Toxicity to vapor exposure and topical application of essential oils and monoterpenes on Musca domestica (Diptera: Muscidae).

The medical and veterinary pest Musca domestica L. has developed resistance to most insecticides used against it. For this reason, there is a constant search for new alternative control tools. The aims of this study were (1) to evaluate the toxicological effects caused by the fumigant activity and the topical application of five essential oils and five monoterpenes in M. domestica adult males and (2) to study the variation of the fumigant activity of the essential oils and monoterpenes according to the solvent used (acetone or a silicone base). Houses flies were exposed to vapors delivered by filter paper treated with 200 microl of essential oil or monoterpene (10%) in acetone or a silicone base. The knockdown time 50% (KT50) values obtained for essential oils (expressed in minutes) were 3.3 (eucalyptus); 10.1 (orange); 10.4 (mint); 10.9 (lavender); and 17.7 (geranium). The KT50 values obtained for monoterpenes (expressed in minutes) were 2.3 (eucalyptol); 7.5 (limonene); 7.6 (linalool); 19.0 (menthone); and 22.6 (menthyl acetate). In all cases, a delay in the onset of poisoning symptoms was observed when a silicone base vehicle was used. When topically applied, the lethal dose 50% (LD50) values for essential oils (expressed in micrograms of oil/insect) were 0.07 (geranium); 0.09 (mint); 0.13 (lavender); 0.14 (eucalyptus); and 0.16 (orange). The LD50 values for monoterpenes (expressed in micrograms of monoterpene/insect) were 0.04 (linalool); 0.09 (menthyl acetate); 0.10 (limonene); 0.11 (menthone); and 0.13 (eucalyptol). These results suggest that the studied essential oils and monoterpenes are potential tools for controlling M. domestica.

Fumigant insecticidal activity and repellent effect of five essential oils and seven monoterpenes on first-instar nymphs of Rhodnius prolixus.

The aim of this study was to evaluate the fumigant and repellent activity of five essential oils (from eucalyptus, geranium, lavender, mint, and orange oil) and seven monoterpenes (eucalyptol, geraniol, limonene, linalool, menthone, linalyl acetate, and menthyl acetate) on first-instar nymphs of the bloodsucking bug Rhodnius prolixus Stahl (vector of Chagas disease in several Latin American countries). Fumigant activity was evaluated by exposing the nymphs to the vapors emitted by 100 microl of essential oil or monoterpene in a closed recipient. The knockdown time 50% (KT50) for eucalyptus essential oil was 215.6 min (seven times less toxic than dichlorvos, a volatile organophosphorus insecticide used as a positive control). The remaining essential oils showed a poor fumigant activity: < 50% of nymphs were knocked down after 540 min of exposure. The KT50 values for monoterpenes, expressed in minutes, were as follows: 117.2 (eucalyptol), 408.7 (linalool), 474.0 (menthone), and 484.2 (limonene). Eucalyptol was 3.5 times less toxic than dichlorvos. No affected nymphs were observed after 540 min of exposure to geraniol, linalyl acetate, or menthyl acetate. Repellency was quantified using a video tracking system. Two concentrations of essential oils or monoterpenes were studied (40 and 400 microg/cm2). Only mint and lavender essential oils produced a light repellent effect at 400 microg/cm2. Geraniol and menthyl acetate produced a repellent effect at both tested concentrations and menthone only elicited an effect at 400 microg/cm2. In all cases, the repellent effect was lesser than that produced by the broad-spectrum insect repellent N,N-diethyl-3-methylbenzamide (DEET).

Anticholinesterase and pediculicidal activities of monoterpenoids.

The repetitive and inadequate application of pediculicidal products frequently results in the development of resistance to these compounds. Essential oils are a promising alternative to synthetic insecticides, although their mode of action remains to be explored. It has been proposed that one possible target of the essential oils is the inhibition of acetylcholinesterase (AChE). The role of monoterpenoids as possible AChE inhibitors and their relationship with the toxicity was investigated both in vitro and in vivo. Inhibition of electric eel AChE activity showed that the most effective inhibitor was 1,8-cineole with IC(50) 6 x 10(-3) M. The inhibition of AChE activity of head louse homogenate by 1,8-cineole showed IC(50) 7.7 x 10(-2) M. The intoxication symptoms of head lice exposed to vapors of 1,8-cineole was recorded before the in vivo head louse AChE inhibition assay. No correlation was found between neurotoxic symptoms and inhibition of AChE activity.

Volatile compounds secreted by Brindley's glands of adult Triatoma infestans: identification and biological activity of previously unidentified compounds.

Volatile emissions of adult male Triatoma infestans were collected on non-polar SPME fibers and analyzed by gas chromatography linked to a mass spectrometer. A complex mixture of 16 short-chain esters and acids were identified. The composition of short-chain aliphatic acids (ethanoic to nonanoic acids) was similar to previously reported results. The most abundant aliphatic acid was 2-methylpropanoic acid, constituting 18% of the total volatile content. Also abundant were the esters 2- and 3-methylbutyl 2-methylpropanoate, which constituted 30% and 22%, respectively, of the total volatile content. A similar pattern of compounds was observed in the volatiles secreted by dissected male Brindley's glands; however, in this case, 2- and 3-methylbutan-1-ol were detected which were not found in live insect volatile emissions. Large variability in volatile composition was also observed among the glands excised from different insects. Electroantennographic (EAG) evaluation of the components of Brindley's gland showed significant responses for 2- and 3-methylbutyl 2-methylpropanoate compared to controls. The mixture of volatiles secreted by excised Brindley's glands and the isolated 2- and 3-methylbutyl 2-methylpropanoate had repellent effects on both male and female T. infestans, possibly associated with a defensive strategy.

Permethrin-resistant head lice (Anoplura: Pediculidae) in Argentina are susceptible to spinosad.

The insecticidal activity of spinosad was evaluated against susceptible and permethrin-resistant human lice. In a permethrin-susceptible strain of the body louse, Pediculus humanus humanus L. (Anoplura: Pediculidae), the toxicity of spinosad was similar to that established for permethrin, with an LD50 value of 1.2 ng/insect and 2.4 ng/insect, respectively. Topical application of spinosad to populations of permethrin-resistant head louse, Pediculus humanus capitis De Geer (Anoplura: Pediculidae), showed that susceptibility to spinosad was independent of resistance to permethrin. The effectiveness of spinosad against human lice and the low mammalian toxicity reported in the literature suggest that spinosad could be useful for the management of permethrin-resistant human lice.

Identification of volatile emissions from Platypus mutatus (=sulcatus) (Coleoptera: Platypodidae) and their behavioral activity.

We report here the identification and behavioral activity of volatile compounds emitted by male Platypus mutatus (=sulcatus) Chapuis while boring galleries in living poplar, Populus deltoides Marshall, trees. Headspace analysis using solid phase microextraction techniques showed the presence of 6-methyl-5-hepten-2-ol (sulcatol) and 6-methyl-5-hepten-2-one (sulcatone). Only one enantiomer of sulcatol, retusol, was found to be part of the volatile emission. Behavioral assays showed that females are more attracted than males to galleries with boring males inside. Both sulcatol and sulcatone elicited electroantennographic responses by female P. mutatus. Furthermore, behavioral bioassays showed that both sulcatol and sulcatone elicit behaviorally attractive responses by females. These results suggest that male P. mutatus releases a sex pheromone composed mainly of retusol and sulcatone.

Increased monooxygenase activity associated with resistance to permethrin in Pediculus humanus capitis (Anoplura: Pediculidae) from Argentina.

We studied the profile of permethrin resistance in populations of head lice infesting children 6-12 yr old in schools and their homes in and around Buenos Aires, Argentina. Five permethrin-resistant populations with different levels of resistance were collected: Hogar Loyola (HL), Republica de Turquia (RT), Hogar Mitre (HM), Guardia de Honor (GH), and Ricardo Guiraldes (RG). One susceptible population, Bandera Argentina (BA), also was collected. Their level of resistance was evaluated, and results showed resistance ratios of 13 for HL, 16 for RT, 22 for HM, 61 for GH, and 69 for RG. To elucidate the possible involvement of the cytochrome P450 monooxygenase system in conferring permethrin resistance, ethoxycoumarin-O-deethylase (ECOD) activity was measured in abdomens of individual third instars and adults by using a fluorometric assay. The ECOD activity was lower in the susceptible BA population (4.7 ng per louse) than in the resistant ones (13.7 ng per louse for RG, 12.3 ng per louse for GH, 8.6 ng per louse for RT, and 8.2 ng per louse for HL). ECOD activity was significantly correlated with the level of resistance in the field populations (r = 0.97, P = 0.0009), suggesting a role for cytochrome monooxygenase P450 system in permethrin resistance by head louse, Pediculus humanus capitis De Geer.

Behavioural response of Triatoma infestans (Klug) (Hemiptera: Reduviidae) to quinazolines.

The behavioural responses of the haematophagous bug Triatoma infestans towards some previously identified components of its faeces: 4-methylquinazoline, 2,4- dimethylquinazoline and their mixtures were evaluated using a video tracking system. Fifth instar nymphs and females but not males were significantly attracted to polyethylene glycol formulations of 4-methyl + 2,4-dimethylquinazoline (50 microg each). Fifth instar nymphs were also attracted to 4-methylquinazoline alone (50 microg) but females were only attracted by the mixture of both methyl quinazolines (50 microg each). Syntheses of both methyl quinazolines were carried out starting from 2-aminoacetophenone by modifying the conditions of reported procedures.

Prevalence and levels of permethrin resistance in Pediculus humanus capitis De Geer (Anoplura: Pediculidae) from Buenos Aires, Argentina.

Permethrin has been used extensively for control of Pediculus humanus capitis De Geer (Anoplura: Pediculidae) in Argentina since 1990, resulting in the development of resistance to this and other pyrethroids. This resistance was first detected in some field populations in 1997. A survey for resistance in Buenos Aires in 2001 revealed significant resistance levels in lice on children at 24 of 26 (92.3%) schools. When compared with a previously unexposed reference population, resistance ratios (RRs) obtained by exposing the insects to filter papers impregnated with permethrin ranged from 2 to 60 in 10 (39%) of the schools. RRs in the remaining 14 (61%) populations were too great to not be measured with the filter paper method (RR > 88.7). As an alternative, we used topical applications of 0.1 microl of acetone solution of permethrin on the dorsal abdomen of adults and third instars. This topical method, which has not been previously reported for head lice, was capable of quantifying higher levels of resistance. Highly resistant populations had RRs from 162.5 to 655.2. When applied to populations with low and intermediate levels of resistance, results from the filter paper and topical application methods were highly correlated, and RRs from topical application were higher than those from the filter paper method. Results from the combination of the two methods indicated that head lice among Buenos Aries school children are highly resistant to permethrin, and the resistance is widespread.

Toxic effect of aliphatic alcohols against susceptible and permethrin-resistant Pediculus humanus capitis (Anoplura: Pediculidae).

The effectiveness of 1-octanol, 1-nonanol, 1-decanol, 1-undecanol, and 1-dodecanol was evaluated by immersion method against susceptible and permethrin-resistant head lice, Pediculus humanus capitis De Geer, from Buenos Aires, Argentina. All the tested alcohols showed knockdown effect at 10 min and mortality 18 h after treatment. The highest activity was found for the 1-dodecanol (KC50 2.55%, LC50 2.28%) and the lowest for 1-octanol (KC50 8%, LC50 4.46%). The toxicity to the head lice systematically increased with the increase in carbon atoms in the n-aliphatic alcohol moiety, and with the octanol:water coefficient (r2 = 0.94). The pediculicidal activity of 1-dodecanol was not correlated with resistance to permethrin, because no significant difference was observed between toxicity parameters in the susceptible (MAR) and the permethrin-resistant populations which had different resistant levels (RR 5.77 x for E49 population, RR 9.5 x for HL population and RR > 35.3 x for GH population). The pediculicidal effect of aliphatic alcohols demonstrated in this study and the lack of correlation with the permethrin resistance may prove to have a practical value for use in susceptible and permethrin-resistant head lice control.

Residualidad de un formulado sólido del insecticida microbiano Bti (H-14) en el control de larvas de Aedes aegypti (Diptera: Culicidae) / Residuality of a microbial insecticide solid formulated Bti (H-14) in the

El Bacillus thuringiensis var. israelensis (Bti ó Bti H-14) produce una toxina proteica, la cual constituye una herramienta de gran utilidad para el control de larvas de Aedes aegypti. Una suspensión líquida de la toxina y las esporas (Bti) ha sido utilizada en campañas nacionales de control del mosquito vector del Dengue en Argentina, pero la falta de efecto residual constituye elmayor inconveniente para su aplicación. En la búsqueda de nuevas estrategias para el control de vectores de enfermedades, este trabajo presenta algunos resultados sobre una nueva formulación de liberación controlada de Bti, denominada ôMosquito Dunks®õ. Se utilizaron larvas de Ae. aegypti del III estadio tardío ó IV temprano (cepa susceptible CIPEIN) que en ensayos de laboratorio demostraron una actividad residual a dosis de uso recomendadas por el fabricante (14,4 mg/cm2) de al menos 180 días con mortalidad total luego de 48 horas de exposición. Los resultados que se presentan en este trabajo sugieren que la nueva formulación sólida de liberación lenta de Bti, debido a su residualidad representa una buena alternativa para el reemplazo del Temefos en el control de larvas de Aedes aegypti, especialmente en aquellos casos donde se detecte resistencia al larvicida organofosforado.

The entomopathogenous agent Bacillus thuringensis var. israelensis (Bti or Bti H-14), produces a proteic toxin that is a promissory tool to use in chemical control of larvae of Aedes aegypti. Bti (H-14) as a liquid suspension, has been used in national campaigns against Ae. aegypti , vector of Dengue disease, but its lack of residual effect is a major inconvenient. In the search of new strategies in vector control, this paper presents some results of an alternative slow release formulation of Bti (H-14), the ôMosquito Dunksõ . III instar or young IV instar larvae of a susceptible strain of Aedes aegypti (CIPEIN strain) was used in laboratory assays. The ôMosquito Dunksõ at recommended use dosis (14,4 mg / 100 cm2 surface) were assayed for residual activity. 100% mortality was found for at least 180 days (6 month) with 48 hs larvae exposition. The results presented in this work suggests that this new solid slow release formulation of Bti with more residual activity could be a good alternative for Temephos for Aedes aegypti larvae control, specially for those cases in which resistance to the organophosphorus compound is reported.

Past and present of Chagas vector control and future needs

It presents a historical background, and information on insecticides used, insecticide formulation, innovative tools, insecticide resistance, and future needs. Document in pdf format; Acrobat Reader required.

Third instar nymphs of Rhodnius prolixus exposed to alpha-cyanopyrethroids: from hyperactivity to death.

The hyperactivity, incoordination, recovery, and mortality produced by four alpha-cyanopyrethroids usually used for Chagas disease vector control (beta-cypermethrin, beta-cyfluthrin, lambda-cyhalothrin, and deltamethrin) were evaluated on third instar nymphs of Rhodnius prolixus. All pyrethroids modified the locomotor activity of the nymphs, which increased linearly as a function of the log of insecticide concentration. lambda-Cyhalothrin showed the lowest values of Effective Concentration 50%, Lethal Concentration 50%, Effective Time 50%, and Lethal Time 50% when insecticides were applied by contact with treated filter papers. Recovery from incoordination was observed after topical application of the insecticides. The recovery was inhibited by the simultaneous application of piperonyl butoxide, suggesting that biotransformation by mixed-function microsomal oxidases is involved in the process of recovery.

Behavioral response of fifth instar nymphs of Triatoma infestans (Hemiptera: Reduviidae) to pyrethroids.

The hyperactivity (an increase in locomotor activity) and repellency produced by eight pyrethroids, applied as films on filter paper, were evaluated on fifth instar nymphs of Triatoma infestans (Hemiptera: Reduviidae) using a video tracking technique. All the pyrethroids studied produced hyperactivity. As a trend, hyperactivity produced by cyanopyrethroids was higher than that produced by non-cyanopyrethroids. Hyperactivity was not observed when nymphs were pretreated with the sulphydryl reagent N-ethylmaleimide before exposure to the pyrethroids. The eight pyrethroids failed to produce repellency. No repellency was also observed for the flowable formulation of deltamethrin at the concentration recommended for T. infestans control.

[The use of insecticide fumigant canister to protect an insectarium and its residual effect against triatomine bugs, in laboratory conditions]. / Emprego de pote fumígeno para proteção de insetário e sua ação residual contra triatomíneos, em condições de laboratório.

An insecticide fumigant canister based on synthetic pyrethroids and dichlorvos was employed against cockroaches and ants which were invading an insectarium used for rearing triatominae. After removal of the Triatominae, the canister was activated and found to kill all the invading insects within 48 hours. Possible residual action against triatomines was then monitored by a 24-hour exposure of eggs, nymphs and adults of Triatoma infestans, Panstrongylus megistus and Rhodnius neglectus in the treated insectarium. No ovicidal action was observed but some mortality of adults and nymphs of the three species was observed up to 72 hours after the fumigation.

Resistance to insecticides and effect of synergists on permethrin toxicity in Pediculus capitis (Anoplura: Pediculidae) from Buenos Aires.

Permethrin-resistant colonies of Pediculus capitis (De Geer) from Buenos Aires were used to establish a resistance profile and to examine resistance mechanisms. All permethrin-resistant head lice (resistance ratio from 52.8 to > 88.7) were also resistant to d-phenothrin (resistance ratio from 40.86 to > 48.39) and deltamethrin (resistance ratio from 16.24 to 38.06). No cross-resistance to carbaryl was found in any of the pyrethroid-resistant P. capitis tested. Otherwise, all resistant colonies showed low to high levels of resistance to beta-cypermethrin. This pyrethroid had never been applied as a pediculicide in Argentina; however, the high level of resistance found in these permethrin-resistant colonies (resistance ratio from 9.74 to 50.97) demonstrated that pyrethroid cross-resistance occurred to this novel insecticide. Treatment with piperonyl butoxide (PBO) or triphenylphosphate (TPP) significantly decreased the toxicity of permethrin in the four colonies tested. The esterase inhibitor TPP produced lower enhancement of toxicity than the multifunction oxidase inhibitor PBO in the colonies having the highest resistance levels. Results presented here concerning the cross-resistance profile and synergism by enzyme inhibitors in permethrin-resistant head lice demonstrated that enhanced metabolism was involved in the pyrethroid resistance. However, the substantial degree of resistance that remained after synergism suggested the presence of another resistance mechanism. Cross-resistance to pyrethroid and susceptibility to the carbamate carbaryl suggested a common action mechanism.

Antifeedant activity of withanolides from Salpichroa origanifolia on Musca domestica.

The antifeedant effect of several salpichrolides on larvae of Musca domestica was investigated. Three naturally occurring compounds, salpichrolide A (1), salpichrolide C (2), and salpichrolide G (3), previously isolated from Salpichroa origanifolia, and two known (4, 6) and three new (5, 7, 8) synthetic analogues were tested. The maximal effect on development was observed for salpichrolide A (1), while salpichrolide G (3) was the most toxic. The content of the salpichrolides in S. origanifolia was monitored by HPLC during plant development, reaching a maximum during summer.