María Inés Picollo

High Resistance to Pyrethroid Insecticides Associated with Ineffective Field Treatments in Triatoma infestans (Hemiptera: Reduviidae) from Northern Argentina

Abstract Field populations of Triatoma infestans Klug were collected during 2002 from four villages in northern Argentina (El Chorro, La Toma, El Sauzal, and Salvador Mazza), after application of deltamethrin and other pyrethroids was ineffective. High levels of resistance to the pyrethroid insecticides deltamethrin, β-cypermethrin, β-cyfluthrin, and lambda-cyhalothrin were detected in all of the evaluated populations. The resistance ratio to pyrethroids determined by topical application ranged from 50.5 (deltamethrin, El Sauzal) to 667.6 (β-cyfluthrin, Salvador Mazza). None of the pyrethroid-resistant insects was resistant to the organophosphorus insecticide fenitrothion. Topical application of piperonyl butoxide to the most deltamethrin-resistant population (Salvador Mazza) led to slight reduction in levels of resistance. Activity of P450 monooxygenase, measured in individual insects through ethoxycoumarine-O-deethylase, showed a slight but noticeable difference in the distribution of activities between susceptible and resistant populations. The total percentage of insects below 0.48 pmol of 7-OH coumarine/min/insect was 36.4 for Salvador Mazza population and 64.3 pmol of 7-OH coumarine/min/insect for CIPEIN strain. Whereas a low level of resistance to deltamethrin was previously related to monooxygenase activity in T. infestans, the high levels of resistance shown by these populations seem to involve monooxygenase in combination with other resistance mechanisms, for example, insensitivity of nervous membrane. Research on T. infestans resistance is in progress to improve Chagas vector control programs in Latin America and to implement resistance management strategies.

Fumigant and Repellent Properties of Essential Oils and Component Compounds Against Permethrin-Resistant Pediculus humanus capitis (Anoplura: Pediculidae) from Argentina

Abstract The repeated use of permethrin and other insecticides for the control of head lice, Pediculus humanus capitis De Geer (Anoplura: Pediculidae), during past decades has resulted in the development of marked levels of resistance. Thus, new alternative insecticides are needed for the control of head lice. We studied the fumigant and repellent properties of essential oils from 16 native and exotic plants in Argentina, and 21 chemical components against permethrin-resistant head lice from Argentina. With a direct vapor-exposure bioassay, the most effective oil was from the native Myrcianthes cisplatensis Cambess (Myrtaceae) with a time to 50% knockdown (KT50) of 1.3 min, followed by exotic species, Eucalyptus cinerea F.V. Muell., Eucalyptus viminalis Labill., and Eucalyptus saligna Smith. with KT50 values of 12.0, 14.9, and 17.4 min, respectively. The most effective components were 1,8-cineole and anisole, with KT50 values of 11.1 and 12.7 min, respectively. Regression analysis of KT50 values and vapor pressures and water-partition coefficients for the essential oil components revealed that the most effective fumigants were among the more volatile components. Repellency assays indicated that the essential oil from Mentha pulegium L. and its benzyl alcohol component were the most effective repellents, having repellency indices of 75.5 and 57.8%, respectively. Thus, some Argentinean plants contain essential oils and components that function as fumigants or as repellents and thereby show potential for development of new control products for head lice.

Differential patterns of insecticide resistance in eggs and first instars of Triatoma infestans (Hemiptera: Reduviidae) from Argentina and Bolivia.

Abstract Previous work at our laboratory has indicated high resistance levels to deltamethrin correlated with failures of chemical control in field populations of Triatoma infestans (Klug, 1834) (Hemiptera: Reduviidae) in northern Argentina and southern Bolivia. The aim of the present work was to study the resistance patterns in eggs and first instars of T. infestans in populations from Argentina and Bolivia. At the egg stage, a population from Salvador Mazza, Argentina, showed the highest resistance ratio to deltamethrin and lambda-cyhalothrin, and it was susceptible to fipronil and fenitrothion. A population from Mataral, Bolivia, showed very low resistance ratios to deltamethrin and lambda-cyhalothrin, and it was susceptible to fipronil and fenitrothion. A Sucre population was susceptible to deltamethrin and fenitrothion, and it showed very low resistant ratios to lambda-cyhalothrin and fipronil. A Yacuiba population was susceptible to deltamethrin. At the first instar, the Salvador Mazza population was susceptible to fipronil, whereas the Mataral and Sucre populations were susceptible to fenitrothion, and they showed very low resistance ratios to lambda-cyhalothrin but the high resistance to fipronil. The Salvador Mazza population was resistant to deltamethrin at the larval stage. Remarkable differences were found in the resistance profile to fipronil in first instars and eggs from Sucre and Mataral. These indicated that the expression of insecticide resistance in eggs varies between populations and that the pyrethroid resistance diagnosed in T. infestans first instars is not indicative of resistance in the egg stage.

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.

Relative Contribution of Monooxygenase and Esterase to Pyrethroid Resistance in Triatoma infestans (Hemiptera: Reduviidae) from Argentina and Bolivia

Abstract Recently, high resistance to pyrethroid insecticides has been associated with ineffective field treatments against Triatoma infestans (Klug) (Hemiptera: Reduviidae) in northern Argentina. Samples were collected from two areas in Argentina (Salta and La Rioja) and one are in Bolivia (Yacuiba), and they were subjected to toxicological and biochemical assays. All populations were resistant to deltamethrin, but they showed different profiles to nonpyrethroid insecticides. The Salta population showed high resistance ratios (RRs) to deltamethrin and only slight differences in the susceptibility to fenitrothion and fipronil compared with the reference strain. Otherwise, the La Rioja population showed a lower RR to deltamethrin and no resistance to fenitrothion or fipronil. Finally, the Yacuiba population had high a RR to deltamethrin, but it was susceptibility to fenitrothion and fipronil. In several cases, deltamethrin-resistant populations had higher susceptibility to bendiocarb than the reference strain. Measured activity of P450 monooxygenase in individual insects (based on ethoxycoumarine-O-deethylase), tended to be higher in the deltamethrin-resistant populations, but the differences were not statistically significant. Activity of specific esterases determined by the hydrolysis of 7-coumaryl permethrate demonstrated an increase in the percentage of insects with higher esterase activity in the Salta and La Rioja populations. Unexpectedly, the Yacuiba population showed lower pyrethroid esterase activity than the reference strain. The different pyrethroid resistance patterns found in T. infestans from three geographical regions within Argentina and in Bolivia suggests that enzyme-based pyrethroid resistance in this species has multiple origins. Nevertheless, because nerve insensitivity (related to the presence of the kdr gene) is also an important mechanism related to pyrethroid resistance, further studies on the kdr gene should be carried to clarify the relative contribution of each pyrethroid-associated mechanism in deltamethrin-resistant populations of T. infestans.

Prevalence and Levels of Permethrin Resistance in Pediculus humanus capitis De Geer (Anoplura: Pediculidae) from Buenos Aires, Argentina

Abstract 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 μl 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.

Role of enhanced detoxication in a deltamethrin-resistant population of Triatoma infestans (Hemiptera, Reduviidae) from Argentina

Deltamethrin and other pyrethroids have been extensively used in Argentina since 1980, for the chemical control of Triatoma infestans Klug (Hemiptera: Reduviidae). Recently, resistance to deltamethrin was detected in field populations by the survival of bugs exposed by topical application to the diagnostic dose estimated on the CIPEIN susceptible strain. Results of the current study showed low resistant ratios (RRs) to deltamethrin for the resistant populations (RR ranged from 2.0 for San Luis colony to 7.9 for Salta colony). Biochemical studies were made on the most resistant colony (Salta) and the susceptible strain (CIPEIN), in order to establish the importance of degradative mechanisms as a cause of the detected resistance. Esterase activity was measured on 3 days old first instars through phenylthioacetate and a-naphtyl acetate activities. The results showed a significant difference in no cholinesterase esterase activity from susceptible (7.6 +/- 0,7 micro M S./i.min.) and Salta resistant colony (9.5 +/- 0.8 microM S./i.min.). Cytochrome p450 mono-oxygenase (p450) activity was measured on individual insects through ethoxycoumarine deethylase (ECOD) activity using a fluorescence microplate reader. The dependence of ECOD activity on age and body region of the nymphs, and pH and time of incubation were studied in order to optimize the measurement. As a result, comparative studies were performed on abdomens of 2 days old first instars at pH 7.2 and 4 h incubation time. ECOD activity of first nymphs was significantly lower in the susceptible colony (61.3 +/- 9.08 pg ECOD/ insect) than in the resistant one (108.1+/- 5.7 pg ECOD/ insect). These results suggest that degradative esterases (no-cholinesterase) and mono-oxygenases cytochrome p450, play an important role in the resistance to deltamethrin in Salta colony from Argentina.

Interspecific hybridization of Eucalyptus as a potential tool to improve the bioactivity of essential oils against permethrin-resistant head lice from Argentina

The essential oils extracted from Eucalyptus grandis, Eucalyptus camaldulensis, Eucalyptus tereticornis, and the hybrids E. grandisxE. camaldulensis, and E. grandisxE. tereticornis were analyzed by GC-MS, and evaluated for their fumigant and repellent effects on permethrin-resistant head lice. Fumigant activity of both hybrids was higher than that for pure species. E. grandisxE. tereticornis and E. grandisxE. camaldulensis showed KT50 values of 12.99 and 13.63min, respectively. E. grandis, E. camaldulensis, and E. tereticornis showed KT50 values of 25.57, 35.01, and 31.31, respectively. A simple regression analysis revealed a significant correlation between KT50 data and % of 1,8-cineole in these essential oils. Repellency varied from 47.80+/-16% to 80.69+/-6% for the five Eucalyptus essential oils tested. Interspecific hybridization improves the pediculicidal activity of Eucalyptus essential oils.

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

Abstract 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× for E49 population, RR 9.5× for HL population and RR > 35.3× 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.

Evidence of Pyrethroid Resistance in Eggs of Pediculus humanus capitis (Phthiraptera: Pediculidae) from Argentina

Abstract Insecticide resistance in Pediculus humanus capitis De Geer 1778 (Phthiraptera: Pediculidae) from different countries has been well documented in the last years. Otherwise, little research has been reported about insecticide resistance in insect embryos and none in human louse embryos. In this work, we studied insecticide resistance in eggs of three head lice populations whose pyrethroid resistance was shown in adults and nymphs compared with a susceptible laboratory body louse strain. All head louse populations showed high permethrin resistance in eggs. Levels of permethrin resistance (LCRs) assessed in eggs by immersion technique were higher than those previously reported for the corresponding populations of adults by topical application. Comparison of LCR values for different populations showed that there was a direct relationship between the resistance levels assessed in eggs and those in adults. All permethrin-resistant eggs showed high resistance to d-phenothrin and dichloro-diphenyl-trichloroethane (DDT) and low resistance to carbaryl, which is in common with the resistance profile established for adults and nymphs. The results concerning the high resistance to pyrethroid (permethrin and d-phenothrin) and the cross-resistance to DDT and carbaryl detected in head louse eggs and adults suggested similar resistance mechanisms in eggs and adults of head louse populations from adults.