Pyrethroid resistance distribution in Triatoma infestans and environmental association along the Argentine endemic zone.

Insecticide resistance is considered a barrier to chemical control of Triatoma infestans, the main vector of Chagas disease in the Southern Cone of South America. Although initiatives to reduce the incidence of the disease in the region have integrated different strategies, they have mainly relied on vector elimination using pyrethroid insecticides such as deltamethrin. Reports of pyrethroid resistance in connection with T. infestans control failures first emerged in northern Argentina and southern Bolivia. Recently, a mosaic pyrethroid-resistant focus has been described in the center of the Argentine Gran Chaco (Department of General Güemes, Chaco Province), characterized by the presence of susceptible and very highly resistant populations in the same area. The involvement of different resistance mechanisms has been proposed, together with the contribution of environmental variables that promote the toxicological heterogeneity described. In the endemic zone of Argentina, however, new questions arise: Are there any other clusters of resistance? Is there a relationship between the distribution of resistance and environmental variables (as has been observed at smaller scale)? We studied toxicological data from insects collected and analyzed at 224 localities between 2010 and 2020 as part of the resistance monitoring conducted by the Chagas National Program. The sites were classified according to the survival rate of insects exposed to a discriminant dose of deltamethrin: 0-0.19 were considered susceptible, 0.2-0.79 low-resistance, and 0.8-1 high-resistance. Localities were georeferenced to describe the spatial distribution of resistance and to identify environmental variables (demographics, land use, urbanization, connectivity, and climate) potentially associated with resistance. We used Generalized Linear Models (GLMs) to examine the association between resistance and environmental predictors, selecting error distributions based on the response variable definition. For the entire period, 197 susceptible localities were distributed across the endemic zone. Localities with different survival rates were found throughout the area; 9 high-resistance localities circled the two previously identified resistant foci, and 18 low-resistance in 6 provinces, highlighting their relevance for control planning. Precipitation variables were linked to resistance in all the GLMs evaluated. Presence/absence models were the most accurate, with precipitation, distance from the capital city, and land use contributing to the distribution of resistance. This information could be valuable for improving T. infestans control strategies in future scenarios characterized by unpredictable changes in land use and precipitation.

Fluctuating asymmetry and exposure to pyrethroid insecticides in Triatoma infestans populations in northeastern Argentina

Exposure to sublethal doses of insecticide may affect biological traits in triatomines. We investigated the effects of toxicological phenotype (pyrethroid resistance status) and exposure to sublethal doses of deltamethrin on two traits of Triatoma infestans Klug (Heteroptera: Reduviidae) using a phenotypic plasticity experimental design. First-instar nymphs from 14 and 10 full-sib families from pyrethroid-susceptible and pyrethroid-resistant populations, respectively, were used. For the susceptible population, we treated first instars topically with acetone (control) or deltamethrin (treatment) once. For the resistant population, instars were treated once, twice and three times as first, third or fifth-instar nymphs, respectively. We measured cuticle thickness, wing size and wing shape of 484 emerging adults, and tested for treatment effects using mixed ANOVA and MANOVA models. Toxicological phenotype, exposure to deltamethrin and full-sib family exerted significant effects on cuticle thickness, wing size and wing shape. Adult triatomines previously treated with deltamethrin developed significantly thicker cuticles than control triatomines only in the resistant population and significantly bigger wings in both populations. Mean cuticle thickness and wing size increased with increasing exposures to deltamethrin. Exposure to sublethal doses of deltamethrin generated morphological modifications that may affect insect survival and flight dispersal, and hence may have evolutionary and epidemiological consequences.

Estado de la resistencia a insecticidas en Triatoma infestans de Argentina

Los insectos triatominos son los vectores de la enfermedad de Chagas. El control químico de estos insectos es la principal herramienta utilizada para reducir la incidencia de la enfermedad. Los piretroides son los principales insecticidas utilizados para el control de los triatominos en todos los países endémicos. La evolución de resistencia a insecticidas convierte en inefciente una exitosa estrategia de control químico. En Argentina se han detectado diferentes focos de resistencia en Triatoma infestans, el vector más importante en el Cono Sur de Sudamérica, asociados a defciencias en el control. Los estudios realizados sobre esta especie muestran que la resistencia es un problema complejo, ya que evolucionó en varias zonas de la distribución geográfca de la especie, se identifcaron diferentes mecanismos de resistencia, está asociada a modifcaciones en diversos procesos biológicos, está vinculada a variaciones ambientales y hay escasez de alternativas de control. En este manuscrito presentamos una revisión del conocimiento sobre la resistencia a piretroides en T. infestans en Argentina.

Genetic structure of deltamethrin-resistant populations of Triatoma infestans (Hemiptera: Reduviidae) in the Gran Chaco.

The genetic structure of natural populations offers insight into the complexities of their dynamics, information that can be relevant to vector control strategies. Microsatellites are useful neutral markers to investigate the genetic structure and gene flow in Triatoma infestans, one of the main vectors of Chagas disease in South America. Recently, a heterogeneous pyrethroid-resistant hotspot was found in the Argentine Gran Chaco, characterized by the highest levels of deltamethrin resistance found at the present time. We applied population genetics analyses to microsatellite and village data and search for associations between the genetic variability and the heterogeneous toxicological pattern previously found. We genotyped 10 microsatellite loci in 67 T. infestans from 6 villages with no, low, and high pyrethroid resistance. The most genetically diverse populations were those susceptible or with low values of resistance. In contrast, high-resistance populations had lower herozygosity and some monomorphic loci. A negative association was found between variability and resistant ratios. Global and pairwise FSTs indicated significant differentiation between populations. The only susceptible population was discriminated in all the performed studies. Low-resistance populations were also differentiated by a discriminant analysis of principal components (DAPC) and were composed mostly by the same two genetic clusters according to STRUCTURE Bayesian algorithm. Individuals from the high-resistance populations were overlapped in the DAPC and shared significant proportions of a genetic cluster. These observations suggest that the resistant populations might have a common origin, although more genetic markers and samples are required to test this hypothesis more rigorously.

Insecticide Resistance Mechanisms in Triatoma infestans (Reduviidae: Triatominae): The Putative Role of Enhanced Detoxification and Knockdown Resistance (kdr) Allele in a Resistant Hotspot From the Argentine Chaco.

Chagas disease affects around 6 million people in the world, and in Latin America, it is mainly transmitted by the kissing bug. Chemical control of the vector with pyrethroid insecticides has been the most frequently used tool to reduce the disease incidence. Failures of field control have been detected in areas of the Argentinian Gran Chaco that correlate with high levels of insecticide resistance. Here, we provide evidence of the mechanisms involved in the resistance to insecticides of field populations of T. infestans from General Güemes Department (Chaco Province, Argentina). The biochemical analysis suggests the increase in the activity of the degradative enzymes P450 oxidases and esterases as a minor contributive mechanism in low-resistance populations. The molecular study revealed high frequencies of the kdr L925I mutation at the voltage-gated sodium channel as responsible for the high resistance ratios detected. This knowledge contributes to the generation of comprehensive vector control strategies that reduce the incidence of the disease.

Insecticide Resistance Mechanisms in Triatoma infestans (Reduviidae: Triatominae): The Putative Role of Enhanced Detoxification and Knockdown Resistance (kdr) Allele in a Resistant Hotspot From the Argentine Chaco / Mecanismos de resistência a inseticidas em Triatoma infestans (Reduviidae: Triatominae): o papel putativo do alelo de desintoxicação aprimorada e resistência à queda ( kdr ) em um hotspot resistente do Chaco argentino

Chagas disease affects around 6 million people in the world, and in Latin America, it is mainly transmitted by the kissing bug. Chemical control of the vector with pyrethroid insecticides has been the most frequently used tool to reduce the disease incidence. Failures of field control have been detected in areas of the Argentinian Gran Chaco that correlate with high levels of insecticide resistance. Here, we provide evidence of the mechanisms involved in the resistance to insecticides of field populations of T. infestans from General Güemes Department (Chaco Province, Argentina). The biochemical analysis suggests the increase in the activity of the degradative enzymes P450 oxidases and esterases as a minor contributive mechanism in low-resistance populations. The molecular study revealed high frequencies of the kdr L925I mutation at the voltage-gated sodium channel as responsible for the high resistance ratios detected. This knowledge contributes to the generation of comprehensive vector control strategies that reduce the incidence of the disease.

Genetic structure of deltamethrin-resistant populations of Triatoma infestans (Hemiptera: Reduviidae) in the Gran Chaco / Estrutura genética de populações de Triatoma infestans (Hemiptera: Reduviidae) resistentes à deltametrina no Gran Chaco

The genetic structure of natural populations offers insight into the complexities of their dynamics, information that can be relevant to vector control strategies. Microsatellites are useful neutral markers to investigate the genetic structure and gene flow in Triatoma infestans, one of the main vectors of Chagas disease in South America. Recently, a heterogeneous pyrethroidresistant hotspot was found in the Argentine Gran Chaco, characterized by the highest levels of deltamethrin resistance found at the present time. We applied population genetics analyses to microsatellite and village data and search for associations between the genetic variability and the heterogeneous toxicological pattern previously found. We genotyped 10 microsatellite loci in 67 T. infestans from 6 villages with no, low, and high pyrethroid resistance. The most genetically diverse populations were those susceptible or with low values of resistance. In contrast, high-resistance populations had lower herozygosity and some monomorphic loci. A negative association was found between variability and resistant ratios. Global and pairwise FSTs indicated significant differentiation between populations. The only susceptible population was discriminated in all the performed studies. Low-resistance populations were also differentiated by a discriminant analysis of principal components (DAPC) and were composed mostly by the same two genetic clusters according to STRUCTURE Bayesian algorithm. Individuals from the high-resistance populations were overlapped in the DAPC and shared significant proportions of a genetic cluster. These observations suggest that the resistant populations might have a common origin, although more genetic markers and samples are required to test this hypothesis more rigorously.

Excretion/defecation patterns in Triatoma infestans populations that are, respectively, susceptible and resistant to deltamethrin

Pyrethroid resistance has been detected in Triatoma infestans (Klug) (Hemiptera: Reduviidae) specimens from different areas of Argentina and Bolivia. Genes conferring resistance can have a pleiotropic effect with epidemiological and evolutionary consequences. This research studied excretion/defecation patterns in deltamethrin-resistant T. infestans in order to elucidate its biological performance, adaptive consequences and role in the transmission of Chagas' disease. One deltamethrin-susceptible strain and two deltamethrin-resistant strains were used. Fifth-instar nymphs were fed ad libitum and their defecations recorded during and after the first or second feeding in the stadium. Resistant insects began to defecate later, defecated less, showed a lower proportion of defecating individuals and lower defecation indices compared with susceptible insects during the first hour after feeding. The number of bloodmeals in the stadium did not affect the main variables determining the pattern of defecation. The present study suggests that alterations in the excretion/defecation pattern in resistant insects entail an adaptive cost and, considering only this pattern, determine a lower capacity for transmission of Trypanosoma cruzi (Kinetoplastida: Trypanosomatidae) compared with susceptible insects.

Modulation of the behavioral and electrical responses to the repellent DEET elicited by the pre-exposure to the same compound in Blattella germanica.

Insects under different stimuli from the environment modify behavioural responses due to changes in the sensitivity of neurons at the peripheral and/or at the central level of the nervous system. This phenomenon is called neuronal plasticity, and sensory adaptation is an example of it. An insect repellent is a chemical that produces oriented movements of the insects away from its source. In this work we studied the modulation of the behavioural and electrical response to the repellent N, N-diethyl-3-methylbenzamide (DEET) in males of the German cockroach B. germanica produced by previous exposure to the same repellent. Methods. We determined repellency using a circular arena, one half of which was treated with DEET. The time spent by insects in each half of the arena was measured, and a repellency coefficient (RC) was calculated. The RCs of pre-exposed and non-pre-exposed insects were compared. To determine a possible role of nitric oxide in the modulation of the response to DEET after pre-exposure, the nitric oxide donor S-nitroso-acetyl-cysteine (SNAC) was applied on cockroaches' antennae. The electrical activity of the cockroaches' antennae in response to DEET was recorded using electroantennogram (EAG) technique. The response to DEET was recorded also after a long stimulation with the same repellent, and after topical application of SNAC and dbcGMP (a cGMP analogue) on the antennae. Results. We found that previous exposure of B. germanica males to the repellent DEET produced an increase of the repellency at the behavioural level, measured as RC. A possible role of nitric oxide (NO) in the transduction pathway of this phenomenon is suggested, since treatment of the cockroaches with the NO donor SNAC also produced an increase of the repellency elicited by DEET. On the other hand, the response of the cockroaches' antennae exposed to DEET was determined electrophysiologically. The electrical activity in response to DEET decreased when the insects' antennae were stimulated with a long pulse of the repellent. The activity of the antennae was restored after 10 min. Treatment of the antennae either with SNAC or dbGMPc also produced a decrease in the response of the antennae to the repellent. Discussion.The previous exposure to a chemical stimulus can modify the behaviour associated to the same stimulus, increasing or decreasing the behavioural response. In the case of DEET we found that pre-exposure increased DEET repellency in male cockroaches. We also found NO involvement in a similar phenomenon. On the other hand, the test showed that DEET is perceived by insects' antennae as an odour. A long exposure of the antennae to DEET caused a transient decrease in the response of the antennae to the same compound. The same effect was achieved by treating the antennae with SNAC or dbcGMP, suggesting the involvement of the NO/cGMP system in the transduction pathway of the sensory adaptation phenomenon elicited by an odour in this species.

Locomotor behaviour of Blattella germanica modified by DEET.

N,N-diethyl-3-methylbenzamide (DEET) is the active principle of most insect repellents used worldwide. However, its toxicity on insects has not been widely studied. The aim of this work is to study the effects of DEET on the locomotor activity of Blattella germanica. DEET has a dose-dependent repellent activity on B. germanica. Locomotor activity was significantly lower when insects were pre-exposed to 700 µg/cm(2) of DEET for 20 or 30 minutes, but it did not change when pre-exposure was shorter. Locomotor activity of insects that were pre-exposed to 2.000 µg/cm(2) of DEET for 10 minutes was significantly lower than the movement registered in controls. No differences were observed when insects were pre-exposed to lower concentrations of DEET. A 30-minute pre-exposure to 700 µg/cm(2) of DEET caused a significant decrease in locomotor activity. Movement was totally recovered 24 h later. The locomotor activity measured during the exposure to different concentrations of DEET remained unchanged. Insects with decreased locomotor activity were repelled to the same extent than control insects by the same concentration of DEET. We demonstrated that the repellency and modification of locomotor activity elicited by DEET are non-associated phenomena. We also suggested that the reduction in locomotor activity indicates toxicity of DEET, probably to insect nervous system.

Microgeographical study of insecticide resistance in Triatoma infestans from Argentina.

Chagas disease is a chronic parasitic infection restricted to America where it is currently estimated that 90 million people are at risk of acquiring the infection. Chemical control with pyrethroid insecticides has been effective to reduce disease transmission in several areas of the Southern Cone, although insecticide resistance has evolved and diminished the campaigns' results. Considering previous reports on the different levels of resistance between Triatoma infestans from different geographical areas, the objective of this work was to determine if T. infestans populations are toxicologically structured within localities. Response to the insecticide was measured and compared between houses of two Argentine localities. Different toxicity of deltamethrin was detected between dwellings of Chaco province, accounting for both susceptible and resistant houses within the same locality. However no difference was found among houses of Salta province. The results obtained in this work suggest that geographical structure is present not only at the between localities level, but also at the microgeograhical level.