Insights sobre a evolução e dispersão da resistência aos piretróides entre Triatoma infestans andinos silvestres da Bolívia

Abstract: Sylvatic populations of Triatoma infestans represent a challenge to Chagas disease control as they are not targeted by vector control activities and may play a key role in post-spraying house reinfestation. Understanding sylvatic foci distribution and gene flow between sylvatic and domestic populations is crucial to optimize vector control interventions and elucidate the development and spread of insecticide resistance. Herein, the genetic profiles of five Andean T. infestans populations from Bolivia with distinct insecticide susceptibility profiles were compared. Multilocus genotypes based on eight microsatellites and the DNA sequence of a fragment of the cytochrome B (cytB) gene were obtained for 92 individuals. CytB haplotypes were analyzed with previously reported Bolivian T. infestans haplotypes to evaluate putative historical gene flow among populations. Each specimen was also screened for two nucleotide mutations in the sodium channel gene (kdr), related to pyrethroid resistance (L1014 and L9251). Significant genetic differentiation was observed among all populations, although individuals of admixed origin were detected in four of them. Notably, the genetic profiles of adjacent domestic and sylvatic populations of Mataral, characterized by higher levels of insecticide resistance, support their common ancestry. Only one sylvatic individual from Mataral carried the kdr mutation L1014, suggesting that this mechanism is unlikely to cause the altered insecticide susceptibility observed in these populations. However, as the resistance mutation is present in the area, it has the potential to be selected under insecticidal pressure. Genetic comparisons of these populations suggest that insecticide resistance is likely conferred by ancient trait(s) in T. infestans sylvatic populations, which are capable of invading domiciles. These results emphasize the need for stronger entomological surveillance in the region, including early detection of house invasion, particularly post-spraying, monitoring for resistance to pyrethroids and the design of integrative control actions that consider sylvatic foci around domestic settings and their dispersion dynamics.

Improved vector control of Triatoma infestans limited by emerging pyrethroid resistance across an urban-to-rural gradient in the Argentine Chaco

Background: The sustainable elimination of Triatoma infestans in the Gran Chaco region represents an enduring challenge. Following the limited effects of a routine pyrethroid insecticide spraying campaign conducted over 2011­2013 (first period) in Avia Terai, an endemic municipality with approximately 2300 houses, we implemented a rapid-impact intervention package to suppress house infestation across the urban-to-rural gradient over 2015­2019 (second period). Here, we assess their impacts and whether persisting infestations were associated with pyrethroid resistance. Methods: The 2011­2013 campaign achieved a limited detection and spray coverage across settings (< 68%), more so during the surveillance phase. Following community mobilization and school-based interventions, the 2015­2019 program assessed baseline house infestation using a stratified sampling strategy; sprayed all rural houses with suspension concentrate beta-cypermethrin, and selectively sprayed infested and adjacent houses in urban and peri-urban settings; and monitored house infestation and performed selective treatments over the follow-up. Results: Over the first period, house infestation returned to pre-intervention levels within 3­4 years. The adjusted relative odds of house infestation between 2011­2013 and 2015­2016 differed very little (adj. OR: 1.17, 95% CI 0.91­1.51). Over the second period, infestation decreased significantly between 0 and 1 year post-spraying (YPS) (adj. OR: 0.36, 95% CI 0.28­0.46), with heterogeneous effects across the gradient. Mean bug abundance also dropped between 0 and 1 YPS and thereafter remained stable in rural and peri-urban areas. Using multiple regression models, house infestation and bug abundance at 1 YPS were 3­4 times higher if the house had been infested before treatment, or was scored as high-risk or non-participating. No low-risk house was ever infested. Persistent foci over two successive surveys increased from 30.0 to 59.3% across the gradient. Infestation was more concentrated in peridomestic rather than domestic habitats. Discriminating-dose bioassays showed incipient or moderate pyrethroid resistance in 7% of 28 triatomine populations collected over 2015­2016 and in 83% of 52 post-spraying populations. Conclusions: The intervention package was substantially more effective than the routine insecticide spraying campaign, though the effects were lower than predicted due to unexpected incipient or moderate pyrethroid resistance. Increased awareness and diagnosis of vector control failures in the Gran Chaco, including appropriate remedial actions, are greatly needed.

Pyrethroid Resistance Associated With a Decreased DEET Repellency in the Common Bed Bug (Hemiptera: Cimicidae)

The global resurgence of the bed bug (Cimex lectularius L.) and the widespread resistance of this insect to pyrethroid insecticides have created the need to find alternatives to chemical control. Thus, synthetic or natural repellents have been considered as a suitable choice to control bed bug infestations. Repellents not only keep insects away from their hosts, but also allow a decrease in insecticide selection pressure. The aim of this study was to evaluate the repellence effectivity of DEET against two bed bug colonies from Argentina (a susceptible one and a fieldcollected pyrethroid-resistant one), under two starving periods. We found different repellent effects of DEET on the two C. lectularius colonies and no differences between the two starving periods. In fact, DEET had a lower effect on the insecticide-resistant colony. The methodology used in this study proved to be useful to test the variation of the effect of DEET between pyrethroid-susceptible and -resistant bed bugs.

Beating the odds: Sustained Chagas disease vector control in remote indigenous communities of the Argentine Chaco over a seven-year period / Superando as probabilidades: controle sustentado do vetor da doença de Chagas em comunidades indígenas remotas do Chaco argentino durante um período de sete anos

Background: Rapid reinfestation of insecticide-treated dwellings hamper the sustained elimination of Triatoma infestans, the main vector of Chagas disease in the Gran Chaco region. We conducted a seven-year longitudinal study including community-wide spraying with pyrethroid insecticides combined with periodic vector surveillance to investigate the house reinfestation process in connection with baseline pyrethroid resistance, housing quality and household mobility in a rural section of Pampa del Indio mainly inhabited by deprived indigenous people (Qom). Methodology/principal findings: Despite evidence of moderate pyrethroid resistance in local T. infestans populations, house infestation dropped from 31.9% at baseline to 0.7% at 10 months post-spraying (MPS), with no triatomine found at 59 and 78 MPS. Household-based surveillance corroborated the rare occurrence of T. infestans and the house invasion of other four triatomine species. The annual rates of loss of initially occupied houses and of household mobility were high (4.6-8.0%). Housing improvements did not translate into a significant reduction of mud-walled houses and refuges for triatomines because most households kept the former dwelling or built new ones with mud walls. Conclusions/significance: Our results refute the assumption that vector control actions performed in marginalized communities of the Gran Chaco are doomed to fail. The larger-than-expected impacts of the intervention program were likely associated with the combined effects of high-coverage, professional insecticide spraying followed by systematic vector surveillance-and-response, broad geographic coverage creating a buffer zone, frequent housing replacement and residential mobility. The dynamical interactions among housing quality, mobility and insecticide-based control largely affect the chances of vector elimination.

Toxicological, Enzymatic, and Molecular Assessment of the Insecticide Susceptibility Profile of Triatoma infestans (Hemiptera: Reduviidae, Triatominae) Populations From Rural Communities of Santa Cruz, Bolivia

A wide range of insecticide resistance profiles has been reported across Bolivian domestic and sylvatic populations of Triatoma infestans (Klug, 1834) (Hemiptera, Reduviidae), including some with levels proven to be a threat for vector control. In this work, the insecticide profile of domestic T. infestans was studied with standardized toxicological bioassays, in an area that has not undergone consistent vector control. F1 first-instar nymphs hatched in laboratory from bugs captured in three communities from the Santa Cruz Department were evaluated with different insecticides. Moreover, the enzymatic activity of esterases and cytochrome P450 monooxygenases was measured in individual insects to evaluate the possible mechanism of metabolic resistance to pyrethroids. In addition, the DNA sequence of sodium channel gene (kdr) was screened for two point mutations associated with pyrethroid resistance previously reported in T. infestans. All populations showed reduced susceptibility to deltamethrin and α-cypermethrin, albeit the RR50 values varied significantly among them. Increased P450 monooxygenases and permethrate esterases suggest the contribution, as detoxifying mechanisms, to the observed resistance to deltamethrin in all studied populations. No individuals presented either mutation associated to resistance in the kdr gene. The level of susceptibility to α-cypermethrin, the insecticide used by the local vector control program, falls within an acceptable range to continue its use in these populations. However, the observed RR50 values evidence the possibility of selection for resistance to pyrethroids, especially to deltamethrin. Consequently, the use of pyrethroid insecticides should be closely monitored in these communities, which should be kept under entomological surveillance and sustained interventions.

Analysing deltamethrin susceptibility and pyrethroid esterase activity variations in sylvatic and domestic Triatoma infestans at the embryonic stage

The aim of the present work was to study the deltamethrin susceptibility of eggs from Triatoma infestans populations and the contribution of pyrethroid esterases to deltamethrin degradation. Insects were collected from sylvatic areas, including Veinte de Octubre and Kirus-Mayu (Bolivia) and from domiciliary areas, including El Palmar (Bolivia) and La Pista (Argentina). Deltamethrin susceptibility was determined by dose-response bioassays. Serial dilutions of deltamethrin (0.0005-1 mg/mL) were topically applied to 12-day-old eggs. Samples from El Palmar had the highest lethal dose ratio (LDR) value (44.90) compared to the susceptible reference strain (NFS), whereas the Veinte de Octubre samples had the lowest value (0.50). Pyrethroid esterases were evaluated using 7-coumaryl permethrate (7-CP) on individually homogenised eggs from each population and from NFS. The El Palmar and La Pista samples contained 40.11 and 36.64 pmol/min/mg protein, respectively, and these values were statistically similar to NFS (34.92 pmol/min/mg protein) and different from Kirus-Mayu and Veinte de Octubre (27.49 and 22.69 pmol/min/mg protein, respectively). The toxicological data indicate that the domestic populations were resistant to deltamethrin, but no statistical contribution of 7-CP esterases was observed. The sylvatic populations had similar LDR values to NFS, but lower 7-CP esterase activities. Moreover, this is the first study of the pyrethroid esterases on T. infestans eggs employing a specific substrate (7-CP).

Toxicological and biochemical analysis of the susceptibility of sylvatic Triatoma infestans from the Andean Valley of Bolivia to organophosphate insecticide

To increase our knowledge of the natural susceptibility of Triatoma infestans to an organophosphate insecticide, we performed toxicological and biochemical studies on three sylvatic populations from Bolivia and two populations from domestic dwellings from Bolivia and Argentina. Fifty-per-cent lethal doses (LD50) were determined based on the topical application of fenitrothion on first instar nymphs and mortality was assessed at 24 h. Both type of populations exhibited LD50ratios significantly higher than 1 with a range of the values (1.42-2.47); the maximum value were found in a sylvatic (-S) population, Veinte de Octubre-S. Samples were biochemically analysed using a glutathione S-transferase activity assay. The highest significant activity was obtained for Veinte de Octubre-S and the lowest activity was obtained for the reference population (102.69 and 54.23 pmol per minute per mg of protein respectively). Two out of the three sylvatic populations (Veinte de Octubre-S and Kirus Mayu-S) exhibited significantly higher glutathione S-transferase activity than that of the reference population. Based on this analysis of the natural susceptibility of this organism to organophosphate insecticides, continental and focal surveys of organophosphate susceptibility should be conducted to evaluate the evolution and distribution of this phenomenon.

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 æM S./i.min.) and Salta resistant colony (9.5 ± 0.8 æM S./i.min.). Cytochrome P450 mono-oxygenase (P450) activity was measured on individual insects through ethoxycoumarine deethylase (ECOD) activity using a fluorescence micro plate 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.

Resistencia a insecticidas en Blattella Germánica de Buenos Aires / Resistance to insecticides in Blatella germanica from Buenos Aires

Se determinó la línea base de susceptibilidad a delta deltametrina, cipermetrina, cxlorpirifós y fenitrotión en ninfas del primer estadio de la cucaracha alemana, Blatella germanica, provenintes de una población susceptible a insecticidas mantenida en laboratorio (cepa CIPEIN). En base a los valores de DL50, se observó el siguiente orden decrteciente de toxicidad: deltametrina > clorpirifós > cipewrmetrina fenitrotión. La DL99 de cada uno de estos insecticiodas fue aplicada a la descendencia de individuos recolectados en cuatro sitios de Buenos Aires y a individuos de una cepa resistente a deltametrina criada en laboratorio desde 1994. En todos los casos, al menos un insecticida produjop menor mortalidad que la observada en la cepa susceptible, sigiriendo la excistencia de resistencia. Se obtuvieron valores de DL50 y los Grados de Resistencia (GR) a deltametrina para cada muestra. En todos los casos se observó resistencia a este insecticida, con GR que variaron entre 7,3 y 75,4.

Efectividad de lociones capilares sobre poblaciones de Pediculus capitis resistentes a insecticidas / Effectiveness of capilar lotions against insecticide resistant Pediculus capitis populations

Se estudio la influencia de los distintos componentes en la efectividad de lociones experimentales sobre piojos resistentes a insecticidas piretroides. Se encontró que el aumento de la concentración del insecticida D-fenotrina desde 0,2 por ciento (concentración recomendada en lociones comerciales) hasta 0,5 por ciento, no produjo diferencias significativas (p=0,495) en la actividad pediculicida. Por el contrario se encontró un aumento significativo de efectividad de lociones experimentales con d-fenotrina o,2 por ciento al agregar alcohol alifático dodecanol en concentración 10 por ciento. El volteo de piojos expuestos se incrementó desde 23,+- 4,7 por ciento hasta 92,0+- 7,5 por ciento respectivamente. Se encontró además que las lociones conteniendo sólo dodecanol (sin d-fenotrina) producian un volteo significativo de los piojos expuestos (35+-8,5). Se comprobó que el volteo producido por la loción con dodecanol mostraba recuperación parcial a las 18 horas, mientras que la efectividad de las lociones que combinan dodecanol con insecticidas aumentaba al cabo de 18 horas alcanzando 100 por ciento de mortalidad. Se postula que la potenciación de actividad pediculicida demostrada para lociones que combinan dodecanol y d-fenotrina podría deberse a daño cuticular producido por el alcohol que favorecería la penetración del insecticida para ejercer su efecto tóxico