Pyrethroid susceptibility reversal in Aedes aegypti: A longitudinal study in Tapachula, Mexico.

Pyrethroid resistance in Aedes aegypti has become widespread after almost two decades of frequent applications to reduce the transmission of mosquito-borne diseases. Because few insecticide classes are available for public health use, insecticide resistance management (IRM) is proposed as a strategy to retain their use. A key hypothesis of IRM assumes that negative fitness is associated with resistance, and when insecticides are removed from use, susceptibility is restored. In Tapachula, Mexico, pyrethroids (PYRs) were used exclusively by dengue control programs for 15 years, thereby contributing to selection for high PYR resistance in mosquitoes and failure in dengue control. In 2013, PYRs were replaced by organophosphates-insecticides from a class with a different mode of action. To test the hypothesis that PYR resistance is reversed in the absence of PYRs, we monitored Ae. aegypti's PYR resistance from 2016 to 2021 in Tapachula. We observed significant declining rates in the lethal concentration 50 (LC50), for permethrin and deltamethrin. For each month following the discontinuation of PYR use by vector control programs, we observed increases in the odds of mosquitoes dying by 1.5% and 8.4% for permethrin and deltamethrin, respectively. Also, knockdown-resistance mutations (kdr) in the voltage-gated sodium channel explained the variation in the permethrin LC50s, whereas variation in the deltamethrin LC50s was only explained by time. This trend was rapidly offset by application of a mixture of neonicotinoid and PYRs by vector control programs. Our results suggest that IRM strategies can be used to reverse PYR resistance in Ae. aegypti; however, long-term commitment by operational and community programs will be required for success.

Evaluation of commercial aerosol insecticides for control of Aedes aegypti susceptible or resistant to pyrethroids.

OBJECTIVE: To evaluate indoor use of commercial aerosols for dengue vector mosquito control, and estimate the number of treatable houses per can. MATERIALS AND METHODS: Four aerosol products containing combinations of pyrethroids (two containing propoxur and one containing synergists too), were evaluated with mosquitoes in a room of a Tapachulastyle house. Eight cages containing 20 insecticide susceptible or resistant females were hung from tripods, another set was placed in sheltered areas of the room. From the entrance of the room, one of 4-9 concentrations was sprayed for each aerosol, leaving the mosquitoes for 30 min after sprayed. Mortality was recorded after 24 h and lethal concentrations were calculated. RESULTS: Aerosol A had the highest LC50, with 0.308 g for mosquitoes hanging from tripods and 0.453 g for sheltered mosquitoes; followed by aerosols C, D and B, with statistical differences between types of exposure. CONCLUSIONS: Aerosols B-D could spray 20-25 3-room houses (56 m3-room), killing all resistant mosquitoes. Aerosols may become a good tool for indoor mosquito control, if the optimal concentration and correct spray method are used.

Aedes aegypti, Ae. albopictus and Culex quinquefasciatus Adults Found Coexisting in Urban and Semiurban Dwellings of Southern Chiapas, Mexico.

Tapachula, Mexico, a tropical city, is an endemic area for dengue, in addition to several outbreaks in the last decade with chikungunya and zika. As part of the migratory corridor from Central to North America and the risks of scattered infectious diseases that this implies, the identification and distribution of potential disease vectors in and around residential areas are essential in terms of entomological surveillance for the prevention of disease outbreaks. The identification of mosquito species of medical importance coexisting in houses and cemeteries in Tapachula and two semiurban sites in southern Chiapas was investigated. Adult mosquitoes were collected from May to December 2018, resting inside and outside houses and in the tombstones and fallen tree leaves in cemeteries. A total of 10,883 mosquitoes belonging to three vector species were collected across 20 sites; 6738 were from neighborhood houses, of which 55.4% were Culex quinquefasciatus, 41.6% Aedes aegypti, and 2.9% Ae. albopictus. Aedes aegypti was the most common mosquito resting inside houses (56.7%), while Ae. albopictus and Cx. quinquefasciatus were mostly found resting outside houses (75.7%). In the cemeteries, Cx. quinquefasciatus (60.8%) and Ae. albopictus (37.3%) were the most abundant, while Ae. aegypti (1.9%) was the least abundant. This is the first report to identify adults of three major disease vector species coexisting in the domestic environment of urban and semiurban sites and Ae. albopictus adult resting inside of urban houses in Mexico. It would be opportune to consider comprehensive strategies that can be applied in this region to control the three species at the same time and avoid outbreaks of the diseases they transmit.

Resistance to Pyrethroids in the Malaria Vector Anopheles albimanus in Two Important Villages in the Soconusco Region of Chiapas, Mexico, 2022.

Chiapas State comprises the largest malaria foci from Mexico, and 57% of the autochthonous cases in 2021, all with Plasmodium vivax infections, were reported in this State. Southern Chiapas is at constant risk of cases imported due to migratory human flow. Since chemical control of vector mosquitoes is the main entomological action implemented for the prevention and control of vector-borne diseases, this work aimed to investigate the susceptibility of Anopheles albimanus to insecticides. To this end, mosquitoes were collected in cattle in two villages in southern Chiapas in July-August 2022. Two methods were used to evaluate the susceptibility: the WHO tube bioassay and the CDC bottle bioassay. For the latter, diagnostic concentrations were calculated. The enzymatic resistance mechanisms were also analyzed. CDC diagnostic concentrations were obtained; 0.7 µg/mL deltamethrin, 12 µg/mL permethrin, 14.4 µg/mL malathion, and 2 µg/mL chlorpyrifos. Mosquitoes from Cosalapa and La Victoria were susceptible to organophosphates and to bendiocarb, but resistant to pyrethroids, with mortalities between 89% and 70% (WHO), and 88% and 78% (CDC), for deltamethrin and permethrin, respectively. High esterase levels are suggested as the resistance mechanism involved in the metabolism of pyrethroids in mosquitoes from both villages. Mosquitoes from La Victoria might also involve cytochrome P450. Therefore, organophosphates and carbamates are suggested to currently control An. albimanus. Its use might reduce the frequency of resistance genes to pyrethroids and vector abundance and may impede the transmission of malaria parasites.

Current enzyme-mediated insecticide resistance status of Aedes aegypti populations from a dengue-endemic city in southern Mexico.

OBJECTIVE: To identify the enzyme-mediated insecticide resistance in Aedes aegypti in Tapachula, Mexico. MATERIALS AND METHODS: Biochemical assays were undertaken to determine the enzyme levels in mosquitoes from 22 sites collected in 2018 and 2020 in Tapachula. Results of 2018 were correlated with the resistance to insecticides pub-lished. RESULTS: Mosquitoes had higher levels than those of the susceptible strain in 2018 and 2020 respectively of α-esterases in 15 and 12 sites; ß-esterases in 7 and 6 sites; glutathione-S-transferases in 11 and 19 sites; ρNPA-esterases in 21 and 17 sites; and cytochromes P450 in 20 and 22 sites. In mosquitoes of 2018, there was a moderate correlation between previously documented Malathion resistance ratios and the insensitive acetylcholinesterase (r=0.459, p= 0.03). CONCLUSIONS: The elevated enzyme levels found indicate its contribution to the resistance to pyrethroids and organo-phosphates already published in mosquitoes from Tapachula. Bioassays using enzyme inhibitors resulted in greater mor-tality, confirming that metabolism contributes to resistance.

Insecticide resistance in Aedes aegypti from Tapachula, Mexico: Spatial variation and response to historical insecticide use.

BACKGROUND: Insecticide use continues as the main strategy to control Aedes aegypti, the vector of dengue, Zika, chikungunya, and yellow fever. In the city of Tapachula, Mexico, mosquito control programs switched from pyrethroids to organophosphates for outdoor spatial spraying in 2013. Additionally, the spraying scheme switched from total coverage to focused control, prioritizing areas with higher entomological-virological risk. Five years after this strategy had been implemented, we evaluated the status and variability of insecticide resistance among Ae. aegypti collected at 26 sites in Tapachula. METHODOLOGY/PRINCIPAL FINDINGS: We determined the lethal concentrations at 50% of the tested populations (LC50) using a bottle bioassay, and then, we calculated the resistance ratio (RR) relative to the susceptible New Orleans strain. Permethrin and deltamethrin (pyrethroids), chlorpyrifos and malathion (organophosphates), and bendiocarb (carbamate) were tested. The frequencies of the substitutions V1016I and F1534C, which are in the voltage-gated sodium channel and confer knockdown-resistance (kdr) to pyrethroid insecticides, were calculated. Despite 5 years having passed since the removal of pyrethroids from the control programs, Ae. aegypti remained highly resistant to permethrin and deltamethrin (RR > 10-fold). In addition, following 5 years of chlorpyrifos use, mosquitoes at 15 of 26 sites showed moderate resistance to chlorpyrifos (5- to 10-fold), and the mosquitoes from one site were highly resistant. All sites had low resistance to malathion (< 5-fold). Resistance to bendiocarb was low at 19 sites, moderate at five, and high at two. Frequencies of the V1016I ranged from 0.16-0.71, while C1534 approached fixation at 23 sites (0.8-1). Resistance profiles and kdr allele frequencies varied across Tapachula. The variability was not associated with a spatial pattern at the scale of the sampling. CONCLUSION/SIGNIFICANCE: Mosquito populations respond to selection pressure at a focal scale in the field. Spatial variation across sites highlights the importance of testing multiple sites within geographical regions.

Permethrin resistance in Aedes aegypti: Genomic variants that confer knockdown resistance, recovery, and death.

Pyrethroids are one of the few classes of insecticides available to control Aedes aegypti, the major vector of dengue, chikungunya, and Zika viruses. Unfortunately, evolving mechanisms of pyrethroid resistance in mosquito populations threaten our ability to control disease outbreaks. Two common pyrethroid resistance mechanisms occur in Ae. aegypti: 1) knockdown resistance, which involves amino acid substitutions at the pyrethroid target site-the voltage-gated sodium channel (VGSC)-and 2) enhanced metabolism by detoxification enzymes. When a heterogeneous population of mosquitoes is exposed to pyrethroids, different responses occur. During exposure, a proportion of mosquitoes exhibit immediate knockdown, whereas others are not knocked-down and are designated knockdown resistant (kdr). When these individuals are removed from the source of insecticide, the knocked-down mosquitoes can either remain in this status and lead to dead or recover within a few hours. The proportion of these phenotypic responses is dependent on the pyrethroid concentration and the genetic background of the population tested. In this study, we sequenced and performed pairwise genome comparisons between kdr, recovered, and dead phenotypes in a pyrethroid-resistant colony from Tapachula, Mexico. We identified single-nucleotide polymorphisms (SNPs) associated with each phenotype and identified genes that are likely associated with the mechanisms of pyrethroid resistance, including detoxification, the cuticle, and insecticide target sites. We identified high association between kdr and mutations at VGSC and moderate association with additional insecticide target site, detoxification, and cuticle protein coding genes. Recovery was associated with cuticle proteins, the voltage-dependent calcium channel, and a different group of detoxification genes. We provide a list of detoxification genes under directional selection in this field-resistant population. Their functional roles in pyrethroid metabolism and their potential uses as genomic markers of resistance require validation.

Permethrin Resistance Status and Associated Mechanisms in Aedes albopictus (Diptera: Culicidae) From Chiapas, Mexico.

There are major public health concerns regarding the spread of mosquito-borne diseases such as dengue, Zika, and chikungunya, which are mainly controlled by using insecticides against the vectors, Aedes aegypti (Linnaeus) and Aedes albopictus (Skuse). Pyrethroids are the primary class of insecticides used for vector control, due to their rapid knockdown effect and low toxicity to vertebrates. Unfortunately, continued use of pyrethroids has led to widespread insecticide resistance in Ae. aegypti; however, we lack information for Ae. albopictus-a sympatric species in Chiapas since 2002. In this study, we evaluated the permethrin resistance status of Ae. albopictus collected from Mexico and Texas. We also selected for permethrin resistance in the laboratory and investigated the potential mechanisms conferring resistance in this species. Knockdown resistance mutations, specifically F1534C, in the voltage-gated sodium channel gene, and increased activity of detoxifying enzymes were evaluated. Low levels of permethrin resistance (<2.4-fold) were observed in our field populations of Ae. albopictus and the F1534C mutation was not detected in any of the sites. Low levels of resistance were also observed in the artificially selected strain. There was significantly higher cytochrome P450 activity in our permethrin-selected and nonselected strains from Mexico compared to the control strain. Our results suggest the Ae. albopictus sampled from 2016 are mostly susceptible to pyrethroids. These results contrast with the high levels of permethrin resistance (>58-fold) found in Ae. aegypti from the same sites in Mexico. This research indicates the importance of continued monitoring of Ae. albopictus populations to prevent resistance from developing in the future.

Resistencia a insecticidas en Aedes aegypti y Aedes albopictus (Diptera: Culicidae) de Tapachula, Chiapas, México / Insecticide resistance in Aedes aegypti and Ae. albopictus (Diptera: Culicidae) populations from Tapachula, Chiapas, Mexico

Resumen: Objetivo: Determinar la resistencia a insecticidas en Ae. aegypti y Ae. albopictus de Tapachula, Chiapas, México. Material y métodos: Se utilizaron ovitrampas para obtener huevos de mosquitos Aedes y se realizaron pruebas de susceptibilidad (CDC) y ensayos enzimáticos con la primera generación. Resultados: Aedes aegypti mostró resistencia a deltametrina, permetrina, malatión, clorpirifos, temefos y a bendiocarb (CARB), mientras que Aedes albopictus a malatión y en menor grado a cloripirifos, temefos, permetrina y deltametrina. Ambas especies mostraron altos niveles de enzimas como citocomo P450 y glutatión S-tranferasa, mientras que los niveles de esterasas variaron por especie y sitio muestreado. Se detectó acetilcolinesterasa insensible a insecticidas en ambas especies. Conclusión: En un hábitat urbano de Tapachula, Chiapas, México donde se aplica control con insecticidas Ae. aegypti y Ae. albopictus sólo son susceptibles al propoxur.

Abstract: Objective: To determine the insecticide resistance status of Ae. aegypti and Ae. albopictus from Tapachula, México. Materials and methods: Mosquito eggs were collected with the use of ovitraps and CDC susceptibility bioassays and biochemical assays were conducted to determine resistance levels and resistance mechanisms, respectively. Results: Ae. aegypti showed resistance to deltamethrin and permethrin (PYRs), malathion, chlorpyrifos and temephos (OP), and to bendiocarb (CARB), while Ae. albopictus showed resistance to malathion and to a lesser intensity to chlorypirifos, temephos, permethrin and deltamethrin. Both species showed high levels of P450 and GSTs, while levels of esterases varied by species and collection site. Altered acethilcholinesterase was detected in both species. Conclusion: In an urban habitat from Tapachula, Chiapas, Mexico where vector control using insecticides takes place, Ae. aegypti and Ae. albopictus are only susceptible to propoxur.

[Insecticide resistance in Aedes aegypti and Ae. albopictus (Diptera: Culicidae) populations from Tapachula, Chiapas, Mexico]. / Resistencia a insecticidas en Aedes aegypti y Aedes albopictus (Diptera: Culicidae) de Tapachula, Chiapas, México.

OBJECTIVE: To determine the insecticide resistance status of Ae. aegypti and Ae. albopictus from Tapachula, México. MATERIALS AND METHODS: Mosquito eggs were collected with the use of ovitraps and CDC susceptibility bioassays and biochemical assays were conducted to determine resistance levels and resistance mechanisms, respectively. RESULTS: Ae. aegypti showed resistance to deltamethrin and permethrin (PYRs), malathion, chlorpyrifos and temephos (OP), and to bendiocarb (CARB), while Ae. albopictus showed resistance to malathion and to a lesser intensity to chlorypirifos, temephos, permethrin and deltamethrin. Both species showed high levels of P450 and GSTs, while levels of esterases varied by species and collection site. Altered acethilcholinesterase was detected in both species. CONCLUSIONS: In an urban habitat from Tapachula, Chiapas, Mexico where vector control using insecticides takes place, Ae. aegypti and Ae. albopictus are only susceptible to propoxur.

OBJETIVO: Determinar la resistencia a insecticidas en Ae. aegypti y Ae. albopictus de Tapachula, Chiapas, México. MATERIAL Y MÉTODOS: Se utilizaron ovitrampas para obtener huevos de mosquitos Aedes y se realizaron pruebas de susceptibilidad (CDC) y ensayos enzimáticos con la primera generación. RESULTADOS: Aedes aegypti mostró resistencia a deltametrina, permetrina, malatión, clorpirifos, temefos y a bendiocarb (CARB), mientras que Aedes albopictus a malatión y en menor grado a cloripirifos, temefos, permetrina y deltametrina. Ambas especies mostraron altos niveles de enzimas como citocomo P450 y glutatión S-tranferasa, mientras que los niveles de esterasas variaron por especie y sitio muestreado. Se detectó acetilcolinesterasa insensible a insecticidas en ambas especies. CONCLUSIONES: En un hábitat urbano de Tapachula, Chiapas, México donde se aplica control con insecticidas Ae. aegypti y Ae. albopictus sólo son susceptibles al propoxur.

Global genetic diversity of Aedes aegypti.

Mosquitoes, especially Aedes aegypti, are becoming important models for studying invasion biology. We characterized genetic variation at 12 microsatellite loci in 79 populations of Ae. aegypti from 30 countries in six continents, and used them to infer historical and modern patterns of invasion. Our results support the two subspecies Ae. aegypti formosus and Ae. aegypti aegypti as genetically distinct units. Ae. aegypti aegypti populations outside Africa are derived from ancestral African populations and are monophyletic. The two subspecies co-occur in both East Africa (Kenya) and West Africa (Senegal). In rural/forest settings (Rabai District of Kenya), the two subspecies remain genetically distinct, whereas in urban settings, they introgress freely. Populations outside Africa are highly genetically structured likely due to a combination of recent founder effects, discrete discontinuous habitats and low migration rates. Ancestral populations in sub-Saharan Africa are less genetically structured, as are the populations in Asia. Introduction of Ae. aegypti to the New World coinciding with trans-Atlantic shipping in the 16th to 18th centuries was followed by its introduction to Asia in the late 19th century from the New World or from now extinct populations in the Mediterranean Basin. Aedes mascarensis is a genetically distinct sister species to Ae. aegypti s.l. This study provides a reference database of genetic diversity that can be used to determine the likely origin of new introductions that occur regularly for this invasive species. The genetic uniqueness of many populations and regions has important implications for attempts to control Ae. aegypti, especially for the methods using genetic modification of populations.