The TIRS trial: Enrollment procedures and baseline characterization of a pediatric cohort to quantify the epidemiologic impact of targeted indoor residual spraying on Aedes-borne viruses in Merida, Mexico.

Aedes mosquito-borne viruses (ABVs) place a substantial strain on public health resources in the Americas. Vector control of Aedes mosquitoes is an important public health strategy to decrease or prevent spread of ABVs. The ongoing Targeted Indoor Residual Spraying (TIRS) trial is an NIH-sponsored clinical trial to study the efficacy of a novel, proactive vector control technique to prevent dengue virus (DENV), Zika virus (ZIKV), and chikungunya virus (CHIKV) infections in the endemic city of Merida, Yucatan, Mexico. The primary outcome of the trial is laboratory-confirmed ABV infections in neighborhood clusters. Despite the difficulties caused by the COVID-19 pandemic, by early 2021 the TIRS trial completed enrollment of 4,792 children aged 2-15 years in 50 neighborhood clusters which were allocated to control or intervention arms via a covariate-constrained randomization algorithm. Here, we describe the makeup and ABV seroprevalence of participants and mosquito population characteristics in both arms before TIRS administration. Baseline surveys showed similar distribution of age, sex, and socio-economic factors between the arms. Serum samples from 1,399 children were tested by commercially available ELISAs for presence of anti-ABV antibodies. We found that 45.1% of children were seropositive for one or more flaviviruses and 24.0% were seropositive for CHIKV. Of the flavivirus-positive participants, most were positive for ZIKV-neutralizing antibodies by focus reduction neutralization testing which indicated a higher proportion of participants with previous ZIKV than DENV infections within the cohort. Both study arms had statistically similar seroprevalence for all viruses tested, similar socio-demographic compositions, similar levels of Ae. aegypti infestation, and similar observed mosquito susceptibility to insecticides. These findings describe a population with a high rate of previous exposure to ZIKV and lower titers of neutralizing antibodies against DENV serotypes, suggesting susceptibility to future outbreaks of flaviviruses is possible, but proactive vector control may mitigate these risks.

Insecticide susceptibility status of Anopheles albimanus populations in historical malaria foci in Quintana Roo, Mexico.

BACKGROUND: Mexico has experienced a significant reduction in malaria cases over the past two decades. Certification of localities as malaria-free areas (MFAs) has been proposed as a steppingstone before elimination is achieved throughout the country. The Mexican state of Quintana Roo is a candidate for MFA certification. Monitoring the status of insecticide susceptibility of major vectors is crucial for MFA certification. This study describes the susceptibility status of Anopheles albimanus, main malaria vector, from historically important malaria foci in Quintana Roo, using both phenotypic and genotypic approaches. METHODS: Adult mosquito collections were carried out at three localities: Palmar (Municipality of Othon P. Blanco), Buenavista (Bacalar) and Puerto Morelos (Puerto Morelos). Outdoor human-landing catches were performed by pairs of trained staff from 18:00 to 22:00 during 3-night periods at each locality during the rainy season of 2022. Wild-caught female mosquitoes were exposed to diagnostic doses of deltamethrin, permethrin, malathion, pirimiphos-methyl or bendiocarb using CDC bottle bioassays. Mortality was registered at the diagnostic time and recovery was assessed 24 h after exposure. Molecular analyses targeting the Voltage-Gated Sodium Channel (vgsc) gene and acetylcholinesterase (ace-1) gene were used to screen for target site polymorphisms. An SNP analysis was carried out to identify mutations at position 995 in the vgsc gene and at position 280 in the ace-1 gene. RESULTS: A total of 2828 anophelines were collected. The main species identified were Anopheles albimanus (82%) and Anopheles vestitipennis (16%). Mortalities in the CDC bottle bioassay ranged from 99% to 100% for all the insecticides and mosquito species. Sequence analysis was performed on 35 An. albimanus across the three localities; of those, 25 were analysed for vgsc and 10 for ace-1 mutations. All individuals showed wild type alleles. CONCLUSION: The results demonstrated that An. albimanus populations from historical malaria foci in Quintana Roo are susceptible to the main insecticides used by the Ministry of Health.

Residual efficacy of the neonicotinoid insecticide clothianidin against pyrethroid-resistant Aedes aegypti.

BACKGROUND: Here we report the residual efficacy of the neonicotinoid insecticide clothianidin against pyrethroid-resistant Aedes aegypti. We first conducted a range-finding evaluation of clothianidin on three different substrates (wall, wood, cloth) using three doses (100, 300 and 600 mg a.i. m-2 ) and conducting World Health Organization (WHO) cone bioassays to assess acute (24 h) and delayed (up to 7 days) mortality. In experimental houses located in Merida (Mexico) and using free-flying pyrethroid-resistant Ae. aegypti females, we quantified the acute and delayed mortality after a 24-h exposure to the targeted indoor residual spraying (TIRS) of two clothianidin doses (100 and 300 mg a.i. m-2 ). RESULTS: Range-finding studies with WHO cones showed low (<50%) acute mortality for all surfaces, doses and times post spraying. Delayed mortality was higher, with average values above or close to the 60% mark (and 95% confidence interval estimates crossing 80% for the 600 mg a.i. m-2 dose). In experimental houses, a similar low acute mortality was quantified (range of mortality across 12 months was 2-44% for 100 mg a.i. m-2 and 8-61% for 300 mg a.i/m2 ). However, delayed mortality showed a strong effect of clothianidin on free-flying Ae. aegypti, with values above 80% up to 7 months post-TIRS. CONCLUSION: Novel residual insecticide molecules have a promising outlook for Ae. aegypti control and can contribute to the expansion and adoption of TIRS in urban areas. clothianidin can contribute to the control of resistant Ae. aegypti and provide residual control for up to 7 months after application. © 2022 Society of Chemical Industry.

Preventive residual insecticide applications successfully controlled Aedes aegypti in Yucatan, Mexico.

Insecticide-based approaches remain a key pillar for Aedes-borne virus (ABV, dengue, chikungunya, Zika) control, yet they are challenged by the limited effect of traditional outdoor insecticide campaigns responding to reported arboviral cases and by the emergence of insecticide resistance in mosquitoes. A three-arm Phase II unblinded entomological cluster randomized trial was conducted in Merida, Yucatan State, Mexico, to quantify the entomological impact of targeted indoor residual spraying (TIRS, application of residual insecticides in Ae. aegypti indoor resting sites) applied preventively 2 months before the beginning of the arbovirus transmission season. Trial arms involved the use of two insecticides with unrelated modes of action (Actellic 300CS, pirimiphos-methyl, and SumiShield 50WG, clothianidin) and a control arm where TIRS was not applied. Entomological impact was quantified by Prokopack adult collections performed indoors during 10 min per house. Regardless of the insecticide, conducting a preventive TIRS application led to significant reductions in indoor Ae. aegypti densities, which were maintained at the same levels as in the low arbovirus transmission period (Actellic 300CS reduced Ae. aegypti density up to 8 months, whereas SumiShield 50WG up to 6 months). The proportional reduction in Ae. aegypti abundance in treatment houses compared to control houses was 50-70% for Actellic 300CS and 43-63% for SumiShield 50WG. Total operational costs including insecticide ranged from US$4.2 to US$10.5 per house, depending on the insecticide cost. Conducting preventive residual insecticide applications can maintain Ae. aegypti densities at low levels year-round with important implications for preventing ABVs in the Americas and beyond.

Insecticide Susceptibility Tests of Aedes taeniorhynchus in Yucatan, Mexico.

We report results of susceptibility tests for Aedes taeniorhynchus from 2 localities of Yucatan State, Mexico, to different insecticides. The Centers for Disease Control and Prevention bottle bioassays were performed using the active ingredients of 3 pyrethroids, 2 organophosphates, and 1 carbamate: permethrin (15 µg/ml), deltamethrin (10 µg/ml), alpha-cypermethrin (10 µg/ml), malathion (50 µg/ml), chlorpyrifos (85 µg/ml), and bendiocarb (12.5 µg/ml). The mortality recorded at the diagnostic time of exposure (30 min) was 100% with all insecticides evaluated and for both populations. These results suggest complete susceptibility to the 3 chemical groups generally used for urban Ae. aegypti mosquito control.

Efficacy of targeted indoor residual spraying with the pyrrole insecticide chlorfenapyr against pyrethroid-resistant Aedes aegypti.

BACKGROUND: There is an increased need to mitigate the emergence of insecticide resistance and incorporate new formulations and modes of application to control the urban vector Aedes aegypti. Most research and development of insecticide formulations for the control of Ae. aegypti has focused on their peridomestic use as truck-mounted ULV-sprays or thermal fogs despite the widespread knowledge that most resting Ae. aegypti are found indoors. A recent modification of indoor residual spraying (IRS), termed targeted IRS (TIRS) works by restricting applications to 1.5 m down to the floor and on key Ae. aegypti resting sites (under furniture). TIRS also opens the possibility of evaluating novel residual insecticide formulations currently being developed for malaria IRS. METHODS: We evaluated the residual efficacy of chlorfenapyr, formulated as Sylando 240SC, for 12 months on free-flying field-derived pyrethroid-resistant Ae. aegypti using a novel experimental house design in Merida, Mexico. On a monthly basis, 600 female Ae. aegypti were released into the houses and left indoors with access to sugar solution for 24 hours. After the exposure period, dead and alive mosquitoes were counted in houses treated with chlorfenapyr as well as untreated control houses to calculate 24-h mortality. An evaluation for these exposed cohorts of surviving mosquitoes was extended up to seven days under laboratory conditions to quantify "delayed mortality". RESULTS: Mean acute (24-h) mortality of pyrethroid-resistant Ae. aegypti ranged 80-97% over 5 months, dropping below 30% after 7 months post-TIRS. If delayed mortality was considered (quantifying mosquito mortality up to 7 days after exposure), residual efficacy was above 90% for up to 7 months post-TIRS application. Generalized Additive Mixed Models quantified a residual efficacy of chlorfenapyr of 225 days (ca. 7.5 months). CONCLUSIONS: Chlorfenapyr represents a new option for TIRS control of Ae. aegypti in urban areas, providing a highly-effective time of protection against indoor Ae. aegypti females of up to 7 months.

Abundance and Seasonality of Aedes aegypti (Diptera: Culicidae) in Two Suburban Localities of South Mexico, With Implications for Wolbachia (Rickettsiales: Rickettsiaceae)-Carrying Male Releases for Population Suppression.

We conducted a baseline characterization of the abundance and seasonality of Aedes aegypti (Linnaeus, 1762)-a vector of dengue, chikungunya, and Zika-in two suburban localities of Yucatan, Mexico, as the first step in the implementation of an integrated vector management (IVM) plan combining 'traditional Aedes control' (source reduction/truck-mounted ultra-low volume [ULV] spraying) and incompatible insect technique/sterile insect technique for population suppression in Yucatan, Mexico. Weekly entomological collections with ovitraps and BG-sentinel traps were performed in 1-ha quadrants of both localities for 1 yr. Three distinct periods/phases were identified, closely associated with precipitation: 1) a phase of low population abundance during the dry season (weekly average of Aedes eggs per ovitrap and adults per BG trap = 15.51 ± 0.71 and 10.07 ± 0.88, respectively); 2) a phase of population growth and greatest abundance of Aedes (49.03 ± 1.48 eggs and 25.69 ± 1.31 adults) during the rainy season; and finally 3) a phase of decline among populations (20.91 ± 0.97 eggs and 3.24 ± 0.21 adults) after the peak of the rainy season. Seasonal abundance and dynamics of Ae. aegypti populations suggest that it is feasible to develop and implement time-specific actions as part of an IVM approach incorporating integrating novel technologies (such as rear-and-release of Wolbachia-infected males) with classic (insecticide-based) approaches implemented routinely for vector control. In agreement with the local vector control program, we propose a pilot IVM strategy structured in a preparation phase, an attack phase with traditional vector control, and a suppression phase with inundative releases, which are described in this paper.

Detection of Aedes (Stegomyia) albopictus (Skuse) in ovitraps of Mérida city, México / Detección de Aedes (Stegomyia) albopictus (Skuse) en ovitrampas en Mérida, México

Introduction: The vector-borne diseases program in México has an established network of ovitraps for entomological surveillance of Aedes spp. In response to reports of Aedes albopictus in the periphery of Mérida, the state capital of Yucatán, the Ministry of Health increased the specificity of this surveillance. Objective: To describe the presence and distribution of Ae. albopictus in Mérida and its relative abundance compared to Aedes aegypti in ovitraps of the vector control program. Materials and methods: During October, 2019, 91 ovitraps were randomly selected from 31 neighborhoods of Mérida. Mosquitoes were reared at the insectary of the Collaborative Unit for Entomological Bioassays of the Autonomous University of Yucatán from eggs collected in the field. Relative abundance was determined for adult individuals of each identified species and neighborhood. Results: 32 % of the neighborhoods were positive for Ae. albopictus and 100 % for Ae. aegypti. A total of 28 adults of Ae. albopictus (10 females and 18 males) were obtained from ovitraps. No correlation was observed between the abundance of Ae. aegypti and Ae. albopictus for both adults and females (p>0.05) at the neighborhood level. Conclusions: The results confirm that Ae. albopictus coexisted with Ae. aegypti in Mérida at the time of the study. The low relative abundance suggests that Ae. albopictus was in the initial phase of invasion.

Introducción. El programa de enfermedades transmitidas por vectores en México tiene una red establecida de ovitrampas para la vigilancia entomológica de Aedes spp. Los servicios de salud del estado de Yucatán, en respuesta a reportes de Aedes albopictus en la periferia de Mérida, capital del estado, incrementaron la especificidad de dicha vigilancia. Objetivo. Describir la presencia y distribución de Ae. albopictus en Mérida y su abundancia relativa comparada con Aedes aegypti, en ovitrampas del programa de control de vectores. Materiales y métodos. Durante octubre de 2019, se seleccionaron al azar 91 ovitrampas en 31 barrios de Mérida. Los mosquitos adultos se obtuvieron del insectario de la Unidad Colaborativa para Bioensayos Entomológicos de la Universidad Autónoma de Yucatán a partir de huevos recolectados en campo. Se determinó la abundancia relativa de individuos adultos de cada especie identificada y por barrios evaluados. Resultados. En el 32 % de los barrios muestreados, se detectó Ae. albopictus y, en todos ellos, Ae. aegypti. Se recolectaron 28 adultos de Ae. albopictus (10 hembras y 18 machos) en las ovitrampas. No se observó correlación entre la abundancia de adultos ni de hembras Ae. aegypti y Ae. albopictus por barrio (p>0,05). Conclusiones. Los resultados confirmaron que Ae. albopictus estaba coexistiendo con Ae. aegypti en Mérida en el momento del estudio. La baja abundancia relativa sugiere que Ae. albopictus se encontraba en la fase inicial de invasión.

Detección de Aedes (Stegomyia) albopictus (Skuse) en ovitrampas en Mérida, México / Detection of Aedes (Stegomyia) albopictus (Skuse) in ovitraps of Mérida city, México

Resumen | Introducción. El programa de enfermedades transmitidas por vectores en México tiene una red establecida de ovitrampas para la vigilancia entomológica de Aedes spp. Los servicios de salud del estado de Yucatán, en respuesta a reportes de Aedes albopictus en la periferia de Mérida, capital del estado, incrementaron la especificidad de dicha vigilancia. Objetivo. Describir la presencia y distribución de Ae. albopictus en Mérida y su abundancia relativa comparada con Aedes aegypti, en ovitrampas del programa de control de vectores. Materiales y métodos. Durante octubre de 2019, se seleccionaron al azar 91 ovitrampas en 31 barrios de Mérida. Los mosquitos adultos se obtuvieron del insectario de la Unidad Colaborativa para Bioensayos Entomológicos de la Universidad Autónoma de Yucatán a partir de huevos recolectados en campo. Se determinó la abundancia relativa de individuos adultos de cada especie identificada y por barrios evaluados. Resultados. En el 32 % de los barrios muestreados, se detectó Ae. albopictus y, en todos ellos, Ae. aegypti. Se recolectaron 28 adultos de Ae. albopictus (10 hembras y 18 machos) en las ovitrampas. No se observó correlación entre la abundancia de adultos ni de hembras Ae. aegypti y Ae. albopictus por barrio (p>0,05). Conclusiones. Los resultados confirmaron que Ae. albopictus estaba coexistiendo con Ae.aegypti en Mérida en el momento del estudio. La baja abundancia relativa sugiere que Ae.albopictus se encontraba en la fase inicial de invasión.

Abstract | Introduction:The vector-borne diseases program in México has an established network of ovitraps for entomological surveillance of Aedes spp. In response to reports of Aedes albopictus in the periphery of Mérida, the state capital of Yucatán, the Ministry of Health increased the specificity of this surveillance. Objective: To describe the presence and distribution of Ae. albopictus in Mérida and its relative abundance compared to Aedesaegypti in ovitraps of the vector control program. Materials and methods: During October, 2019, 91 ovitraps were randomly selected from 31 neighborhoods of Mérida. Mosquitoes were reared at the insectary of the Collaborative Unit for Entomological Bioassays of the Autonomous University of Yucatán from eggs collected in the field. Relative abundance was determined for adult individuals of each identified species and neighborhood. Results: 32 % of the neighborhoods were positive for Ae. albopictus and 100 % for Ae. aegypti. A total of 28 adults of Ae. albopictus (10 females and 18 males) were obtained from ovitraps. No correlation was observed between the abundance of Ae. aegypti and Ae. albopictus for both adults and females (p>0.05) at the neighborhood level. Conclusions: The results confirm that Ae. albopictus coexisted with Ae. aegypti in Mérida at the time of the study. The low relative abundance suggests that Ae. albopictus was in the initial phase of invasion.

The entomological impact of passive metofluthrin emanators against indoor Aedes aegypti: A randomized field trial.

BACKGROUND: In the absence of vaccines or drugs, insecticides are the mainstay of Aedes-borne disease control. Their utility is challenged by the slow deployment of resources, poor community compliance and inadequate household coverage. Novel application methods are required. METHODOLOGY AND PRINCIPAL FINDINGS: A 10% w/w metofluthrin "emanator" that passively disseminates insecticide from an impregnated net was evaluated in a randomized trial of 200 houses in Mexico. The devices were introduced at a rate of 1 per room and replaced at 3-week intervals. During each of 7 consecutive deployment cycles, indoor resting mosquitoes were sampled using aspirator collections. Assessments of mosquito landing behaviours were made in a subset of houses. Pre-treatment, there were no differences in Aedes aegypti indices between houses recruited to the control and treatment arms. Immediately after metofluthrin deployment, the entomological indices between the trial arms diverged. Averaged across the trial, there were significant reductions in Abundance Rate Ratios for total Ae. aegypti, female abundance and females that contained blood meals (2.5, 2.4 and 2.3-times fewer mosquitoes respectively; P<0.001). Average efficacy was 60.2% for total adults, 58.3% for females, and 57.2% for blood-fed females. The emanators also reduced mosquito landings by 90% from 12.5 to 1.2 per 10-minute sampling period (P<0.05). Homozygous forms of the pyrethroid resistant kdr alleles V410L, V1016L and F1534C were common in the target mosquito population; found in 39%, 24% and 95% of mosquitoes collected during the trial. CONCLUSIONS/SIGNIFICANCE: This is the first randomized control trial to evaluate the entomological impact of any volatile pyrethroid on urban Ae. aegypti. It demonstrates that volatile pyrethroids can have a sustained impact on Ae. aegypti population densities and human-vector contact indoors. These effects occur despite the presence of pyrethroid-resistant alleles in the target population. Formulations like these may have considerable utility for public health vector control responses.

Evaluating Over-the-Counter Household Insecticide Aerosols for Rapid Vector Control of Pyrethroid-Resistant Aedes aegypti.

Vector control methods that mobilize and impact rapidly during dengue, Zika, and chikungunya outbreaks are urgently needed in urban contexts. We investigated whether one person using a handheld aerosolized insecticide could achieve efficacy levels comparable to targeted indoor residual spraying (TIRS), using pyrethroid-resistant Aedes aegypti in a semi-field setting with experimental houses in Mexico. The insecticide product (H24, a carbamate and pyrethroid mixture), available over-the-counter locally, was sprayed only on known Ae. aegypti-resting surfaces, for example, walls less than 1.5 m and dark hidden areas. In six identical houses with paired bedrooms, one bedroom was treated, and the other remained an untreated control. Each week for 8 weeks, 100 female pyrethroid-resistant Ae. aegypti were released in each bedroom and followed up daily. Mortality rates in treated bedrooms exceeded 90% for at least 2 weeks, and more than 80% (89.2; 95% CI: 79.98-98.35) for 3 weeks or more. Mortality rates in control houses were zero. Results demonstrate that the immediate impact of TIRS can be delivered by one person using existing products, at an estimated cost for the average household in Mexico of under US$3 per month. Triggered by early outbreak signs, dissemination via community hubs and mass/social media of instructions to treat the home immediately, with monthly re-treatment thereafter, provides a simple means to engage and empower householders. Compatible with integrated vector management strategies, it enables self-protection even if existing agencies falter, a situation exemplified by the potential impact on vector control of the restrictions imposed during the 2020 COVID-19 pandemic.

Field Efficacy Trials of Aerial Ultra-Low-Volume Application of Insecticides Against Caged Aedes aegypti in Mexico.

We evaluated the efficacy of aerial ultra-low-volume (ULV) insecticide spraying in field bioassays with caged Aedes aegypti in May 2017 in Puerto Vallarta, Jalisco, Mexico. The insecticides tested included an organophosphate (Mosquitocida UNO ULV) and a neonicotinoid-pyrethroid combination (Cielo). Two Ae. aegypti populations were evaluated: a field pyrethroid-resistant local strain (Puerto Vallarta) and an insecticide-susceptible laboratory strain (New Orleans). Knockdown after 1 h by both products was ≥97.0%, and mortality after 24 h was ≥98% for the susceptible laboratory strain. Knockdown of the local Puerto Vallarta field strain by both products after 1 h was ≥96.5%; and mosquito mortality after 24 h was also very high (≥98%). Meteorological conditions during this evaluation were favorable for aerial mosquito control and represented conditions that typically occur during adulticide space spray applications. Temperature oscillated between 24°C and 26°C with winds between 6 and 10 km/h. The majority of droplets met the droplet distribution criteria required for the insecticides. The evaluation demonstrated an acceptable performance of both products for Ae. aegypti control when applied undiluted at a rate of 199.4 ml/ha and 73.07 ml/ha for Mosquitocida UNO ULV and Cielo, respectively. The volume median diameter (VMD) droplet size was characterized at 31.3 µm and 37.3 µm, respectively.

Experimental evaluation of the impact of household aerosolized insecticides on pyrethroid resistant Aedes aegypti.

The extensive reliance on insecticides to control Aedes aegypti mosquitoes and disrupt transmission of dengue, chikungunya and Zika has fueled the emergence of widespread resistance to insecticides. Mismatch between the frequency of pyrethroid resistance in mosquitoes and the occurrence of pyrethroid-based insecticide applications for vector control is often hypothesized to be due to household use of commercial insecticide products. We experimentally quantified phenotypic and genotypic responses of four Ae. aegypti strains (three field, pyrethroid resistant, and one laboratory, pyrethroid susceptible) after exposure to two commonly used household aerosol insecticide products (a space spray and a residual spray formulation) containing pyrethroid active ingredients. Experiments were performed within homes of Mérida, Mexico. After exposure to the products, all three pyrethroid resistant field Ae. aegypti strains had significantly lower mortality rates (averaging 41% and 50% for the two products, respectively) than the controls (99%). Applying insecticides as surface sprays led to a significant increase in the frequency of I1016 kdr homozygotes in surviving Ae. aegypti, suggesting strong selection pressure for this allele. Given the large-scale use of household aerosol insecticide products in areas that are endemic for Ae. aegypti-transmitted diseases, their role as a pyrethroid resistance selection source, particularly when used as space sprays, should be taken into consideration when designing resistance management plans.