Mosquito-transmitted viruses such as dengue are a global and growing public health challenge. Without widely available vaccines, mosquito control is the primary tool for fighting the spread of these viruses. New mosquito control technologies are needed to complement existing methods, given current challenges with scalability, acceptability, and effectiveness. A field trial was conducted in collaboration with the Communities Organized to Prevent Arboviruses project in Ponce, Puerto Rico, to measure entomological and epidemiological effects of reducing Aedes aegypti populations using Wolbachia incompatible insect technique. We packed and shipped Wolbachia-males from California and released them into 19 treatment clusters from September 2020 to December 2020. Preliminary evaluation revealed sub-optimal Wolbachia-male densities and impact on the wild-type population. In 2021, we shifted to a phased release strategy starting in four clusters, reducing the mosquito population by 49% (CI 29-63%). We describe the investigation into male quality and other factors that may have limited the impact of Wolbachia-male releases. Laboratory assays showed a small but significant impact of packing and shipping on male fitness. However, mark-release-recapture assessments suggest that male daily survival rates in the field may have been significantly impacted. We compared induced-sterility levels and suppression of the wild population and found patterns consistent with mosquito population compensation in response to our intervention. Analysis of epidemiological impact was not possible due to very low viral transmission rates during the intervention period. Our entomological impact data provide evidence that Wolbachia incompatible-male releases reduced Ae. aegypti populations, although efficacy will be maximized when releases are part of an integrated control program. With improvement of shipping vessels and shipped male fitness, packing and shipping male mosquitoes could provide a key solution for expanding access to this technology. Our project underscores the challenges involved in large and complex field effectiveness assessments of novel vector control methods.

Mass-trapping has been used to control outbreaks of Aedes aegypti (Linnaeus) (Diptera: Culicidae) in Puerto Rico since 2011. We investigated the effect of multi-year, insecticide-free mass trapping had on the insecticide susceptibility profile of Ae. aegypti. Eggs collected in southern Puerto Rico were used to generate F1 populations that were tested for susceptibility to permethrin, sumethrin, bifenthrin, deltamethrin, and malathion according to CDC bottle bioassays protocols. All populations of Ae. aegypti were resistant to the synthetic pyrethroids and mosquitoes from two locations were partially resistant to malathion. Population genetic analysis, using a double digest restriction sites associated DNA sequencing (ddRADseq) approach, indicated a large amount of migration between study sites effectively homogenizing the mosquito populations. Mass-trapping using noninsecticidal autocidal gravid ovitraps did not restore susceptibility to five active ingredients that are found in commercial insecticides. Migration between communities was high and would have brought outside alleles, including resistant alleles to the treatment communities. Further investigation suggests that household use of commercially available insecticide products may continue to select for resistance in absence of public health space spraying of insecticides.

To improve detection and assessment of Aedes aegypti abundance, we investigated whether microhabitat factors of the location of autocidal gravid ovitraps (AGO traps) influenced captures of gravid females in 2 locations in southern Puerto Rico. One location had been under vector control for several years using mass AGO trapping (intervention site), where Ae. aegypti abundance was several times lower than in the other study site without mosquito control (nonintervention site). We observed 10 environmental factors describing trap microhabitat location, and monitored water volume and minimum, maximum, and average temperature in AGO traps. Air temperature, relative humidity, and rainfall were recorded at each site. We conducted a hot-spot analysis of AGO traps to understand whether trap captures were influenced by the local abundance of mosquitoes rather than or in addition to trap microhabitat factors. AGO traps were classified using a 2-step cluster analysis based on attributes of trap microhabitats, water temperature, and water volume. Captures of female Ae. aegypti in each cluster per site were compared between resulting clusters to determine whether trap microhabitat factors defining the clusters were associated with trap captures. Trap captures in both study sites were mostly correlated with captures in nearby traps regardless of trap microhabitat factors, possibly reflecting the influence of the spatial aggregation of mosquitoes coming from nearby aquatic habitats or the concentration of dispersing adults. These results indicated that AGO traps can be located at places that can be easily reached during periodic inspections, such as in front of houses, without much regard to local microhabitat conditions.