use of altered male mosquitoes to dec dengue transmission

 a recent study found that mating Aedes aegypti female mosquitoes that can carry dengue virus with Wolbachia-infected males dramatically decreased human dengue infections  (see dengue mosquitos infected with other dec dengue transmission in dropbox, or DOI: 10.1056/NEJMoa2503304)

Details:

-- this study in Singapore, a tropical city-state, was designed to make female A. aegypti mosquitoes infertile by releasing male infected with the wAlbB strain of wolbachia bacteria, in 2022 during weeks 30 to 37. The trial was concluded in week 37 in 2024. (these weeks were considered to be "epidemiologic weeks" to compare data over time without calendar date fluctuations)

-- this was a cluster-randomized trial with test-negative controls performed in 15 geographic population clusters. All clusters were in urban areas; 8 clusters received deployments of male wolbachia-infected mosquitoes (intervention clusters) and 7 clusters received no deployments (control clusters). this study was led by the Environmental Health Institute of the National Environment Agency

    -- 393,236 residents lived in the intervention clusters, and 331,192 lived in the control clusters

    -- although these clusters were randomized to the groups getting the wolbachia-infected mosquitoes, this was a constrained randomization to prevent chance imbalance in the historical risk of dengue at baseline

-- the trial relied on a nationally representative, passively monitored database of patients (ranging in age from newborn to 104 years old) who had been tested for suspected dengue virus infection.

-- the researchers generated a localized wolbachia-infected A. aegypti mosquito line by crossing a Singapore wild-type line with a wolbachia-infected line imported from the United States over six generations, which resulted in complete maternal transmission and complete cytoplasmic incompatibility (ie, inability for the females to produce live offspring that could continue the spread of dengue)

-- the half-life of released male A. aegypti mosquitoes is 4 days, so the protocol called for twice weekly release of one to six mosquitoes per human resident per week on weekday mornings between 6:30 a.m. and 11 a.m. in the intervention clusters. Male wolbachia-infected mosquitoes were also released twice a week on the ground floors of high- and low-rise housing estates within the designated buffer areas.

-- Adult A. aegypti populations in the intervention and control clusters were monitored with the use of Gravitraps (containers that are designed to attract and capture female mosquitoes), with an average of six Gravitraps deployed per apartment block. The abundance of mosquitoes was quantified by means of the weekly Gravitrap A. aegypti index (GAI), which was defined as the total number of female adult A. aegypti mosquitoes that were caught in functional Gravitraps divided by the number of functional Gravitraps.

-- Epidemiologic efficacy was assessed through the use of two nationally representative datasets:

    -- the first dataset consisted of data from all major diagnostic laboratories through primary care clinics and hospitals used by the residents with suspected dengue illness and were tested for dengue 

    -- the second dataset was from the national dengue surveillance system, in which all cases of dengue are legally mandated to be reported to the Ministry of Health.

-- The primary end points were the diagnosis of symptomatic dengue virus infection of any severity caused by any of the 4 serotypes of the virus, as measured by the odds ratio for the distribution of wolbachia exposure among laboratory-confirmed dengue cases as compared with test negative controls, and the overall  dengue incidence

Results:

-- 189,670 residents with suspected dengue virus infection (dengue-testing database) were analyzed at the Environmental Health Institute, from hospital laboratories and major commercial diagnostic laboratories, through consultations at primary care clinics or hospitals

-- more than 30% of dengue positive samples were serotyped at the trial sites during the trial period: the predominant serotype was dengue virus type 3

-- the baseline average abundance of the mosquitoes (number of adult female mosquitoes trapped divided by number of traps):

    -- intervention clusters (with wolbachia-infected mosquitoes): 0.18

    -- control clusters (no  wolbachia-infected mosquitoes released): 0.19

        -- ie, no difference

-- from 3 months after the initiation of the intervention until the end of the 24-month trial period, the average abundance was:

    -- intervention clusters: 0.041

    -- control clusters: 0.277

        -- ie, huge difference, as per figure A below

figure A shows the abundance of Aedes aegypti mosquitoes in randomized clusters portraying a really dramatic decrease in mosquitoes in the intervention sites vs the control sites over time, as measured by the weekly Gravitrap findings (using the GAI index, as noted above)i; figure B just shows where the different interventions were frequently close to each other in each of the intervention clusters, thereby decreasing the likelihood of contamination of the intervention clusters by the control clusters

-- testing for suspected dengue virus infection was done in 8245 residents in the intervention clusters and in 10,344 residents in the control clusters

-- in the intention-to-treat analysis at 6 months or more, the percentage of residents who were who were dengue-positive:

    -- intervention clusters: 354 of 5722 tests [6%] 

    -- control clusters: 1519 of 7080 tests [21%]

        -- the percentage of residents who tested positive for dengue viral infection was lower in the intervention clusters than in the control clusters across all periods of exposure to the wolbachia-infected mosquitoes

-- the percentages of positive cases ranged from 6 to 10% in the intervention clusters, as compared with 21 to 23% in the control clusters.

-- the estimated overall protective efficacy with 3 to 12 months or more of the intervention ranged from 71 to 72% (represented by odds ratios of 0.28 to 0.29)

    -- the protective efficacy remained fairly constant across all exposure durations, with protective effects of:

        -- 0+ months: 65% (47-77%), 10% (787/8245) in intervention groups and 23% (2402/10344) in controls

        -- 3+ months: 71% (58-81%), 7% (457/6743) in intervention groups and 21% (1717/8125) in controls

        -- 6+ months: 72% (58-82%), 6% (354/5722) in intervention groups and 21% (1519/7080) in controls

        -- 9+ months: 71% (55-82%), 7% (317/4668) in intervention groups and 21% (1403/6118) in controls

        -- 12+ months: 71% (51-84%), 7% (256/3724) in intervention groups and 21% (1037/4986) in controls

-- protective efficacy was consistent across all age and sex subgroups, as well as across each calendar year

-- similarly, the protective efficacy in general remained stable in all subgroups as the exposure time increased, and it remained consistent after test results determined with IgM assays were removed from the analysis (IgM assays are 71% to 100% valid from day 4-7 of infection but are less accurate for secondary infections and have false-positives with other flaviviruses such as Zika)

Commentary:

-- dengue is a global virus causing potentially severe morbidity and mortality. The spread of dengue has increased considerably because of global warming, such that in 2025 there were >3.6 million infections and almost 2000 deaths across 100 countries (and these numbers seem to increase to new records each year)

-- in the US, there were cases reported in 49 continental US states, though most were from travelers infected elsewhere

-- local spread (indigenous cases) was largely reported in Florida, Hawaii, Texas, Arizona and California

    -- for indigenous cases: overall there were 1046 cases in 2022, 1462 in 2023, 6605 cases in 2024, and 4034 in 2025 per https://www.cdc.gov/dengue/data-research/facts-stats/historic-data.html

    -- the incidence of dengue in a climate warming study in the Americas and Asia in "a dataset covering 21 countries in Asia and the Americas found a nonlinear relationship between temperature and dengue incidence with the largest impact of warming at lower temperatures (below about 20°C), peak incidence at 27.8°C, and subsequent declines at higher temperatures. Using this inferred temperature response, we estimate that historical climate change has increased dengue incidence by 18% (11 – 27%) on average across our study countries, and that future warming could further increase it by 49% (16 – 136%) to 76% (27 – 239%) by mid-century for low or high emissions scenarios, respectively, with some cooler regions projected to see dengue doubling due to warming and other currently hot regions seeing no impact or even small declines" in this preprint study (https://pmc.ncbi.nlm.nih.gov/articles/PMC10802639/) and with similar projections in the US itself.

    -- another study assessed the global effects of climate change: "a significant increase in global dengue cases from 1990 to 2019 and a positive correlation between temperature and dengue incidence. The association between annual mean minimum temperatures and dengue incidence strengthened at temperatures exceeding 21°C. Central and eastern sub-Saharan Africa, as well as Oceania, were identified as the regions most sensitive to dengue; males and individuals aged 15–19 or 70–84 years were the most susceptible to dengue under rising temperatures. Our projections suggest that global dengue incidence will substantially increase by 2050 and 2100. By 2100, regions including Africa, the Arabian Peninsula, the southern United States, southern China, and island countries in the Pacific and Indian Oceans are projected to become year-round dengue-endemic under a high-emission climate scenario": https://agupubs.onlinelibrary.wiley.com/doi/full/10.1029/2024GH001059

-- there have been a few attempts at dengue vaccines since 2010. the CYD-TDV vaccine is anticipated to cease production in the next few years (a study found that it increased the risk of severe dengue in those getting breakthrough infections), some vaccines have lacked effect on some dengue subtypes (especially serotype 2, the most severe dengue subtype). the TAK-003 vaccine is promising but "the Strategic Advisory Group of Experts on Immunization states that 'although potential risk of severe dengue in seronegative vaccinated individuals following DENV-3 (dengue serotype 3) or DENV-4 (serotype 4) infection cannot be ruled out based on the available data, TAK-003 be considered for inclusion into existing national immunization programs in high transmission areas'"

-- over the past few years there has been research on Aedes aegypti mosquitoes that had been infected with Wolbachia pipientis bacteria, though the studies have had design problems (eg involving transmission to male and female mosquitoes, and some of the females were fertile), though the concept and data overall have found this technique to be a really viable possibility

    -- the mechanism for infecting the males with the wAlbB strain of wolbachia bacteria is that it creates cytoplasmic incompatibility in the females, and the offspring of the infected of wolbachia-infected bacteria males and wild-type females are not viable. Hence, the mosquito-to-mosquito chain is interrupted over time.

-- this current study was notable in that the number of A.aegypti mosquitoes were greatly reduced by the intervention of introducing A. aegypti mosquitoes infected with the wAlbB strain of wolbachia bacteria into male mosquitoes in different urban areas, with an attendant reduction of 71-72% of dengue cases vs the control areas after at least 3 months of exposure to these variant mosquitoes, an effect across all ages and sex subgroups

-- a Brazilian study using Sterile Insect Technology (SIT), whereby millions of pathogen-free Ae. aegypti eggs were hatched and incubated in an RNA solution (PTB2-dsRNA) for one hour, the males and females were separated, and the male pupae were soaked in a thiotepa solution for 1 hour, then washed and fed 10% sugar solutions. there was quality control to detect the presence of contaminating females and verifying the sterility of the males. Comparing the incidence of dengue before the study, the researchers found up to a 98.7% suppression of live progeny of field Ae. aegypti mosquitoes over time. And when comparing the 2020 and 2022 dengue outbreaks that occurred in the region, the incidence of dengue in the area of this study was 97% lower when compared to the control cities: https://www.thelancet.com/journals/lanam/article/PIIS2667-193X(23)00072-8/fulltext

-- one advantage of this current study is that through their methodology there should also be protection against other diseases mediated through adult A. aegypti mosquitoes: zika, chikungunya, and yellow fever

-- though Dengue serotype 3 was the most common one in Singapore, the results should be generalizable to the other serotypes since the transmission process for the dengue virus is fundamentally disrupted at the mosquito level

-- though there are high costs for this intervention because of the need for sustained releases of huge numbers of altered male mosquitoes over time, as well as sex separation and irradiation of the wolbachia mosquitoes prior to their release, the high efficacy of the process far exceeds the researchers' 40% calculated minimum to make the intervention cost-effective

Limitations:

-- there could well be some contamination in the study whereby fertile females carrying dengue could possibly be in the area of the intervention, perhaps by residents exposed to dengue when visiting other areas, thereby decreasing the apparent efficacy of the wolbachia mosquitoes in their own region. also, it is still possible that some fertile females made it into the intervention region although the clusters were at least 700 meters apart; a larger more inclusive application of this study methodology might eliminate this problem since in the current study design perhaps high winds might have might have transferred fertile female mosquitoes into adjacent areas??? The above Brazilian trial supports this likelihood that a larger study would report even more benefit

    -- we do not have specific data on the accuracy of eliminating female mosquitoes from the group in the clusters, and the fertile females might have spread dengue

-- the epidemiology of dengue globally is dependent on the quality and extent of the public health systems of each country and likely do not have consistent accurate data in many of the countries with high rates of dengue infections, especially the milder cases. to the extent this is true, the above global dengue incidence numbers are undercounted

so,

-- this study adds very impressive information that introducing altered male mosquitoes pretty dramatically decreases the spread of dengue

-- it is reasonable to think that more massive application of this approach would further decrease dengue, since in this study carried out in several urban areas in Singapore there may well have been "contamination" whereby some fertile female mosquitoes made it into the study areas and transmitted dengue. 

-- the exploits of dengue vaccines have gone on for >15 years, with none clearly being safe and effective for all of the 4 dengue serotypes

-- the efficacy of insect repellants is in the 20-40% range, and these chemicals that are often pretty toxic to us and our environment. And perhaps there would have been higher inset repellant efficacy if people replenished their application of these chemicals every few hours as suggested, which would also increase their adverse effects on us and our environment....

-- and the appeal of altering the female A. aegypti mosquitoes' ability to transmit dengue and not relying on human vaccines is quite appealing, especially given the track record of the several vaccine attempts so far

    -- the technique used in this study with targeting the males seems to be great in reducing the increasing scourge of dengue with our presently unabated climate change

    -- and, perhaps one of the most important outcomes is the high likelihood that this intervention could also lead to huge decreases in other diseases mediated through adult A. aegypti mosquitoes: zika, chikungunya, and yellow fever), and perhaps a modification of this approach might be useful in other mosquito-borne infections such as malaria, Eastern Equine Encephalitis or West Nile virus or ......

geoff

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