The range of the mosquito Aedes aegypti continues to expand, putting more than two billion people at risk of arboviral infection. The sterile insect technique (SIT) has been used to successfully combat agricultural pests at large scale, but not mosquitoes, mainly because of challenges with consistent production and distribution of high-quality male mosquitoes. We describe automated processes to rear and release millions of competitive, sterile male Wolbachia-infected mosquitoes, and use of these males in a large-scale suppression trial in Fresno County, California. In 2018, we released 14.4 million males across three replicate neighborhoods encompassing 293 hectares. At peak mosquito season, the number of female mosquitoes was 95.5% lower (95% CI, 93.6-96.9) in release areas compared to non-release areas, with the most geographically isolated neighborhood reaching a 99% reduction. This work demonstrates the high efficacy of mosquito SIT in an area ninefold larger than in previous similar trials, supporting the potential of this approach in public health and nuisance-mosquito eradication programs.

Efficient production of male Wolbachia-infected Aedes aegypti mosquitoes enables large-scale suppression of wild populations.

Living in the endosymbiotic world of Wolbachia: A centennial review.

Wolbachia is an endosymbiotic Alphaproteobacteria that can suppress insect-borne diseases through decreasing host virus transmission (population replacement) or through decreasing host population density (population suppression). We contrast natural Wolbachia infections in insect populations with Wolbachia transinfections in mosquitoes to gain insights into factors potentially affecting the long-term success of Wolbachia releases. Natural Wolbachia infections can spread rapidly, whereas the slow spread of transinfections is governed by deleterious effects on host fitness and demographic factors. Cytoplasmic incompatibility (CI) generated by Wolbachia is central to both population replacement and suppression programs, but CI in nature can be variable and evolve, as can Wolbachia fitness effects and virus blocking. Wolbachia spread is also influenced by environmental factors that decrease Wolbachia titer and reduce maternal Wolbachia transmission frequency. More information is needed on the interactions between Wolbachia and host nuclear/mitochondrial genomes, the interaction between invasion success and local ecological factors, and the long-term stability of Wolbachia-mediated virus blocking.

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Evolutionary Ecology of Wolbachia Releases for Disease Control

Background 
Important arboviral diseases, such as dengue, chikungunya, and Zika virus infections, are transmitted mainly by the Aedes aegypti vector. So far, controlling this vector species with current tools and strategies has not demonstrated sustainable and significant impacts. Our main objective was to evaluate whether open field release of sterile males, produced from combining the sterile insect technique using radiation with the insect incompatible technique through Wolbachia-induced incompatibility (SIT/IIT), could suppress natural populations of Ae. aegypti in semi-rural village settings in Thailand.


Methodology/Principal findings 
Irradiated Wolbachia-infected Aedes aegypti males produced by the SIT/IIT approach were completely sterile and were able to compete with the wild fertile ones. Open field release of these sterile males was conducted in an ecologically isolated village in Chachoengsao Province, eastern Thailand. House-to-house visit and media reports resulted in community acceptance and public awareness of the technology. During intervention, approximately 100–200 sterile males were released weekly in each household. After 6 months of sterile male release, a significant reduction (p<0.05) of the mean egg hatch rate (84%) and the mean number of females per household (97.30%) was achieved in the treatment areas when compared to the control ones.


Conclusions/Significance 
Our study represents the first open field release of sterile Ae. aegypti males developed from a combined SIT/IIT approach. Entomological assessment using ovitraps, adult sticky traps, and portable vacuum aspirators confirmed the success in reducing natural populations of Ae. aegypti females in treated areas. Public awareness through media resulted in positive support for practical use of this strategy in wider areas. Further study using a systematic randomized trial is needed to determine whether this approach could have a significant impact on the diseases transmitted by Ae. aegypti vector.

/pdf/combined-sterile-insect-technique-and-incompatible-insect-37ct59e2ma.pdf

Combined sterile insect technique and incompatible insect technique: The first proof-of-concept to suppress Aedes aegypti vector populations in semi-rural settings in Thailand.

Cytoplasmic incompatibility (CI) is the most common symbiont-induced reproductive manipulation. Specifically, symbiont-induced sperm modifications cause catastrophic mitotic defects in the fertilized embryo and ensuing lethality in crosses between symbiotic males and either aposymbiotic females or females harboring a different symbiont strain. However, if the female carries the same symbiont strain, then embryos develop properly, thereby imparting a relative fitness benefit to symbiont-transmitting mothers. Thus, CI drives maternally-transmitted bacteria to high frequencies in arthropods worldwide. In the past two decades, CI experienced a boom in interest due to its (i) deployment in worldwide efforts to curb mosquito-borne diseases, (ii) causation by bacteriophage genes, cifA and cifB, that modify sexual reproduction, and (iii) important impacts on arthropod speciation. This review serves as a gateway to experimental, conceptual, and quantitative themes of CI and outlines significant gaps in understanding CI's mechanism that are ripe for investigation from diverse subdisciplines in the life sciences.

Symbiont-mediated cytoplasmic incompatibility: what have we learned in 50 years?

The radiation-based sterile insect technique (SIT) has successfully suppressed field populations of several insect pest species, but its effect on mosquito vector control has been limited. The related incompatible insect technique (IIT)-which uses sterilization caused by the maternally inherited endosymbiotic bacteria Wolbachia-is a promising alternative, but can be undermined by accidental release of females infected with the same Wolbachia strain as the released males. Here we show that combining incompatible and sterile insect techniques (IIT-SIT) enables near elimination of field populations of the world's most invasive mosquito species, Aedes albopictus. Millions of factory-reared adult males with an artificial triple-Wolbachia infection were released, with prior pupal irradiation of the released mosquitoes to prevent unintentionally released triply infected females from successfully reproducing in the field. This successful field trial demonstrates the feasibility of area-wide application of combined IIT-SIT for mosquito vector control.

Incompatible and sterile insect techniques combined eliminate mosquitoes

Stable Introduction of Plant-Virus-Inhibiting Wolbachia into Planthoppers for Rice Protection.

Combined incompatible and sterile insect technique (IIT-SIT) has been considered to be an effective and safe approach to control mosquito populations. Immobilization of male adults by chilling is a crucial process required for the packing, transportation and release of the mosquitoes during the implementation of IIT-SIT for mosquito control. In this study, effects of chilling on the Aedes albopictus males with triple Wolbachia infections (HC line), a powerful weapon to fight against the wild type Ae. albopictus population via IIT-SIT, were evaluated under both laboratory and field conditions. Irradiated HC (IHC) males were exposed to 1, 5 and 10°C for 1, 2, 3, 6 and 24 h. The survival rate of the post-chilled IHC males was then monitored. Longevity of post-chilled IHC males was compared to non-chilled males under laboratory and semi-field conditions. Mating competitiveness of IHC/HC males after exposure to 5 or 10°C for 0, 3 and 24 h was then evaluated. Effects of compaction and transportation under chilled conditions on the survival rate of IHC males were also monitored. The optimal chilling conditions for handling IHC males were temperatures between 5 and 10°C for a duration of less than 3 h with no negative impacts on survival rate, longevity and mating competitiveness when compared to non-chilled males. However, the overall quality of post-chilled IHC/HC males decreased when exposed to low temperatures for 24 h. Reduced survival was observed when IHC males were stored at 5°C under a compaction height of 8 cm. Transportation with chilling temperatures fluctuating from 8 to 12°C has no negative impact on the survival of IHC males. This study identified the optimal chilling temperature and duration for the handling and transportation of Ae. albopictus IHC male adults without any detrimental effect on their survival, longevity and mating competitiveness. Further studies are required to develop drone release systems specific for chilled mosquitoes to improve release efficiency, as well as to compare the population suppression efficiency between release of post-chilled and non-chilled males in the field.

/pdf/toward-implementation-of-combined-incompatible-and-sterile-3k7065xi20.pdf

Toward implementation of combined incompatible and sterile insect techniques for mosquito control: Optimized chilling conditions for handling Aedes albopictus male adults prior to release.

Yongkang Liang

Papers

Incompatible and sterile insect techniques combined eliminate mosquitoes

Stable Introduction of Plant-Virus-Inhibiting Wolbachia into Planthoppers for Rice Protection.

Toward implementation of combined incompatible and sterile insect techniques for mosquito control: Optimized chilling conditions for handling Aedes albopictus male adults prior to release.