Dpto. Proyectos, Teoría y Técnica del Diseño y la Arquitectura

Aedes aegypti is the primary vector of arthropod-borne viruses including dengue, chikungunya and Zika. Vector population control methods are reviving to impede disease transmission. An efficient sex separation for male-only releases is crucial for area-wide mosquito population suppression strategies. Here, we report on the construction of two genetic sexing strains using red- and white-eye colour mutations as selectable markers. Quality control analysis showed that the Red-eye genetic sexing strains (GSS) is better and more genetically stable than the White-eye GSS. The introduction of an irradiation-induced inversion (Inv35) increases genetic stability and reduces the probability of female contamination of the male release batches. Bi-weekly releases of irradiated males of both the Red-eye GSS and the Red-eye GSS/Inv35 fully suppressed target laboratory cage populations within six and nine weeks, respectively. An image analysis algorithm allowing sex determination based on eye colour identification at the pupal stage was developed. The next step is to automate the Red-eye-based genetic sexing and validate it in pilot trials prior to its integration in large-scale population suppression programmes.

Successful implementation of the sterile insect technique (SIT) against Aedes albopictus and Anopheles arabiensis relies on a continuous supply of sterilemales. Tomeet this requirement, optimization of the mass-rearing techniques is needed. This study, therefore, aims to assess a new mass-rearing cage (MRC) in terms of egg production efficiency and egg hatch rate (quality). In addition, adult survival was evaluated based on a cage adult-index for Ae. albopictus. Moreover, the cage’s suitability for use in mass An. arabiensis egg production was compared to that of the FAO/IAEA Anopheles reference cage. In Ae. albopictus rearing, the new MRC produced 1,112,110 eggs per cage following six blood meals, with minimum loss of eggs in the egging water. Furthermore, the adult index gave a good proxy of daily mortality rates in Ae. albopictus. In An. arabiensis rearing, about 130,000 eggs per egg batch were collected both from the new and the reference MRC. These findings suggest that the new MRC prototype is efficient in terms of egg production and can be used for mass-rearing in SIT programs targeting Ae. albopictus as well as An. arabiensis. The adult index was also positively validated for the detection of unusual mortality rates in Ae. albopictus mass-rearing facilities. Overall, the new MRC has shown several advantages; however, further improvements are necessary to minimize escapes during the egg collection processes.

Introduction: The widespread emergence of resistance to insecticides used to control adult Aedes mosquitoes has made traditional control strategies inadequate for the reduction of various vector populations. Therefore, complementary vector control methods, such as the Sterile Insect Technique, are needed to enhance existing efforts. The technique relies on the rearing and release of large numbers of sterile males, and the development of efficient and standardized mass-rearing procedures and tools is essential for its application against medically important mosquitoes. Methods: In the effort to reduce the cost of the rearing process, a prototype low-cost plexiglass massrearing cage has been developed and tested for egg production and egg hatch rate in comparison to the current Food and Agriculture Organization/International Atomic Energy Agency (FAO/IAEA) stainless-steel cage. Additionally, an adult-index was validated and used as a proxy to estimate the mosquito survival rates by counting the number of male and female mosquitoes that were resting within each of the 6 squares at a given point of time each day in the cage. Results: The study has shown that the prototype mass-rearing cage is cheap and is as efficient as the FAO/IAEA stainless-steel cage in terms of egg production, with even better overall egg hatch rate. The mean numbers of eggs per cage, after seven cycles of blood feeding and egg collection, were 969,789 ± 138,101 and 779,970 ± 123,042, corresponding to 81 ± 11 and 65 ± 10 eggs per female over her lifespan, in the prototype and the stainless-steelmass-rearing cages, respectively. The longevity of adult male and female mosquitoes was not affected by cage type and, the adult-index could be considered as an appropriate proxy for survival. Moreover, the mass-rearing cage prototype is easy to handle and transport and improves economic and logistic efficiency. Conclusion: The low-cost mass-rearing prototype cage can be recommended to produce Ae. aegypti in the context of rear and release techniques. The proposed adult-index can be used as a quick proxy of mosquito survival rates in mass-rearing settings.

Background: Several mosquito population suppression strategies based on the rearing and release of sterile males have provided promising results. However, the lack of an efficient male selection method has hampered the expansion of these approaches into large-scale operational programmes. Currently, most of these programmes targeting Aedes mosquitoes rely on sorting methods based on the sexual size dimorphism (SSD) at the pupal stage. The currently available sorting methods have not been developed based on biometric analysis, and there is therefore potential for improvement. We applied an automated pupal size estimator developed by Grupo Tragsa with laboratory samples of Anopheles arabiensis, Aedes albopictus, Ae. polynesiensis, and three strains of Ae. aegypti. The frequency distribution of the pupal size was analyzed. We propose a general model for the analysis of the frequency distribution of mosquito pupae in the context of SSD-sorting methods, which is based on a Gaussian mixture distribution functions, thus making possible the analysis of performance (% males recovery) and purity (% males on the sorted sample). Results: For the three Aedes species, the distribution of the pupae size can be modeled by a mixture of two Gaussian distribution functions and the proposed model fitted the experimental data. For a given population, each size threshold is linked to a specific outcome of male recovery. Two dimensionless parameters that measure the suitability for SSD-based sorting of a specific batch of pupae are provided. The optimal sorting results are predicted for the highest values of SSD and lowest values of intra-batch variance. Rearing conditions have a strong influence in the performance of the SSD-sorting methods and non-standard rearing can lead to increase pupae size heterogeneity. Conclusions: Sex sorting of pupae based on size dimorphism can be achieved with a high performance (% males recovery) and a reasonably high purity (% males on the sorted sample) for the different Aedes species and strains. The purity and performance of a sex sorting operation in the tested Aedes species are linked parameters whose relation can be modeled. The conclusions of this analysis are applicable to all the existing SSD-sorting methods. The efficiency of the SSDsorting methods can be improved by reducing the heterogeneity of pupae size within rearing containers. The heterogeneity between batches does not strongly affect the quality of the sex sorting, as long as a specific separation threshold is not pre-set before the sorting process. For new developments, we recommend using adaptive and precise threshold selection methods applied individually to each batch or to a mix of batches. Adaptive and precise thresholds will allow the sex-sorting of mixed batches in operational conditions maintaining the target purity at the cost of a reduction in performance. We also recommend a strategy whereby an acceptable level of purity is pre-selected and remains constant across the different batches of pupae while the performance varies from batch to batch to fit with the desired purity.

Vector-borne diseases are responsible for more than one million deaths per year. Alternative methods of mosquito control to insecticides such as genetic control techniques are thus urgently needed. In genetic techniques involving the release of sterile insects, it is critical to release insects of high quality. Sterile males must be able to disperse, survive and compete with wild males in order to inseminate wild females. There is currently no standardized, fast-processing method to assess mosquito male quality. Since male competitiveness is linked to their ability to fly, we developed a flight test device that aimed to measure the quality of sterile male mosquitoes via their capacity to escape a series of flight tubes within two hours and compared it to two other reference methods (survival rate and mating propensity). This comparison was achieved in three different stress treatment settings usually encountered when applying the sterile insect technique, i.e. irradiation, chilling and compaction. In all treatments, survival and insemination rates could be predicted by the results of a flight test, with over 80% of the inertia predicted. This novel tool could become a standardised quality control method to evaluate cumulative stress throughout the processes related to genetic control of mosquitoes.