Limited impact of vector control on the population genetic structure of Glossina fuscipes fuscipes from the sleeping sickness focus of Maro, Chad.

Tsetse flies (genus Glossina) transmit deadly trypanosomes to human populations and domestic animals in sub-Saharan Africa. Some foci of Human African Trypanosomiasis due to Trypanosoma brucei gambiense (g-HAT) persist in southern Chad, where a program of tsetse control was implemented against the local vector Glossina fuscipes fuscipes in 2018 in Maro. We analyzed the population genetics of G. f. fuscipes from the Maro focus before control (T0), one year (T1), and 18 months (T2) after the beginning of control efforts. Most flies captured displayed a local genetic profile (local survivors), but a few flies displayed outlier genotypes. Moreover, disturbance of isolation by distance signature (increase of genetic distance with geographic distance) and effective population size estimates, absence of any genetic signature of a bottleneck, and an increase of genetic diversity between T0 and T2 strongly suggest gene flows from various origins, and a limited impact of the vector control efforts on this tsetse population. Continuous control and surveillance of g-HAT transmission is thus recommended in Maro. Particular attention will need to be paid to the border with the Central African Republic, a country where the entomological and epidemiological status of g-HAT is unknown.

Title: Impact limité de la lutte antivectorielle sur la structure des populations de Glossina fuscipes fuscipes dans le foyer de la maladie du sommeil de Maro, Tchad. Abstract: Les mouches tsé-tsé (genre Glossina) transmettent des trypanosomes mortels aux populations humaines ainsi qu'aux animaux domestiques en Afrique sub-saharienne. Certains foyers de la trypanosomiase humaine Africaine due à Trypanosoma brucei gambiense (THA-g) persistent au sud du Tchad, où un programme de lutte antivectorielle a été mis en place contre le vecteur local de la maladie, Glossina fuscipes fuscipes, en particulier à Maro en 2018. Nous avons analysé la structure génétique des populations de G. f. fuscipes de ce foyer à T0 (avant lutte), une année après le début de la lutte (T1), et 18 mois après (T2). La plupart des mouches capturées après le début de la lutte ont montré un profil génétique local (survivants locaux), mais quelques-unes d'entre elles présentaient des génotypes d'individus atypiques. Par ailleurs, la présence de perturbations des signatures d'isolement par la distance (augmentation de la distance génétique avec la distance géographique), l'absence de signature génétique d'un goulot d'étranglement, et un accroissement de la diversité génétique entre T0 et T2 sont des arguments forts en faveur de la recolonisation de la zone par des mouches d'origines variées, tout en témoignant des effets limités de la campagne de lutte dans ce foyer. Ces résultats conduisent à recommander une lutte et une surveillance continues dans le foyer de Maro. Une attention particulière devra par ailleurs être prêtée à l'autre côté de la rive, située côté République Centre Africaine, dont le statut épidémiologique reste inconnu concernant les tsé-tsé et la THA-g.

Animal trypanosomosis eliminated in a major livestock production region in Senegal following the eradication of a tsetse population.

African animal trypanosomosis (AAT) was one of the main disease-related constraints to the development of intensive livestock production systems in the Niayes region of Senegal, a 30 km wide strip of land along the coast between Dakar and Saint-Louis. To overcome this constraint, the Government of Senegal initiated an area-wide integrated pest management programme combining chemical control tactics with the sterile insect technique to eradicate a population of the tsetse fly Glossina palpalis gambiensis Vanderplank, 1949 (Diptera, Glossinidae) in this area. The project was implemented following a phased conditional approach, and the target area was divided into three blocks treated sequentially. This study aims to assess the temporal dynamics of the prevalence of Trypanosoma spp. during the implementation of this programme. Between 2009 and 2022, 4,359 blood samples were collected from cattle and screened for trypanosomes using both the buffy coat and ELISA techniques, and PCR tests since 2020. The seroprevalence decreased from 18.9% (95%CI: 11.2-26.5) in 2009 to 0% in 2017-2022 in block 1, and from 92.9% (95%CI: 88.2-97) in 2010 to 0% in 2021 in block 2. The parasitological and serological data confirm the entomological monitoring results, i.e., that there is a high probability that the population of G. p. gambiensis has been eradicated from the Niayes and that the transmission of AAT has been interrupted in the treated area. These results indicate the effectiveness of the adopted approach and show that AAT can be sustainably removed through the creation of a zone free of G. p. gambiensis.

Title: Trypanosomose animale éliminée dans une importante région de production d'élevage au Sénégal suite à l'éradication d'une population de glossines. Abstract: La trypanosomose animale africaine (TAA) était l'une des principales contraintes pathologiques au développement de systèmes de production animale intensifs dans les Niayes du Sénégal, une bande de terre large de 30 km longeant la côte entre Dakar et Saint-Louis. Pour surmonter cette contrainte, le Gouvernement du Sénégal a lancé un programme de lutte intégrée à l'échelle de la zone combinant lutte chimique et technique de l'insecte stérile pour éradiquer une population de Glossina palpalis gambiensis Vanderplank, 1949 (Diptera, Glossinidae). Le projet a été mis en œuvre selon une approche conditionnelle progressive, et la zone cible a été divisée en trois blocs, traités de manière séquentielle. L'objectif de cette étude était d'évaluer la dynamique temporelle de la prévalence de Trypanosoma spp. au cours de la mise en œuvre du programme. Entre 2009 et 2022, 4 359 échantillons de sang ont été prélevés sur des bovins et ont fait l'objet d'un dépistage des trypanosomes à l'aide des techniques du buffy-coat et ELISA, ainsi que de test PCR depuis 2020. Dans le bloc 1, la séroprévalence est passée de 18,9 % (IC 95 % : 11,2­26,5) en 2009 à 0 % entre 2017­2022 et de 92,9 % (IC 95 % : 88,2-97) en 2010 à 0 % en 2021 pour le block 2. Les données parasitologiques et sérologiques confirment les résultats du suivi entomologique selon lesquels il est très probable que la population de Glossina palpalis gambiensis soit éradiquée des Niayes, et que la transmission de la TAA a été interrompue dans la zone traitée. Elles indiquent l'efficacité de l'approche adoptée, et montrent que la TAA peut être durablement éliminée grâce à la création d'une zone exempte de G. p. gambiensis.

Sterile Insect Technique (SIT) field trial targeting the suppression of Aedes albopictus in Greece.

The sterile insect technique (SIT) involves releasing large numbers of sterile males to outcompete wild males in mating with females, leading to a decline in pest populations. In the current study, we conducted a suppression trial in Greece against the invasive dengue vector mosquito Aedes albopictus (Skuse) through the weekly release of sterile males for 22 weeks from June to September 2019. Our approach included the long-distance transport of sterile mosquitoes, and their release at a density of 2,547 ± 159 sterile males per hectare per week as part of an area-wide integrated pest management strategy (AW-IPM). The repeated releases of sterile males resulted in a gradual reduction in egg density, reaching 78% from mid-June to early September. This reduction remained between 70% and 78% for four weeks after the end of the releases. Additionally, in the SIT intervention area, the ovitrap index, representing the percentage of traps containing eggs, remained lower throughout the trial than in the control area. This trial represents a significant advance in the field of mosquito control, as it explores the viability and efficacy of producing and transporting sterile males from a distant facility to the release area. Our results provide valuable insights for future SIT programmes targeting Ae. Albopictus, and the methodology we employed can serve as a starting point for developing more refined and effective release protocols, including the transportation of sterile males over long distances from production units to intervention areas.

Title: Essai sur le terrain de la Technique de l'Insecte Stérile (TIS) ciblant la suppression d'Aedes albopictus en Grèce. Abstract: La technique de l'insecte stérile (TIS) consiste à libérer un grand nombre de mâles stériles pour supplanter les mâles sauvages lors de l'accouplement avec les femelles, entraînant ainsi un déclin des populations de nuisibles. Dans la présente étude, nous avons mené un essai de suppression en Grèce contre le moustique vecteur invasif de la dengue, Aedes albopictus (Skuse), par le biais de la libération hebdomadaire de mâles stériles pendant 22 semaines de juin à septembre 2019. Notre approche comprenait le transport sur de longues distances de moustiques stériles, et leur lâcher à une densité de 2 547 ± 159 mâles stériles par hectare et par semaine dans le cadre d'une stratégie de lutte intégrée contre les nuisibles à l'échelle de la zone (AW-IPM). Les lâchers répétés de mâles stériles ont entraîné une réduction progressive de la densité des œufs, atteignant 78 % de la mi-juin au début septembre. Cette réduction est restée entre 70 % et 78 % pendant quatre semaines après la fin des lâchers. De plus, dans la zone d'intervention de la TIS, l'indice d'oviposition, représentant le pourcentage de pièges contenant des œufs, est resté plus faible que dans la zone témoin tout au long de l'essai. Cet essai représente une avancée significative dans le domaine de la lutte contre les moustiques, car il explore la viabilité et l'efficacité de la production et du transport de mâles stériles depuis une installation éloignée vers la zone de lâcher. Nos résultats fournissent des informations précieuses pour les futurs programmes de TIS ciblant Ae. albopictus et la méthodologie que nous avons utilisée pourra servir de point de départ pour développer des protocoles de libération plus raffinés et plus efficaces, y compris le transport de mâles stériles sur de longues distances depuis les unités de production jusqu'aux zones d'intervention.

Mating harassment may boost the effectiveness of the sterile insect technique for Aedes mosquitoes.

The sterile insect technique is based on the overflooding of a target population with released sterile males inducing sterility in the wild female population. It has proven to be effective against several insect pest species of agricultural and veterinary importance and is under development for Aedes mosquitoes. Here, we show that the release of sterile males at high sterile male to wild female ratios may also impact the target female population through mating harassment. Under laboratory conditions, male to female ratios above 50 to 1 reduce the longevity of female Aedes mosquitoes by reducing their feeding success. Under controlled conditions, blood uptake of females from an artificial host or from a mouse and biting rates on humans are also reduced. Finally, in a field trial conducted in a 1.17 ha area in China, the female biting rate is reduced by 80%, concurrent to a reduction of female mosquito density of 40% due to the swarming of males around humans attempting to mate with the female mosquitoes. This suggests that the sterile insect technique does not only suppress mosquito vector populations through the induction of sterility, but may also reduce disease transmission due to increased female mortality and lower host contact.

Two invasions at once: update on the introduction of the invasive species Aedes aegypti and Aedes albopictus in Cyprus - a call for action in Europe.

Aedes aegypti, the yellow fever mosquito and Aedes albopictus, the tiger mosquito, continue to expand their geographical distribution, reshaping the European epidemiological risks for mosquito-borne diseases. The reintroduction of Aedes aegypti near the airport and port in Larnaka and the detection of Aedes albopictus near the marina and old port of the Limassol area in Cyprus are reported herein. The measures taken to investigate these events included (i) communication to health authorities, (ii) expert on-site visits and verification of findings, (iii) enhanced active surveillance, and (iv) development of an Emergency Action Plan followed by a Contingency Plan. These emergency action plans were developed to delimitate the infested areas and to prevent the spreading of the mosquito populations into new areas. The general principles are presented along with their rationale to serve as guidelines for other geographical regions targeting suppression/eradication with a sterile insect technique component. In parallel, this manuscript serves as a call for action at the European level to impede the further spread of these species and support the activities being undertaken in Cyprus to combat the incursions of Aedes invasive species.

Title: Deux invasions à la fois : le point sur l'introduction des espèces envahissantes Aedes aegypti et Aedes albopictus à Chypre ­ un appel à l'action en Europe. Abstract: Aedes aegypti, le moustique de la fièvre jaune, et Aedes albopictus, le moustique tigre, continuent à étendre leurs aires de distribution, redessinant les risques épidémiologiques des maladies transmises par les moustiques en Europe. La réintroduction d'Aedes aegypti près de l'aéroport et du port de Larnaka, et la détection d'Aedes albopictus près de la marina et du vieux port de la zone de Limassol à Chypre sont rapportées dans cette étude. Les mesures prises pour investiguer ces évènements ont inclus : (i) la communication avec les autorités sanitaires, (ii) des visites d'experts sur site et la vérification des résultats, (iii) une surveillance active augmentée et (iv) le développement d'un Plan d'Actions d'Urgence suivi d'un Plan de Contingence. Des plans d'action d'urgence ont été développés pour délimiter les zones infestées et prévenir l'expansion des populations de moustiques à d'autres zones. Les principes généraux sont présentés, en même temps que leur justification, pour guider d'autres zones géographiques visant la suppression / éradication avec une composante de technique de l'insecte stérile. En parallèle, cet article sert d'appel, au niveau européen, à prévenir la diffusion toujours plus importante de ces espèces et pour appuyer les activités à mener à Chypre pour combattre les incursions des espèces invasives d'Aedes.

Effects of radiation on the fitness, sterility and arbovirus susceptibility of a Wolbachia-free Aedes albopictus strain for use in the sterile insect technique.

BACKGROUND: The sterile insect technique (SIT) is a green and species-specific insect pest control technique that suppresses target populations by releasing factory-reared, radiosterilized males into the wild. Once released, it is important to be able to distinguish the released males from the wild males for monitoring purposes. Several methods to mark the sterile males exist. However, most have limitations due to monetary, process efficiency, or insect quality. Aedes albopictus is naturally infected with Wolbachia at a high prevalence, therefore the elimination of Wolbachia can serve as a biomarker to distinguish factory-reared male mosquitoes from wild conspecifics. RESULTS: In this study, a Wolbachia-free Ae. albopictus GT strain was developed and its fitness evaluated, which was found to be comparable to the wild GUA strain. In addition, GT male mosquitoes were irradiated at the adult stage and a dose of 20 Gy or more induced over 99% sterility. Moreover, a dose of 30 Gy (almost completely sterilizing male and female mosquitoes) had limited effects on the mating competitiveness of GT males and the vector competence of GT females, respectively. However, radiation reduced mosquito longevity, regardless of sex. CONCLUSION: Our results indicate that the Ae. albopictus GT strain can be distinguished from wild mosquitoes based on Wolbachia status and shows similar fitness, radio-sensitivity and arbovirus susceptibility to the GUA strain, indicating that it is feasible to use the GT strain to suppress Ae. albopictus populations for SIT programmes. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Combining two genetic sexing strains allows sorting of non-transgenic males for Aedes genetic control.

Chemical control of disease vectoring mosquitoes Aedes albopictus and Aedes aegypti is costly, unsustainable, and increasingly ineffective due to the spread of insecticide resistance. The Sterile Insect Technique is a valuable alternative but is limited by slow, error-prone, and wasteful sex-separation methods. Here, we present four Genetic Sexing Strains (two for each Aedes species) based on fluorescence markers linked to the m and M sex loci, allowing for the isolation of transgenic males. Furthermore, we demonstrate how combining these sexing strains enables the production of non-transgenic males. In a mass-rearing facility, 100,000 first instar male larvae could be sorted in under 1.5 h with an estimated 0.01-0.1% female contamination on a single machine. Cost-efficiency analyses revealed that using these strains could result in important savings while setting up and running a mass-rearing facility. Altogether, these Genetic Sexing Strains should enable a major upscaling in control programmes against these important vectors.

Gamma-radiation of Glossina palpalis gambiensis revisited: effect on fertility and mating competitiveness.

African animal trypanosomoses are vector-borne diseases that cause enormous livestock losses in sub-Saharan Africa, with drastic socio-economic impacts. Vector control in the context of an area-wide integrated pest management program with a sterile insect technique component requires the production of high-quality sterile male tsetse flies. In our study, we evaluated the effect of irradiation on the fecundity of Glossina palpalis gambiensis to identify the optimal dose that will induce maximum sterility while maintaining biological performance as much as possible. In addition, male mating performance was evaluated in semi-field cages. The irradiation doses used were 90, 100, 110, 120, 130, 140, and 150 Gy, and untreated males were used as the control. The results showed that pupal production and emergence rates were higher in batches of females that had mated with fertile males than in those that had mated with irradiated males with any experimental dose. A dose of 120 Gy administered to male flies induced 97-99% sterility after mating with virgin females. For the semi-field cage experiments, males irradiated with 120 Gy showed good sexual competitiveness as compared to fertile males and those irradiated with 140 Gy, considering the level of filling of spermatheca and the number of pairs formed. The optimal radiation dose of 120 Gy found in this study is slightly different from the traditional dose of 110 Gy that has been used in several eradication programmes in the past. The potential reasons for this difference are discussed, and an argument is made for the inclusion of reliable dosimetry systems in these types of studies.

Title: Le rayonnement gamma pour Glossina palpalis gambiensis revisité : effet sur la fertilité et la compétitivité sexuelle. Abstract: Les trypanosomoses animales africaines sont des maladies à transmission vectorielle qui causent d'énormes pertes de bétail en Afrique subsaharienne, avec des impacts socio-économiques importants. La lutte antivectorielle dans le cadre d'un programme de lutte intégrée contre les ravageurs à l'échelle d'une zone avec une composante de technique d'insectes stériles nécessite la production de glossines mâles stériles de haute qualité. Dans notre étude, nous avons évalué l'effet de l'irradiation sur la fécondité de Glossina palpalis gambiensis afin d'identifier la dose optimale qui induira une stérilité maximale tout en maintenant au maximum les performances biologiques. De plus, les performances d'accouplement des mâles ont été évaluées en cages de semi-terrain. Les doses d'irradiation utilisées étaient de 90, 100, 110, 120, 130, 140 et 150 Gy, et des mâles non traités ont été utilisés comme contrôle. Les résultats ont montré que les taux de production et d'émergence de pupes étaient plus élevés dans les lots de femelles qui s'étaient accouplées avec des mâles fertiles que dans les lots de celles accouplées avec des mâles irradiés, avec n'importe quelle dose expérimentale. Une dose de 120 Gy administrée à des mouches mâles a induit une stérilité de 97 à 99 % après accouplement avec des femelles vierges. Pour les expériences en cages de semi-terrain, les mâles irradiés à 120 Gy ont montré une bonne compétitivité sexuelle par rapport aux mâles fertiles et à ceux irradiés à 140 Gy, en considérant le niveau de remplissage de leur spermathèque et le nombre de couples formés. La dose de rayonnement optimale de 120 Gy trouvée dans cette étude est légèrement différente de la dose traditionnelle de 110 Gy qui a été utilisée dans plusieurs programmes d'éradication dans le passé. Les raisons potentielles de cette différence sont discutées et un argument est avancé pour l'inclusion de systèmes de dosimétrie fiables dans ce type d'études.

Vector competence of sterile male Glossina fuscipes fuscipes for Trypanosoma brucei brucei: implications for the implementation of the sterile insect technique in a sleeping sickness focus in Chad.

BACKGROUND: Human African trypanosomiasis (HAT) is a neglected tropical disease caused by Trypanosoma brucei gambiense transmitted by tsetse flies in sub-Saharan West Africa. In southern Chad the most active and persistent focus is the Mandoul focus, with 98% of the reported human cases, and where African animal trypanosomosis (AAT) is also present. Recently, a control project to eliminate tsetse flies (Glossina fuscipes fuscipes) in this focus using the sterile insect technique (SIT) was initiated. However, the release of large numbers of sterile males of G. f. fuscipes might result in a potential temporary increase in transmission of trypanosomes since male tsetse flies are also able to transmit the parasite. The objective of this work was therefore to experimentally assess the vector competence of sterile males treated with isometamidium for Trypanosoma brucei brucei. METHODS: An experimental infection was set up in the laboratory, mimicking field conditions: the same tsetse species that is present in Mandoul was used. A T. b. brucei strain close to T. b. gambiense was used, and the ability of the sterile male tsetse flies fed on blood with and without a trypanocide to acquire and transmit trypanosomes was measured. RESULTS: Only 2% of the experimentally infected flies developed an immature infection (midgut) while none of the flies developed a metacyclic infection of T. b. brucei in the salivary glands. We did not observe any effect of the trypanocide used (isometamidium chloride at 100 mg/l) on the development of infection in the flies. CONCLUSIONS: Our results indicate that sterile males of the tested strain of G. f. fuscipes were unable to cyclically transmit T. b. brucei and might even be refractory to the infection. The data of the research indicate that the risk of cyclical transmission of T. brucei by sterile male G. f. fuscipes of the strain colonized at IAEA for almost 40 years appears to be small.

From the Lab to the Field: Long-Distance Transport of Sterile Aedes Mosquitoes.

Pilot programs of the sterile insect technique (SIT) against Aedes aegypti may rely on importing significant and consistent numbers of high-quality sterile males from a distant mass rearing factory. As such, long-distance mass transport of sterile males may contribute to meet this requirement if their survival and quality are not compromised. This study therefore aimed to develop and assess a novel method for long-distance shipments of sterile male mosquitoes from the laboratory to the field. Different types of mosquito compaction boxes in addition to a simulation of the transport of marked and unmarked sterile males were assessed in terms of survival rates/recovery rates, flight ability and morphological damage to the mosquitoes. The novel mass transport protocol allowed long-distance shipments of sterile male mosquitoes for up to four days with a nonsignificant impact on survival (>90% for 48 h of transport and between 50 and 70% for 96 h depending on the type of mosquito compaction box), flight ability, and damage. In addition, a one-day recovery period for transported mosquitoes post-transport increased the escaping ability of sterile males by more than 20%. This novel system for the long-distance mass transport of mosquitoes may therefore be used to ship sterile males worldwide for journeys of two to four days. This study demonstrated that the protocol can be used for the standard mass transport of marked or unmarked chilled Aedes mosquitoes required for the SIT or other related genetic control programs.

Radiation dose-fractionation in adult Aedes aegypti mosquitoes.

Balancing process efficiency and adult sterile male biological quality is one of the challenges in the success of the sterile insect technique (SIT) against insect pest populations. For the SIT against mosquitoes, many stress factors need to be taken into consideration when producing sterile males that require high biological quality to remain competitive once released in the field. Pressures of mass rearing, sex sorting, irradiation treatments, packing, transport and release including handling procedures for each step, add to the overall stress budget of the sterile male post-release. Optimizing the irradiation step to achieve maximum sterility while keeping off-target somatic damage to a minimum can significantly improve male mating competitiveness. It is therefore worth examining various protocols that have been found to be effective in other insect species, such as dose fractionation. A fully sterilizing dose of 70 Gy was administered to Aedes aegypti males as one acute dose or fractionated into either two equal doses of 35 Gy, or one low dose of 10 Gy followed by a second dose of 60 Gy. The two doses were separated by either 1- or 2-day intervals. Longevity, flight ability, and mating competitiveness tests were performed to identify beneficial effects of the various treatments. Positive effects of fractionating dose were seen in terms of male longevity and mating competitiveness. Although applying split doses generally improved male quality parameters, the benefits may not outweigh the added labor in SIT programmes for the management of mosquito vectors.

Title: Fractionnement de la dose d'irradiation chez les moustiques Aedes aegypti adultes. Abstract: Équilibrer l'efficacité du processus et la qualité biologique des mâles adultes stériles est l'un des défis du succès de la technique des insectes stériles (TIS) contre les populations d'insectes nuisibles. Pour la TIS contre les moustiques, de nombreux facteurs de stress sont à prendre en compte lors de la production de mâles stériles qui nécessitent une haute qualité biologique pour rester compétitifs une fois relâchés au champ. Les pressions de l'élevage en masse, du triage par sexe, des traitements d'irradiation, de l'emballage, du transport et de la libération, y compris les procédures de manipulation pour chaque étape, s'ajoutent au budget de stress global du mâle stérile après la libération. L'optimisation de l'étape d'irradiation pour atteindre une stérilité maximale tout en minimisant les dommages somatiques hors cible peut améliorer considérablement la compétitivité de l'accouplement des mâles et il est donc important d'examiner divers protocoles qui se sont révélés efficaces chez d'autres espèces d'insectes, comme le fractionnement de dose. Une dose entièrement stérilisante de 70 Gy a été administrée aux mâles Aedes aegypti en une dose unique ou fractionnée en deux doses égales de 35 Gy, ou une faible dose de 10 Gy suivie d'une seconde dose de 60 Gy. Les deux doses étaient séparées par des intervalles de 1 ou 2 jours. Des tests de longévité, d'aptitude au vol et de compétitivité à l'accouplement ont été réalisés pour identifier les effets bénéfiques des différents traitements. Des effets positifs de la dose de fractionnement ont été observés en termes de longévité des mâles et de compétitivité à l'accouplement. Bien que l'application de doses fractionnées améliore généralement les paramètres de qualité des mâles, les avantages peuvent ne pas compenser le travail supplémentaire dans les programmes TIS pour la gestion des moustiques vecteurs.

When less is more: accounting for overcompensation in mosquito SIT projects.

Compensation and overcompensation under field conditions are confirmed in Aedes mosquitoes recently by Evans et al.: equal or increased densities of emerging adults may thus result from reduced larval densities. Here the consequences when applying the sterile insect technique and provide recommendations to avoid counterproductive effects are discussed.

Suitability of Raycell MK2 Blood X-ray Irradiator for the Use in the Sterile Insect Technique: Dose Response in Fruit Flies, Tsetse Flies and Mosquitoes.

The sterile insect technique (SIT) is based on the inundatory field release of a target pest following their reproductive sterilization via exposure to radiation. Until recently, gamma irradiation from isotopic sources has been the most widely used in SIT programs. As isotopic sources are becoming increasingly expensive, especially for small programs, and regulations surrounding their procurement and shipment increasingly strict, irradiation capacity is one of the limiting factors in smaller or newly developing SIT projects. For this reason, the possibility of using X-ray irradiators has been evaluated in the recent decade. The availability of "off-the-shelf" blood X-ray irradiators that meet the technical requirements for insect irradiation can provide irradiation capacity for those SIT projects in which the acquisition of gamma ray irradiators is not feasible. Following the recent technical characterization of a Raycell MK2 X-ray blood irradiator, it was found in this study, that MK2 instruments were suitable for the sterilization of fruit flies, tsetse flies and mosquitoes, inducing comparable, even slightly higher, sterility levels compared to those achieved by gamma ray irradiation. This, together with its estimated processing efficiency, shows that MK2 irradiators are suitable for small- to mid-sized SIT programs.

Optimization of oviposition trap settings to monitor populations of Aedes mosquitoes, vectors of arboviruses in La Reunion.

Several dengue epidemics recently occurred in La Reunion, an island harboring two dengue viruses (DVs) vectors: Aedes albopictus, and Ae. aegypti, the former being the main local DV vector. Aedes aegypti shows a peculiar ecology, compared to other tropical populations of the same species. This study aimed to provide researchers and public-health users with locally validated oviposition traps (ovitraps) to monitor Aedes populations. A field experiment was performed in Saint-Joseph to assess the effect of different settings on the detection probability and apparent density of Aedes mosquitoes. Black plastic ovitraps were identified as the best choice. Vacoa trees (Pandanus utilis) were the only observed breeding sites for Ae. aegypti, shared with Ae. albopictus. They were the experimental units in a Latin square design with three factors: trap position in the trees (ground vs canopy), oviposition surface in the trap (blotting paper vs. vacoa leaf), and addition of organic matter to the trap water. The latter factor was found unimportant. On the ground, Ae. aegypti eggs were only found with vacoa leaves as the oviposition surface. Their detection and apparent density increased when ovitraps were located in the tree canopy. The main factor for Ae. albopictus was the oviposition surface, with a preference for blotting paper. In all trap settings, their detection was close to 100%. Larval survival was lower for a high egg density, combined with blotting paper as the oviposition surface. When monitoring mixed Aedes populations in La Reunion, we recommend using black plastic ovitraps, placed at 1.50-to-2.00-m high in vacoa trees, with vacoa leaves as the oviposition surface.

Optimization of adult mosquito trap settings to monitor populations of Aedes and Culex mosquitoes, vectors of arboviruses in La Reunion.

Competent arbovirus vectors are found in the culicid mosquito fauna of south-west Indian Ocean (SWIO) islands. In La Reunion, Aedes albopictus and Aedes aegypti mosquitoes are known vectors of dengue and chikungunya viruses. Culex quinquefasciatus is a potential vector of Rift Valley fever and West Nile viruses. To prepare a vector-control field trial against Ae. aegypti, this study aimed at identifying the best trapping strategy to catch adult Ae. aegypti, using BG-Sentinel traps (Biogents, Germany). It was implemented in two sites in southern La Reunion. Catches of Ae. albopictus and Cx. quinquefasciatus mosquitoes were also recorded. A Latin square design was used to estimate the detection probability and the apparent daily density-according to the BG-Sentinel trapping strategy: none, carbon dioxide (CO2), a commercial attractant-BG-Lure (Biogents, Germany), or both. The use of CO2 alone was associated with a higher detection probability for Ae. aegypti and Cx. quinquefasciatus mosquitoes, as well as a large increase in their apparent density. Traps with BG-Lure-alone or in combination with CO2, did not improve the detection probability of Ae. aegypti and Cx. quinquefasciatus mosquitoes. The same result was found for male Ae. albopictus. For females, baiting BG-Sentinel traps with CO2 or BG-Lure had no significant effect. The same apparent densities were found for Ae. aegypti and Ae. albopictus mosquitoes in both study sites-where Ae. aegypti mosquitoes were found at very low densities during previous surveys.

A Mark-Release-Recapture Study to Estimate Field Performance of Imported Radio-Sterilized Male Aedes albopictus in Albania.

The pathogen transmitting Aedes albopictus mosquito is spreading rapidly in Europe, putting millions of humans and animals at risk. This species is well-established in Albania since its first detection in 1979. The sterile insect technique (SIT) is increasingly gaining momentum worldwide as a component of area-wide-integrated pest management. However, estimating how the sterile males will perform in the field and the size of target populations is crucial for better decision-making, designing and elaborating appropriate SIT pilot trials, and subsequent large-scale release strategies. A mark-release-recapture (MRR) experiment was carried out in Albania within a highly urbanized area in the city of Tirana. The radio-sterilized adults of Ae. albopictus Albania strain males were transported by plane from Centro Agricoltura Ambiente (CAA) mass-production facility (Bologna, Italy), where they were reared. In Albania, sterile males were sugar-fed, marked with fluorescent powder, and released. The aim of this study was to estimate, under field conditions, their dispersal capacity, probability of daily survival and competitiveness, and the size of the target population. In addition, two adult mosquito collection methods were also evaluated: BG-Sentinel traps baited with BG-Lure and CO2, (BGS) versus human landing catch (HLC). The overall recapture rates did not differ significantly between the two methods (2.36% and 1.57% of the total male released were recaptured respectively by BGS and HLC), suggesting a similar trapping efficiency under these conditions. Sterile males traveled a mean distance of 93.85 ± 42.58 m and dispersed up to 258 m. Moreover, they were observed living in the field up to 15 days after release with an average life expectancy of 4.26 ± 0.80 days. Whether mosquitoes were marked with green, blue, yellow, or pink, released at 3.00 p.m. or 6.00 p.m., there was no significant difference in the recapture, dispersal, and survival rates in the field. The Fried competitiveness index was estimated at 0.28. This mark-release-recapture study provided important data for better decision-making and planning before moving to pilot SIT trials in Albania. Moreover, it also showed that both BG-traps and HLC were successful in monitoring adult mosquitoes and provided similar estimations of the main entomological parameters needed.

Response of male adult Aedes mosquitoes to gamma radiation in different nitrogen environments.

The developmental stage of the mosquito is one of the main factors that affect its response to ionizing radiation. Irradiation of adults has been reported to have beneficial effects. However, the main challenge is to immobilize and compact a large number of adult male mosquitoes for homogenous irradiation with minimal deleterious effects on their quality. The present study investigates the use of nitrogen in the irradiation of adult Aedes albopictus and Ae. aegypti. Irradiation in nitrogen (N2) and in air after being treated with nitrogen (PreN2) were compared with irradiation in air at gamma radiation doses of 0, 55, 70, 90, 110, and 125 Gy. In both species, approximately 0% egg hatch rate was observed following doses above 55 Gy in air versus 70 Gy in PreN2 and 90 Gy in N2. Males irradiated at a high mosquito density showed similar egg hatch rates as those irradiated at a low density. Nitrogen treatments showed beneficial effects on the longevity of irradiated males for a given dose, revealing the radioprotective effect of anoxia. However, irradiation in N2 or PreN2 slightly reduced the male flight ability. Nitrogen treatment was found to be a reliable method for adult mosquito immobilization. Overall, our results demonstrated that nitrogen may be useful in adult Aedes mass irradiation. The best option seems to be PreN2 since it reduces the immobilization duration and requires a lower dose than that required in the N2 environment to achieve full sterility but with similar effects on male quality. However, further studies are necessary to develop standardized procedures including containers, time and pressure for flushing with nitrogen, immobilization duration considering mosquito species, age, and density.

Life-history traits of a fluorescent Anopheles arabiensis genetic sexing strain introgressed into South African genomic background.

BACKGROUND: South Africa has set a mandate to eliminate local malaria transmission by 2023. In pursuit of this objective a Sterile Insect Technique programme targeting the main vector Anopheles arabiensis is currently under development. Significant progress has been made towards operationalizing the technology. However, one of the main limitations being faced is the absence of an efficient genetic sexing system. This study is an assessment of an An. arabiensis (AY-2) strain carrying the full Y chromosome from Anopheles gambiae, including a transgenic red fluorescent marker, being introgressed into a South African genetic background as a potential tool for a reliable sexing system. METHODS: Adult, virgin males from the An. arabiensis AY-2 strain were outcrossed to virgin females from the South African, Kwazulu-Natal An. arabiensis (KWAG strain) over three generations. Anopheles arabiensis AY-2 fluorescent males were sorted as first instar larvae (L1) using the Complex Object Parametric Analyzer and Sorter (COPAS) and later screened as pupae to verify the sex. Life history traits of the novel hybrid KWAG-AY2 strain were compared to the original fluorescent AY-2 strain, the South African wild-type KWAG strain and a standard laboratory An. arabiensis (Dongola reference strain). RESULTS: The genetic stability of the sex-linked fluorescent marker and the integrity and high level of sexing efficiency of the system were confirmed. No recombination events in respect to the fluorescent marker were detected over three rounds of introgression crosses. KWAG-AY2 had higher hatch rates and survival of L1 to pupae and L1 to adult than the founding strains. AY-2 showed faster development time of immature stages and larger adult body size, but lower larval survival rates. Adult KWAG males had significantly higher survival rates. There was no significant difference between the strains in fecundity and proportion of males. KWAG-AY2 males performed better than reference strains in flight ability tests. CONCLUSION: The life history traits of KWAG-AY2, its rearing efficiency under laboratory conditions, the preservation of the sex-linked fluorescence and perfect sexing efficiency after three rounds of introgression crosses, indicate that it has potential for mass rearing. The potential risks and benefits associated to the use of this strain within the Sterile Insect Technique programme in South Africa are discussed.

Standardization of the FAO/IAEA Flight Test for Quality Control of Sterile Mosquitoes.

Successful implementation of the sterile insect technique (SIT) against Aedes aegypti and Aedes albopictus relies on maintaining a consistent release of high-quality sterile males. Affordable, rapid, practical quality control tools based on the male's flight ability (ability to escape from a flight device) may contribute to meeting this requirement. Therefore, this study aims to standardize the use of the original FAO/IAEA rapid quality control flight test device (FTD) (version 1.0), while improving handling conditions and reducing the device's overall cost by assessing factors that could impact the subsequent flight ability of Aedes mosquitoes. The new FTD (version 1.1) is easier to use. The most important factors affecting escape rates were found to be tube color (or "shade"), the combined use of a lure and fan, mosquito species, and mosquito age and density (25; 50; 75; 100 males). Other factors measured but found to be less important were the duration of the test (30, 60, 90, 120 min), fan speed (normal 3000 rpm vs. high 6000 rpm), and mosquito strain origin. In addition, a cheaper version of the FTD (version 2.0) that holds eight individual tubes instead of 40 was designed and successfully validated against the new FTD (version 1.1). It was sensitive enough to distinguish between the effects of cold stress and high irradiation dose. Therefore, the eight-tube FTD may be used to assess Aedes' flight ability. This study demonstrated that the new designs (versions 1.1 and 2.0) of the FTD could be used for standard routine quality assessments of Aedes mosquitoes required for an SIT and other male release-based programs.

Field Performance Assessment of Irradiated Aedes albopictus Males Through Mark-Release-Recapture Trials With Multiple Release Points.

Mark-release-recapture (MRR) trials have been conducted in Northern Italy to evaluate the capacity of radio-substerilized Aedes albopictus males to survive, disperse, and engage in mating in the field. Two MRR sessions with the human landing collection method (HLC) were conducted with the simultaneous release of irradiated males marked with four different pigment colors. The survival and dispersal rates seem to be influenced more by environmental factors such as barriers, shading, and vegetation rather than weather parameters. In this study, we confirmed a positive linear relationship between the sterile adult male's daily survival rate and the relative humidity previously reported in similar experimental conditions and a different dispersal capacity of the released A. albopictus males in low- (NDVI index <0.4) and high (NDVI index >0.4)-vegetated areas. Consistent with previous studies, A. albopictus males have their maximal dispersion in the first days after release, while in the following days the males become more stationary. The similar field performances obtained with marked and unmarked radio-sterilized and untreated A. albopictus males on similar environments confirm the negligible effects of irradiation and marking procedures on the quality of the males released. The similar sterile to wild (S/W) male ratio measured in high- and low-vegetation areas in the release sites indicates a similar distribution pattern for the wild and the released sterile males. According to the MRR data collected, the Lincoln index estimated different A. albopictus mean population densities in the study areas equal to 7,000 and 3,000 male/ha, respectively.