Targeting mosquitoes to target malaria: Anopheline mosquitoes transmit the malaria parasite, but the insects themselves are prone to their own infections. Lovett et al. engineered a specific fungal pathogen of anophelines to carry insect-selective toxins. The effectiveness of this fungus for controlling mosquitoes was trialed in near-field conditions in Burkina Faso in a setup called MosquitoSphere. Approximately 75% of wild insecticide-resistant mosquitoes released into the environment became infected with the transgenic fungus, causing population collapse within 45 days. Science , this issue p. 894

Malaria control demands the development of a wide range of complementary strategies. We describe the properties of a naturally occurring, non–genetically modified symbiotic bacterium, Delftia tsuruhatensis TC1, which was isolated from mosquitoes incapable of sustaining the development of Plasmodium falciparum parasites. D. tsuruhatensis TC1 inhibits early stages of Plasmodium development and subsequent transmission by the Anopheles mosquito through secretion of a small-molecule inhibitor. We have identified this inhibitor to be the hydrophobic molecule harmane. We also found that, on mosquito contact, harmane penetrates the cuticle, inhibiting Plasmodium development. D. tsuruhatensis TC1 stably populates the mosquito gut, does not impose a fitness cost on the mosquito, and inhibits Plasmodium development for the mosquito’s life. Contained field studies in Burkina Faso and modeling showed that D. tsuruhatensis TC1 has the potential to complement mosquito-targeted malaria transmission control.

Genetically enhanced Metarhizium pingshaense are being developed for malaria vector control in Burkina Faso. However, not much is known about the local prevalence and pathogenicity of this fungus, so we prospected mosquitoes and plant roots (a common habitat for Metarhizium spp.) for entomopathogenic fungi. Our investigations showed that Metarhizium spp. represented between 29–74% of fungi isolated from plant root rhizospheres in diverse collection sites. At low spore dosages (1 × 106 conidia/ml), two mosquito-derived M. pingshaense isolates (Met_S26 and Met_S10) showed greater virulence against Anopheles coluzzii (LT80 of ~7 days) than isolates tested in previous studies (LT80 of ~10 days). In addition, the local isolates did not cause disease in non-target insects (honeybees and cockroaches). Our work provides promising findings for isolating local Metarhizium strains for application in mosquito biological control and for future transgenic biocontrol strategies in Burkina Faso.

Transgenic Metarhizium pingshaense expressing the spider neurotoxin Hybrid (Met-Hybrid) kill mosquitoes faster and at lower spore doses than wild-type strains. In this study, we demonstrate that this approach dovetails with the cornerstone of current malaria control: pyrethroid-insecticides, which are the cornerstone of current malaria control. We used World Health Organization (WHO) tubes, to compare the impact on insecticide resistance of Met-Hybrid with red fluorescent M. pingshaense (Met-RFP), used as a proxy for the wild-type fungus. Insecticides killed less than 20% of Anopheles coluzzii and Anopheles gambiae s.s. mosquitoes collected in a malaria endemic region of Burkina Faso where pyrethroid use is common. Seven days post-infection, mortality for insecticide-sensitive and resistant mosquitoes averaged 94% with Met-Hybrid and 64% with Met-RFP, with LT₈₀ values of 5.32±0.199 days and 7.76±0.183 days, respectively. Eighty nine percent of insecticide-resistant mosquitoes exposed to permethrin five days post-infection with Met-Hybrid died within 24 hours: only 22% died from Met-Hybrid alone over this 24-hour period. Compared to Met-RFP, Met-Hybrid also significantly reduced flight capacity of mosquitoes 3 to 5 days post-infection. Based on WHOPES phase I laboratory susceptibility bioassays, transgenic Met-Hybrid provides effective biological control for adult African malaria vectors that may be used to synergistically manage insecticide resistance with current methods.

Background: Malaria, a disease transmitted by Anopheles mosquitoes, is a major public health problem causing millions of deaths worldwide, mostly among children under the age of 5 years. Biotechnological interventions targeting parasite-vector interactions have shown that the microsporidian symbiont Microsporidia MB has the potential to disrupt and block Plasmodium transmission. Methods: A prospective cross-sectional survey was conducted in Zinder City (Zinder), Niger, from August to September 2022, using the CDC light trap technique to collect adult mosquitoes belonging to the Anopheles gambiae complex. The survey focused on collecting mosquitoes from three neighborhoods of Zinder (Birni, Kangna and Garin Malan, located in communes I, II and IV, respectively). Collected mosquitoes were sorted and preserved in 70% ethanol. PCR was used to identify host species and detect the presence of Microsporidia MB and Plasmodium falciparum infection. Results: Of the 257 Anopheles mosquitoes collected and identified by PCR, Anopheles coluzzii was the most prevalent species, accounting for 97.7% of the total. Microsporidia MB was exclusively detected in A. coluzzii, with a prevalence of 6.8% (17/251) among the samples. No significant difference in prevalence was found among the three neighborhoods. Only one An. coluzzii mosquito tested PCR-positive for P. falciparum. Conclusions: The results confirm the presence of Microsporidia MB in Anopheles mosquitoes in Zinder, Niger, indicating its potential use as a biotechnological intervention against malaria transmission. However, further studies are needed to determine the efficacy of Microsporidia MB to disrupt Plasmodium transmission as well as its impact on vector fitness. Graphical

Using bacteria to express and deliver anti-parasite molecules in mosquitoes is among the list of genetic tools to control malaria. The introduction and spread of transgenic bacteria through wild adult mosquitoes is one of the major challenges of this strategy. In prospect of future field experiments, an open field study with blank (without bacteria) attractive sugar bait (ASB) was performed under the assumption that transgenic bacteria would be spread to all sugar fed mosquitoes. Two types of ASB stations were developed, one with clay pots (CP) placed at mosquito resting sites and one with window entry traps (WET) placed inside inhabited houses. The ASB consisted in either glucose, honey or fruit cocktail solutions. In addition, mark-release-recapture (MRR) experiment of mosquitoes after feeding them with glucose was also conducted to check the proportion of the mosquito population that can be reached by the two ASB stations as well as its suitability to complement the ASB stations for disseminating bacteria. Overall, 88% of the mosquitoes were collected in the WET_ASB. The CP_ASB stations were much less attractive with the highest average of 82 ± 11 mosquitoes/day in the CP near the wood piles. The proportions of sugar fed mosquitoes upon ASB were low in both type of ASB stations, ~ 2% and ~ 14% in WET and CP, respectively. Honey solution was the most attractive solution compared to the glucose and the fruit cocktail solutions. The recapture rate in the MRR experiment was low: ~ 4.1% over 7 days. The WET_ASB looks promising to disseminate transgenic bacteria to endophilic West Africa Anopheles mosquito. However, this feeding station may not be fully effective and could be combined with the CP_ASB to also target outdoor resting mosquitoes. Overall, efforts are needed to improve the mosquito-feeding rates upon ASB.