Project Summary: Wolbachia endosymbionts hold strong potential for the control of vector-borne infections. Indeed, these bacteria are currently used to control dengue virus transmission by Aedes aegypti, where they combine the ability to spread through populations via Cytoplasmic Incompatibility (CI) with efficient blocking of arboviral infections. For decades, however, scientists have been puzzled by the apparent lack of both natural and artificial Wolbachia infections in Anopheles, the vectors of human malaria, with only a handful of examples reported. Wolbachia are genetically intractable, making functional study of both their reproductive manipulations and this host tropism difficult. However, my previous work suggests that a Wolbachia-derived CI-inducing factor, CifB, causes severe abnormalities in females (rescued by a different gene, cifA), that may in part prevent Wolbachia from establishing natural infections in Anopheles mosquitoes. I will elucidate the mechanisms regulating Wolbachia infections in Anopheles, while gaining general mechanistic insight into CI in insects. Using a combination of genetics, cell biology, molecular, genomic and bioinformatics approaches to address key questions related to Wolbachia toxicity and rescue across different insects, these studies will push the field of insect-endosymbiont interactions into new areas of inquiry and will also generate tools for the generation of Wolbachia-based strategies for the control of malaria-transmitting Anopheles populations. I will conduct the K99 phase of this award as a post-doctoral researcher in Dr. Flaminia Catteruccia’s group at the Harvard T.H. Chan School of Public Health. The laboratory is exceptionally well-equipped to establish transgenic insects and is one of the only vector biology labs with a robotic micro-injector and a COPAS Biosorter in addition to a world-class Plasmodium infection facility. Dr. Catteruccia is supported by the Howard Hughes Medical Institute, which allows flexibility of research funds which can be drawn upon for this project, and facilitates access to state-of-the-art equipment, facilitating expansive research in vector biology. In addition to Dr. Catteruccia’s expertise in mosquito reproduction, mosquito transgenesis, and host-parasite interactions, I will be supported by Dr. Zhiyong Xi, an expert at the forefront of Wolbachia-based vector control implementation in Aedes mosquitoes, and in establishing Wolbachia infections in Anopheles. Further, I will collaborate with Dr. John Beckmann, a molecular biologist and biochemist with expertise working with Wolbachia’s CI-inducing factors, and Dr. Lauren Childs, a mathematical modeling specialist. Additional mentors will include local researchers Dr. Daniel Neafsey, who has vast experience with functional genomics in Wolbachia, Anopheles, and Plasmodium, and Dr. Paul Garrity whose group excels at genetic engineering of various Dipterans. This combination of expertise will train me to have a unique perspective and skillset to apply to research questions related to tripartite interactions between Wolbachia, Anopheles and Plasmodium, with which I will be well- positioned to establish an independent laboratory with a distinct set of research capabilities and questions.

Project Narrative: Associations between malaria-transmitting Anopheles mosquitoes and Wolbachia bacteria are both rare in nature and challenging to establish in laboratory settings, precluding the utility of Wolbachia in malaria control. It is not understood why Anopheles seem to be largely incompatible with Wolbachia infections, despite these bacteria infecting an estimated 60% of insect species, including many other mosquitoes. In this proposal, I outline a plan to investigate the basis of this incompatibility and overcome it, while simultaneously pursuing an alternate use of Wolbachia-induced Cytoplasmic Incompatibility in a gene drive approach.